Publications by authors named "Stephen Baker"

573 Publications

The genomic epidemiology of multi-drug resistant invasive non-typhoidal in selected sub-Saharan African countries.

BMJ Glob Health 2021 08;6(8)

University of Antananarivo, Antananarivo, Madagascar.

Background: Invasive non-typhoidal (iNTS) is one of the leading causes of bacteraemia in sub-Saharan Africa. We aimed to provide a better understanding of the genetic characteristics and transmission patterns associated with multi-drug resistant (MDR) iNTS serovars across the continent.

Methods: A total of 166 iNTS isolates collected from a multi-centre surveillance in 10 African countries (2010-2014) and a fever study in Ghana (2007-2009) were genome sequenced to investigate the geographical distribution, antimicrobial genetic determinants and population structure of iNTS serotypes-genotypes. Phylogenetic analyses were conducted in the context of the existing genomic frameworks for various iNTS serovars. Population-based incidence of MDR-iNTS disease was estimated in each study site.

Results: Typhimurium sequence-type (ST) 313 and Enteritidis ST11 were predominant, and both exhibited high frequencies of MDR; Dublin ST10 was identified in West Africa only. Mutations in the gene (fluoroquinolone resistance) were identified in . Enteritidis and . Typhimurium in Ghana; an ST313 isolate carrying was found in Kenya. International transmission of MDR ST313 (lineage II) and MDR ST11 (West African clade) was observed between Ghana and neighbouring West African countries. The incidence of MDR-iNTS disease exceeded 100/100 000 person-years-of-observation in children aged <5 years in several West African countries.

Conclusions: We identified the circulation of multiple MDR iNTS serovar STs in the sampled sub-Saharan African countries. Investment in the development and deployment of iNTS vaccines coupled with intensified antimicrobial resistance surveillance are essential to limit the impact of these pathogens in Africa.
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http://dx.doi.org/10.1136/bmjgh-2021-005659DOI Listing
August 2021

Predicting wildfire particulate matter and hypothetical re-emission of radiological Cs-137 contamination incidents.

Sci Total Environ 2021 Jul 5;795:148872. Epub 2021 Jul 5.

Missoula Fire Sciences Laboratory, Rocky Mountain Research Station, US Forest Service, Missoula, MT, USA.

Radiological release incidents can potentially contaminate widespread areas with radioactive materials and decontamination efforts are typically focused on populated areas, which means radionuclides may be left in forested areas for long periods of time. Large wildfires in contaminated forested areas have the potential to reintroduce these radionuclides into the atmosphere and cause exposure to first responders and downwind communities. One important radionuclide contaminant released from radiological incidents is radiocesium (Cs) due to high yields and its long half-life of 30.2 years. An Eulerian 3D photochemical transport model was used to estimate potential ambient impacts of Cs re-emission due to wildfire following hypothetical radiological release scenarios. The Community Multiscale Air Quality (CMAQ) model did well at predicting levels and periods of increased PM2.5 carbon due to wildfire smoke at routine surface monitors in California during the summer of 2016. The model also did well at capturing the extent of the surface mixing layer compared to aerosol lidar measurements. Emissions from a large hypothetical wildfire were introduced into the wildland-urban interface (WUI) impacted by a hypothetical radiological release event. While ambient concentrations tended to be highest near the fire, the highest population committed effective dose equivalent by inhalation to an adult from Cs over an hour was downwind where wind flows moved smoke to high population areas. Seasonal variations in meteorology (wind flows) can result in differential population impacts even in the same metropolitan area. Modeled post-incident ambient levels of Cs both near these wildfires and further downwind in nearby urban areas were well below levels that would necessitate population evacuation or warrant other protective action recommendations such as shelter-in-place. These results suggest that 1) the modeling system captures local to regional scale transport and levels of PM2.5 from wildfire and 2) first responders and downwind population would not be expected to be at elevated risk from the initial inhalathion exposure of Cs re-emission.
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http://dx.doi.org/10.1016/j.scitotenv.2021.148872DOI Listing
July 2021

Quantitative analysis of clot density, fibrin fiber radius, and protofibril packing in acute phase myocardial infarction.

Thromb Res 2021 Jul 8;205:110-119. Epub 2021 Jul 8.

Leeds Thrombosis Collective, Discovery and Translational Science Department, Leeds Institute of Cardiovascular and Metabolic Medicine, University of Leeds, Leeds, UK. Electronic address:

Introduction: Coronary artery disease is associated with impaired clot structure. The aim of this study was to investigate acute phase myocardial infarction (AMI) and provide detailed quantitative analysis of clot ultrastructure.

Materials And Methods: Clot formation and breakdown, pore size, fiber density, fiber radius and protofibril packing were investigated in plasma clots from AMI patients. These data were compared to those from healthy controls.

Results: Analysis on clot formation using turbidity showed increased lag time, suggesting changes in protofibril packing and increased fiber size for AMI patients compared to healthy controls. Additionally, increased average rate of clotting and decreased time to maximum absorbance in AMI patients suggest that clots formed more quickly. Moreover, we observed increased time from max OD to max rate of lysis. Increased fibrinogen and decreased plasminogen in AMI patients were accounted for in represented significant differences. AMI samples showed increased time to 25% and 50% lysis, but no change in 75% lysis, representative of delayed lysis onset, but expediated lysis once initiated. These data suggest that AMI patients formed less porous clots made from more densely packed fibers with decreased numbers of protofibrils, which was confirmed using decreased permeation and increased fiber density, and decreased turbidimetry.

Conclusions: AMI plasma formed clots that were denser, less permeable, and lysed more slowly than healthy controls. These findings were confirmed by detailed analysis of clot ultrastructure, fiber size, and protofibril packing. Dense clot structures that are resistant to lysis may contribute to a prothrombotic milieu in AMI.
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http://dx.doi.org/10.1016/j.thromres.2021.06.024DOI Listing
July 2021

A tightly clustered hepatitis E virus genotype 1a is associated with endemic and outbreak infections in Bangladesh.

PLoS One 2021 22;16(7):e0255054. Epub 2021 Jul 22.

Oxford University Clinical Research Unit, Wellcome Asia Programme, The Hospital for Tropical Diseases, Ho Chi Minh City, Vietnam.

Background: Hepatitis E virus (HEV) infection is endemic in Bangladesh and there are occasional outbreaks. The molecular characteristics and pathogenesis of endemic and outbreak HEV strains are poorly understood. We compared the genetic relatedness and virulence associated mutations of endemic HEV strains with outbreak strains.

Methods: We analyzed systematically collected serum samples from HEV immunoglobulin M (IgM) positive patients attended at Bangabandhu Sheikh Mujib Medical University, Dhaka from August 2013 to June 2015. HEV RNA positive samples were subjected to whole genome sequencing. Genotype and subtype of the strains were determined by phylogenetic analysis. Virulence associated mutations e.g. acute viral hepatitis (AVH), fulminant hepatic failure (FHF), chronic hepatitis, ribavirin treatment failure (RTF), B and T cell neutralization epitopes were determined.

Results: 92 HEV immunoglobulin M (IgM) antibody positive plasma samples (43 in 2013-2014 and 49 in 2014-2015) were studied. 77.1% (70/92) of the samples were HEV RNA positive. A 279 bp open reading frame (ORF) 2 and ORF 3 sequence was obtained from 54.2% (38/70) of the strains. Of these 38 strains, whole genome sequence (WGS) was obtained from 21 strains. In phylogenetic analysis of 38 (279 bp) sequence all HEV sequences belonged to genotype 1 and subtype 1a. Further phylogenetic analysis of 21 HEV WGS, Bangladeshi HEV sequences clustered with genotype 1a sequences from neighboring countries. Within genotype 1a cluster, Bangladesh HEV strains formed a separate cluster with the 2010 HEV outbreak strains from northern Bangladesh. 80.9 to 100% of the strains had A317T, T735I, L1120I, L1110F, P259S, V1479I, G1634K mutations associates AVH, FHF and RTF. Mutations in T cell recognition epitope T3, T5, T7 was observed in 76.1%, 100% and 100% of the strains respectively.

Conclusion: Strains of HEV genotype 1a are dominant in Bangladesh and are associated with endemic and outbreak of HEV infection. HEV isolates in Bangladesh have high prevalence of virulence associated mutations and mutation which alters antigenicity to B and T cell epitopes.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0255054PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8297744PMC
July 2021

The emergence of azithromycin-resistant Typhi in Nepal.

JAC Antimicrob Resist 2020 Dec 21;2(4):dlaa109. Epub 2020 Dec 21.

Oxford University Clinical Research Unit, Patan Academy of Health Sciences, Kathmandu, Nepal.

Background: Typhoid fever remains a significant cause of morbidity and mortality in Asia and Africa. The emergence of azithromycin resistance in South Asia is concerning, as azithromycin is one of the last effective oral drugs for treating typhoid.

Objectives: To describe the molecular mechanism and phylogenetics of azithromycin-resistant (Azith) Typhi isolates from Patan Hospital, Kathmandu, Nepal.

Methods: Whole-genome sequences of three Azith  . Typhi isolates (MIC >256 mg/L) were analysed and compared with a global collection to investigate the azithromycin resistance mechanism and phylogenetic structure. Clinical information is reported for one of the three patients infected with Azith  . Typhi.

Results: The three Azith isolates belonged to the H58 lineage and were genetically identical; they were distantly related to contemporaneous . Typhi from Nepal and Azith  . Typhi recently described in Bangladesh. Azithromycin resistance was mediated by a non-synonymous mutation in the gene (R717L). The three Azith isolates showed reduced susceptibility to ciprofloxacin (double mutation in the : S83F and D87G), and were susceptible to ampicillin, chloramphenicol and co-trimoxazole. Clinical information from one patient suggested non-response to azithromycin treatment.

Conclusions: This is the first molecular description of Azith  . Typhi in Nepal. These organisms showed no phylogenetic link to Azith  . Typhi in Bangladesh. Our data suggest that increasing use of azithromycin may pose a strong selective pressure driving the emergence of Azith  . Typhi in South Asia. Further investigations are needed to evaluate treatment responses to azithromycin, predict evolutionary trajectories, and track the transmission of these organisms.
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http://dx.doi.org/10.1093/jacamr/dlaa109DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8210228PMC
December 2020

Elimination of fibrin γ-chain cross-linking by FXIIIa increases pulmonary embolism arising from murine inferior vena cava thrombi.

Proc Natl Acad Sci U S A 2021 Jul;118(27)

Leeds Thrombosis Collective, Discovery & Translational Science Department, Leeds Institute of Cardiovascular and Metabolic Medicine, University of Leeds, Leeds LS2 9NL, United Kingdom;

The onset of venous thromboembolism, including pulmonary embolism, represents a significant health burden affecting more than 1 million people annually worldwide. Current treatment options are based on anticoagulation, which is suboptimal for preventing further embolic events. In order to develop better treatments for thromboembolism, we sought to understand the structural and mechanical properties of blood clots and how this influences embolism in vivo. We developed a murine model in which fibrin γ-chain cross-linking by activated Factor XIII is eliminated (FGG3X) and applied methods to study thromboembolism at whole-body and organ levels. We show that FGG3X mice have a normal phenotype, with overall coagulation parameters and platelet aggregation and function largely unaffected, except for total inhibition of fibrin γ-chain cross-linking. Elimination of fibrin γ-chain cross-linking resulted in thrombi with reduced strength that were prone to fragmentation. Analysis of embolism in vivo using Xtreme optical imaging and light sheet microscopy demonstrated that the elimination of fibrin γ-chain cross-linking resulted in increased embolization without affecting clot size or lysis. Our findings point to a central previously unrecognized role for fibrin γ-chain cross-linking in clot stability. They also indirectly indicate mechanistic targets for the prevention of thrombosis through selective modulation of fibrin α-chain but not γ-chain cross-linking by activated Factor XIII to reduce thrombus size and burden, while maintaining clot stability and preventing embolism.
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http://dx.doi.org/10.1073/pnas.2103226118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8271579PMC
July 2021

Bactericidal activities and post-antibiotic effects of ofloxacin and ceftriaxone against drug-resistant Salmonella enterica serovar Typhi.

J Antimicrob Chemother 2021 Jun 28. Epub 2021 Jun 28.

Wellcome Trust Major Overseas Programme, Oxford University Clinical Research Unit, Hospital for Tropical Diseases, Vo Van Kiet, District 5, Ho Chi Minh City, Vietnam.

Background: The clinical response to ceftriaxone in patients with typhoid fever is significantly slower than with ofloxacin, despite infection with Salmonella enterica serovar Typhi (S. Typhi) isolates with similar susceptibilities (MIC 0.03-0.12 mg/L). The response to ofloxacin is slower if the isolate has intermediate susceptibility (MIC 0.25-1.0 mg/L).

Objectives: To determine the bactericidal activity and post-antibiotic effect (PAE) of ceftriaxone and ofloxacin against S. Typhi.

Methods: The mean time to reach a 99.9% reduction in log10 count (bactericidal activity) and PAE of ceftriaxone and ofloxacin were determined for 18 clinical isolates of S. Typhi in time-kill experiments (MIC range for ofloxacin 0.06-1.0 mg/L and for ceftriaxone 0.03-0.12 mg/L).

Results: The mean (SD) bactericidal activity of ofloxacin was 33.1 (15.2) min and 384.4 (60) min for ceftriaxone. After a 30 min exposure to ofloxacin, the mean (SD) duration of PAE was 154.7 (52.6) min. There was no detectable PAE after 1 h of exposure to ceftriaxone. For ofloxacin, bactericidal activity and PAE did not significantly differ between isolates with full or intermediate susceptibility provided ofloxacin concentrations were maintained at 4×MIC.

Conclusions: Infections with S. Typhi with intermediate ofloxacin susceptibility may respond to doses that maintain ofloxacin concentrations at 4×MIC at the site of infection. The slow bactericidal activity of ceftriaxone and absent PAE may explain the slow clinical response in typhoid.
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http://dx.doi.org/10.1093/jac/dkab215DOI Listing
June 2021

Determining the burden of fungal infections in Zimbabwe.

Sci Rep 2021 06 24;11(1):13240. Epub 2021 Jun 24.

Institute of Immunology and Infection Research, University of Edinburgh, Ashworth Laboratories, King's Buildings, Charlotte Auerbach Road, Edinburgh, EH9 3FL, UK.

Zimbabwe currently faces several healthcare challenges, most notably HIV and associated infections including tuberculosis (TB), malaria and recently outbreaks of cholera, typhoid fever and COVID-19. Fungal infections, which are also a major public health threat, receive considerably less attention. Consequently, there is dearth of data regarding the burden of fungal diseases in the country. We estimated the burden of fungal diseases in Zimbabwe based on published literature and 'at-risk' populations (HIV/AIDS patients, survivors of pulmonary TB, cancer, chronic obstructive pulmonary disease, asthma and patients receiving critical care) using previously described methods. Where there was no data for Zimbabwe, regional, or international data was used. Our study revealed that approximately 14.9% of Zimbabweans suffer from fungal infections annually, with 80% having tinea capitis. The annual incidence of cryptococcal meningitis and Pneumocystis jirovecii pneumonia in HIV/AIDS were estimated at 41/100,000 and 63/100,000, respectively. The estimated prevalence of recurrent vulvovaginal candidiasis (RVVC) was 2,739/100,000. The estimated burden of fungal diseases in Zimbabwe is high in comparison to other African countries, highlighting the urgent need for increased awareness and surveillance to improve diagnosis and management.
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http://dx.doi.org/10.1038/s41598-021-92605-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8225815PMC
June 2021

Inhibitory Concentrations of Ciprofloxacin Induce an Adaptive Response Promoting the Intracellular Survival of Salmonella enterica Serovar Typhimurium.

mBio 2021 06 22;12(3):e0109321. Epub 2021 Jun 22.

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

Antimicrobial resistance (AMR) is a pressing global health crisis, which has been fueled by the sustained use of certain classes of antimicrobials, including fluoroquinolones. While the genetic mutations responsible for decreased fluoroquinolone (ciprofloxacin) susceptibility are known, the implications of ciprofloxacin exposure on bacterial growth, survival, and interactions with host cells are not well described. Aiming to understand the influence of inhibitory concentrations of ciprofloxacin we subjected three clinical isolates of Salmonella enterica serovar Typhimurium to differing concentrations of ciprofloxacin, dependent on their MICs, and assessed the impact on bacterial growth, morphology, and transcription. We further investigated the differential morphology and transcription that occurred following ciprofloxacin exposure and measured the ability of ciprofloxacin-treated bacteria to invade and replicate in host cells. We found that ciprofloxacin-exposed Typhimurium is able to recover from inhibitory concentrations of ciprofloxacin and that the drug induces specific morphological and transcriptional signatures associated with the bacterial SOS response, DNA repair, and intracellular survival. In addition, ciprofloxacin-treated Typhimurium has increased capacity for intracellular replication in comparison to that of untreated organisms. These data suggest that Typhimurium undergoes an adaptive response under ciprofloxacin perturbation that promotes cellular survival, a consequence that may justify more measured use of ciprofloxacin for Salmonella infections. The combination of multiple experimental approaches provides new insights into the collateral effects that ciprofloxacin and other antimicrobials have on invasive bacterial pathogens. Antimicrobial resistance is a critical concern in global health. In particular, there is rising resistance to fluoroquinolones, such as ciprofloxacin, a first-line antimicrobial for many Gram-negative pathogens. We investigated the adaptive response of clinical isolates of Salmonella enterica serovar Typhimurium to ciprofloxacin, finding that the bacteria adapt in short timespans to high concentrations of ciprofloxacin in a way that promotes intracellular survival during early infection. Importantly, by studying three clinically relevant isolates, we were able to show that individual isolates respond differently to ciprofloxacin and that for each isolate, there was a heterogeneous response under ciprofloxacin treatment. The heterogeneity that arises from ciprofloxacin exposure may drive survival and proliferation of Salmonella during treatment and lead to drug resistance.
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http://dx.doi.org/10.1128/mBio.01093-21DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8262899PMC
June 2021

Serial Excision for Treatment of Non-melanoma Skin Cancer.

Plast Reconstr Surg Glob Open 2021 Jun 10;9(6):e3607. Epub 2021 Jun 10.

Department of Plastic and Reconstructive Surgery, MedStar Georgetown University Hospital, Washington, D.C.

Mohs micrographic surgery (MMS) has become the predominant modality of excision for non-melanoma skin cancers (NMSC). Patients are referred for MMS under the assumption that it is the most effective procedure for definitive removal of the cancer while also allowing for maximal tissue preservation to achieve optimal cosmesis. The objective of this study was to investigate outcomes of serial excision (SE) as an alternative excision modality for NMSC.

Methods: Patients undergoing SE for basal cell carcinoma or squamous cell carcinoma by the senior author from 2009 to 2020 were retrospectively reviewed. Patient demographics, lesion characteristics, and excision characteristics were recorded. The primary outcome was the number of excisions required to achieve negative margins.

Results: In total, 129 patients with 205 NMSC lesions were retrospectively reviewed. An estimated 69 lesions (33.7%) were located in high risk areas, as defined by the National Comprehensive Cancer Network. Negative margins were achieved in 191 (93.2%) lesions. In 88.3% of lesions (n = 181/205), negative margins were achieved in 2 or less excisions. 12 lesions (5.9%) were referred for MMS.

Conclusions: Our results demonstrate that SE is an effective modality for definitive removal of NMSC. Recent research reveals that SE is much less expensive than MMS, and therefore places a smaller financial burden on the patient and the healthcare system as a whole. Relative to MMS, SE offers similar if not increased benefits for lower cost. Our findings highlight the need to critically reassess the select indications for MMS.
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http://dx.doi.org/10.1097/GOX.0000000000003607DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8191689PMC
June 2021

Risk factors for the development of neonatal sepsis in a neonatal intensive care unit of a tertiary care hospital of Nepal.

BMC Infect Dis 2021 Jun 9;21(1):546. Epub 2021 Jun 9.

Oxford University Clinical Research Unit, Patan Academy of Health Sciences, Kathmandu, Nepal.

Background: Sepsis is an overwhelming and life-threatening response to bacteria in bloodstream and a major cause of neonatal morbidity and mortality. Understanding the etiology and potential risk factors for neonatal sepsis is urgently required, particularly in low-income countries where burden of infection is high and its epidemiology is poorly understood.

Methods: A prospective observational cohort study was conducted between April 2016 and October 2017 in a level three NICU at a tertiary care hospital in Nepal to determine the bacterial etiology and potential risk factors for neonatal sepsis.

Results: Among 142 NICU admitted neonates, 15% (21/142) and 32% (46/142) developed blood culture-positive and -negative neonatal sepsis respectively. Klebsiella pneumoniae (34%, 15/44) and Enterobacter spp. (25%, 11/44) were the most common isolates. The antimicrobial resistance of isolates to ampicillin (100%, 43/43), cefotaxime (74%, 31/42) and ampicillin-sulbactam (55%, 21/38) were the highest. Bla (53%, 18/34) and bla (46%, 13/28) were the commonest ESBL and carbapenemase genes respectively. In univariate logistic regression, the odds of sepsis increased with each additional day of use of invasive procedures such as mechanical ventilation (OR 1.086, 95% CI 1.008-1.170), umbilical artery catheter (OR 1.375, 95% CI 1.049-1.803), intravenous cannula (OR 1.140, 95% CI 1.062-1.225); blood transfusion events (OR 3.084, 95% CI 1.407-6.760); NICU stay (OR 1.109, 95% CI 1.040-1.182) and failure to breast feed (OR 1.130, 95% CI 1.060-1.205). Sepsis odds also increased with leukopenia (OR 1.790, 95% CI 1.04-3.082), increase in C-reactive protein (OR 1.028, 95% CI 1.016-1.040) and decrease in platelets count (OR 0.992, 95% CI 0.989-0.994). In multivariate analysis, increase in IV cannula insertion days (OR 1.147, 95% CI 1.039-1.267) and CRP level (OR 1.028, 95% CI 1.008-1.049) increased the odds of sepsis.

Conclusions: Our study indicated various nosocomial risk factors and underscored the need to improve local infection control measures so as to reduce the existing burden of sepsis. We have highlighted certain sepsis associated laboratory parameters along with identification of antimicrobial resistance genes, which can guide for early and better therapeutic management of sepsis. These findings could be extrapolated to other low-income settings within the region.
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http://dx.doi.org/10.1186/s12879-021-06261-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8191200PMC
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
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8269201PMC
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

Global population structure and genotyping framework for genomic surveillance of the major dysentery pathogen, Shigella sonnei.

Nat Commun 2021 05 11;12(1):2684. Epub 2021 May 11.

Department of Infectious Diseases, Central Clinical School, Monash University, Melbourne, VIC, Australia.

Shigella sonnei is the most common agent of shigellosis in high-income countries, and causes a significant disease burden in low- and middle-income countries. Antimicrobial resistance is increasingly common in all settings. Whole genome sequencing (WGS) is increasingly utilised for S. sonnei outbreak investigation and surveillance, but comparison of data between studies and labs is challenging. Here, we present a genomic framework and genotyping scheme for S. sonnei to efficiently identify genotype and resistance determinants from WGS data. The scheme is implemented in the software package Mykrobe and tested on thousands of genomes. Applying this approach to analyse >4,000 S. sonnei isolates sequenced in public health labs in three countries identified several common genotypes associated with increased rates of ciprofloxacin resistance and azithromycin resistance, confirming intercontinental spread of highly-resistant S. sonnei clones and demonstrating the genomic framework can facilitate monitoring the spread of resistant clones, including those that have recently emerged, at local and global scales.
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http://dx.doi.org/10.1038/s41467-021-22700-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8113504PMC
May 2021

Commentary on: Reflecting on Your Reflection: Examining the Effect of a Non-Reversing Mirror on Self Perception.

Authors:
Stephen B Baker

Aesthet Surg J 2021 Apr 13. Epub 2021 Apr 13.

Department of Plastic Surgery, Medstar Georgetown University Hospital, Washington, DC.

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http://dx.doi.org/10.1093/asj/sjab187DOI Listing
April 2021

Commentary on: Key Anatomical Clarifications for the Marginal Mandibular Branch of the Facial Nerve: Clinical Significance for the Plastic Surgeon.

Authors:
Stephen B Baker

Aesthet Surg J 2021 Apr 2. Epub 2021 Apr 2.

MedStar Georgetown University Hospital, Department of Plastic Surgery, Washington, DC, USA.

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http://dx.doi.org/10.1093/asj/sjab088DOI Listing
April 2021

: High Prevalence and Possibly Chronic Shedding in Human Respiratory Tract, But No Zoonotic Transmission.

Viruses 2021 03 24;13(4). Epub 2021 Mar 24.

Emerging Infection Group, Oxford University Clinical Research Unit, Ho Chi Minh City 7000, Vietnam.

is a recently discovered DNA virus family consisting of two species, and . Here we used PCR amplification and sequencing to characterize redondoviruses in nasal/throat swabs collected longitudinally from a cohort of 58 individuals working with animals in Vietnam. We additionally analyzed samples from animals to which redondovirus DNA-positive participants were exposed. Redondoviruses were detected in approximately 60% of study participants, including 33% (30/91) of samples collected during episodes of acute respiratory disease and in 50% (29/58) of baseline samples (with no respiratory symptoms). Vientovirus (73%; 24/33) was detected more frequently in samples than brisaviruses (27%; 9/33). In the 23 participants with at least 2 redondovirus-positive samples among their longitudinal samples, 10 (43.5%) had identical redondovirus replication-gene sequences detected (sampling duration: 35-132 days). We found no identical redondovirus replication genes in samples from different participants, and no redondoviruses were detected in 53 pooled nasal/throat swabs collected from domestic animals. Phylogenetic analysis described no large-scale geographical clustering between viruses from Vietnam, the US, Spain, and China, indicating that redondoviruses are highly genetically diverse and have a wide geographical distribution. Collectively, our study provides novel insights into the family in humans, describing a high prevalence, potentially associated with chronic shedding in the respiratory tract with lack of evidence of zoonotic transmission from close animal contacts. The tropism and potential pathogenicity of this viral family remain to be determined.
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http://dx.doi.org/10.3390/v13040533DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8063800PMC
March 2021

Evolutionary histories and antimicrobial resistance in Shigella flexneri and Shigella sonnei in Southeast Asia.

Commun Biol 2021 03 19;4(1):353. Epub 2021 Mar 19.

Oxford University Clinical Research Unit, Ho Chi Minh City, Vietnam.

Conventional disease surveillance for shigellosis in developing country settings relies on serotyping and low-resolution molecular typing, which fails to contextualise the evolutionary history of the genus. Here, we interrogated a collection of 1,804 Shigella whole genome sequences from organisms isolated in four continental Southeast Asian countries (Thailand, Vietnam, Laos, and Cambodia) over three decades to characterise the evolution of both S. flexneri and S. sonnei. We show that S. sonnei and each major S. flexneri serotype are comprised of genetically diverse populations, the majority of which were likely introduced into Southeast Asia in the 1970s-1990s. Intranational and regional dissemination allowed widespread propagation of both species across the region. Our data indicate that the epidemiology of S. sonnei and the major S. flexneri serotypes were characterised by frequent clonal replacement events, coinciding with changing susceptibility patterns against contemporaneous antimicrobials. We conclude that adaptation to antimicrobial pressure was pivotal to the recent evolutionary trajectory of Shigella in Southeast Asia.
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http://dx.doi.org/10.1038/s42003-021-01905-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7979695PMC
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

Applying to Residency in the COVID-19 Era: Virtual Interview Tips for Success.

Plast Reconstr Surg Glob Open 2021 Jan 15;9(1):e3389. Epub 2020 Dec 15.

Department of Plastic and Reconstructive Surgery, MedStar Georgetown University Hospital, Washington, D.C.

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http://dx.doi.org/10.1097/GOX.0000000000003389DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7862738PMC
January 2021

The Well-rounded Applicant in a Square Hole: Social Media during the COVID-19 Application Cycle.

Plast Reconstr Surg Glob Open 2021 Jan 23;9(1):e3147. Epub 2020 Dec 23.

Department of Plastic and Reconstructive Surgery, MedStar Georgetown University Hospital, Washington, D.C.

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http://dx.doi.org/10.1097/GOX.0000000000003147DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7858710PMC
January 2021

Spontaneous Emergence of Azithromycin Resistance in Independent Lineages of Salmonella Typhi in Northern India.

Clin Infect Dis 2021 03;72(5):e120-e127

Department of Medical Microbiology, Postgraduate Institute of Medical Education and Research, Chandigarh, India.

Background: The emergence and spread of antimicrobial resistance (AMR) pose a major threat to the effective treatment and control of typhoid fever. The ongoing outbreak of extensively drug-resistant Salmonella Typhi (S. Typhi) in Pakistan has left azithromycin as the only remaining broadly efficacious oral antimicrobial for typhoid in South Asia. Ominously, azithromycin-resistant S. Typhi organisms have been subsequently reported in Bangladesh, Pakistan, and Nepal.

Methods: Here, we aimed to understand the molecular basis of AMR in 66 S. Typhi organisms isolated in a cross-sectional study performed in a suburb of Chandigarh in Northern India using whole-genome sequencing and phylogenetic analysis.

Results: We identified 7 S. Typhi organisms with the R717Q mutation in the acrB gene that was recently found to confer resistance to azithromycin in Bangladesh. Six out of the seven azithromycin-resistant S. Typhi isolates also exhibited triple mutations in gyrA (S83F and D87N) and parC (S80I) genes and were resistant to ciprofloxacin. These contemporary ciprofloxacin/azithromycin-resistant isolates were phylogenetically distinct from each other and from those reported from Bangladesh, Pakistan, and Nepal.

Conclusions: The independent emergence of azithromycin-resistant typhoid in Northern India reflects an emerging broader problem across South Asia and illustrates the urgent need for the introduction of typhoid conjugate vaccines in the region.
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http://dx.doi.org/10.1093/cid/ciaa1773DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7935384PMC
March 2021

Colonization with and causes infections in a Vietnamese intensive care unit.

Microb Genom 2021 02;7(2)

Oxford University Clinical Research Unit, Ho Chi Minh City, Vietnam.

Pre-existing colonization with or has been found to increase the risk of infection in intensive care patients. We previously conducted a longitudinal study to characterize colonization of these two organisms in patients admitted to intensive care in a hospital in southern Vietnam. Here, using genomic and phylogenetic analyses, we aimed to assess the contribution these colonizing organisms made to infections. We found that in the majority of patients infected with or , the sequence type of the disease-causing (infecting) isolate was identical to that of corresponding colonizing organisms in the respective patient. Further in-depth analysis revealed that in patients infected by ST188 and by ST17, ST23, ST25 and ST86, the infecting isolate was closely related to and exhibited limited genetic variation relative to pre-infection colonizing isolates. Multidrug-resistant ST188 was identified as the predominant agent of colonization and infection. Colonization and infection by were characterized by organisms with limited antimicrobial resistance profiles but extensive repertoires of virulence genes. Our findings augment the understanding of the link between bacterial colonization and infection in a low-resource setting, and could facilitate the development of novel evidence-based approaches to prevent and treat infections in high-risk patients in intensive care.
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http://dx.doi.org/10.1099/mgen.0.000514DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8208697PMC
February 2021

Challenges and opportunities in antimicrobial resistance research.

Authors:
Stephen Baker

Microbiology (Reading) 2021 01;167(1)

Cambridge Institute of Therapeutic Immunology and Infectious Disease, University of Cambridge School of Clinical Medicine, Cambridge Biomedical Campus, Cambridge, UK.

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http://dx.doi.org/10.1099/mic.0.000999DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8116779PMC
January 2021

GPVI (Glycoprotein VI) Interaction With Fibrinogen Is Mediated by Avidity and the Fibrinogen αC-Region.

Arterioscler Thromb Vasc Biol 2021 03 21;41(3):1092-1104. Epub 2021 Jan 21.

Discovery and Translational Science Department, Institute of Cardiovascular and Metabolic Medicine (R.-G.X., J.S.G., S.R.B., H.R.M., C.D., R.A.S.A.).

Objective: GPVI (glycoprotein VI) is a key molecular player in collagen-induced platelet signaling and aggregation. Recent evidence indicates that it also plays important role in platelet aggregation and thrombus growth through interaction with fibrin(ogen). However, there are discrepancies in the literature regarding whether the monomeric or dimeric form of GPVI binds to fibrinogen at high affinity. The mechanisms of interaction are also not clear, including which region of fibrinogen is responsible for GPVI binding. We aimed to gain further understanding of the mechanisms of interaction at molecular level and to identify the regions on fibrinogen important for GPVI binding. Approach and Results: Using multiple surface- and solution-based protein-protein interaction methods, we observe that dimeric GPVI binds to fibrinogen with much higher affinity and has a slower dissociation rate constant than the monomer due to avidity effects. Moreover, our data show that the highest affinity interaction of GPVI is with the αC-region of fibrinogen. We further show that GPVI interacts with immobilized fibrinogen and fibrin variants at a similar level, including a nonpolymerizing fibrin variant, suggesting that GPVI binding is independent of fibrin polymerization.

Conclusions: Based on the above findings, we conclude that the higher affinity of dimeric GPVI over the monomer for fibrinogen interaction is achieved by avidity. The αC-region of fibrinogen appears essential for GPVI binding. We propose that fibrin polymerization into fibers during coagulation will cluster GPVI through its αC-region, leading to downstream signaling, further activation of platelets, and potentially stimulating clot growth. Graphic Abstract: A graphic abstract is available for this article.
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http://dx.doi.org/10.1161/ATVBAHA.120.315030DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7901536PMC
March 2021

Enhanced biofilm and extracellular matrix production by chronic carriage versus acute isolates of Salmonella Typhi.

PLoS Pathog 2021 01 19;17(1):e1009209. Epub 2021 Jan 19.

Center for Microbial Pathogenesis, Research Institute at Nationwide Children's Hospital, Columbus, Ohio, United States of America.

Salmonella Typhi is the primary causative agent of typhoid fever; an acute systemic infection that leads to chronic carriage in 3-5% of individuals. Chronic carriers are asymptomatic, difficult to treat and serve as reservoirs for typhoid outbreaks. Understanding the factors that contribute to chronic carriage is key to development of novel therapies to effectively resolve typhoid fever. Herein, although we observed no distinct clustering of chronic carriage isolates via phylogenetic analysis, we demonstrated that chronic isolates were phenotypically distinct from acute infection isolates. Chronic carriage isolates formed significantly thicker biofilms with greater biomass that correlated with significantly higher relative levels of extracellular DNA (eDNA) and DNABII proteins than biofilms formed by acute infection isolates. Importantly, extracellular DNABII proteins include integration host factor (IHF) and histone-like protein (HU) that are critical to the structural integrity of bacterial biofilms. In this study, we demonstrated that the biofilm formed by a chronic carriage isolate in vitro, was susceptible to disruption by a specific antibody against DNABII proteins, a successful first step in the development of a therapeutic to resolve chronic carriage.
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http://dx.doi.org/10.1371/journal.ppat.1009209DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7815147PMC
January 2021
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