Publications by category
Journal articles
Thornton CR, Davies GE, Dougherty L (In Press). Development of a Monoclonal Antibody and a Lateral-Flow Device for the Rapid Serological Detection of a Mucorales-specific Biomarker. Frontiers in Cellular and Infection Microbiology
Thornton CR (2023). The Potential for Rapid Antigen Testing for Mucormycosis in the Context of COVID-19.
Expert Review of Molecular DiagnosticsAbstract:
The Potential for Rapid Antigen Testing for Mucormycosis in the Context of COVID-19
Introduction: Mucormycosis is a highly aggressive angio-invasive disease of humans caused by Mucorales fungi. Prior to the COVID-19 pandemic, mucormycosis was a rare mycosis typically seen in immunocompromised patients with hematological malignancies or in transplant recipients. During the second wave of the pandemic, there was a dramatic increase in the disease, especially in India, where a unique set of circumstances led to large numbers of life-threatening and disfiguring rhino-orbital-
cerebral mucormycosis (ROCM) infections.
Areas covered: the review examines mucormycosis as a super-infection of COVID-19 patients, and the risk factors for COVID-19-associated mucormycosis (CAM) that drove the ROCM epidemic in India. The limitations of current diagnostic procedures are identified, and the measures needed to improve the speed and accuracy of detection discussed.
Expert opinion: Despite increased awareness, global healthcare systems remain unprepared for further outbreaks of ROCM. Current diagnosis of the disease is slow and inaccurate, negatively impacting on patient survival. This is most evident in low- to middle income countries which lack suitably equipped diagnostic facilities for rapid identification of the infecting pathogens. Rapid antigen testing using point-of-care lateral-flow assays could potentially have aided in the quick and accurate diagnosis of the disease, allowing earlier intervention with surgery and Mucorales-active antifungal drugs.
Abstract.
Schwenk J, Maurer A, Beziere N, Fiz F, Boschetti F, Geistlich S, Seyfried D, Gunzer M, Reischl G, Wehrmueller JE, et al (2022). Antibody-guided Molecular Imaging of Aspergillus Lung Infections in Leukemia Patients. Journal of Nuclear Medicine, 63, 1450-1451.
Parslow BY, Thornton CR (2022). Continuing Shifts in Epidemiology and Antifungal Susceptibility Highlight the Need for Improved Disease Management of Invasive Candidiasis.
Microorganisms,
10(6).
Abstract:
Continuing Shifts in Epidemiology and Antifungal Susceptibility Highlight the Need for Improved Disease Management of Invasive Candidiasis
Invasive candidiasis (IC) is a systemic life-threatening infection of immunocompromised humans, but remains a relatively neglected disease among public health authorities. Ongoing assessments of disease epidemiology are needed to identify and map trends of importance that may necessitate improvements in disease management and patient care. Well established incidence increases, largely due to expanding populations of patients with predisposing risk factors, has led to increased clinical use and pressures on antifungal drugs. This has been exacerbated by a lack of fast, accurate diagnostics that have led treatment guidelines to often recommend preventative strategies in the absence of proven infection, resulting in unnecessary antifungal use in many instances. The consequences of this are multifactorial but a contribution to emerging drug resistance is of primary concern, with high levels of antifungal use heavily implicated in global shifts to more resistant Candida strains. Preserving and expanding the utility and number of antifungals should therefore be of the highest priority. This may be achievable through the development and use of biomarker tests, bringing about a new era in improved antifungal stewardship, as well as novel antifungals that offer favourable profiles by targeting Candida pathogenesis mechanisms over cell viability.
Abstract.
Davies GE, Thornton CR (2022). Development of a Monoclonal Antibody and a Serodiagnostic Lateral-Flow Device Specific to Rhizopus arrhizus (Syn. R. oryzae), the Principal Global Agent of Mucormycosis in Humans.
Journal of Fungi,
8(7).
Abstract:
Development of a Monoclonal Antibody and a Serodiagnostic Lateral-Flow Device Specific to Rhizopus arrhizus (Syn. R. oryzae), the Principal Global Agent of Mucormycosis in Humans
Mucormycosis is a highly aggressive angio-invasive disease of humans caused by fungi in the zygomycete order, Mucorales. Though a number of different species can cause mucormycosis, the principal agent of the disease worldwide is Rhizopus arrhizus, which accounts for the majority of rhino-orbital-cerebral, pulmonary, and disseminated infections in immunocompromised individuals. It is also the main cause of life-threatening infections in patients with poorly controlled diabetes mellitus, and in corticosteroid-treated patients with SARS-CoV-2 infection, where it causes the newly described disease, COVID-19-associated mucormycosis (CAM). Diagnosis currently relies on non-specific CT, a lengthy and insensitive culture from invasive biopsy, and a time-consuming histopathology of tissue samples. At present, there are no rapid antigen tests for the disease that detect biomarkers of infection, and which allow point-of-care diagnosis. Here, we report the development of an IgG1 monoclonal antibody (mAb), KC9, which is specific to Rhizopus arrhizus var. arrhizus (syn. Rhizopus oryzae) and Rhizopus arrhizus var. delemar (Rhizopus delemar), and which binds to a 15 kDa extracellular polysaccharide (EPS) antigen secreted during hyphal growth of the pathogen. Using the mAb, we have developed a competitive lateral-flow device (LFD) that allows rapid (30 min) and sensitive (~50 ng/mL running buffer) detection of the EPS biomarker, and which is compatible with human serum (limit of detection of ~500 ng/mL) and bronchoalveolar lavage fluid (limit of detection of ~100 ng/mL). The LFD, therefore, provides a potential novel opportunity for the non-invasive detection of mucormycosis caused by Rhizopus arrhizus.
Abstract.
Parslow BY, Thornton CR (2022). Distribution and Diagnostics of Invasive Candidiasis.
Scholarly Community EncyclopediaAbstract:
Distribution and Diagnostics of Invasive Candidiasis
Invasive candidiasis (IC) is a systemic life-threatening infection of immunocompromised humans, but remains a relatively neglected disease among public health authorities. Ongoing assessments of disease epidemiology are needed to identify and map trends of importance that may necessitate improvements in disease management and patient care. Well-established incidence increases, largely due to expanding populations of patients with pre-disposing risk factors, has led to increased clinical use and pressures on antifungal drugs. This has been exacerbated by a lack of fast, accurate diagnostics that have led treatment guidelines to often recommend preventative strategies in the absence of proven infection, resulting in unnecessary antifungal use in many instances. The consequences of this are multifactorial, but a contribution to emerging drug resistance is of primary concern, with high levels of antifungal use heavily implicated in global shifts to more resistant Candida strains.
Abstract.
Henneberg S, Hasenberg A, Maurer A, Neumann F, Bornemann L, Gonzales-Menendez I, Kraus A, Hasenberg M, Thornton CR, Pichler BJ, et al (2021). Antibody-guided in vivo imaging of Aspergillus fumigatus lung infections during anti-fungal azole treatment.
Nature Communications,
12Abstract:
Antibody-guided in vivo imaging of Aspergillus fumigatus lung infections during anti-fungal azole treatment
Invasive pulmonary aspergillosis (IPA) is a life-threatening lung disease of immunocompromised humans, caused by the opportunistic fungal pathogen Aspergillus fumigatus. Inadequacies in current diagnostic procedures mean that early diagnosis of the disease, critical to patient survival, remains a major clinical challenge, and is leading to the empiric use of antifungal drugs and emergence of azole resistance. A non-invasive procedure that allows both unambiguous detection of IPA, and its response to azole treatment, is therefore needed. Here, we show that a humanised Aspergillus-specific monoclonal antibody, dual labelled with a radionuclide and fluorophore, can be used in immunoPET/MRI in vivo and 3D light sheet fluorescence microscopy ex vivo to quantify early A. fumigatus lung infections and to monitor the efficacy of azole therapy. Our antibody-guided approach reveals that early drug intervention is critical to prevent complete invasion of the lungs by the fungus, and demonstrates the power of molecular imaging as a non-invasive procedure for tracking IPA in vivo.
Abstract.
Davies G, Singh O, Prattes J, Hoenigl M, Sheppard PW, Thornton C (2021). Aspergillus fumigatus and its Allergenic Ribotoxin Asp f I: Old Enemies but New Opportunities for Urine-based Detection of Invasive Pulmonary Aspergillosis using Lateral-Flow Technology.
Journal of Fungi,
7(1).
Abstract:
Aspergillus fumigatus and its Allergenic Ribotoxin Asp f I: Old Enemies but New Opportunities for Urine-based Detection of Invasive Pulmonary Aspergillosis using Lateral-Flow Technology
Invasive pulmonary aspergillosis (IPA) caused by Aspergillus fumigatus is a life-threatening lung disease of immunocompromised patients. Diagnosis currently relies on non-specific chest CT, culture of the fungus from invasive lung biopsy, and detection of the cell wall carbohydrate galactomannan (GM) in serum or in BAL fluids recovered during invasive bronchoscopy. Urine provides an ideal bodily fluid for the non-invasive detection of pathogen biomarkers, with current urine-based immunodiagnostics for IPA focused on GM. Surrogate protein biomarkers might serve to improve disease detection. Here, we report the development of a monoclonal antibody (mAb), PD7, which is specific to A. fumigatus and related species in the Section Fumigati, and which binds to its 18 kDa ribotoxin Asp f I. Using PD7, we show that the protein is secreted during hyphal development, and so represents an ideal candidate for detecting invasive growth. We have developed a lateral-flow device (Afu-LFD®) incorporating the mAb which has a limit of detection of ~15 ng Asp f I/mL urine. Preliminary evidence of the test’s diagnostic potential is demonstrated with urine from a patient with acute lymphoid leukaemia with probable IPA. The Afu-LFD® therefore provides a potential novel opportunity for non-invasive urine-based detection of IPA caused by A. fumigatus.
Abstract.
Amich J, Mokhtari Z, Strobel M, Vialetto E, Sheta D, Yu Y, Hartweg J, Kalleda N, Jarick K, Brede C, et al (2020). 3D light sheet fluorescence microscopy of lungs to dissect local host immune - Aspergillus fumigatus interactions.
mBio,
11Abstract:
3D light sheet fluorescence microscopy of lungs to dissect local host immune - Aspergillus fumigatus interactions
Aspergillus fumigatus is an opportunistic fungal pathogen that can cause life-threatening invasive lung infections in immunodeficient patients. The cellular and molecular processes of infection during onset, establishment and progression are highly complex and depend on both fungal attributes and the immune status of the host. Therefore, preclinical animal models are paramount to investigate and gain better insight into the infection process. Yet, despite their extensive use, commonly employed murine models of invasive pulmonary aspergillosis are not well understood due to analytical limitations. Here we present quantitative light sheet fluorescence microscopy (LSFM) to describe fungal growth and the local immune response in whole lungs at cellular resolution within its anatomical context. We analyzed three very common murine models of pulmonary aspergillosis based on immunosuppression with corticosteroids, chemotherapy-induced leukopenia or myeloablative irradiation. LSFM uncovered distinct architectures of fungal growth and degrees of tissue invasion in each model. Furthermore, LSFM revealed the spatial distribution, interaction and activation of two key immune cell populations in antifungal defense: alveolar macrophages and polymorphonuclear neutrophils. Interestingly, the patterns of fungal growth correlated with the detected effects of the immunosuppressive regimens on the local immune cell populations. Moreover, LSFM demonstrates that the commonly used intranasal route of spore administration did not result in a complete intra-alveolar deposition, as more than 60% of fungal growth occurred outside of the alveolar space. Hence, LSFM allows for more rigorous characterization than previously used methods of murine models of invasive pulmonary aspergillosis and pinpointing their strengths and limitations.
Abstract.
Gunzer M, Thornton CR, Beziere N (2020). Advances in the in vivo molecular imaging of invasive pulmonary aspergillosis.
Journal of Fungi,
6(4).
Abstract:
Advances in the in vivo molecular imaging of invasive pulmonary aspergillosis
Invasive pulmonary aspergillosis (IPA) is a life-threatening infection of immunocompromised patients with Aspergillus fumigatus, a ubiquitous environmental mould. While there are numerous functioning antifungal therapies, their high cost, substantial side effects and fear of overt resistance development preclude permanent prophylactic medication of risk-patients. Hence, a fast and definitive diagnosis of IPA is desirable, to quickly identify those patients that really require aggressive antimycotic treatment and to follow the course of the therapeutic intervention. Yet, despite decades of research into this issue such a diagnostic procedure is still not available. Here we discuss the array of currently available methods for IPA detection and their limits. We then show that molecular imaging using positron-emission-tomography (PET) combined with morphological computed tomography or magnetic imaging is highly promising to become a future non-invasive approach for IPA diagnosis and therapy monitoring, albeit still requiring thorough validation and relying on further acceptance and dissemination of the approach. Thereby our approach using the A. fumigatus-specific humanized monoclonal antibody hJF5 labelled with 64Cu as PET-tracer has proven highly effective in pre-clinical models and hence bears high potential for human application.
Abstract.
Thornton CR (2020). Detection of the 'big five' mold killers of humans: Aspergillus, Fusarium, Lomentospora, Scedosporium and Mucormycetes.
Advances in Applied Microbiology,
110, 1-61.
Abstract:
Detection of the 'big five' mold killers of humans: Aspergillus, Fusarium, Lomentospora, Scedosporium and Mucormycetes
Fungi are an important but frequently overlooked cause of morbidity and mortality in humans. Life-threatening fungal infections mainly occur in immunocompromised patients, and are typically caused by environmental opportunists that take advantage of a weakened immune system. The filamentous fungus Aspergillus fumigatus is the most important and well-documented mould pathogen of humans, causing a number of complex respiratory diseases, including invasive pulmonary aspergillosis, an often fatal disease in patients with acute leukemia or in immunosuppressed bone marrow or solid organ transplant recipients. However, non-Aspergillus moulds are increasingly reported as agents of disseminated diseases, with Fusarium, Scedosporium, Lomentospora and mucormycete species now firmly established as pathogens of immunosuppressed and immunocompetent individuals. Despite well-documented risk factors for invasive fungal diseases, and increased awareness of the risk factors for life-threatening infections, the number of deaths attributable to moulds is likely to be severely underestimated driven, to a large extent, by the lack of readily accessible, cheap, and accurate tests that allow detection and differentiation of infecting species. Early diagnosis is critical to patient survival but, unlike Aspergillus diseases, where a number of CE-marked or FDA-approved biomarker tests are now available for clinical diagnosis, similar tests for fusariosis, scedosporiosis and mucormycosis remain experimental, with detection reliant on insensitive and slow culture of pathogens from invasive bronchoalveolar lavage fluid, tissue biopsy, or from blood. This review examines the ecology, epidemiology, and contemporary methods of detection of these mould pathogens, and the obstacles to diagnostic test development and translation of novel biomarkers to the clinical setting.
Abstract.
Amich J, Mokhtari Z, Strobel M, Vialetto E, Kalleda N, Jarick KJ, Brede C, Jordan-Garrote A-L, Thusek S, Schmiedgen K, et al (2019). 3D light sheet fluorescence microscopy of lungs to dissect local host immune - Aspergillus fumigatus interactions.
bioRxiv, 1-26.
Abstract:
3D light sheet fluorescence microscopy of lungs to dissect local host immune - Aspergillus fumigatus interactions
Aspergillus fumigatus is an opportunistic fungal pathogen that can cause life-threatening invasive lung infections in immunodeficient patients. The cellular and molecular processes of infection during onset, establishment and progression are highly complex and depend on both fungal attributes and the immune status of the host. Therefore, preclinical animal models are paramount to investigate and gain better insight into the infection process. Yet, despite their extensive use, commonly employed murine models of invasive pulmonary aspergillosis are not well understood due to analytical limitations. Here we present quantitative light sheet fluorescence microscopy (LSFM) to describe fungal growth and the local immune response in whole lungs at cellular resolution within its anatomical context. We analyzed three very common murine models of pulmonary aspergillosis based on immunosuppression with corticosteroids, chemotherapy-induced leukopenia or myeloablative irradiation. LSFM uncovered distinct architectures of fungal growth and degrees of tissue invasion in each model. Furthermore, LSFM revealed the spatial distribution, interaction and activation of two key immune cell populations in antifungal defense: alveolar macrophages and polymorphonuclear neutrophils. Interestingly, the patterns of fungal growth correlated with the detected effects of the immunosuppressive regimens on the local immune cell populations. Moreover, LSFM demonstrates that the commonly used intranasal route of spore administration did not result in the desired intra-alveolar deposition, as more than 60% of fungal growth occurred outside of the alveolar space. Hence, LSFM allows for more rigorous characterization of murine models of invasive pulmonary aspergillosis and pinpointing their strengths and limitations.
Abstract.
Thornton CR (2018). Molecular Imaging of Invasive Pulmonary Aspergillosis using ImmunoPET/MRI: the Future Looks Bright.
Frontiers in Microbiology,
9Abstract:
Molecular Imaging of Invasive Pulmonary Aspergillosis using ImmunoPET/MRI: the Future Looks Bright
Invasive pulmonary aspergillosis (IPA) is a life-threatening lung disease of immuno-compromised humans caused by the ubiquitous environmental mould Aspergillus. Biomarker tests for the disease lack sensitivity and specificity, and culture of the fungus from invasive lung biopsy is slow, insensitive, and undesirable in critically ill patients. A Computed Tomogram (CT) of the chest offers a simple non-intrusive diagnostic procedure for rapid decision-making, and so is used in many haematology units to drive antifungal treatment. However, radiological indicators that raise the suspicion of IPA are either transient signs in the early stages of the disease, or are not specific for Aspergillus infection, with other angio-invasive moulds or bacterial pathogens producing comparable radiological manifestations in a chest CT. Improvements to the specificity of radiographic imaging of IPA have been attempted by coupling CT and Positron Emission Tomography (PET) with [18F]FDG, a marker of metabolic activity well-suited to cancer imaging, but with limited use in invasive fungal disease diagnostics due to its inability to differentiate between infectious etiologies, cancer, and inflammation. Bioluminescence imaging using single genetically modified strains of Aspergillus fumigatus has enabled in vivo monitoring of IPA in animal models of disease. For in vivo detection of Aspergillus lung infections in humans, radiolabelled Aspergillus-specific monoclonal antibodies, and iron siderophores, hold enormous potential for clinical diagnosis. This review examines the different experimental technologies used to image IPA, and recent advances in state-of-the-art molecular imaging of IPA using antibody-guided Positron Emission Tomography/Magnetic Resonance Imaging (immunoPET/MRI).
Abstract.
Box H, Negri C, Livermore J, Whalley S, Johnson A, McEntee L, Alastruey-Izquierdo A, Meis J, Thornton CR, Hope W, et al (2018). Pharmacodynamics of Voriconazole for Invasive Pulmonary Scedosporiosis.
Antimicrobial Agents and Chemotherapy,
62(5).
Abstract:
Pharmacodynamics of Voriconazole for Invasive Pulmonary Scedosporiosis
Scedosporium apiospermum is a medically important fungal pathogen that causes a wide range of infections in humans. There are relatively few antifungal agents that are active against Scedosporium spp. Little is known about the pharmacodynamics of voriconazole against Scedosporium. Both static and dynamic in vitro models of invasive scedosporiosis were developed. Monoclonal antibodies that target a soluble cell wall antigen secreted by Scedosporium apiospermum were used to describe the pharmacodynamics of voriconazole. Mathematical pharmacokinetic-pharmacodynamic models were fitted to the data to estimate the drug exposure required to suppress the release of fungal antigen. The experimental results were bridged to humans using Monte Carlo simulation. All 3 strains of S. apiospermum tested invaded through the cellular bilayer of the in vitro models and liberated antigen. There was a concentration-dependent decline in antigen with near maximal antifungal activity in all 3 strains with 10 mg/L. Similarly, there was a drug exposure dependent decline in circulating antigen in the dynamic model and complete suppression of antigen with an AUC of approximately 80 mg.h/L. A regression of the AUC:MIC versus area under the antigen time curve showed that near maximal effect was obtained with AUC:MIC of approximately 100. Monte Carlo simulation suggested that only isolates with an MIC of 0.5 mg/L enable pharmacodynamic targets to be acheived with a standard regimen of voriconazole. Isolates with higher MICs may need higher drug exposure targets than are currently recommended for other fungi.
Abstract.
Morad HOJ, Wild A-M, Wiehr S, Davies G, Maurer A, Pichler BJ, Thornton CR (2018). Pre-Clinical Imaging of Invasive Candidiasis using ImmunoPET/MR.
Frontiers in Microbiology,
9, 1-15.
Abstract:
Pre-Clinical Imaging of Invasive Candidiasis using ImmunoPET/MR
The human commensal yeast Candida is the 4th most common cause of hospital-acquired bloodstream infections, with C. albicans accounting for the majority of the >400,000 life-threatening infections annually. Diagnosis of invasive candidiasis (IC), a disease encompassing candidemia (blood-borne yeast infection) and deep-seated organ infections, is a major challenge since clinical manifestations of the disease are indistinguishable from viral, bacterial and other fungal diseases, and diagnostic tests for biomarkers in the bloodstream such as PCR, ELISA and pan-fungal β-D-glucan lack either standardisation, sensitivity or specificity. Blood culture remains the gold standard for diagnosis, but test sensitivity is poor and turn-around time slow. Furthermore, cultures can only be obtained when the yeast resides in the bloodstream, with samples recovered from hematogenous infections often yielding negative results. Consequently, there is a pressing need for a diagnostic test that allows the identification of metastatic foci in deep-seated Candida infections, without the need for invasive biopsy. Here, we report the development of a highly specific mouse IgG3 monoclonal antibody (MC3) that binds to a putative β-1,2-mannan epitope present in high molecular weight mannoproteins and phospholipomannans on the surface of yeast and hyphal morphotypes of C. albicans, and its use as a [64Cu]NODAGA-labeled tracer for whole-body pre-clinical imaging of deep-seated C. albicans infections using antibody-guided positron emission tomography and magnetic resonance imaging (immunoPET/MRI). When used in a mouse intravenous (i.v.) challenge model that faithfully mimics disseminated C. albicans infections in humans, the [64Cu]NODAGA-MC3 tracer accurately detects infections of the kidney, the principal site of blood-borne candidiasis. Using a strain of the emerging human pathogen Candida auris that reacts with MC3 in vitro, but which is non-infective in i.v. challenged mice, we demonstrate the accuracy of the tracer in diagnosing invasive infections in vivo. This pre-clinical study demonstrates the principle of. antibody-guided molecular imaging for detection of deep organ infections in IC, without the need for invasive tissue biopsy.
Abstract.
Liew N, Mazon Moya MJ, Wierzbicki C, Hollinshead M, Dillon MJ, Thornton CR, Ellison A, Cable J, Fisher MC, Mostowy S, et al (2017). Chytrid fungus infection in zebrafish demonstrates that the pathogen can parasitise non-amphibian vertebrate hosts.
Nature Communications,
8, 15048-15048.
Abstract:
Chytrid fungus infection in zebrafish demonstrates that the pathogen can parasitise non-amphibian vertebrate hosts
Aquatic chytrid fungi threaten amphibian biodiversity worldwide owing to their ability to rapidly expand their geographical distributions and to infect a wide range of hosts. Combating this risk requires an understanding of chytrid host range to identify potential reservoirs of infection and to safeguard uninfected regions through enhanced biosecurity. Here we extend our knowledge on the host range of the chytrid Batrachochytrium dendrobatidis by demonstrating infection of a non-amphibian vertebrate host, the zebrafish. We observe dose-dependent mortality and show that chytrid can infect and proliferate on zebrafish tissue. We also show that infection phenotypes (fin erosion, cell apoptosis and muscle degeneration) are direct symptoms of infection. Successful infection is dependent on disrupting the zebrafish microbiome, highlighting that, as is widely found in amphibians, commensal bacteria confer protection against this pathogen. Collectively, our findings greatly expand the limited tool kit available to study pathogenesis and host response to chytrid infection.
Abstract.
Hoenigl M, Eigl S, Heldt S, Duettmann W, Thornton C, Prattes J (2017). Clinical Evaluation of the Newly Formatted Lateral-Flow Device for Invasive Pulmonary Aspergillosis.
Mycoses,
00, 1-4.
Abstract:
Clinical Evaluation of the Newly Formatted Lateral-Flow Device for Invasive Pulmonary Aspergillosis
The study evaluated the newly formatted Aspergillus-specific lateral-flow-device (LFD), and compared its performance to the original prototype “old” LFD test using BALF samples from 28 patients (14 patients with probable/proven invasive pulmonary aspergillosis [IPA] and 14 patients with no evidence for IPA). A total of 10/14 (71%) of BALF samples from patients with probable/proven IPA resulted positive with the new LFD, including 8/9 with true-positive and 2/5 with false-negative results with the old LFD. All 14 samples from patients without IPA resulted negative with the new LFD; specificity of the new LFD was significantly improved compared to the old LFD.
Abstract.
Duvaux L, Shiller J, Vandeputte P, Duge de Bernoville T, Thornton CR, Papon N, Le Cam B, Bouchara J-P, Gasteboise A (2017). Draft genome sequence of the human pathogenic fungus Scedosporium boydii.
Genome Announcements,
5, e00871-17-e00871-17.
Abstract:
Draft genome sequence of the human pathogenic fungus Scedosporium boydii
The opportunistic fungal pathogen Scedosporium boydii is the most common Scedosporium species in French patients with cystic fibrosis. Here we present the first genome report for S. boydii, providing a resource which may enable the elucidation of the pathogenic mechanisms in this species.
Abstract.
Pateau V, Razafimandimby B, Vandeputte P, Guillemette T, Thornton CR, Bouchara J-P, Giraud S (2017). Gene disruption in Scedosporium aurantiacum: proof of concept with the disruption of SODC gene encoding a cytosolic Cu,Zn-superoxide dismutase.
Mycopathologia,
183(1), 241-249.
Abstract:
Gene disruption in Scedosporium aurantiacum: proof of concept with the disruption of SODC gene encoding a cytosolic Cu,Zn-superoxide dismutase
Scedosporium species are opportunistic pathogens responsible for a large variety of infections in humans. An increasing occurrence was observed in patients with underlying conditions such as immunosuppression or cystic fibrosis. Indeed, the genus Scedosporium ranks the second among the filamentous fungi colonizing the respiratory tracts of the CF patients. To date, there is very scarce information on the pathogenic mechanisms, at least in part because of the limited genetic tools available. In the present study, we successfully developed an efficient transformation and targeted gene disruption approach on the species Scedosporium aurantiacum. The disruption cassette was constructed using double-joint PCR procedure, and resistance to hygromycin B as the selection marker. This proof of concept was performed on the functional gene SODC encoding the Cu,Zn-superoxide dismutase. Disruption of the SODC gene improved susceptibility of the fungus to oxidative stress. This technical advance should open new research areas and help to better understand the biology of Scedosporium species.
Abstract.
Shah A, Abdolrasouli A, Schelenz S, Thornton C, Ni MZ, Devaraj A, Devic N, Ward L, Carby M, Reed A, et al (2017). Latent class modelling for pulmonary aspergillosis diagnosis in lung transplant recipients.
Thorax,
72(3), A13-A14.
Abstract:
Latent class modelling for pulmonary aspergillosis diagnosis in lung transplant recipients
Rationale Timely, accurate diagnosis of invasive aspergillosis (IA) is key to enable initiation of antifungal therapy in lung transplantation. Despite promising novel fungal biomarkers, the lack of a diagnostic gold-standard creates difficulty in determining utility.
Objectives This study aimed to use latent class modelling of fungal diagnostics to classify lung transplant recipients (LTR) with IA in a large single centre.
Methods Regression models were used to compare composite biomarker testing of bronchoalveolar lavage to clinical and EORTC-MSG guideline-based diagnosis of IA with mortality used as a surrogate primary outcome measure. Bootstrap anal- ysis identified radiological features associated with IA. Bayesian latent class modelling was used to define IA.
Measurements and Main Results a clinical diagnosis of fungal infection (P =
Abstract.
Davies G, Rolle A-M, Maurer A, Spycher PR, Schillinger C, Solouk-Saran D, Hasenberg M, Weski J, Foslet J, Dubois A, et al (2017). Towards Translational ImmunoPET/MR Imaging of Invasive Pulmonary Aspergillosis: the Humanised Monoclonal Antibody JF5 Detects Aspergillus Lung Infections in Vivo.
Theranostics,
7, 3398-3414.
Abstract:
Towards Translational ImmunoPET/MR Imaging of Invasive Pulmonary Aspergillosis: the Humanised Monoclonal Antibody JF5 Detects Aspergillus Lung Infections in Vivo
Invasive pulmonary aspergillosis (IPA) is a life-threatening lung disease of hematological malignancy and bone marrow transplant patients caused by the ubiquitous environmental fungus Aspergillus fumigatus. Current diagnostic tests for the disease lack sensitivity as well as specificity, and culture of the fungus from invasive lung biopsy, considered the gold standard for IPA detection, is slow and often not possible in critically ill patients. In a previous study, we reported the development of a novel non-invasive procedure for IPA diagnosis based on antibody-guided positron emission tomography and magnetic resonance imaging (immunoPET/MRI) using a [64Cu]DOTA-labeled mouse monoclonal antibody (mAb), mJF5, specific to Aspergillus. To enable translation of the tracer to the clinical setting, we report here the development of a humanised version of the antibody (hJF5), and pre-clinical imaging of lung infection using a [64Cu]NODAGA-hJF5 tracer. The humanised antibody tracer shows a significant increase in in vivo biodistribution in A. fumigatus infected lungs compared to its radiolabeled murine counterpart [64Cu]NODAGA-mJF5. Using reverse genetics of the pathogen, we show that the antibody binds to the antigenic determinant 1,5-galactofuranose (Galf) present in a diagnostic mannoprotein antigen released by the pathogen during invasive growth in the lung. The absence of the epitope Galf in mammalian carbohydrates, coupled with the enhanced imaging capabilities of the hJF5 antibody, means that the [64Cu]NODAGA-hJF5 tracer developed here represents an ideal candidate for the diagnosis of IPA and translation to the clinical setting.
Abstract.
Dillon MJ, Bowkett AE, Bungard MJ, Beckman KM, O'Brien MF, Bates K, Fisher MC, Stevens JR, Thornton CR (2017). Tracking the amphibian pathogens Batrachochytrium dendrobatidis and Batrachochytrium salamandrivorans using a highly specific monoclonal antibody and lateral-flow technology.
Microbial Biotechnology,
10(2), 381-394.
Abstract:
Tracking the amphibian pathogens Batrachochytrium dendrobatidis and Batrachochytrium salamandrivorans using a highly specific monoclonal antibody and lateral-flow technology
The fungus Batrachochytrium dendrobatidis (Bd) causes chytridiomycosis, a lethal epizootic disease of amphibians. Rapid identification of the pathogen and biosecurity is essential to prevent its spread, but current laboratory-based tests are time-consuming and require specialist equipment. Here, we describe the generation of an IgM monoclonal antibody (mAb), 5C4, specific to Bd as well as the related salamander and newt pathogen Batrachochytrium salamandrivorans (Bsal). The mAb, which binds to a glycoprotein antigen present on the surface of zoospores, sporangia and zoosporangia, was used to develop a lateral-flow assay (LFA) for rapid (15 min) detection of the pathogens. The LFA detects known lineages of Bd and also Bsal, as well as the closely related fungus Homolaphlyctis polyrhiza, but does not detect a wide range of related and unrelated fungi and oomycetes likely to be present in amphibian habitats. When combined with a simple swabbing procedure, the LFA was 100% accurate in detecting the water-soluble 5C4 antigen present in skin, foot and pelvic samples from frogs, newts and salamanders naturally infected with Bd or Bsal. Our results demonstrate the potential of the portable LFA as a rapid qualitative assay for tracking these amphibian pathogens and as an adjunct test to nucleic acid-based detection methods.
Abstract.
Escudero N, Ferreira SR, Lopez-Moya F, Naranjo-Ortiz MA, Marin-Ortiz AI, Thornton CR, Lopez-Llorca LV (2016). Chitosan enhances parasitism of Meloidogyne javanica eggs by the nematophagous fungus Pochonia chlamydosporia.
Fungal Biology,
120(4), 572-585.
Abstract:
Chitosan enhances parasitism of Meloidogyne javanica eggs by the nematophagous fungus Pochonia chlamydosporia
Pochonia chlamydosporia (Pc), a nematophagous fungus and root endophyte, uses appressoria and extracellular enzymes, principally proteases, to degrade the eggs of plant parasitic nematodes (PPN). Unlike other fungi, Pc is resistant to chitosan, a deacetylated form of chitin, used in agriculture as a biopesticide to control plant pathogens. In the present work, we show that chitosan increases pathogenicity of Pc to Meloidogyne javanica by stimulating appressorial differentiation and parasitism of eggs. Using antibodies specific to the Pc enzymes VCP1 (a subtilisin), and SCP1 (a serine carboxypeptidase), we demonstrate chitosan elicitation of the fungal proteases during the parasitic process. Chitosan increases VCP1 immuno-labelling in the cells walls of Pc conidia, hyphal tips of germinating spores, and in appressoria on infected M. javanica eggs. These results support the role of proteases in pathogenicity of the fungus and their activation by chitosan. Phylogenetic analysis of the Pc genome reveals a large diversity of subtilisins (S8) and serine carboxypeptidases (S10). The VCP1 group in the S8 tree shows evidence of gene duplication indicating recent adaptations to nutrient sources. Our results show that chitosan enhances Pc pathogenicity through increased proteolytic activities and can be used to improve the efficacy of M. javanica biocontrol.
Abstract.
Al-Maqtoofi M, Thornton CR (2016). Detection of human pathogenic Fusarium species in hospital and communal sink biofilms by using a highly specific monoclonal antibody.
Environmental Microbiology,
18, 3620-3634.
Abstract:
Detection of human pathogenic Fusarium species in hospital and communal sink biofilms by using a highly specific monoclonal antibody
The fungus Fusarium is well known as a plant pathogen, but has recently emerged as an opportunistic pathogen of humans. Habitats providing direct human exposure to infectious propagules are largely unknown, but there is growing evidence that plumbing systems are sources of human pathogenic strains in the Fusarium solani species complex (FSSC) and Fusarium oxysporum species complex (FOSC), the most common groups infecting humans. Here, we use a newly developed Fusarium-specific monoclonal antibody (mAb ED7) to track FSSC and FOSC strains in sink drain biofilms by detecting its target antigen, an extracellular 200kDa carbohydrate, in saline swabs. The antigen was detectable in 52% of swab samples collected from sinks across a University campus and a tertiary care hospital. The mAb was 100% accurate in detecting FSSC, FOSC and F. dimerum species complex (FDSC) strains that were present, as mixed fungal communities, in 83% of sink drain biofilms. Specificity of the ELISA was confirmed by sequencing of the internally transcribed spacer 1 (ITS1)-5.8S-ITS2 rRNA-encoding regions of culturable yeasts and molds that were recovered using mycological culture, while translation elongation factor (TEF)-1α analysis of Fusarium isolates included FSSC 1-a, FOSC 33 and FDSC ET-gr, the most common clinical pathotypes in each group.
Abstract.
Sharpe RA, Le Cocq K, Nikolaou V, Osborne NJ, Thornton CR (2016). Identifying risk factors for exposure to culturable allergenic fungi in energy efficient homes by using highly specific monoclonal antibodies.
Environmental Research,
144, 32-42.
Abstract:
Identifying risk factors for exposure to culturable allergenic fungi in energy efficient homes by using highly specific monoclonal antibodies
The aim of this study was to determine the accuracy of monoclonal antibodies (mAbs) in identifying culturable allergenic fungi present in visible mould growth in energy efficient homes, and to identify risk factors for exposure to these known allergenic fungi. Swabs were taken from fungal contaminated surfaces and culturable yeasts and moulds isolated by using mycological culture. Soluble antigens from cultures were tested by ELISA using mAbs specific to the culturable allergenic fungi Aspergillus and Penicillium spp. Ulocladium, Alternaria, and Epicoccum spp. Cladosporium spp. Fusarium spp. and Trichoderma spp. Diagnostic accuracies of the ELISA tests were determined by sequencing of the internally transcribed spacer 1 (ITS1)-5.8S-ITS2-encoding regions of recovered fungi following ELISA. There was 100% concordance between the two methods, with ELISAs providing genus-level identity and ITS sequencing providing species-level identities (210 out of 210 tested). Species of Aspergillus/Penicillium, Cladosporium, Ulocladium/Alternaria/Epicoccum, Fusarium and Trichoderma were detected in 82% of the samples. The presence of condensation was associated with an increased risk of surfaces being contaminated by Aspergillus/Penicillium spp. and Cladosporium spp. whereas moisture within the building fabric (water ingress/rising damp) was only associated with increased risk of Aspergillus/Penicillium spp. Property type and energy efficiency levels were found to moderate the risk of indoor surfaces becoming contaminated with Aspergillus/Penicillium and Cladosporium which in turn was modified by the presence of condensation, water ingress and rising damp, consistent with previous literature.
Abstract.
Rolle AM, Hasenberg M, Thornton CR, Solouk-Saran D, Männ L, Weski J, Maurer A, Fischer E, Spycher PR, Schibli R, et al (2016). ImmunoPET/MR imaging allows specific detection of Aspergillus fumigatus lung infection in vivo.
Proceedings of the National Academy of Sciences of the United States of America,
113(8), E1026-E1033.
Abstract:
ImmunoPET/MR imaging allows specific detection of Aspergillus fumigatus lung infection in vivo
Invasive pulmonary aspergillosis (IPA) is a life-threatening lung disease caused by the fungus Aspergillus fumigatus, and is a leading cause of invasive fungal infection-related mortality and morbidity in patients with hematological malignancies and bone marrow transplants. We developed and tested a novel probe for noninvasive detection of A. fumigatus lung infection based on antibody-guided positron emission tomography and magnetic resonance (immunoPET/MR) imaging. Administration of a [64Cu]DOTA-labeled A. fumigatus-specific monoclonal antibody (mAb), JF5, to neutrophil-depleted A. fumigatus-infected mice allowed specific localization of lung infection when combined with PET. Optical imaging with a fluorochrome-labeled version of the mAb showed colocalization with invasive hyphae. The mAbbased newly developed PET tracer [64Cu]DOTA-JF5 distinguished IPA from bacterial lung infections and, in contrast to [18F]FDG-PET, discriminated IPA from a general increase in metabolic activity associated with lung inflammation. To our knowledge, this is the first time that antibody-guided in vivo imaging has been used for noninvasive diagnosis of a fungal lung disease (IPA) of humans, an approach with enormous potential for diagnosis of infectious diseases and with potential for clinical translation.
Abstract.
Wiehr S, Warnke P, Rolle A-M, Schutz M, Kohlhofer U, Quintanilla-Martinez de Fend L, Maurer A, Thornton C, Boschetti F, Reischl G, et al (2016). New pathogen-specific immunoPET/MR tracer for molecular imaging of a systemic bacterial infection.
Oncotarget,
7, 10990-11001.
Abstract:
New pathogen-specific immunoPET/MR tracer for molecular imaging of a systemic bacterial infection
The specific and rapid detection of Enterobacteriaceae, the most frequent cause of gram-negative bacterial infections in humans, remains a major challenge. We developed a non-invasive method to rapidly detect systemic Yersinia enterocolitica infections using immunoPET (antibody-targeted positron emission tomography) with [64Cu]NODAGA-labeled Yersinia-specific polyclonal antibodies targeting the outer membrane protein YadA. In contrast to the tracer [18F]FDG, [64Cu]NODAGA-YadA uptake co-localized in a dose dependent manner with bacterial lesions of Yersinia-infected mice, as detected by magnetic resonance (MR) imaging. This was accompanied by elevated uptake of [64Cu]NODAGA-YadA in infected tissues, in ex vivo biodistribution studies, whereas reduced uptake was observed following blocking with unlabeled anti-YadA antibody. We show, for the first time, a bacteria-specific, antibody-based, in vivo imaging method for the diagnosis of a Gram-negative enterobacterial infection as a proof of concept, which may provide new insights into pathogen-host interactions.
Abstract.
Al-Laaeiby A, Kershaw M, Penn T, Thornton CR (2016). Targeted disruption of melanin biosynthesis genes in the human pathogenic fungus Lomentospora prolificans and its consequences for pathogen survival.
International Journal of Molecular Sciences,
17(4), 1-18.
Abstract:
Targeted disruption of melanin biosynthesis genes in the human pathogenic fungus Lomentospora prolificans and its consequences for pathogen survival
The dematiaceous (melanised) fungus Lomentospora (Scedosporium) prolificans is a life-threatening opportunistic pathogen of immunocompromised humans, resistant to anti-fungal drugs. Melanin has been shown to protect human pathogenic fungi against antifungal drugs, oxidative killing and environmental stresses. To determine the protective role of melanin in L. prolificans to oxidative killing (H2O2), UV radiation and the polyene anti-fungal drug amphotericin B, targeted gene disruption was used to generate mutants of the pathogen lacking the dihydroxynaphthalene (DHN)-melanin biosynthetic enzymes polyketide synthase (PKS1), tetrahydroxynapthalene reductase (4HNR) and scytalone dehydratase (SCD1). Infectious propagules (spores) of the wild-type strain 3.1 were black/brown, whereas spores of the PKS-deficient mutant ΔLppks1::hph were white. Complementation of the albino mutant ΔLppks1::hph restored the black-brown spore pigmentation, while the 4HNR-deficient mutant ΔLp4hnr::hph and SCD-deficient mutant ΔLpscd1::hph both produced orange-yellow spores. The mutants ΔLppks1::hph and ΔLp4hnr::hph showed significant reductions in spore survival following H2O2 treatment, while spores of ΔLpscd1::hph and the ΔLppks1::hph complemented strain ΔLppks1::hph:PKS showed spore survivals similar to strain 3.1. Spores of the mutants ΔLp4hnr::hph and ΔLpscd1::hph and complemented strain ΔLppks1::hph:PKS showed spore survivals similar to 3.1 following exposure to UV radiation, but survival of ΔLppks1::hph spores was significantly reduced compared to the wild-type strain. Strain 3.1 and mutants ΔLp4hnr::hph and ΔLppks1::hph:PKS were resistant to amphotericin B while, paradoxically, the PKS1- and SCD1-deficient mutants showed significant increases in growth in the presence of the antifungal drug. Taken together, these results show that while melanin plays a protective role in the survival of the pathogen to oxidative killing and UV radiation, melanin does not contribute to its resistance to amphotericin B.
Abstract.
Shaw S, Le Cocq K, Paszkiewicz K, Moore K, Winsbury R, Studholme D, Salmon D, Thornton CR, Grant MR (2016). Transcriptional reprogramming underpins enhanced plant
growth promotion by the biocontrol fungus Trichoderma hamatum GD12
during antagonistic interactions with Sclerotinia sclerotiorum in
soil.
Molecular Plant Pathology,
17, 1425-1441.
Abstract:
Transcriptional reprogramming underpins enhanced plant
growth promotion by the biocontrol fungus Trichoderma hamatum GD12
during antagonistic interactions with Sclerotinia sclerotiorum in
soil
The free-living soil fungus Trichoderma hamatum strain GD12 is notable amongst Trichoderma strains in both controlling plant diseases and in stimulating plant growth, a property enhanced during its antagonistic interactions with pathogens in soil. These attributes, alongside its markedly expanded genome and proteome compared to other biocontrol and plant growth promoting Trichoderma strains, imply a rich potential for sustainable alternatives to synthetic pesticides and fertilisers for controlling plant disease and increasing yields. The purpose of this study was to investigate the transcriptional responses of GD12 underpinning its biocontrol and plant growth promotion capabilities during antagonistic interactions with the pathogen Sclerotinia sclerotiorum in soil. Using an extensive mRNA-seq study capturing different time points during the pathogen-antagonist interaction in soil, we show that dynamic and biphasic signatures in the GD12 transcriptome underpin its biocontrol and plant (lettuce) growth promotional activities. Functional predictions of differentially expressed genes demonstrate the enrichment of transcripts encoding proteins involved in transportation and oxidation-reduction reactions during both processes and an over-representation of siderophores. We identify a biphasic response during biocontrol characterised by a significant induction of transcripts encoding small-secreted cysteine rich proteins, secondary metabolite producing gene clusters and genes unique to GD12. These data support the hypothesis that Sclerotinia biocontrol is mediated by the synthesis and secretion of antifungal compounds and that GD12's unique reservoir of uncharacterised genes is actively recruited during effective biological control of a plurivorous plant pathogen.
Abstract.
Solouk D, Bergmueller E, Buffen K, Rolle A-M, Thornton C, Weski J, Gunzer M, Hasenberg M (2016). Viability of a promising antibody (JF5) for the fast detection of invasive pulmonary aspergillosis (IPA).
MYCOSES,
59, 32-32.
Author URL.
Prattes J, Lackner M, Eigl S, Reischies F, Raggam RB, Koidl C, Flick H, Wolfler A, Neumeister P, Thornton CR, et al (2015). Diagnostic accuracy of the Aspergillus specific Bronchoalveolar Lavage Lateral-Flow Assay in Haematological Malignancy Patients. Mycoses: diagnosis, therapy and prophylaxis of fungal diseases, 58(8), 461-469.
Sharpe R, Thornton CR, Nikolaou V, Osborne N (2015). Fuel poverty increases risk of mould contamination, regardless of adult risk perception & ventilation in social housing properties. Environment International, 79, 115-129.
Eigl S, Prattes J, Reischies FM, Raggam RB, Spiess B, Reinwald M, Buchheidt D, Thornton CR, Krause R, Flick H, et al (2015). Galactomannan Testing and Aspergillus PCR in same-day bronchoalveolar lavage and blood samples obtained from patients with hematological malignancies at risk for invasive mould infection.
MYCOSES,
58, 157-157.
Author URL.
Sharpe R, Thornton CR, Nikolaou V, Osborne N (2015). Higher energy efficient homes are associated with increased risk of doctor diagnosed asthma in a UK sub population. Environment International, 75, 234-244.
Thornton CR, Ryder LS, Le Cocq K, Soanes DM (2015). Identifying the emerging human pathogen Scedosporium prolificans by using a species-specific monoclonal antibody that binds to the melanin biosynthetic enzyme tetrahydroxynaphthalene reductase. Environmental Microbiology, 17(4), 1023-1048.
Bustin SA, Thornton CR, Shannon M, Agrawal SG, Lass-Floerl C, Mutschlechner W, Roberts D, Johnson G (2015). Immuno PCR-based assays for the early detection of Aspergillus species.
MYCOSES,
58, 28-28.
Author URL.
Thornton CR, Wills OE (2015). Immunodetection of Fungal and Oomycete Pathogens: Established and Emerging Threats to Human Health, Animal Welfare and Global Food Security. Critical Reviews in Microbiology, 41(1), 27-51.
Osborne NJ, Thornton CR, Sharpe RA (2015). Indoor Fungal Exposure and Allergic Respiratory Disease.
Current Allergy and Asthma Reports,
15(12).
Abstract:
Indoor Fungal Exposure and Allergic Respiratory Disease
A gathering body of evidence has repeatedly revealed associations between indoor fungi and initiation, promotion, and exacerbation of allergic respiratory disease. The relationship between the exposure and outcome are complicated by the difficulties in measuring both exposure and outcome, the multifactorial nature of the disease, and the wide range of potential confounders. New technologies are becoming available that may enable better measurement of exposure and tighter case definitions so as to build more confidence in the associations discovered. The growing strength of the evidence base will aid the design of future public health interventions and generate new hypotheses on the cause of the rapid increase in allergic respiratory disease prevalence.
Abstract.
Sharpe RA, Bearman N, Thornton CR, Husk K, Osborne NJ (2015). Indoor fungal diversity and asthma: a meta-analysis and systematic review of risk factors.
Journal of Allergy and Clinical Immunology,
135(1), 110-122.
Abstract:
Indoor fungal diversity and asthma: a meta-analysis and systematic review of risk factors
Background Indoor dampness increases the risk of indoor fungal growth. A complex interaction between occupant behaviors and the built environment are thought to affect indoor fungal concentrations and species diversity, which are believed to increase the risk of having asthma, exacerbation of asthma symptoms, or both. To date, no systematic review has investigated this relationship.
Abstract.
Sharpe RA, Bearman N, Thornton CR, Husk K, Osborne NJ (2015). Indoor fungal diversity and asthma: a metaanalysis and systematic review of risk factors. Journal of Allergy and Clinical Immunology, 135(1), 110-122.
Eigl S, Prattes J, Reinwald M, Thornton CR, Reischies F, Spiess B, Neumeister P, Zollner-Schwetz I, Raggam RB, Flick H, et al (2015). Influence of Mould-active Antifungal Treatment on the Performance of the Aspergillus specific Bronchoalveolar Lavage Lateral-Flow Device Test. International Journal of Antimicrobial Agents, 46, 401-405.
Eigl S, Prattes J, Lackner M, Willinger B, Spiess B, Selitsch B, Meilinger M, Reischies F, Neumeister P, Wolfler A, et al (2015). Multicenter evaluation of a Lateral-Flow Device Test for Diagnosing Invasive Pulmonary Aspergillosis in ICU Patients. Critical Care, 19, 1-9.
Miceli M, Goggins MI, Chander P, Sekaran AK, Kizy AE, Samuel L, Jiang H, Thornton CR, Ramesh M, Alangaden G, et al (2015). Performance of Lateral Flow Device and Galactomannan for the Detection of Aspergillus Species in Bronchoalveolar Fluid of Patients at Risk for Invasive Pulmonary Aspergillosis. Mycoses: diagnosis, therapy and prophylaxis of fungal diseases, 58, 368-374.
Alcantara C, Thornton CR, Perez de-Luque A, Le Cocq K, Pedraza V, Murray PJ (2015). The Free-Living Rhizosphere Fungus Trichoderma hamatum GD12 Enhances Clover Productivity in Clover-Ryegrass Mixtures.
Plant and Soil: international journal on plant-soil relationships,
398, 165-180.
Abstract:
The Free-Living Rhizosphere Fungus Trichoderma hamatum GD12 Enhances Clover Productivity in Clover-Ryegrass Mixtures
Aim: a principal goal of grassland management is to
minimize the use of artificial fertilizers by maximising
the productivity of nitrogen-fixing leguminous plants
such as clovers. The objective of this study was to
investigate whether a plant-growth-promoting strain of
the free-living rhizosphere fungus Trichoderma
hamatum (GD12) could be used as a natural and sustainable
means of enhancing the competitiveness of
white clover (Trifolium repens) while allowing increased
productivity of both clover and ryegrass
(Lolium perenne) in mixed species systems.
Methods: an assay was conducted in rhizotrons with
white clover and ryegrass sown alone and in mixture
and in soils inoculated and non-inoculated with GD12.
Plant height, growing rate, phenological stage, number
of Rhizobium nodules and biomass were assessed. A
histological study of Rhizobium nodules and a stable
isotopes analysis was conducted to determine the N
fixation capacity of white clover.
Results: When introduced as a soil inoculant, the fungus
increased biomass production of both plant species and
shortened their phenological cycles. Furthermore, in
clover, GD12 enhanced plant height and growth rate
and stimulated Rhizobium nodulation, while 15N stable
isotope analysis demonstrated increased N2 fixation.
Conclusion: This shows that soil amendment with a
beneficial strain of saprotrophic fungus bestows a competitive
advantage to white clover in clover-ryegrass
mixtures and provides a sustainable mechanism for
improving the mixture productivity.
Abstract.
Solouk D, Bergmueller E, Weski J, Rolle A-M, Wiehr S, Thornton C, Gunzer M, Hasenberg M, Solouk D (2015). Towards the establishment of intravital mouse lung imaging via two Photon Microscopy for the design of a novel antibody-based aspergillosis detection system and the observation of pathogen-immune cell interactions.
MYCOSES,
58, 56-56.
Author URL.
Sharpe RA, Thornton CR, Tyrell J, Nikolaou V, Osborne NJ (2015). Variable risk of atopic disease due to indoor fungal exposure in NHANES 2005-2006. Clinical and Experimental Allergy, 45(10), 1566-1578.
Bergmueller E, Solouk D, Weski J, Thornton C, Gunzer M, Hasenberg M (2015). Where do spores go? Characterization of fungal distribution patterns among different murine Aspergillus fumigatus infection models using light sheet microscopy.
MYCOSES,
58, 9-9.
Author URL.
Hoenigl M, Prattes J, Eigl S, Lass-Flörl C, Willinger B, Reischies F, Posch V, Lackner M, Flick H, Hönigl K, et al (2014). 1462Bronchoalveolar Lavage Lateral-Flow Device Test for Diagnosing Invasive Pulmonary Aspergillosis in ICU patients: a multicenter study. Open Forum Infectious Diseases, 1(Suppl 1), s385-s386.
Thornton CR (2014). Breaking the Mould - Novel Diagnostic and Therapeutic Strategies for Invasive Pulmonary Aspergillosis in the Immune Deficient Patient. Expert Review of Clinical Immunology, 6(10), 771-780.
Hoenigl M, Thornton CR, Duettmann W, Posch V, Seeber K, Krause R, Lackner M, Lass-Florl C (2014). Bronchoalveolar Lavage Lateral-Flow Device Test for Diagnosing Invasive Pulmonary Aspergillosis in Pulmonology and ICU Patients. American Journal of Respiratory and Critical Care Medicine, 189
Willinger B, Lackner M, Lass-Florl C, Prattes J, Posch V, Selitsch B, Eschertzhuber S, Hoenigl K, Koidl C, Sereinigg M, et al (2014). Bronchoalveolar lavage lateral-flow device test for invasive pulmonary aspergillosis in solid organ transplant patients: a semi-prospective multicenter study. Transplantation, 98(8), 898-902.
Ghamrawi S, Renier G, Saulnier P, Cuenot S, Zykwinska A, Dutilh BE, Thornton CR, Faure S, Bouchara J-P (2014). Cell wall modifications during conidial maturation of the human pathogenic fungus Pseudallescheria boydii. PLoS One, 9(6).
Prattes J, Prueller F, Koidl C, Raggam R, Zollner-Schwetz I, Eigl S, Posch V, Hoenigl K, Duettmann W, Flick H, et al (2014). Diagnostics for invasive Pulmonary Aspergillosis in Bronchoalveolar Lavage Fluid of Patients with Underlying Pulmonary Diseases: Comparison of Aspergillus Lateral Flow Device, Galactomannan-Antigen-Test, Beta-D-Glucan-Tests and Conventional Culture.
MYCOSES,
57, 15-16.
Author URL.
Davies GE, Thornton CR (2014). Differentiation of the emerging human pathogens Trichosporon asahii and Trichosporon asteroides from other pathogenic yeasts and moulds by using species-specific monoclonal antibodies. PLoS One, 9(1).
Vandeputte P, Ghamrawi S, Rechenmann M, Iltis A, Giraud S, Fleury M, Thornton CR, Delhaes L, Meyer W, Papon N, et al (2014). Draft genome sequence of the pathogenic fungus Scedosporium apiospermum. Genome Announcements, 2(5).
Sharpe R, Thornton CR, Osborne NJ (2014). Modifiable Factors Governing Indoor Fungal Diversity and Risk of Asthma. Clinical and Experimental Allergy, 44, 631-641.
Ghamrawi S, Gilles R, Saulnier P, Cuenot S, Zykwinska A, Bas ED, Thornton C, Faure S, Bouchara J-P (2014). Modifications de la paroi au cours de la maturation des conidies chez Pseudallescheria boydii. Journal de Mycologie Médicale, 24(3).
Prattes J, Prueller F, Koidl C, Raggam RB, Palfner M, Eigl S, Flick H, Buzina W, Zollner-Schwetz I, Thornton CR, et al (2014). Novel Tests for Diagnosis of Invasive Pulmonary Aspergillosis in Patients with Underlying Respiratory Diseases. American Journal of Respiratory and Critical Care Medicine, 190(8), 922-929.
Hoenigl M, Prattes J, Spiess B, Wagner J, Prueller F, Raggam RB, Posch V, Duettmann W, Hoenigl K, Wofler A, et al (2014). Performance of Galactomannan, Beta-D-Glucan,
Aspergillus Lateral-Flow Device, Conventional Culture and PCR tests for Diagnosis of Invasive Pulmonary Aspergillosis in Bronchoalveolar Lavage Fluid. Journal of Clinical Microbiology, 52, 2039-2045.
Kobayashi M, Thornton CR (2014). Serological-diagnosis of chronic pulmonary aspergillosis (CPA) and Aspergillus lateral-flow device (LFD). European Respiratory Journal, 44, 2501-2501.
Al-Maqtoofi MY, Thornton CR (2014). Tracking the human pathogenic fungus Fusarium by using a highly specific monoclonal antibody.
IMMUNOLOGY,
143, 169-169.
Author URL.
White PL, Parr C, Thornton CR, Barnes RA (2013). An Evaluation of real-time PCR, Galactomannan ELISA and a novel Lateral-Flow Device for the diagnosis of invasive aspergillosis. Journal of Clinical Microbiology, 51(5), 1510-1516.
Held J, Schmidt T, Thornton CR, Kotter E, Bertz H (2013). Comparison of a Novel Aspergillus Lateral-Flow Device and the Platelia Galactomannan Assay for Diagnosis of Invasive Aspergillosis following Haematopoietic Stem Cell Transplantation. Infection, 41(6), 1163-1169.
Held J, Schmidt T, Thornton CR, Kotter E, Bertz H (2013). Comparison of a novel Aspergillus Antigen Lateral Flow Device and the Platelia (R) Aspergillus Antigen ELISA in patients after hematopoietic stem cell transplantation.
MYCOSES,
56, 18-18.
Author URL.
Wiederhold NP, Najvar LK, Bocanegra RA, Kirkpatrick WR, Patterson TF, Thornton CR (2013). Inter-Laboratory and Inter-Study Reproducibility of a Novel Lateral-Flow Device and the Influence of Antifungal Therapy on the Detection of Invasive Pulmonary Aspergillosis.
Journal of Clinical Microbiology,
51, 459-465.
Abstract:
Inter-Laboratory and Inter-Study Reproducibility of a Novel Lateral-Flow Device and the Influence of Antifungal Therapy on the Detection of Invasive Pulmonary Aspergillosis
Lateral-flow devices (LFD) have gained interest as potential point-of-care assays for the diagnosis of infectious diseases. Our objective was to evaluate the inter-laboratory and inter-study reproducibility and the effects of antifungal therapy on a LFD developed for invasive pulmonary aspergillosis (IPA) detection. An established neutropenic guinea pig model of IPA caused by Aspergillus fumigatus was used. At predetermined time points (1 hr, 3, 5, and 7 days post-inoculation) blood and BAL fluid were collected from infected and uninfected animals. In a separate experiment, guinea pigs were treated with posaconazole (10 mg/kg PO BID), voriconazole (10 mg/kg PO BID), liposomal amphotericin B (10 mg/kg IP QD), or caspofungin (2 mg/kg IP QD), and samples were collected on days 7 and 11. Each laboratory independently evaluated the IgG monoclonal antibody-based LFD. Galactomannan and (1→3)-β-D-glucan were also measured using commercially available kits. Good inter-laboratory agreement was observed with the LFD as 96.7 % (29/30) of the serum and 73.3% (22/30) of the BAL samples from infected animals were in agreement. Good inter-study agreement was also observed. The serum sensitivity of each surrogate marker assay was reduced in animals treated with antifungals. In contrast, these markers remained elevated within the BAL fluids of treated animals, which was consistent with the fungal burden and histopathology results. These results demonstrate that the LFD assay is reproducible between different laboratories and studies. However, the sensitivity of this assay and other markers of IPA may be reduced within the serum in the presence of antifungal therapy
Abstract.
Studholme D, Harris B, Le Cocq K, Winsbury R, Perera V, Ryder L, Beale M, Ward J, Thornton CR, Grant M, et al (2013). Investigating the Beneficial Traits of Trichoderma hamatum GD12 for Sustainable Agriculture - Insights from Genomics.
Frontiers in Plant-Microbe Interactions,
4, 1-13.
Abstract:
Investigating the Beneficial Traits of Trichoderma hamatum GD12 for Sustainable Agriculture - Insights from Genomics
Trichoderma hamatum strain GD12 is unique in that it can promote plant growth, activate biocontrol against pre- and post-emergence soil pathogens and can induce systemic resistance to foliar pathogens. This study extends previous work in lettuce to demonstrate that GD12 can confer beneficial agronomic traits to other plants, providing examples of plant growth promotion in the model dicot, Arabidopsis thaliana and induced foliar resistance to Magnaporthe oryzae in the model monocot rice. We further characterize the lettuce-T. hamatum interaction to show that bran extracts from GD12 and a N-acetyl-β-D-glucosamindase-deficient mutant differentially promote growth in a concentration dependent manner, and these differences correlate with differences in the small molecule secretome. We show that GD12 mycoparasitises a range of isolates of the pre-emergent soil pathogen Sclerotinia sclerotiorum and that this interaction induces a further increase in plant growth promotion above that conferred by GD12. To understand the genetic potential encoded by T. hamatum GD12 and to facilitate its use as a model beneficial to study plant growth promotion, induced systemic resistance and mycoparasitism we present de novo genome sequence data. We compare GD12 with other published Trichoderma genomes and show that T. hamatum GD12 contains unique genomic regions with the potential to encode novel bioactive metabolites that may contribute to GD12’s agrochemically important traits.
Abstract.
Thornton CR (2013). Lateral-Flow Device for Diagnosis of Fungal Infection. Current Fungal Infection Reports, 7(3), 244-251.
Ryder, LS, Mentlak TA, Dagdas YF, Kershaw MJ, Thornton CR, Chen J, Wang Z, Talbot, NJ (2013). NADPH oxidases regulate septin-mediated cytoskeletal re-modeling during plant infection by the rice blast fungus. Proceedings of the National Academy of Sciences of USA, 110, 3179-3184.
Johnson GL, Shannon M, Thornton CR, Agrawal SG, Bustin SA (2013). Proximity ligation assay for the sensitive, specific and early diagnosis of invasive fungal disease.
MYCOSES,
56, 48-49.
Author URL.
Wiederhold NP, Thornton CR (2013). Reply to “Interlaboratory and Interstudy Reproducibility of a Novel Lateral-Flow Device: a Statistical Issue”. Journal of Clinical Microbiology, 51(5).
Hoenigl M, Koidl C, Duettmann W, Seeber K, Wagner J, Wölfler A, Raggam RB, Thornton CR, Krause R (2012). Bronchoalveolar lavage lateral-flow device test for invasive pulmonary aspergillosis diagnosis in haematological malignancy and solid organ transplant patients.
Journal of Infection,
65(6), 588-591.
Abstract:
Bronchoalveolar lavage lateral-flow device test for invasive pulmonary aspergillosis diagnosis in haematological malignancy and solid organ transplant patients
Due to the crude mortality of 80-90% in absence of adequate treatment, timely diagnosis and early start of antifungal therapy are key factors in the successful treatment of invasive pulmonary aspergillosis (IPA). Diagnosis of IPA, however, remains difficult as clinical signs and symptoms as well as radiological findings are often unspecific and conventional culture methods lack sensitivity. In recent years antigen testing has therefore become one of the cornerstones of IFI diagnostics, even though tests are limited by varying turnaround time as well as sensitivity and specificity issues. These limitations may be overcome by the Lateral-Flow Device (LFD) test, a single sample point-of-care test that is based on the detection of an Aspergillus extracellular glycoprotein antigen by monoclonal antibody JF5. This retrospective study evaluates the LFD test by using bronchoalveolar lavage (BAL) samples from patients with haematological malignancies and patients after solid organ transplantation (SOT).
Thirty-nine BAL samples from 37 patients were included (29 samples haematological malignancies and 10 samples SOT; 12 probable IPA, 9 possible IPA, 16 no IPA). Sensitivity and specificity of BAL LFD test for probable IPA were 100% and 81% respectively. GM levels in cases with negative LFD were significantly lower than in patients with positive LFD (P
Abstract.
Thornton CR, Johnson G, Agrawal S (2012). Detection of invasive pulmonary aspergillosis in haematological malignancy patients by using lateral-flow technology. Journal of Visualized Experiments, 61
Romão-Dumaresq AS, de Araújo WL, Talbot NJ, Thornton CR (2012). RNA interference of endochitinases in the sugarcane endophyte Trichoderma virens 223 reduces its fitness as a biocontrol agent of pineapple disease.
PLoS One,
7(10).
Abstract:
RNA interference of endochitinases in the sugarcane endophyte Trichoderma virens 223 reduces its fitness as a biocontrol agent of pineapple disease
The sugarcane root endophyte Trichoderma virens 223 holds enormous potential as a sustainable alternative to chemical pesticides in the control of sugarcane diseases. Its efficacy as a biocontrol agent is thought to be associated with its production of chitinase enzymes, including N-acetyl-β-D-glucosaminidases, chitobiosidases and endochitinases. We used targeted gene deletion and RNA-dependent gene silencing strategies to disrupt N-acetyl-β-D-glucosaminidase and endochitinase activities of the fungus, and to determine their roles in the biocontrol of soil-borne plant pathogens. The loss of N-acetyl-β-D-glucosaminidase activities was dispensable for biocontrol of the plurivorous damping-off pathogens Rhizoctonia solani and Sclerotinia sclerotiorum, and of the sugarcane pathogen Ceratocystis paradoxa, the causal agent of pineapple disease. Similarly, suppression of endochitinase activities had no effect on R. solani and S. sclerotiorum disease control, but had a pronounced effect on the ability of T. virens 223 to control pineapple disease. Our work demonstrates a critical requirement for T. virens 223 endochitinase activity in the biocontrol of C. paradoxa sugarcane disease, but not for general antagonism of other soil pathogens. This may reflect its lifestyle as a sugarcane root endophyte.
Abstract.
Ryder LS, Harris BD, Soanes DM, Kershaw MJ, Talbot NJ, Thornton CR (2012). Saprotrophic competitiveness and biocontrol fitness of a genetically modified strain of the plant-growth-promoting fungus Trichoderma hamatum GD12. Microbiology, 158(Special issue of Trichoderma: from basic biology to biotechnology), 84-97.
Ryder LS, Harris BD, Soanes DM, Kershaw MJ, Talbot NJ, Thornton CR (2012). Saprotrophic competitiveness and biocontrol fitness of a genetically modified strain of the plant-growth-promoting fungus Trichoderma hamatum GD12.
Microbiology (Reading),
158(Pt 1), 84-97.
Abstract:
Saprotrophic competitiveness and biocontrol fitness of a genetically modified strain of the plant-growth-promoting fungus Trichoderma hamatum GD12.
Trichoderma species are ubiquitous soil fungi that hold enormous potential for the development of credible alternatives to agrochemicals and synthetic fertilizers in sustainable crop production. In this paper, we show that substantial improvements in plant productivity can be met by genetic modification of a plant-growth-promoting and biocontrol strain of Trichoderma hamatum, but that these improvements are obtained in the absence of disease pressure only. Using a quantitative monoclonal antibody-based ELISA, we show that an N-acetyl-β-d-glucosaminidase-deficient mutant of T. hamatum, generated by insertional mutagenesis of the corresponding gene, has impaired saprotrophic competitiveness during antagonistic interactions with Rhizoctonia solani in soil. Furthermore, its fitness as a biocontrol agent of the pre-emergence damping-off pathogen Sclerotinia sclerotiorum is significantly reduced, and its ability to promote plant growth is constrained by the presence of both pathogens. This work shows that while gains in T. hamatum-mediated plant-growth-promotion can be met through genetic manipulation of a single beneficial trait, such a modification has negative impacts on other aspects of its biology and ecology that contribute to its success as a saprotrophic competitor and antagonist of soil-borne pathogens. The work has important implications for fungal morphogenesis, demonstrating a clear link between hyphal architecture and secretory potential. Furthermore, it highlights the need for a holistic approach to the development of genetically modified Trichoderma strains for use as crop stimulants and biocontrol agents in plant agriculture.
Abstract.
Author URL.
Johnson GL, Bibby D, Bustin B, Thornton CR, Agrawal S (2011). Detection of Aspergillus in broncho-alveolar lavage fluid using two biological assays; evidence of active infection?. Mycoses, 54, 130-131.
Thornton CR (2010). Detection of Invasive Aspergillosis. Advances in Applied Microbiology, 70, 187-216.
Thornton CR, Slaughter DC, Davis RM (2010). Detection of the sour-rot pathogen Geotrichum candidum in tomato fruit and juice by using a highly specific monoclonal antibody-based ELISA. International Journal of Food Microbiology, 143(3), 166-172.
Thornton CR (2010). Environmental fungi as a cause of human mycoses. Culture, 31(2), 1-5.
Wiederhold NP, Thornton CR, Najvar LK, Kirkpatrick WR, Bocanegra R, Patterson TF (2009). Comparison of Lateral Flow Technology and Galactomannan and (1→3)-β-D-Glucan Assays for Detection of Invasive Pulmonary Aspergillosis. Clinical and Vaccine Immunology, 16(12), 1844-1846.
Richards TA, Soanes DM, Foster PG, Leonard G, Thornton CR, Talbot NJ (2009). Phylogenomic analysis demonstrates a pattern of rare and ancient horizontal gene transfer between plants and fungi. The Plant Cell, 21, 1897-1911.
Thornton CR (2009). Tracking the emerging human pathogen Pseudallescheria boydii by using highly specific monoclonal antibodies.
Clinical and Vaccine Immunology,
16(5), 756-764.
Abstract:
Tracking the emerging human pathogen Pseudallescheria boydii by using highly specific monoclonal antibodies
Pseudallescheria boydii has long been known to cause white grain mycetoma in immunocompetent humans, but it has recently emerged as an opportunistic pathogen of humans, causing potentially fatal invasive infections in immunocompromised individuals and evacuees of natural disasters, such as tsunamis and hurricanes. The diagnosis of P. boydii is problematic since it exhibits morphological characteristics similar to those of other hyaline fungi that cause infectious diseases, such as Aspergillus fumigatus and Scedosporium prolificans. This paper describes the development of immunoglobulin M (IgM) and IgG1 kappa-light chain monoclonal antibodies (MAbs) specific to P. boydii and certain closely related fungi. The MAbs bind to an immunodominant carbohydrate epitope on an extracellular 120-kDa antigen present in the spore and hyphal cell walls of P. boydii and Scedosporium apiospermum. The MAbs do not react with S. prolificans, Scedosporium dehoogii, or a large number of clinically relevant fungi, including A. fumigatus, Candida albicans, Cryptococcus neoformans, Fusarium solani, and Rhizopus oryzae. The MAbs were used in immunofluorescence and double-antibody sandwich enzyme-linked immunosorbent assays (DAS-ELISAs) to accurately differentiate P. boydii from other infectious fungi and to track the pathogen in environmental samples. Specificity of the DAS-ELISA was confirmed by sequencing of the internally transcribed spacer 1 (ITS1)-5.8S-ITS2 rRNA-encoding regions of environmental isolates.
Abstract.
Thornton CR (2009). Use of monoclonal antibody-based diagnostic assays for the detection of invasive mycoses. Microbiologist, 10(2), 49-50.
Thornton CR (2008). Development of an immunochromatographic lateral-flow device for rapid serodiagnosis of invasive aspergillosis.
Clinical and Vaccine Immunology,
15(7), 1095-1105.
Abstract:
Development of an immunochromatographic lateral-flow device for rapid serodiagnosis of invasive aspergillosis
Aspergillus fumigatus is a cosmopolitan saprotrophic fungus that is second only to Candida species as a cause of invasive fungal infections in immunocompromised humans. Current immunodiagnostic tests for invasive aspergillosis (IA) are based on the detection of circulating galactomannan ( GM) in a patient's serum by using a rat monoclonal antibody (MAb), EB-A2, that binds to tetra (1 -> 5)-beta-D-galactofuranoside, the immunodominant epitope in GM. The potential cross-reactivity of MAb EB-A2 with non-Aspergillus fungi, with contaminating GM in beta-lactam antibiotics and foodstuffs, and with bacterial lipoteichoic acids has prompted efforts to discover non-GM antigens that can act as surrogate markers for the diagnosis of IA. This paper describes the development of a mouse MAb, JF5, that binds to a protein epitope present on an extracellular glycoprotein antigen secreted constitutively during the active growth of A. fumigatus. The MAb was used to develop an immunochromatographic lateral-flow device (LFD) for the rapid (15-min) detection of Aspergillus antigens in human serum. The test is highly specific, reacting with antigens from Aspergillus species but not with antigens from a large number of clinically important fungi, including Candida species, Cryptococcus neoformans, Fusarium solani, Penicillium marneffei, Pseudallescheria boydii, and Rhizopus oryzae. The LFD was able to detect circulating antigen in serum samples from patients suspected of having or shown to have IA on the basis of their clinical symptoms and results from tests for GM and fungal (1 -> 3)-beta-D-glucan. The ease of use of the LFD provides a diagnostic platform for the routine testing of vulnerable patients who have an elevated risk of IA.
Abstract.
Thornton, C.R. (2008). Tracking fungi in soil with monoclonal antibodies. European Journal of Plant Pathology, 121, 347-353.
Gilbert MJ, Thornton CR, Wakley GE, Talbot NJ (2006). A P-type ATPase required for rice blast disease and induction of host resistance. Nature, 440(7083), 535-539.
Richards TA, Dacks JB, Jenkinson JM, Thornton CR, Talbot NJ (2006). Evolution of filamentous plant pathogens: gene exchange across eukaryotic kingdoms. Current Biology, 16(18), 1857-1864.
Thornton CR, Talbot, N.J. (2006). Immunofluorescence microscopy and immunogold EM for investigating fungal infection of plants. Nature Protocols, 1, 2506-2511.
Kershaw, M.J. Wakley, G.E. Talbot, N.J. Thornton CR (2005). Four conserved intra-molecular disulfide linkages are required for secretion and cell wall localization of a hydrophobin during fungal morphogenesis. Molecular Microbiology, 56(1), 117-125.
Thornton CR (2005). Use of monoclonal antibodies to quantify the dynamics of α-galactosidase and endo-1,4-β-glucanase production by Trichoderma hamatum during saprotrophic growth and sporulation in peat. Environmental Microbiology, 7(5), 737-749.
Thornton, CR, Groenhof AC, Forrest, R & Lamotte, R (2004). A One-Step, Immunochromatographic Lateral Flow Device Specific to Rhizoctonia solani and Certain Related Species, and its use to Detect and Quantify R. solani in Soil. Phytopathology, 94, 280-288.
Tucker SL, Thornton CR, Tasker K, Jacob C, Giles G, Egan M, Talbot NJ (2004). A fungal metallothionein is required for pathogenicity of Magnaporthe grisea. The Plant Cell, 16(6), 1575-1588.
Thornton CR (2004). An immunological approach to quantifying the saprotrophic growth dynamics of Trichoderma species during antagonistic interactions with Rhizoctonia solani in a soil-less mix. Environmental Microbiology, 6(4), 323-334.
Wang ZY, Thornton CR, Kershaw MJ, Debao L, Talbot NJ (2003). The glyoxylate cycle is required for temporal regulation of virulence by the plant pathogenic fungus Magnaporthe grisea. Molecular Microbiology, 47(6), 1601-1612.
Thornton CR, Pitt, D. Wakley, G.E. Talbot, N.J. (2002). Production of a monoclonal antibody specific to the genus Trichoderma and closely related fungi, and its use to detect Trichoderma spp. in naturally infested composts. Microbiology, 148, 1263-1279.
Bailey DJ, Thornton CR, Dewey FM, Gilligan CA (2001). A non-destructive immunoblotting technique for visualisation and analysis of the growth dynamics of Rhizoctonia solani.
Mycological Research,
105(8), 983-990.
Abstract:
A non-destructive immunoblotting technique for visualisation and analysis of the growth dynamics of Rhizoctonia solani
Immunoblotting combined with computer imaging and a simple, non-linear mathematical model were used to demonstrate the potential of a technique for non-destructive visualisation and analysis of fungal growth of Rhizoctonia solani over the surface of non-sterile sand. Immunoblotting detected actively growing regions of mycelium enabling visualisation of individual hyphae at the colony edge. A zone of active growth was detected expanding radially over time. Active growth did not continue in the centre of the fungal colony leading to the development of a ring of mycelium surrounding the inoculum. Change in the density of actively growing mycelium with distance from the inoculum unit was summarised for each colony at each time by a Gaussian function, describing a wave of actively growing mycelium, symmetrical in density about its centre but differing amongst replicate colonies. The effectiveness of the immunoblotting technique to detect differences in colony growth was tested by comparing the growth of replicate colonies for two contrasting isolates of R. solani. When both isolates of R. solani were grown at 23 °C the amplitude of the wave increased to a maximum and then decayed over time, the location of the centre of the wave moved outwards at a constant rate, whilst the width of the wave increased. Increasing the temperature to 28 °, accelerated this intrinsic growth process for one isolate, but retarded growth of the other.
Abstract.
Thornton CR, O'Neill TM, Hilton G, Gilligan CA (1999). Detection and recovery of Rhizoctonia solani in naturally infested glasshouse soils using a combined baiting, double monoclonal antibody ELISA.
Plant Pathology,
48(5), 627-634.
Author URL.
Thornton CR, Gilligan CA (1999). Quantification of the effect of the hyperparasite Trichoderma harzianum on the saprotrophic growth dynamics of Rhizoctonia solani in compost using a monoclonal antibody-based ELISA.
Mycological Research,
103, 443-448.
Author URL.
Thornton CR, Dewey FM, Gilligan CA (1997). Production and characterization of a monoclonal antibody raised against surface antigens from mycelium of Gaeumannomyces graminis var. tritici evidence for an extracellular polyphenol oxidase.
Phytopathology,
87(1), 123-131.
Abstract:
Production and characterization of a monoclonal antibody raised against surface antigens from mycelium of Gaeumannomyces graminis var. tritici evidence for an extracellular polyphenol oxidase
A murine monoclonal antibody (MAb) of immunoglobulin class M (IgM) was raised against surface antigens from Gaeumannomyces graminis var. tritici and, by enzyme-linked immunosorbent assay, recognized isolates of G. graminis var. tritici, G. graminis var. avenae and G. graminis var. graminis. Characterization of the antigen by heat and protease treatments showed that the epitope recognized by the MAb was a protein. Antigen production was detected only in live mycelia. Immunofluorescence studies showed that the antigen was associated with both the broad melanized macrohyphae and hyaline mycelia of G. graminis var. tritici. Secretion of antigen into an aqueous minimal medium was promoted only by exposure of live mycelia to certain pbenolic substrates, including monophenols ortho-, para-, and meta-cresol; 3,4,5-trihydroxybenzoic acid (gallic acid); and phenolic amino acid L-3- (3,4-dihydroxyphenyl) alanine (L-DOPA). Antigen secretion was not promoted by 3-(4-hydroxyphenyl) alanine (L-tyrosine). The MAb reacted strongly with purified enzyme laccase (polyphenol oxidase, EC 1.10.3.2) but did not recognize purified tyrosinase (monophenol oxidase, EC 1.14.18.1). Moreover, chemicals that bind to copper and inhibit copper-containing enzymes such as laccase completely inhibited antigen secretion in response to L-DOPA. The MAb was tested for specificity against a wide range of fungi, common yeast species, and grain positive and negative bacteria. It did not recognize antigens from a broad range of unrelated fungi, including Gliocladium roseum, Fusarium sp. Phoma exigua, Phialophora fastigiata, Penicillium crustosum, Pythium ultimum, Rhizopus stolonifer, Rhizoctonia carotae, R. oryzae, R. tuliparum, and Trichoderma viride, nor did it recognize surface antigens from yeasts or bacteria. The MAb cross-reacted with antigens from Botrytis spp. Chaetomium globosum, R. cerealis, and R. solani. However, secretion of antigen by R. solani and R. cerealis was not promoted by L-DOPA, and secretion by C. globosum in response to the phenolic amino acid was significantly less compared to G. graminis var. tritici.
Abstract.
Otten W, Gilligan CA, Thornton CR (1997). Quantification of fungal antigens in soil with a monoclonal antibody-based ELISA: Analysis and reduction of soil-specific bias.
Phytopathology,
87(7), 730-736.
Abstract:
Quantification of fungal antigens in soil with a monoclonal antibody-based ELISA: Analysis and reduction of soil-specific bias
This paper describes methods to improve the use of immunoassays for quantification of soilborne fungal antigens. Calibration curves, prepared by diluting known quantities of an antigen into soil extracts and into soil, were described by a four-parameter logistic curve from which two principal criteria, the lower detection limit and the horizontal locational parameter, were used to summarize the sensitivity and bias of an immunoassay. We identify two sources of bias, retention of the antigen in soil due to bonding and interference of soluble soil components in plate-trapped-antigen, enzyme-linked immunosorbent assays. Using a monoclonalantibody that recognizes a putative catechol oxidase secreted by hyphae of Rhizoctonia solani, we show that bias due to retention of the antigen in soil is substantially greater than bias due to interference. Three soils were compared: a sand, a clay, and a loam. The degree of retention varied with soil type, with more than a 1,000-fold reduction in sensitivity in the clay soil. Addition of CuSO4 to the extraction solution and optimizing the volume of extractant reduced the bias and increased the sensitivity of the assay for all three soils. Possible mechanisms for the effect due to Cu2+ and the implications for the design and use of calibration curves for assays involving quantification of fungal antigens in soil are discussed.
Abstract.
Dewey FM, Thornton CR, Gilligan CA (1997). Use of Monoclonal Antibodies to Detect, Quantify and Visualize Fungi in Soils. Advances in Botanical Research, 24(C), 275-308.
Thornton CR (1996). Detection and quantification of Rhizoctonia solani in soil by monoclonal antibody-based immuno-magnetic bead assay.
Soil Biology and Biochemistry,
28(4-5), 527-532.
Abstract:
Detection and quantification of Rhizoctonia solani in soil by monoclonal antibody-based immuno-magnetic bead assay
Murine monoclonal antibodies (MAbs) of the immunoglobulin classes IgM and IgA, from two different hybridoma cell lines, and rabbit (polyclonal) antiserum raised against mycelial antigens from an AG4 isolate of R. solani were used to develop magnetic microsphere-enzyme immunoassays (MM-EIAs) for the detection and quantification of the pathogen in soil. The assay can also be used to differentiate R. solani anastomosis group (AG) 2-2 isolates from other AG isolates grown in vitro. Mycelial antigens were solubilised in saline buffer containing detergent and following high speed centrifugation soluble antigen was delineated by the addition either of a mixture of specific murine MAbs of different immunoglobulin classes or a mixture of murine MAb supernatant and rabbit antiserum. Due to the low concentration of reactants a soluble immune complex was formed. The soluble complex was isolated by the addition of magnetic beads coated with antibodies that specifically recognised the murine IgM MAbs. The bound antibody antigen complex was then detected using a commercial antibody enzyme conjugate and chromogen.
Abstract.
Thornton CR, Dewey FM (1996). Detection of phialoconidia of Trichoderma harzianum in peat-bran by monoclonal antibody-based enzyme-linked immunosorbent assay.
Mycological Research,
100(2), 217-222.
Abstract:
Detection of phialoconidia of Trichoderma harzianum in peat-bran by monoclonal antibody-based enzyme-linked immunosorbent assay
Murine monoclonal antibodies (MAbs) were raised against phialoconidia of a Trichoderma harzianum isolate T95. A hybridoma cell line. GH5, was raised that produced MAbs of the immunoglobulin class IgM that recognized, by enzyme-linked immunosorbent assay (ELISA), antigens from conidia of T95. They did not react with antigens from mycelium or chlamydospores of this isolate but recognized antigens from conidia of a number of other T. harzianum and T. viride isolates. Heat, protease and periodate treatment of T95 conidial antigens showed that GH5 MAbs recognized a protein epitope that formed part of a larger glycoprotein molecule. GH5 MAbs were used to develop an ELISA for the detection of phialoconidia of T. harzianum in peat-bran.
Abstract.
Thornton CR, Dewey FM, Gilligan CA (1994). Development of a monoclonal antibody-based enzyme-linked immunosorbent assay for the detection of live propagules of Trichoderma harzianum in a peat-bran medium.
Soil Biology and Biochemistry,
26(7), 909-920.
Abstract:
Development of a monoclonal antibody-based enzyme-linked immunosorbent assay for the detection of live propagules of Trichoderma harzianum in a peat-bran medium
Mouse monoclonal antibodies (MAbs) and mouse and rabbit (polyclonal) antisera were raised against a Trichoderma harzianum isolate T95. A cell line, Th.HD33, raised from splenocytes from a mouse immunized with lyophilized mycelium and Quil a adjuvant, produced MAbs of the immunoglobulin sub-class IgG2a that were specific recognizing, by ELISA, antigens from a number of Trichoderma species and species from the related genus Hypocrea but not from a range of other soil-borne fungi. These MAbs were used to develop an enzyme-linked immunosorbent assay (ELISA) for the detection of live propagules of Trichoderma harzianum in a peat-bran medium. Heat, protease and periodate treatment of the T95 antigen and binding on Western blots indicate that the Th.HD33 MAbs recognized a protein epitope of ca Mr 18,500 which forms part of a larger glycoprotein molecule. © 1994.
Abstract.
THORNTON CR, DEWEY FM, GILLIGAN CA (1993). Development of monoclonal antibody‐based immunological assays for the detection of live propagules of Rhizoctonia solani in soil.
Plant Pathology,
42(5), 763-773.
Abstract:
Development of monoclonal antibody‐based immunological assays for the detection of live propagules of Rhizoctonia solani in soil
Mouse monoclonal antibodies (mAbs) and rabbit (polyclonal) antiserum were used to develop DIAGNOSTIC‐ELISA, double‐antibody‐sandwich‐ELISA (DAS‐ELISA), DIP‐STICK and immuno‐fluorescence colony staining immunoassays for the specific detection of Rhizoctonia solani in soil. mAbs were raised against an anastomosis group 4 isolate of R. solani. Mice were immunized using either phosphate‐buffered saline (PBS) suspensions of lyophilized mycelium plus Quil a adjuvant, or with a solubilized acetone precipitate prepared from cell‐free surface washings from solid slant cultures. Polyclonal antisera were raised in rabbits using PBS suspensions of lyophilized mycelium and Quil a adjuvant. Hybridoma supernatants and rabbit antisera were screened by ELISA. Four of the cell lines raised produced mAbs that were species‐specific. They recognized antigens from R. solani by ELISA and immunofluorescence, but not other related or unrelated species of soil‐borne fungi. The remaining cell line produced mAbs that cross‐reacted slightly, by ELISA, with antigens from R. cerealis. These mAbs did not recognize R. cerealis by immunofluorescence, or other related or unrelated soil‐borne fungi, by ELISA. Copyright © 1993, Wiley Blackwell. All rights reserved
Abstract.
Thornton CR, Jarvis B, Cooke R (1991). A chitin assay for the enumeration of Plasmodiophora brassicae resting spores in clubroot tissue.
Mycological Research,
95, 879-882.
Author URL.
Chapters
Thornton CR (2023). Serological Techniques for Diagnosis. In Lane CR, Beales PA, Hughes KJD (Eds.)
Fungal Plant Pathogens Applied Techniques, Wallingford, Oxfordshire, UK: Cabi, 169-182.
Abstract:
Serological Techniques for Diagnosis
Abstract.
Thornton CR (2012). Serological techniques for diagnosis. In Lane C, Beales P, Hughes K (Eds.) Fungal Plant Pathogens, Principles and Protocols, CABI, 159-177.
Thornton CR (2009). Production of monoclonal antibodies to plant pathogens. In (Ed)
, 63-74.
Abstract:
Production of monoclonal antibodies to plant pathogens.
Abstract.
Author URL.
Thornton CR (2007). Tracking fungi in soil with monoclonal antibodies. In (Ed) Sustainable disease management in a European context, 347-353.
Thornton, C.R. (2001). Immunological methods for fungi. In Talbot NJ (Ed) Molecular and Cell Biology of Filamentous Fungi: a Practical Approach, UK: Oxford University Press, 227-257.
Conferences
Thornton C (2019). Bench-to-Bedside Diagnosis of Invasive Pulmonary Aspergillosis. 9th Trends in Medical Mycology. 11th - 14th Oct 2019.
Abstract:
Bench-to-Bedside Diagnosis of Invasive Pulmonary Aspergillosis
Abstract.
Maurer A, Beziere N, Seyfried D, Wehrmueller J, Spycher P, Schwenck J, Wild A-M, Davies G, Boschetti F, Wiehr S, et al (2019). Development and preclinical evaluation of an immunoPET tracer for clinical imaging of Aspergillosis.
Author URL.
Schwenck J, Beziere N, Maurer A, Wild AM, Henneberg H, Davies G, Reischl G, Spycher PR, Wehrmüller JE, Boschetti F, et al (2019). Translational immunoPET/MR imaging of invasive pulmonary aspergillosis. NuklearMedizin 2019. 3rd - 6th Apr 2019.
Abstract:
Translational immunoPET/MR imaging of invasive pulmonary aspergillosis
Abstract.
Henneberg S, Thornton C, Wiehr S, Rolle AM, Pichler B, Hasenberg M, Weski J, Hasenberg A, Gunzer M (2017). Intravital lung imaging to study the interactions between immune cells and Aspergillus fumigatus.
Author URL.
Thornton CR (2017). Monoclonal antibody JF5 for aspergillosis: from BALF to immunoPET/MRI. Trends in Medical Mycology. 6th - 9th Oct 2017.
Abstract:
Monoclonal antibody JF5 for aspergillosis: from BALF to immunoPET/MRI
Abstract.
Author URL.
Sharpe RA, Thornton CR, Nikolaou V, Osborne NJ (2015). Household energy efficiency, fungi and allergic asthma.
Author URL.
Shah A, Abdolrasouli A, Soresi S, Herbst S, Reed A, Carby M, Thornton CR, Drumright L, Shaunak S, Armstrong-James D, et al (2015). The Utility of Novel Multi-Stage Testing for the Diagnosis of Pulmonary Aspergillosis in a Cohort of Lung Transplant Recipients.
Author URL.
Hoenigl M, Thornton C, Duettmann W, Posch V, Seeber K, Krause R, Lackner M, Lass-Florl C (2014). Bronchoalveolar Lavage Lateral-Flow Device Test for Diagnosing Invasive Pulmonary Aspergillosis in Pulmonology and Icu Patients.
Author URL.
Sereinigg M, Lass-Floerl C, Willinger B, Prattes J, Posch V, Selitsch B, Lackner M, Eschertzhuber S, Drescher M, Koidl C, et al (2014). Bronchoalveolar Lavage Lateral-Flow Device Test for Invasive Pulmonary Aspergillosis in Solid Organ Transplant Patients: a Semi-Prospective Multicenter Study.
Author URL.
Sereinigg M, Lass-Floerl C, Willinger B, Prattes J, Posch V, Selitsch B, Lackner M, Eschertzhuber S, Drescher M, Koidl C, et al (2014). Bronchoalveolar Lavage Lateral-Flow Device Test for Invasive Pulmonary Aspergillosis in Solid Organ Transplant Patients: a Semi-Prospective Multicenter Study.
Author URL.
Hoenigl M, Prattes J, Eigl S, Lass-Florl C, Willinger B, Reiches F, Posch V, Lackner M, Flick H, Hoenigl K, et al (2014). Bronchoalveolar lavage lateral-flow device test for diagnosing invasive pulmonary aspergillosis in ICU patients: a multicenter study. IDWeek2014. 8th - 12th Oct 2014.
Miceli M, Goggins MI, Chander P, Sekaran AK, Kizy AE, Samuel L, Jiang H, Thornton CR, Ramesh M, Alangeden G, et al (2014). Diagnostic performance of lateral flow device and galactomannan for the detection of Aspergillus in bronchoalveolar lavage of patients at high risk for. invasive aspergillosis. 18th Symposium on Infections in the Immunocompromised Host. 15th - 17th Jun 2014.
McDonagh L, Thornton C, Wallman JF, Stevens JR (2009). Development of an antigen-based rapid diagnostic test for the identification of blowfly (Calliphoridae) species of forensic significance.
Abstract:
Development of an antigen-based rapid diagnostic test for the identification of blowfly (Calliphoridae) species of forensic significance.
Abstract.
Author URL.
Waller PL, Thornton CR, Farley D, Groenhof A (2008). Pathogens and other fungi in growing media constituents.
Abstract:
Pathogens and other fungi in growing media constituents
Abstract.
Thornton CR (2008). Tracking fungi in soil with monoclonal antibodies.
Abstract:
Tracking fungi in soil with monoclonal antibodies
Abstract.
Thornton CR, Dewey FM, Gilligan CA (1994). Development of monoclonal antibody-based immunological assays for the detection of live propagules of Rhizoctonia solani in soil.
Publications by year
In Press
Thornton CR, Davies GE, Dougherty L (In Press). Development of a Monoclonal Antibody and a Lateral-Flow Device for the Rapid Serological Detection of a Mucorales-specific Biomarker. Frontiers in Cellular and Infection Microbiology
2023
Thornton CR (2023). Serological Techniques for Diagnosis. In Lane CR, Beales PA, Hughes KJD (Eds.)
Fungal Plant Pathogens Applied Techniques, Wallingford, Oxfordshire, UK: Cabi, 169-182.
Abstract:
Serological Techniques for Diagnosis
Abstract.
Thornton CR (2023). The Potential for Rapid Antigen Testing for Mucormycosis in the Context of COVID-19.
Expert Review of Molecular DiagnosticsAbstract:
The Potential for Rapid Antigen Testing for Mucormycosis in the Context of COVID-19
Introduction: Mucormycosis is a highly aggressive angio-invasive disease of humans caused by Mucorales fungi. Prior to the COVID-19 pandemic, mucormycosis was a rare mycosis typically seen in immunocompromised patients with hematological malignancies or in transplant recipients. During the second wave of the pandemic, there was a dramatic increase in the disease, especially in India, where a unique set of circumstances led to large numbers of life-threatening and disfiguring rhino-orbital-
cerebral mucormycosis (ROCM) infections.
Areas covered: the review examines mucormycosis as a super-infection of COVID-19 patients, and the risk factors for COVID-19-associated mucormycosis (CAM) that drove the ROCM epidemic in India. The limitations of current diagnostic procedures are identified, and the measures needed to improve the speed and accuracy of detection discussed.
Expert opinion: Despite increased awareness, global healthcare systems remain unprepared for further outbreaks of ROCM. Current diagnosis of the disease is slow and inaccurate, negatively impacting on patient survival. This is most evident in low- to middle income countries which lack suitably equipped diagnostic facilities for rapid identification of the infecting pathogens. Rapid antigen testing using point-of-care lateral-flow assays could potentially have aided in the quick and accurate diagnosis of the disease, allowing earlier intervention with surgery and Mucorales-active antifungal drugs.
Abstract.
2022
Schwenk J, Maurer A, Beziere N, Fiz F, Boschetti F, Geistlich S, Seyfried D, Gunzer M, Reischl G, Wehrmueller JE, et al (2022). Antibody-guided Molecular Imaging of Aspergillus Lung Infections in Leukemia Patients. Journal of Nuclear Medicine, 63, 1450-1451.
Parslow BY, Thornton CR (2022). Continuing Shifts in Epidemiology and Antifungal Susceptibility Highlight the Need for Improved Disease Management of Invasive Candidiasis.
Microorganisms,
10(6).
Abstract:
Continuing Shifts in Epidemiology and Antifungal Susceptibility Highlight the Need for Improved Disease Management of Invasive Candidiasis
Invasive candidiasis (IC) is a systemic life-threatening infection of immunocompromised humans, but remains a relatively neglected disease among public health authorities. Ongoing assessments of disease epidemiology are needed to identify and map trends of importance that may necessitate improvements in disease management and patient care. Well established incidence increases, largely due to expanding populations of patients with predisposing risk factors, has led to increased clinical use and pressures on antifungal drugs. This has been exacerbated by a lack of fast, accurate diagnostics that have led treatment guidelines to often recommend preventative strategies in the absence of proven infection, resulting in unnecessary antifungal use in many instances. The consequences of this are multifactorial but a contribution to emerging drug resistance is of primary concern, with high levels of antifungal use heavily implicated in global shifts to more resistant Candida strains. Preserving and expanding the utility and number of antifungals should therefore be of the highest priority. This may be achievable through the development and use of biomarker tests, bringing about a new era in improved antifungal stewardship, as well as novel antifungals that offer favourable profiles by targeting Candida pathogenesis mechanisms over cell viability.
Abstract.
Davies GE, Thornton CR (2022). Development of a Monoclonal Antibody and a Serodiagnostic Lateral-Flow Device Specific to Rhizopus arrhizus (Syn. R. oryzae), the Principal Global Agent of Mucormycosis in Humans.
Journal of Fungi,
8(7).
Abstract:
Development of a Monoclonal Antibody and a Serodiagnostic Lateral-Flow Device Specific to Rhizopus arrhizus (Syn. R. oryzae), the Principal Global Agent of Mucormycosis in Humans
Mucormycosis is a highly aggressive angio-invasive disease of humans caused by fungi in the zygomycete order, Mucorales. Though a number of different species can cause mucormycosis, the principal agent of the disease worldwide is Rhizopus arrhizus, which accounts for the majority of rhino-orbital-cerebral, pulmonary, and disseminated infections in immunocompromised individuals. It is also the main cause of life-threatening infections in patients with poorly controlled diabetes mellitus, and in corticosteroid-treated patients with SARS-CoV-2 infection, where it causes the newly described disease, COVID-19-associated mucormycosis (CAM). Diagnosis currently relies on non-specific CT, a lengthy and insensitive culture from invasive biopsy, and a time-consuming histopathology of tissue samples. At present, there are no rapid antigen tests for the disease that detect biomarkers of infection, and which allow point-of-care diagnosis. Here, we report the development of an IgG1 monoclonal antibody (mAb), KC9, which is specific to Rhizopus arrhizus var. arrhizus (syn. Rhizopus oryzae) and Rhizopus arrhizus var. delemar (Rhizopus delemar), and which binds to a 15 kDa extracellular polysaccharide (EPS) antigen secreted during hyphal growth of the pathogen. Using the mAb, we have developed a competitive lateral-flow device (LFD) that allows rapid (30 min) and sensitive (~50 ng/mL running buffer) detection of the EPS biomarker, and which is compatible with human serum (limit of detection of ~500 ng/mL) and bronchoalveolar lavage fluid (limit of detection of ~100 ng/mL). The LFD, therefore, provides a potential novel opportunity for the non-invasive detection of mucormycosis caused by Rhizopus arrhizus.
Abstract.
Parslow BY, Thornton CR (2022). Distribution and Diagnostics of Invasive Candidiasis.
Scholarly Community EncyclopediaAbstract:
Distribution and Diagnostics of Invasive Candidiasis
Invasive candidiasis (IC) is a systemic life-threatening infection of immunocompromised humans, but remains a relatively neglected disease among public health authorities. Ongoing assessments of disease epidemiology are needed to identify and map trends of importance that may necessitate improvements in disease management and patient care. Well-established incidence increases, largely due to expanding populations of patients with pre-disposing risk factors, has led to increased clinical use and pressures on antifungal drugs. This has been exacerbated by a lack of fast, accurate diagnostics that have led treatment guidelines to often recommend preventative strategies in the absence of proven infection, resulting in unnecessary antifungal use in many instances. The consequences of this are multifactorial, but a contribution to emerging drug resistance is of primary concern, with high levels of antifungal use heavily implicated in global shifts to more resistant Candida strains.
Abstract.
Eseola A (2022). Spatial control of organelle dynamics during appressorium-mediated plant infection by Magnaporthe oryzae.
Abstract:
Spatial control of organelle dynamics during appressorium-mediated plant infection by Magnaporthe oryzae
Magnaporthe oryzae, the pathogen responsible for the rice blast disease, produces a specialised infection structure called an appressorium that uses massive turgor to break the tough outer cuticle of the rice leaf. Appressorium development is a tightly regulated process that requires surface recognition of a hard hydrophobic surface, successful traversal of cell cycle checkpoints, and autophagic conidial cell death. It is however unknown how organelle trafficking is regulated and spatially controlled in parallel with autophagy and cell cycle progression. I developed molecular markers and a quantitative technique to monitor the trafficking of specific organelles in M. oryzae wild-type strain Guy11 and an ∆atg8 autophagic mutant. Live-cell imaging and quantitative analysis enabled us to characterise the regulated trafficking of 10 organelles within the three-celled conidium during appressorium development. High-resolution live-cell imaging using a photoactivatable green fluorescent protein indicates that germination establishes a separate developmental programme for each conidium cell, permitting organelle trafficking from a single conidium cell into the appressorium while targeting the remaining two cells for autophagy. We discovered that organelle trafficking occurs independently of cell cycle checkpoints for transport into the appressorium. I have quantified the temporal sequence of organelle movement and de novo organelle biogenesis in the incipient appressorium using photoconvertible fluorescent localisation microscopy. Our study shed light on the spatial control of organelle dynamics associated with fungal infection-related morphogenesis.
Abstract.
2021
Henneberg S, Hasenberg A, Maurer A, Neumann F, Bornemann L, Gonzales-Menendez I, Kraus A, Hasenberg M, Thornton CR, Pichler BJ, et al (2021). Antibody-guided in vivo imaging of Aspergillus fumigatus lung infections during anti-fungal azole treatment.
Nature Communications,
12Abstract:
Antibody-guided in vivo imaging of Aspergillus fumigatus lung infections during anti-fungal azole treatment
Invasive pulmonary aspergillosis (IPA) is a life-threatening lung disease of immunocompromised humans, caused by the opportunistic fungal pathogen Aspergillus fumigatus. Inadequacies in current diagnostic procedures mean that early diagnosis of the disease, critical to patient survival, remains a major clinical challenge, and is leading to the empiric use of antifungal drugs and emergence of azole resistance. A non-invasive procedure that allows both unambiguous detection of IPA, and its response to azole treatment, is therefore needed. Here, we show that a humanised Aspergillus-specific monoclonal antibody, dual labelled with a radionuclide and fluorophore, can be used in immunoPET/MRI in vivo and 3D light sheet fluorescence microscopy ex vivo to quantify early A. fumigatus lung infections and to monitor the efficacy of azole therapy. Our antibody-guided approach reveals that early drug intervention is critical to prevent complete invasion of the lungs by the fungus, and demonstrates the power of molecular imaging as a non-invasive procedure for tracking IPA in vivo.
Abstract.
Davies G, Singh O, Prattes J, Hoenigl M, Sheppard PW, Thornton C (2021). Aspergillus fumigatus and its Allergenic Ribotoxin Asp f I: Old Enemies but New Opportunities for Urine-based Detection of Invasive Pulmonary Aspergillosis using Lateral-Flow Technology.
Journal of Fungi,
7(1).
Abstract:
Aspergillus fumigatus and its Allergenic Ribotoxin Asp f I: Old Enemies but New Opportunities for Urine-based Detection of Invasive Pulmonary Aspergillosis using Lateral-Flow Technology
Invasive pulmonary aspergillosis (IPA) caused by Aspergillus fumigatus is a life-threatening lung disease of immunocompromised patients. Diagnosis currently relies on non-specific chest CT, culture of the fungus from invasive lung biopsy, and detection of the cell wall carbohydrate galactomannan (GM) in serum or in BAL fluids recovered during invasive bronchoscopy. Urine provides an ideal bodily fluid for the non-invasive detection of pathogen biomarkers, with current urine-based immunodiagnostics for IPA focused on GM. Surrogate protein biomarkers might serve to improve disease detection. Here, we report the development of a monoclonal antibody (mAb), PD7, which is specific to A. fumigatus and related species in the Section Fumigati, and which binds to its 18 kDa ribotoxin Asp f I. Using PD7, we show that the protein is secreted during hyphal development, and so represents an ideal candidate for detecting invasive growth. We have developed a lateral-flow device (Afu-LFD®) incorporating the mAb which has a limit of detection of ~15 ng Asp f I/mL urine. Preliminary evidence of the test’s diagnostic potential is demonstrated with urine from a patient with acute lymphoid leukaemia with probable IPA. The Afu-LFD® therefore provides a potential novel opportunity for non-invasive urine-based detection of IPA caused by A. fumigatus.
Abstract.
2020
Amich J, Mokhtari Z, Strobel M, Vialetto E, Sheta D, Yu Y, Hartweg J, Kalleda N, Jarick K, Brede C, et al (2020). 3D light sheet fluorescence microscopy of lungs to dissect local host immune - Aspergillus fumigatus interactions.
mBio,
11Abstract:
3D light sheet fluorescence microscopy of lungs to dissect local host immune - Aspergillus fumigatus interactions
Aspergillus fumigatus is an opportunistic fungal pathogen that can cause life-threatening invasive lung infections in immunodeficient patients. The cellular and molecular processes of infection during onset, establishment and progression are highly complex and depend on both fungal attributes and the immune status of the host. Therefore, preclinical animal models are paramount to investigate and gain better insight into the infection process. Yet, despite their extensive use, commonly employed murine models of invasive pulmonary aspergillosis are not well understood due to analytical limitations. Here we present quantitative light sheet fluorescence microscopy (LSFM) to describe fungal growth and the local immune response in whole lungs at cellular resolution within its anatomical context. We analyzed three very common murine models of pulmonary aspergillosis based on immunosuppression with corticosteroids, chemotherapy-induced leukopenia or myeloablative irradiation. LSFM uncovered distinct architectures of fungal growth and degrees of tissue invasion in each model. Furthermore, LSFM revealed the spatial distribution, interaction and activation of two key immune cell populations in antifungal defense: alveolar macrophages and polymorphonuclear neutrophils. Interestingly, the patterns of fungal growth correlated with the detected effects of the immunosuppressive regimens on the local immune cell populations. Moreover, LSFM demonstrates that the commonly used intranasal route of spore administration did not result in a complete intra-alveolar deposition, as more than 60% of fungal growth occurred outside of the alveolar space. Hence, LSFM allows for more rigorous characterization than previously used methods of murine models of invasive pulmonary aspergillosis and pinpointing their strengths and limitations.
Abstract.
Gunzer M, Thornton CR, Beziere N (2020). Advances in the in vivo molecular imaging of invasive pulmonary aspergillosis.
Journal of Fungi,
6(4).
Abstract:
Advances in the in vivo molecular imaging of invasive pulmonary aspergillosis
Invasive pulmonary aspergillosis (IPA) is a life-threatening infection of immunocompromised patients with Aspergillus fumigatus, a ubiquitous environmental mould. While there are numerous functioning antifungal therapies, their high cost, substantial side effects and fear of overt resistance development preclude permanent prophylactic medication of risk-patients. Hence, a fast and definitive diagnosis of IPA is desirable, to quickly identify those patients that really require aggressive antimycotic treatment and to follow the course of the therapeutic intervention. Yet, despite decades of research into this issue such a diagnostic procedure is still not available. Here we discuss the array of currently available methods for IPA detection and their limits. We then show that molecular imaging using positron-emission-tomography (PET) combined with morphological computed tomography or magnetic imaging is highly promising to become a future non-invasive approach for IPA diagnosis and therapy monitoring, albeit still requiring thorough validation and relying on further acceptance and dissemination of the approach. Thereby our approach using the A. fumigatus-specific humanized monoclonal antibody hJF5 labelled with 64Cu as PET-tracer has proven highly effective in pre-clinical models and hence bears high potential for human application.
Abstract.
Thomas G (2020). Characterisation of the Volatile Chemical Signalling from the Beneficial Soil Fungus Trichoderma hamatum.
Abstract:
Characterisation of the Volatile Chemical Signalling from the Beneficial Soil Fungus Trichoderma hamatum
Trichoderma hamatum GD12 is a beneficial soil fungus attracting agricultural interest due to its ability to promote plant growth and elicit biocontrol against a range of fungal pathogens. Evidence suggests volatile organic compounds (VOCs) produced by beneficial fungi can play a role in these biological activities. Two previously generated mutants of T. hamatum GD12 demonstrate enhanced plant growth promotion and antagonism against a range of pathogens, respectively, for which differences in VOC production could be responsible. The N-acetyl-β-D-glucosaminidase deficient mutant of T. hamatum GD12 (ΔThnag::hph) showed enhanced growth promotion of lettuce, relative to T hamatum GD12. Moreover, ΔThnag::hph. demonstrated a loss of biocontrol activity against the fungal pathogen Sclerotinia sclerotiorum. Deletion of Heterochromatin Remodelling Protein 1 (ΔThhepA::hph) showed enhanced antagonism against a range of fungal pathogens, showing chemical zones of antibiosis against the pathogens, suggesting enhanced production of inhibitory compounds. However, the involvement of VOCs in these biological activities is currently unknown.
VOC analysis demonstrated quantitative and qualitative differences in VOC production across the three strains of T. hamatum, which could relate to their unique biological activities. Moreover, during the physical interaction of T. hamatum with S. sclerotiorum, significant changes in VOC production were observed relative to controls, including the de novo production of VOCs which were absent in controls, indicating ordinarily silent secondary metabolite clusters in T. hamatum GD12 are activated by the presence of S. sclerotiorum. To date, studies investigating microbial VOC production sample VOCs from fungi inoculated in vitro, which does not reflect the heterogenous environment where soil fungi usually reside. Here, plastic tubing coated in a sorptive fibre (polydimethylsiloxane (PDMS)) was effective at capturing T. hamatum VOCs directly from peat microcosms. During the interaction of T. hamatum with S. sclerotiorum in soil microcosms, PDMS tubing detected quantitative changes in VOC production, showing increases in the production of VOCs in co-cultured microcosms relative to axenic microcosms. Finally, a novel VOC is overproduced by the ΔThnag::hph mutant relative to GD12 and ΔThhepA::hph, potentially playing a role in the enhanced plant growth promotion observed. Taken together, this thesis provides a clearer understanding of the VOCs involved in the enhanced biocontrol and plant growth promoting capabilities of T. hamatum GD12.
Abstract.
Thornton CR (2020). Detection of the 'big five' mold killers of humans: Aspergillus, Fusarium, Lomentospora, Scedosporium and Mucormycetes.
Advances in Applied Microbiology,
110, 1-61.
Abstract:
Detection of the 'big five' mold killers of humans: Aspergillus, Fusarium, Lomentospora, Scedosporium and Mucormycetes
Fungi are an important but frequently overlooked cause of morbidity and mortality in humans. Life-threatening fungal infections mainly occur in immunocompromised patients, and are typically caused by environmental opportunists that take advantage of a weakened immune system. The filamentous fungus Aspergillus fumigatus is the most important and well-documented mould pathogen of humans, causing a number of complex respiratory diseases, including invasive pulmonary aspergillosis, an often fatal disease in patients with acute leukemia or in immunosuppressed bone marrow or solid organ transplant recipients. However, non-Aspergillus moulds are increasingly reported as agents of disseminated diseases, with Fusarium, Scedosporium, Lomentospora and mucormycete species now firmly established as pathogens of immunosuppressed and immunocompetent individuals. Despite well-documented risk factors for invasive fungal diseases, and increased awareness of the risk factors for life-threatening infections, the number of deaths attributable to moulds is likely to be severely underestimated driven, to a large extent, by the lack of readily accessible, cheap, and accurate tests that allow detection and differentiation of infecting species. Early diagnosis is critical to patient survival but, unlike Aspergillus diseases, where a number of CE-marked or FDA-approved biomarker tests are now available for clinical diagnosis, similar tests for fusariosis, scedosporiosis and mucormycosis remain experimental, with detection reliant on insensitive and slow culture of pathogens from invasive bronchoalveolar lavage fluid, tissue biopsy, or from blood. This review examines the ecology, epidemiology, and contemporary methods of detection of these mould pathogens, and the obstacles to diagnostic test development and translation of novel biomarkers to the clinical setting.
Abstract.
2019
Amich J, Mokhtari Z, Strobel M, Vialetto E, Kalleda N, Jarick KJ, Brede C, Jordan-Garrote A-L, Thusek S, Schmiedgen K, et al (2019). 3D light sheet fluorescence microscopy of lungs to dissect local host immune - Aspergillus fumigatus interactions.
bioRxiv, 1-26.
Abstract:
3D light sheet fluorescence microscopy of lungs to dissect local host immune - Aspergillus fumigatus interactions
Aspergillus fumigatus is an opportunistic fungal pathogen that can cause life-threatening invasive lung infections in immunodeficient patients. The cellular and molecular processes of infection during onset, establishment and progression are highly complex and depend on both fungal attributes and the immune status of the host. Therefore, preclinical animal models are paramount to investigate and gain better insight into the infection process. Yet, despite their extensive use, commonly employed murine models of invasive pulmonary aspergillosis are not well understood due to analytical limitations. Here we present quantitative light sheet fluorescence microscopy (LSFM) to describe fungal growth and the local immune response in whole lungs at cellular resolution within its anatomical context. We analyzed three very common murine models of pulmonary aspergillosis based on immunosuppression with corticosteroids, chemotherapy-induced leukopenia or myeloablative irradiation. LSFM uncovered distinct architectures of fungal growth and degrees of tissue invasion in each model. Furthermore, LSFM revealed the spatial distribution, interaction and activation of two key immune cell populations in antifungal defense: alveolar macrophages and polymorphonuclear neutrophils. Interestingly, the patterns of fungal growth correlated with the detected effects of the immunosuppressive regimens on the local immune cell populations. Moreover, LSFM demonstrates that the commonly used intranasal route of spore administration did not result in the desired intra-alveolar deposition, as more than 60% of fungal growth occurred outside of the alveolar space. Hence, LSFM allows for more rigorous characterization of murine models of invasive pulmonary aspergillosis and pinpointing their strengths and limitations.
Abstract.
Thornton C (2019). Bench-to-Bedside Diagnosis of Invasive Pulmonary Aspergillosis. 9th Trends in Medical Mycology. 11th - 14th Oct 2019.
Abstract:
Bench-to-Bedside Diagnosis of Invasive Pulmonary Aspergillosis
Abstract.
Maurer A, Beziere N, Seyfried D, Wehrmueller J, Spycher P, Schwenck J, Wild A-M, Davies G, Boschetti F, Wiehr S, et al (2019). Development and preclinical evaluation of an immunoPET tracer for clinical imaging of Aspergillosis.
Author URL.
Schwenck J, Beziere N, Maurer A, Wild AM, Henneberg H, Davies G, Reischl G, Spycher PR, Wehrmüller JE, Boschetti F, et al (2019). Translational immunoPET/MR imaging of invasive pulmonary aspergillosis. NuklearMedizin 2019. 3rd - 6th Apr 2019.
Abstract:
Translational immunoPET/MR imaging of invasive pulmonary aspergillosis
Abstract.
2018
Thornton CR (2018). Molecular Imaging of Invasive Pulmonary Aspergillosis using ImmunoPET/MRI: the Future Looks Bright.
Frontiers in Microbiology,
9Abstract:
Molecular Imaging of Invasive Pulmonary Aspergillosis using ImmunoPET/MRI: the Future Looks Bright
Invasive pulmonary aspergillosis (IPA) is a life-threatening lung disease of immuno-compromised humans caused by the ubiquitous environmental mould Aspergillus. Biomarker tests for the disease lack sensitivity and specificity, and culture of the fungus from invasive lung biopsy is slow, insensitive, and undesirable in critically ill patients. A Computed Tomogram (CT) of the chest offers a simple non-intrusive diagnostic procedure for rapid decision-making, and so is used in many haematology units to drive antifungal treatment. However, radiological indicators that raise the suspicion of IPA are either transient signs in the early stages of the disease, or are not specific for Aspergillus infection, with other angio-invasive moulds or bacterial pathogens producing comparable radiological manifestations in a chest CT. Improvements to the specificity of radiographic imaging of IPA have been attempted by coupling CT and Positron Emission Tomography (PET) with [18F]FDG, a marker of metabolic activity well-suited to cancer imaging, but with limited use in invasive fungal disease diagnostics due to its inability to differentiate between infectious etiologies, cancer, and inflammation. Bioluminescence imaging using single genetically modified strains of Aspergillus fumigatus has enabled in vivo monitoring of IPA in animal models of disease. For in vivo detection of Aspergillus lung infections in humans, radiolabelled Aspergillus-specific monoclonal antibodies, and iron siderophores, hold enormous potential for clinical diagnosis. This review examines the different experimental technologies used to image IPA, and recent advances in state-of-the-art molecular imaging of IPA using antibody-guided Positron Emission Tomography/Magnetic Resonance Imaging (immunoPET/MRI).
Abstract.
Box H, Negri C, Livermore J, Whalley S, Johnson A, McEntee L, Alastruey-Izquierdo A, Meis J, Thornton CR, Hope W, et al (2018). Pharmacodynamics of Voriconazole for Invasive Pulmonary Scedosporiosis.
Antimicrobial Agents and Chemotherapy,
62(5).
Abstract:
Pharmacodynamics of Voriconazole for Invasive Pulmonary Scedosporiosis
Scedosporium apiospermum is a medically important fungal pathogen that causes a wide range of infections in humans. There are relatively few antifungal agents that are active against Scedosporium spp. Little is known about the pharmacodynamics of voriconazole against Scedosporium. Both static and dynamic in vitro models of invasive scedosporiosis were developed. Monoclonal antibodies that target a soluble cell wall antigen secreted by Scedosporium apiospermum were used to describe the pharmacodynamics of voriconazole. Mathematical pharmacokinetic-pharmacodynamic models were fitted to the data to estimate the drug exposure required to suppress the release of fungal antigen. The experimental results were bridged to humans using Monte Carlo simulation. All 3 strains of S. apiospermum tested invaded through the cellular bilayer of the in vitro models and liberated antigen. There was a concentration-dependent decline in antigen with near maximal antifungal activity in all 3 strains with 10 mg/L. Similarly, there was a drug exposure dependent decline in circulating antigen in the dynamic model and complete suppression of antigen with an AUC of approximately 80 mg.h/L. A regression of the AUC:MIC versus area under the antigen time curve showed that near maximal effect was obtained with AUC:MIC of approximately 100. Monte Carlo simulation suggested that only isolates with an MIC of 0.5 mg/L enable pharmacodynamic targets to be acheived with a standard regimen of voriconazole. Isolates with higher MICs may need higher drug exposure targets than are currently recommended for other fungi.
Abstract.
Morad HOJ, Wild A-M, Wiehr S, Davies G, Maurer A, Pichler BJ, Thornton CR (2018). Pre-Clinical Imaging of Invasive Candidiasis using ImmunoPET/MR.
Frontiers in Microbiology,
9, 1-15.
Abstract:
Pre-Clinical Imaging of Invasive Candidiasis using ImmunoPET/MR
The human commensal yeast Candida is the 4th most common cause of hospital-acquired bloodstream infections, with C. albicans accounting for the majority of the >400,000 life-threatening infections annually. Diagnosis of invasive candidiasis (IC), a disease encompassing candidemia (blood-borne yeast infection) and deep-seated organ infections, is a major challenge since clinical manifestations of the disease are indistinguishable from viral, bacterial and other fungal diseases, and diagnostic tests for biomarkers in the bloodstream such as PCR, ELISA and pan-fungal β-D-glucan lack either standardisation, sensitivity or specificity. Blood culture remains the gold standard for diagnosis, but test sensitivity is poor and turn-around time slow. Furthermore, cultures can only be obtained when the yeast resides in the bloodstream, with samples recovered from hematogenous infections often yielding negative results. Consequently, there is a pressing need for a diagnostic test that allows the identification of metastatic foci in deep-seated Candida infections, without the need for invasive biopsy. Here, we report the development of a highly specific mouse IgG3 monoclonal antibody (MC3) that binds to a putative β-1,2-mannan epitope present in high molecular weight mannoproteins and phospholipomannans on the surface of yeast and hyphal morphotypes of C. albicans, and its use as a [64Cu]NODAGA-labeled tracer for whole-body pre-clinical imaging of deep-seated C. albicans infections using antibody-guided positron emission tomography and magnetic resonance imaging (immunoPET/MRI). When used in a mouse intravenous (i.v.) challenge model that faithfully mimics disseminated C. albicans infections in humans, the [64Cu]NODAGA-MC3 tracer accurately detects infections of the kidney, the principal site of blood-borne candidiasis. Using a strain of the emerging human pathogen Candida auris that reacts with MC3 in vitro, but which is non-infective in i.v. challenged mice, we demonstrate the accuracy of the tracer in diagnosing invasive infections in vivo. This pre-clinical study demonstrates the principle of. antibody-guided molecular imaging for detection of deep organ infections in IC, without the need for invasive tissue biopsy.
Abstract.
2017
Liew N, Mazon Moya MJ, Wierzbicki C, Hollinshead M, Dillon MJ, Thornton CR, Ellison A, Cable J, Fisher MC, Mostowy S, et al (2017). Chytrid fungus infection in zebrafish demonstrates that the pathogen can parasitise non-amphibian vertebrate hosts.
Nature Communications,
8, 15048-15048.
Abstract:
Chytrid fungus infection in zebrafish demonstrates that the pathogen can parasitise non-amphibian vertebrate hosts
Aquatic chytrid fungi threaten amphibian biodiversity worldwide owing to their ability to rapidly expand their geographical distributions and to infect a wide range of hosts. Combating this risk requires an understanding of chytrid host range to identify potential reservoirs of infection and to safeguard uninfected regions through enhanced biosecurity. Here we extend our knowledge on the host range of the chytrid Batrachochytrium dendrobatidis by demonstrating infection of a non-amphibian vertebrate host, the zebrafish. We observe dose-dependent mortality and show that chytrid can infect and proliferate on zebrafish tissue. We also show that infection phenotypes (fin erosion, cell apoptosis and muscle degeneration) are direct symptoms of infection. Successful infection is dependent on disrupting the zebrafish microbiome, highlighting that, as is widely found in amphibians, commensal bacteria confer protection against this pathogen. Collectively, our findings greatly expand the limited tool kit available to study pathogenesis and host response to chytrid infection.
Abstract.
Hoenigl M, Eigl S, Heldt S, Duettmann W, Thornton C, Prattes J (2017). Clinical Evaluation of the Newly Formatted Lateral-Flow Device for Invasive Pulmonary Aspergillosis.
Mycoses,
00, 1-4.
Abstract:
Clinical Evaluation of the Newly Formatted Lateral-Flow Device for Invasive Pulmonary Aspergillosis
The study evaluated the newly formatted Aspergillus-specific lateral-flow-device (LFD), and compared its performance to the original prototype “old” LFD test using BALF samples from 28 patients (14 patients with probable/proven invasive pulmonary aspergillosis [IPA] and 14 patients with no evidence for IPA). A total of 10/14 (71%) of BALF samples from patients with probable/proven IPA resulted positive with the new LFD, including 8/9 with true-positive and 2/5 with false-negative results with the old LFD. All 14 samples from patients without IPA resulted negative with the new LFD; specificity of the new LFD was significantly improved compared to the old LFD.
Abstract.
Duvaux L, Shiller J, Vandeputte P, Duge de Bernoville T, Thornton CR, Papon N, Le Cam B, Bouchara J-P, Gasteboise A (2017). Draft genome sequence of the human pathogenic fungus Scedosporium boydii.
Genome Announcements,
5, e00871-17-e00871-17.
Abstract:
Draft genome sequence of the human pathogenic fungus Scedosporium boydii
The opportunistic fungal pathogen Scedosporium boydii is the most common Scedosporium species in French patients with cystic fibrosis. Here we present the first genome report for S. boydii, providing a resource which may enable the elucidation of the pathogenic mechanisms in this species.
Abstract.
Pateau V, Razafimandimby B, Vandeputte P, Guillemette T, Thornton CR, Bouchara J-P, Giraud S (2017). Gene disruption in Scedosporium aurantiacum: proof of concept with the disruption of SODC gene encoding a cytosolic Cu,Zn-superoxide dismutase.
Mycopathologia,
183(1), 241-249.
Abstract:
Gene disruption in Scedosporium aurantiacum: proof of concept with the disruption of SODC gene encoding a cytosolic Cu,Zn-superoxide dismutase
Scedosporium species are opportunistic pathogens responsible for a large variety of infections in humans. An increasing occurrence was observed in patients with underlying conditions such as immunosuppression or cystic fibrosis. Indeed, the genus Scedosporium ranks the second among the filamentous fungi colonizing the respiratory tracts of the CF patients. To date, there is very scarce information on the pathogenic mechanisms, at least in part because of the limited genetic tools available. In the present study, we successfully developed an efficient transformation and targeted gene disruption approach on the species Scedosporium aurantiacum. The disruption cassette was constructed using double-joint PCR procedure, and resistance to hygromycin B as the selection marker. This proof of concept was performed on the functional gene SODC encoding the Cu,Zn-superoxide dismutase. Disruption of the SODC gene improved susceptibility of the fungus to oxidative stress. This technical advance should open new research areas and help to better understand the biology of Scedosporium species.
Abstract.
Henneberg S, Thornton C, Wiehr S, Rolle AM, Pichler B, Hasenberg M, Weski J, Hasenberg A, Gunzer M (2017). Intravital lung imaging to study the interactions between immune cells and Aspergillus fumigatus.
Author URL.
Shah A, Abdolrasouli A, Schelenz S, Thornton C, Ni MZ, Devaraj A, Devic N, Ward L, Carby M, Reed A, et al (2017). Latent class modelling for pulmonary aspergillosis diagnosis in lung transplant recipients.
Thorax,
72(3), A13-A14.
Abstract:
Latent class modelling for pulmonary aspergillosis diagnosis in lung transplant recipients
Rationale Timely, accurate diagnosis of invasive aspergillosis (IA) is key to enable initiation of antifungal therapy in lung transplantation. Despite promising novel fungal biomarkers, the lack of a diagnostic gold-standard creates difficulty in determining utility.
Objectives This study aimed to use latent class modelling of fungal diagnostics to classify lung transplant recipients (LTR) with IA in a large single centre.
Methods Regression models were used to compare composite biomarker testing of bronchoalveolar lavage to clinical and EORTC-MSG guideline-based diagnosis of IA with mortality used as a surrogate primary outcome measure. Bootstrap anal- ysis identified radiological features associated with IA. Bayesian latent class modelling was used to define IA.
Measurements and Main Results a clinical diagnosis of fungal infection (P =
Abstract.
Thornton CR (2017). Monoclonal antibody JF5 for aspergillosis: from BALF to immunoPET/MRI. Trends in Medical Mycology. 6th - 9th Oct 2017.
Abstract:
Monoclonal antibody JF5 for aspergillosis: from BALF to immunoPET/MRI
Abstract.
Author URL.
Davies G, Rolle A-M, Maurer A, Spycher PR, Schillinger C, Solouk-Saran D, Hasenberg M, Weski J, Foslet J, Dubois A, et al (2017). Towards Translational ImmunoPET/MR Imaging of Invasive Pulmonary Aspergillosis: the Humanised Monoclonal Antibody JF5 Detects Aspergillus Lung Infections in Vivo.
Theranostics,
7, 3398-3414.
Abstract:
Towards Translational ImmunoPET/MR Imaging of Invasive Pulmonary Aspergillosis: the Humanised Monoclonal Antibody JF5 Detects Aspergillus Lung Infections in Vivo
Invasive pulmonary aspergillosis (IPA) is a life-threatening lung disease of hematological malignancy and bone marrow transplant patients caused by the ubiquitous environmental fungus Aspergillus fumigatus. Current diagnostic tests for the disease lack sensitivity as well as specificity, and culture of the fungus from invasive lung biopsy, considered the gold standard for IPA detection, is slow and often not possible in critically ill patients. In a previous study, we reported the development of a novel non-invasive procedure for IPA diagnosis based on antibody-guided positron emission tomography and magnetic resonance imaging (immunoPET/MRI) using a [64Cu]DOTA-labeled mouse monoclonal antibody (mAb), mJF5, specific to Aspergillus. To enable translation of the tracer to the clinical setting, we report here the development of a humanised version of the antibody (hJF5), and pre-clinical imaging of lung infection using a [64Cu]NODAGA-hJF5 tracer. The humanised antibody tracer shows a significant increase in in vivo biodistribution in A. fumigatus infected lungs compared to its radiolabeled murine counterpart [64Cu]NODAGA-mJF5. Using reverse genetics of the pathogen, we show that the antibody binds to the antigenic determinant 1,5-galactofuranose (Galf) present in a diagnostic mannoprotein antigen released by the pathogen during invasive growth in the lung. The absence of the epitope Galf in mammalian carbohydrates, coupled with the enhanced imaging capabilities of the hJF5 antibody, means that the [64Cu]NODAGA-hJF5 tracer developed here represents an ideal candidate for the diagnosis of IPA and translation to the clinical setting.
Abstract.
Dillon MJ, Bowkett AE, Bungard MJ, Beckman KM, O'Brien MF, Bates K, Fisher MC, Stevens JR, Thornton CR (2017). Tracking the amphibian pathogens Batrachochytrium dendrobatidis and Batrachochytrium salamandrivorans using a highly specific monoclonal antibody and lateral-flow technology.
Microbial Biotechnology,
10(2), 381-394.
Abstract:
Tracking the amphibian pathogens Batrachochytrium dendrobatidis and Batrachochytrium salamandrivorans using a highly specific monoclonal antibody and lateral-flow technology
The fungus Batrachochytrium dendrobatidis (Bd) causes chytridiomycosis, a lethal epizootic disease of amphibians. Rapid identification of the pathogen and biosecurity is essential to prevent its spread, but current laboratory-based tests are time-consuming and require specialist equipment. Here, we describe the generation of an IgM monoclonal antibody (mAb), 5C4, specific to Bd as well as the related salamander and newt pathogen Batrachochytrium salamandrivorans (Bsal). The mAb, which binds to a glycoprotein antigen present on the surface of zoospores, sporangia and zoosporangia, was used to develop a lateral-flow assay (LFA) for rapid (15 min) detection of the pathogens. The LFA detects known lineages of Bd and also Bsal, as well as the closely related fungus Homolaphlyctis polyrhiza, but does not detect a wide range of related and unrelated fungi and oomycetes likely to be present in amphibian habitats. When combined with a simple swabbing procedure, the LFA was 100% accurate in detecting the water-soluble 5C4 antigen present in skin, foot and pelvic samples from frogs, newts and salamanders naturally infected with Bd or Bsal. Our results demonstrate the potential of the portable LFA as a rapid qualitative assay for tracking these amphibian pathogens and as an adjunct test to nucleic acid-based detection methods.
Abstract.
2016
Escudero N, Ferreira SR, Lopez-Moya F, Naranjo-Ortiz MA, Marin-Ortiz AI, Thornton CR, Lopez-Llorca LV (2016). Chitosan enhances parasitism of Meloidogyne javanica eggs by the nematophagous fungus Pochonia chlamydosporia.
Fungal Biology,
120(4), 572-585.
Abstract:
Chitosan enhances parasitism of Meloidogyne javanica eggs by the nematophagous fungus Pochonia chlamydosporia
Pochonia chlamydosporia (Pc), a nematophagous fungus and root endophyte, uses appressoria and extracellular enzymes, principally proteases, to degrade the eggs of plant parasitic nematodes (PPN). Unlike other fungi, Pc is resistant to chitosan, a deacetylated form of chitin, used in agriculture as a biopesticide to control plant pathogens. In the present work, we show that chitosan increases pathogenicity of Pc to Meloidogyne javanica by stimulating appressorial differentiation and parasitism of eggs. Using antibodies specific to the Pc enzymes VCP1 (a subtilisin), and SCP1 (a serine carboxypeptidase), we demonstrate chitosan elicitation of the fungal proteases during the parasitic process. Chitosan increases VCP1 immuno-labelling in the cells walls of Pc conidia, hyphal tips of germinating spores, and in appressoria on infected M. javanica eggs. These results support the role of proteases in pathogenicity of the fungus and their activation by chitosan. Phylogenetic analysis of the Pc genome reveals a large diversity of subtilisins (S8) and serine carboxypeptidases (S10). The VCP1 group in the S8 tree shows evidence of gene duplication indicating recent adaptations to nutrient sources. Our results show that chitosan enhances Pc pathogenicity through increased proteolytic activities and can be used to improve the efficacy of M. javanica biocontrol.
Abstract.
Al-Maqtoofi M, Thornton CR (2016). Detection of human pathogenic Fusarium species in hospital and communal sink biofilms by using a highly specific monoclonal antibody.
Environmental Microbiology,
18, 3620-3634.
Abstract:
Detection of human pathogenic Fusarium species in hospital and communal sink biofilms by using a highly specific monoclonal antibody
The fungus Fusarium is well known as a plant pathogen, but has recently emerged as an opportunistic pathogen of humans. Habitats providing direct human exposure to infectious propagules are largely unknown, but there is growing evidence that plumbing systems are sources of human pathogenic strains in the Fusarium solani species complex (FSSC) and Fusarium oxysporum species complex (FOSC), the most common groups infecting humans. Here, we use a newly developed Fusarium-specific monoclonal antibody (mAb ED7) to track FSSC and FOSC strains in sink drain biofilms by detecting its target antigen, an extracellular 200kDa carbohydrate, in saline swabs. The antigen was detectable in 52% of swab samples collected from sinks across a University campus and a tertiary care hospital. The mAb was 100% accurate in detecting FSSC, FOSC and F. dimerum species complex (FDSC) strains that were present, as mixed fungal communities, in 83% of sink drain biofilms. Specificity of the ELISA was confirmed by sequencing of the internally transcribed spacer 1 (ITS1)-5.8S-ITS2 rRNA-encoding regions of culturable yeasts and molds that were recovered using mycological culture, while translation elongation factor (TEF)-1α analysis of Fusarium isolates included FSSC 1-a, FOSC 33 and FDSC ET-gr, the most common clinical pathotypes in each group.
Abstract.
Sharpe RA, Le Cocq K, Nikolaou V, Osborne NJ, Thornton CR (2016). Identifying risk factors for exposure to culturable allergenic fungi in energy efficient homes by using highly specific monoclonal antibodies.
Environmental Research,
144, 32-42.
Abstract:
Identifying risk factors for exposure to culturable allergenic fungi in energy efficient homes by using highly specific monoclonal antibodies
The aim of this study was to determine the accuracy of monoclonal antibodies (mAbs) in identifying culturable allergenic fungi present in visible mould growth in energy efficient homes, and to identify risk factors for exposure to these known allergenic fungi. Swabs were taken from fungal contaminated surfaces and culturable yeasts and moulds isolated by using mycological culture. Soluble antigens from cultures were tested by ELISA using mAbs specific to the culturable allergenic fungi Aspergillus and Penicillium spp. Ulocladium, Alternaria, and Epicoccum spp. Cladosporium spp. Fusarium spp. and Trichoderma spp. Diagnostic accuracies of the ELISA tests were determined by sequencing of the internally transcribed spacer 1 (ITS1)-5.8S-ITS2-encoding regions of recovered fungi following ELISA. There was 100% concordance between the two methods, with ELISAs providing genus-level identity and ITS sequencing providing species-level identities (210 out of 210 tested). Species of Aspergillus/Penicillium, Cladosporium, Ulocladium/Alternaria/Epicoccum, Fusarium and Trichoderma were detected in 82% of the samples. The presence of condensation was associated with an increased risk of surfaces being contaminated by Aspergillus/Penicillium spp. and Cladosporium spp. whereas moisture within the building fabric (water ingress/rising damp) was only associated with increased risk of Aspergillus/Penicillium spp. Property type and energy efficiency levels were found to moderate the risk of indoor surfaces becoming contaminated with Aspergillus/Penicillium and Cladosporium which in turn was modified by the presence of condensation, water ingress and rising damp, consistent with previous literature.
Abstract.
Rolle AM, Hasenberg M, Thornton CR, Solouk-Saran D, Männ L, Weski J, Maurer A, Fischer E, Spycher PR, Schibli R, et al (2016). ImmunoPET/MR imaging allows specific detection of Aspergillus fumigatus lung infection in vivo.
Proceedings of the National Academy of Sciences of the United States of America,
113(8), E1026-E1033.
Abstract:
ImmunoPET/MR imaging allows specific detection of Aspergillus fumigatus lung infection in vivo
Invasive pulmonary aspergillosis (IPA) is a life-threatening lung disease caused by the fungus Aspergillus fumigatus, and is a leading cause of invasive fungal infection-related mortality and morbidity in patients with hematological malignancies and bone marrow transplants. We developed and tested a novel probe for noninvasive detection of A. fumigatus lung infection based on antibody-guided positron emission tomography and magnetic resonance (immunoPET/MR) imaging. Administration of a [64Cu]DOTA-labeled A. fumigatus-specific monoclonal antibody (mAb), JF5, to neutrophil-depleted A. fumigatus-infected mice allowed specific localization of lung infection when combined with PET. Optical imaging with a fluorochrome-labeled version of the mAb showed colocalization with invasive hyphae. The mAbbased newly developed PET tracer [64Cu]DOTA-JF5 distinguished IPA from bacterial lung infections and, in contrast to [18F]FDG-PET, discriminated IPA from a general increase in metabolic activity associated with lung inflammation. To our knowledge, this is the first time that antibody-guided in vivo imaging has been used for noninvasive diagnosis of a fungal lung disease (IPA) of humans, an approach with enormous potential for diagnosis of infectious diseases and with potential for clinical translation.
Abstract.
Wiehr S, Warnke P, Rolle A-M, Schutz M, Kohlhofer U, Quintanilla-Martinez de Fend L, Maurer A, Thornton C, Boschetti F, Reischl G, et al (2016). New pathogen-specific immunoPET/MR tracer for molecular imaging of a systemic bacterial infection.
Oncotarget,
7, 10990-11001.
Abstract:
New pathogen-specific immunoPET/MR tracer for molecular imaging of a systemic bacterial infection
The specific and rapid detection of Enterobacteriaceae, the most frequent cause of gram-negative bacterial infections in humans, remains a major challenge. We developed a non-invasive method to rapidly detect systemic Yersinia enterocolitica infections using immunoPET (antibody-targeted positron emission tomography) with [64Cu]NODAGA-labeled Yersinia-specific polyclonal antibodies targeting the outer membrane protein YadA. In contrast to the tracer [18F]FDG, [64Cu]NODAGA-YadA uptake co-localized in a dose dependent manner with bacterial lesions of Yersinia-infected mice, as detected by magnetic resonance (MR) imaging. This was accompanied by elevated uptake of [64Cu]NODAGA-YadA in infected tissues, in ex vivo biodistribution studies, whereas reduced uptake was observed following blocking with unlabeled anti-YadA antibody. We show, for the first time, a bacteria-specific, antibody-based, in vivo imaging method for the diagnosis of a Gram-negative enterobacterial infection as a proof of concept, which may provide new insights into pathogen-host interactions.
Abstract.
Al-Laaeiby A, Kershaw M, Penn T, Thornton CR (2016). Targeted disruption of melanin biosynthesis genes in the human pathogenic fungus Lomentospora prolificans and its consequences for pathogen survival.
International Journal of Molecular Sciences,
17(4), 1-18.
Abstract:
Targeted disruption of melanin biosynthesis genes in the human pathogenic fungus Lomentospora prolificans and its consequences for pathogen survival
The dematiaceous (melanised) fungus Lomentospora (Scedosporium) prolificans is a life-threatening opportunistic pathogen of immunocompromised humans, resistant to anti-fungal drugs. Melanin has been shown to protect human pathogenic fungi against antifungal drugs, oxidative killing and environmental stresses. To determine the protective role of melanin in L. prolificans to oxidative killing (H2O2), UV radiation and the polyene anti-fungal drug amphotericin B, targeted gene disruption was used to generate mutants of the pathogen lacking the dihydroxynaphthalene (DHN)-melanin biosynthetic enzymes polyketide synthase (PKS1), tetrahydroxynapthalene reductase (4HNR) and scytalone dehydratase (SCD1). Infectious propagules (spores) of the wild-type strain 3.1 were black/brown, whereas spores of the PKS-deficient mutant ΔLppks1::hph were white. Complementation of the albino mutant ΔLppks1::hph restored the black-brown spore pigmentation, while the 4HNR-deficient mutant ΔLp4hnr::hph and SCD-deficient mutant ΔLpscd1::hph both produced orange-yellow spores. The mutants ΔLppks1::hph and ΔLp4hnr::hph showed significant reductions in spore survival following H2O2 treatment, while spores of ΔLpscd1::hph and the ΔLppks1::hph complemented strain ΔLppks1::hph:PKS showed spore survivals similar to strain 3.1. Spores of the mutants ΔLp4hnr::hph and ΔLpscd1::hph and complemented strain ΔLppks1::hph:PKS showed spore survivals similar to 3.1 following exposure to UV radiation, but survival of ΔLppks1::hph spores was significantly reduced compared to the wild-type strain. Strain 3.1 and mutants ΔLp4hnr::hph and ΔLppks1::hph:PKS were resistant to amphotericin B while, paradoxically, the PKS1- and SCD1-deficient mutants showed significant increases in growth in the presence of the antifungal drug. Taken together, these results show that while melanin plays a protective role in the survival of the pathogen to oxidative killing and UV radiation, melanin does not contribute to its resistance to amphotericin B.
Abstract.
Shaw S, Le Cocq K, Paszkiewicz K, Moore K, Winsbury R, Studholme D, Salmon D, Thornton CR, Grant MR (2016). Transcriptional reprogramming underpins enhanced plant
growth promotion by the biocontrol fungus Trichoderma hamatum GD12
during antagonistic interactions with Sclerotinia sclerotiorum in
soil.
Molecular Plant Pathology,
17, 1425-1441.
Abstract:
Transcriptional reprogramming underpins enhanced plant
growth promotion by the biocontrol fungus Trichoderma hamatum GD12
during antagonistic interactions with Sclerotinia sclerotiorum in
soil
The free-living soil fungus Trichoderma hamatum strain GD12 is notable amongst Trichoderma strains in both controlling plant diseases and in stimulating plant growth, a property enhanced during its antagonistic interactions with pathogens in soil. These attributes, alongside its markedly expanded genome and proteome compared to other biocontrol and plant growth promoting Trichoderma strains, imply a rich potential for sustainable alternatives to synthetic pesticides and fertilisers for controlling plant disease and increasing yields. The purpose of this study was to investigate the transcriptional responses of GD12 underpinning its biocontrol and plant growth promotion capabilities during antagonistic interactions with the pathogen Sclerotinia sclerotiorum in soil. Using an extensive mRNA-seq study capturing different time points during the pathogen-antagonist interaction in soil, we show that dynamic and biphasic signatures in the GD12 transcriptome underpin its biocontrol and plant (lettuce) growth promotional activities. Functional predictions of differentially expressed genes demonstrate the enrichment of transcripts encoding proteins involved in transportation and oxidation-reduction reactions during both processes and an over-representation of siderophores. We identify a biphasic response during biocontrol characterised by a significant induction of transcripts encoding small-secreted cysteine rich proteins, secondary metabolite producing gene clusters and genes unique to GD12. These data support the hypothesis that Sclerotinia biocontrol is mediated by the synthesis and secretion of antifungal compounds and that GD12's unique reservoir of uncharacterised genes is actively recruited during effective biological control of a plurivorous plant pathogen.
Abstract.
Solouk D, Bergmueller E, Buffen K, Rolle A-M, Thornton C, Weski J, Gunzer M, Hasenberg M (2016). Viability of a promising antibody (JF5) for the fast detection of invasive pulmonary aspergillosis (IPA).
MYCOSES,
59, 32-32.
Author URL.
2015
Prattes J, Lackner M, Eigl S, Reischies F, Raggam RB, Koidl C, Flick H, Wolfler A, Neumeister P, Thornton CR, et al (2015). Diagnostic accuracy of the Aspergillus specific Bronchoalveolar Lavage Lateral-Flow Assay in Haematological Malignancy Patients. Mycoses: diagnosis, therapy and prophylaxis of fungal diseases, 58(8), 461-469.
Sharpe R, Thornton CR, Nikolaou V, Osborne N (2015). Fuel poverty increases risk of mould contamination, regardless of adult risk perception & ventilation in social housing properties. Environment International, 79, 115-129.
Eigl S, Prattes J, Reischies FM, Raggam RB, Spiess B, Reinwald M, Buchheidt D, Thornton CR, Krause R, Flick H, et al (2015). Galactomannan Testing and Aspergillus PCR in same-day bronchoalveolar lavage and blood samples obtained from patients with hematological malignancies at risk for invasive mould infection.
MYCOSES,
58, 157-157.
Author URL.
Sharpe R, Thornton CR, Nikolaou V, Osborne N (2015). Higher energy efficient homes are associated with increased risk of doctor diagnosed asthma in a UK sub population. Environment International, 75, 234-244.
Sharpe RA, Thornton CR, Nikolaou V, Osborne NJ (2015). Household energy efficiency, fungi and allergic asthma.
Author URL.
Thornton CR, Ryder LS, Le Cocq K, Soanes DM (2015). Identifying the emerging human pathogen Scedosporium prolificans by using a species-specific monoclonal antibody that binds to the melanin biosynthetic enzyme tetrahydroxynaphthalene reductase. Environmental Microbiology, 17(4), 1023-1048.
Bustin SA, Thornton CR, Shannon M, Agrawal SG, Lass-Floerl C, Mutschlechner W, Roberts D, Johnson G (2015). Immuno PCR-based assays for the early detection of Aspergillus species.
MYCOSES,
58, 28-28.
Author URL.
Thornton CR, Wills OE (2015). Immunodetection of Fungal and Oomycete Pathogens: Established and Emerging Threats to Human Health, Animal Welfare and Global Food Security. Critical Reviews in Microbiology, 41(1), 27-51.
Osborne NJ, Thornton CR, Sharpe RA (2015). Indoor Fungal Exposure and Allergic Respiratory Disease.
Current Allergy and Asthma Reports,
15(12).
Abstract:
Indoor Fungal Exposure and Allergic Respiratory Disease
A gathering body of evidence has repeatedly revealed associations between indoor fungi and initiation, promotion, and exacerbation of allergic respiratory disease. The relationship between the exposure and outcome are complicated by the difficulties in measuring both exposure and outcome, the multifactorial nature of the disease, and the wide range of potential confounders. New technologies are becoming available that may enable better measurement of exposure and tighter case definitions so as to build more confidence in the associations discovered. The growing strength of the evidence base will aid the design of future public health interventions and generate new hypotheses on the cause of the rapid increase in allergic respiratory disease prevalence.
Abstract.
Sharpe RA, Bearman N, Thornton CR, Husk K, Osborne NJ (2015). Indoor fungal diversity and asthma: a meta-analysis and systematic review of risk factors.
Journal of Allergy and Clinical Immunology,
135(1), 110-122.
Abstract:
Indoor fungal diversity and asthma: a meta-analysis and systematic review of risk factors
Background Indoor dampness increases the risk of indoor fungal growth. A complex interaction between occupant behaviors and the built environment are thought to affect indoor fungal concentrations and species diversity, which are believed to increase the risk of having asthma, exacerbation of asthma symptoms, or both. To date, no systematic review has investigated this relationship.
Abstract.
Sharpe RA, Bearman N, Thornton CR, Husk K, Osborne NJ (2015). Indoor fungal diversity and asthma: a metaanalysis and systematic review of risk factors. Journal of Allergy and Clinical Immunology, 135(1), 110-122.
Eigl S, Prattes J, Reinwald M, Thornton CR, Reischies F, Spiess B, Neumeister P, Zollner-Schwetz I, Raggam RB, Flick H, et al (2015). Influence of Mould-active Antifungal Treatment on the Performance of the Aspergillus specific Bronchoalveolar Lavage Lateral-Flow Device Test. International Journal of Antimicrobial Agents, 46, 401-405.
Eigl S, Prattes J, Lackner M, Willinger B, Spiess B, Selitsch B, Meilinger M, Reischies F, Neumeister P, Wolfler A, et al (2015). Multicenter evaluation of a Lateral-Flow Device Test for Diagnosing Invasive Pulmonary Aspergillosis in ICU Patients. Critical Care, 19, 1-9.
Miceli M, Goggins MI, Chander P, Sekaran AK, Kizy AE, Samuel L, Jiang H, Thornton CR, Ramesh M, Alangaden G, et al (2015). Performance of Lateral Flow Device and Galactomannan for the Detection of Aspergillus Species in Bronchoalveolar Fluid of Patients at Risk for Invasive Pulmonary Aspergillosis. Mycoses: diagnosis, therapy and prophylaxis of fungal diseases, 58, 368-374.
Box H, Livermore J, McEntee L, Johnson A, Whalley S, Goodwin J, Birch M, Thornton C (2015). Pharmacodynamics of a novel antifungal agent, F901318 against Pseudallescheria boydii and Scedosporium apiospermum.
Abstract:
Pharmacodynamics of a novel antifungal agent, F901318 against Pseudallescheria boydii and Scedosporium apiospermum
Background: F901318 is a novel antifungal agent currently in phase I clinical development. F901318 has a unique mechanism of action. Pseudallescheria boydii and Scedosporium apiospermum are rare and emerging human fungal pathogens that can cause lethal infections. These infections are difficult to treat with existing antifungal agents. The pharmacokinetic-pharmacodynamic relationships of F901318 against these opportunistic pathogens were analysed in this study. Methods: Static and dynamic in vitro models of the human alveolus were developed to describe the pharmacodynamics of F901318. Isolates of S. apiospermum and P. boydii were studied in a static model. Three isolates of S. apiospermum were studied in a dynamic model. Concentration time profiles for F901318 were generated. Minimum inhibitory concentrations (MIC) were determined using both CLSI and EUCAST methodologies. Recently developed monoclonal antibodies specific for soluble cell wall antigens were used as biomarkers for the estimation of fungal growth and the antifungal drug effect. Results: the modal MICs for P.boydii and S. apiospermum isolates ranged from 0.03-0.25mg/L for both EUCAST and CLSI methodologies. In static models a dose-dependent decline in antigen concentrations was observed. Concentrations of approximately 0.4mg/L were sufficient to suppress growth of all species. Exposure response relationships were observed in the dynamic model for isolates of S. apiospermum. The trough concentrations associated with near maximal suppression of growth were approximately 0.2mg/L. Average area under curve: MIC values in the range of approximately 6-11 and 11-22 were associated with complete suppression of growth for EUCAST and CLSI methodologies respectively. Conclusions: F901318 demonstrates potent in vitro activity against isolates of P. boydii and S. apiospermum. These results may be bridged to humans in order to inform dosage recommendations for the potential treatment of patients with these infections.
Abstract.
Alcantara C, Thornton CR, Perez de-Luque A, Le Cocq K, Pedraza V, Murray PJ (2015). The Free-Living Rhizosphere Fungus Trichoderma hamatum GD12 Enhances Clover Productivity in Clover-Ryegrass Mixtures.
Plant and Soil: international journal on plant-soil relationships,
398, 165-180.
Abstract:
The Free-Living Rhizosphere Fungus Trichoderma hamatum GD12 Enhances Clover Productivity in Clover-Ryegrass Mixtures
Aim: a principal goal of grassland management is to
minimize the use of artificial fertilizers by maximising
the productivity of nitrogen-fixing leguminous plants
such as clovers. The objective of this study was to
investigate whether a plant-growth-promoting strain of
the free-living rhizosphere fungus Trichoderma
hamatum (GD12) could be used as a natural and sustainable
means of enhancing the competitiveness of
white clover (Trifolium repens) while allowing increased
productivity of both clover and ryegrass
(Lolium perenne) in mixed species systems.
Methods: an assay was conducted in rhizotrons with
white clover and ryegrass sown alone and in mixture
and in soils inoculated and non-inoculated with GD12.
Plant height, growing rate, phenological stage, number
of Rhizobium nodules and biomass were assessed. A
histological study of Rhizobium nodules and a stable
isotopes analysis was conducted to determine the N
fixation capacity of white clover.
Results: When introduced as a soil inoculant, the fungus
increased biomass production of both plant species and
shortened their phenological cycles. Furthermore, in
clover, GD12 enhanced plant height and growth rate
and stimulated Rhizobium nodulation, while 15N stable
isotope analysis demonstrated increased N2 fixation.
Conclusion: This shows that soil amendment with a
beneficial strain of saprotrophic fungus bestows a competitive
advantage to white clover in clover-ryegrass
mixtures and provides a sustainable mechanism for
improving the mixture productivity.
Abstract.
Shah A, Abdolrasouli A, Soresi S, Herbst S, Reed A, Carby M, Thornton CR, Drumright L, Shaunak S, Armstrong-James D, et al (2015). The Utility of Novel Multi-Stage Testing for the Diagnosis of Pulmonary Aspergillosis in a Cohort of Lung Transplant Recipients.
Author URL.
Solouk D, Bergmueller E, Weski J, Rolle A-M, Wiehr S, Thornton C, Gunzer M, Hasenberg M, Solouk D (2015). Towards the establishment of intravital mouse lung imaging via two Photon Microscopy for the design of a novel antibody-based aspergillosis detection system and the observation of pathogen-immune cell interactions.
MYCOSES,
58, 56-56.
Author URL.
Sharpe RA, Thornton CR, Tyrell J, Nikolaou V, Osborne NJ (2015). Variable risk of atopic disease due to indoor fungal exposure in NHANES 2005-2006. Clinical and Experimental Allergy, 45(10), 1566-1578.
Bergmueller E, Solouk D, Weski J, Thornton C, Gunzer M, Hasenberg M (2015). Where do spores go? Characterization of fungal distribution patterns among different murine Aspergillus fumigatus infection models using light sheet microscopy.
MYCOSES,
58, 9-9.
Author URL.
2014
Hoenigl M, Prattes J, Eigl S, Lass-Flörl C, Willinger B, Reischies F, Posch V, Lackner M, Flick H, Hönigl K, et al (2014). 1462Bronchoalveolar Lavage Lateral-Flow Device Test for Diagnosing Invasive Pulmonary Aspergillosis in ICU patients: a multicenter study. Open Forum Infectious Diseases, 1(Suppl 1), s385-s386.
Thornton CR (2014). Breaking the Mould - Novel Diagnostic and Therapeutic Strategies for Invasive Pulmonary Aspergillosis in the Immune Deficient Patient. Expert Review of Clinical Immunology, 6(10), 771-780.
Hoenigl M, Thornton C, Duettmann W, Posch V, Seeber K, Krause R, Lackner M, Lass-Florl C (2014). Bronchoalveolar Lavage Lateral-Flow Device Test for Diagnosing Invasive Pulmonary Aspergillosis in Pulmonology and Icu Patients.
Author URL.
Hoenigl M, Thornton CR, Duettmann W, Posch V, Seeber K, Krause R, Lackner M, Lass-Florl C (2014). Bronchoalveolar Lavage Lateral-Flow Device Test for Diagnosing Invasive Pulmonary Aspergillosis in Pulmonology and ICU Patients. American Journal of Respiratory and Critical Care Medicine, 189
Sereinigg M, Lass-Floerl C, Willinger B, Prattes J, Posch V, Selitsch B, Lackner M, Eschertzhuber S, Drescher M, Koidl C, et al (2014). Bronchoalveolar Lavage Lateral-Flow Device Test for Invasive Pulmonary Aspergillosis in Solid Organ Transplant Patients: a Semi-Prospective Multicenter Study.
Author URL.
Sereinigg M, Lass-Floerl C, Willinger B, Prattes J, Posch V, Selitsch B, Lackner M, Eschertzhuber S, Drescher M, Koidl C, et al (2014). Bronchoalveolar Lavage Lateral-Flow Device Test for Invasive Pulmonary Aspergillosis in Solid Organ Transplant Patients: a Semi-Prospective Multicenter Study.
Author URL.
Hoenigl M, Prattes J, Eigl S, Lass-Florl C, Willinger B, Reiches F, Posch V, Lackner M, Flick H, Hoenigl K, et al (2014). Bronchoalveolar lavage lateral-flow device test for diagnosing invasive pulmonary aspergillosis in ICU patients: a multicenter study. IDWeek2014. 8th - 12th Oct 2014.
Willinger B, Lackner M, Lass-Florl C, Prattes J, Posch V, Selitsch B, Eschertzhuber S, Hoenigl K, Koidl C, Sereinigg M, et al (2014). Bronchoalveolar lavage lateral-flow device test for invasive pulmonary aspergillosis in solid organ transplant patients: a semi-prospective multicenter study. Transplantation, 98(8), 898-902.
Ghamrawi S, Renier G, Saulnier P, Cuenot S, Zykwinska A, Dutilh BE, Thornton CR, Faure S, Bouchara J-P (2014). Cell wall modifications during conidial maturation of the human pathogenic fungus Pseudallescheria boydii. PLoS One, 9(6).
Miceli M, Goggins MI, Chander P, Sekaran AK, Kizy AE, Samuel L, Jiang H, Thornton CR, Ramesh M, Alangeden G, et al (2014). Diagnostic performance of lateral flow device and galactomannan for the detection of Aspergillus in bronchoalveolar lavage of patients at high risk for. invasive aspergillosis. 18th Symposium on Infections in the Immunocompromised Host. 15th - 17th Jun 2014.
Prattes J, Prueller F, Koidl C, Raggam R, Zollner-Schwetz I, Eigl S, Posch V, Hoenigl K, Duettmann W, Flick H, et al (2014). Diagnostics for invasive Pulmonary Aspergillosis in Bronchoalveolar Lavage Fluid of Patients with Underlying Pulmonary Diseases: Comparison of Aspergillus Lateral Flow Device, Galactomannan-Antigen-Test, Beta-D-Glucan-Tests and Conventional Culture.
MYCOSES,
57, 15-16.
Author URL.
Davies GE, Thornton CR (2014). Differentiation of the emerging human pathogens Trichosporon asahii and Trichosporon asteroides from other pathogenic yeasts and moulds by using species-specific monoclonal antibodies. PLoS One, 9(1).
Vandeputte P, Ghamrawi S, Rechenmann M, Iltis A, Giraud S, Fleury M, Thornton CR, Delhaes L, Meyer W, Papon N, et al (2014). Draft genome sequence of the pathogenic fungus Scedosporium apiospermum. Genome Announcements, 2(5).
Sharpe R, Thornton CR, Osborne NJ (2014). Modifiable Factors Governing Indoor Fungal Diversity and Risk of Asthma. Clinical and Experimental Allergy, 44, 631-641.
Ghamrawi S, Gilles R, Saulnier P, Cuenot S, Zykwinska A, Bas ED, Thornton C, Faure S, Bouchara J-P (2014). Modifications de la paroi au cours de la maturation des conidies chez Pseudallescheria boydii. Journal de Mycologie Médicale, 24(3).
Prattes J, Prueller F, Koidl C, Raggam RB, Palfner M, Eigl S, Flick H, Buzina W, Zollner-Schwetz I, Thornton CR, et al (2014). Novel Tests for Diagnosis of Invasive Pulmonary Aspergillosis in Patients with Underlying Respiratory Diseases. American Journal of Respiratory and Critical Care Medicine, 190(8), 922-929.
Hoenigl M, Prattes J, Spiess B, Wagner J, Prueller F, Raggam RB, Posch V, Duettmann W, Hoenigl K, Wofler A, et al (2014). Performance of Galactomannan, Beta-D-Glucan,
Aspergillus Lateral-Flow Device, Conventional Culture and PCR tests for Diagnosis of Invasive Pulmonary Aspergillosis in Bronchoalveolar Lavage Fluid. Journal of Clinical Microbiology, 52, 2039-2045.
Kobayashi M, Thornton CR (2014). Serological-diagnosis of chronic pulmonary aspergillosis (CPA) and Aspergillus lateral-flow device (LFD). European Respiratory Journal, 44, 2501-2501.
Al-Maqtoofi MY, Thornton CR (2014). Tracking the human pathogenic fungus Fusarium by using a highly specific monoclonal antibody.
IMMUNOLOGY,
143, 169-169.
Author URL.
2013
White PL, Parr C, Thornton CR, Barnes RA (2013). An Evaluation of real-time PCR, Galactomannan ELISA and a novel Lateral-Flow Device for the diagnosis of invasive aspergillosis. Journal of Clinical Microbiology, 51(5), 1510-1516.
Held J, Schmidt T, Thornton CR, Kotter E, Bertz H (2013). Comparison of a Novel Aspergillus Lateral-Flow Device and the Platelia Galactomannan Assay for Diagnosis of Invasive Aspergillosis following Haematopoietic Stem Cell Transplantation. Infection, 41(6), 1163-1169.
Held J, Schmidt T, Thornton CR, Kotter E, Bertz H (2013). Comparison of a novel Aspergillus Antigen Lateral Flow Device and the Platelia (R) Aspergillus Antigen ELISA in patients after hematopoietic stem cell transplantation.
MYCOSES,
56, 18-18.
Author URL.
Wiederhold NP, Najvar LK, Bocanegra RA, Kirkpatrick WR, Patterson TF, Thornton CR (2013). Inter-Laboratory and Inter-Study Reproducibility of a Novel Lateral-Flow Device and the Influence of Antifungal Therapy on the Detection of Invasive Pulmonary Aspergillosis.
Journal of Clinical Microbiology,
51, 459-465.
Abstract:
Inter-Laboratory and Inter-Study Reproducibility of a Novel Lateral-Flow Device and the Influence of Antifungal Therapy on the Detection of Invasive Pulmonary Aspergillosis
Lateral-flow devices (LFD) have gained interest as potential point-of-care assays for the diagnosis of infectious diseases. Our objective was to evaluate the inter-laboratory and inter-study reproducibility and the effects of antifungal therapy on a LFD developed for invasive pulmonary aspergillosis (IPA) detection. An established neutropenic guinea pig model of IPA caused by Aspergillus fumigatus was used. At predetermined time points (1 hr, 3, 5, and 7 days post-inoculation) blood and BAL fluid were collected from infected and uninfected animals. In a separate experiment, guinea pigs were treated with posaconazole (10 mg/kg PO BID), voriconazole (10 mg/kg PO BID), liposomal amphotericin B (10 mg/kg IP QD), or caspofungin (2 mg/kg IP QD), and samples were collected on days 7 and 11. Each laboratory independently evaluated the IgG monoclonal antibody-based LFD. Galactomannan and (1→3)-β-D-glucan were also measured using commercially available kits. Good inter-laboratory agreement was observed with the LFD as 96.7 % (29/30) of the serum and 73.3% (22/30) of the BAL samples from infected animals were in agreement. Good inter-study agreement was also observed. The serum sensitivity of each surrogate marker assay was reduced in animals treated with antifungals. In contrast, these markers remained elevated within the BAL fluids of treated animals, which was consistent with the fungal burden and histopathology results. These results demonstrate that the LFD assay is reproducible between different laboratories and studies. However, the sensitivity of this assay and other markers of IPA may be reduced within the serum in the presence of antifungal therapy
Abstract.
Studholme D, Harris B, Le Cocq K, Winsbury R, Perera V, Ryder L, Beale M, Ward J, Thornton CR, Grant M, et al (2013). Investigating the Beneficial Traits of Trichoderma hamatum GD12 for Sustainable Agriculture - Insights from Genomics.
Frontiers in Plant-Microbe Interactions,
4, 1-13.
Abstract:
Investigating the Beneficial Traits of Trichoderma hamatum GD12 for Sustainable Agriculture - Insights from Genomics
Trichoderma hamatum strain GD12 is unique in that it can promote plant growth, activate biocontrol against pre- and post-emergence soil pathogens and can induce systemic resistance to foliar pathogens. This study extends previous work in lettuce to demonstrate that GD12 can confer beneficial agronomic traits to other plants, providing examples of plant growth promotion in the model dicot, Arabidopsis thaliana and induced foliar resistance to Magnaporthe oryzae in the model monocot rice. We further characterize the lettuce-T. hamatum interaction to show that bran extracts from GD12 and a N-acetyl-β-D-glucosamindase-deficient mutant differentially promote growth in a concentration dependent manner, and these differences correlate with differences in the small molecule secretome. We show that GD12 mycoparasitises a range of isolates of the pre-emergent soil pathogen Sclerotinia sclerotiorum and that this interaction induces a further increase in plant growth promotion above that conferred by GD12. To understand the genetic potential encoded by T. hamatum GD12 and to facilitate its use as a model beneficial to study plant growth promotion, induced systemic resistance and mycoparasitism we present de novo genome sequence data. We compare GD12 with other published Trichoderma genomes and show that T. hamatum GD12 contains unique genomic regions with the potential to encode novel bioactive metabolites that may contribute to GD12’s agrochemically important traits.
Abstract.
Thornton CR (2013). Lateral-Flow Device for Diagnosis of Fungal Infection. Current Fungal Infection Reports, 7(3), 244-251.
Ryder, LS, Mentlak TA, Dagdas YF, Kershaw MJ, Thornton CR, Chen J, Wang Z, Talbot, NJ (2013). NADPH oxidases regulate septin-mediated cytoskeletal re-modeling during plant infection by the rice blast fungus. Proceedings of the National Academy of Sciences of USA, 110, 3179-3184.
Johnson GL, Shannon M, Thornton CR, Agrawal SG, Bustin SA (2013). Proximity ligation assay for the sensitive, specific and early diagnosis of invasive fungal disease.
MYCOSES,
56, 48-49.
Author URL.
Wiederhold NP, Thornton CR (2013). Reply to “Interlaboratory and Interstudy Reproducibility of a Novel Lateral-Flow Device: a Statistical Issue”. Journal of Clinical Microbiology, 51(5).
2012
Hoenigl M, Koidl C, Duettmann W, Seeber K, Wagner J, Wölfler A, Raggam RB, Thornton CR, Krause R (2012). Bronchoalveolar lavage lateral-flow device test for invasive pulmonary aspergillosis diagnosis in haematological malignancy and solid organ transplant patients.
Journal of Infection,
65(6), 588-591.
Abstract:
Bronchoalveolar lavage lateral-flow device test for invasive pulmonary aspergillosis diagnosis in haematological malignancy and solid organ transplant patients
Due to the crude mortality of 80-90% in absence of adequate treatment, timely diagnosis and early start of antifungal therapy are key factors in the successful treatment of invasive pulmonary aspergillosis (IPA). Diagnosis of IPA, however, remains difficult as clinical signs and symptoms as well as radiological findings are often unspecific and conventional culture methods lack sensitivity. In recent years antigen testing has therefore become one of the cornerstones of IFI diagnostics, even though tests are limited by varying turnaround time as well as sensitivity and specificity issues. These limitations may be overcome by the Lateral-Flow Device (LFD) test, a single sample point-of-care test that is based on the detection of an Aspergillus extracellular glycoprotein antigen by monoclonal antibody JF5. This retrospective study evaluates the LFD test by using bronchoalveolar lavage (BAL) samples from patients with haematological malignancies and patients after solid organ transplantation (SOT).
Thirty-nine BAL samples from 37 patients were included (29 samples haematological malignancies and 10 samples SOT; 12 probable IPA, 9 possible IPA, 16 no IPA). Sensitivity and specificity of BAL LFD test for probable IPA were 100% and 81% respectively. GM levels in cases with negative LFD were significantly lower than in patients with positive LFD (P
Abstract.
Thornton CR, Johnson G, Agrawal S (2012). Detection of invasive pulmonary aspergillosis in haematological malignancy patients by using lateral-flow technology. Journal of Visualized Experiments, 61
Romão-Dumaresq AS, de Araújo WL, Talbot NJ, Thornton CR (2012). RNA interference of endochitinases in the sugarcane endophyte Trichoderma virens 223 reduces its fitness as a biocontrol agent of pineapple disease.
PLoS One,
7(10).
Abstract:
RNA interference of endochitinases in the sugarcane endophyte Trichoderma virens 223 reduces its fitness as a biocontrol agent of pineapple disease
The sugarcane root endophyte Trichoderma virens 223 holds enormous potential as a sustainable alternative to chemical pesticides in the control of sugarcane diseases. Its efficacy as a biocontrol agent is thought to be associated with its production of chitinase enzymes, including N-acetyl-β-D-glucosaminidases, chitobiosidases and endochitinases. We used targeted gene deletion and RNA-dependent gene silencing strategies to disrupt N-acetyl-β-D-glucosaminidase and endochitinase activities of the fungus, and to determine their roles in the biocontrol of soil-borne plant pathogens. The loss of N-acetyl-β-D-glucosaminidase activities was dispensable for biocontrol of the plurivorous damping-off pathogens Rhizoctonia solani and Sclerotinia sclerotiorum, and of the sugarcane pathogen Ceratocystis paradoxa, the causal agent of pineapple disease. Similarly, suppression of endochitinase activities had no effect on R. solani and S. sclerotiorum disease control, but had a pronounced effect on the ability of T. virens 223 to control pineapple disease. Our work demonstrates a critical requirement for T. virens 223 endochitinase activity in the biocontrol of C. paradoxa sugarcane disease, but not for general antagonism of other soil pathogens. This may reflect its lifestyle as a sugarcane root endophyte.
Abstract.
Ryder LS, Harris BD, Soanes DM, Kershaw MJ, Talbot NJ, Thornton CR (2012). Saprotrophic competitiveness and biocontrol fitness of a genetically modified strain of the plant-growth-promoting fungus Trichoderma hamatum GD12. Microbiology, 158(Special issue of Trichoderma: from basic biology to biotechnology), 84-97.
Ryder LS, Harris BD, Soanes DM, Kershaw MJ, Talbot NJ, Thornton CR (2012). Saprotrophic competitiveness and biocontrol fitness of a genetically modified strain of the plant-growth-promoting fungus Trichoderma hamatum GD12.
Microbiology (Reading),
158(Pt 1), 84-97.
Abstract:
Saprotrophic competitiveness and biocontrol fitness of a genetically modified strain of the plant-growth-promoting fungus Trichoderma hamatum GD12.
Trichoderma species are ubiquitous soil fungi that hold enormous potential for the development of credible alternatives to agrochemicals and synthetic fertilizers in sustainable crop production. In this paper, we show that substantial improvements in plant productivity can be met by genetic modification of a plant-growth-promoting and biocontrol strain of Trichoderma hamatum, but that these improvements are obtained in the absence of disease pressure only. Using a quantitative monoclonal antibody-based ELISA, we show that an N-acetyl-β-d-glucosaminidase-deficient mutant of T. hamatum, generated by insertional mutagenesis of the corresponding gene, has impaired saprotrophic competitiveness during antagonistic interactions with Rhizoctonia solani in soil. Furthermore, its fitness as a biocontrol agent of the pre-emergence damping-off pathogen Sclerotinia sclerotiorum is significantly reduced, and its ability to promote plant growth is constrained by the presence of both pathogens. This work shows that while gains in T. hamatum-mediated plant-growth-promotion can be met through genetic manipulation of a single beneficial trait, such a modification has negative impacts on other aspects of its biology and ecology that contribute to its success as a saprotrophic competitor and antagonist of soil-borne pathogens. The work has important implications for fungal morphogenesis, demonstrating a clear link between hyphal architecture and secretory potential. Furthermore, it highlights the need for a holistic approach to the development of genetically modified Trichoderma strains for use as crop stimulants and biocontrol agents in plant agriculture.
Abstract.
Author URL.
Thornton CR (2012). Serological techniques for diagnosis. In Lane C, Beales P, Hughes K (Eds.) Fungal Plant Pathogens, Principles and Protocols, CABI, 159-177.
2011
Johnson GL, Bibby D, Bustin B, Thornton CR, Agrawal S (2011). Detection of Aspergillus in broncho-alveolar lavage fluid using two biological assays; evidence of active infection?. Mycoses, 54, 130-131.
2010
Thornton CR (2010). Detection of Invasive Aspergillosis. Advances in Applied Microbiology, 70, 187-216.
Thornton CR, Slaughter DC, Davis RM (2010). Detection of the sour-rot pathogen Geotrichum candidum in tomato fruit and juice by using a highly specific monoclonal antibody-based ELISA. International Journal of Food Microbiology, 143(3), 166-172.
Thornton CR (2010). Environmental fungi as a cause of human mycoses. Culture, 31(2), 1-5.
Thornton CR (2010). MONOCLONAL ANTIBODY-BASED ELISA DETECTION OF SOUR ROT PATHOGEN IN TOMATO JUICE.
2009
Wiederhold NP, Thornton CR, Najvar LK, Kirkpatrick WR, Bocanegra R, Patterson TF (2009). Comparison of Lateral Flow Technology and Galactomannan and (1→3)-β-D-Glucan Assays for Detection of Invasive Pulmonary Aspergillosis. Clinical and Vaccine Immunology, 16(12), 1844-1846.
McDonagh L, Thornton C, Wallman JF, Stevens JR (2009). Development of an antigen-based rapid diagnostic test for the identification of blowfly (Calliphoridae) species of forensic significance.
Abstract:
Development of an antigen-based rapid diagnostic test for the identification of blowfly (Calliphoridae) species of forensic significance.
Abstract.
Author URL.
Thornton CR (2009). New Method of Detecting and Diagnosing Invasive Aspergillosis.
Richards TA, Soanes DM, Foster PG, Leonard G, Thornton CR, Talbot NJ (2009). Phylogenomic analysis demonstrates a pattern of rare and ancient horizontal gene transfer between plants and fungi. The Plant Cell, 21, 1897-1911.
Thornton CR (2009). Production of monoclonal antibodies to plant pathogens. In (Ed)
, 63-74.
Abstract:
Production of monoclonal antibodies to plant pathogens.
Abstract.
Author URL.
Thornton CR (2009). Tracking the emerging human pathogen Pseudallescheria boydii by using highly specific monoclonal antibodies.
Clinical and Vaccine Immunology,
16(5), 756-764.
Abstract:
Tracking the emerging human pathogen Pseudallescheria boydii by using highly specific monoclonal antibodies
Pseudallescheria boydii has long been known to cause white grain mycetoma in immunocompetent humans, but it has recently emerged as an opportunistic pathogen of humans, causing potentially fatal invasive infections in immunocompromised individuals and evacuees of natural disasters, such as tsunamis and hurricanes. The diagnosis of P. boydii is problematic since it exhibits morphological characteristics similar to those of other hyaline fungi that cause infectious diseases, such as Aspergillus fumigatus and Scedosporium prolificans. This paper describes the development of immunoglobulin M (IgM) and IgG1 kappa-light chain monoclonal antibodies (MAbs) specific to P. boydii and certain closely related fungi. The MAbs bind to an immunodominant carbohydrate epitope on an extracellular 120-kDa antigen present in the spore and hyphal cell walls of P. boydii and Scedosporium apiospermum. The MAbs do not react with S. prolificans, Scedosporium dehoogii, or a large number of clinically relevant fungi, including A. fumigatus, Candida albicans, Cryptococcus neoformans, Fusarium solani, and Rhizopus oryzae. The MAbs were used in immunofluorescence and double-antibody sandwich enzyme-linked immunosorbent assays (DAS-ELISAs) to accurately differentiate P. boydii from other infectious fungi and to track the pathogen in environmental samples. Specificity of the DAS-ELISA was confirmed by sequencing of the internally transcribed spacer 1 (ITS1)-5.8S-ITS2 rRNA-encoding regions of environmental isolates.
Abstract.
Thornton CR (2009). Use of monoclonal antibody-based diagnostic assays for the detection of invasive mycoses. Microbiologist, 10(2), 49-50.
2008
Thornton CR (2008). Development of an immunochromatographic lateral-flow device for rapid serodiagnosis of invasive aspergillosis.
Clinical and Vaccine Immunology,
15(7), 1095-1105.
Abstract:
Development of an immunochromatographic lateral-flow device for rapid serodiagnosis of invasive aspergillosis
Aspergillus fumigatus is a cosmopolitan saprotrophic fungus that is second only to Candida species as a cause of invasive fungal infections in immunocompromised humans. Current immunodiagnostic tests for invasive aspergillosis (IA) are based on the detection of circulating galactomannan ( GM) in a patient's serum by using a rat monoclonal antibody (MAb), EB-A2, that binds to tetra (1 -> 5)-beta-D-galactofuranoside, the immunodominant epitope in GM. The potential cross-reactivity of MAb EB-A2 with non-Aspergillus fungi, with contaminating GM in beta-lactam antibiotics and foodstuffs, and with bacterial lipoteichoic acids has prompted efforts to discover non-GM antigens that can act as surrogate markers for the diagnosis of IA. This paper describes the development of a mouse MAb, JF5, that binds to a protein epitope present on an extracellular glycoprotein antigen secreted constitutively during the active growth of A. fumigatus. The MAb was used to develop an immunochromatographic lateral-flow device (LFD) for the rapid (15-min) detection of Aspergillus antigens in human serum. The test is highly specific, reacting with antigens from Aspergillus species but not with antigens from a large number of clinically important fungi, including Candida species, Cryptococcus neoformans, Fusarium solani, Penicillium marneffei, Pseudallescheria boydii, and Rhizopus oryzae. The LFD was able to detect circulating antigen in serum samples from patients suspected of having or shown to have IA on the basis of their clinical symptoms and results from tests for GM and fungal (1 -> 3)-beta-D-glucan. The ease of use of the LFD provides a diagnostic platform for the routine testing of vulnerable patients who have an elevated risk of IA.
Abstract.
Waller PL, Thornton CR, Farley D, Groenhof A (2008). Pathogens and other fungi in growing media constituents.
Abstract:
Pathogens and other fungi in growing media constituents
Abstract.
Thornton, C.R. (2008). Tracking fungi in soil with monoclonal antibodies. European Journal of Plant Pathology, 121, 347-353.
Thornton CR (2008). Tracking fungi in soil with monoclonal antibodies.
Abstract:
Tracking fungi in soil with monoclonal antibodies
Abstract.
2007
Thornton CR (2007). Tracking fungi in soil with monoclonal antibodies. In (Ed) Sustainable disease management in a European context, 347-353.
2006
Gilbert MJ, Thornton CR, Wakley GE, Talbot NJ (2006). A P-type ATPase required for rice blast disease and induction of host resistance. Nature, 440(7083), 535-539.
Richards TA, Dacks JB, Jenkinson JM, Thornton CR, Talbot NJ (2006). Evolution of filamentous plant pathogens: gene exchange across eukaryotic kingdoms. Current Biology, 16(18), 1857-1864.
Thornton CR, Talbot, N.J. (2006). Immunofluorescence microscopy and immunogold EM for investigating fungal infection of plants. Nature Protocols, 1, 2506-2511.
2005
Kershaw, M.J. Wakley, G.E. Talbot, N.J. Thornton CR (2005). Four conserved intra-molecular disulfide linkages are required for secretion and cell wall localization of a hydrophobin during fungal morphogenesis. Molecular Microbiology, 56(1), 117-125.
Thornton CR (2005). Use of monoclonal antibodies to quantify the dynamics of α-galactosidase and endo-1,4-β-glucanase production by Trichoderma hamatum during saprotrophic growth and sporulation in peat. Environmental Microbiology, 7(5), 737-749.
2004
Thornton, CR, Groenhof AC, Forrest, R & Lamotte, R (2004). A One-Step, Immunochromatographic Lateral Flow Device Specific to Rhizoctonia solani and Certain Related Species, and its use to Detect and Quantify R. solani in Soil. Phytopathology, 94, 280-288.
Tucker SL, Thornton CR, Tasker K, Jacob C, Giles G, Egan M, Talbot NJ (2004). A fungal metallothionein is required for pathogenicity of Magnaporthe grisea. The Plant Cell, 16(6), 1575-1588.
Thornton CR (2004). An immunological approach to quantifying the saprotrophic growth dynamics of Trichoderma species during antagonistic interactions with Rhizoctonia solani in a soil-less mix. Environmental Microbiology, 6(4), 323-334.
2003
Wang ZY, Thornton CR, Kershaw MJ, Debao L, Talbot NJ (2003). The glyoxylate cycle is required for temporal regulation of virulence by the plant pathogenic fungus Magnaporthe grisea. Molecular Microbiology, 47(6), 1601-1612.
2002
Thornton CR, Pitt, D. Wakley, G.E. Talbot, N.J. (2002). Production of a monoclonal antibody specific to the genus Trichoderma and closely related fungi, and its use to detect Trichoderma spp. in naturally infested composts. Microbiology, 148, 1263-1279.
2001
Bailey DJ, Thornton CR, Dewey FM, Gilligan CA (2001). A non-destructive immunoblotting technique for visualisation and analysis of the growth dynamics of Rhizoctonia solani.
Mycological Research,
105(8), 983-990.
Abstract:
A non-destructive immunoblotting technique for visualisation and analysis of the growth dynamics of Rhizoctonia solani
Immunoblotting combined with computer imaging and a simple, non-linear mathematical model were used to demonstrate the potential of a technique for non-destructive visualisation and analysis of fungal growth of Rhizoctonia solani over the surface of non-sterile sand. Immunoblotting detected actively growing regions of mycelium enabling visualisation of individual hyphae at the colony edge. A zone of active growth was detected expanding radially over time. Active growth did not continue in the centre of the fungal colony leading to the development of a ring of mycelium surrounding the inoculum. Change in the density of actively growing mycelium with distance from the inoculum unit was summarised for each colony at each time by a Gaussian function, describing a wave of actively growing mycelium, symmetrical in density about its centre but differing amongst replicate colonies. The effectiveness of the immunoblotting technique to detect differences in colony growth was tested by comparing the growth of replicate colonies for two contrasting isolates of R. solani. When both isolates of R. solani were grown at 23 °C the amplitude of the wave increased to a maximum and then decayed over time, the location of the centre of the wave moved outwards at a constant rate, whilst the width of the wave increased. Increasing the temperature to 28 °, accelerated this intrinsic growth process for one isolate, but retarded growth of the other.
Abstract.
Thornton, C.R. (2001). Immunological methods for fungi. In Talbot NJ (Ed) Molecular and Cell Biology of Filamentous Fungi: a Practical Approach, UK: Oxford University Press, 227-257.
1999
Thornton CR, O'Neill TM, Hilton G, Gilligan CA (1999). Detection and recovery of Rhizoctonia solani in naturally infested glasshouse soils using a combined baiting, double monoclonal antibody ELISA.
Plant Pathology,
48(5), 627-634.
Author URL.
Thornton CR, Gilligan CA (1999). Quantification of the effect of the hyperparasite Trichoderma harzianum on the saprotrophic growth dynamics of Rhizoctonia solani in compost using a monoclonal antibody-based ELISA.
Mycological Research,
103, 443-448.
Author URL.
1997
Thornton CR, Dewey FM, Gilligan CA (1997). Production and characterization of a monoclonal antibody raised against surface antigens from mycelium of Gaeumannomyces graminis var. tritici evidence for an extracellular polyphenol oxidase.
Phytopathology,
87(1), 123-131.
Abstract:
Production and characterization of a monoclonal antibody raised against surface antigens from mycelium of Gaeumannomyces graminis var. tritici evidence for an extracellular polyphenol oxidase
A murine monoclonal antibody (MAb) of immunoglobulin class M (IgM) was raised against surface antigens from Gaeumannomyces graminis var. tritici and, by enzyme-linked immunosorbent assay, recognized isolates of G. graminis var. tritici, G. graminis var. avenae and G. graminis var. graminis. Characterization of the antigen by heat and protease treatments showed that the epitope recognized by the MAb was a protein. Antigen production was detected only in live mycelia. Immunofluorescence studies showed that the antigen was associated with both the broad melanized macrohyphae and hyaline mycelia of G. graminis var. tritici. Secretion of antigen into an aqueous minimal medium was promoted only by exposure of live mycelia to certain pbenolic substrates, including monophenols ortho-, para-, and meta-cresol; 3,4,5-trihydroxybenzoic acid (gallic acid); and phenolic amino acid L-3- (3,4-dihydroxyphenyl) alanine (L-DOPA). Antigen secretion was not promoted by 3-(4-hydroxyphenyl) alanine (L-tyrosine). The MAb reacted strongly with purified enzyme laccase (polyphenol oxidase, EC 1.10.3.2) but did not recognize purified tyrosinase (monophenol oxidase, EC 1.14.18.1). Moreover, chemicals that bind to copper and inhibit copper-containing enzymes such as laccase completely inhibited antigen secretion in response to L-DOPA. The MAb was tested for specificity against a wide range of fungi, common yeast species, and grain positive and negative bacteria. It did not recognize antigens from a broad range of unrelated fungi, including Gliocladium roseum, Fusarium sp. Phoma exigua, Phialophora fastigiata, Penicillium crustosum, Pythium ultimum, Rhizopus stolonifer, Rhizoctonia carotae, R. oryzae, R. tuliparum, and Trichoderma viride, nor did it recognize surface antigens from yeasts or bacteria. The MAb cross-reacted with antigens from Botrytis spp. Chaetomium globosum, R. cerealis, and R. solani. However, secretion of antigen by R. solani and R. cerealis was not promoted by L-DOPA, and secretion by C. globosum in response to the phenolic amino acid was significantly less compared to G. graminis var. tritici.
Abstract.
Otten W, Gilligan CA, Thornton CR (1997). Quantification of fungal antigens in soil with a monoclonal antibody-based ELISA: Analysis and reduction of soil-specific bias.
Phytopathology,
87(7), 730-736.
Abstract:
Quantification of fungal antigens in soil with a monoclonal antibody-based ELISA: Analysis and reduction of soil-specific bias
This paper describes methods to improve the use of immunoassays for quantification of soilborne fungal antigens. Calibration curves, prepared by diluting known quantities of an antigen into soil extracts and into soil, were described by a four-parameter logistic curve from which two principal criteria, the lower detection limit and the horizontal locational parameter, were used to summarize the sensitivity and bias of an immunoassay. We identify two sources of bias, retention of the antigen in soil due to bonding and interference of soluble soil components in plate-trapped-antigen, enzyme-linked immunosorbent assays. Using a monoclonalantibody that recognizes a putative catechol oxidase secreted by hyphae of Rhizoctonia solani, we show that bias due to retention of the antigen in soil is substantially greater than bias due to interference. Three soils were compared: a sand, a clay, and a loam. The degree of retention varied with soil type, with more than a 1,000-fold reduction in sensitivity in the clay soil. Addition of CuSO4 to the extraction solution and optimizing the volume of extractant reduced the bias and increased the sensitivity of the assay for all three soils. Possible mechanisms for the effect due to Cu2+ and the implications for the design and use of calibration curves for assays involving quantification of fungal antigens in soil are discussed.
Abstract.
Dewey FM, Thornton CR, Gilligan CA (1997). Use of Monoclonal Antibodies to Detect, Quantify and Visualize Fungi in Soils. Advances in Botanical Research, 24(C), 275-308.
1996
Thornton CR (1996). Detection and quantification of Rhizoctonia solani in soil by monoclonal antibody-based immuno-magnetic bead assay.
Soil Biology and Biochemistry,
28(4-5), 527-532.
Abstract:
Detection and quantification of Rhizoctonia solani in soil by monoclonal antibody-based immuno-magnetic bead assay
Murine monoclonal antibodies (MAbs) of the immunoglobulin classes IgM and IgA, from two different hybridoma cell lines, and rabbit (polyclonal) antiserum raised against mycelial antigens from an AG4 isolate of R. solani were used to develop magnetic microsphere-enzyme immunoassays (MM-EIAs) for the detection and quantification of the pathogen in soil. The assay can also be used to differentiate R. solani anastomosis group (AG) 2-2 isolates from other AG isolates grown in vitro. Mycelial antigens were solubilised in saline buffer containing detergent and following high speed centrifugation soluble antigen was delineated by the addition either of a mixture of specific murine MAbs of different immunoglobulin classes or a mixture of murine MAb supernatant and rabbit antiserum. Due to the low concentration of reactants a soluble immune complex was formed. The soluble complex was isolated by the addition of magnetic beads coated with antibodies that specifically recognised the murine IgM MAbs. The bound antibody antigen complex was then detected using a commercial antibody enzyme conjugate and chromogen.
Abstract.
Thornton CR, Dewey FM (1996). Detection of phialoconidia of Trichoderma harzianum in peat-bran by monoclonal antibody-based enzyme-linked immunosorbent assay.
Mycological Research,
100(2), 217-222.
Abstract:
Detection of phialoconidia of Trichoderma harzianum in peat-bran by monoclonal antibody-based enzyme-linked immunosorbent assay
Murine monoclonal antibodies (MAbs) were raised against phialoconidia of a Trichoderma harzianum isolate T95. A hybridoma cell line. GH5, was raised that produced MAbs of the immunoglobulin class IgM that recognized, by enzyme-linked immunosorbent assay (ELISA), antigens from conidia of T95. They did not react with antigens from mycelium or chlamydospores of this isolate but recognized antigens from conidia of a number of other T. harzianum and T. viride isolates. Heat, protease and periodate treatment of T95 conidial antigens showed that GH5 MAbs recognized a protein epitope that formed part of a larger glycoprotein molecule. GH5 MAbs were used to develop an ELISA for the detection of phialoconidia of T. harzianum in peat-bran.
Abstract.
1994
Thornton CR, Dewey FM, Gilligan CA (1994). Development of a monoclonal antibody-based enzyme-linked immunosorbent assay for the detection of live propagules of Trichoderma harzianum in a peat-bran medium.
Soil Biology and Biochemistry,
26(7), 909-920.
Abstract:
Development of a monoclonal antibody-based enzyme-linked immunosorbent assay for the detection of live propagules of Trichoderma harzianum in a peat-bran medium
Mouse monoclonal antibodies (MAbs) and mouse and rabbit (polyclonal) antisera were raised against a Trichoderma harzianum isolate T95. A cell line, Th.HD33, raised from splenocytes from a mouse immunized with lyophilized mycelium and Quil a adjuvant, produced MAbs of the immunoglobulin sub-class IgG2a that were specific recognizing, by ELISA, antigens from a number of Trichoderma species and species from the related genus Hypocrea but not from a range of other soil-borne fungi. These MAbs were used to develop an enzyme-linked immunosorbent assay (ELISA) for the detection of live propagules of Trichoderma harzianum in a peat-bran medium. Heat, protease and periodate treatment of the T95 antigen and binding on Western blots indicate that the Th.HD33 MAbs recognized a protein epitope of ca Mr 18,500 which forms part of a larger glycoprotein molecule. © 1994.
Abstract.
Thornton CR, Dewey FM, Gilligan CA (1994). Development of monoclonal antibody-based immunological assays for the detection of live propagules of Rhizoctonia solani in soil.
1993
THORNTON CR, DEWEY FM, GILLIGAN CA (1993). Development of monoclonal antibody‐based immunological assays for the detection of live propagules of Rhizoctonia solani in soil.
Plant Pathology,
42(5), 763-773.
Abstract:
Development of monoclonal antibody‐based immunological assays for the detection of live propagules of Rhizoctonia solani in soil
Mouse monoclonal antibodies (mAbs) and rabbit (polyclonal) antiserum were used to develop DIAGNOSTIC‐ELISA, double‐antibody‐sandwich‐ELISA (DAS‐ELISA), DIP‐STICK and immuno‐fluorescence colony staining immunoassays for the specific detection of Rhizoctonia solani in soil. mAbs were raised against an anastomosis group 4 isolate of R. solani. Mice were immunized using either phosphate‐buffered saline (PBS) suspensions of lyophilized mycelium plus Quil a adjuvant, or with a solubilized acetone precipitate prepared from cell‐free surface washings from solid slant cultures. Polyclonal antisera were raised in rabbits using PBS suspensions of lyophilized mycelium and Quil a adjuvant. Hybridoma supernatants and rabbit antisera were screened by ELISA. Four of the cell lines raised produced mAbs that were species‐specific. They recognized antigens from R. solani by ELISA and immunofluorescence, but not other related or unrelated species of soil‐borne fungi. The remaining cell line produced mAbs that cross‐reacted slightly, by ELISA, with antigens from R. cerealis. These mAbs did not recognize R. cerealis by immunofluorescence, or other related or unrelated soil‐borne fungi, by ELISA. Copyright © 1993, Wiley Blackwell. All rights reserved
Abstract.
1991
Thornton CR, Jarvis B, Cooke R (1991). A chitin assay for the enumeration of Plasmodiophora brassicae resting spores in clubroot tissue.
Mycological Research,
95, 879-882.
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