Publications by category
Journal articles
Masoli JAH, Jeffries A, Temperton B, Auckland C, Michelsen M, Warwick-Dugdale J, Manley R, Farbos A, Ellard S, Knight B, et al (In Press). Viral genetic sequencing identifies staff transmission of COVID-19 is important in a community hospital outbreak.
Abstract:
Viral genetic sequencing identifies staff transmission of COVID-19 is important in a community hospital outbreak
AbstractBackgroundWe have successfully used whole-genome sequencing to provide additional information for transmission pathways in infectious spread. We report and interpret genomic sequencing results in clinical context from a large outbreak of COVID-19 with 46 cases across staff and patients in a community hospital in the UK.MethodsFollowing multiple symptomatic cases within a two-week period, all staff and patients were screened by RT-PCR and staff subsequently had serology tests.ResultsThirty staff (25%) and 16 patients (62%) tested positive for COVID-19. Genomic sequencing data showed significant overlap of viral haplotypes in staff who had overlapping shift patterns. Patient haplotypes were more distinct from each other but had overlap with staff haplotypes.ConclusionsThis study includes clinical and genomic epidemiological detail that demonstrates the value of a combined approach. Viral genetic sequencing has identified that staff transmission of COVID-19 was important in this community hospital outbreak.Key pointsDetailed analysis of a large community hospital outbreak in older adults and staff with concurrent clinical and genomic data, including working patterns.Staff transmission was important in this community hospital outbreak.We found plausible associations between staff and patient cases.
Abstract.
Cotton S, McHugh MP, Dewar R, Haas JG, Templeton K, Consortium TCGU, Robson SC, Connor TR, Loman NJ, Golubchik T, et al (2023). Investigation of hospital discharge cases and SARS-CoV-2 introduction into Lothian care homes. Journal of Hospital Infection, 135, 28-36.
Kennedy NA, Janjua M, Chanchlani N, Lin S, Bewshea C, Nice R, McDonald TJ, Auckland C, Harries LW, Davies M, et al (2023). Vaccine escape, increased breakthrough and reinfection in infliximab-treated patients with IBD during the Omicron wave of the SARS-CoV-2 pandemic.
Gut,
72(2), 295-305.
Abstract:
Vaccine escape, increased breakthrough and reinfection in infliximab-treated patients with IBD during the Omicron wave of the SARS-CoV-2 pandemic.
OBJECTIVE: Antitumour necrosis factor (TNF) drugs impair serological responses following SARS-CoV-2 vaccination. We sought to assess if a third dose of a messenger RNA (mRNA)-based vaccine substantially boosted anti-SARS-CoV-2 antibody responses and protective immunity in infliximab-treated patients with IBD. DESIGN: Third dose vaccine induced anti-SARS-CoV-2 spike (anti-S) receptor-binding domain (RBD) antibody responses, breakthrough SARS-CoV-2 infection, reinfection and persistent oropharyngeal carriage in patients with IBD treated with infliximab were compared with a reference cohort treated with vedolizumab from the impaCt of bioLogic therApy on saRs-cov-2 Infection and immuniTY (CLARITY) IBD study. RESULTS: Geometric mean (SD) anti-S RBD antibody concentrations increased in both groups following a third dose of an mRNA-based vaccine. However, concentrations were lower in patients treated with infliximab than vedolizumab, irrespective of whether their first two primary vaccine doses were ChAdOx1 nCoV-19 (1856 U/mL (5.2) vs 10 728 U/mL (3.1), p
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Author URL.
Baker S, Dougan G, Hess C, Kingston N, Lehner PJ, Lyons PA, Matheson NJ, Owehand WH, Saunders C, Summers C, et al (2022). Author Correction: SARS-CoV-2 evolution during treatment of chronic infection. Nature, 608(7922), e23-e23.
Baker S, Dougan G, Hess C, Kingston N, Lehner PJ, Lyons PA, Matheson NJ, Owehand WH, Saunders C, Summers C, et al (2022). Author Correction: Sensitivity of SARS-CoV-2 B.1.1.7 to mRNA vaccine-elicited antibodies. Nature, 608(7922), e24-e24.
Kläser K, Molteni E, Graham M, Canas LS, Österdahl MF, Antonelli M, Chen L, Deng J, Murray B, Kerfoot E, et al (2022). COVID-19 due to the B.1.617.2 (Delta) variant compared to B.1.1.7 (Alpha) variant of SARS-CoV-2: a prospective observational cohort study.
Scientific Reports,
12(1).
Abstract:
COVID-19 due to the B.1.617.2 (Delta) variant compared to B.1.1.7 (Alpha) variant of SARS-CoV-2: a prospective observational cohort study
The Delta (B.1.617.2) variant was the predominant UK circulating SARS-CoV-2 strain between May and December 2021. How Delta infection compares with previous variants is unknown. This prospective observational cohort study assessed symptomatic adults participating in the app-based COVID Symptom Study who tested positive for SARS-CoV-2 from May 26 to July 1, 2021 (Delta overwhelmingly the predominant circulating UK variant), compared (1:1, age- and sex-matched) with individuals presenting from December 28, 2020 to May 6, 2021 (Alpha (B.1.1.7) the predominant variant). We assessed illness (symptoms, duration, presentation to hospital) during Alpha- and Delta-predominant timeframes; and transmission, reinfection, and vaccine effectiveness during the Delta-predominant period. 3581 individuals (aged 18 to 100 years) from each timeframe were assessed. The seven most frequent symptoms were common to both variants. Within the first 28 days of illness, some symptoms were more common with Delta versus Alpha infection (including fever, sore throat, and headache) and some vice versa (dyspnoea). Symptom burden in the first week was higher with Delta versus Alpha infection; however, the odds of any given symptom lasting ≥ 7 days was either lower or unchanged. Illness duration ≥ 28 days was lower with Delta versus Alpha infection, though unchanged in unvaccinated individuals. Hospitalisation for COVID-19 was unchanged. The Delta variant appeared more (1.49) transmissible than Alpha. Re-infections were low in all UK regions. Vaccination markedly reduced the risk of Delta infection (by 69-84%). We conclude that COVID-19 from Delta or Alpha infections is similar. The Delta variant is more transmissible than Alpha; however, current vaccines showed good efficacy against disease. This research framework can be useful for future comparisons with new emerging variants.
Abstract.
Aggarwal D, Page AJ, Schaefer U, Savva GM, Myers R, Volz E, Ellaby N, Platt S, Groves N, Gallagher E, et al (2022). Genomic assessment of quarantine measures to prevent SARS-CoV-2 importation and transmission. Nature Communications, 13(1).
Aggarwal D, Warne B, Jahun AS, Hamilton WL, Fieldman T, du Plessis L, Hill V, Blane B, Watkins E, Wright E, et al (2022). Genomic epidemiology of SARS-CoV-2 in a UK university identifies dynamics of transmission.
Nature Communications,
13(1).
Abstract:
Genomic epidemiology of SARS-CoV-2 in a UK university identifies dynamics of transmission
AbstractUnderstanding SARS-CoV-2 transmission in higher education settings is important to limit spread between students, and into at-risk populations. In this study, we sequenced 482 SARS-CoV-2 isolates from the University of Cambridge from 5 October to 6 December 2020. We perform a detailed phylogenetic comparison with 972 isolates from the surrounding community, complemented with epidemiological and contact tracing data, to determine transmission dynamics. We observe limited viral introductions into the university; the majority of student cases were linked to a single genetic cluster, likely following social gatherings at a venue outside the university. We identify considerable onward transmission associated with student accommodation and courses; this was effectively contained using local infection control measures and following a national lockdown. Transmission clusters were largely segregated within the university or the community. Our study highlights key determinants of SARS-CoV-2 transmission and effective interventions in a higher education setting that will inform public health policy during pandemics.
Abstract.
Nickbakhsh S, Hughes J, Christofidis N, Griffiths E, Shaaban S, Enright J, Smollett K, Nomikou K, Palmalux N, Tong L, et al (2022). Genomic epidemiology of SARS-CoV-2 in a university outbreak setting and implications for public health planning.
Scientific Reports,
12(1).
Abstract:
Genomic epidemiology of SARS-CoV-2 in a university outbreak setting and implications for public health planning
Whole genome sequencing of SARS-CoV-2 has occurred at an unprecedented scale, and can be exploited for characterising outbreak risks at the fine-scale needed to inform control strategies. One setting at continued risk of COVID-19 outbreaks are higher education institutions, associated with student movements at the start of term, close living conditions within residential halls, and high social contact rates. Here we analysed SARS-CoV-2 whole genome sequences in combination with epidemiological data to investigate a large cluster of student cases associated with University of Glasgow accommodation in autumn 2020, Scotland. We identified 519 student cases of SARS-CoV-2 infection associated with this large cluster through contact tracing data, with 30% sequencing coverage for further analysis. We estimated at least 11 independent introductions of SARS-CoV-2 into the student population, with four comprising the majority of detected cases and consistent with separate outbreaks. These four outbreaks were curtailed within a week following implementation of control measures. The impact of student infections on the local community was short-term despite an underlying increase in community infections. Our study highlights the need for context-specific information in the formation of public health policy for higher educational settings.
Abstract.
Twohig KA, Nyberg T, Zaidi A, Thelwall S, Sinnathamby MA, Aliabadi S, Seaman SR, Harris RJ, Hope R, Lopez-Bernal J, et al (2022). Hospital admission and emergency care attendance risk for SARS-CoV-2 delta (B.1.617.2) compared with alpha (B.1.1.7) variants of concern: a cohort study.
The Lancet Infectious Diseases,
22(1), 35-42.
Abstract:
Hospital admission and emergency care attendance risk for SARS-CoV-2 delta (B.1.617.2) compared with alpha (B.1.1.7) variants of concern: a cohort study
Background: the SARS-CoV-2 delta (B.1.617.2) variant was first detected in England in March, 2021. It has since rapidly become the predominant lineage, owing to high transmissibility. It is suspected that the delta variant is associated with more severe disease than the previously dominant alpha (B.1.1.7) variant. We aimed to characterise the severity of the delta variant compared with the alpha variant by determining the relative risk of hospital attendance outcomes. Methods: This cohort study was done among all patients with COVID-19 in England between March 29 and May 23, 2021, who were identified as being infected with either the alpha or delta SARS-CoV-2 variant through whole-genome sequencing. Individual-level data on these patients were linked to routine health-care datasets on vaccination, emergency care attendance, hospital admission, and mortality (data from Public Health England's Second Generation Surveillance System and COVID-19-associated deaths dataset; the National Immunisation Management System; and NHS Digital Secondary Uses Services and Emergency Care Data Set). The risk for hospital admission and emergency care attendance were compared between patients with sequencing-confirmed delta and alpha variants for the whole cohort and by vaccination status subgroups. Stratified Cox regression was used to adjust for age, sex, ethnicity, deprivation, recent international travel, area of residence, calendar week, and vaccination status. Findings: Individual-level data on 43 338 COVID-19-positive patients (8682 with the delta variant, 34 656 with the alpha variant; median age 31 years [IQR 17–43]) were included in our analysis. 196 (2·3%) patients with the delta variant versus 764 (2·2%) patients with the alpha variant were admitted to hospital within 14 days after the specimen was taken (adjusted hazard ratio [HR] 2·26 [95% CI 1·32–3·89]). 498 (5·7%) patients with the delta variant versus 1448 (4·2%) patients with the alpha variant were admitted to hospital or attended emergency care within 14 days (adjusted HR 1·45 [1·08–1·95]). Most patients were unvaccinated (32 078 [74·0%] across both groups). The HRs for vaccinated patients with the delta variant versus the alpha variant (adjusted HR for hospital admission 1·94 [95% CI 0·47–8·05] and for hospital admission or emergency care attendance 1·58 [0·69–3·61]) were similar to the HRs for unvaccinated patients (2·32 [1·29–4·16] and 1·43 [1·04–1·97]; p=0·82 for both) but the precision for the vaccinated subgroup was low. Interpretation: This large national study found a higher hospital admission or emergency care attendance risk for patients with COVID-19 infected with the delta variant compared with the alpha variant. Results suggest that outbreaks of the delta variant in unvaccinated populations might lead to a greater burden on health-care services than the alpha variant. Funding: Medical Research Council; UK Research and Innovation; Department of Health and Social Care; and National Institute for Health Research.
Abstract.
Fletcher J, Porter R, Boulton Z, Brown L, Knight B, Romanczuk L, Aiken S, Delury C, Michell S (2022). In vitro efficacy of antibiotic loaded calcium sulfate beads (Stimulan Rapid Cure) against polymicrobial communities and individual bacterial strains derived from diabetic foot infections.
J Med Microbiol,
71(5).
Abstract:
In vitro efficacy of antibiotic loaded calcium sulfate beads (Stimulan Rapid Cure) against polymicrobial communities and individual bacterial strains derived from diabetic foot infections.
Introduction. Diabetic foot infection (DFI) is the main reason for diabetes-related hospitalisation and is a major cause of diabetes-related amputation. DFIs are often complicated by ischaemia in the affected limb, the presence of polymicrobial biofilms and increasingly the occurrence of antibiotic resistant bacteria.Hypothesis/Gap statement. Antibiotic loaded beads could inhibit the growth of polymicrobial DFI communities with differing compositions in vitro.Aim. This study investigates the in vitro efficacy of antibiotic loaded calcium sulfate beads (Stimulan Rapid Cure, Biocomposites Ltd. UK) against polymicrobial DFI communities and individual bacterial strains derived from DFIs.Methodology. Debrided tissue obtained from the base of infected diabetic foot ulcers was homogenised and spread over the surface of Columbia blood agar (CBA) and fastidious anaerobe agar (FAA) plates. Calcium sulfate beads containing a combination of vancomycin and gentamicin were then placed on the surface of the agar and following incubation, zones of inhibition (ZOI) were measured. For individual bacterial strains isolated from the infected tissue, calcium sulfate beads containing vancomycin, gentamicin, flucloxacillin or rifampicin and beads containing a combination of vancomycin and gentamicin or flucloxacillin and rifampicin were tested for their ability to inhibit growth.Results. Calcium sulfate beads loaded with a combination of vancomycin and gentamicin were able to inhibit bacterial growth from all polymicrobial tissue homogenates tested, with ZOI diameters ranging from 15 to 40 mm. In the case of individual bacterial strains, beads containing combinations of vancomycin and gentamicin or flucloxacillin and rifampicin were able to produce ZOI with Gram-positive facultatitive anaerobic strains such as Staphylococcus aureus and Enterococcus faecalis, Gram-negative facultative anaerobic strains such as Pseudomonas aeruginosa and obligate anaerobic strains such as Finegoldia magna even where acquired resistance to one of the antibiotics in the combination was evidenced.Conclusion. The local use of calcium sulfate beads containing a combination of two antibiotics demonstrated high efficacy against polymicrobial DFI communities and individual DFI bacterial strains in in vitro zone of inhibition tests. These results show promise for clinical application, but further research and clinical studies are required.
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Author URL.
Robson SC, Connor TR, Loman NJ, Golubchik T, Martinez Nunez RT, Bonsall D, Rambaut A, Snell LB, Ludden C, Corden S, et al (2022). Publisher Correction: Genomic reconstruction of the SARS CoV-2 epidemic in England. Nature, 606(7915), e18-e18.
Willett BJ, Grove J, MacLean OA, Wilkie C, De Lorenzo G, Furnon W, Cantoni D, Scott S, Logan N, Ashraf S, et al (2022). SARS-CoV-2 Omicron is an immune escape variant with an altered cell entry pathway.
Nature Microbiology,
7(8), 1161-1179.
Abstract:
SARS-CoV-2 Omicron is an immune escape variant with an altered cell entry pathway
Vaccines based on the spike protein of SARS-CoV-2 are a cornerstone of the public health response to COVID-19. The emergence of hypermutated, increasingly transmissible variants of concern (VOCs) threaten this strategy. Omicron (B.1.1.529), the fifth VOC to be described, harbours multiple amino acid mutations in spike, half of which lie within the receptor-binding domain. Here we demonstrate substantial evasion of neutralization by Omicron BA.1 and BA.2 variants in vitro using sera from individuals vaccinated with ChAdOx1, BNT162b2 and mRNA-1273. These data were mirrored by a substantial reduction in real-world vaccine effectiveness that was partially restored by booster vaccination. The Omicron variants BA.1 and BA.2 did not induce cell syncytia in vitro and favoured a TMPRSS2-independent endosomal entry pathway, these phenotypes mapping to distinct regions of the spike protein. Impaired cell fusion was determined by the receptor-binding domain, while endosomal entry mapped to the S2 domain. Such marked changes in antigenicity and replicative biology may underlie the rapid global spread and altered pathogenicity of the Omicron variant.
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Ashford F, Best A, Dunn SJ, Ahmed Z, Siddiqui H, Melville J, Wilkinson S, Mirza J, Cumley N, Stockton J, et al (2022). SARS-CoV-2 Testing in the Community: Testing Positive Samples with the TaqMan SARS-CoV-2 Mutation Panel to Find Variants in Real Time. Journal of Clinical Microbiology, 60(4), e02408-e02421.
Volz E, Mishra S, Chand M, Barrett JC, Johnson R, Geidelberg L, Hinsley WR, Laydon DJ, Dabrera G, Toole AO, et al (2021). Assessing transmissibility of SARS-CoV-2 lineage B.1.1.7 in England.
NATURE,
593(7858), 266-+.
Author URL.
Graham MS, Sudre CH, May A, Antonelli M, Murray B, Varsavsky T, Kläser K, Canas LS, Molteni E, Modat M, et al (2021). Changes in symptomatology, reinfection, and transmissibility associated with the SARS-CoV-2 variant B.1.1.7: an ecological study. The Lancet Public Health, 6(5), e335-e345.
Volz E, Hill V, McCrone JT, Price A, Jorgensen D, O’Toole Á, Southgate J, Johnson R, Jackson B, Nascimento FF, et al (2021). Evaluating the Effects of SARS-CoV-2 Spike Mutation D614G on Transmissibility and Pathogenicity. Cell, 184(1), 64-75.e11.
Elliott P, Haw D, Wang H, Eales O, Walters CE, Ainslie KEC, Atchison C, Fronterre C, Diggle PJ, Page AJ, et al (2021). Exponential growth, high prevalence of SARS-CoV-2, and vaccine effectiveness associated with the Delta variant.
Science,
374(6574).
Abstract:
Exponential growth, high prevalence of SARS-CoV-2, and vaccine effectiveness associated with the Delta variant.
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections were rising during early summer 2021 in many countries as a result of the Delta variant. We assessed reverse transcription polymerase chain reaction swab positivity in the Real-time Assessment of Community Transmission–1 (REACT-1) study in England. During June and July 2021, we observed sustained exponential growth with an average doubling time of 25 days, driven by complete replacement of the Alpha variant by Delta and by high prevalence at younger, less-vaccinated ages. Prevalence among unvaccinated people [1.21% (95% credible interval 1.03%, 1.41%)] was three times that among double-vaccinated people [0.40% (95% credible interval 0.34%, 0.48%)]. However, after adjusting for age and other variables, vaccine effectiveness for double-vaccinated people was estimated at between ~50% and ~60% during this period in England. Increased social mixing in the presence of Delta had the potential to generate sustained growth in infections, even at high levels of vaccination.
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Vöhringer HS, Sanderson T, Sinnott M, De Maio N, Nguyen T, Goater R, Schwach F, Harrison I, Hellewell J, Ariani CV, et al (2021). Genomic reconstruction of the SARS-CoV-2 epidemic in England.
Nature,
600(7889), 506-511.
Abstract:
Genomic reconstruction of the SARS-CoV-2 epidemic in England
AbstractThe evolution of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus leads to new variants that warrant timely epidemiological characterization. Here we use the dense genomic surveillance data generated by the COVID-19 Genomics UK Consortium to reconstruct the dynamics of 71 different lineages in each of 315 English local authorities between September 2020 and June 2021. This analysis reveals a series of subepidemics that peaked in early autumn 2020, followed by a jump in transmissibility of the B.1.1.7/Alpha lineage. The Alpha variant grew when other lineages declined during the second national lockdown and regionally tiered restrictions between November and December 2020. A third more stringent national lockdown suppressed the Alpha variant and eliminated nearly all other lineages in early 2021. Yet a series of variants (most of which contained the spike E484K mutation) defied these trends and persisted at moderately increasing proportions. However, by accounting for sustained introductions, we found that the transmissibility of these variants is unlikely to have exceeded the transmissibility of the Alpha variant. Finally, B.1.617.2/Delta was repeatedly introduced in England and grew rapidly in early summer 2021, constituting approximately 98% of sampled SARS-CoV-2 genomes on 26 June 2021.
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Kemp SA, Collier DA, Datir RP, Ferreira IATM, Gayed S, Jahun A, Hosmillo M, Rees-Spear C, Mlcochova P, Lumb IU, et al (2021). SARS-CoV-2 evolution during treatment of chronic infection.
Nature,
592(7853), 277-282.
Abstract:
SARS-CoV-2 evolution during treatment of chronic infection
The spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is critical for virus infection through the engagement of the human ACE2 protein1 and is a major antibody target. Here we show that chronic infection with SARS-CoV-2 leads to viral evolution and reduced sensitivity to neutralizing antibodies in an immunosuppressed individual treated with convalescent plasma, by generating whole-genome ultra-deep sequences for 23 time points that span 101 days and using in vitro techniques to characterize the mutations revealed by sequencing. There was little change in the overall structure of the viral population after two courses of remdesivir during the first 57 days. However, after convalescent plasma therapy, we observed large, dynamic shifts in the viral population, with the emergence of a dominant viral strain that contained a substitution (D796H) in the S2 subunit and a deletion (ΔH69/ΔV70) in the S1 N-terminal domain of the spike protein. As passively transferred serum antibodies diminished, viruses with the escape genotype were reduced in frequency, before returning during a final, unsuccessful course of convalescent plasma treatment. In vitro, the spike double mutant bearing both ΔH69/ΔV70 and D796H conferred modestly decreased sensitivity to convalescent plasma, while maintaining infectivity levels that were similar to the wild-type virus.The spike substitution mutant D796H appeared to be the main contributor to the decreased susceptibility to neutralizing antibodies, but this mutation resulted in an infectivity defect. The spike deletion mutant ΔH69/ΔV70 had a twofold higher level of infectivity than wild-type SARS-CoV-2, possibly compensating for the reduced infectivity of the D796H mutation. These data reveal strong selection on SARS-CoV-2 during convalescent plasma therapy, which is associated with the emergence of viral variants that show evidence of reduced susceptibility to neutralizing antibodies in immunosuppressed individuals.
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Collier DA, De Marco A, Ferreira IATM, Meng B, Datir RP, Walls AC, Kemp SA, Bassi J, Pinto D, Silacci-Fregni C, et al (2021). Sensitivity of SARS-CoV-2 B.1.1.7 to mRNA vaccine-elicited antibodies.
NATURE,
593(7857), 136-+.
Author URL.
de Silva TI, Liu G, Lindsey BB, Dong D, Moore SC, Hsu NS, Shah D, Wellington D, Mentzer AJ, Angyal A, et al (2021). The impact of viral mutations on recognition by SARS-CoV-2 specific T cells.
iScience,
24(11).
Abstract:
The impact of viral mutations on recognition by SARS-CoV-2 specific T cells
We identify amino acid variants within dominant SARS-CoV-2 T cell epitopes by interrogating global sequence data. Several variants within nucleocapsid and ORF3a epitopes have arisen independently in multiple lineages and result in loss of recognition by epitope-specific T cells assessed by IFN-γ and cytotoxic killing assays. Complete loss of T cell responsiveness was seen due to Q213K in the A∗01:01-restricted CD8+ ORF3a epitope FTSDYYQLY207-215; due to P13L, P13S, and P13T in the B∗27:05-restricted CD8+ nucleocapsid epitope QRNAPRITF9-17; and due to T362I and P365S in the A∗03:01/A∗11:01-restricted CD8+ nucleocapsid epitope KTFPPTEPK361-369. CD8+ T cell lines unable to recognize variant epitopes have diverse T cell receptor repertoires. These data demonstrate the potential for T cell evasion and highlight the need for ongoing surveillance for variants capable of escaping T cell as well as humoral immunity.
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Hubert CL, Michell SL (2020). A universal oyster infection model demonstrates that Vibrio vulnificus Type 6 secretion systems have antibacterial activity in vivo.
Environ Microbiol,
22(10), 4381-4393.
Abstract:
A universal oyster infection model demonstrates that Vibrio vulnificus Type 6 secretion systems have antibacterial activity in vivo.
With the rapid increase of aquaculture contributing to sustainable food security, comes the need to better understand seafood associated diseases. One of the major aquatic bacterial genera responsible for human infections from seafood is Vibrio, especially from oysters. Currently, in vivo study of bacterial interactions within oysters is limited by the inability to promote high-level uptake of bacteria by oysters. This study has therefore evolved current natural marine snow protocols to generate 'artificial' marine snow, into which bacteria can be incorporated to facilitate extensive uptake by oysters. This presents an adaptable model for bacterial study within filter-feeding shellfish. Using this model, we demonstrate for the first time the antibacterial activity of Vibrio vulnificus Type 6 secretion systems in vivo, revealing an important role for the T6SS in V. vulnificus ecology.
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Aanensen DM, Abudahab K, Adams A, Afifi S, Alam MT, Alderton A, Alikhan N-F, Allan J, Almsaud M, Alrezaihi A, et al (2020). An integrated national scale SARS-CoV-2 genomic surveillance network.
LANCET MICROBE,
1(3), E99-E100.
Author URL.
Bradshaw WJ, Bruxelle J-F, Kovacs-Simon A, Harmer NJ, Janoir C, Péchiné S, Acharya KR, Michell SL (2019). Molecular features of lipoprotein CD0873: a potential vaccine against the human pathogen Clostridioides difficile.
J Biol Chem,
294(43), 15850-15861.
Abstract:
Molecular features of lipoprotein CD0873: a potential vaccine against the human pathogen Clostridioides difficile.
Clostridioides difficile is the primary cause of antibiotic-associated diarrhea and colitis, a healthcare-associated intestinal disease resulting in a significant fatality rate. Colonization of the gut is critical for C. difficile pathogenesis. The bacterial molecules essential for efficient colonization therefore offer great potential as vaccine candidates. Here we present findings demonstrating that the C. difficile immunogenic lipoprotein CD0873 plays a critical role in pathogen success in vivo We found that in a dixenic colonization model, a CD0873-positive strain of C. difficile significantly outcompeted a CD0873-negative strain. Immunization of mice with recombinant CD0873 prevented long-term gut colonization and was correlated with a strong secretory IgA immune response. We further present high-resolution crystal structures of CD0873, at 1.35-2.50 Å resolutions, offering a first view of the ligand-binding pocket of CD0873 and provide evidence that this lipoprotein adhesin is part of a tyrosine import system, an amino acid key in C. difficile infection. These findings suggest that CD0873 could serve as an effective component in a vaccine against C. difficile.
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Author URL.
Fletcher J, Porter R, Aiken S, Delury C, Michell S (2019). The effect of local release antibiotic beads on in-vitro bacterial growth from tissue taken from infected diabetic foot ulcers. Access Microbiology, 1(1A).
Church SR, Lux T, Baker-Austin C, Buddington SP, Michell SL (2016). Vibrio vulnificus Type 6 Secretion System 1 Contains Anti-Bacterial Properties.
PLoS One,
11(10).
Abstract:
Vibrio vulnificus Type 6 Secretion System 1 Contains Anti-Bacterial Properties.
Vibrio vulnificus is a bacterium responsible for severe gastroenteritis, sepsis and wound infections. Gastroenteritis and sepsis are commonly associated with the consumption of raw oysters, whereas wound infection is often associated with the handling of contaminated fish. Although classical virulence factors of this emerging pathogen are well characterised, there remains a paucity of knowledge regarding the general biology of this species. To investigate the presence of previously unreported virulence factors, we applied whole genome sequencing to a panel of ten V. vulnificus strains with varying virulence potentials. This identified two novel type 6 secretion systems (T6SSs), systems that are known to have a role in bacterial virulence and population dynamics. By utilising a range of molecular techniques and assays we have demonstrated the functionality of one of these T6SSs. Furthermore, we have shown that this system is subject to thermoregulation and is negatively regulated by increasing salinity concentrations. This secretion system was also shown to be involved in the killing of V. vulnificus strains that did not possess this system and a model is proposed as to how this interaction may contribute to population dynamics within V. vulnificus strains. In addition to this intra-species killing, this system also contributes to the killing of inter bacterial species and may have a role in the general composition of Vibrio species in the environment.
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Author URL.
Nandi T, Holden MTG, Didelot X, Mehershahi K, Boddey JA, Beacham I, Peak I, Harting J, Baybayan P, Guo Y, et al (2015). Burkholderia pseudomallei sequencing identifies genomic clades with distinct recombination, accessory, and epigenetic profiles.
Genome Res,
25(4).
Author URL.
Nandi T, Holden MTG, Didelot X, Mehershahi K, Boddey JA, Beacham I, Peak I, Harting J, Baybayan P, Guo Y, et al (2015). Burkholderia pseudomallei sequencing identifies genomic clades with distinct recombination, accessory, and epigenetic profiles.
Genome Res,
25(1), 129-141.
Abstract:
Burkholderia pseudomallei sequencing identifies genomic clades with distinct recombination, accessory, and epigenetic profiles.
Burkholderia pseudomallei (Bp) is the causative agent of the infectious disease melioidosis. To investigate population diversity, recombination, and horizontal gene transfer in closely related Bp isolates, we performed whole-genome sequencing (WGS) on 106 clinical, animal, and environmental strains from a restricted Asian locale. Whole-genome phylogenies resolved multiple genomic clades of Bp, largely congruent with multilocus sequence typing (MLST). We discovered widespread recombination in the Bp core genome, involving hundreds of regions associated with multiple haplotypes. Highly recombinant regions exhibited functional enrichments that may contribute to virulence. We observed clade-specific patterns of recombination and accessory gene exchange, and provide evidence that this is likely due to ongoing recombination between clade members. Reciprocally, interclade exchanges were rarely observed, suggesting mechanisms restricting gene flow between clades. Interrogation of accessory elements revealed that each clade harbored a distinct complement of restriction-modification (RM) systems, predicted to cause clade-specific patterns of DNA methylation. Using methylome sequencing, we confirmed that representative strains from separate clades indeed exhibit distinct methylation profiles. Finally, using an E. coli system, we demonstrate that Bp RM systems can inhibit uptake of non-self DNA. Our data suggest that RM systems borne on mobile elements, besides preventing foreign DNA invasion, may also contribute to limiting exchanges of genetic material between individuals of the same species. Genomic clades may thus represent functional units of genetic isolation in Bp, modulating intraspecies genetic diversity.
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Nandi T, Holden MTG, Didelot X, Mehershahi K, Boddey JA, Beacham I, Peak I, Harting J, Baybayan P, Guo Y, et al (2015). Errata: Burkholderia pseudomallei sequencing identifies genomic clades with distinct recombination, accessory, and epigenetic profiles (Genome Research (2015) 25 (129-141)). Genome Research, 25(4).
Aiken SS, Cooper JJ, Florance H, Robinson MT, Michell S (2015). Local Release of Antibiotics for Surgical Site Infection Management Using High-Purity Calcium Sulfate: an in Vitro Elution Study.
SURGICAL INFECTIONS,
16(1), 54-61.
Author URL.
Charlton TM, Kovacs-Simon A, Michell SL, Fairweather NF, Tate EW (2015). Quantitative Lipoproteomics in Clostridium difficile Reveals a Role for Lipoproteins in Sporulation.
CHEMISTRY & BIOLOGY,
22(11), 1562-1573.
Author URL.
Peano C, Chiaramonte F, Motta S, Pietrelli A, Jaillon S, Rossi E, Consolandi C, Champion OL, Michell SL, Freddi L, et al (2014). Gene and protein expression in response to different growth temperatures and oxygen availability in Burkholderia thailandensis.
PLoS One,
9(3).
Abstract:
Gene and protein expression in response to different growth temperatures and oxygen availability in Burkholderia thailandensis.
Burkholderia thailandensis, although normally avirulent for mammals, can infect macrophages in vitro and has occasionally been reported to cause pneumonia in humans. It is therefore used as a model organism for the human pathogen B. pseudomallei, to which it is closely related phylogenetically. We characterized the B. thailandensis clinical isolate CDC2721121 (BtCDC272) at the genome level and studied its response to environmental cues associated with human host colonization, namely, temperature and oxygen limitation. Effects of the different growth conditions on BtCDC272 were studied through whole genome transcription studies and analysis of proteins associated with the bacterial cell surface. We found that growth at 37°C, compared to 28°C, negatively affected cell motility and flagella production through a mechanism involving regulation of the flagellin-encoding fliC gene at the mRNA stability level. Growth in oxygen-limiting conditions, in contrast, stimulated various processes linked to virulence, such as lipopolysaccharide production and expression of genes encoding protein secretion systems. Consistent with these observations, BtCDC272 grown in oxygen limitation was more resistant to phagocytosis and strongly induced the production of inflammatory cytokines from murine macrophages. Our results suggest that, while temperature sensing is important for regulation of B. thailandensis cell motility, oxygen limitation has a deeper impact on its physiology and constitutes a crucial environmental signal for the production of virulence factors.
Abstract.
Author URL.
Kovacs-Simon A, Leuzzi R, Kasendra M, Minton N, Titball RW, Michell SL (2014). Lipoprotein CD0873 is a novel adhesin of Clostridium difficile.
J Infect Dis,
210(2), 274-284.
Abstract:
Lipoprotein CD0873 is a novel adhesin of Clostridium difficile.
Clostridium difficile is a cause of antibiotic-associated diarrhea and colitis, a healthcare-associated intestinal disease. Colonization of the gut is a critical step in the course of infection. The C. difficile lipoprotein CD0873 was identified as a putative adhesin through a bioinformatics approach. Surface exposure of CD0873 was confirmed and a CD0873 mutant was generated. The CD0873 mutant showed a significant reduction in adherence to Caco-2 cells and wild-type bacteria preincubated with anti-CD0873 antibodies showed significantly decreased adherence to Caco-2 cells. In addition, we demonstrated that purified recombinant CD0873 protein alone associates with Caco-2 cells. This is the first definitive identification of a C. difficile adhesin, which now allows work to devise improved measures for preventing and treating disease.
Abstract.
Author URL.
Butt E, Foster JAH, Keedwell E, Bell JEA, Titball RW, Bhangu A, Michell SL, Sheridan R (2013). Derivation and validation of a simple, accurate and robust prediction rule for risk of mortality in patients with Clostridium difficile infection.
BMC Infect Dis,
13Abstract:
Derivation and validation of a simple, accurate and robust prediction rule for risk of mortality in patients with Clostridium difficile infection.
BACKGROUND: Clostridium difficile infection poses a significant healthcare burden. However, the derivation of a simple, evidence based prediction rule to assist patient management has not yet been described. METHOD: Univariate, multivariate and decision tree procedures were used to deduce a prediction rule from over 186 variables; retrospectively collated from clinical data for 213 patients. The resulting prediction rule was validated on independent data from a cohort of 158 patients described by Bhangu et al. (Colorectal Disease, 12(3):241-246, 2010). RESULTS: Serum albumin levels (g/L) (P = 0.001), respiratory rate (resps /min) (P = 0.002), C-reactive protein (mg/L) (P = 0.034) and white cell count (mcL) (P = 0.049) were predictors of all-cause mortality. Threshold levels of serum albumin ≤ 24.5 g/L, C- reactive protein >228 mg/L, respiratory rate >17 resps/min and white cell count >12 × 10(3) mcL were associated with an increased risk of all-cause mortality. A simple four variable prediction rule was devised based on these threshold levels and when tested on the initial data, yield an area under the curve score of 0.754 (P
Abstract.
Author URL.
He M, Miyajima F, Roberts P, Ellison L, Pickard DJ, Martin MJ, Connor TR, Harris SR, Fairley D, Bamford KB, et al (2013). Emergence and global spread of epidemic healthcare-associated Clostridium difficile.
Nature Genetics,
45(1), 109-113.
Abstract:
Emergence and global spread of epidemic healthcare-associated Clostridium difficile
Epidemic C. difficile (027/BI/NAP1) has rapidly emerged in the past decade as the leading cause of antibiotic-associated diarrhea worldwide. However, the key events in evolutionary history leading to its emergence and the subsequent patterns of global spread remain unknown. Here, we define the global population structure of C. difficile 027/BI/NAP1 using whole-genome sequencing and phylogenetic analysis. We show that two distinct epidemic lineages, FQR1 and FQR2, not one as previously thought, emerged in North America within a relatively short period after acquiring the same fluoroquinolone resistance-conferring mutation and a highly related conjugative transposon. The two epidemic lineages showed distinct patterns of global spread, and the FQR2 lineage spread more widely, leading to healthcare-associated outbreaks in the UK, continental Europe and Australia. Our analysis identifies key genetic changes linked to the rapid transcontinental dissemination of epidemic C. difficile 027/BI/NAP1 and highlights the routes by which it spreads through the global healthcare system. © 2013 Nature America, Inc. All rights reserved.
Abstract.
Gourlay LJ, Peri C, Ferrer-Navarro M, Conchillo-Solé O, Gori A, Rinchai D, Thomas RJ, Champion OL, Michell SL, Kewcharoenwong C, et al (2013). Exploiting the Burkholderia pseudomallei acute phase antigen BPSL2765 for structure-based epitope discovery/design in structural vaccinology.
Chem Biol,
20(9), 1147-1156.
Abstract:
Exploiting the Burkholderia pseudomallei acute phase antigen BPSL2765 for structure-based epitope discovery/design in structural vaccinology.
We solved the crystal structure of Burkholderia pseudomallei acute phase antigen BPSL2765 in the context of a structural vaccinology study, in the area of melioidosis vaccine development. Based on the structure, we applied a recently developed method for epitope design that combines computational epitope predictions with in vitro mapping experiments and successfully identified a consensus sequence within the antigen that, when engineered as a synthetic peptide, was selectively immunorecognized to the same extent as the recombinant protein in sera from melioidosis-affected subjects. Antibodies raised against the consensus peptide were successfully tested in opsonization bacterial killing experiments and antibody-dependent agglutination tests of B. pseudomallei. Our strategy represents a step in the development of immunodiagnostics, in the production of specific antibodies and in the optimization of antigens for vaccine development, starting from structural and physicochemical principles.
Abstract.
Author URL.
Michell SL (2012). A better understanding of what makes some proteins "fat".
J Bacteriol,
194(9), 2129-2130.
Author URL.
He M, Miyajima F, Roberts P, Ellison L, Pickard DJ, Martin MJ, Connor TR, Harris SR, Fairley D, Bamford KB, et al (2012). Emergence and global spread of epidemic healthcare-associated Clostridium difficile. Nature Genetics
Foster JAH, Butt JEC, Bell J, Goff A, Morgan C, Hancock J, Carmichael C, Keedwell EC, Michell SLI, Sheridan RP, et al (2012). IMPROVING CLINICAL MANAGEMENT IN CLOSTRIDIUM DIFFICILE: FAECAL CALPROTECTIN DOES NOT PREDICT SEVERITY, RECURRENCE OR MORTALITY.
AGE AND AGEING,
41, 72-72.
Author URL.
Thomas RM, Twine SM, Fulton KM, Tessier L, Kilmury SLN, Ding W, Harmer N, Michell SL, Oyston PCF, Titball RW, et al (2011). Glycosylation of DsbA in Francisella tularensis subsp. tularensis.
J Bacteriol,
193(19), 5498-5509.
Abstract:
Glycosylation of DsbA in Francisella tularensis subsp. tularensis.
In Francisella tularensis subsp. tularensis, DsbA has been shown to be an essential virulence factor and has been observed to migrate to multiple protein spots on two-dimensional electrophoresis gels. In this work, we show that the protein is modified with a 1,156-Da glycan moiety in O-linkage. The results of mass spectrometry studies suggest that the glycan is a hexasaccharide, comprised of N-acetylhexosamines, hexoses, and an unknown monosaccharide. Disruption of two genes within the FTT0789-FTT0800 putative polysaccharide locus, including a galE homologue (FTT0791) and a putative glycosyltransferase (FTT0798), resulted in loss of glycan modification of DsbA. The F. tularensis subsp. tularensis ΔFTT0798 and ΔFTT0791::Cm mutants remained virulent in the murine model of subcutaneous tularemia. This indicates that glycosylation of DsbA does not play a major role in virulence under these conditions. This is the first report of the detailed characterization of the DsbA glycan and putative role of the FTT0789-FTT0800 gene cluster in glycan biosynthesis.
Abstract.
Author URL.
Kovacs-Simon A, Titball RW, Michell SL (2011). Lipoproteins of bacterial pathogens.
Infect Immun,
79(2), 548-561.
Abstract:
Lipoproteins of bacterial pathogens.
Bacterial lipoproteins are a set of membrane proteins with many different functions. Due to this broad-ranging functionality, these proteins have a considerable significance in many phenomena, from cellular physiology through cell division and virulence. Here we give a general overview of lipoprotein biogenesis and highlight examples of the roles of lipoproteins in bacterial disease caused by a selection of medically relevant Gram-negative and Gram-positive pathogens: Mycobacterium tuberculosis, Streptococcus pneumoniae, Borrelia burgdorferi, and Neisseria meningitidis. Lipoproteins have been shown to play key roles in adhesion to host cells, modulation of inflammatory processes, and translocation of virulence factors into host cells. As such, a number of lipoproteins have been shown to be potential vaccines. This review provides a summary of some of the reported roles of lipoproteins and of how this knowledge has been exploited in some cases for the generation of novel countermeasures to bacterial diseases.
Abstract.
Author URL.
Wand ME, Müller CM, Titball RW, Michell SL (2011). Macrophage and Galleria mellonella infection models reflect the virulence of naturally occurring isolates of B. pseudomallei, B. thailandensis and B. oklahomensis.
BMC Microbiol,
11(1).
Abstract:
Macrophage and Galleria mellonella infection models reflect the virulence of naturally occurring isolates of B. pseudomallei, B. thailandensis and B. oklahomensis.
BACKGROUND: Burkholderia pseudomallei is the causative agent of melioidosis, a tropical disease of humans with a variable and often fatal outcome. In murine models of infection, different strains exhibit varying degrees of virulence. In contrast, two related species, B. thailandensis and B. oklahomensis, are highly attenuated in mice. Our aim was to determine whether virulence in mice is reflected in macrophage or wax moth larvae (Galleria mellonella) infection models. RESULTS: B. pseudomallei strains 576 and K96243, which have low median lethal dose (MLD) values in mice, were able to replicate and induce cellular damage in macrophages and caused rapid death of G. mellonella. In contrast, B. pseudomallei strain 708a, which is attenuated in mice, showed reduced replication in macrophages, negligible cellular damage and was avirulent in G. mellonella larvae. B. thailandensis isolates were less virulent than B. pseudomallei in all of the models tested. However, we did record strain dependent differences. B. oklahomensis isolates were the least virulent isolates. They showed minimal ability to replicate in macrophages, were unable to evoke actin-based motility or to form multinucleated giant cells and were markedly attenuated in G. mellonella compared to B. thailandensis. CONCLUSIONS: We have shown that the alternative infection models tested here, namely macrophages and Galleria mellonella, are able to distinguish between strains of B. pseudomallei, B. thailandensis and B. oklahomensis and that these differences reflect the observed virulence in murine infection models. Our results indicate that B. oklahomensis is the least pathogenic of the species investigated. They also show a correlation between isolates of B. thailandensis associated with human infection and virulence in macrophage and Galleria infection models.
Abstract.
Author URL.
Wang W, Hale C, Goulding D, Haslam SM, Tissot B, Lindsay C, Michell S, Titball R, Yu J, Toribio AL, et al (2011). Mannosidase 2, alpha 1 deficiency is associated with ricin resistance in embryonic stem (ES) cells.
PLoS One,
6(8).
Abstract:
Mannosidase 2, alpha 1 deficiency is associated with ricin resistance in embryonic stem (ES) cells.
Host gene products required for mediating the action of toxins are potential targets for reversing or controlling their pathogenic impact following exposure. To identify such targets libraries of insertional gene-trap mutations generated with a PiggyBac transposon in Blm-deficient embryonic stem cells were exposed to the plant toxin, ricin. Resistant clones were isolated and genetically characterised and one was found to be a homozygous mutant of the mannosidase 2, alpha 1 (Man2α1) locus with a matching defect in the homologous allele. The causality of the molecular lesion was confirmed by removal of the transposon following expression of PB-transposase. Comparative glycomic and lectin binding analysis of the Man2α1 (-/-) ricin resistant cells revealed an increase in the levels of hybrid glycan structures and a reduction in terminal β-galactose moieties, potential target receptors for ricin. Furthermore, naïve ES cells treated with inhibitors of the N-linked glycosylation pathway at the mannosidase 2, alpha 1 step exhibited either full or partial resistance to ricin. Therefore, we conclusively identified mannosidase 2, alpha 1 deficiency to be associated with ricin resistance.
Abstract.
Author URL.
Vanaporn M, Wand M, Michell SL, Sarkar-Tyson M, Ireland P, Goldman S, Kewcharoenwong C, Rinchai D, Lertmemongkolchai G, Titball RW, et al (2011). Superoxide dismutase C is required for intracellular survival and virulence of Burkholderia pseudomallei.
Microbiology (Reading),
157(Pt 8), 2392-2400.
Abstract:
Superoxide dismutase C is required for intracellular survival and virulence of Burkholderia pseudomallei.
Burkholderia pseudomallei is an intracellular pathogen and the causative agent of melioidosis, a life-threatening disease of humans. Within host cells, superoxide is an important mediator of pathogen killing. In this study, we have identified the B. pseudomallei K96243 sodC gene, shown that it has superoxide dismutase activity, and constructed an allelic deletion mutant of this gene. Compared with the wild-type, the mutant was more sensitive to killing by extracellular superoxide, but not to superoxide generated intracellularly. The sodC mutant showed a markedly decreased survival in J774A.1 mouse macrophages, and reduced numbers of bacteria were recovered from human polymorphonuclear neutrophils (PMNs) when compared with the wild-type. The numbers of wild-type or mutant bacteria recovered from human diabetic neutrophils were significantly lower than from normal human neutrophils. The sodC mutant was attenuated in BALB/c mice. Our results indicate that SodC plays a key role in the virulence of B. pseudomallei, but that diabetics are not more susceptible to infection because of a reduced ability of PMNs to kill by superoxide.
Abstract.
Author URL.
Michell SL, Dean RE, Eyles JE, Hartley MG, Waters E, Prior JL, Titball RW, Oyston PCF (2010). Deletion of the Bacillus anthracis capB homologue in Francisella tularensis subspecies tularensis generates an attenuated strain that protects mice against virulent tularaemia.
J Med Microbiol,
59(Pt 11), 1275-1284.
Abstract:
Deletion of the Bacillus anthracis capB homologue in Francisella tularensis subspecies tularensis generates an attenuated strain that protects mice against virulent tularaemia.
As there is currently no licensed vaccine against Francisella tularensis, the causative agent of tularaemia, the bacterium is an agent of concern as a potential bioweapon. Although F. tularensis has a low infectious dose and high associated mortality, it possesses few classical virulence factors. An analysis of the F. tularensis subspecies tularensis genome sequence has revealed the presence of a region containing genes with low sequence homology to part of the capBCADE operon of Bacillus anthracis. We have generated an isogenic capB mutant of F. tularensis subspecies tularensis SchuS4 and shown it to be attenuated. Furthermore, using BALB/c mice, we have demonstrated that this capB strain affords protection against significant homologous challenge with the wild-type strain. These data have important implications for the development of a defined and efficacious tularaemia vaccine.
Abstract.
Author URL.
Forslund A-L, Salomonsson EN, Golovliov I, Kuoppa K, Michell S, Titball R, Oyston P, Noppa L, Sjöstedt A, Forsberg A, et al (2010). The type IV pilin, PilA, is required for full virulence of Francisella tularensis subspecies tularensis.
BMC Microbiol,
10Abstract:
The type IV pilin, PilA, is required for full virulence of Francisella tularensis subspecies tularensis.
BACKGROUND: all four Francisella tularensis subspecies possess gene clusters with potential to express type IV pili (Tfp). These clusters include putative pilin genes, as well as pilB, pilC and pilQ, required for secretion and assembly of Tfp. A hallmark of Tfp is the ability to retract the pilus upon surface contact, a property mediated by the ATPase PilT. Interestingly, out of the two major human pathogenic subspecies only the highly virulent type a strains have a functional pilT gene. RESULTS: in a previous study, we were able to show that one pilin gene, pilA, was essential for virulence of a type B strain in a mouse infection model. In this work we have examined the role of several Tfp genes in the virulence of the pathogenic type a strain SCHU S4. pilA, pilC, pilQ, and pilT were mutated by in-frame deletion mutagenesis. Interestingly, when mice were infected with a mixture of each mutant strain and the wild-type strain, the pilA, pilC and pilQ mutants were out-competed, while the pilT mutant was equally competitive as the wild-type. CONCLUSIONS: This suggests that expression and surface localisation of PilA contribute to virulence in the highly virulent type a strain, while PilT was dispensable for virulence in the mouse infection model.
Abstract.
Author URL.
Champion MD, Zeng Q, Nix EB, Nano FE, Keim P, Kodira CD, Borowsky M, Young S, Koehrsen M, Engels R, et al (2009). Comparative genomic characterization of Francisella tularensis strains belonging to low and high virulence subspecies.
PLoS Pathog,
5(5).
Abstract:
Comparative genomic characterization of Francisella tularensis strains belonging to low and high virulence subspecies.
Tularemia is a geographically widespread, severely debilitating, and occasionally lethal disease in humans. It is caused by infection by a gram-negative bacterium, Francisella tularensis. In order to better understand its potency as an etiological agent as well as its potential as a biological weapon, we have completed draft assemblies and report the first complete genomic characterization of five strains belonging to the following different Francisella subspecies (subsp.): the F. tularensis subsp. tularensis FSC033, F. tularensis subsp. holarctica FSC257 and FSC022, and F. tularensis subsp. novicida GA99-3548 and GA99-3549 strains. Here, we report the sequencing of these strains and comparative genomic analysis with recently available public Francisella sequences, including the rare F. tularensis subsp. mediasiatica FSC147 strain isolate from the Central Asian Region. We report evidence for the occurrence of large-scale rearrangement events in strains of the holarctica subspecies, supporting previous proposals that further phylogenetic subdivisions of the Type B clade are likely. We also find a significant enrichment of disrupted or absent ORFs proximal to predicted breakpoints in the FSC022 strain, including a genetic component of the Type I restriction-modification defense system. Many of the pseudogenes identified are also disrupted in the closely related rarely human pathogenic F. tularensis subsp. mediasiatica FSC147 strain, including modulator of drug activity B (mdaB) (FTT0961), which encodes a known NADPH quinone reductase involved in oxidative stress resistance. We have also identified genes exhibiting sequence similarity to effectors of the Type III (T3SS) and components of the Type IV secretion systems (T4SS). One of the genes, msrA2 (FTT1797c), is disrupted in F. tularensis subsp. mediasiatica and has recently been shown to mediate bacterial pathogen survival in host organisms. Our findings suggest that in addition to the duplication of the Francisella Pathogenicity Island, and acquisition of individual loci, adaptation by gene loss in the more recently emerged tularensis, holarctica, and mediasiatica subspecies occurred and was distinct from evolutionary events that differentiated these subspecies, and the novicida subspecies, from a common ancestor. Our findings are applicable to future studies focused on variations in Francisella subspecies pathogenesis, and of broader interest to studies of genomic pathoadaptation in bacteria.
Abstract.
Author URL.
Richards MI, Michell SL, Oyston PCF (2008). An intracellularly inducible gene involved in virulence and polyphosphate production in Francisella.
J Med Microbiol,
57(Pt 10), 1183-1192.
Abstract:
An intracellularly inducible gene involved in virulence and polyphosphate production in Francisella.
Francisella tularensis is an intracellular pathogen capable of multiplying to high levels in macrophages. By protein analysis, only a few proteins have been shown previously to be expressed at high levels in macrophages relative to bacteria grown in culture media. To identify additional genes that show increased expression during intracellular growth, we developed a plasmid for use in Francisella based on the induction of expression of green fluorescent protein. Clones of F. tularensis subsp. novicida were identified that were fluorescent only intracellularly and not when grown in vitro. Sequencing identified a range of genes comprising some such as dnaK that are already known to be expressed intracellularly and some novel targets. One of these newly identified regulated genes, FTN1472/FTT1564, was selected for further study. Isogenic mutants were generated in F. tularensis subsp. novicida and subsp. tularensis by allelic replacement. Inactivation of the gene resulted in abolition of polyphosphate production by F. novicida, strongly supporting the bioinformatic analysis, which had suggested that the gene may encode a polyphosphate kinase. The mutants exhibited defects for intracellular growth in macrophages and were attenuated in mice, indicating a key role for the putative polyphosphate kinase in the virulence of Francisella.
Abstract.
Author URL.
Michell, S.L. Diaper, H. Wikstrom, P. Titball, RW (2007). A 55 kDa hypothetical membrane protein is an iron-regulated virulence factor of Francisella tularensis subspecies novicida U112. Journal of Medical Microbiology, 56, 1268-1276.
Isherwood, K.E. Michell, S.L. Diaper, H. Titball, RW (2007). A Francisella tularensis subspecies novicida purF mutant, but not a purA mutant, induces protective immunity to tularemia in mice. Vaccine, 25, 2011-2018.
sm202, Grice ID, Griffin K, Hitchen PG (2007). The immunologically distinct O-antigens from Francisella tularensis subspecies tuarensis and Francisella novicida are both virulence determinants and protective antigens. Infection and Immunity, 75(1), 371-378.
sm202, Bystrom A, Forslund AL, Johansson A (2006). Direct repeat-mediated deletion of a type IV pilin gene results in major virulence attenuation of Francisella tularensis. Molecular Microbiology, 59(6), 1818-1830.
RT101, Andersson SGE, Chain P, Chu MC (2005). The complete genome sequence of Francisella tularensis, the causative agent of tularemia. Nature Genetics, 37(2), 153-159.
Michell SL, Whelan AO, Wheeler PR, Panico M, Easton RL, Etienne AT, Haslam SM, Dell A, Morris HR, Reason AJ, et al (2003). The MPB83 antigen from Mycobacterium bovis contains O-linked mannose and (1->3)-mannibiose moieties. Journal of Biological Chemistry, 278(18), 16423-16432.
Wooff EE, Michell SL, Gordon SV, Chambers MA, Bardarov A, Jacobs Jr WR, Hewinson RG, Wheeler PR (2002). Functional genomics reveals the sole sulphate transporter of the mycobacterium tuberculosis complex and its relevance to the acquisition of sulphate in vivo. Molecular Microbiology, 43(3), 653-663.
Wiker HG, Michell SL, Hewinson RG, Spierings E, Nagai S, Harboe M (1999). Cloning, expression and significance of MPT53 for identification of secreted proteins of Mycobacterium tuberculosis.
Microbial Pathogenesis,
26(4), 207-219.
Abstract:
Cloning, expression and significance of MPT53 for identification of secreted proteins of Mycobacterium tuberculosis
Based on our N-terminal amino acid sequence of MPT53 and a deduced DNA sequence, we searched for the corresponding gene in the Mycobacterium tuberculosis genomic sequence at the Sanger centre, localizing mpt53 close to mpt70 and mpt83. The gene was cloned and expressed, followed by purification of MPT53 to homogeneity from recombinant M. smegmatis culture fluid. In MPT53 there is 60% identity with the active site of thioredoxin of M. tuberculosis (MPT46) with two cysteins in a CXXC motif, but MPT53 could not serve as an alternative substrate for thioredoxin reductase. Testing for IgM and IgG1 anti-MPT53 in cattle sera showed that MPT53 is immunogenic following natural and experimental infection with M. bovis. Cloning of mpt53 represents cloning of the last of the 10 proteins originally defined as 'secreted proteins' of M. tuberculosis and M. bovis based on determination of their 'Localization index' (LI). The need for a precise definition of the term 'secreted protein' is discussed. So far we have observed full concordance between occurrence of an LI value indicating secretion of a protein and occurrence of a signal sequence in the corresponding gene. Signal sequence independent protein secretion in mycobacteria may occur for a limited number of proteins and remains to be established.
Abstract.
Hewinson RG, Michell SL, Russell WP, McAdam RA, Jacobs WR (1996). Molecular characterization of MPT83: a seroreactive antigen of Mycobacterium tuberculosis with homology to MPT70.
Scand J Immunol,
43(5), 490-499.
Abstract:
Molecular characterization of MPT83: a seroreactive antigen of Mycobacterium tuberculosis with homology to MPT70.
The Mycobacterium bovis antigens MPB70 and MPB83 are homologous cross-reactive proteins. It has been reported previously that MPB83 is glycosylated and exists in two forms with apparent molecular masses of 23kDa and 25kDa, whereas the apparent molecular mass of MPB70 is 22kDa. Using a monoclonal antibody, SB10, which recognizes an epitope common to both MPB70 and MPB83, we compared the expression of these proteins in M. bovis BCG, virulent M. bovis and virulent Mycobacterium tuberculosis by Western blotting of bacterial lysates. The previously described pattern of high and low producing substrains of BCG for MPB70 was also applicable for MPB83. Virulent M. bovis was found to express high levels of MPB70 and MPB83. Immunoblotting experiments using sera from Balb/c mice infected with live M. tuberculosis H37Rv revealed that although the MPB83 homologue of M. tuberculosis, MPT83, is expressed at low levels in M. tuberculosis when grown in vitro, the protein is highly immunogenic during infection with live bacteria. A clone from a mycobacterial shuttle cosmid library of M. tuberculosis H37Rv was isolated which expressed both MPT70 and MPT83. Genetic analysis of this cosmid revealed that MPT70 and MPT83 were encoded by separate genes with the gene encoding MPT83 situated 2.4kb upstream of mpt70. Both genes are transcribed in the same direction. The gene encoding MPT83 was cloned and DNA sequencing revealed an open reading frame of 660bp encoding a protein with a predicted molecular mass of 22kDa. Recombinant MPT83 was expressed in Escherichia coli from the native AUG initiation codon by translational coupling. In E. coli MPT83 was expressed as a 23kDa antigen whereas in the rapid growing mycobacterium Mycobacterium smegmatis the protein was expressed as a 25kDa protein indicating post-translational modification of the protein by M. smegmatis. In recombinant M. smegmatis MPT83 was predominantly cell associated whereas MPT70 was secreted into the culture medium. Amino acid sequence comparison between MPT83 and MPT70 revealed a 61% identity between the proteins, although little homology was apparent at the amino terminus. In MPT83 this region contained a typical lipoprotein signal peptide cleavage motif and a putative signal motif for O glycosylation. Both these motifs were absent from the amino acid sequence of MPT70.
Abstract.
Author URL.
Chapters
Michell SL, Griffin, K.F. Titball, R.W. (2006). Tularemia Pathogenesis and Immunity. In Friedman H, Anderson B (Eds.) Microbial Infection and Bioterrorism, Kluwer Academic Publishers, 121-138.
Conferences
Fraser D, Keedwell EC, Michell S, Sheridan R (2019). EMOCS: Evolutionary Multi-objective Optimisation for Clinical Scorecard Generation. Genetic and Evolutionary Computation Conference, GECCO 2019. 13th - 17th Jul 2019.
Publications by year
In Press
Masoli JAH, Jeffries A, Temperton B, Auckland C, Michelsen M, Warwick-Dugdale J, Manley R, Farbos A, Ellard S, Knight B, et al (In Press). Viral genetic sequencing identifies staff transmission of COVID-19 is important in a community hospital outbreak.
Abstract:
Viral genetic sequencing identifies staff transmission of COVID-19 is important in a community hospital outbreak
AbstractBackgroundWe have successfully used whole-genome sequencing to provide additional information for transmission pathways in infectious spread. We report and interpret genomic sequencing results in clinical context from a large outbreak of COVID-19 with 46 cases across staff and patients in a community hospital in the UK.MethodsFollowing multiple symptomatic cases within a two-week period, all staff and patients were screened by RT-PCR and staff subsequently had serology tests.ResultsThirty staff (25%) and 16 patients (62%) tested positive for COVID-19. Genomic sequencing data showed significant overlap of viral haplotypes in staff who had overlapping shift patterns. Patient haplotypes were more distinct from each other but had overlap with staff haplotypes.ConclusionsThis study includes clinical and genomic epidemiological detail that demonstrates the value of a combined approach. Viral genetic sequencing has identified that staff transmission of COVID-19 was important in this community hospital outbreak.Key pointsDetailed analysis of a large community hospital outbreak in older adults and staff with concurrent clinical and genomic data, including working patterns.Staff transmission was important in this community hospital outbreak.We found plausible associations between staff and patient cases.
Abstract.
2023
Cotton S, McHugh MP, Dewar R, Haas JG, Templeton K, Consortium TCGU, Robson SC, Connor TR, Loman NJ, Golubchik T, et al (2023). Investigation of hospital discharge cases and SARS-CoV-2 introduction into Lothian care homes. Journal of Hospital Infection, 135, 28-36.
Kennedy NA, Janjua M, Chanchlani N, Lin S, Bewshea C, Nice R, McDonald TJ, Auckland C, Harries LW, Davies M, et al (2023). Vaccine escape, increased breakthrough and reinfection in infliximab-treated patients with IBD during the Omicron wave of the SARS-CoV-2 pandemic.
Gut,
72(2), 295-305.
Abstract:
Vaccine escape, increased breakthrough and reinfection in infliximab-treated patients with IBD during the Omicron wave of the SARS-CoV-2 pandemic.
OBJECTIVE: Antitumour necrosis factor (TNF) drugs impair serological responses following SARS-CoV-2 vaccination. We sought to assess if a third dose of a messenger RNA (mRNA)-based vaccine substantially boosted anti-SARS-CoV-2 antibody responses and protective immunity in infliximab-treated patients with IBD. DESIGN: Third dose vaccine induced anti-SARS-CoV-2 spike (anti-S) receptor-binding domain (RBD) antibody responses, breakthrough SARS-CoV-2 infection, reinfection and persistent oropharyngeal carriage in patients with IBD treated with infliximab were compared with a reference cohort treated with vedolizumab from the impaCt of bioLogic therApy on saRs-cov-2 Infection and immuniTY (CLARITY) IBD study. RESULTS: Geometric mean (SD) anti-S RBD antibody concentrations increased in both groups following a third dose of an mRNA-based vaccine. However, concentrations were lower in patients treated with infliximab than vedolizumab, irrespective of whether their first two primary vaccine doses were ChAdOx1 nCoV-19 (1856 U/mL (5.2) vs 10 728 U/mL (3.1), p
Abstract.
Author URL.
2022
Baker S, Dougan G, Hess C, Kingston N, Lehner PJ, Lyons PA, Matheson NJ, Owehand WH, Saunders C, Summers C, et al (2022). Author Correction: SARS-CoV-2 evolution during treatment of chronic infection. Nature, 608(7922), e23-e23.
Baker S, Dougan G, Hess C, Kingston N, Lehner PJ, Lyons PA, Matheson NJ, Owehand WH, Saunders C, Summers C, et al (2022). Author Correction: Sensitivity of SARS-CoV-2 B.1.1.7 to mRNA vaccine-elicited antibodies. Nature, 608(7922), e24-e24.
Kläser K, Molteni E, Graham M, Canas LS, Österdahl MF, Antonelli M, Chen L, Deng J, Murray B, Kerfoot E, et al (2022). COVID-19 due to the B.1.617.2 (Delta) variant compared to B.1.1.7 (Alpha) variant of SARS-CoV-2: a prospective observational cohort study.
Scientific Reports,
12(1).
Abstract:
COVID-19 due to the B.1.617.2 (Delta) variant compared to B.1.1.7 (Alpha) variant of SARS-CoV-2: a prospective observational cohort study
The Delta (B.1.617.2) variant was the predominant UK circulating SARS-CoV-2 strain between May and December 2021. How Delta infection compares with previous variants is unknown. This prospective observational cohort study assessed symptomatic adults participating in the app-based COVID Symptom Study who tested positive for SARS-CoV-2 from May 26 to July 1, 2021 (Delta overwhelmingly the predominant circulating UK variant), compared (1:1, age- and sex-matched) with individuals presenting from December 28, 2020 to May 6, 2021 (Alpha (B.1.1.7) the predominant variant). We assessed illness (symptoms, duration, presentation to hospital) during Alpha- and Delta-predominant timeframes; and transmission, reinfection, and vaccine effectiveness during the Delta-predominant period. 3581 individuals (aged 18 to 100 years) from each timeframe were assessed. The seven most frequent symptoms were common to both variants. Within the first 28 days of illness, some symptoms were more common with Delta versus Alpha infection (including fever, sore throat, and headache) and some vice versa (dyspnoea). Symptom burden in the first week was higher with Delta versus Alpha infection; however, the odds of any given symptom lasting ≥ 7 days was either lower or unchanged. Illness duration ≥ 28 days was lower with Delta versus Alpha infection, though unchanged in unvaccinated individuals. Hospitalisation for COVID-19 was unchanged. The Delta variant appeared more (1.49) transmissible than Alpha. Re-infections were low in all UK regions. Vaccination markedly reduced the risk of Delta infection (by 69-84%). We conclude that COVID-19 from Delta or Alpha infections is similar. The Delta variant is more transmissible than Alpha; however, current vaccines showed good efficacy against disease. This research framework can be useful for future comparisons with new emerging variants.
Abstract.
Aggarwal D, Page AJ, Schaefer U, Savva GM, Myers R, Volz E, Ellaby N, Platt S, Groves N, Gallagher E, et al (2022). Genomic assessment of quarantine measures to prevent SARS-CoV-2 importation and transmission. Nature Communications, 13(1).
Aggarwal D, Warne B, Jahun AS, Hamilton WL, Fieldman T, du Plessis L, Hill V, Blane B, Watkins E, Wright E, et al (2022). Genomic epidemiology of SARS-CoV-2 in a UK university identifies dynamics of transmission.
Nature Communications,
13(1).
Abstract:
Genomic epidemiology of SARS-CoV-2 in a UK university identifies dynamics of transmission
AbstractUnderstanding SARS-CoV-2 transmission in higher education settings is important to limit spread between students, and into at-risk populations. In this study, we sequenced 482 SARS-CoV-2 isolates from the University of Cambridge from 5 October to 6 December 2020. We perform a detailed phylogenetic comparison with 972 isolates from the surrounding community, complemented with epidemiological and contact tracing data, to determine transmission dynamics. We observe limited viral introductions into the university; the majority of student cases were linked to a single genetic cluster, likely following social gatherings at a venue outside the university. We identify considerable onward transmission associated with student accommodation and courses; this was effectively contained using local infection control measures and following a national lockdown. Transmission clusters were largely segregated within the university or the community. Our study highlights key determinants of SARS-CoV-2 transmission and effective interventions in a higher education setting that will inform public health policy during pandemics.
Abstract.
Nickbakhsh S, Hughes J, Christofidis N, Griffiths E, Shaaban S, Enright J, Smollett K, Nomikou K, Palmalux N, Tong L, et al (2022). Genomic epidemiology of SARS-CoV-2 in a university outbreak setting and implications for public health planning.
Scientific Reports,
12(1).
Abstract:
Genomic epidemiology of SARS-CoV-2 in a university outbreak setting and implications for public health planning
Whole genome sequencing of SARS-CoV-2 has occurred at an unprecedented scale, and can be exploited for characterising outbreak risks at the fine-scale needed to inform control strategies. One setting at continued risk of COVID-19 outbreaks are higher education institutions, associated with student movements at the start of term, close living conditions within residential halls, and high social contact rates. Here we analysed SARS-CoV-2 whole genome sequences in combination with epidemiological data to investigate a large cluster of student cases associated with University of Glasgow accommodation in autumn 2020, Scotland. We identified 519 student cases of SARS-CoV-2 infection associated with this large cluster through contact tracing data, with 30% sequencing coverage for further analysis. We estimated at least 11 independent introductions of SARS-CoV-2 into the student population, with four comprising the majority of detected cases and consistent with separate outbreaks. These four outbreaks were curtailed within a week following implementation of control measures. The impact of student infections on the local community was short-term despite an underlying increase in community infections. Our study highlights the need for context-specific information in the formation of public health policy for higher educational settings.
Abstract.
Twohig KA, Nyberg T, Zaidi A, Thelwall S, Sinnathamby MA, Aliabadi S, Seaman SR, Harris RJ, Hope R, Lopez-Bernal J, et al (2022). Hospital admission and emergency care attendance risk for SARS-CoV-2 delta (B.1.617.2) compared with alpha (B.1.1.7) variants of concern: a cohort study.
The Lancet Infectious Diseases,
22(1), 35-42.
Abstract:
Hospital admission and emergency care attendance risk for SARS-CoV-2 delta (B.1.617.2) compared with alpha (B.1.1.7) variants of concern: a cohort study
Background: the SARS-CoV-2 delta (B.1.617.2) variant was first detected in England in March, 2021. It has since rapidly become the predominant lineage, owing to high transmissibility. It is suspected that the delta variant is associated with more severe disease than the previously dominant alpha (B.1.1.7) variant. We aimed to characterise the severity of the delta variant compared with the alpha variant by determining the relative risk of hospital attendance outcomes. Methods: This cohort study was done among all patients with COVID-19 in England between March 29 and May 23, 2021, who were identified as being infected with either the alpha or delta SARS-CoV-2 variant through whole-genome sequencing. Individual-level data on these patients were linked to routine health-care datasets on vaccination, emergency care attendance, hospital admission, and mortality (data from Public Health England's Second Generation Surveillance System and COVID-19-associated deaths dataset; the National Immunisation Management System; and NHS Digital Secondary Uses Services and Emergency Care Data Set). The risk for hospital admission and emergency care attendance were compared between patients with sequencing-confirmed delta and alpha variants for the whole cohort and by vaccination status subgroups. Stratified Cox regression was used to adjust for age, sex, ethnicity, deprivation, recent international travel, area of residence, calendar week, and vaccination status. Findings: Individual-level data on 43 338 COVID-19-positive patients (8682 with the delta variant, 34 656 with the alpha variant; median age 31 years [IQR 17–43]) were included in our analysis. 196 (2·3%) patients with the delta variant versus 764 (2·2%) patients with the alpha variant were admitted to hospital within 14 days after the specimen was taken (adjusted hazard ratio [HR] 2·26 [95% CI 1·32–3·89]). 498 (5·7%) patients with the delta variant versus 1448 (4·2%) patients with the alpha variant were admitted to hospital or attended emergency care within 14 days (adjusted HR 1·45 [1·08–1·95]). Most patients were unvaccinated (32 078 [74·0%] across both groups). The HRs for vaccinated patients with the delta variant versus the alpha variant (adjusted HR for hospital admission 1·94 [95% CI 0·47–8·05] and for hospital admission or emergency care attendance 1·58 [0·69–3·61]) were similar to the HRs for unvaccinated patients (2·32 [1·29–4·16] and 1·43 [1·04–1·97]; p=0·82 for both) but the precision for the vaccinated subgroup was low. Interpretation: This large national study found a higher hospital admission or emergency care attendance risk for patients with COVID-19 infected with the delta variant compared with the alpha variant. Results suggest that outbreaks of the delta variant in unvaccinated populations might lead to a greater burden on health-care services than the alpha variant. Funding: Medical Research Council; UK Research and Innovation; Department of Health and Social Care; and National Institute for Health Research.
Abstract.
Fletcher J, Porter R, Boulton Z, Brown L, Knight B, Romanczuk L, Aiken S, Delury C, Michell S (2022). In vitro efficacy of antibiotic loaded calcium sulfate beads (Stimulan Rapid Cure) against polymicrobial communities and individual bacterial strains derived from diabetic foot infections.
J Med Microbiol,
71(5).
Abstract:
In vitro efficacy of antibiotic loaded calcium sulfate beads (Stimulan Rapid Cure) against polymicrobial communities and individual bacterial strains derived from diabetic foot infections.
Introduction. Diabetic foot infection (DFI) is the main reason for diabetes-related hospitalisation and is a major cause of diabetes-related amputation. DFIs are often complicated by ischaemia in the affected limb, the presence of polymicrobial biofilms and increasingly the occurrence of antibiotic resistant bacteria.Hypothesis/Gap statement. Antibiotic loaded beads could inhibit the growth of polymicrobial DFI communities with differing compositions in vitro.Aim. This study investigates the in vitro efficacy of antibiotic loaded calcium sulfate beads (Stimulan Rapid Cure, Biocomposites Ltd. UK) against polymicrobial DFI communities and individual bacterial strains derived from DFIs.Methodology. Debrided tissue obtained from the base of infected diabetic foot ulcers was homogenised and spread over the surface of Columbia blood agar (CBA) and fastidious anaerobe agar (FAA) plates. Calcium sulfate beads containing a combination of vancomycin and gentamicin were then placed on the surface of the agar and following incubation, zones of inhibition (ZOI) were measured. For individual bacterial strains isolated from the infected tissue, calcium sulfate beads containing vancomycin, gentamicin, flucloxacillin or rifampicin and beads containing a combination of vancomycin and gentamicin or flucloxacillin and rifampicin were tested for their ability to inhibit growth.Results. Calcium sulfate beads loaded with a combination of vancomycin and gentamicin were able to inhibit bacterial growth from all polymicrobial tissue homogenates tested, with ZOI diameters ranging from 15 to 40 mm. In the case of individual bacterial strains, beads containing combinations of vancomycin and gentamicin or flucloxacillin and rifampicin were able to produce ZOI with Gram-positive facultatitive anaerobic strains such as Staphylococcus aureus and Enterococcus faecalis, Gram-negative facultative anaerobic strains such as Pseudomonas aeruginosa and obligate anaerobic strains such as Finegoldia magna even where acquired resistance to one of the antibiotics in the combination was evidenced.Conclusion. The local use of calcium sulfate beads containing a combination of two antibiotics demonstrated high efficacy against polymicrobial DFI communities and individual DFI bacterial strains in in vitro zone of inhibition tests. These results show promise for clinical application, but further research and clinical studies are required.
Abstract.
Author URL.
Robson SC, Connor TR, Loman NJ, Golubchik T, Martinez Nunez RT, Bonsall D, Rambaut A, Snell LB, Ludden C, Corden S, et al (2022). Publisher Correction: Genomic reconstruction of the SARS CoV-2 epidemic in England. Nature, 606(7915), e18-e18.
Willett BJ, Grove J, MacLean OA, Wilkie C, De Lorenzo G, Furnon W, Cantoni D, Scott S, Logan N, Ashraf S, et al (2022). SARS-CoV-2 Omicron is an immune escape variant with an altered cell entry pathway.
Nature Microbiology,
7(8), 1161-1179.
Abstract:
SARS-CoV-2 Omicron is an immune escape variant with an altered cell entry pathway
Vaccines based on the spike protein of SARS-CoV-2 are a cornerstone of the public health response to COVID-19. The emergence of hypermutated, increasingly transmissible variants of concern (VOCs) threaten this strategy. Omicron (B.1.1.529), the fifth VOC to be described, harbours multiple amino acid mutations in spike, half of which lie within the receptor-binding domain. Here we demonstrate substantial evasion of neutralization by Omicron BA.1 and BA.2 variants in vitro using sera from individuals vaccinated with ChAdOx1, BNT162b2 and mRNA-1273. These data were mirrored by a substantial reduction in real-world vaccine effectiveness that was partially restored by booster vaccination. The Omicron variants BA.1 and BA.2 did not induce cell syncytia in vitro and favoured a TMPRSS2-independent endosomal entry pathway, these phenotypes mapping to distinct regions of the spike protein. Impaired cell fusion was determined by the receptor-binding domain, while endosomal entry mapped to the S2 domain. Such marked changes in antigenicity and replicative biology may underlie the rapid global spread and altered pathogenicity of the Omicron variant.
Abstract.
Ashford F, Best A, Dunn SJ, Ahmed Z, Siddiqui H, Melville J, Wilkinson S, Mirza J, Cumley N, Stockton J, et al (2022). SARS-CoV-2 Testing in the Community: Testing Positive Samples with the TaqMan SARS-CoV-2 Mutation Panel to Find Variants in Real Time. Journal of Clinical Microbiology, 60(4), e02408-e02421.
2021
Volz E, Mishra S, Chand M, Barrett JC, Johnson R, Geidelberg L, Hinsley WR, Laydon DJ, Dabrera G, Toole AO, et al (2021). Assessing transmissibility of SARS-CoV-2 lineage B.1.1.7 in England.
NATURE,
593(7858), 266-+.
Author URL.
Graham MS, Sudre CH, May A, Antonelli M, Murray B, Varsavsky T, Kläser K, Canas LS, Molteni E, Modat M, et al (2021). Changes in symptomatology, reinfection, and transmissibility associated with the SARS-CoV-2 variant B.1.1.7: an ecological study. The Lancet Public Health, 6(5), e335-e345.
Volz E, Hill V, McCrone JT, Price A, Jorgensen D, O’Toole Á, Southgate J, Johnson R, Jackson B, Nascimento FF, et al (2021). Evaluating the Effects of SARS-CoV-2 Spike Mutation D614G on Transmissibility and Pathogenicity. Cell, 184(1), 64-75.e11.
Elliott P, Haw D, Wang H, Eales O, Walters CE, Ainslie KEC, Atchison C, Fronterre C, Diggle PJ, Page AJ, et al (2021). Exponential growth, high prevalence of SARS-CoV-2, and vaccine effectiveness associated with the Delta variant.
Science,
374(6574).
Abstract:
Exponential growth, high prevalence of SARS-CoV-2, and vaccine effectiveness associated with the Delta variant.
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections were rising during early summer 2021 in many countries as a result of the Delta variant. We assessed reverse transcription polymerase chain reaction swab positivity in the Real-time Assessment of Community Transmission–1 (REACT-1) study in England. During June and July 2021, we observed sustained exponential growth with an average doubling time of 25 days, driven by complete replacement of the Alpha variant by Delta and by high prevalence at younger, less-vaccinated ages. Prevalence among unvaccinated people [1.21% (95% credible interval 1.03%, 1.41%)] was three times that among double-vaccinated people [0.40% (95% credible interval 0.34%, 0.48%)]. However, after adjusting for age and other variables, vaccine effectiveness for double-vaccinated people was estimated at between ~50% and ~60% during this period in England. Increased social mixing in the presence of Delta had the potential to generate sustained growth in infections, even at high levels of vaccination.
Abstract.
Author URL.
Vöhringer HS, Sanderson T, Sinnott M, De Maio N, Nguyen T, Goater R, Schwach F, Harrison I, Hellewell J, Ariani CV, et al (2021). Genomic reconstruction of the SARS-CoV-2 epidemic in England.
Nature,
600(7889), 506-511.
Abstract:
Genomic reconstruction of the SARS-CoV-2 epidemic in England
AbstractThe evolution of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus leads to new variants that warrant timely epidemiological characterization. Here we use the dense genomic surveillance data generated by the COVID-19 Genomics UK Consortium to reconstruct the dynamics of 71 different lineages in each of 315 English local authorities between September 2020 and June 2021. This analysis reveals a series of subepidemics that peaked in early autumn 2020, followed by a jump in transmissibility of the B.1.1.7/Alpha lineage. The Alpha variant grew when other lineages declined during the second national lockdown and regionally tiered restrictions between November and December 2020. A third more stringent national lockdown suppressed the Alpha variant and eliminated nearly all other lineages in early 2021. Yet a series of variants (most of which contained the spike E484K mutation) defied these trends and persisted at moderately increasing proportions. However, by accounting for sustained introductions, we found that the transmissibility of these variants is unlikely to have exceeded the transmissibility of the Alpha variant. Finally, B.1.617.2/Delta was repeatedly introduced in England and grew rapidly in early summer 2021, constituting approximately 98% of sampled SARS-CoV-2 genomes on 26 June 2021.
Abstract.
Kemp SA, Collier DA, Datir RP, Ferreira IATM, Gayed S, Jahun A, Hosmillo M, Rees-Spear C, Mlcochova P, Lumb IU, et al (2021). SARS-CoV-2 evolution during treatment of chronic infection.
Nature,
592(7853), 277-282.
Abstract:
SARS-CoV-2 evolution during treatment of chronic infection
The spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is critical for virus infection through the engagement of the human ACE2 protein1 and is a major antibody target. Here we show that chronic infection with SARS-CoV-2 leads to viral evolution and reduced sensitivity to neutralizing antibodies in an immunosuppressed individual treated with convalescent plasma, by generating whole-genome ultra-deep sequences for 23 time points that span 101 days and using in vitro techniques to characterize the mutations revealed by sequencing. There was little change in the overall structure of the viral population after two courses of remdesivir during the first 57 days. However, after convalescent plasma therapy, we observed large, dynamic shifts in the viral population, with the emergence of a dominant viral strain that contained a substitution (D796H) in the S2 subunit and a deletion (ΔH69/ΔV70) in the S1 N-terminal domain of the spike protein. As passively transferred serum antibodies diminished, viruses with the escape genotype were reduced in frequency, before returning during a final, unsuccessful course of convalescent plasma treatment. In vitro, the spike double mutant bearing both ΔH69/ΔV70 and D796H conferred modestly decreased sensitivity to convalescent plasma, while maintaining infectivity levels that were similar to the wild-type virus.The spike substitution mutant D796H appeared to be the main contributor to the decreased susceptibility to neutralizing antibodies, but this mutation resulted in an infectivity defect. The spike deletion mutant ΔH69/ΔV70 had a twofold higher level of infectivity than wild-type SARS-CoV-2, possibly compensating for the reduced infectivity of the D796H mutation. These data reveal strong selection on SARS-CoV-2 during convalescent plasma therapy, which is associated with the emergence of viral variants that show evidence of reduced susceptibility to neutralizing antibodies in immunosuppressed individuals.
Abstract.
Collier DA, De Marco A, Ferreira IATM, Meng B, Datir RP, Walls AC, Kemp SA, Bassi J, Pinto D, Silacci-Fregni C, et al (2021). Sensitivity of SARS-CoV-2 B.1.1.7 to mRNA vaccine-elicited antibodies.
NATURE,
593(7857), 136-+.
Author URL.
de Silva TI, Liu G, Lindsey BB, Dong D, Moore SC, Hsu NS, Shah D, Wellington D, Mentzer AJ, Angyal A, et al (2021). The impact of viral mutations on recognition by SARS-CoV-2 specific T cells.
iScience,
24(11).
Abstract:
The impact of viral mutations on recognition by SARS-CoV-2 specific T cells
We identify amino acid variants within dominant SARS-CoV-2 T cell epitopes by interrogating global sequence data. Several variants within nucleocapsid and ORF3a epitopes have arisen independently in multiple lineages and result in loss of recognition by epitope-specific T cells assessed by IFN-γ and cytotoxic killing assays. Complete loss of T cell responsiveness was seen due to Q213K in the A∗01:01-restricted CD8+ ORF3a epitope FTSDYYQLY207-215; due to P13L, P13S, and P13T in the B∗27:05-restricted CD8+ nucleocapsid epitope QRNAPRITF9-17; and due to T362I and P365S in the A∗03:01/A∗11:01-restricted CD8+ nucleocapsid epitope KTFPPTEPK361-369. CD8+ T cell lines unable to recognize variant epitopes have diverse T cell receptor repertoires. These data demonstrate the potential for T cell evasion and highlight the need for ongoing surveillance for variants capable of escaping T cell as well as humoral immunity.
Abstract.
2020
Hubert CL, Michell SL (2020). A universal oyster infection model demonstrates that Vibrio vulnificus Type 6 secretion systems have antibacterial activity in vivo.
Environ Microbiol,
22(10), 4381-4393.
Abstract:
A universal oyster infection model demonstrates that Vibrio vulnificus Type 6 secretion systems have antibacterial activity in vivo.
With the rapid increase of aquaculture contributing to sustainable food security, comes the need to better understand seafood associated diseases. One of the major aquatic bacterial genera responsible for human infections from seafood is Vibrio, especially from oysters. Currently, in vivo study of bacterial interactions within oysters is limited by the inability to promote high-level uptake of bacteria by oysters. This study has therefore evolved current natural marine snow protocols to generate 'artificial' marine snow, into which bacteria can be incorporated to facilitate extensive uptake by oysters. This presents an adaptable model for bacterial study within filter-feeding shellfish. Using this model, we demonstrate for the first time the antibacterial activity of Vibrio vulnificus Type 6 secretion systems in vivo, revealing an important role for the T6SS in V. vulnificus ecology.
Abstract.
Author URL.
Aanensen DM, Abudahab K, Adams A, Afifi S, Alam MT, Alderton A, Alikhan N-F, Allan J, Almsaud M, Alrezaihi A, et al (2020). An integrated national scale SARS-CoV-2 genomic surveillance network.
LANCET MICROBE,
1(3), E99-E100.
Author URL.
Hubert C (2020). Characterising the role of Vibrio vulnificus type 6 secretion systems 1 and 2 in an in vivo oyster model.
Abstract:
Characterising the role of Vibrio vulnificus type 6 secretion systems 1 and 2 in an in vivo oyster model
Vibrio vulnificus is a significant human pathogen commonly isolated from temperate marine environments, where it is particularly abundant within filter-feeding shellfish. V. vulnificus is currently increasing in prevalence, theorised to be due to climate change facilitating V. vulnificus growth in previously inhospitable environments. Infection of susceptible individuals with V. vulnificus typically results in either primary septicaemia or necrotic wound infection, depending upon the route of entry, and frequently results in death if not treated rapidly.
Two type 6 secretion systems (T6SS) have been identified in V. vulnificus, termed the T6SS1 and the T6SS2. The T6SS is a molecular syringe utilised to inject cytotoxic effector proteins into neighbouring cells. Whilst the T6SS2 is present in all sequenced
V. vulnificus strains, only a subset possesses the T6SS1. Previous bacterial co-culture killing assays between T6SS1+ and T6SS1- V. vulnificus strains demonstrated thermoregulated T6SS1-mediated killing of T6SS1- strains. This study further characterised the role of both the T6SS1 and the T6SS2 in vitro. In vitro co-culture assays demonstrated that both the T6SS1 and the T6SS2 have antibacterial killing activity at the environmentally representative temperature of 21 °C. This is the first characterised role for the T6SS2 of V. vulnificus. No anti-eukaryotic activity was observed following co-culture with the phagocytic amoeba, Dictyostelium discoideum, suggesting that T6SS activity is purely antibacterial.
In vitro bacterial co-culture assays were replicated in vivo using an oyster model. To facilitate high-level uptake of bacterial strains of interest by oysters, an artificial marine snow model was developed where bacteria were incorporated into easily ingested phytoplankton aggregates. Uptake of bacteria from artificial marine snow was extremely successful, resulting in bacterial loads within oysters significantly greater than achieved by any study to date. Using this model, this study was able to demonstrate that V. vulnificus utilises both the T6SS1 and the T6SS2 to target and kill neighbouring bacteria, in both an intra and inter-species manner. This data suggests that the T6SSs of V. vulnificus play a key role in V. vulnificus ecology and the dynamics between bacterial populations in vivo.
Abstract.
2019
Fraser D, Keedwell EC, Michell S, Sheridan R (2019). EMOCS: Evolutionary Multi-objective Optimisation for Clinical Scorecard Generation. Genetic and Evolutionary Computation Conference, GECCO 2019. 13th - 17th Jul 2019.
Bradshaw WJ, Bruxelle J-F, Kovacs-Simon A, Harmer NJ, Janoir C, Péchiné S, Acharya KR, Michell SL (2019). Molecular features of lipoprotein CD0873: a potential vaccine against the human pathogen Clostridioides difficile.
J Biol Chem,
294(43), 15850-15861.
Abstract:
Molecular features of lipoprotein CD0873: a potential vaccine against the human pathogen Clostridioides difficile.
Clostridioides difficile is the primary cause of antibiotic-associated diarrhea and colitis, a healthcare-associated intestinal disease resulting in a significant fatality rate. Colonization of the gut is critical for C. difficile pathogenesis. The bacterial molecules essential for efficient colonization therefore offer great potential as vaccine candidates. Here we present findings demonstrating that the C. difficile immunogenic lipoprotein CD0873 plays a critical role in pathogen success in vivo We found that in a dixenic colonization model, a CD0873-positive strain of C. difficile significantly outcompeted a CD0873-negative strain. Immunization of mice with recombinant CD0873 prevented long-term gut colonization and was correlated with a strong secretory IgA immune response. We further present high-resolution crystal structures of CD0873, at 1.35-2.50 Å resolutions, offering a first view of the ligand-binding pocket of CD0873 and provide evidence that this lipoprotein adhesin is part of a tyrosine import system, an amino acid key in C. difficile infection. These findings suggest that CD0873 could serve as an effective component in a vaccine against C. difficile.
Abstract.
Author URL.
Fletcher J, Porter R, Aiken S, Delury C, Michell S (2019). The effect of local release antibiotic beads on in-vitro bacterial growth from tissue taken from infected diabetic foot ulcers. Access Microbiology, 1(1A).
2016
Church SR, Lux T, Baker-Austin C, Buddington SP, Michell SL (2016). Vibrio vulnificus Type 6 Secretion System 1 Contains Anti-Bacterial Properties.
PLoS One,
11(10).
Abstract:
Vibrio vulnificus Type 6 Secretion System 1 Contains Anti-Bacterial Properties.
Vibrio vulnificus is a bacterium responsible for severe gastroenteritis, sepsis and wound infections. Gastroenteritis and sepsis are commonly associated with the consumption of raw oysters, whereas wound infection is often associated with the handling of contaminated fish. Although classical virulence factors of this emerging pathogen are well characterised, there remains a paucity of knowledge regarding the general biology of this species. To investigate the presence of previously unreported virulence factors, we applied whole genome sequencing to a panel of ten V. vulnificus strains with varying virulence potentials. This identified two novel type 6 secretion systems (T6SSs), systems that are known to have a role in bacterial virulence and population dynamics. By utilising a range of molecular techniques and assays we have demonstrated the functionality of one of these T6SSs. Furthermore, we have shown that this system is subject to thermoregulation and is negatively regulated by increasing salinity concentrations. This secretion system was also shown to be involved in the killing of V. vulnificus strains that did not possess this system and a model is proposed as to how this interaction may contribute to population dynamics within V. vulnificus strains. In addition to this intra-species killing, this system also contributes to the killing of inter bacterial species and may have a role in the general composition of Vibrio species in the environment.
Abstract.
Author URL.
2015
Nandi T, Holden MTG, Didelot X, Mehershahi K, Boddey JA, Beacham I, Peak I, Harting J, Baybayan P, Guo Y, et al (2015). Burkholderia pseudomallei sequencing identifies genomic clades with distinct recombination, accessory, and epigenetic profiles.
Genome Res,
25(4).
Author URL.
Nandi T, Holden MTG, Didelot X, Mehershahi K, Boddey JA, Beacham I, Peak I, Harting J, Baybayan P, Guo Y, et al (2015). Burkholderia pseudomallei sequencing identifies genomic clades with distinct recombination, accessory, and epigenetic profiles.
Genome Res,
25(1), 129-141.
Abstract:
Burkholderia pseudomallei sequencing identifies genomic clades with distinct recombination, accessory, and epigenetic profiles.
Burkholderia pseudomallei (Bp) is the causative agent of the infectious disease melioidosis. To investigate population diversity, recombination, and horizontal gene transfer in closely related Bp isolates, we performed whole-genome sequencing (WGS) on 106 clinical, animal, and environmental strains from a restricted Asian locale. Whole-genome phylogenies resolved multiple genomic clades of Bp, largely congruent with multilocus sequence typing (MLST). We discovered widespread recombination in the Bp core genome, involving hundreds of regions associated with multiple haplotypes. Highly recombinant regions exhibited functional enrichments that may contribute to virulence. We observed clade-specific patterns of recombination and accessory gene exchange, and provide evidence that this is likely due to ongoing recombination between clade members. Reciprocally, interclade exchanges were rarely observed, suggesting mechanisms restricting gene flow between clades. Interrogation of accessory elements revealed that each clade harbored a distinct complement of restriction-modification (RM) systems, predicted to cause clade-specific patterns of DNA methylation. Using methylome sequencing, we confirmed that representative strains from separate clades indeed exhibit distinct methylation profiles. Finally, using an E. coli system, we demonstrate that Bp RM systems can inhibit uptake of non-self DNA. Our data suggest that RM systems borne on mobile elements, besides preventing foreign DNA invasion, may also contribute to limiting exchanges of genetic material between individuals of the same species. Genomic clades may thus represent functional units of genetic isolation in Bp, modulating intraspecies genetic diversity.
Abstract.
Author URL.
Nandi T, Holden MTG, Didelot X, Mehershahi K, Boddey JA, Beacham I, Peak I, Harting J, Baybayan P, Guo Y, et al (2015). Errata: Burkholderia pseudomallei sequencing identifies genomic clades with distinct recombination, accessory, and epigenetic profiles (Genome Research (2015) 25 (129-141)). Genome Research, 25(4).
Aiken SS, Cooper JJ, Florance H, Robinson MT, Michell S (2015). Local Release of Antibiotics for Surgical Site Infection Management Using High-Purity Calcium Sulfate: an in Vitro Elution Study.
SURGICAL INFECTIONS,
16(1), 54-61.
Author URL.
Charlton TM, Kovacs-Simon A, Michell SL, Fairweather NF, Tate EW (2015). Quantitative Lipoproteomics in Clostridium difficile Reveals a Role for Lipoproteins in Sporulation.
CHEMISTRY & BIOLOGY,
22(11), 1562-1573.
Author URL.
2014
Peano C, Chiaramonte F, Motta S, Pietrelli A, Jaillon S, Rossi E, Consolandi C, Champion OL, Michell SL, Freddi L, et al (2014). Gene and protein expression in response to different growth temperatures and oxygen availability in Burkholderia thailandensis.
PLoS One,
9(3).
Abstract:
Gene and protein expression in response to different growth temperatures and oxygen availability in Burkholderia thailandensis.
Burkholderia thailandensis, although normally avirulent for mammals, can infect macrophages in vitro and has occasionally been reported to cause pneumonia in humans. It is therefore used as a model organism for the human pathogen B. pseudomallei, to which it is closely related phylogenetically. We characterized the B. thailandensis clinical isolate CDC2721121 (BtCDC272) at the genome level and studied its response to environmental cues associated with human host colonization, namely, temperature and oxygen limitation. Effects of the different growth conditions on BtCDC272 were studied through whole genome transcription studies and analysis of proteins associated with the bacterial cell surface. We found that growth at 37°C, compared to 28°C, negatively affected cell motility and flagella production through a mechanism involving regulation of the flagellin-encoding fliC gene at the mRNA stability level. Growth in oxygen-limiting conditions, in contrast, stimulated various processes linked to virulence, such as lipopolysaccharide production and expression of genes encoding protein secretion systems. Consistent with these observations, BtCDC272 grown in oxygen limitation was more resistant to phagocytosis and strongly induced the production of inflammatory cytokines from murine macrophages. Our results suggest that, while temperature sensing is important for regulation of B. thailandensis cell motility, oxygen limitation has a deeper impact on its physiology and constitutes a crucial environmental signal for the production of virulence factors.
Abstract.
Author URL.
Kovacs-Simon A, Leuzzi R, Kasendra M, Minton N, Titball RW, Michell SL (2014). Lipoprotein CD0873 is a novel adhesin of Clostridium difficile.
J Infect Dis,
210(2), 274-284.
Abstract:
Lipoprotein CD0873 is a novel adhesin of Clostridium difficile.
Clostridium difficile is a cause of antibiotic-associated diarrhea and colitis, a healthcare-associated intestinal disease. Colonization of the gut is a critical step in the course of infection. The C. difficile lipoprotein CD0873 was identified as a putative adhesin through a bioinformatics approach. Surface exposure of CD0873 was confirmed and a CD0873 mutant was generated. The CD0873 mutant showed a significant reduction in adherence to Caco-2 cells and wild-type bacteria preincubated with anti-CD0873 antibodies showed significantly decreased adherence to Caco-2 cells. In addition, we demonstrated that purified recombinant CD0873 protein alone associates with Caco-2 cells. This is the first definitive identification of a C. difficile adhesin, which now allows work to devise improved measures for preventing and treating disease.
Abstract.
Author URL.
2013
Butt E, Foster JAH, Keedwell E, Bell JEA, Titball RW, Bhangu A, Michell SL, Sheridan R (2013). Derivation and validation of a simple, accurate and robust prediction rule for risk of mortality in patients with Clostridium difficile infection.
BMC Infect Dis,
13Abstract:
Derivation and validation of a simple, accurate and robust prediction rule for risk of mortality in patients with Clostridium difficile infection.
BACKGROUND: Clostridium difficile infection poses a significant healthcare burden. However, the derivation of a simple, evidence based prediction rule to assist patient management has not yet been described. METHOD: Univariate, multivariate and decision tree procedures were used to deduce a prediction rule from over 186 variables; retrospectively collated from clinical data for 213 patients. The resulting prediction rule was validated on independent data from a cohort of 158 patients described by Bhangu et al. (Colorectal Disease, 12(3):241-246, 2010). RESULTS: Serum albumin levels (g/L) (P = 0.001), respiratory rate (resps /min) (P = 0.002), C-reactive protein (mg/L) (P = 0.034) and white cell count (mcL) (P = 0.049) were predictors of all-cause mortality. Threshold levels of serum albumin ≤ 24.5 g/L, C- reactive protein >228 mg/L, respiratory rate >17 resps/min and white cell count >12 × 10(3) mcL were associated with an increased risk of all-cause mortality. A simple four variable prediction rule was devised based on these threshold levels and when tested on the initial data, yield an area under the curve score of 0.754 (P
Abstract.
Author URL.
He M, Miyajima F, Roberts P, Ellison L, Pickard DJ, Martin MJ, Connor TR, Harris SR, Fairley D, Bamford KB, et al (2013). Emergence and global spread of epidemic healthcare-associated Clostridium difficile.
Nature Genetics,
45(1), 109-113.
Abstract:
Emergence and global spread of epidemic healthcare-associated Clostridium difficile
Epidemic C. difficile (027/BI/NAP1) has rapidly emerged in the past decade as the leading cause of antibiotic-associated diarrhea worldwide. However, the key events in evolutionary history leading to its emergence and the subsequent patterns of global spread remain unknown. Here, we define the global population structure of C. difficile 027/BI/NAP1 using whole-genome sequencing and phylogenetic analysis. We show that two distinct epidemic lineages, FQR1 and FQR2, not one as previously thought, emerged in North America within a relatively short period after acquiring the same fluoroquinolone resistance-conferring mutation and a highly related conjugative transposon. The two epidemic lineages showed distinct patterns of global spread, and the FQR2 lineage spread more widely, leading to healthcare-associated outbreaks in the UK, continental Europe and Australia. Our analysis identifies key genetic changes linked to the rapid transcontinental dissemination of epidemic C. difficile 027/BI/NAP1 and highlights the routes by which it spreads through the global healthcare system. © 2013 Nature America, Inc. All rights reserved.
Abstract.
Gourlay LJ, Peri C, Ferrer-Navarro M, Conchillo-Solé O, Gori A, Rinchai D, Thomas RJ, Champion OL, Michell SL, Kewcharoenwong C, et al (2013). Exploiting the Burkholderia pseudomallei acute phase antigen BPSL2765 for structure-based epitope discovery/design in structural vaccinology.
Chem Biol,
20(9), 1147-1156.
Abstract:
Exploiting the Burkholderia pseudomallei acute phase antigen BPSL2765 for structure-based epitope discovery/design in structural vaccinology.
We solved the crystal structure of Burkholderia pseudomallei acute phase antigen BPSL2765 in the context of a structural vaccinology study, in the area of melioidosis vaccine development. Based on the structure, we applied a recently developed method for epitope design that combines computational epitope predictions with in vitro mapping experiments and successfully identified a consensus sequence within the antigen that, when engineered as a synthetic peptide, was selectively immunorecognized to the same extent as the recombinant protein in sera from melioidosis-affected subjects. Antibodies raised against the consensus peptide were successfully tested in opsonization bacterial killing experiments and antibody-dependent agglutination tests of B. pseudomallei. Our strategy represents a step in the development of immunodiagnostics, in the production of specific antibodies and in the optimization of antigens for vaccine development, starting from structural and physicochemical principles.
Abstract.
Author URL.
2012
Michell SL (2012). A better understanding of what makes some proteins "fat".
J Bacteriol,
194(9), 2129-2130.
Author URL.
He M, Miyajima F, Roberts P, Ellison L, Pickard DJ, Martin MJ, Connor TR, Harris SR, Fairley D, Bamford KB, et al (2012). Emergence and global spread of epidemic healthcare-associated Clostridium difficile. Nature Genetics
Foster JAH, Butt JEC, Bell J, Goff A, Morgan C, Hancock J, Carmichael C, Keedwell EC, Michell SLI, Sheridan RP, et al (2012). IMPROVING CLINICAL MANAGEMENT IN CLOSTRIDIUM DIFFICILE: FAECAL CALPROTECTIN DOES NOT PREDICT SEVERITY, RECURRENCE OR MORTALITY.
AGE AND AGEING,
41, 72-72.
Author URL.
2011
Thomas RM, Twine SM, Fulton KM, Tessier L, Kilmury SLN, Ding W, Harmer N, Michell SL, Oyston PCF, Titball RW, et al (2011). Glycosylation of DsbA in Francisella tularensis subsp. tularensis.
J Bacteriol,
193(19), 5498-5509.
Abstract:
Glycosylation of DsbA in Francisella tularensis subsp. tularensis.
In Francisella tularensis subsp. tularensis, DsbA has been shown to be an essential virulence factor and has been observed to migrate to multiple protein spots on two-dimensional electrophoresis gels. In this work, we show that the protein is modified with a 1,156-Da glycan moiety in O-linkage. The results of mass spectrometry studies suggest that the glycan is a hexasaccharide, comprised of N-acetylhexosamines, hexoses, and an unknown monosaccharide. Disruption of two genes within the FTT0789-FTT0800 putative polysaccharide locus, including a galE homologue (FTT0791) and a putative glycosyltransferase (FTT0798), resulted in loss of glycan modification of DsbA. The F. tularensis subsp. tularensis ΔFTT0798 and ΔFTT0791::Cm mutants remained virulent in the murine model of subcutaneous tularemia. This indicates that glycosylation of DsbA does not play a major role in virulence under these conditions. This is the first report of the detailed characterization of the DsbA glycan and putative role of the FTT0789-FTT0800 gene cluster in glycan biosynthesis.
Abstract.
Author URL.
Kovacs-Simon A, Titball RW, Michell SL (2011). Lipoproteins of bacterial pathogens.
Infect Immun,
79(2), 548-561.
Abstract:
Lipoproteins of bacterial pathogens.
Bacterial lipoproteins are a set of membrane proteins with many different functions. Due to this broad-ranging functionality, these proteins have a considerable significance in many phenomena, from cellular physiology through cell division and virulence. Here we give a general overview of lipoprotein biogenesis and highlight examples of the roles of lipoproteins in bacterial disease caused by a selection of medically relevant Gram-negative and Gram-positive pathogens: Mycobacterium tuberculosis, Streptococcus pneumoniae, Borrelia burgdorferi, and Neisseria meningitidis. Lipoproteins have been shown to play key roles in adhesion to host cells, modulation of inflammatory processes, and translocation of virulence factors into host cells. As such, a number of lipoproteins have been shown to be potential vaccines. This review provides a summary of some of the reported roles of lipoproteins and of how this knowledge has been exploited in some cases for the generation of novel countermeasures to bacterial diseases.
Abstract.
Author URL.
Wand ME, Müller CM, Titball RW, Michell SL (2011). Macrophage and Galleria mellonella infection models reflect the virulence of naturally occurring isolates of B. pseudomallei, B. thailandensis and B. oklahomensis.
BMC Microbiol,
11(1).
Abstract:
Macrophage and Galleria mellonella infection models reflect the virulence of naturally occurring isolates of B. pseudomallei, B. thailandensis and B. oklahomensis.
BACKGROUND: Burkholderia pseudomallei is the causative agent of melioidosis, a tropical disease of humans with a variable and often fatal outcome. In murine models of infection, different strains exhibit varying degrees of virulence. In contrast, two related species, B. thailandensis and B. oklahomensis, are highly attenuated in mice. Our aim was to determine whether virulence in mice is reflected in macrophage or wax moth larvae (Galleria mellonella) infection models. RESULTS: B. pseudomallei strains 576 and K96243, which have low median lethal dose (MLD) values in mice, were able to replicate and induce cellular damage in macrophages and caused rapid death of G. mellonella. In contrast, B. pseudomallei strain 708a, which is attenuated in mice, showed reduced replication in macrophages, negligible cellular damage and was avirulent in G. mellonella larvae. B. thailandensis isolates were less virulent than B. pseudomallei in all of the models tested. However, we did record strain dependent differences. B. oklahomensis isolates were the least virulent isolates. They showed minimal ability to replicate in macrophages, were unable to evoke actin-based motility or to form multinucleated giant cells and were markedly attenuated in G. mellonella compared to B. thailandensis. CONCLUSIONS: We have shown that the alternative infection models tested here, namely macrophages and Galleria mellonella, are able to distinguish between strains of B. pseudomallei, B. thailandensis and B. oklahomensis and that these differences reflect the observed virulence in murine infection models. Our results indicate that B. oklahomensis is the least pathogenic of the species investigated. They also show a correlation between isolates of B. thailandensis associated with human infection and virulence in macrophage and Galleria infection models.
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Author URL.
Wang W, Hale C, Goulding D, Haslam SM, Tissot B, Lindsay C, Michell S, Titball R, Yu J, Toribio AL, et al (2011). Mannosidase 2, alpha 1 deficiency is associated with ricin resistance in embryonic stem (ES) cells.
PLoS One,
6(8).
Abstract:
Mannosidase 2, alpha 1 deficiency is associated with ricin resistance in embryonic stem (ES) cells.
Host gene products required for mediating the action of toxins are potential targets for reversing or controlling their pathogenic impact following exposure. To identify such targets libraries of insertional gene-trap mutations generated with a PiggyBac transposon in Blm-deficient embryonic stem cells were exposed to the plant toxin, ricin. Resistant clones were isolated and genetically characterised and one was found to be a homozygous mutant of the mannosidase 2, alpha 1 (Man2α1) locus with a matching defect in the homologous allele. The causality of the molecular lesion was confirmed by removal of the transposon following expression of PB-transposase. Comparative glycomic and lectin binding analysis of the Man2α1 (-/-) ricin resistant cells revealed an increase in the levels of hybrid glycan structures and a reduction in terminal β-galactose moieties, potential target receptors for ricin. Furthermore, naïve ES cells treated with inhibitors of the N-linked glycosylation pathway at the mannosidase 2, alpha 1 step exhibited either full or partial resistance to ricin. Therefore, we conclusively identified mannosidase 2, alpha 1 deficiency to be associated with ricin resistance.
Abstract.
Author URL.
Vanaporn M, Wand M, Michell SL, Sarkar-Tyson M, Ireland P, Goldman S, Kewcharoenwong C, Rinchai D, Lertmemongkolchai G, Titball RW, et al (2011). Superoxide dismutase C is required for intracellular survival and virulence of Burkholderia pseudomallei.
Microbiology (Reading),
157(Pt 8), 2392-2400.
Abstract:
Superoxide dismutase C is required for intracellular survival and virulence of Burkholderia pseudomallei.
Burkholderia pseudomallei is an intracellular pathogen and the causative agent of melioidosis, a life-threatening disease of humans. Within host cells, superoxide is an important mediator of pathogen killing. In this study, we have identified the B. pseudomallei K96243 sodC gene, shown that it has superoxide dismutase activity, and constructed an allelic deletion mutant of this gene. Compared with the wild-type, the mutant was more sensitive to killing by extracellular superoxide, but not to superoxide generated intracellularly. The sodC mutant showed a markedly decreased survival in J774A.1 mouse macrophages, and reduced numbers of bacteria were recovered from human polymorphonuclear neutrophils (PMNs) when compared with the wild-type. The numbers of wild-type or mutant bacteria recovered from human diabetic neutrophils were significantly lower than from normal human neutrophils. The sodC mutant was attenuated in BALB/c mice. Our results indicate that SodC plays a key role in the virulence of B. pseudomallei, but that diabetics are not more susceptible to infection because of a reduced ability of PMNs to kill by superoxide.
Abstract.
Author URL.
2010
Michell SL, Dean RE, Eyles JE, Hartley MG, Waters E, Prior JL, Titball RW, Oyston PCF (2010). Deletion of the Bacillus anthracis capB homologue in Francisella tularensis subspecies tularensis generates an attenuated strain that protects mice against virulent tularaemia.
J Med Microbiol,
59(Pt 11), 1275-1284.
Abstract:
Deletion of the Bacillus anthracis capB homologue in Francisella tularensis subspecies tularensis generates an attenuated strain that protects mice against virulent tularaemia.
As there is currently no licensed vaccine against Francisella tularensis, the causative agent of tularaemia, the bacterium is an agent of concern as a potential bioweapon. Although F. tularensis has a low infectious dose and high associated mortality, it possesses few classical virulence factors. An analysis of the F. tularensis subspecies tularensis genome sequence has revealed the presence of a region containing genes with low sequence homology to part of the capBCADE operon of Bacillus anthracis. We have generated an isogenic capB mutant of F. tularensis subspecies tularensis SchuS4 and shown it to be attenuated. Furthermore, using BALB/c mice, we have demonstrated that this capB strain affords protection against significant homologous challenge with the wild-type strain. These data have important implications for the development of a defined and efficacious tularaemia vaccine.
Abstract.
Author URL.
Forslund A-L, Salomonsson EN, Golovliov I, Kuoppa K, Michell S, Titball R, Oyston P, Noppa L, Sjöstedt A, Forsberg A, et al (2010). The type IV pilin, PilA, is required for full virulence of Francisella tularensis subspecies tularensis.
BMC Microbiol,
10Abstract:
The type IV pilin, PilA, is required for full virulence of Francisella tularensis subspecies tularensis.
BACKGROUND: all four Francisella tularensis subspecies possess gene clusters with potential to express type IV pili (Tfp). These clusters include putative pilin genes, as well as pilB, pilC and pilQ, required for secretion and assembly of Tfp. A hallmark of Tfp is the ability to retract the pilus upon surface contact, a property mediated by the ATPase PilT. Interestingly, out of the two major human pathogenic subspecies only the highly virulent type a strains have a functional pilT gene. RESULTS: in a previous study, we were able to show that one pilin gene, pilA, was essential for virulence of a type B strain in a mouse infection model. In this work we have examined the role of several Tfp genes in the virulence of the pathogenic type a strain SCHU S4. pilA, pilC, pilQ, and pilT were mutated by in-frame deletion mutagenesis. Interestingly, when mice were infected with a mixture of each mutant strain and the wild-type strain, the pilA, pilC and pilQ mutants were out-competed, while the pilT mutant was equally competitive as the wild-type. CONCLUSIONS: This suggests that expression and surface localisation of PilA contribute to virulence in the highly virulent type a strain, while PilT was dispensable for virulence in the mouse infection model.
Abstract.
Author URL.
2009
Champion MD, Zeng Q, Nix EB, Nano FE, Keim P, Kodira CD, Borowsky M, Young S, Koehrsen M, Engels R, et al (2009). Comparative genomic characterization of Francisella tularensis strains belonging to low and high virulence subspecies.
PLoS Pathog,
5(5).
Abstract:
Comparative genomic characterization of Francisella tularensis strains belonging to low and high virulence subspecies.
Tularemia is a geographically widespread, severely debilitating, and occasionally lethal disease in humans. It is caused by infection by a gram-negative bacterium, Francisella tularensis. In order to better understand its potency as an etiological agent as well as its potential as a biological weapon, we have completed draft assemblies and report the first complete genomic characterization of five strains belonging to the following different Francisella subspecies (subsp.): the F. tularensis subsp. tularensis FSC033, F. tularensis subsp. holarctica FSC257 and FSC022, and F. tularensis subsp. novicida GA99-3548 and GA99-3549 strains. Here, we report the sequencing of these strains and comparative genomic analysis with recently available public Francisella sequences, including the rare F. tularensis subsp. mediasiatica FSC147 strain isolate from the Central Asian Region. We report evidence for the occurrence of large-scale rearrangement events in strains of the holarctica subspecies, supporting previous proposals that further phylogenetic subdivisions of the Type B clade are likely. We also find a significant enrichment of disrupted or absent ORFs proximal to predicted breakpoints in the FSC022 strain, including a genetic component of the Type I restriction-modification defense system. Many of the pseudogenes identified are also disrupted in the closely related rarely human pathogenic F. tularensis subsp. mediasiatica FSC147 strain, including modulator of drug activity B (mdaB) (FTT0961), which encodes a known NADPH quinone reductase involved in oxidative stress resistance. We have also identified genes exhibiting sequence similarity to effectors of the Type III (T3SS) and components of the Type IV secretion systems (T4SS). One of the genes, msrA2 (FTT1797c), is disrupted in F. tularensis subsp. mediasiatica and has recently been shown to mediate bacterial pathogen survival in host organisms. Our findings suggest that in addition to the duplication of the Francisella Pathogenicity Island, and acquisition of individual loci, adaptation by gene loss in the more recently emerged tularensis, holarctica, and mediasiatica subspecies occurred and was distinct from evolutionary events that differentiated these subspecies, and the novicida subspecies, from a common ancestor. Our findings are applicable to future studies focused on variations in Francisella subspecies pathogenesis, and of broader interest to studies of genomic pathoadaptation in bacteria.
Abstract.
Author URL.
2008
Richards MI, Michell SL, Oyston PCF (2008). An intracellularly inducible gene involved in virulence and polyphosphate production in Francisella.
J Med Microbiol,
57(Pt 10), 1183-1192.
Abstract:
An intracellularly inducible gene involved in virulence and polyphosphate production in Francisella.
Francisella tularensis is an intracellular pathogen capable of multiplying to high levels in macrophages. By protein analysis, only a few proteins have been shown previously to be expressed at high levels in macrophages relative to bacteria grown in culture media. To identify additional genes that show increased expression during intracellular growth, we developed a plasmid for use in Francisella based on the induction of expression of green fluorescent protein. Clones of F. tularensis subsp. novicida were identified that were fluorescent only intracellularly and not when grown in vitro. Sequencing identified a range of genes comprising some such as dnaK that are already known to be expressed intracellularly and some novel targets. One of these newly identified regulated genes, FTN1472/FTT1564, was selected for further study. Isogenic mutants were generated in F. tularensis subsp. novicida and subsp. tularensis by allelic replacement. Inactivation of the gene resulted in abolition of polyphosphate production by F. novicida, strongly supporting the bioinformatic analysis, which had suggested that the gene may encode a polyphosphate kinase. The mutants exhibited defects for intracellular growth in macrophages and were attenuated in mice, indicating a key role for the putative polyphosphate kinase in the virulence of Francisella.
Abstract.
Author URL.
2007
Michell, S.L. Diaper, H. Wikstrom, P. Titball, RW (2007). A 55 kDa hypothetical membrane protein is an iron-regulated virulence factor of Francisella tularensis subspecies novicida U112. Journal of Medical Microbiology, 56, 1268-1276.
Isherwood, K.E. Michell, S.L. Diaper, H. Titball, RW (2007). A Francisella tularensis subspecies novicida purF mutant, but not a purA mutant, induces protective immunity to tularemia in mice. Vaccine, 25, 2011-2018.
sm202, Grice ID, Griffin K, Hitchen PG (2007). The immunologically distinct O-antigens from Francisella tularensis subspecies tuarensis and Francisella novicida are both virulence determinants and protective antigens. Infection and Immunity, 75(1), 371-378.
2006
sm202, Bystrom A, Forslund AL, Johansson A (2006). Direct repeat-mediated deletion of a type IV pilin gene results in major virulence attenuation of Francisella tularensis. Molecular Microbiology, 59(6), 1818-1830.
Michell SL, Griffin, K.F. Titball, R.W. (2006). Tularemia Pathogenesis and Immunity. In Friedman H, Anderson B (Eds.) Microbial Infection and Bioterrorism, Kluwer Academic Publishers, 121-138.
2005
RT101, Andersson SGE, Chain P, Chu MC (2005). The complete genome sequence of Francisella tularensis, the causative agent of tularemia. Nature Genetics, 37(2), 153-159.
2003
Michell SL, Whelan AO, Wheeler PR, Panico M, Easton RL, Etienne AT, Haslam SM, Dell A, Morris HR, Reason AJ, et al (2003). The MPB83 antigen from Mycobacterium bovis contains O-linked mannose and (1->3)-mannibiose moieties. Journal of Biological Chemistry, 278(18), 16423-16432.
2002
Wooff EE, Michell SL, Gordon SV, Chambers MA, Bardarov A, Jacobs Jr WR, Hewinson RG, Wheeler PR (2002). Functional genomics reveals the sole sulphate transporter of the mycobacterium tuberculosis complex and its relevance to the acquisition of sulphate in vivo. Molecular Microbiology, 43(3), 653-663.
1999
Wiker HG, Michell SL, Hewinson RG, Spierings E, Nagai S, Harboe M (1999). Cloning, expression and significance of MPT53 for identification of secreted proteins of Mycobacterium tuberculosis.
Microbial Pathogenesis,
26(4), 207-219.
Abstract:
Cloning, expression and significance of MPT53 for identification of secreted proteins of Mycobacterium tuberculosis
Based on our N-terminal amino acid sequence of MPT53 and a deduced DNA sequence, we searched for the corresponding gene in the Mycobacterium tuberculosis genomic sequence at the Sanger centre, localizing mpt53 close to mpt70 and mpt83. The gene was cloned and expressed, followed by purification of MPT53 to homogeneity from recombinant M. smegmatis culture fluid. In MPT53 there is 60% identity with the active site of thioredoxin of M. tuberculosis (MPT46) with two cysteins in a CXXC motif, but MPT53 could not serve as an alternative substrate for thioredoxin reductase. Testing for IgM and IgG1 anti-MPT53 in cattle sera showed that MPT53 is immunogenic following natural and experimental infection with M. bovis. Cloning of mpt53 represents cloning of the last of the 10 proteins originally defined as 'secreted proteins' of M. tuberculosis and M. bovis based on determination of their 'Localization index' (LI). The need for a precise definition of the term 'secreted protein' is discussed. So far we have observed full concordance between occurrence of an LI value indicating secretion of a protein and occurrence of a signal sequence in the corresponding gene. Signal sequence independent protein secretion in mycobacteria may occur for a limited number of proteins and remains to be established.
Abstract.
1996
Hewinson RG, Michell SL, Russell WP, McAdam RA, Jacobs WR (1996). Molecular characterization of MPT83: a seroreactive antigen of Mycobacterium tuberculosis with homology to MPT70.
Scand J Immunol,
43(5), 490-499.
Abstract:
Molecular characterization of MPT83: a seroreactive antigen of Mycobacterium tuberculosis with homology to MPT70.
The Mycobacterium bovis antigens MPB70 and MPB83 are homologous cross-reactive proteins. It has been reported previously that MPB83 is glycosylated and exists in two forms with apparent molecular masses of 23kDa and 25kDa, whereas the apparent molecular mass of MPB70 is 22kDa. Using a monoclonal antibody, SB10, which recognizes an epitope common to both MPB70 and MPB83, we compared the expression of these proteins in M. bovis BCG, virulent M. bovis and virulent Mycobacterium tuberculosis by Western blotting of bacterial lysates. The previously described pattern of high and low producing substrains of BCG for MPB70 was also applicable for MPB83. Virulent M. bovis was found to express high levels of MPB70 and MPB83. Immunoblotting experiments using sera from Balb/c mice infected with live M. tuberculosis H37Rv revealed that although the MPB83 homologue of M. tuberculosis, MPT83, is expressed at low levels in M. tuberculosis when grown in vitro, the protein is highly immunogenic during infection with live bacteria. A clone from a mycobacterial shuttle cosmid library of M. tuberculosis H37Rv was isolated which expressed both MPT70 and MPT83. Genetic analysis of this cosmid revealed that MPT70 and MPT83 were encoded by separate genes with the gene encoding MPT83 situated 2.4kb upstream of mpt70. Both genes are transcribed in the same direction. The gene encoding MPT83 was cloned and DNA sequencing revealed an open reading frame of 660bp encoding a protein with a predicted molecular mass of 22kDa. Recombinant MPT83 was expressed in Escherichia coli from the native AUG initiation codon by translational coupling. In E. coli MPT83 was expressed as a 23kDa antigen whereas in the rapid growing mycobacterium Mycobacterium smegmatis the protein was expressed as a 25kDa protein indicating post-translational modification of the protein by M. smegmatis. In recombinant M. smegmatis MPT83 was predominantly cell associated whereas MPT70 was secreted into the culture medium. Amino acid sequence comparison between MPT83 and MPT70 revealed a 61% identity between the proteins, although little homology was apparent at the amino terminus. In MPT83 this region contained a typical lipoprotein signal peptide cleavage motif and a putative signal motif for O glycosylation. Both these motifs were absent from the amino acid sequence of MPT70.
Abstract.
Author URL.
