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Interpreting the phenotypes of bla SHV alleles in Klebsiella pneumoniae genomes is complex. Whilst all strains are expected to carry a chromosomal copy conferring resistance to ampicillin, they may also carry mutations in chromosomal bla SHV alleles or additional plasmid-borne bla SHV alleles that have extended-spectrum β-lactamase (ESBL) activity and/or β-lactamase inhibitor (BLI) resistance activity. In addition, the role of individual mutations/a changes is not completely documented or understood. This has led to confusion in the literature and in antimicrobial resistance (AMR) gene databases [e.g. the National Center for Biotechnology Information (NCBI) Reference Gene Catalog and the β-lactamase database (BLDB)] over the specific functionality of individual sulfhydryl variable (SHV) protein variants. Therefore, the identification of ESBL-producing strains from K. pneumoniae genome data is complicated. Here, we reviewed the experimental evidence for the expansion of SHV enzyme function associated with specific aa substitutions. We then systematically assigned SHV alleles to functional classes (WT, ESBL and BLI resistant) based on the presence of these mutations. This resulted in the re-classification of 37 SHV alleles compared with the current assignments in the NCBI's Reference Gene Catalog and/or BLDB (21 to WT, 12 to ESBL and 4 to BLI resistant). Phylogenetic and comparative genomic analyses support that (i) SHV-1 (encoded by bla SHV-1 ) is the ancestral chromosomal variant, (ii) ESBL- and BLI-resistant variants have evolved multiple times through parallel substitution mutations, (iii) ESBL variants are mostly mobilized to plasmids and (iv) BLI-resistant variants mostly result from mutations in chromosomal bla SHV . We used matched genome-phenotype data from the KlebNET-GSP AMR Genotype-Phenotype Group to identify 3999 K . pneumoniae isolates carrying one or more bla SHV alleles but no other acquired β-lactamases to assess genotype-phenotype relationships for bla SHV . This collection includes human, animal and environmental isolates collected between 2001 and 2021 from 24 countries. Our analysis supports that mutations at Ambler sites 238 and 179 confer ESBL activity, whilst most omega-loop substitutions do not. Our data also provide support for the WT assignment of 67 protein variants, including 8 that were noted in public databases as ESBL. These eight variants were reclassified as WT because they lack ESBL-associated mutations, and our phenotype data support susceptibility to third-generation cephalosporins (SHV-27, SHV-38, SHV-40, SHV-41, SHV-42, SHV-65, SHV-164 and SHV-187). The approach and results outlined here have been implemented in Kleborate v2.4.1 (a software tool for genotyping K. pneumoniae ), whereby known and novel bla SHV alleles are classified based on causative mutations. Kleborate v2.4.1 was updated to include ten novel protein variants from the KlebNET-GSP dataset and all alleles in public databases as of November 2023. This study demonstrates the power of sharing AMR phenotypes alongside genome data to improve the understanding of resistance mechanisms.

Background: Patients with prolonged hospitalisation have a significant risk of carriage of and subsequent infection with extended spectrum β-lactamase (ESBL)-producing and carbapenemase-producing Klebsiella pneumoniae. However, the distinctive roles of the community and hospital environments in the transmission of ESBL-producing or carbapenemase-producing K pneumoniae remain elusive. We aimed to investigate the prevalence and transmission of K pneumoniae within and between the two tertiary hospitals in Hanoi, Viet Nam, using whole-genome sequencing., Methods: We did a prospective cohort study of 69 patients in intensive care units (ICUs) from two hospitals in Hanoi, Viet Nam. Patients were included if they were aged 18 years or older, admitted for longer than the mean length of stay in their ICU, and cultured K pneumoniae from their clinical samples. Longitudinally collected samples from patients (collected weekly) and the ICU environment (collected monthly) were cultured on selective media, and whole-genome sequences from K pneumoniae colonies analysed. We did phylogenetic analyses and correlated phenotypic antimicrobial susceptibility testing with genotypic features of K pneumoniae isolates. We constructed transmission networks of patient samples, relating ICU admission times and locations with genetic similarity of infecting K pneumoniae., Findings: Between June 1, 2017, and Jan 31, 2018, 69 patients were in the ICUs and eligible for inclusion, and a total of 357 K pneumoniae isolates were cultured and successfully sequenced. 228 (64%) of K pneumoniae isolates carried between two and four different ESBL-encoding and carbapenemase-encoding genes, with 164 (46%) isolates carrying genes encoding both, with high minimum inhibitory concentrations. We found a novel co-occurrence of bla KPC-2 and bla NDM-1 in 46·6% of samples from the globally successful ST15 lineage. Despite being physically and clinically separated, the two hospitals shared closely related strains carrying the same array of antimicrobial resistance genes., Interpretation: These results highlight the high prevalence of ESBL-positive carbapenem-resistant K pneumoniae in ICUs in Viet Nam. Through studying K pneumoniae ST15 in detail, we showed how important resistance genes are contained within these strains that are carried broadly by patients entering the two hospitals directly or through referral., Funding: Medical Research Council Newton Fund, Ministry of Science and Technology, Wellcome Trust, Academy of Medical Sciences, Health Foundation, and National Institute for Health and Care Research Cambridge Biomedical Research Centre., Competing Interests: Declaration of interests JP is a paid consultant for Next Gen Diagnostics. 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.)

Background: Viet Nam has high rates of antimicrobial resistance (AMR) but little capacity for genomic surveillance. This study used whole genome sequencing to examine the prevalence and transmission of three key AMR pathogens in two intensive care units (ICUs) in Hanoi, Viet Nam., Methods: A prospective surveillance study of all adults admitted to ICUs at the National Hospital for Tropical Diseases and Bach Mai Hospital was done between June 19, 2017, and Jan 16, 2018. Clinical and environmental samples were cultured on selective media, characterised with MALDI TOF mass spectrometry, and sequenced with Illumina. Phylogenies based on the de-novo assemblies (SPAdes) were constructed with MAFFT (PARsnp), Gubbins, and RAxML. Resistance genes were detected with Abricate against the US National Center for Biotechnology Information database., Findings: 3153 Escherichia coli, Klebsiella pneumoniae, and Acinetobacter baumannii isolates from 369 patients were analysed. Phylogenetic analysis revealed predominant lineages within A baumannii (global clone 2, sequence types ST2 and ST571) and K pneumoniae (ST15, ST16, ST656, ST11, and ST147) isolates. Isolation from stool was most common with E coli (87·0%) followed by K pneumoniae (62·5%). Of the E coli, 85·0% carried a bla CTX-M variant, while 81·8% of K pneumoniae isolates carried bla NDM (54·4%), or bla KPC (45·1%), or both. Transmission analysis with single nucleotide polymorphisms identified 167 clusters involving 251 (68%) of 369 patients, in some cases involving patients from both ICUs. There were no clear differences between the lineages or AMR genes recovered between the two ICUs., Interpretation: This study represents the largest prospective surveillance study of key AMR pathogens in Vietnamese ICUs. Clusters of closely related isolates in patients across both ICUs suggests recent transmission before ICU admission in other health-care settings or in the community., Funding: UK Medical Research Council Newton Fund, Viet Nam Ministry of Science and Technology, Wellcome Trust, Academy of Medical Sciences, Health Foundation, and UK National Institute for Health and Care Research Cambridge Biomedical Research Centre., Competing Interests: Declaration of interests LWR reports travel fees from European Molecular Biology Laboratory, European Bioinformatics Institute (EMBL-EBI), and is on the Microbial Genomics Early Career Microbiologists board of reviewers, outside of the submitted work. HRvD is a board member of the Surveillance and Epidemiology of Drug Resistant Infections Consortium, outside of the submitted work. JP reports grants from the National Institute for Health and Care Research and Wellcome Trust and consulting fees and stock options from Next Gen Diagnostics, outside of the submitted work. ZI reports grants from Global Challenges Research Fund, UK Research and Innovation, and National Institute for Health and Care Research Health Protection Research Unit, travel fees from the US Centers for Disease Control and Prevention, and is part of the advisory board for Laboratory of Molecular Infection Medicine Sweden and CLIMB-BIG-DATA, outside of the submitted work. MET reports book royalties from Oxford University Press and teaching honoraria from Wellcome Sanger Institute, outside of the submitted work. All other authors declare no competing interests., (Copyright © 2022 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.)

The significant bottlenecks in determining bacterial species are much more time-consuming and the biology specialist's long-term experience requirements. Specifically, it takes more than half a day to cultivate a bacterium, and then a skilled microbiologist and a costly specialized machine are utilized to analyze the genes and classify the bacterium according to its nucleotide sequence. To overcome these issues as well as get higher recognition accuracy, we proposed applying convolutional neural networks (CNNs) architectures to automatically classify bacterial species based on some key characteristics of bacterial colonies. Our experiment confirmed that the classification of three bacterial colonies could be performed with the highest accuracy (97.19%) using a training set of 5000 augmented images derived from the 40 original photos taken in the Hanoi Medical University laboratory in Vietnam.

In Vietnam, the public health burden of rickettsial infections continues to be underestimated due to knowledge gaps in the epidemiology of these diseases. We conducted a systematic study among 27 hospitals from 26 provinces in eight ecological regions throughout Vietnam to investigate the prevalence, distribution, and clinical characteristics of rickettsial diseases. We recruited 1834 patients in the study from April 2018 to October 2019. The findings showed that rickettsial diseases were common among undifferentiated febrile patients, with 564 (30.8%) patients positive by qPCR for scrub typhus, murine typhus or spotted fever. Scrub typhus (484, 85.8%) was the most common rickettsial disease, followed by murine typhus (67, 11.9%) and spotted fever (10, 1.8%). Rickettsial diseases were widely distributed in all regions of Vietnam and presented with nonspecific clinical manifestations.

Scrub typhus has been documented since 1932 in Vietnam, however, the disease burden of scrub typhus remains poorly understood in the country. We conducted this study to describe the phylogenetic analysis of the 56-kDa type-specific antigen (TSA) gene of Orientia tsutsugamushi associated with PCR positive cases of scrub typhus. Of 116 positive samples, 65 type-specific antigen gene sequences were obtained and classified into 3 genogroups: Karp, Kato and Gilliam. The Karp genogroup was the most frequently detected phylogenetic cluster in the study with 30 samples (46%), followed by Kato and Gilliam with 20 (31%) and 15 (23%), respectively. All sequences showed 94-100% nucleotide similarity to reference sequences collected in the central part of Vietnam in 2017. Patients infected with Karp genogroup were more likely to have significant thrombocytopenia than the other genogroups. These results suggest that any scrub typhus vaccine considered for use in Vietnam should provide protection against each of these 3 genogroups., Competing Interests: The authors have declared that no competing interests exist.

Rickettsioses are endemic to Vietnam; however, only a limited number of clinical studies have been performed on these vectorborne bacteria. We conducted a prospective hospital-based study at 2 national referral hospitals in Hanoi to describe the clinical characteristics of scrub typhus and murine typhus in northern Vietnam and to assess the diagnostic applicability of quantitative real-time PCR assays to diagnose rickettsial diseases. We enrolled 302 patients with acute undifferentiated fever and clinically suspected rickettsiosis during March 2015-March 2017. We used a standardized case report form to collect clinical information and laboratory results at the time of admission and during treatment. We confirmed scrub typhus in 103 (34.1%) patients and murine typhus in 12 (3.3%) patients. These results highlight the need for increased emphasis on training for healthcare providers for earlier recognition, prevention, and treatment of rickettsial diseases in Vietnam.

AbstractRickettsial infections are recognized as important causes of fever throughout southeast Asia. Herein, we determined the seroprevalence to rickettsioses within rural and urban populations of northern Vietnam. Prevalence of individuals with evidence of prior rickettsial infections (IgG positive) was surprisingly low, with 9.14% (83/908) testing positive to the three major rickettsial serogroups thought to circulate in the region. Prevalence of typhus group rickettsiae (TG)-specific antibodies (6.5%, 58/908) was significantly greater than scrub typhus group orientiae (STG)- or spotted fever group rickettsiae (SFG)-specific antibodies ( P < 0.05). The majority of TG seropositives were observed among urban rather than rural residents ( P < 0.05). In contrast, overall antibody prevalence to STG and SFG were both very low (1.1%, 10/908 for STG; 1.7%, 15/908 for SFG), with no significant differences between rural and urban residents. These results provide data on baseline population characteristics that may help inform development of Rickettsia serological testing criteria in future clinical studies.