Publications by authors named "Gordon Dougan"

395 Publications

Functional analysis of colonization factor antigen I positive enterotoxigenic identifies genes implicated in survival in water and host colonization.

Microb Genom 2021 Jun;7(6)

Department of Medicine, University of Cambridge, Cambridge, UK.

Enterotoxigenic (ETEC) expressing the colonization pili CFA/I are common causes of diarrhoeal infections in humans. Here, we use a combination of transposon mutagenesis and transcriptomic analysis to identify genes and pathways that contribute to ETEC persistence in water environments and colonization of a mammalian host. ETEC persisting in water exhibit a distinct RNA expression profile from those growing in richer media. Multiple pathways were identified that contribute to water survival, including lipopolysaccharide biosynthesis and stress response regulons. The analysis also indicated that ETEC growing in mice encounter a bottleneck driving down the diversity of colonizing ETEC populations.
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http://dx.doi.org/10.1099/mgen.0.000554DOI Listing
June 2021

Longitudinal analysis reveals that delayed bystander CD8+ T cell activation and early immune pathology distinguish severe COVID-19 from mild disease.

Immunity 2021 06 16;54(6):1257-1275.e8. Epub 2021 May 16.

R&D Department, Hycult Biotech, 5405 PD Uden, the Netherlands.

The kinetics of the immune changes in COVID-19 across severity groups have not been rigorously assessed. Using immunophenotyping, RNA sequencing, and serum cytokine analysis, we analyzed serial samples from 207 SARS-CoV2-infected individuals with a range of disease severities over 12 weeks from symptom onset. An early robust bystander CD8 T cell immune response, without systemic inflammation, characterized asymptomatic or mild disease. Hospitalized individuals had delayed bystander responses and systemic inflammation that was already evident near symptom onset, indicating that immunopathology may be inevitable in some individuals. Viral load did not correlate with this early pathological response but did correlate with subsequent disease severity. Immune recovery is complex, with profound persistent cellular abnormalities in severe disease correlating with altered inflammatory responses, with signatures associated with increased oxidative phosphorylation replacing those driven by cytokines tumor necrosis factor (TNF) and interleukin (IL)-6. These late immunometabolic and immune defects may have clinical implications.
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http://dx.doi.org/10.1016/j.immuni.2021.05.010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8125900PMC
June 2021

High-Content Imaging to Phenotype Antimicrobial Effects on Individual Bacteria at Scale.

mSystems 2021 May 18;6(3). Epub 2021 May 18.

Cambridge Institute of Therapeutic Immunology & Infectious Disease, University of Cambridge Department of Medicine, Jeffrey Cheah Biomedical Centre, Cambridge, United Kingdom

High-content imaging (HCI) is a technique for screening multiple cells in high resolution to detect subtle morphological and phenotypic variation. The method has been commonly deployed on model eukaryotic cellular systems, often for screening new drugs and targets. HCI is not commonly utilized for studying bacterial populations but may be a powerful tool in understanding and combatting antimicrobial resistance. Consequently, we developed a high-throughput method for phenotyping bacteria under antimicrobial exposure at the scale of individual bacterial cells. Imaging conditions were optimized on an Opera Phenix confocal microscope (Perkin Elmer), and novel analysis pipelines were established for both Gram-negative bacilli and Gram-positive cocci. The potential of this approach was illustrated using isolates of , serovar Typhimurium, and HCI enabled the detection and assessment of subtle morphological characteristics, undetectable through conventional phenotypical methods, that could reproducibly distinguish between bacteria exposed to different classes of antimicrobials with distinct modes of action (MOAs). In addition, distinctive responses were observed between susceptible and resistant isolates. By phenotyping single bacterial cells, we observed intrapopulation differences, which may be critical in identifying persistence or emerging resistance during antimicrobial treatment. The work presented here outlines a comprehensive method for investigating morphological changes at scale in bacterial populations under specific perturbation. High-content imaging (HCI) is a microscopy technique that permits the screening of multiple cells simultaneously in high resolution to detect subtle morphological and phenotypic variation. The power of this methodology is that it can generate large data sets comprised of multiple parameters taken from individual cells subjected to a range of different conditions. We aimed to develop novel methods for using HCI to study bacterial cells exposed to a range of different antibiotic classes. Using an Opera Phenix confocal microscope (Perkin Elmer) and novel analysis pipelines, we created a method to study the morphological characteristics of , serovar Typhimurium, and when exposed to antibacterial drugs with differing modes of action. By imaging individual bacterial cells at high resolution and scale, we observed intrapopulation differences associated with different antibiotics. The outlined methods are highly relevant for how we begin to better understand and combat antimicrobial resistance.
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http://dx.doi.org/10.1128/mSystems.00028-21DOI Listing
May 2021

Mapping Rora expression in resting and activated CD4+ T cells.

PLoS One 2021 18;16(5):e0251233. Epub 2021 May 18.

EMBL-European Bioinformatics Institute, Wellcome Genome Campus, Hinxton, Cambridge, United Kingdom.

The transcription factor Rora has been shown to be important for the development of ILC2 and the regulation of ILC3, macrophages and Treg cells. Here we investigate the role of Rora across CD4+ T cells in general, but with an emphasis on Th2 cells, both in vitro as well as in the context of several in vivo type 2 infection models. We dissect the function of Rora using overexpression and a CD4-conditional Rora-knockout mouse, as well as a RORA-reporter mouse. We establish the importance of Rora in CD4+ T cells for controlling lung inflammation induced by Nippostrongylus brasiliensis infection, and have measured the effect on downstream genes using RNA-seq. Using a systematic stimulation screen of CD4+ T cells, coupled with RNA-seq, we identify upstream regulators of Rora, most importantly IL-33 and CCL7. Our data suggest that Rora is a negative regulator of the immune system, possibly through several downstream pathways, and is under control of the local microenvironment.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0251233PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8130942PMC
May 2021

A global resource for genomic predictions of antimicrobial resistance and surveillance of Salmonella Typhi at pathogenwatch.

Nat Commun 2021 05 17;12(1):2879. Epub 2021 May 17.

Centre for Genomic Pathogen Surveillance, Wellcome Genome Campus, Hinxton, Cambridgeshire, UK.

As whole-genome sequencing capacity becomes increasingly decentralized, there is a growing opportunity for collaboration and the sharing of surveillance data within and between countries to inform typhoid control policies. This vision requires free, community-driven tools that facilitate access to genomic data for public health on a global scale. Here we present the Pathogenwatch scheme for Salmonella enterica serovar Typhi (S. Typhi), a web application enabling the rapid identification of genomic markers of antimicrobial resistance (AMR) and contextualization with public genomic data. We show that the clustering of S. Typhi genomes in Pathogenwatch is comparable to established bioinformatics methods, and that genomic predictions of AMR are highly concordant with phenotypic susceptibility data. We demonstrate the public health utility of Pathogenwatch with examples selected from >4,300 public genomes available in the application. Pathogenwatch provides an intuitive entry point to monitor of the emergence and spread of S. Typhi high risk clones.
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http://dx.doi.org/10.1038/s41467-021-23091-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8128892PMC
May 2021

Long-read-sequenced reference genomes of the seven major lineages of enterotoxigenic Escherichia coli (ETEC) circulating in modern time.

Sci Rep 2021 Apr 29;11(1):9256. Epub 2021 Apr 29.

Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, Solna, Sweden.

Enterotoxigenic Escherichia coli (ETEC) is an enteric pathogen responsible for the majority of diarrheal cases worldwide. ETEC infections are estimated to cause 80,000 deaths annually, with the highest rates of burden, ca 75 million cases per year, amongst children under 5 years of age in resource-poor countries. It is also the leading cause of diarrhoea in travellers. Previous large-scale sequencing studies have found seven major ETEC lineages currently in circulation worldwide. We used PacBio long-read sequencing combined with Illumina sequencing to create high-quality complete reference genomes for each of the major lineages with manually curated chromosomes and plasmids. We confirm that the major ETEC lineages all harbour conserved plasmids that have been associated with their respective background genomes for decades, suggesting that the plasmids and chromosomes of ETEC are both crucial for ETEC virulence and success as pathogens. The in-depth analysis of gene content, synteny and correct annotations of plasmids will elucidate other plasmids with and without virulence factors in related bacterial species. These reference genomes allow for fast and accurate comparison between different ETEC strains, and these data will form the foundation of ETEC genomics research for years to come.
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http://dx.doi.org/10.1038/s41598-021-88316-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8085198PMC
April 2021

Evaluation of Typhoid Conjugate Vaccine Effectiveness in Ghana (TyVEGHA) Using a Cluster-Randomized Controlled Phase IV Trial: Trial Design and Population Baseline Characteristics.

Vaccines (Basel) 2021 Mar 19;9(3). Epub 2021 Mar 19.

International Vaccine Institute, Seoul 08826, Korea.

Typhoid fever remains a significant health problem in sub-Saharan Africa, with incidence rates of >100 cases per 100,000 person-years of observation. Despite the prequalification of safe and effective typhoid conjugate vaccines (TCV), some uncertainties remain around future demand. Real-life effectiveness data, which inform public health programs on the impact of TCVs in reducing typhoid-related mortality and morbidity, from an African setting may help encourage the introduction of TCVs in high-burden settings. Here, we describe a cluster-randomized trial to investigate population-level protection of TYPBAR-TCV, a Vi-polysaccharide conjugated to a tetanus-toxoid protein carrier (Vi-TT) against blood-culture-confirmed typhoid fever, and the synthesis of health economic evidence to inform policy decisions. A total of 80 geographically distinct clusters are delineated within the Agogo district of the Asante Akim region in Ghana. Clusters are randomized to the intervention arm receiving Vi-TT or a control arm receiving the meningococcal A conjugate vaccine. The primary study endpoint is the total protection of Vi-TT against blood-culture-confirmed typhoid fever. Total, direct, and indirect protection are measured as secondary outcomes. Blood-culture-based enhanced surveillance enables the estimation of incidence rates in the intervention and control clusters. Evaluation of the real-world impact of TCVs and evidence synthesis improve the uptake of prequalified/licensed safe and effective typhoid vaccines in public health programs of high burden settings. This trial is registered at the Pan African Clinical Trial Registry, accessible at Pan African Clinical Trials Registry (ID: PACTR202011804563392).
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http://dx.doi.org/10.3390/vaccines9030281DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8003794PMC
March 2021

Spatiotemporal persistence of multiple, diverse clades and toxins of Corynebacterium diphtheriae.

Nat Commun 2021 03 8;12(1):1500. Epub 2021 Mar 8.

Department of Medicine, Cambridge Institute of Therapeutic Immunology & Infectious Disease (CITIID), University of Cambridge, Cambridge, UK.

Diphtheria is a respiratory disease caused by the bacterium Corynebacterium diphtheriae. Although the development of a toxin-based vaccine in the 1930s has allowed a high level of control over the disease, cases have increased in recent years. Here, we describe the genomic variation of 502 C. diphtheriae isolates across 16 countries and territories over 122 years. We generate a core gene phylogeny and determine the presence of antimicrobial resistance genes and variation within the tox gene of 291 tox isolates. Numerous, highly diverse clusters of C. diphtheriae are observed across the phylogeny, each containing isolates from multiple countries, regions and time of isolation. The number of antimicrobial resistance genes, as well as the breadth of antibiotic resistance, is substantially greater in the last decade than ever before. We identified and analysed 18 tox gene variants, with mutations estimated to be of medium to high structural impact.
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http://dx.doi.org/10.1038/s41467-021-21870-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7940655PMC
March 2021

Genomic epidemiology of COVID-19 in care homes in the east of England.

Elife 2021 03 2;10. Epub 2021 Mar 2.

Wellcome Sanger Institute, Hinxton, United Kingdom.

COVID-19 poses a major challenge to care homes, as SARS-CoV-2 is readily transmitted and causes disproportionately severe disease in older people. Here, 1167 residents from 337 care homes were identified from a dataset of 6600 COVID-19 cases from the East of England. Older age and being a care home resident were associated with increased mortality. SARS-CoV-2 genomes were available for 700 residents from 292 care homes. By integrating genomic and temporal data, 409 viral clusters within the 292 homes were identified, indicating two different patterns - outbreaks among care home residents and independent introductions with limited onward transmission. Approximately 70% of residents in the genomic analysis were admitted to hospital during the study, providing extensive opportunities for transmission between care homes and hospitals. Limiting viral transmission within care homes should be a key target for infection control to reduce COVID-19 mortality in this population.
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http://dx.doi.org/10.7554/eLife.64618DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7997667PMC
March 2021

The Rab32/BLOC-3-dependent pathway mediates host defense against different pathogens in human macrophages.

Sci Adv 2021 Jan 15;7(3). Epub 2021 Jan 15.

Institute of Medical Sciences, University of Aberdeen, Foresterhill, Aberdeen AB252ZD, UK.

Macrophages provide a first line of defense against microorganisms, and while some mechanisms to kill pathogens such as the oxidative burst are well described, others are still undefined or unknown. Here, we report that the Rab32 guanosine triphosphatase and its guanine nucleotide exchange factor BLOC-3 (biogenesis of lysosome-related organelles complex-3) are central components of a trafficking pathway that controls both bacterial and fungal intracellular pathogens. This host-defense mechanism is active in both human and murine macrophages and is independent of well-known antimicrobial mechanisms such as the NADPH (reduced form of nicotinamide adenine dinucleotide phosphate)-dependent oxidative burst, production of nitric oxide, and antimicrobial peptides. To survive in human macrophages, Typhi actively counteracts the Rab32/BLOC-3 pathway through its pathogenicity island-1-encoded type III secretion system. These findings demonstrate that the Rab32/BLOC-3 pathway is a novel and universal host-defense pathway and protects mammalian species from various pathogens.
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http://dx.doi.org/10.1126/sciadv.abb1795DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7810368PMC
January 2021

Ventilator-associated pneumonia in critically ill patients with COVID-19.

Crit Care 2021 01 11;25(1):25. Epub 2021 Jan 11.

Division of Anaesthesia, Department of Medicine, University of Cambridge, Level 4, Addenbrooke's Hospital, Hills Road, Cambridge, CB2 0QQ, UK.

Background: Pandemic COVID-19 caused by the coronavirus SARS-CoV-2 has a high incidence of patients with severe acute respiratory syndrome (SARS). Many of these patients require admission to an intensive care unit (ICU) for invasive ventilation and are at significant risk of developing a secondary, ventilator-associated pneumonia (VAP).

Objectives: To study the incidence of VAP and bacterial lung microbiome composition of ventilated COVID-19 and non-COVID-19 patients.

Methods: In this retrospective observational study, we compared the incidence of VAP and secondary infections using a combination of microbial culture and a TaqMan multi-pathogen array. In addition, we determined the lung microbiome composition using 16S RNA analysis in a subset of samples. The study involved 81 COVID-19 and 144 non-COVID-19 patients receiving invasive ventilation in a single University teaching hospital between March 15th 2020 and August 30th 2020.

Results: COVID-19 patients were significantly more likely to develop VAP than patients without COVID (Cox proportional hazard ratio 2.01 95% CI 1.14-3.54, p = 0.0015) with an incidence density of 28/1000 ventilator days versus 13/1000 for patients without COVID (p = 0.009). Although the distribution of organisms causing VAP was similar between the two groups, and the pulmonary microbiome was similar, we identified 3 cases of invasive aspergillosis amongst the patients with COVID-19 but none in the non-COVID-19 cohort. Herpesvirade activation was also numerically more frequent amongst patients with COVID-19.

Conclusion: COVID-19 is associated with an increased risk of VAP, which is not fully explained by the prolonged duration of ventilation. The pulmonary dysbiosis caused by COVID-19, and the causative organisms of secondary pneumonia observed are similar to that seen in critically ill patients ventilated for other reasons.
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http://dx.doi.org/10.1186/s13054-021-03460-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7797892PMC
January 2021

A blueprint for the implementation of a validated approach for the detection of SARS-Cov2 in clinical samples in academic facilities.

Wellcome Open Res 2020 21;5:110. Epub 2020 Oct 21.

Cambridge Institute of Therapeutic Immunology & Infectious Disease (CITIID), Department of Medicine, University of Cambridge, Cambridge, UK.

The COVID-19 pandemic is expanding at an unprecedented rate. As a result, diagnostic services are stretched to their limit, and there is a clear need for the provision of additional diagnostic capacity. Academic laboratories, many of which are closed due to governmental lockdowns, may be in a position to support local screening capacity by adapting their current laboratory practices. Here, we describe the process of developing a SARS-Cov2 diagnostic workflow in a conventional academic Containment Level 2 laboratory. Our outline includes simple SARS-Cov2 deactivation upon contact, the method for a quantitative real-time reverse transcriptase PCR detecting SARS-Cov2, a description of process establishment and validation, and some considerations for establishing a similar workflow elsewhere. This was achieved under challenging circumstances through the collaborative efforts of scientists, clinical staff, and diagnostic staff to mitigate to the ongoing crisis. Within 14 days, we created a validated COVID-19 diagnostics service for healthcare workers in our local hospital. The described methods are not exhaustive, but we hope may offer support to other academic groups aiming to set up something comparable in a short time frame.
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http://dx.doi.org/10.12688/wellcomeopenres.15937.2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7590889PMC
October 2020

Gallbladder carriage generates genetic variation and genome degradation in Salmonella Typhi.

PLoS Pathog 2020 10 21;16(10):e1008998. Epub 2020 Oct 21.

Cambridge Institute of Therapeutic Immunology & Infectious Disease (CITIID) Department of Medicine, University of Cambridge, Cambridge, United Kingdom.

Despite recent advances in typhoid fever control, asymptomatic carriage of Salmonella Typhi in the gallbladder remains poorly understood. Aiming to understand if S. Typhi becomes genetically adapted for long-term colonisation in the gallbladder, we performed whole genome sequencing on a collection of S. Typhi isolated from the gallbladders of typhoid carriers. These sequences were compared to contemporaneously sampled sequences from organisms isolated from the blood of acute patients within the same population. We found that S. Typhi carriage was not restricted to any particular genotype or conformation of antimicrobial resistance genes, but was largely reflective of S. Typhi circulating in the general population. However, gallbladder isolates showed a higher genetic variability than acute isolates, with median pairwise SNP distances of 21 and 13 SNPs (p = 2.8x10-9), respectively. Within gallbladder isolates of the predominant H58 genotype, variation was associated with a higher prevalence of nonsense mutations. Notably, gallbladder isolates displayed a higher frequency of non-synonymous mutations in genes encoding hypothetical proteins, membrane lipoproteins, transport/binding proteins, surface antigens, and carbohydrate degradation. Specifically, we identified several gallbladder-specific non-synonymous mutations involved in LPS synthesis and modification, with some isolates lacking the Vi capsular polysaccharide vaccine target due to the 134Kb deletion of SPI-7. S. Typhi is under strong selective pressure in the human gallbladder, which may be reflected phylogenetically by long terminal branches that may distinguish organisms from chronic and acute infections. Our work shows that selective pressures asserted by the hostile environment of the human gallbladder generate new antigenic variants and raises questions regarding the role of carriage in the epidemiology of typhoid fever.
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http://dx.doi.org/10.1371/journal.ppat.1008998DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7605710PMC
October 2020

Multiple Introductions of Salmonella enterica Serovar Typhi H58 with Reduced Fluoroquinolone Susceptibility into Chile.

Emerg Infect Dis 2020 11;26(11):2736-2740

Salmonella enterica serovar Typhi H58, an antimicrobial-resistant lineage, is globally disseminated but has not been reported in Latin America. Genomic analysis revealed 3 independent introductions of Salmonella Typhi H58 with reduced fluoroquinolone susceptibility into Chile. Our findings highlight the utility of enhanced genomic surveillance for typhoid fever in this region.
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http://dx.doi.org/10.3201/eid2611.201676DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7588544PMC
November 2020

Prophage exotoxins enhance colonization fitness in epidemic scarlet fever-causing Streptococcus pyogenes.

Nat Commun 2020 10 6;11(1):5018. Epub 2020 Oct 6.

Australian Infectious Diseases Research Centre and School of Chemistry and Molecular Biosciences, The University of Queensland, St. Lucia, QLD, Australia.

The re-emergence of scarlet fever poses a new global public health threat. The capacity of North-East Asian serotype M12 (emm12) Streptococcus pyogenes (group A Streptococcus, GAS) to cause scarlet fever has been linked epidemiologically to the presence of novel prophages, including prophage ΦHKU.vir encoding the secreted superantigens SSA and SpeC and the DNase Spd1. Here, we report the molecular characterization of ΦHKU.vir-encoded exotoxins. We demonstrate that streptolysin O (SLO)-induced glutathione efflux from host cellular stores is a previously unappreciated GAS virulence mechanism that promotes SSA release and activity, representing the first description of a thiol-activated bacterial superantigen. Spd1 is required for resistance to neutrophil killing. Investigating single, double and triple isogenic knockout mutants of the ΦHKU.vir-encoded exotoxins, we find that SpeC and Spd1 act synergistically to facilitate nasopharyngeal colonization in a mouse model. These results offer insight into the pathogenesis of scarlet fever-causing GAS mediated by prophage ΦHKU.vir exotoxins.
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http://dx.doi.org/10.1038/s41467-020-18700-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7538557PMC
October 2020

Genomics of the Argentinian cholera epidemic elucidate the contrasting dynamics of epidemic and endemic Vibrio cholerae.

Nat Commun 2020 10 1;11(1):4918. Epub 2020 Oct 1.

Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, CB10 1SA, UK.

In order to control and eradicate epidemic cholera, we need to understand how epidemics begin, how they spread, and how they decline and eventually end. This requires extensive sampling of epidemic disease over time, alongside the background of endemic disease that may exist concurrently with the epidemic. The unique circumstances surrounding the Argentinian cholera epidemic of 1992-1998 presented an opportunity to do this. Here, we use 490 Argentinian V. cholerae genome sequences to characterise the variation within, and between, epidemic and endemic V. cholerae. We show that, during the 1992-1998 cholera epidemic, the invariant epidemic clone co-existed alongside highly diverse members of the Vibrio cholerae species in Argentina, and we contrast the clonality of epidemic V. cholerae with the background diversity of local endemic bacteria. Our findings refine and add nuance to our genomic definitions of epidemic and endemic cholera, and are of direct relevance to controlling current and future cholera epidemics.
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http://dx.doi.org/10.1038/s41467-020-18647-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7530988PMC
October 2020

Make it new: reformism and British public health.

Lancet Microbe 2020 Oct 3;1(6):e231-e232. Epub 2020 Sep 3.

Department of Medicine, University of Cambridge, Cambridge, UK.

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http://dx.doi.org/10.1016/S2666-5247(20)30143-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7470782PMC
October 2020

Point of Care Nucleic Acid Testing for SARS-CoV-2 in Hospitalized Patients: A Clinical Validation Trial and Implementation Study.

Cell Rep Med 2020 Aug 15;1(5):100062. Epub 2020 Jul 15.

Department of Infectious Diseases, Cambridge University NHS Hospitals Foundation Trust, Cambridge CB2 0QQ, UK.

There is an urgent need for rapid SARS-CoV-2 testing in hospitals to limit nosocomial spread. We report an evaluation of point of care (POC) nucleic acid amplification testing (NAAT) in 149 participants with parallel combined nasal and throat swabbing for POC versus standard lab RT-PCR testing. Median time to result is 2.6 (IQR 2.3-4.8) versus 26.4 h (IQR 21.4-31.4, p < 0.001), with 32 (21.5%) positive and 117 (78.5%) negative. Cohen's κ correlation between tests is 0.96 (95% CI 0.91-1.00). When comparing nearly 1,000 tests pre- and post-implementation, the median time to definitive bed placement from admission is 23.4 (8.6-41.9) versus 17.1 h (9.0-28.8), p = 0.02. Mean length of stay on COVID-19 "holding" wards is 58.5 versus 29.9 h (p < 0.001). POC testing increases isolation room availability, avoids bed closures, allows discharge to care homes, and expedites access to hospital procedures. POC testing could mitigate the impact of COVID-19 on hospital systems.
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http://dx.doi.org/10.1016/j.xcrm.2020.100062DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7362826PMC
August 2020

Protection conferred by typhoid fever against recurrent typhoid fever in urban Kolkata.

PLoS Negl Trop Dis 2020 08 17;14(8):e0008530. Epub 2020 Aug 17.

International Centre for Diarrheal Disease Research, Bangladesh, Dhaka, Bangladesh.

We evaluated the protection conferred by a first documented visit for clinical care of typhoid fever against recurrent typhoid fever prompting a visit. This study takes advantage of multi-year follow-up of a population with endemic typhoid participating in a cluster-randomized control trial of Vi capsular polysaccharide typhoid vaccine in Kolkata, India. A population of 70,566 individuals, of whom 37,673 were vaccinated with one dose of either Vi vaccine or a control (Hepatitis A) vaccine, were observed for four years. Surveillance detected 315 first typhoid visits, among whom 4 developed subsequent typhoid, 3 due to reinfection, defined using genomic criteria and corresponding to -124% (95% CI: -599, 28) protection by the initial illness. Point estimates of protection conferred by an initial illness were negative or negligible in both vaccinated and non-vaccinated subjects, though confidence intervals around the point estimates were wide. These data provide little support for a protective immunizing effect of clinically treated typhoid illness, though modest levels of protection cannot be excluded.
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http://dx.doi.org/10.1371/journal.pntd.0008530DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7430703PMC
August 2020

High relatedness of invasive multi-drug resistant non-typhoidal Salmonella genotypes among patients and asymptomatic carriers in endemic informal settlements in Kenya.

PLoS Negl Trop Dis 2020 08 3;14(8):e0008440. Epub 2020 Aug 3.

Cambridge Institute for Therapeutic Immunology & Infectious Disease, Department of Medicine, Cambridge University, Cambridge, United Kingdom.

Invasive Non-typhoidal Salmonella (iNTS) disease is a major public health challenge, especially in Sub-Saharan Africa (SSA). In Kenya, mortality rates are high (20-25%) unless prompt treatment is instituted. The most common serotypes are Salmonella enterica serotype Typhimurium (S. Typhimurium) and Salmonella enterica serotype Enteritidis (S. Enteritidis). In a 5 year case-control study in children residing in the Mukuru informal settlement in Nairobi, Kenya, a total of 4201 blood cultures from suspected iNTS cases and 6326 fecal samples from age-matched controls were studied. From the laboratory cultures we obtained a total of 133 S. Typhimurium isolates of which 83(62.4%) came from cases (53 blood and 30 fecal) and 50(37.6%) from controls (fecal). A total of 120 S. Enteritidis consisted of 70(58.3%) from cases (43 blood and 27 fecal) and 50(41.7%) from controls (fecal). The S. Typhimurium population fell into two distinct ST19 lineages constituting 36.1%, as well as ST313 lineage I (27.8%) and ST313 lineage II (36.1%) isolates. The S. Enteritidis isolates fell into the global epidemic lineage (46.6%), the Central/Eastern African lineage (30.5%), a novel Kenyan-specific lineage (12.2%) and a phylogenetically outlier lineage (10.7%). Detailed phylogenetic analysis revealed a high level of relatedness between NTS from blood and stool originating from cases and controls, indicating a common source pool. Multidrug resistance was common throughout, with 8.5% of such isolates resistant to extended spectrum beta lactams. The high rate of asymptomatic carriage in the population is a concern for transmission to vulnerable individuals and this group could be targeted for vaccination if an iNTS vaccine becomes available.
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http://dx.doi.org/10.1371/journal.pntd.0008440DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7425985PMC
August 2020

Tenacious Endemic Typhoid Fever in Samoa.

Clin Infect Dis 2020 07;71(Suppl 2):S120-S126

Center for Vaccine Development and Global Health, University of Maryland School of Medicine, Baltimore, Maryland, USA.

Background: Typhoid fever has been endemic on the island nation of Samoa (2016 population, 195 979) since the 1960s and has persisted through 2019, despite economic development and improvements in water supply and sanitation.

Methods: Salmonella enterica serovar Typhi isolates from the 2 hospitals with blood culture capability and matched patient demographic and clinical data from January 2008 through December 2019 were analyzed. Denominators to calculate incidence by island, region, and district came from 2011 and 2016 censuses and from 2017-2019 projections from Samoa's Bureau of Statistics. Data were analyzed to describe typhoid case burden and incidence from 2008 to 2019 by time, place, and person.

Results: In sum, 53-193 blood culture-confirmed typhoid cases occurred annually from 2008 to 2019, without apparent seasonality. Typhoid incidence was low among children age < 48 months (17.6-27.8/105), rose progressively in ages 5-9 years (54.0/105), 10-19 years (60.7-63.4/105), and 20-34 years (61.0-79.3/105), and then tapered off; 93.6% of cases occurred among Samoans < 50 years of age. Most typhoid cases and the highest incidence occurred in Northwest Upolu, but Apia Urban Area (served by treated water supplies) also exhibited moderate incidence. The proportion of cases from short-cycle versus long-cycle transmission is unknown. Samoan S. Typhi are pansusceptible to traditional first-line antibiotics. Nevertheless, enhanced surveillance in 2019 detected 4 (2.9%) deaths among 140 cases.

Conclusions: Typhoid has been endemic in Samoa in the period 2008-2019. Interventions, including mass vaccination with a Vi-conjugate vaccine coadministered with measles vaccine are planned.
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http://dx.doi.org/10.1093/cid/ciaa314DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7388710PMC
July 2020

The architecture and stabilisation of flagellotropic tailed bacteriophages.

Nat Commun 2020 07 27;11(1):3748. Epub 2020 Jul 27.

Infection & Immunity Program, Biomedicine Discovery Institute & Department of Biochemistry and Molecular Biology, Monash University, Clayton, VIC, Australia.

Flagellotropic bacteriophages engage flagella to reach the bacterial surface as an effective means to increase the capture radius for predation. Structural details of these viruses are of great interest given the substantial drag forces and torques they face when moving down the spinning flagellum. We show that the main capsid and auxiliary proteins form two nested chainmails that ensure the integrity of the bacteriophage head. Core stabilising structures are conserved in herpesviruses suggesting their ancestral origin. The structure of the tail also reveals a robust yet pliable assembly. Hexameric rings of the tail-tube protein are braced by the N-terminus and a β-hairpin loop, and interconnected along the tail by the splayed β-hairpins. By contrast, we show that the β-hairpin has an inhibitory role in the tail-tube precursor, preventing uncontrolled self-assembly. Dyads of acidic residues inside the tail-tube present regularly-spaced motifs well suited to DNA translocation into bacteria through the tail.
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http://dx.doi.org/10.1038/s41467-020-17505-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7385642PMC
July 2020

A novel therapeutic antibody screening method using bacterial high-content imaging reveals functional antibody binding phenotypes of Escherichia coli ST131.

Sci Rep 2020 07 24;10(1):12414. Epub 2020 Jul 24.

Department of Medicine, Cambridge Institute for Therapeutic Immunology & Infectious Disease, University of Cambridge, Cambridge, UK.

The increase of antimicrobial resistance (AMR), and lack of new classes of licensed antimicrobials, have made alternative treatment options for AMR pathogens increasingly attractive. Recent studies have demonstrated anti-bacterial efficacy of a humanised monoclonal antibody (mAb) targeting the O25b O-antigen of Escherichia coli ST131. To evaluate the phenotypic effects of antibody binding to diverse clinical E. coli ST131 O25b bacterial isolates in high-throughput, we designed a novel mAb screening method using high-content imaging (HCI) and image-based morphological profiling to screen a mAb targeting the O25b O-antigen. Screening the antibody against a panel of 86 clinical E. coli ST131 O25:H4 isolates revealed 4 binding phenotypes: no binding (18.60%), weak binding (4.65%), strong binding (69.77%) and strong agglutinating binding (6.98%). Impaired antibody binding could be explained by the presence of insertion sequences or mutations in O-antigen or lipopolysaccharide core biosynthesis genes, affecting the amount, structure or chain length of the O-antigen. The agglutinating binding phenotype was linked with lower O-antigen density, enhanced antibody-mediated phagocytosis and increased serum susceptibly. This study highlights the need to screen candidate mAbs against large panels of clinically relevant isolates, and that HCI can be used to evaluate mAb binding affinity and potential functional efficacy against AMR bacteria.
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http://dx.doi.org/10.1038/s41598-020-69300-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7382476PMC
July 2020

Pathogen genomic surveillance of typhoidal infection in adults and children reveals no association between clinical outcomes and infecting genotypes.

Trop Med Health 2020 13;48:58. Epub 2020 Jul 13.

Oxford Vaccine Group, Department of Paediatrics, University of Oxford and the NIHR Oxford Biomedical Research Centre, Oxford, OX3 7LE UK.

Background: India is endemic for enteric fever, and it is not known whether the variations in clinical manifestations between patients are due to host, environmental or pathogen factors.Blood culture surveillance was conducted at St. John's Medical College Hospital, Bangalore, between July 2016 and June 2017. Clinical, laboratory and demographic data were collected from each case, and bacterial isolates were subjected to whole genome sequencing. Comparative analysis between adults and paediatric patients was carried out to ascertain differences between adult and paediatric disease.

Results: Among the 113 cases of blood culture-confirmed enteric fever, young adults (16-30 years) and children < 15 years accounted for 47% and 37% of cases, respectively. Anaemia on presentation was seen in 46% of cases, and 19% had an abnormal leucocyte count on presentation. The majority received treatment as inpatients (70%), and among these, adults had a significantly longer duration of admission when compared with children ( = 0.002). There were atypical presentations including arthritis, acute haemolysis and a case of repeated typhoid infection with two separate Typhi genotypes. There was no association between infecting genotype/serovar and treatment status (outpatient vs inpatient), month of isolation, duration of admission, patient age (adult or child), antimicrobial susceptibility, Widal positivity or haematologic parameters.

Conclusions: Amidst the many public health concerns of South India, enteric fever continues to contribute substantially to hospital burden with non-specific as well as uncommon clinical features in both paediatric and adult populations likely driven by host and environmental factors. Robust clinical surveillance as well monitoring of pathogen population structure is required to inform treatment and preventive strategies.
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http://dx.doi.org/10.1186/s41182-020-00247-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7359007PMC
July 2020

Rapid implementation of SARS-CoV-2 sequencing to investigate cases of health-care associated COVID-19: a prospective genomic surveillance study.

Lancet Infect Dis 2020 11 14;20(11):1263-1271. Epub 2020 Jul 14.

Department of Pathology, University of Cambridge, Cambridge, UK. Electronic address:

Background: The burden and influence of health-care associated severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections is unknown. We aimed to examine the use of rapid SARS-CoV-2 sequencing combined with detailed epidemiological analysis to investigate health-care associated SARS-CoV-2 infections and inform infection control measures.

Methods: In this prospective surveillance study, we set up rapid SARS-CoV-2 nanopore sequencing from PCR-positive diagnostic samples collected from our hospital (Cambridge, UK) and a random selection from hospitals in the East of England, enabling sample-to-sequence in less than 24 h. We established a weekly review and reporting system with integration of genomic and epidemiological data to investigate suspected health-care associated COVID-19 cases.

Findings: Between March 13 and April 24, 2020, we collected clinical data and samples from 5613 patients with COVID-19 from across the East of England. We sequenced 1000 samples producing 747 high-quality genomes. We combined epidemiological and genomic analysis of the 299 patients from our hospital and identified 35 clusters of identical viruses involving 159 patients. 92 (58%) of 159 patients had strong epidemiological links and 32 (20%) patients had plausible epidemiological links. These results were fed back to clinical, infection control, and hospital management teams, leading to infection-control interventions and informing patient safety reporting.

Interpretation: We established real-time genomic surveillance of SARS-CoV-2 in a UK hospital and showed the benefit of combined genomic and epidemiological analysis for the investigation of health-care associated COVID-19. This approach enabled us to detect cryptic transmission events and identify opportunities to target infection-control interventions to further reduce health-care associated infections. Our findings have important implications for national public health policy as they enable rapid tracking and investigation of infections in hospital and community settings.

Funding: COVID-19 Genomics UK funded by the Department of Health and Social Care, UK Research and Innovation, and the Wellcome Sanger Institute.
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http://dx.doi.org/10.1016/S1473-3099(20)30562-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7806511PMC
November 2020

Effective control of SARS-CoV-2 transmission between healthcare workers during a period of diminished community prevalence of COVID-19.

Elife 2020 06 19;9. Epub 2020 Jun 19.

Department of Medicine, University of Cambridge, Cambridge, United Kingdom.

Previously, we showed that 3% (31/1032)of asymptomatic healthcare workers (HCWs) from a large teaching hospital in Cambridge, UK, tested positive for SARS-CoV-2 in April 2020. About 15% (26/169) HCWs with symptoms of coronavirus disease 2019 (COVID-19) also tested positive for SARS-CoV-2 (Rivett et al., 2020). Here, we show that the proportion of both asymptomatic and symptomatic HCWs testing positive for SARS-CoV-2 rapidly declined to near-zero between 25th April and 24th May 2020, corresponding to a decline in patient admissions with COVID-19 during the ongoing UK 'lockdown'. These data demonstrate how infection prevention and control measures including staff testing may help prevent hospitals from becoming independent 'hubs' of SARS-CoV-2 transmission, and illustrate how, with appropriate precautions, organizations in other sectors may be able to resume on-site work safely.
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http://dx.doi.org/10.7554/eLife.59391DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7326489PMC
June 2020

Factors associated with occurrence of salmonellosis among children living in Mukuru slum, an urban informal settlement in Kenya.

BMC Infect Dis 2020 Jun 17;20(1):422. Epub 2020 Jun 17.

Centre for Microbiology Research, Kenya Medical Research Institute, Off Mbagathi Road, PO Box 54840-00200, Nairobi, Kenya.

Background: In Kenya, typhoid fever and invasive non-typhoidal salmonellosis present a huge burden of disease, especially in poor-resource settings where clean water supply and sanitation conditions are inadequate. The epidemiology of both diseases is poorly understood in terms of severity and risk factors. The aim of the study was to determine the disease burden and spatial distribution of salmonellosis, as well as socioeconomic and environmental risk factors for these infections, in a large informal settlement near the city of Nairobi, from 2013 to 2017.

Methods: Initially, a house-to-house baseline census of 150,000 population in Mukuru informal settlement was carried out and relevant socioeconomic, demographic, and healthcare utilization information was collected using structured questionnaires. Salmonella bacteria were cultured from the blood and faeces of children < 16 years of age who reported at three outpatient facilities with fever alone or fever and diarrhea. Tests of association between specific Salmonella serotypes and risk factors were conducted using Pearson Chi-Square (χ) test.

Results: A total of 16,236 children were recruited into the study. The prevalence of bloodstream infections by Non-Typhoidal Salmonella (NTS), consisting of Salmonella Typhimurium/ Enteriditis, was 1.3%; Salmonella Typhi was 1.4%, and this was highest among children < 16 years of age. Occurrence of Salmonella Typhimurium/ Enteriditis was not significantly associated with rearing any domestic animals. Rearing chicken was significantly associated with high prevalence of S. Typhi (2.1%; p = 0.011). The proportion of children infected with Salmonella Typhimurium/ Enteriditis was significantly higher in households that used water pots as water storage containers compared to using water directly from the tap (0.6%). Use of pit latrines and open defecation were significant risk factors for S. Typhi infection (1.6%; p = 0.048). The proportion of Salmonella Typhimurium/ Enteriditis among children eating street food 4 or more times per week was higher compared to 1 to 2 times/week on average (1.1%; p = 0.032).

Conclusion: Typhoidal and NTS are important causes of illness in children in Mukuru informal settlement, especially among children less than 16 years of age. Improving Water, Sanitation and Hygiene (WASH) including boiling water, breastfeeding, hand washing practices, and avoiding animal contact in domestic settings could contribute to reducing the risk of transmission of Salmonella disease from contaminated environments.
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http://dx.doi.org/10.1186/s12879-020-05134-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7302364PMC
June 2020