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In developing countries, Salmonella enterica serovar Enteritidis causes substantial illness and death, and drug resistance is increasing. Isolates from the United Kingdom containing virulence-resistance plasmids were characterized. They mainly caused invasive infections in adults linked to Africa. The common features in isolates from these continents indicate the role of human travel in their spread. [ABSTRACT FROM AUTHOR]

Objectives: In England, group B streptococci (GBS; Streptococcus agalactiae) are considered universally susceptible to penicillin. Reports from Africa, Asia, North America and a few European countries have described GBS isolates with penicillin MICs above the epidemiological cut-off (0.125 mg/L). Our aim was to characterize a penicillin-resistant GBS (PRGBS) isolate recovered in 2016 from a patient treated with long-term antimicrobials in the UK., Methods: Antibiotic susceptibility of a referred isolate from a discharging sinus overlying a chronic prosthetic joint infection was determined using gradient strip testing for seven antibiotics. Illumina short read sequencing was carried out using a HiSeq 2500 platform to determine MLST, capsular type, to detect mutations in the pbp genes, and to compare the isolate with contemporaneous GBS isolates circulating in the UK., Results: The GBS isolate belonged to capsular type Ia and MLST 144. We observed resistance to penicillin (MIC = 1 mg/L) and tetracycline (32 mg/L) with susceptibility to linezolid (1 mg/L), erythromycin (0.064 mg/L), clindamycin (0.064 mg/L), teicoplanin (0.064 mg/L) and vancomycin (0.25 mg/L). Deduced amino acid sequences revealed substitutions and non-synonymous changes in PBP2x and PBP2b. Genomic analysis of contemporaneous cases (n = 34) from across the UK identified single nucleotide polymorphism (SNP) variation ranged from 153-6596 SNPs., Conclusions: We confirm the first identification of a PRGBS isolate amongst referrals to the UK's national reference laboratory. Substitutions in pbp1a, pbp2a, pbp2x and pbp2b were identified that likely developed in the face of long-term beta-lactam antibiotic use., (© The Author(s) 2024. Published by Oxford University Press on behalf of British Society for Antimicrobial Chemotherapy.)

Promoting vaccine acceptance and demand is an essential, yet often underrecognized component of ensuring that everyone has access to the full benefits of immunization. Convened by the Sabin Vaccine Institute, the Vaccination Acceptance Research Network (VARN) is a global network of multidisciplinary stakeholders driving strengthened vaccination acceptance, demand, and delivery. VARN works to advance and apply social and behavioral science insights, research, and expertise to the challenges and opportunities facing vaccination decision-makers. The second annual VARN conference, When Communities Lead, Global Immunization Succeeds, was held June 13-15, 2023, in Bangkok, Thailand. VARN2023 provided a space for the exploration and dissemination of a growing body of evidence, knowledge, and practice for driving action across the vaccination acceptance, demand, and delivery ecosystem. VARN2023 was co-convened by Sabin and UNICEF and co-sponsored by Gavi, the Vaccine Alliance. VARN2023 brought together 231 global, regional, national, sub-national, and community-level representatives from 47 countries. The conference provided a forum to share learnings and solutions from work conducted across 40+ countries. This article is a synthesis of evidence-based insights from the VARN2023 Conference within four key recommendations: (1) Make vaccine equity and inclusion central to programming to improve vaccine confidence, demand, and delivery; (2) Prioritize communities in immunization service delivery through people-centered approaches and tools that amplify community needs to policymakers, build trust, and combat misinformation; (3) Encourage innovative community-centric solutions for improved routine immunization coverage; and (4) Strengthen vaccination across the life course through building vaccine demand, service integration, and improving the immunization service experience. Insights from VARN can be applied to positively impact vaccination acceptance, demand, and uptake around the world., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Rupali Limaye reports a relationship with Sabin Vaccine Institute that includes: funding grants. Eve Dube reports a relationship with Sabin Vaccine Institute that includes: travel reimbursement. Holly Seale reports a relationship with Sabin Vaccine Institute that includes: travel reimbursement. Rubina Qasim reports a relationship with Sabin Vaccine Institute that includes: travel reimbursement. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Community engagement is vital to the development of people-centered, successful vaccination programs. The diverse Vaccination Acceptance Research Network (VARN) community brings together interdisciplinary professionals from across the immunization ecosystem who play a crucial role in vaccination acceptance, demand, and delivery. Over the course of the VARN2023 conference, researchers and practitioners alike shared ideas and experiences focused on strategies and approaches to building trust between communities and health systems to increase equity in vaccination. Health professionals and community members must have equal value in the design and delivery of community-centered immunization services, while key vaccination decision-makers must also consider community experiences, concerns, and expertise in program design and policymaking. Therefore, strategies for community engagement and cultivating trust with communities are crucial for the success of any immunization program. Furthermore, health workers need additional skills, support, and resources to effectively communicate complex information about immunization, including effective strategies for countering misinformation. This article summarizes three skills-building sessions offered at the VARN2023 conference, focused on human-centered design, motivational interviewing, and engaging with journalists to leverage the voices of communities. These sessions offered practical, evidence-based tools for use across geographic and social settings that can be used by practitioners, researchers, and other stakeholders to increase vaccination demand and uptake in their communities., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: [NPR, KLH, TS, and AG are Sabin employees. TU is a consultant for the Sabin Vaccine Institute (Sabin). GL, CK, DRP, and EL are UNICEF employees. LM, ABC, PK, JS, NR, and MK were VARN2023 delegates who spoke on related topics and/or co-organized and/or facilitated the VARN2023 side events on human-centered design, motivational interviewing, and journalist engagement.]., (Copyright © 2024. Published by Elsevier Ltd.)

Streptococcus mitis is a leading cause of infective endocarditis (IE). However, our understanding of the genomic epidemiology and pathogenicity of IE-associated S. mitis is hampered by low IE incidence. Here we use whole genome sequencing of 129 S. mitis bloodstream infection (BSI) isolates collected between 2001-2016 from clinically diagnosed IE cases in the UK to investigate genetic diversity, antimicrobial resistance, and pathogenicity. We show high genetic diversity of IE-associated S. mitis with virtually all isolates belonging to distinct lineages indicating no predominance of specific lineages. Additionally, we find a highly variable distribution of known pneumococcal virulence genes among the isolates, some of which are overrepresented in disease when compared to carriage strains. Our findings suggest that S. mitis in patients with clinically diagnosed IE is not primarily caused by specific hypervirulent or antimicrobial resistant lineages, highlighting the accidental pathogenic nature of S. mitis in patients with clinically diagnosed IE., (© 2024. The Author(s).)

Background: The COVID-19 pandemic has had a significant impact on different countries because of which various health and safety measures were implemented, with digital media playing a pivotal role. However, digital media also pose significant concerns such as misinformation and lack of direction., Objective: We aimed to explore the effects of COVID-19-related infodemics through digital, social, and electronic media on the vaccine-related attitudes of caregivers and health care providers in Pakistan., Methods: This study employs a qualitative exploratory study design with purposive sampling strategies, and it was conducted at 3 primary health care facilities in the province of Sindh, Pakistan. Seven focus group discussions with health care providers and 60 in-depth interviews with caregivers were conducted using semistructured interviews through virtual platforms (ConnectOnCall and Zoom). Transcripts were analyzed through thematic analysis., Results: Our study reveals the pivotal role of electronic media, mobile health (mHealth), and social media during the COVID-19 pandemic. Four major themes were identified: (1) sources of information on COVID-19 and its vaccination, (2) electronic media value and misleading communication, (3) mHealth leveraging and limitations during COVID-19, and (4) social media influence and barriers during COVID-19. Health care providers and caregivers reported that the common sources of information were electronic media and mHealth, followed by social media. Some participants also used global media for more reliable information related to COVID-19. mHealth solutions such as public awareness messages, videos, call ringtones, and helplines promoted COVID-19 prevention techniques and vaccine registration. However, the overwhelming influx of news and sociobehavioral narratives, including misinformation/disinformation through social media such as WhatsApp, Facebook, and Twitter, were found to be the primary enablers of vaccine-related infodemics. Electronic media and mHealth were utilized more widely to promote information and communication on the COVID-19 pandemic and vaccination. However, social media and electronic media-driven infodemics were identified as the major factors for misinformation related to COVID-19 and vaccine hesitancy. Further, we found a digital divide between the urban and rural populations, with the use of electronic media in rural settings and social media in urban settings., Conclusions: In a resource-constrained setting like Pakistan, the usage of mHealth, social media, and electronic media for information spread (both factual and mis/disinformation) related to COVID-19 and its vaccination had a significant impact on attitudes toward COVID-19 vaccination. Based on the qualitative findings, we generated a model of digital communications and information dissemination to increase knowledge about COVID-19 and its prevention measures, including vaccination, which can be replicated in similar settings for other disease burdens and related infodemics. Further, to mitigate the infodemics, both digital and nondigital interventions are needed at a larger scale., (©Abdul Momin Kazi, Nazia Ahsan, Rawshan Jabeen, Raheel Allana, Saima Jamal, Muhammad Ayub Khan Mughal, Kathryn L Hopkins, Fauzia Aman Malik. Originally published in JMIR Infodemiology (https://infodemiology.jmir.org), 04.09.2024.)

Misinformation related to vaccines has been shown to potentially negatively impact public perceptions and intentions to vaccinate in many contexts including COVID-19 vaccination in Ghana. Psychological inoculation - where recipients are warned about the misleading techniques used in misinformation - is a potential intervention which could preemptively boost public resistance against misinformation. Cranky Uncle Vaccine is an interactive, digital game that applies inoculation, offering a scalable tool building public resilience against vaccine misinformation and promoting positive health-related behaviors. In this study, we document the process of developing and testing a West African version of Cranky Uncle Vaccine , with co-design workshops and a pilot test conducted in urban and peri-urban areas of the Greater Accra region of Ghana with 829 young people who had access to mobile and computer devices. The average age was 21.8 and participants were highly educated (median education level "Some/all university") with slightly more females (51.2%) than males (48.4%). Pilot participants filled out surveys before and after playing the game, measuring vaccine attitudes (pre-game M  = 3.4, post-game M  = 3.6), intent to get vaccinated (pre-game M  = 3.5, post-game M  = 3.6), and discernment between vaccine facts and fallacies (pre-game AUC = 0.72, post-game AUC = 0.75). We observed a significant improvement in attitudes toward vaccines, with players demonstrating increased likelihood to get vaccinated after completing the game. Among players who indicated that they were unlikely to get vaccinated in the pre-game survey ( N  = 52, or 6.3% of participants), just over half of these participants (53%) switched to likely to get vaccinated after playing the game. Perceived reliability of vaccine facts remained the same, while perceived reliability of vaccine fallacies significantly decreased, indicating improved ability to spot misleading arguments about vaccines. These results demonstrate the effectiveness of a digital game in building public resilience against vaccine misinformation as well as improving vaccine attitudes and intent to get vaccinated.

Background: Carbapenemase-producing Enterobacterales (CPE) are challenging in healthcare, with resistance to multiple classes of antibiotics. This study describes the emergence of imipenemase (IMP)-encoding CPE among diverse Enterobacterales species between 2016 and 2019 across a London regional network., Methods: We performed a network analysis of patient pathways, using electronic health records, to identify contacts between IMP-encoding CPE-positive patients. Genomes of IMP-encoding CPE isolates were overlaid with patient contacts to imply potential transmission events., Results: Genomic analysis of 84 Enterobacterales isolates revealed diverse species (predominantly Klebsiella spp, Enterobacter spp, and Escherichia coli); 86% (72 of 84) harbored an IncHI2 plasmid carrying blaIMP and colistin resistance gene mcr-9 (68 of 72). Phylogenetic analysis of IncHI2 plasmids identified 3 lineages showing significant association with patient contacts and movements between 4 hospital sites and across medical specialties, which was missed in initial investigations., Conclusions: Combined, our patient network and plasmid analyses demonstrate an interspecies, plasmid-mediated outbreak of blaIMPCPE, which remained unidentified during standard investigations. With DNA sequencing and multimodal data incorporation, the outbreak investigation approach proposed here provides a framework for real-time identification of key factors causing pathogen spread. Plasmid-level outbreak analysis reveals that resistance spread may be wider than suspected, allowing more interventions to stop transmission within hospital networks.SummaryThis was an investigation, using integrated pathway networks and genomics methods, of the emergence of imipenemase-encoding carbapenemase-producing Enterobacterales among diverse Enterobacterales species between 2016 and 2019 in patients across a London regional hospital network, which was missed on routine investigations., Competing Interests: Potential conflicts of interest. A. C. M. is a majority shareholder in a NEXT Q company that is developing technology support systems for IPC. M. Gi. has received honoraria from Pfizer and Menarini Pharmaceuticals. J. F. T. holds shares in Oxford Nanopore Technologies. K. L. H. has received grant money from Shionogi and consulting fees from Cepheid. J. A. O. has received consulting fees from Gama Healthcare, Biointerations, Spectrum X, and Ondine. All other authors report no potential conflicts. Funding to pay the Open Access publication charges for this article was provided by Imperial College London. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed., (© The Author(s) 2024. Published by Oxford University Press on behalf of Infectious Diseases Society of America.)

OXA-244, an R214G variant of OXA-48, is silently spreading worldwide likely because of difficulties in detection using classical screening media. Here, we characterized two clinical isolates of Escherichia coli and Citrobacter youngae that displayed reduced susceptibility to carbapenems but were lacking significant carbapenemase activity as revealed by negative Carba NP test results. However, positive test results were seen for OXA-48-like enzymes by lateral flow immunoassays. WGS revealed the presence of a bla OXA-181-like gene that codes for OXA-484, an R214G variant of OXA-181. Bla OXA-484 gene was located on a 58.4-kb IncP1-like plasmid (pN-OXA-484), that upon transfer into E. coli HB4 with impaired permeability, conferred carbapenem and temocillin resistance (MICs > 32 mg/L). E. coli TOP10 (pTOPO-OXA-484) revealed reduced MICs in most substrates as compared to E. coli TOP10 (pTOPO-OXA-181), especially for imipenem (0.25 mg/L versus 0.75 mg/L) and temocillin (16 mg/L versus 1028 mg/L). Catalytic efficiencies of OXA-484 were reduced as compared to OXA-181 for most ß-lactams including imipenem and temocillin with 27.5- and 21.7-fold reduction, respectively. Molecular modeling confirmed that the salt bridges between R214, D159, and the R1 substituent's carboxylate group of temocillin were not possible with G214 in OXA-484, explaining the reduced affinity for temocillin. In addition, changes in active site's water network may explain the decrease in hydrolysis rate of carbapenems. OXA-484 has weak imipenem and temocillin hydrolytic activities, which may lead to silent spread due to underdetection using selective screening media or biochemical imipenem hydrolysis confirmatory tests.

Introduction: The periodic evaluation of knowledge, attitudes, and practices (KAP) of healthcare workers, including vaccinators, concerning expanded programs on immunization (EPI) is very crucial for a better healthcare system. This study was carried out to assess the KAP of vaccinators about the EPI, including cold storage of vaccines and their practices related to vaccine cold chain management., Method: A cross-sectional study was conducted from January 2022 to June 2022 among registered vaccinators in the twin cities (Islamabad and Rawalpindi) of Pakistan. A structured self-administered questionnaire (English and Urdu) was developed as per the Pakistan national EPI policy and strategic guidelines 2022 and World Health Organization (WHO) guidelines, as well as from earlier studies (Cronbach's alpha value of 0.734). The final questionnaire consisted of closed-ended questions in four sections, including sociodemographic information, knowledge (with dichotomous variables of yes/no), attitudes (with a 5-point Likert scale ranging from strongly agree to strongly disagree), and handling of vaccines and cold chain management. Completed questionnaires were entered into Microsoft Excel and then imported into SPSS version 25 for statistical analysis., Results: A total of 186 vaccinators completely filled out their questionnaires, with a 97.9% response rate. More than half of the participants (57.5%) had no training related to EPI. Most of the respondents had a moderate to poor level of knowledge regarding EPI. The overall attitude was positive, and 57% of the participants strongly agreed that the national immunization programs can significantly contribute to the decrease in morbidity and mortality rates among children. In the current study, participants showed good practices toward EPI, vaccine storage, and cold chain management. The majority (93.5%) of the participants checked the expiry of vaccines at regular intervals to maintain the first expiry first out (FEFO) in their healthcare setting., Discussion: In conclusion, most of the vaccinators had moderate to poor knowledge, a positive attitude, and good practices toward EPI, vaccine cold storage, and cold chain management. Lack of training among vaccinators on EPI was also observed. These findings have suggested that continuous training, education, and regular supervision of vaccinators in EPI are important for maximum immunization effectiveness and coverage., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Azhar, Rashid, Islam, Akhtar, Hopkins, Sommers, Ikram, Anwer, Maqbool, Khan, Ahmed and Akhtar.)

Background: Hospital sinks are environmental reservoirs that harbour healthcare-associated (HCA) pathogens. Selective pressures in sink environments, such as antibiotic residues, nutrient waste and hardness ions, may promote antibiotic resistance gene (ARG) exchange between bacteria. However, cheap and accurate sampling methods to characterize these factors are lacking., Aims: To validate a workflow to detect antibiotic residues and evaluate water chemistry using dipsticks. Secondarily, to validate boric acid to preserve the taxonomic and ARG ('resistome') composition of sink trap samples for metagenomic sequencing., Methods: Antibiotic residue dipsticks were validated against serial dilutions of ampicillin, doxycycline, sulfamethoxazole and ciprofloxacin, and water chemistry dipsticks against serial dilutions of chemical calibration standards. Sink trap aspirates were used for a 'real-world' pilot evaluation of dipsticks. To assess boric acid as a preservative of microbial diversity, the impact of incubation with and without boric acid at ∼22 °C on metagenomic sequencing outputs was evaluated at Day 2 and Day 5 compared with baseline (Day 0)., Findings: The limits of detection for each antibiotic were: 3 μg/L (ampicillin), 10 μg/L (doxycycline), 20 μg/L (sulfamethoxazole) and 8 μg/L (ciprofloxacin). The best performing water chemistry dipstick correctly characterized 34/40 (85%) standards in a concentration-dependent manner. One trap sample tested positive for the presence of tetracyclines and sulphonamides. Taxonomic and resistome composition were largely maintained after storage with boric acid at ∼22 °C for up to five days., Conclusions: Dipsticks can be used to detect antibiotic residues and characterize water chemistry in sink trap samples. Boric acid was an effective preservative of trap sample composition, representing a low-cost alternative to cold-chain transport., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Integration of genomic technologies into routine antimicrobial resistance (AMR) surveillance in health-care facilities has the potential to generate rapid, actionable information for patient management and inform infection prevention and control measures in near real time. However, substantial challenges limit the implementation of genomics for AMR surveillance in clinical settings. Through a workshop series and online consultation, international experts from across the AMR and pathogen genomics fields convened to review the evidence base underpinning the use of genomics for AMR surveillance in a range of settings. Here, we summarise the identified challenges and potential benefits of genomic AMR surveillance in health-care settings, and outline the recommendations of the working group to realise this potential. These recommendations include the definition of viable and cost-effective use cases for genomic AMR surveillance, strengthening training competencies (particularly in bioinformatics), and building capacity at local, national, and regional levels using hub and spoke models., Competing Interests: Declaration of interests EJ had partial salary cover from Wellcome Trust over the course of this work. KSB reports funding from the Biotechnology and Biological Sciences Council and Medical Research Council and partial salary cover from Wellcome Trust and the UK Health Security Agency over the course of this work. LYH reports funding from Pfizer and honoraria from BioMerieux for lectures in 2022. BE and INO report receiving funding from the UK Department of Health and Social Care: with a grant managed by the Fleming Fund and work performed under the auspices of the SEQAFRICA project. INO reports funding from the Bill & Melinda Gates Foundation, Joint Programming Initiative on Antimicrobial Resistance, Wellcome Trust, Grand Challenges Africa Award, and UK Medical Research Council, royalties for Genetics: Genes, Genomes and Evolution (Oxford University Press) and Divining Without Seeds and for Antimicrobial Resistance in Developing Countries (Springer), consulting fees from Wellcome Trust, and honoraria for Harvard University seminars and Peter Wildy Lecture Award 2023. SJP is a member of the Scientific Advisory Board of Next Gen Diagnostics and was supported by Illumina to attend the European Congress of Clinical Microbiology and Infectious Diseases conference. NAF reports funding from the Bill & Melinda Gates Foundation, UK Research and Innovation, and National Institute for Health and Care Research. All other authors declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY 4.0 license. Published by Elsevier Ltd.. All rights reserved.)

Historically, epidemiological investigation and surveillance for bacterial antimicrobial resistance (AMR) has relied on low-resolution isolate-based phenotypic analyses undertaken at local and national reference laboratories. Genomic sequencing has the potential to provide a far more high-resolution picture of AMR evolution and transmission, and is already beginning to revolutionise how public health surveillance networks monitor and tackle bacterial AMR. However, the routine integration of genomics in surveillance pipelines still has considerable barriers to overcome. In 2022, a workshop series and online consultation brought together international experts in AMR and pathogen genomics to assess the status of genomic applications for AMR surveillance in a range of settings. Here we focus on discussions around the use of genomics for public health and international AMR surveillance, noting the potential advantages of, and barriers to, implementation, and proposing recommendations from the working group to help to drive the adoption of genomics in public health AMR surveillance. These recommendations include the need to build capacity for genome sequencing and analysis, harmonising and standardising surveillance systems, developing equitable data sharing and governance frameworks, and strengthening interactions and relationships among stakeholders at multiple levels., Competing Interests: Declaration of interests KSB reports funding from the Biotechnology and Biological Sciences Research Council and Medical Research Council (MRC) and partial salary cover from Wellcome Trust and the UK Health Security Agency (UKHSA) over the course of this work. EJ had partial salary cover from Wellcome Trust over the course of this work. SWL has received the Robert Austrian Research Award sponsored by Pfizer in 2022. DGA reports funding from the NIHR. INO reports funding from the Bill & Melinda Gates Foundation, Joint Programming Intiative on Antimicrobial Resistance, Wellcome Trust, Grand Challenges Africa Award, and UK MRC; royalties for Genetics: Genes, Genomes and Evolution (Oxford University Press), and Divining Without Seeds and for Antimicrobial Resistance in Developing Countries (Springer); consulting fees from Wellcome Trust; honoraria for Harvard University seminars; and the Peter Wildy Lecture Award 2023. DGA, BE, RSH, and INO report receiving funding from the UK Department of Health and Social Care in the form of a grant managed by the Fleming Fund and work performed under the auspices of the SEQAFRICA project. LS-B reports funding by Conselleria de Sanitat Universal i Salut Pública, Generalitat Valenciana, Valencia (Spain), under Plan GenT (reference number CDEI-06/20-B). NAF reports funding from the Gates Foundation, UK Research and Innovation, and NIHR. SJP is a member of the Scientific Advisory Board of Next Gen Diagnostics and was supported by Illumina to attend the European Congress of Clinical Microbiology and Infectious Disease conference. All other authors declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY 4.0 license. Published by Elsevier Ltd.. All rights reserved.)

Nearly a century after the beginning of the antibiotic era, which has been associated with unparalleled improvements in human health and reductions in mortality associated with infection, the dwindling pipeline for new antibiotic classes coupled with the inevitable spread of antimicrobial resistance (AMR) poses a major global challenge. Historically, surveillance of bacteria with AMR typically relied on phenotypic analysis of isolates taken from infected individuals, which provides only a low-resolution view of the epidemiology behind an individual infection or wider outbreak. Recent years have seen increasing adoption of powerful new genomic technologies with the potential to revolutionise AMR surveillance by providing a high-resolution picture of the AMR profile of the bacteria causing infections and providing real-time actionable information for treating and preventing infection. However, many barriers remain to be overcome before genomic technologies can be adopted as a standard part of routine AMR surveillance around the world. Accordingly, the Surveillance and Epidemiology of Drug-resistant Infections Consortium convened an expert working group to assess the benefits and challenges of using genomics for AMR surveillance. In this Series, we detail these discussions and provide recommendations from the working group that can help to realise the massive potential benefits for genomics in surveillance of AMR., Competing Interests: Declaration of interests KSB reports funding from the Biotechnology and Biological Sciences Research Council and Medical Research Council and partial salary cover from Wellcome Trust and the UK Health Security Agency (UKHSA) over the course of this work. EJ had partial salary cover from Wellcome Trust over the course of this work. RA reports funding unrelated to this study from Novo Nordisk, Roche, Novartis, and UICC, and honoraria (unrelated to this study) from Merck & Co, Novartis, and F Hoffmann-La Roche. BE and INO report receiving funding from the UK Department of Health and Social Care: with a grant managed by the Fleming Fund and work performed under the auspices of the SEQAFRICA project. INO reports funding from the Bill & Melinda Gates Foundation, Joint Programming Initiative in Antimicrobial Resistance, Wellcome Trust, Grand Challenges Africa Award, and UK Medical Research Council, royalties for Genetics: Genes, Genomes and Evolution (Oxford University Press) and Divining Without Seeds and for Antimicrobial Resistance in Developing Countries (Springer), consulting fees from Wellcome Trust, and honoraria for Harvard University seminars and Peter Wildy Lecture Award 2023. LYH reports funding from Pfizer and honoraria from BioMerieux for a lecture in 2022. DMM reports funding from the British Society for Antimicrobial Chemotherapy. NEW reports funding from Nuclear Threat Initiative, Medical Research Council, Open Philantropy, and Shionogi as well as consulting fees from Nuclear Threat Initiative. DMA reports funding from the National Institute for Health and Care Research. NAF reports funding from the Bill & Melinda Gates Foundation, UK Research and Innovation, and National Institute for Health and Care Research. SJP is a member of the scientific advisory board of Next Gen Diagnostics and was supported by Illumina to attend the European Congress of Clinical Microbiology and Infectious Disease conference. All other authors declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY 4.0 license. Published by Elsevier Ltd.. All rights reserved.)

Several bioinformatics genotyping algorithms are now commonly used to characterize antimicrobial resistance (AMR) gene profiles in whole-genome sequencing (WGS) data, with a view to understanding AMR epidemiology and developing resistance prediction workflows using WGS in clinical settings. Accurately evaluating AMR in Enterobacterales, particularly Escherichia coli , is of major importance, because this is a common pathogen. However, robust comparisons of different genotyping approaches on relevant simulated and large real-life WGS datasets are lacking. Here, we used both simulated datasets and a large set of real E. coli WGS data ( n =1818 isolates) to systematically investigate genotyping methods in greater detail. Simulated constructs and real sequences were processed using four different bioinformatic programs (ABRicate, ARIBA, KmerResistance and SRST2, run with the ResFinder database) and their outputs compared. For simulation tests where 3079 AMR gene variants were inserted into random sequence constructs, KmerResistance was correct for 3076 (99.9 %) simulations, ABRicate for 3054 (99.2 %), ARIBA for 2783 (90.4 %) and SRST2 for 2108 (68.5 %). For simulation tests where two closely related gene variants were inserted into random sequence constructs, KmerResistance identified the correct alleles in 35 338/46 318 (76.3 %) simulations, ABRicate identified them in 11 842/46 318 (25.6 %) simulations, ARIBA identified them in 1679/46 318 (3.6 %) simulations and SRST2 identified them in 2000/46 318 (4.3 %) simulations. In real data, across all methods, 1392/1818 (76 %) isolates had discrepant allele calls for at least 1 gene. In addition to highlighting areas for improvement in challenging scenarios, (e.g. identification of AMR genes at <10× coverage, identifying multiple closely related AMR genes present in the same sample), our evaluations identified some more systematic errors that could be readily soluble, such as repeated misclassification (i.e. naming) of genes as shorter variants of the same gene present within the reference resistance gene database. Such naming errors accounted for at least 2530/4321 (59 %) of the discrepancies seen in real data. Moreover, many of the remaining discrepancies were likely 'artefactual', with reporting of cut-off differences accounting for at least 1430/4321 (33 %) discrepants. Whilst we found that comparing outputs generated by running multiple algorithms on the same dataset could identify and resolve these algorithmic artefacts, the results of our evaluations emphasize the need for developing new and more robust genotyping algorithms to further improve accuracy and performance.

Background: While the COVID-19 pandemic has exposed the vulnerabilities of immunization delivery systems globally, the devastating impact of the pandemic on immunization delivery is most pronounced in low and middle-income countries like Pakistan. We conducted a qualitative study to capture the views and experiences of parents and healthcare workers (HWs) and assess the impact of the COVID-19 pandemic on childhood routine immunization (CRI) and COVID-19 vaccination in Pakistan., Methods: We used a qualitative research design with a purposive sampling approach. Semi-structured interviews (via telephone) and focus group discussions (via Zoom) were conducted with parents/child caregivers and HWs, respectively. All qualitative interviews were conducted between February and July 2021 from three sites (two urban and one rural) in Sindh, Pakistan. Interviews were audio-recorded, transcribed, and coded for a team-based thematic analysis., Results: Overall, most parents and HWs indicated a strong trust in the benefits of CRI; nonetheless, a substantial disruption in the delivery and uptake of these services was also reported. The barriers towards CRI included closed vaccination centers, drastic reduction in outreach programs, lack of information for parents/child caregivers on vaccine availability, fear in the community regarding vaccine safety, limited vaccine supply, and a lack of healthcare staff. For COVID-19 vaccines, challenges cited included skepticism about the reality of the pandemic and confusion over COVID-19 vaccines due to conflicting (or mis-or-dis) information. Both participant groups showed a willingness to integrate COVID-19 vaccination into Pakistan's Expanded Program for Immunization if required in the future., Conclusion: During the COVID-19 pandemic, disruptions of regular immunization delivery in Pakistan were not due to parental unwillingness to vaccinate, but rather to social and logistical challenges caused by a rapidly changing context and difficulties in providing vaccination services safely. Barriers to vaccine access and concerns about COVID-19 exposure during clinic visits also contributed to uncertainty regarding immunization services early in the pandemic. For catchup campaigns and future pandemics, more than focusing interventions on persuading people, strategic approaches to building resilience through system-based interventions, such as investing in surge capacity in the immunization workforce to bounce back quickly after the first shock are required., (© 2023. The Author(s).)

Misinformation can decrease public confidence in vaccines, and reduce vaccination intent and uptake. One strategy for countering these negative impacts comes from inoculation theory. Similar to biological vaccination, inoculation theory posits that exposure to a weakened form of misinformation can develop cognitive immunity, reducing the likelihood of being misled. Online games offer an interactive, technology-driven, and scalable solution using an active form of inoculation that engages and incentivizes players to build resilience against misinformation. We document the development of the critical thinking game Cranky Uncle Vaccine . The game applies research findings from inoculation theory, critical thinking, humor in science communication, and serious games. The game content was iterated through a series of co-design workshops in Kampala (Uganda), Kitale (Kenya), and Kigali (Rwanda). Workshop participants offered feedback on cartoon character design, gameplay experience, and the game's content, helping to make the game more culturally relevant and avoid unintended consequences in East African countries. Our co-design methodology offers an approach for further adaptation of the Cranky Uncle Vaccine game to other regions, as well as a template for developing locally relevant interventions to counter future infodemics.

The first conference of the Vaccination Acceptance Research Network, VARN2022: Shaping Global Vaccine Acceptance with Localized Knowledge, was held virtually, from March 1 st to 3 rd 2022. This inaugural event brought together a global representation of experts to discuss key priorities and opportunities emerging across the ecosystem of vaccine acceptance and demand, from policies to programs and practice. Convened by the Sabin Vaccine Institute, VARN aims to support dialogue among multidisciplinary stakeholders to enhance the uptake of social and behavioral science-based solutions for vaccination decision-makers and implementers. The conference centered around four key themes: 1) Understanding vaccine acceptance and its drivers; 2) One size does not fit all: community- and context-specific approaches to increase vaccine acceptance and demand; 3) Fighting the infodemic and harnessing social media for good; and 4) Frameworks, data integrity and evaluation of best practices. Across the conference, presenters and participants considered the drivers of and strategies to increase vaccine acceptance and demand relating to COVID-19 vaccination and other vaccines across the life-course and across low-, middle- and high-income settings. VARN2022 provided a wealth of evidence from around the world, highlighting the need for human-centered, multi-sectoral and transdisciplinary approaches to improve vaccine acceptance and demand. This report summarizes insights from the diverse presentations and discussions held at VARN2022, which will form a roadmap for future research, policy making, and interventions to improve vaccine acceptance and demand globally., (© 2023. The Author(s).)

Background: We aimed to describe the UK Pseudomonas aeruginosa population structure amongst people with cystic fibrosis (PWCF), and to examine evidence for cross-infection., Methods: Variable Number Tandem Repeat (VNTR) typing was performed on 4640 isolates from 2619 PWCF received from 55 hospital laboratories between 2017 and 2019. A combination of whole genome sequence (WGS)-based analysis of four clusters from one hospital, and epidemiological analysis of shared strains in twelve hospitals evaluated cross-infection., Results: Of 2619 PWCF, 1324 (51%) harboured common clusters or known transmissible strains, while 1295 carried unique strains/those shared among small numbers of patients. Of the former, 9.5% (250 patients) harboured the Liverpool epidemic strain (LES), followed in prevalence by clone C (7.8%; 205 patients), cluster A (5%;130 patients), and cluster D (3.6%; 94 patients). WGS analysis of 10 LES isolates, 9 of cluster D and 6 isolates each of cluster A and clone C from one hospital revealed LES formed the tightest cluster (between 7 and 205 SNPs), and cluster D the loosest (between 53 and 1531 SNPs). Hospital-specific shared strains were found in some centres, although cross-infection was largely historical, with few new acquisitions. Fifty-nine PWCF (2.3%) harboured "high-risk" clones; one ST235 isolate carried a bla IMP-1 allele., Conclusion: Of 2619 PWCF who had P. aeruginosa isolates submitted for VNTR, 51% harboured either common clusters or known transmissible strains, of which LES was the most common. Limited evidence of recent patient-to-patient strain transmission was found, suggesting cross-infection prevention measures and surveillance effectively reduce transmission., Competing Interests: Declaration of Competing Interest None of the authors have had any conflicts of interest to declare relating to this study., (Copyright © 2023. Published by Elsevier B.V.)

Vaccination is critical to minimize serious illness and death from COVID-19. Yet uptake of COVID-19 vaccines remains highly variable, particularly among marginalized communities. This article shares lessons learned from four UNICEF interventions that supported Governments to generate acceptance and demand for COVID-19 vaccines in Zambia, Iraq, Ghana, and India. In Zambia, community rapid assessment provided invaluable real-time insights around COVID-19 vaccination and allowed the identification of population segments that share beliefs and motivations regarding COVID-19 vaccination. Findings were subsequently used to develop recommendations tailored to the different personas. In Iraq, a new outreach approach (3iS: Intensification of Integrated Immunization) utilized direct community engagement to deliver health messages and encourage service uptake, resulting in over 4.4 million doses of COVID-19 and routine immunization vaccines delivered in just 8 months. In Ghana, a human-centered design initiative was applied to co-develop community-informed strategies to improve COVID-19 vaccination rates. In India, a risk communication and community engagement initiative reached half a million people over six months, translating into a 25% increase in vaccination rates. These shared approaches can be leveraged to improve COVID-19 vaccination coverage and close gaps in routine immunization across diverse and marginalized communities.

Introduction. Combination of PCR and Elek testing to identify toxigenic corynebacteria has revealed organisms described as non-toxigenic toxin-gene bearing (NTTB) Corynebacterium diphtheriae or C. ulcerans (i.e. PCR tox positive; Elek negative). These organisms carry part or all of tox , but are unable to express diphtheria toxin (DT) and present a challenge to clinical and public health case management. Gap analysis/Hypothesis. There are few data on the theoretical risk of NTTB reversion to toxigenicity. This unique cluster and subsequent epidemiologically linked isolates allowed the opportunity to determine any change in DT expression status. Aim. To characterize a cluster of infections due to NTTB in a skin clinic and subsequent cases in two household contacts. Methodology. Epidemiological and microbiological investigations were carried out according to existing national guidance at the time. Susceptibility testing used gradient strips. The tox operon analysis and multi-locus sequence typing (MLST) was derived from whole-genome sequencing. Alignment of the tox operon and phylogenetic analyses were performed using clustalW, mega, the public core-genome MLST (cgMLST) scheme and an in-house bioinformatic single nucleotide polymorphism (SNP) typing pipeline. Results. Isolates of NTTB C. diphtheriae were recovered from four cases (cases 1 to 4) with epidermolysis bullosa attending the clinic. Two further isolates were subsequently recovered from case 4, >18 months later, and from two household contacts (cases 5 and 6) after a further 18 months and 3.5 years, respectively. All eight strains were NTTB C. diphtheriae biovar mitis, belonged to the same sequence type (ST-336) with the same deletion in tox . Phylogenetic analysis showed relatively high diversity between the eight strains with 7-199 SNP and 3-109 cgMLST loci differences between them. The number of SNPs between the three isolates from case 4 and two household contacts (cases 5 and 6) was 44-70 with 28-38 cgMLST loci differences. Conclusions. We report a cluster of NTTB C. diphtheriae cases in a skin clinic and evidence of onward household transmission. We conclude the deletion in the tox was responsible for the non-expression of DT. There was no evidence of reversion to DT expression over the 6.5 year period studied. These data informed revision to guidance in the management of NTTB cases and their contacts in the UK.

Introduction: Acquired carbapenemase-producing Gram-negative bacteria are an increasing public health concern globally and have been mandatory to report in England since October 2020. However, in light of the COVID-19 (SARS-CoV-2) pandemic, the Royal College of Pathologists (RCPath) released new guidance "for reducing the need for screening of CRE (carbapenem-resistant Enterobacterales) […] in low-risk areas", without defining "low risk"., Methods: To assess the impact of the RCPath recommendations on screening of carbapenemase-producing Enterobacterales (CPE), an online Select Survey was sent to all NHS acute hospitals in England. The initial survey distribution was between March and April 2021 and the survey was relaunched between November 2021 and March 2022., Results: In total, 54 hospitals completed the survey, representing 39.1% of 138 eligible Trusts. All hospitals had a CPE screening policy in place, and the majority of these reflect UKHSA's Framework of actions to contain CPE. Of the 23 hospitals who reported a reduction in CPE screening, only three (13.0%) indicated that this was due to the RCPath recommendations, with 21 (91.3%) indicating that there had been a natural reduction in the number of patients admitted to the Trust who would have previously been screened due to the COVID-19 pandemic., Conclusion: For most surveyed hospitals, CPE screening was not reduced due to the RCPath recommendations. However, the results highlighted that there is a large amount of individual variation in CPE screening practices and diagnostic testing between hospitals., (Crown Copyright © 2023 Published by Elsevier Ltd on behalf of The Healthcare Infection Society.)

Empirical data regarding dynamic alterations in illicit drug supply markets in response to the COVID-19 pandemic, including the potential for introduction of novel drug substances and/or increased poly-drug combination use at the "street" level, that is, directly proximal to the point of consumption, are currently lacking. Here, a high-throughput strategy employing ambient ionization-mass spectrometry is described for the trace residue identification, characterization, and longitudinal monitoring of illicit drug substances found within >6,600 discarded drug paraphernalia (DDP) samples collected during a pilot study of an early warning system for illicit drug use in Melbourne, Australia from August 2020 to February 2021, while significant COVID-19 lockdown conditions were imposed. The utility of this approach is demonstrated for the de novo identification and structural characterization of β-U10, a previously unreported naphthamide analog within the "U-series" of synthetic opioid drugs, including differentiation from its α-U10 isomer without need for sample preparation or chromatographic separation prior to analysis. Notably, β-U10 was observed with 23 other drug substances, most commonly in temporally distinct clusters with heroin, etizolam, and diphenhydramine, and in a total of 182 different poly-drug combinations. Longitudinal monitoring of the number and weekly "average signal intensity" (ASI) values of identified substances, developed here as a semi-quantitative proxy indicator of changes in availability, relative purity and compositions of street level drug samples, revealed that increases in the number of identifications and ASI for β-U10 and etizolam coincided with a 50% decrease in the number of positive detections and an order of magnitude decrease in the ASI for heroin., (© 2022 The Authors. Drug Testing and Analysis published by John Wiley & Sons Ltd.)

Female sex workers (FSWs) in South Africa experience a uniquely high prevalence of HIV. We describe the HIV cascade of care (CoC) in FSWs in South Africa, and explored service utilisation at sex work programmes. A cross-sectional, study enrolled FSWs across 12 sites in South Africa. Participants were recruited using chain-referral method. Inclusion criteria: ≥ 18 years, cis-gender female, sold/transacted in sex, HIV positive. 1862 HIV positive FSWs were enrolled. 92% were known positive, 87% were on antiretroviral treatment (ART). Of those on ART, 74% were virally suppressed. Younger FSWs were significantly less likely to be on ART or virally suppressed. Female sex workers using HIV services from specialised programs were 1.4 times more likely to be virally suppressed than non-program users. The pre-COVID-19 pandemic HIV CoC amongst FSWs in South Africa shows striking improvement from previous estimates, and approaches achievement of 90:90:90 goals., (© 2022. The Author(s).)

Introduction . The New Delhi metallo-β-lactamase (NDM) variant NDM-5 was first described in 2011 in an isolate of Escherichia coli . We noted that a high proportion of isolates of E. coli positive for bla NDM carbapenemase genes submitted to the UK Health Security Agency (formerly Public Health England) between 2019 and mid-2021 carried the bla NDM-5 allele, with many co-harbouring rmtB , rendering them highly resistant to aminoglycosides as well as to most β-lactams. Hypothesis/Gap Statement . This observation suggested that a common plasmid may be circulating. Aim . To compare these isolates and describe the plasmids carrying these resistance elements. Methodology . All isolates were sequenced on an Illumina platform, with five also subjected to long-read nanopore sequencing to provide complete assemblies. The locations of bla NDM-5 , rmtB and other associated genetic elements were identified. Susceptibility testing to a wide range of antibiotics was carried out on representative isolates. Results . The 34 isolates co-harbouring bla NDM-5 and rmtB were from 14 hospital groups and six different regions across England and consisted of 11 distinct sequence types. All carried IncF plasmids. Assembly of the NDM plasmids in five isolates revealed that they carried rmtB and bla NDM-5 in an IncF conjugative plasmid ranging in size from 85.5 to 161 kb. All carried a highly conserved region, previously described in E. coli plasmid pHC105-NDM, that included bla TEM-1B and rmtB followed by sequence bounded by two IS 26 elements containing ΔIS Aba125 , bla NDM-5 , ble , trpF and tat followed by IS CR1 and an integron with sul1 , aadA2 and dfrA12 cassettes. This arrangement has been described in isolates from other countries and continents, suggesting that such plasmids are widely distributed, at least in E. coli , with similar plasmids also found in Klebsiella pneumoniae . Tested isolates were resistant to most antibiotics except colistin, fosfomycin and tigecycline. Conclusion . These observations suggest that conjugative plasmids carrying a highly conserved resistance gene segment have become widespread in England and elsewhere. This study highlights the value of routine whole-genome sequencing in identifying genetic elements responsible for resistance dissemination.

Introduction. The 16S rRNA methyltransferase (16S RMTase) gene armA is the most common mechanism conferring high-level aminoglycoside resistance in Acinetobacter baumannii , although rmtA , rmtB , rmtC , rmtD and rmtE have also been reported. Hypothesis/Gap statement. The occurrence of 16S RMTase genes in A. baumannii in the UK and Republic of Ireland is currently unknown. Aim. To identify the occurrence of 16S RMTase genes in A. baumannii isolates from the UK and the Republic of Ireland between 2004 and 2015. Methodology. Five hundred and fifty pan-aminoglycoside-resistant A. baumannii isolates isolated from the UK and the Republic of Ireland between 2004 and 2015 were screened by PCR to detect known 16S RMTase genes, and then whole-genome sequencing was conducted to screen for novel 16S RMTase genes. Results. A total of 96.5 % (531/550) of isolates were positive for 16S RMTase genes, with all but 1 harbouring armA (99.8 %, 530/531). The remaining isolates harboured rmtE3 , a new rmtE variant. Most (89.2 %, 473/530) armA -positive isolates belonged to international clone II (ST2), and the rmtE3 -positive isolate belonged to ST79. rmtE3 shared a similar genetic environment to rmtE2 but lacked an IS CR20 element found upstream of rmtE2 . Conclusion. This is the first report of rmtE in A. baumannii in Europe; the potential for transmission of rmtE3 to other bacterial species requires further research.

Introduction. Increasing numbers of carbapenemase-producing Enterobacterales (CPE), which can be challenging to treat, have been referred to the national reference laboratory in England since the early 2000s. Gap Statement/Aim. Previous studies on CPE in the UK have focussed on localized outbreaks. We applied whole-genome sequencing (WGS) to isolates referred to the national reference laboratory over 30 months to inform our understanding of CPE epidemiology in England. Methodology. The first confirmed CPE from each new patient referred by an English diagnostic laboratory between 1 January 2014 and 30 June 2016 was sequenced on an Illumina HiSeq 2500. Multiple isolates from the same patient were included from either different species or the same species with different carbapenemase genes. The data were analysed using an in-house bioinformatics pipeline that determines species identification, multi-locus sequence typing (MLST) profile and antimicrobial resistance gene content. Results. A total of 2658 non-duplicate CPE were sequenced amongst which three host organisms belonging to diverse sequence types (STs) predominated: Klebsiella pneumoniae (1380/2658, 51.9 %; 177 STs), Escherichia coli (723/2658, 27.2 %; 133 STs) and Enterobacter cloacae (294/2658, 11.1 %; 88 STs). Thirty different carbapenemase gene variants were identified, although bla OXA-48-like (1122/2658, 42.2%), bla NDM (692/2658, 26.0 %), bla KPC CPE referred to the national reference laboratory are diverse, suggesting multiple introductions to England and a role for horizontal transfer of carbapenemase genes in English CPE epidemiology.bla VIM (100/2658, 3.8 %) and bla IMP (33/2658, 1.2 %) predominated. ST/carbapenemase gene pairings represented widely distributed high-risk clones or clusters at a regional or hospital level. Conclusion. CPE referred to the national reference laboratory are diverse, suggesting multiple introductions to England and a role for horizontal transfer of carbapenemase genes in English CPE epidemiology.

16S rRNA methyltransferase (16S RMTase) genes confer high-level aminoglycoside resistance, reducing treatment options for multidrug-resistant Gram-negative bacteria. Pseudomonas aeruginosa isolates (n = 221) exhibiting high-level pan-aminoglycoside resistance (amikacin, gentamicin and tobramycin MICs ≥64, ≥32 and ≥32 mg/L, respectively) were screened for 16S RMTase genes to determine their occurrence among isolates submitted to a national reference laboratory from December 2003 to December 2015. 16S RMTase genes were identified using two multiplex PCRs, and whole-genome sequencing (WGS) was used to identify other antibiotic resistance genes, sequence types (STs) and the genetic environment of 16S RMTase genes. 16S RMTase genes were found in 8.6% (19/221) of isolates, with rmtB4 (47.4%; 9/19) being most common, followed by rmtD3 (21.1%; 4/19), rmtF2 (15.8%; 3/19) and single isolates harbouring rmtB1, rmtC and rmtD1. Carbapenemase genes were found in 89.5% (17/19) of 16S RMTase-positive isolates, with bla VIM (52.9%; 9/17) being most common. 16S RMTase genes were found in 'high-risk' clones known to harbour carbapenemase genes (ST233, ST277, ST357, ST654 and ST773). Analysis of the genetic environment of 16S RMTase genes identified that IS6100 was genetically linked to rmtB1; IS91 to rmtB4, rmtC or rmtD3; ISCR14 to rmtD1; and rmtF2 was linked to Tn3, IS91 or Tn1721. Although 16S RMTase genes explained only 8.6% of pan-aminoglycoside resistance in the P. aeruginosa isolates studied, the association of 16S RMTase genes with carbapenemase-producers and 'high-risk' clones highlights that continued surveillance is required to monitor spread as well as the importance of suppressing the emergence of dually-resistant clones in hospital settings., (Crown Copyright © 2022. Published by Elsevier Ltd. All rights reserved.)

Transferable linezolid resistance due to optrA , poxtA , cfr and cfr -like genes is increasingly detected in enterococci associated with animals and humans globally. We aimed to characterize the genetic environment of optrA in linezolid-resistant Enterococcus faecalis isolates from Scotland. Six linezolid-resistant E. faecalis isolated from urogenital samples were confirmed to carry the optrA gene by PCR. Short read (Illumina) sequencing showed the isolates were genetically distinct (>13900 core SNPs) and belonged to different MLST sequence types. Plasmid contents were examined using hybrid assembly of short and long read (Oxford Nanopore MinION) sequencing technologies. The optrA gene was located on distinct plasmids in each isolate, suggesting that transfer of a single plasmid did not contribute to optrA dissemination in this collection. pTM6294-2, BX5936-1 and pWE0438-1 were similar to optrA -positive plasmids from China and Japan, while the remaining three plasmids had limited similarity to other published examples. We identified the novel Tn 6993 transposon in pWE0254-1 carrying linezolid ( optrA ), macrolide ( ermB ) and spectinomycin [ANT(9)-Ia] resistance genes. OptrA amino acid sequences differed by 0-20 residues. We report multiple variants of optrA on distinct plasmids in diverse strains of E. faecalis . It is important to identify the selection pressures driving the emergence and maintenance of resistance against linezolid to retain the clinical utility of this antibiotic.

Introduction: Sex workers in South Africa experience high levels of trauma and mental health issues, but little is known about their drug and alcohol use. This study assessed the prevalence of substance use and its key risk factors amongst female sex workers (FSWs) at two sites in South Africa., Methods: Two cross-sectional studies were conducted, in Soweto and Klerksdorp, South Africa. Using respondent-driven sampling (RDS) 508 FSWs in Soweto and 156 in Klerksdorp were enrolled. A study-specific survey was used to collect social and demographic information, substance use, mental ill-health, and HIV status. Raw and RDS-adjusted data were analyzed using Chi-squared tests of association. Weighted and unweighted Poisson regression models were used to assess key risk factors for alcohol and drug use at both univariate and multivariate levels., Results: Of the 664 FSWs, 56.2% were binge drinkers and 29.4% reported using drugs within the last year. Living in a home with regular food (RR: 1.2597, 95% CI: 1.1009-1.4413) and being HIV positive (RR: 1.1678, 95% CI: 1.0227-1.3334) were associated with a higher risk of binge drinking. Having symptoms suggestive of post-traumatic stress disorder (RR: 1.1803, 95% CI: 1.0025-1.3895) and past year physical/sexual abuse from either intimate (RR: 1.3648, 95% CI: 1.1522-1.6167) or non-intimate partners (RR: 1.3910, 95% CI: 1.1793-1.6407) were associated with a higher risk of drug use., Discussion: In conclusion, our findings demonstrate a high prevalence of alcohol and drug use among FSWs in Soweto and Klerksdorp with site-specific contextual dynamics driving substance use. Site differences highlight the importance of tailoring site-specific substance use harm mitigation for this key population., Competing Interests: The authors have declared that no competing interests exist.

Background: Globally female sex workers (FSWs) are vulnerable to violence from intimate partners, police and clients due to stigma and criminalisation. In this paper we describe South African FSWs' exposure to violence and factors associated with having been raped in the past year., Methods: We conducted a multi-stage, community-centric, cross-sectional survey of 3005 FSWs linked to sex worker programmes in 12 sites across all nine provinces that had a SW programme. Adult women who sold sex in the preceding six months were recruited for interviews via sex worker networks. Survey tools were developed in consultation with peer educators and FSWs., Results: In the past year, 70.4% of FSWs experienced physical violence and 57.9% were raped: by policemen (14.0%), clients (48.3%), other men (30.2%) and/or and intimate partner (31.9%). Sexual IPV was associated with food insecurity, entering sex work as a child, childhood trauma exposure, post-traumatic stress disorder (PTSD), drinking alcohol to cope with sex work, working more days, partner controlling behaviour, having an ex-client partner, and having no current partner to protect from ex-partners. Rape by a client, other men or policemen was associated with food insecurity, childhood trauma, PTSD, depression, using alcohol and drugs, being homeless or staying in a sex work venue, selling sex on the streets, working more days and having entered sex work as a child and been in sex work for longer., Conclusion: South African FSWs are very vulnerable to rape. Within the social climate of gender inequality, sex work stigma, criminalisation, and repeated victimisation, the key drivers are structural factors, childhood and other trauma exposure, mental ill-health, circumstances of sex work and, for SIPV, partner characteristics. Mostly these are amenable to intervention, with legislative change being foundational for ending abuse by policemen, enhancing safety of indoor venues and providing greater economic options for women., Competing Interests: The authors have no conflicts of interest to declare., (© 2021 The Authors.)

Objectives: To determine the prevalence of 16S rRNA methyltransferase- (16S RMTase-) producing Gram-negative bacteria in patients in the UK and to identify potential risk factors for their acquisition., Methods: A 6 month prospective surveillance study was conducted from 1 May to 31 October 2016, wherein 14 hospital laboratories submitted Acinetobacter baumannii, Enterobacterales and Pseudomonas aeruginosa isolates that displayed high-level amikacin resistance according to their testing methods, e.g. no zone of inhibition with amikacin discs. Isolates were linked to patient travel history, medical care abroad, and previous antibiotic exposure using a surveillance questionnaire. In the reference laboratory, isolates confirmed to grow on Mueller-Hinton agar supplemented with 256 mg/L amikacin were screened by PCR for 16S RMTase genes armA, rmtA-rmtH and npmA, and carbapenemase genes (blaKPC, blaNDM, blaOXA-48-like and blaVIM). STs and total antibiotic resistance gene complement were determined via WGS. Prevalence was determined using denominators for each bacterial species provided by participating hospital laboratories., Results: Eighty-four isolates (44.7%), among 188 submitted isolates, exhibited high-level amikacin resistance (MIC >256 mg/L), and 79 (94.0%) of these harboured 16S RMTase genes. armA (54.4%, 43/79) was the most common, followed by rmtB (17.7%, 14/79), rmtF (13.9%, 11/79), rmtC (12.7%, 10/79) and armA + rmtF (1.3%, 1/79). The overall period prevalence of 16S RMTase-producing Gram-negative bacteria was 0.1% (79/71 063). Potential risk factors identified through multivariate statistical analysis included being male and polymyxin use., Conclusions: The UK prevalence of 16S RMTase-producing Gram-negative bacteria is low, but continued surveillance is needed to monitor their spread and inform intervention strategies., (© Crown copyright 2021.)

Objective: A screening centre in Soweto, South Africa (SA), investigated high-risk human papillomavirus (HR-HPV), HIV, cervical cancer risk amongst women., Methods: This cross-sectional study (June 2018-March 2019) describes screening results (Roche Linear Array HPV test and Pap smear liquid based cytology) and history of screening (known HIV status, antiretroviral therapy [ART] use, previous Pap smears). Data were stratified by age group (18-29, 30+ years), HIV status, Pap smear results and tested for statistical significance., Results: Of 280 women, 20.4% were HIV-positive, 18.2% had abnormal Pap smears, 41.8% had HR-HPV. Of older women, 48.2% (n = 78/162) had never had a Pap smear. Of younger women, 89.0% (n = 105/118) never had a Pap smear, but had significantly more low-grade squamous intraepithelial lesions (LSIL) and other HR-HPV infection than older women (12.7%[n = 15/118] vs 4.9%[n = 8/162], p = 0.0193; and 49.2%[n = 58/118] vs 29.0%[n = 47/162], p = 0.0006; respectively). HIV-positive women had more abnormal cytology results and infection with other HR-HPV types or co-infection with other HR-HPV type(s)/HPV-16 compared to HIV-negative women (35.1%[n = 20/57] vs 13.9%[n = 31/223], p = 0.0002; 56.1%[n = 32/57] vs 32.7%[n = 73/223], p = 0.001; and 12.3%[n = 7/57] vs 4.9%[n = 11/223], p = 0.044; respectively). Of 57 HIV-positive women, 45.6% (n = 26) already knew their HIV status; of which 69.2% were on ART and 34.6% never had a Pap smear., Conclusion: South African women have high rates of HIV, Pap smear abnormalities and HR-HPV, with low cervical cancer screening coverage. SA cervical cancer screening policy excludes (undiagnosed) HIV-positive and HIV-negative women <30 years, both populations found to have high prevalence of HR-HPV. HPV-based primary screening from 25 years could improve outcomes., Competing Interests: The authors have declared that no competing interests exist.

Background: South Africa is disproportionately impacted by non-communicable diseases (NCDs) and HIV/AIDS. We investigated the prevalence of known/unknown NCD risk factors, HIV, and NCD risk factor-HIV comorbidity; and treatment status on known diseases to determine the prevalence of controlled/uncontrolled disease., Methods: This cross-sectional study (June 2018-March 2019) within an integrated testing centre in Soweto, South Africa, screened adults (aged ≥18 years) for body mass index (BMI), hypertension (HT), rapid glucose and cholesterol, and HIV. Results were stratified by age group, sex, HIV-status, and self-reported ART use. Analysis included Fisher's exact, chi-squared, Kruskal Wallis, and Student's T-tests., Findings: Of 780 enrolled participants, 19.2% were HIV-positive, 37.5% were overweight/obese, 18.0% hypertensive, 10.8% hyperglycaemic, and 8.1% had hypercholesterolaemia. Significantly more women had overweight/obese BMI than men (46.8% vs 19.7%; p <0.0001), and women aged 25-34 years had significantly more hypercholesterolaemia than same-aged men (18.2% vs 5.6%; p  = 0.02). HIV-positive participants had significantly more hyperglycaemia than HIV-negative participants (16.1% vs 9.6%; p  = 0.02), and those on ART (63.9%) had significantly more hypercholesterolaemia than those not on ART (21.7% vs. 4.9%; p  = 0.002). Of participants with HT, hyperglycaemia, and hypercholesterolaemia; 72.4%, 96.1%, and 93.3% were newly diagnosed. All participants with previously diagnosed NCDs remained with uncontrolled disease., Interpretation: There is a high burden of HIV, NCD risk factors, and comorbidity in Soweto, and amongst young adults (18-34 years), especially women. Lowering age requirements for glucose/cholesterol screening to 18+ years, regardless of BMI, HIV-status, or ART use, may yield timely NCD diagnosis/management., Competing Interests: All authors declare that this project has been supported by the President's Emergency Plan for AIDS Relief (PEPFAR) through the Centres for Disease Control and Prevention (CDC) under the terms of CoAg 1U2GGH001150 with the South African Medical Research Council (SAMRC). KLH declares that SAMRC sponsored travel to attend two meetings in Cape Town at the SAMRC for writing workshops for this manuscript. JD, TD, and GEG are funded by the SAMRC., (© 2021 The Author(s).)

Background: Early and accurate treatment of infections due to carbapenem-resistant organisms is facilitated by rapid diagnostics, but rare resistance mechanisms can compromise detection. One year after a Guiana Extended-Spectrum (GES)-5 carbapenemase-positive Klebsiella oxytoca infection was identified by whole-genome sequencing (WGS; later found to be part of a cluster of 3 cases), a cluster of 11 patients with GES-5-positive K. oxytoca was identified over 18 weeks in the same hospital., Methods: Bacteria were identified by matrix-assisted laser desorption/ionization-time of flight mass spectrometry, antimicrobial susceptibility testing followed European Committee on Antimicrobial Susceptibility Testing guidelines. Ertapenem-resistant isolates were referred to Public Health England for characterization using polymerase chain reaction (PCR) detection of GES, pulsed-field gel electrophoresis (PFGE), and WGS for the second cluster., Results: The identification of the first GES-5 K. oxytoca isolate was delayed, being identified by WGS. Implementation of a GES-gene PCR informed the occurrence of the second cluster in real time. In contrast to PFGE, WGS phylogenetic analysis refuted an epidemiological link between the 2 clusters; it also suggested a cascade of patient-to-patient transmission in the later cluster. A novel GES-5-encoding plasmid was present in K. oxytoca, Escherichia coli, and Enterobacter cloacae isolates from unlinked patients within the same hospital group and in human and wastewater isolates from 3 hospitals elsewhere in the United Kingdom., Conclusions: Genomic sequencing revolutionized the epidemiological understanding of the clusters; it also underlined the risk of covert plasmid propagation in healthcare settings and revealed the national distribution of the resistance-encoding plasmid. Sequencing results also informed and led to the ongoing use of enhanced diagnostic tests for detecting carbapenemases locally and nationally., (© The Author(s) 2019. Published by Oxford University Press for the Infectious Diseases Society of America.)

Carbapenem resistance in Gram-negative bacteria is a public health concern. Consequently, numerous government and agency reports discuss carbapenem-resistant Enterobacterales (CRE) and carbapenem-resistant organisms (CROs). Unfortunately, these terms are fuzzy. Do they include (1) Proteeae with inherent imipenem resistance; (2) porin-deficient Enterobacterales resistant to ertapenem but not other carbapenems; (3) Enterobacterales with OXA-48-like enzymes that remain "carbapenem susceptible" at breakpoint; and (4) Pseudomonas aeruginosa that merely lack porin OprD? Counting CPE or CPOs is better but still insufficient, because different carbapenemases have differing treatment implications, particularly for new β-lactam/β-lactamase inhibitor combinations. At the least, it is essential for authors, journals, and regulatory agencies to specify the carbapenemases meant. The future may demand even greater precision, for mutations can alter hydrolytic activity, and the ability to confer resistance, within carbapenemase families., (© The Author(s) 2020. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

Introduction: South Africa is the HIV epidemic epicentre; however, non-communicable diseases (NCDs) will be the most common cause of death by 2030. To improve identification and initiation of care for HIV and NCDs, we assessed proportion of clients referred and linked to care (LTC) for abnormal/positive screening results and time to LTC and treatment initiation from a HIV Testing Services (HTS) Centre before and after integrated testing for NCDs with optional peer-navigated linkage to care., Materials and Methods: This two-phase prospective study was conducted at an adult HTS Centre in Soweto, South Africa. Phase 1 (February-June 2018) utilised standard of care (SOC) HTS services (blood pressure [BP], HIV rapid diagnostic testing (RDT), sexually transmitted infections [STI]/Tuberculosis [TB] symptom screening) with passive referral for abnormal/positive results. Phase 2 (June 2018-March 2019) further integrated blood glucose/cholesterol/chlamydia RDT, with optional peer-navigated referral. Enrolled referred clients completed telephonic follow-up surveys confirming LTC/treatment initiation ≤3 months post-screening. Socio-demographics, screening results, time to LTC/treatment initiation, peer-navigated referral uptake were reported. Analysis included Fisher's exact, chi-squared, Kruskal Wallis, and Student's T-tests. Thematic analysis was conducted for open-ended survey responses., Results: Of all 320 referrals, 40.0% were HIV-infections, 11.9% STIs, 6.6% TB, and 28.8% high/low BP. Of Phase 2-only referrals, 29.4% were for glucose and 23.5% cholesterol. Integrated NCD-HTS had significantly more clients LTC for HIV (76.7%[n = 66/86] vs 52.4%[n = 22/42], p = 0.0052) and within a shorter average time (6-8 days [Interquartile range (IQR):1-18.5] vs 8-13 days [IQR:2-32]) as compared to SOC HTS. Integrated NCD-HTS clients initiated HIV/STIs/BP treatment on average more quickly as compared to SOC HTS (5 days for STIs [IQR:1-21], 8 days for HIV/BP [IQR:5-17 and 2-13, respectively] vs 10 days for STIs [IQR: 4-32], 19.5 days for HIV [IQR:6.5-26.5], 8 days for BP [IQR:2-29)]. Participants chose passive over active referral (89.1% vs 10.9%; p<0.0001). Participants rejecting peer-navigated referral preferred to go alone (55.7% [n = 39/70]). Non-LTC was due to being busy (41.1% [n = 39/95]) and not being ready/refusing treatment (31.6% [n = 30/95]). Normalised results assessed at referral clinic (49.7% [n = 98/196]), prescribed lifestyle modification/monitoring (30.9% [n = 61/196]), and poor clinic flow/congestion and/or further testing required (10.7% [n = 21/196]) were associated with non-treatment initiation., Conclusion: Same-day treatment initiation is not achieved across diseases, despite peer-navigated referral. There are psychosocial and health systems barriers at entry to care/treatment initiation. Additional research may identify best strategies for rapid treatment initiation., Competing Interests: The authors have declared that no competing interests exist.

Background: In response to increasing numbers of carbapenemase-producing Enterobacterales (CPE) in England, Public Health England (PHE) launched an electronic reporting system (ERS) for the enhanced surveillance of carbapenemase-producing Gram-negative bacteria. Our study aimed to describe system engagement and the epidemiology of CPE in England., Methods: Engagement with the ERS was assessed by calculating the proportion of referrals submitted this system. ERS data were extracted and cases defined as patients with CPE isolated from a screening or clinical specimen in England between 1 st May 2015 to 31 st March 2019. Descriptive summary statistics for each variable were prepared., Results: The ERS processed 12,656 suspected CPE reports. Uptake of the ERS by local microbiology laboratories varied, with approximately 70% of referrals made via the ERS by April 2016; this steadily decreased after March 2018. Six-thousand eight-hundred and fifty-seven cases were included in the analysis. Most cases were from colonised patients (80.6%) rather than infected, and the majority were inpatients in acute hospital settings (87.3%). Carbapenemases were most frequently detected in Klebsiella pneumoniae (39.1%) and Escherichia coli (30.3%). The most frequently identified carbapenemase families were OXA-48-like (45.1%) and KPC (26.4%). Enhanced data variables were poorly completed., Conclusions: The ERS has provided some insight into the epidemiology of CPE in England. An increasing number of routine diagnostic laboratories have introduced methods to routinely identify acquired carbapenemases and PHE has modified its approach to ensure robust surveillance, which is an essential aspect of an effective response to prevent and control the spread of CPE., (Crown Copyright © 2020 Published by Elsevier Ltd on behalf of The Healthcare Infection Society.)

We report two cases of respiratory toxigenic Corynebacterium diphtheriae infection in fully vaccinated UK born adults following travel to Tunisia in October 2019. Both patients were successfully treated with antibiotics and neither received diphtheria antitoxin. Contact tracing was performed following a risk assessment but no additional cases were identified. This report highlights the importance of maintaining a high index of suspicion for re-emerging infections in patients with a history of travel to high-risk areas outside Europe.

Background: While HIV Testing Services (HTS) have increased, many South Africans have not been tested. Non-communicable diseases (NCDs) are the top cause of death worldwide. Integrated NCD-HTS could be a strategy to control both epidemics. Healthcare service strategies depends partially on positive user experience. We investigated client satisfaction of services and clinic flow time of an integrated NCD-HTS clinic., Methods: This prospective, cross-sectional study evaluated HTS client satisfaction with an HTS clinic at two phases. Phase 1 (February-June 2018) utilised standard HTS services: counsellor-led height/weight/blood pressure measurements, HIV rapid testing, and symptoms screening for sexually transmitted infections/Tuberculosis. Phase 2 (June 2018-March 2019) further integrated counsellor-led obesity screening (body mass index/abdominal circumference measurements), rapid cholesterol/glucose testing; and nurse-led Chlamydia and human papilloma virus (HPV)/cervical cancer screening. Socio-demographics, proportion of repeat clients, clinic flow time, and client survey data (open/closed-ended questions using five-point Likert scale) are reported. Fisher's exact test, chi-square analysis, and Kruskal Wallis test conducted comparisons. Multiple linear regression determined predictors associated with clinic time. Content thematic analysis was conducted for free response data., Results: Two hundred eighty-four and three hundred thirty-three participants were from Phase 1 and 2, respectively (N = 617). Phase 1 participants were significantly older (median age 36.5 (28.0-43.0) years vs. 31.0 (25.0-40.0) years; p = 0.0003), divorced/widowed (6.7%, [n = 19/282] vs. 2.4%, [n = 8/332]; p = 0.0091); had tertiary education (27.9%, [n = 79/283] vs. 20.1%, [n = 67/333]; p = 0.0234); and less female (53.9%, [n = 153/284] vs 67.6%, [n = 225/333]; p = 0.0005), compared to Phase 2. Phase 2 had 10.2% repeat clients (n = 34/333), and 97.9% (n = 320/327) were 'very satisfied' with integrated NCD-HTS, despite standard HTS having significantly shorter median time for counsellor-led HTS (36.5, interquartile range [IQR]: 31.0-45.0 vs. 41.5, IQR: 35.0-51.0; p < 0.0001). Phase 2 associations with longer clinic time were clients living together/married (est = 6.548; p = 0.0467), more tests conducted (est = 3.922; p < 0.0001), higher overall satisfaction score (est = 1.210; p = 0.0201). Those who matriculated experienced less clinic time (est = - 7.250; p = 0.0253)., Conclusions: It is possible to integrate counsellor-led NCD rapid testing into standard HTS within historical HTS timeframes, yielding client satisfaction. Rapid cholesterol/glucose testing should be integrated into standard HTS. Research is required on the impact of cervical cancer/HPV screenings to HTS clinic flow to determine if it could be scaled up within the public sector.

Analysis of sequencing data for 143 bla NDM-1 - and bla OXA-48 -positive Klebsiella pneumoniae isolates from 13 European national collections and the public domain resulted in the identification of 15 previously undetected multi-country transmission clusters. For 10 clusters, cases had prior travel/hospitalisation history in countries outside of the European Union including Egypt, Iran, Morocco, Russia, Serbia, Tunisia and Turkey. These findings highlight the benefit of European whole genome sequencing-based surveillance and data sharing for control of antimicrobial resistance.