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Showing papers on "Typing published in 2022"


Journal ArticleDOI
TL;DR: The current status of the SCCmec types, S CCmec subtypes, rules for nomenclature, and multiple methods for identifying SCCMec types and subtypes were summarized in this review.
Abstract: Staphylococcal cassette chromosome mec (SCCmec) typing was established in the 2000s and has been employed as a tool for the molecular epidemiology of methicillin-resistant Staphylococcus aureus, as well as the evolution investigation of Staphylococcus species. Molecular cloning and the conventional sequencing of SCCmec have been adopted to verify the presence and structure of a novel SCCmec type, while convenient PCR-based SCCmec identification methods have been used in practical settings for many years. In addition, whole-genome sequencing has been widely used, and various SCCmec and similar structures have been recently identified in various species. The current status of the SCCmec types, SCCmec subtypes, rules for nomenclature, and multiple methods for identifying SCCmec types and subtypes were summarized in this review, according to the perspective of the International Working Group on the Classification of Staphylococcal Cassette Chromosome Elements.

36 citations


Journal ArticleDOI
TL;DR: Kaptive 2.0 was applied to a curated dataset of >12 000 public KpSC genomes to explore for the first time, to the best of the authors' knowledge, the distribution of predicted O (sub)types across species, sampling niches and clones, which highlighted key differences in the distributions that warrant further investigation.
Abstract: The outer polysaccharide capsule and lipopolysaccharide (LPS) antigens are key targets for novel control strategies targeting Klebsiella pneumoniae and related taxa from the K. pneumoniae species complex (KpSC), including vaccines, phage and monoclonal antibody therapies. Given the importance and growing interest in these highly diverse surface antigens, we had previously developed Kaptive, a tool for rapidly identifying and typing capsule (K) and outer LPS (O) loci from whole genome sequence data. Here, we report two significant updates, now freely available in Kaptive 2.0 (https://github.com/katholt/kaptive): (i) the addition of 16 novel K locus sequences to the K locus reference database following an extensive search of >17 000 KpSC genomes; and (ii) enhanced O locus typing to enable prediction of the clinically relevant O2 antigen (sub)types, for which the genetic determinants have been recently described. We applied Kaptive 2.0 to a curated dataset of >12 000 public KpSC genomes to explore for the first time, to the best of our knowledge, the distribution of predicted O (sub)types across species, sampling niches and clones, which highlighted key differences in the distributions that warrant further investigation. As the uptake of genomic surveillance approaches continues to expand globally, the application of Kaptive 2.0 will generate novel insights essential for the design of effective KpSC control strategies.

35 citations


Journal ArticleDOI
TL;DR: In this article , the European gonococcal population in 2018 and identify emerging or vanishing lineages associated with antimicrobial resistance and epidemiological characteristics of patients, to elucidate recent changes in AMR and gonorrhoea epidemiology in Europe.
Abstract: Genomic surveillance using quality-assured whole-genome sequencing (WGS) together with epidemiological and antimicrobial resistance (AMR) data is essential to characterise the circulating Neisseria gonorrhoeae lineages and their association to patient groups (defined by demographic and epidemiological factors). In 2013, the European gonococcal population was characterised genomically for the first time. We describe the European gonococcal population in 2018 and identify emerging or vanishing lineages associated with AMR and epidemiological characteristics of patients, to elucidate recent changes in AMR and gonorrhoea epidemiology in Europe.We did WGS on 2375 gonococcal isolates from 2018 (mainly Sept 1-Nov 30) in 26 EU and EEA countries. Molecular typing and AMR determinants were extracted from quality-checked genomic data. Association analyses identified links between genomic lineages, AMR, and epidemiological data.Azithromycin-resistant N gonorrhoeae (8·0% [191/2375] in 2018) is rising in Europe due to the introduction or emergence and subsequent expansion of a novel N gonorrhoeae multi-antigen sequence typing (NG-MAST) genogroup, G12302 (132 [5·6%] of 2375; N gonorrhoeae sequence typing for antimicrobial resistance [NG-STAR] clonal complex [CC]168/63), carrying a mosaic mtrR promoter and mtrD sequence and found in 24 countries in 2018. CC63 was associated with pharyngeal infections in men who have sex with men. Susceptibility to ceftriaxone and cefixime is increasing, as the resistance-associated lineage, NG-MAST G1407 (51 [2·1%] of 2375), is progressively vanishing since 2009-10.Enhanced gonococcal AMR surveillance is imperative worldwide. WGS, linked to epidemiological and AMR data, is essential to elucidate the dynamics in gonorrhoea epidemiology and gonococcal populations as well as to predict AMR. When feasible, WGS should supplement the national and international AMR surveillance programmes to elucidate AMR changes over time. In the EU and EEA, increasing low-level azithromycin resistance could threaten the recommended ceftriaxone-azithromycin dual therapy, and an evidence-based clinical azithromycin resistance breakpoint is needed. Nevertheless, increasing ceftriaxone susceptibility, declining cefixime resistance, and absence of known resistance mutations for new treatments (zoliflodacin, gepotidacin) are promising.European Centre for Disease Prevention and Control, Centre for Genomic Pathogen Surveillance, Örebro University Hospital, Wellcome.

24 citations


Journal ArticleDOI
TL;DR: Standardized, high-end typing tools and integrative strategies are needed to fill the gaps and complete an accurate image of the T. gondii genetic population in Europe.
Abstract: Toxoplasma gondii is a major zoonotic agent which may cause harmful effects mainly in pregnant and immunocompromised hosts. Despite many efforts on its genetic characterization, an entirely clear picture of the population structure in Europe has not been achieved yet. The present study aimed to summarize the available genotyping information and to map the distribution of circulating strains. There is consensus on type II T. gondii genotypes prevailing in Europe, but the absence of harmonization in the use of typing methods limits detailed knowledge. Standardized, high-end typing tools and integrative strategies are needed to fill the gaps and complete an accurate image of the T. gondii genetic population in Europe.

22 citations


Journal ArticleDOI
TL;DR: The potential origin of the genetic diversity of T. cruzi is presented, genomic aspects of DTUs focusing on polymorphisms in multigene families encoding surface antigens that play essential functions for parasite survival both in the insect vector and the mammalian host are described.
Abstract: Trypanosoma cruzi, the agent of Chagas disease (ChD), exhibits remarkable biological and genetic diversity, along with eco-epidemiological complexity. In order to facilitate communication among researchers aiming at the characterisation of biological and epidemiological aspects of T. cruzi, parasite isolates and strains were partitioned into seven discrete typing units (DTUs), TcI-TcVI and TcBat, identifiable by reproducible genotyping protocols. Here we present the potential origin of the genetic diversity of T. cruzi and summarise knowledge about eco-epidemiological associations of DTUs with mammalian reservoirs and vectors. Circumstantial evidence of a connection between T. cruzi genotype and ChD manifestations is also discussed emphasising the role of the host’s immune response in clinical ChD progression. We describe genomic aspects of DTUs focusing on polymorphisms in multigene families encoding surface antigens that play essential functions for parasite survival both in the insect vector and the mammalian host. Such antigens most probably contributed to the parasite success in establishing infections in different hosts and exploring several niches. Gaps in the current knowledge and challenges for future research are pointed out.

18 citations


Journal ArticleDOI
TL;DR: K1 remains the dominant hvKP serotype and is associated with most virulence genes in Taiwan, and further studies are required to elucidate the significance of other virulent serotypes.
Abstract: We conducted a longitudinal epidemiological surveillance of hypervirulent Klebsiella pneumoniae (hvKP) in Taiwan. Bacteremic KP isolates collected from 16 hospitals in Taiwan between 2017 and 2019 were collected, and the virulent serotypes (K1, K2, K20, K54, and K57), antimicrobial susceptibilities, and virulence genes of these isolates were investigated. During the 3-year period, 1,310 bacteremic KP isolates were collected, of which 27.5% belonged to virulent serotypes, including K1 (n = 162), K2 (n = 74), K57 (n = 56), K54 (n = 41), and K20 (n = 27). K1 was the most prevalent capsular serotype, with an annual prevalence of 11–15%, and was equally distributed across the four geographic areas. The prevalence of K2 declined significantly in 2019. According to wzi-K typing results, 87% of K1 isolates were classified as wzi-1. Among K2 isolates, wzi-72 (55.4%) and wzi-2 (41.9%) were the most common, whereas wzi-206 was the most prevalent (48.2%) among K57 isolates, followed by wzi-77 (25.0%). Wzi-115 accounted for 85.4% of the K54 isolates, whereas wzi-95 accounted for 92.6% of K20 isolates. rmpA was present in 99.4% of K1, 98.6% of K2, 89.3% of K57, 78.0% of K54, and 84.0% of K20 isolates. rmpA2 was present in 100% of K1 and 98.6% of K2 isolates but was only present in 64.3% of K57, 58.5% of K54, and 74.1% of K20 isolates. K1 remains the dominant hvKP serotype and is associated with most virulence genes in Taiwan. Further studies are required to elucidate the significance of other virulent serotypes.

16 citations


Journal ArticleDOI
TL;DR: In this article , the authors used the standardized EnteroBase core-genome multilocus sequence typing scheme (cgMLST) to classify Shigella strains into four serogroups and more than 50 serotypes.
Abstract: The laboratory surveillance of bacillary dysentery is based on a standardised Shigella typing scheme that classifies Shigella strains into four serogroups and more than 50 serotypes on the basis of biochemical tests and lipopolysaccharide O-antigen serotyping. Real-time genomic surveillance of Shigella infections has been implemented in several countries, but without the use of a standardised typing scheme. Here, we study over 4000 reference strains and clinical isolates of Shigella, covering all serotypes, with both the current serotyping scheme and the standardised EnteroBase core-genome multilocus sequence typing scheme (cgMLST). The Shigella genomes are grouped into eight phylogenetically distinct clusters, within the E. coli species. The cgMLST hierarchical clustering (HC) analysis at different levels of resolution (HC2000 to HC400) recognises the natural population structure of Shigella. By contrast, the serotyping scheme is affected by horizontal gene transfer, leading to a conflation of genetically unrelated Shigella strains and a separation of genetically related strains. The use of this cgMLST scheme will facilitate the transition from traditional phenotypic typing to routine whole-genome sequencing for the laboratory surveillance of Shigella infections.

15 citations


Journal ArticleDOI
TL;DR: In this paper , the authors used the standardized EnteroBase core-genome multilocus sequence typing scheme (cgMLST) to classify Shigella strains into four serogroups and more than 50 serotypes.
Abstract: The laboratory surveillance of bacillary dysentery is based on a standardised Shigella typing scheme that classifies Shigella strains into four serogroups and more than 50 serotypes on the basis of biochemical tests and lipopolysaccharide O-antigen serotyping. Real-time genomic surveillance of Shigella infections has been implemented in several countries, but without the use of a standardised typing scheme. Here, we study over 4000 reference strains and clinical isolates of Shigella, covering all serotypes, with both the current serotyping scheme and the standardised EnteroBase core-genome multilocus sequence typing scheme (cgMLST). The Shigella genomes are grouped into eight phylogenetically distinct clusters, within the E. coli species. The cgMLST hierarchical clustering (HC) analysis at different levels of resolution (HC2000 to HC400) recognises the natural population structure of Shigella. By contrast, the serotyping scheme is affected by horizontal gene transfer, leading to a conflation of genetically unrelated Shigella strains and a separation of genetically related strains. The use of this cgMLST scheme will facilitate the transition from traditional phenotypic typing to routine whole-genome sequencing for the laboratory surveillance of Shigella infections.

14 citations


Journal ArticleDOI
TL;DR: In this article , the authors used phylogenomic and phylodynamic analyses to characterise the lineages of T pallidum driving the current syphilis epidemic in Australia and found that the current epidemic is driven by multiple lineages, rather than one distinct outbreak.
Abstract: BackgroundThe incidence of syphilis has increased markedly in the past decade in high-income countries, including Australia. To date, however, genomic studies of Treponema pallidum have focused mainly on the northern hemisphere. Here, we aimed to characterise the lineages of T pallidum driving the current syphilis epidemic in Australia.MethodsIn this genomic epidemiological analysis, using phylogenomic and phylodynamic analyses, we analysed 456 high-quality T pallidum genomes collected from clinical samples in Australia between Oct 19, 2005, and Dec 31, 2020, and contextualised this information with publicly available sequence data. We also performed detailed genomic characterisation of putative antimicrobial resistance determinants, in addition to correlating single-locus typing of the TP0548 allele with the T pallidum phylogeny.FindingsPhylogenomic analyses identified four major sublineages circulating in Australia and globally, two belonging to the SS14 lineage, and two belonging to the Nichols lineage. Australian sublineages were further delineated into twelve subgroups, with five of the six largest subgroups associated with men who have sex with men, and the sixth lineage was predominantly associated with heterosexual people. Most Australian T pallidum genomes (398 [87%] of 456) were genotypically macrolide resistant, and TP0548 typing correlated significantly with T pallidum genomic subgroups.InterpretationThese findings show that the current syphilis epidemic in Australia is driven by multiple lineages of T pallidum, rather than one distinct outbreak. Major subgroups of T pallidum in Australia have emerged within the past 30 years, are closely related to global lineages, and circulate across different sexual networks. In conjunction with improved testing and treatment, these data could better inform the control of syphilis in Australia.FundingNational Health and Medical Research Council, Australian Research Council. The incidence of syphilis has increased markedly in the past decade in high-income countries, including Australia. To date, however, genomic studies of Treponema pallidum have focused mainly on the northern hemisphere. Here, we aimed to characterise the lineages of T pallidum driving the current syphilis epidemic in Australia. In this genomic epidemiological analysis, using phylogenomic and phylodynamic analyses, we analysed 456 high-quality T pallidum genomes collected from clinical samples in Australia between Oct 19, 2005, and Dec 31, 2020, and contextualised this information with publicly available sequence data. We also performed detailed genomic characterisation of putative antimicrobial resistance determinants, in addition to correlating single-locus typing of the TP0548 allele with the T pallidum phylogeny. Phylogenomic analyses identified four major sublineages circulating in Australia and globally, two belonging to the SS14 lineage, and two belonging to the Nichols lineage. Australian sublineages were further delineated into twelve subgroups, with five of the six largest subgroups associated with men who have sex with men, and the sixth lineage was predominantly associated with heterosexual people. Most Australian T pallidum genomes (398 [87%] of 456) were genotypically macrolide resistant, and TP0548 typing correlated significantly with T pallidum genomic subgroups. These findings show that the current syphilis epidemic in Australia is driven by multiple lineages of T pallidum, rather than one distinct outbreak. Major subgroups of T pallidum in Australia have emerged within the past 30 years, are closely related to global lineages, and circulate across different sexual networks. In conjunction with improved testing and treatment, these data could better inform the control of syphilis in Australia.

13 citations


Journal ArticleDOI
TL;DR: This review mainly presents a clinical overview and perspective of emerging technologies in blood group testing based on the literature to highlight the most promising strategies and promote blood group typing development to ensure blood transfusion safety.
Abstract: Red blood cell (RBC) transfusion is one of the most frequently performed clinical procedures and therapies to improve tissue oxygen delivery in hospitalized patients worldwide. Generally, the cross-match is the mandatory test in place to meet the clinical needs of RBC transfusion by examining donor-recipient compatibility with antigens and antibodies of blood groups. Blood groups are usually an individual's combination of antigens on the surface of RBCs, typically of the ABO blood group system and the RH blood group system. Accurate and reliable blood group typing is critical before blood transfusion. Serological testing is the routine method for blood group typing based on hemagglutination reactions with RBC antigens against specific antibodies. Nevertheless, emerging technologies for blood group testing may be alternative and supplemental approaches when serological methods cannot determine blood groups. Moreover, some new technologies, such as the evolving applications of blood group genotyping, can precisely identify variant antigens for clinical significance. Therefore, this review mainly presents a clinical overview and perspective of emerging technologies in blood group testing based on the literature. Collectively, this may highlight the most promising strategies and promote blood group typing development to ensure blood transfusion safety.

12 citations


Journal ArticleDOI
TL;DR: The strain responsible for the first recognized outbreak by an NDM-1-producing K. pneumoniae in Portugal is characterized, and the use of FT-IR as an asset to support quick infection control is further supported.
Abstract: The changing epidemiology of carbapenem-resistant Klebsiella pneumoniae in Southern European countries is challenging for infection control, and it is critical to identify and track new genetic entities (genes, carbapenemases, clones) quickly and with high precision. We aimed to characterize the strain responsible for the first recognized outbreak by an NDM-1-producing K. pneumoniae in Portugal, and to elucidate its diffusion in an international context. NDM-1-producing multidrug-resistant K. pneumoniae isolates from hospitalized patients (2018–2019) were characterized using FTIR spectroscopy, molecular typing, whole-genome sequencing, and comparative genomics with available K. pneumoniae ST11 KL105 genomes. FT-IR spectroscopy allowed the rapid (ca. 4 h after incubation) identification of the outbreak strains as ST11 KL105, supporting outbreak control. Epidemiological information supports a community source but without linkage to endemic regions of NDM-1 producers. Whole-genome comparison with previous DHA-1-producing ST11 KL105 strains revealed the presence of different plasmid types and antibiotic resistance traits, suggesting the entry of a new strain. In fact, this ST11 KL105 clade has successfully disseminated in Europe with variable beta-lactamases, but essentially as ESBL or DHA-1 producers. We expand the distribution map of NDM-1-producing K. pneumoniae in Europe, at the expense of a successfully established ST11 KL105 K. pneumoniae clade circulating with variable plasmid backgrounds and beta-lactamases. Our work further supports the use of FT-IR as an asset to support quick infection control.

Journal ArticleDOI
TL;DR: In this article , Capillary electrophoresis (CE)-PCR ribotyping is used for C. difficile typing but lacks the discriminatory power to study transmission and outbreaks in detail.
Abstract: Clostridioides difficile is the most common cause of antibiotic-associated gastrointestinal infections. Capillary electrophoresis (CE)-PCR ribotyping is currently the gold standard for C. difficile typing but lacks the discriminatory power to study transmission and outbreaks in detail.

Journal ArticleDOI
TL;DR: Interdisciplinary efforts including WGS and epidemiological investigations were essential to identifying the source of infection and susceptible individuals should be better informed of the risk of acquiring listeriosis from consuming smoked and graved salmon products.
Abstract: ABSTRACT Invasive listeriosis, caused by Listeria (L.) monocytogenes, is a severe foodborne infection, especially for immunocompromised individuals. The aim of our investigation was the identification and analysis of listeriosis outbreaks in Germany with smoked and graved salmon products as the most likely source of infection using whole-genome sequencing (WGS) and patient interviews. In a national surveillance programme, WGS was used for subtyping and core genome multi locus sequence typing (cgMLST) for cluster detection of L. monocytogenes isolates from listeriosis cases as well as food and environmental samples in Germany. Patient interviews were conducted to complement the molecular typing. We identified 22 independent listeriosis outbreaks occurring between 2010 and 2021 that were most likely associated with the consumption of smoked and graved salmon products. In Germany, 228 cases were identified, of 50 deaths (22%) reported 17 were confirmed to have died from listeriosis. Many of these 22 outbreaks were cross-border outbreaks with further cases in other countries. This report shows that smoked and graved salmon products contaminated with L. monocytogenes pose a serious risk for listeriosis infection in Germany. Interdisciplinary efforts including WGS and epidemiological investigations were essential to identifying the source of infection. Uncooked salmon products are high-risk foods frequently contaminated with L. monocytogenes. In order to minimize the risk of infection for consumers, food producers need to improve hygiene measures and reduce the entry of pathogens into food processing. Furthermore, susceptible individuals should be better informed of the risk of acquiring listeriosis from consuming smoked and graved salmon products.

Journal ArticleDOI
TL;DR: A novel Brucella-wide cgMLST scheme is described that helps in accurately tracking and thus controlling the sources of infection and should represent a valuable resource for laboratories with limited computational resources and bioinformatics expertise.
Abstract: Brucellosis poses a significant burden to human and animal health worldwide. Robust and harmonized molecular epidemiological approaches and population studies that include routine disease screening are needed to efficiently track the origin and spread of Brucella strains. ABSTRACT Brucellosis poses a significant burden to human and animal health worldwide. Robust and harmonized molecular epidemiological approaches and population studies that include routine disease screening are needed to efficiently track the origin and spread of Brucella strains. Core genome multilocus sequence typing (cgMLST) is a powerful genotyping system commonly used to delineate pathogen transmission routes for disease surveillance and control. Except for Brucella melitensis, cgMLST schemes for Brucella species are currently not established. Here, we describe a novel cgMLST scheme that covers multiple Brucella species. We first determined the phylogenetic breadth of the genus using 612 Brucella genomes. We selected 1,764 genes that were particularly well conserved and typeable in at least 98% of these genomes. We tested the new scheme on 600 genomes and found high agreement with the whole-genome-based single nucleotide polymorphism (SNP) analysis. Next, we applied the scheme to reanalyze the genome of Brucella strains from epidemiologically linked outbreaks. We demonstrated the applicability of the new scheme for high-resolution typing required in outbreak investigations as previously reported with whole-genome SNP methods. We also used the novel scheme to define the global population structure of the genus using 1,322 Brucella genomes. Finally, we demonstrated the possibility of tracing distribution of Brucella strains by performing cluster analysis of cgMLST profiles and found nearly identical cgMLST profiles in different countries. Our results show that sequencing depth of more than 40-fold is optimal for allele calling with this scheme. In summary, this study describes a novel Brucella-wide cgMLST scheme that is applicable in Brucella molecular epidemiology and helps in accurately tracking and thus controlling the sources of infection. The scheme is publicly accessible and should represent a valuable resource for laboratories with limited computational resources and bioinformatics expertise.

Journal ArticleDOI
01 Feb 2022-Viruses
TL;DR: This array, designated Riems Influenza A Typing Array version 2 (RitA-2), represents an updated and economized version of the RITA-1 array previously published by Hoffmann et al, and is particularly tailored to subtyping influenza A virus of avian hosts and Eurasian geographic origin.
Abstract: Avian influenza virus (AIV) variants emerge frequently, which challenges rapid diagnosis. Appropriate diagnosis reaching the sub- and pathotype level is the basis of combatting notifiable AIV infections. Real-time RT-PCR (RT-qPCR) has become a standard diagnostic tool. Here, a total of 24 arrayed RT-qPCRs is introduced for full subtyping of 16 hemagglutinin and nine neuraminidase subtypes of AIV. This array, designated Riems Influenza A Typing Array version 2 (RITA-2), represents an updated and economized version of the RITA-1 array previously published by Hoffmann et al. RITA-2 provides improved integration of assays (24 instead of 32 parallel reactions) and reduced assay volume (12.5 µL). The technique also adds RT-qPCRs to detect Newcastle Disease (NDV) and Infectious Bronchitis viruses (IBV). In addition, it maximizes inclusivity (all sequences within one subtype) and exclusivity (no intersubtypic cross-reactions) as shown in validation runs using a panel of 428 AIV reference isolates, 15 reference samples each of NDV and IBV, and 122 clinical samples. The open format of RITA-2 is particularly tailored to subtyping influenza A virus of avian hosts and Eurasian geographic origin. Decoupling and re-arranging selected RT-qPCRs to detect specific AIV variants causing epizootic outbreaks with a temporal and/or geographic restriction is possible.

Journal ArticleDOI
TL;DR: A comprehensive update to the international CPS sequence typing database for A. baumannii is provided, increasing the utility of this resource for prediction of CPS type from WGS to assist with clinical surveillance, and/or the design and application of CPS-targeted therapies.
Abstract: Several novel non-antibiotic therapeutics for the critical priority bacterial pathogen, Acinetobacter baumannii, rely on specificity to the cell-surface capsular polysaccharide (CPS). Hence, prediction of CPS type deduced from genes in whole genome sequence data underpins the development and application of these therapies. In this study, we provide a comprehensive update to the A. baumannii K locus reference sequence database for CPS typing (available in Kaptive v. 2.0.1) to include 145 new KL, providing a total of 237 KL reference sequences. The database was also reconfigured for compatibility with the updated Kaptive v. 2.0.0 code that enables prediction of ‘K type’ from special logic parameters defined by detected combinations of KL and additional genes outside the K locus. Validation of the database against 8994 publicly available A. baumannii genome assemblies from NCBI databases identified the specific KL in 73.45% of genomes with perfect, very high or high confidence. Poor sequence quality or the presence of insertion sequences were the main reasons for lower confidence levels. Overall, 17 KL were overrepresented in available genomes, with KL2 the most common followed by the related KL3 and KL22. Substantial variation in gene content of the central portion of the K locus, that usually includes genes specific to the CPS type, included 34 distinct groups of genes for synthesis of various complex sugars and >400 genes for forming linkages between sugars or adding non-sugar substituents. A repertoire of 681 gene types were found across the 237 KL, with 88.4% found in <5% of KL. 3. Significance as a BioResource to the community New therapies that target the bacterial polysaccharide capsule (CPS) show promise as effective tools to curb the high mortality rates associated with extensively resistant A. baumannii; one of the world’s most troublesome Gram-negative pathogens. As important information about the CPS structure produced by an isolate can be extracted from Whole Genome Sequences (WGS), simple bioinformatic tools and definitive sequence databases are needed to facilitate robust prediction of CPS type from WGS data. Here, we provide a comprehensive update to the international CPS sequence typing database for A. baumannii, increasing the utility of this resource for prediction of CPS type from WGS to assist with clinical surveillance, and/or the design and application of CPS-targeted therapies. This study is expected to further inform epidemiological tracking efforts, as well as the design of therapeutics targeting the CPS, enhancing global efforts to identify, trace and treat infections caused by this pathogen. 4. Data summary The updated A. baumannii KL reference sequence database including 241 fully annotated gene clusters is available for download under Kaptive v. 2.0.1 at https://github.com/katholt/Kaptive. Genome assemblies, short read data, or GenBank records used as representative reference sequence for each K locus are listed in Supplementary Table S1, and are referenced within each entry in the A. baumannii KL reference sequence database. The authors confirm all supporting data, code and protocols have been provided within the article or through supplementary data files.

Journal ArticleDOI
TL;DR: It is found that the isolation of PVL-positive strains has been increasing over the past decade and the molecular epidemiology of CA-MRSA in Japan differs from that in Europe and the United States; thus, it is crucial to monitor the trend of changes in CA- MRSA characteristics in Japan.
Abstract: Community-associated MRSA, which is a multidrug-resistant organism and can cause infections in otherwise-healthy individuals, has become a global problem. This paper describes a nationwide surveillance conducted in Japan to investigate changes in molecular epidemiological characteristics of CA-MRSA over the past decade and provides a detailed review of the characteristics of Panton-Valentine leucocidin (PVL)-positive strains isolated in 2018. ABSTRACT Although community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) has emerged worldwide, no nationwide CA-MRSA surveillance has been conducted in Japan to determine the changes in its molecular characteristics over time. We aimed to characterize the molecular epidemiology of Panton-Valentine leucocidin (PVL)-positive CA-MRSA strains collected from across Japan in the past decade. We isolated 1,770 MRSA strains from the skin and pus samples of outpatients of 244 medical facilities in 31 prefectures between 2010 and 2018 (2010, 2012, 2014, 2016, and 2018). Regions, hospitals, and periods in which strains were isolated and patient age group and sex were tabulated. Staphylococcal cassette chromosome mec (SCCmec) typing, detection of virulence factor genes, and antimicrobial susceptibility testing were performed. Whole-genome analysis was performed for the PVL-positive strains isolated in 2018. All strains harbored the mecA gene. Compared to that in 2010, the percentage of SCCmec type IV increased in 2018, with a corresponding increase in the proportion of PVL-positive strains (10% to 26%). Of the isolates obtained in 2018, clonal complex 8 (CC8) was dominant among PVL-positive strains. Core-genome single-nucleotide polymorphism analysis, using whole-genome sequencing, suggested that the CC8 PVL-positive strains spread throughout Japan over the last decade. Furthermore, a unique ST22 clone carrying both the PVL- and toxic shock syndrome toxin-1-encoding genes has emerged. We demonstrated that the molecular epidemiology of CA-MRSA in Japan differs from that in Europe and the United States; thus, it is crucial to monitor the trend of changes in CA-MRSA characteristics in Japan. IMPORTANCE Community-associated MRSA, which is a multidrug-resistant organism and can cause infections in otherwise-healthy individuals, has become a global problem. This paper describes a nationwide surveillance conducted in Japan to investigate changes in molecular epidemiological characteristics of CA-MRSA over the past decade and provides a detailed review of the characteristics of Panton-Valentine leucocidin (PVL)-positive strains isolated in 2018. Although CA-MRSA is rare in Japan to date, we found that the isolation of PVL-positive strains has been increasing over the past decade. In particular, the PVL-positive strains wherein CC8 was dominant exhibited high interstrain similarity, suggesting that a limited number of clones have spread over the past decade. Furthermore, a unique ST22 clone carrying both PVL-encoding and toxic shock syndrome toxin-1-encoding genes has emerged. This study shows that various changes can be observed when molecular epidemiological analysis, combined with next-generation sequencing, is conducted over a long period.

Journal ArticleDOI
TL;DR: Virulence assays showed that MSSA-PENS exhibited a level of virulence comparable to that of penicillin-resistant MSSA (MSSA- PENR), indicating that more-sensitive strains should not be mistaken for lacking aggressiveness in vivo.
Abstract: ABSTRACT Many countries have reported increasing rates of penicillin-susceptible methicillin-sensitive Staphylococcus aureus (MSSA-PENS). To date, there is relatively little known about the current situation and molecular characteristics of MSSA-PENS in China. In this study, we carried out a laboratory-based multi-region retrospective study to investigate the genomic epidemiology and characterisation of MSSA-PENS isolated from invasive bloodstream infections (BSIs) across 17 provinces. The prevalence of MSSA-PENS isolates increased significantly over the 6-year period, with the proportion increasing from 3.51% in 2014–8.80% in 2019, an average relative increase of 22.14% per year (95% confidence interval 9.67%-34.61%, P for trend <0.001), suggesting that China is experiencing a resurgence of MSSA-PENS. Phylogenetic analysis showed a higher strain diversity occurred; the most frequent clonal complexes (CCs) identified were CC188 (17.14%), CC398 (15.71%) and CC5 (15.71%). Over half of MSSA-PENS strains were pan-susceptible, with erythromycin the most frequent resistance observed. Moreover, 25 isolates were identified as immune evasion cluster negative, including CC15, CC188 and CC1, and 6 strains encoded the Panton-Valentine leucocidin gene. Importantly, virulence assays showed that MSSA-PENS exhibited a level of virulence comparable to that of penicillin-resistant MSSA (MSSA-PENR), indicating that more-sensitive strains should not be mistaken for lacking aggressiveness in vivo. Furthermore, 11 of these isolates were confirmed as blaZ positive but phenotype sensitive, with different amino acid changes in blaZ. Our data support the recommendation to clinicians regarding the usage of penicillin in invasive BSIs caused by MSSA-PENS, which might create a novel opportunity for better antimicrobial stewardship in the future.

Journal ArticleDOI
TL;DR: A high-resolution melting (HRM) method for the rapid screening of DNA and direct assignment into one of the 12 species of the Brucella genus based on single nucleotide polymorphisms (SNPs) based on whole genome SNP analysis based on 988 genomes.
Abstract: Brucella sp. are the causative agents of brucellosis. One of the main characteristics of the Brucella genus concerns its very high genetic homogeneity. To date, classical bacteriology typing is still considered as the gold standard assay for direct diagnosis of Brucella. Molecular approaches are routinely used for the identification of Brucella at the genus level. However, genotyping is more complex, and to date, no method exists to quickly assign a strain into species and biovar levels, and new approaches are required. Next generation sequencing (NGS) opened a new era into the diagnosis of bacterial diseases. In this study, we designed a high-resolution melting (HRM) method for the rapid screening of DNA and direct assignment into one of the 12 species of the Brucella genus. This method is based on 17 relevant single nucleotide polymorphisms (SNPs), identified and selected from a whole genome SNP (wgSNP) analysis based on 988 genomes (complete and drafts). These markers were tested against the collection of the European Reference Laboratory (EU-RL) for brucellosis (1440 DNAs extracted from Brucella strains). The results confirmed the reliability of the panel of 17 SNP markers, allowing the differentiation of each species of Brucella together with biovars 1, 2, and 3 of B. suis and vaccine strain Rev1 (B. melitensis) within 3 h, which is a considerable gain of time for brucellosis diagnosis. Therefore, this genotyping tool provides a new and quick alternative for Brucella identification based on SNPs with the HRM-PCR assay.

Journal ArticleDOI
TL;DR: It is concluded that the risk of exchanging S. aureus/MRSA between humans and dogs is higher than that for S. pseudintermedius.
Abstract: There are uncertainties with respect to the transmission of methicillin-susceptible and methicillin-resistant Staphylococcus aureus (MSSA and MRSA) and Staphylococcus pseudintermedius between dogs and humans. In this study, we investigated concomitant nasal colonization of dogs and humans in three cohorts. Cohort I, households owning dogs: In 42 of 84 households, 66 humans (36.9%) and 10 dogs (8.9%) carried S. aureus. MRSA, attributed to sequence type (ST) 22 and ST130, were detected in two (1.1%) of the humans but in none of the dogs. Typing by means of spa-typing and whole-genome sequencing (WGS) indicated eight transmissions of S. aureus between humans and dogs in 8 of 42 (19.0%) households with human S. aureus carriers, whereas in 11 of 38 (29.0%) households with ≥two persons and S. aureus colonization of humans, 15 human-to-human transmissions were observed (p = 0.43). S. pseudintermedius was isolated from 42 dogs (37.5%), but from only one human (0.6%). In this case, WGS-based typing indicated strong relatedness of this isolate with a canine isolate from the same household. Cohort II, dogs and their owners visiting a veterinary practice: Among 17 humans and 17 dogs attending a veterinary practice, MSSA was detected in three humans and two dogs, and S. pseudintermedius in only six dogs. Cohort III, dogs used for animal-assisted interventions in human healthcare facilities and their owners: MSSA was obtained in 1 of 59 dogs (1.7%) and in 17 of 60 (28.3%) of the dog owners, while S. pseudintermedius was isolated from seven (12%) dogs and one (1.7%) human owner. We conclude that the risk of exchanging S. aureus/MRSA between humans and dogs is higher than that for S. pseudintermedius.

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TL;DR: In this article , the authors conducted a comprehensive study of 23 bushmeat markets in Cameroon and one seizure from a French airport using a multi-gene DNA-typing approach and a dedicated species-assignment pipeline (DNABUSHMEAT).

Journal ArticleDOI
TL;DR: A rapid high-resolution HLA typing assay using RNA-Seq that can provide accurate HLA genotyping and HLA allele-specific transcript expression in 7–8 h, a timeline short enough to perform the assay for deceased donors is demonstrated.
Abstract: HLA typing provides essential results for stem cell and solid organ transplants, as well as providing diagnostic benefits for various rheumatology, gastroenterology, neurology, and infectious diseases. It is becoming increasingly clear that understanding the expression of patient HLA transcripts can provide additional benefits for many of these same patient groups. Our study cohort was evaluated using a long-read RNA sequencing methodology to provide rapid HLA genotyping results and normalized HLA transcript expression. Our assay used NGSEngine to determine the HLA genotyping result and normalized mRNA transcript expression using Athlon2. The assay demonstrated an excellent concordance rate of 99.7%. Similar to previous studies, for the class I loci, patients demonstrated significantly lower expression of HLA-C than HLA-A and -B (Mann–Whitney U, p value = 0.0065 and p value = 0.0154, respectively). In general, the expression of class II transcripts was lower than that of class I transcripts. This study demonstrates a rapid high-resolution HLA typing assay using RNA-Seq that can provide accurate HLA genotyping and HLA allele-specific transcript expression in 7–8 h, a timeline short enough to perform the assay for deceased donors.

Journal ArticleDOI
TL;DR: Wang et al. as discussed by the authors explored the prevalence of blaCTX-M and elucidated the characteristics of transferable blaaCTXM-55 plasmids, and selected three transferable plasmid for whole genome sequencing sequencing and further analysis of genetic context.
Abstract: The aim of this study was to explore the prevalence of blaCTX-M and elucidate the characteristics of transferable blaCTX-M-55 plasmids.Escherichia coli were isolated from 10 large-scale duck farms from nine counties and cities in Guizhou Province. They were identified, subjected to antimicrobial susceptibility testing and screened for antibiotic resistance genes. blaCTX-M-positive strains were subtyped, and blaCTX-M-55-containing strains were further analysed by multilocus sequence typing, conjugation experiments and polymerase chain reaction-based replicon typing. Three transferable plasmids were selected for whole genome sequencing sequencing and further analysis of genetic context.In total, 128 E. coli strains were identified, and 23 were blaCTX-M-positive. Subtype analysis of blaCTX-M-positive strains revealed that 23 were blaCTX-M-55 and one of the strains contained both blaCTX-M-65 and blaCTX-M-55. They were divided into 12 multilocus sequence typing types, and 6 were undetermined. blaCTX-M-55-bearing plasmids in seven were transferred to E. coli C600 at frequencies between 10-6 and 10-9, and blaCTX-M-55 was found to be located on IncN, IncFⅡ and IncFII-N fusion type plasmids. Three plasmids (IncFⅡ plasmids pEC99_Plas3 and pS1_Plas2 and IncFII-N plasmid pD2_plas2) shared a homologous region of IS26-ARGs-IS26-blaTEM-orf477-blaCTX-M-55-ISEcp1△-IS26.Our study demonstrates the environmental presence of blaCTX-M-55 harbouring E. coli in Guizhou. Analysis of the genetic context of blaCTX-M-55 aids in understanding possible dissemination routes of ESBL genes and controlling widespread antibiotic resistance.

Journal ArticleDOI
TL;DR: Food and food production environments may be a source of L. monocytogenes of serogroup IIa with pathogenic potential and the presence of several virulence genes or putative molecular markers playing a role in pathogenesis of listeriosis and involved in survival in adverse environmental conditions is shown.
Abstract: In the present study, 100 L. monocytogenes isolates of serogroup IIa from food and food production environments in Poland were characterized towards the presence of virulence, resistance, and stress response genes using whole-genome sequencing (WGS). The strains were also molecularly typed and compared with multi-locus sequence typing (MLST) and core genome MLST analyses. The present isolates were grouped into 6 sublineages (SLs), with the most prevalent SL155 (33 isolates), SL121 (32 isolates), and SL8 (28 isolates) and classified into six clonal complexes, with the most prevalent CC155 (33 strains), CC121 (32 isolates), and CC8 (28 strains). Furthermore, the strains were grouped to eight sequence types, with the most prevalent ST155 (33 strains), ST121 (30 isolates), and ST8 (28; strains) followed by 60 cgMLST types (CTs). WGS data showed the presence of several virulence genes or putative molecular markers playing a role in pathogenesis of listeriosis and involved in survival of L. monocytogenes in adverse environmental conditions. Some of the present strains were molecularly closely related to L. monocytogenes previously isolated in Poland. The results of the study showed that food and food production environments may be a source of L. monocytogenes of serogroup IIa with pathogenic potential.

Journal ArticleDOI
TL;DR: The methods for typing M. pneumoniae and M. genitalium, including the results of their application in different studies, are summarized and current knowledge regarding the association of typing data with the clinical characteristics of infections is presented.
Abstract: Mycoplasma pneumoniae and Mycoplasma genitalium are cell wall-less bacteria with strongly reduced genome content and close phylogenetic relatedness. In humans, the only known natural host, the microorganisms colonize the respiratory or genitourinary mucosa and may cause a broad range of clinical presentations. Besides fundamental differences in their tissue specificity, transmission route, and ability to cause prevalence peaks, both species share similarities such as the occurrence of asymptomatic carriers, preferred populations for infection, and problems with high rates of antimicrobial resistance. To further understand the epidemiology of these practically challenging bacteria, typing of strains is necessary. Since the cultivation of both pathogens is difficult and not performed outside of specialized laboratories, molecular typing methods with adequate discriminatory power, stability, and reproducibility have been developed. These include the characterization of genes containing repetitive sequences, of variable genome regions without the presence of repetitive sequences, determination of single and multi-locus variable-number tandem repeats, and detection of single nucleotide polymorphisms in different genes, respectively. The current repertoire of procedures allows reliable differentiation of strains circulating in different populations and in different time periods as well as comparison of strains occurring subsequently in individual patients. In this review, the methods for typing M. pneumoniae and M. genitalium, including the results of their application in different studies, are summarized and current knowledge regarding the association of typing data with the clinical characteristics of infections is presented.

Journal ArticleDOI
TL;DR: The developed method allows for the rapid identification of HAdV respiratory pathogens, including recombinants, and bypasses the need for whole genome sequencing for real-time surveillance of circulating adenovirus strains in outbreaks and populations by clinical virologists, public health officials, and epidemiologists.
Abstract: Human adenoviruses (HAdVs) within species B, C, and E are responsible for highly contagious and potentially severe respiratory disease infections. The traditional method to type these pathogens was based on virus neutralization and hemagglutination assays, which are both time-consuming and difficult, particularly due to the nonavailability of reagents. Subsequent molecular typing based on the partial characterization of the hexon gene and/or the restriction enzyme analysis (REA) of the genomes is inadequate, particularly in identifying recombinants. Here, a rapid, simple, and cost-effective method for molecular typing HAdV respiratory pathogens is presented. This incorporates three pairs of universal PCR primers that target the variable regions of the three major capsid genes, i.e., hexon, penton base, and fiber genes, that span the genome. The protocol enables typing and characterization of genotypes within species B, C, and E, as well as of some genotypes within species D and F. To validate this method, we surveyed 100 children with HAdV-associated acute respiratory infections identified by direct immunofluorescence (Hong Kong; July through October, 2014). Throat swab specimens were collected and analyzed by PCR amplification and sequencing; these sequences were characterized by BLAST. HAdVs were detected in 98 out of 100 (98%) samples, distributing as follows: 74 HAdV-B3 (74%); 10 HAdV-E4 (10%); 7 HAdV-C2 (7%); 2 HAdV-C6 (2%); 1 HAdV-B7 (1%); 1 HAdV-C1 (1%); 2 co-infection (2%); and 1 novel recombinant (1%). This study is the first detailed molecular epidemiological survey of HAdVs in Hong Kong. The developed method allows for the rapid identification of HAdV respiratory pathogens, including recombinants, and bypasses the need for whole genome sequencing for real-time surveillance of circulating adenovirus strains in outbreaks and populations by clinical virologists, public health officials, and epidemiologists.

Journal ArticleDOI
TL;DR: This work used variant typing PCR to describe the evolution of SARS-CoV-2 Omicron sublineages between December 2021 and mid-March 2022 and found that the selective advantage of the BA.2 variant over BA.1 is not due to greater nasopharyngeal viral loads.
Abstract: We used variant typing PCR to describe the evolution of SARS-CoV-2 Omicron sublineages between December 2021 and mid-March 2022. The selective advantage of the BA.2 variant over BA.1 is not due to greater nasopharyngeal viral loads.

DOI
01 Jan 2022
TL;DR: In this paper, a type-case construction for intersection type systems is proposed, which combines the union elimination rule of the intersection type system and the typing rules for type-cases of the extension.
Abstract: We extend classic union and intersection type systems with a type-case construction and show that the combination of the union elimination rule of the former and the typing rules for type-cases of our extension encompasses occurrence typing. To apply this system in practice, we define a canonical form for the expressions of our extension, called MSC-form. We show that an expression of the extension is typable if and only if its MSC-form is, and reduce the problem of typing the latter to the one of reconstructing annotations for that term. We provide a sound algorithm that performs this reconstruction and a proof-of-concept implementation

Journal ArticleDOI
TL;DR: PLASTER as mentioned in this paper is a robust data processing pipeline for accurate allele typing of SMRT sequenced amplicons, which can improve drug safety and efficacy through screening prior to drug administration, but it is limited in scale and has employed nascent data processing pipelines.
Abstract: Abstract The CYP2D6 enzyme is estimated to metabolize 25% of commonly used pharmaceuticals and is of intense pharmacogenetic interest due to the polymorphic nature of the CYP2D6 gene. Accurate allele typing of CYP2D6 has proved challenging due to frequent copy number variants (CNVs) and paralogous pseudogenes. SNP-arrays, qPCR and short-read sequencing have been employed to interrogate CYP2D6 , however these technologies are unable to capture longer range information. Long-read sequencing using the PacBio Single Molecule Real Time (SMRT) sequencing platform has yielded promising results for CYP2D6 allele typing. However, previous studies have been limited in scale and have employed nascent data processing pipelines. We present a robust data processing pipeline “PLASTER” for accurate allele typing of SMRT sequenced amplicons. We demonstrate the pipeline by typing CYP2D6 alleles in a large cohort of 377 Solomon Islanders. This pharmacogenetic method will improve drug safety and efficacy through screening prior to drug administration.

Journal ArticleDOI
TL;DR: In this paper , the authors build a collection of Salmonella isolates that are genetically diverse and evaluate their ability to form biofilm under different conditions relevant to a processing environment.
Abstract: Salmonella is extremely diverse, with >2500 serovars that are genetically and phenotypically diverse. The aim of this study was to build a collection of Salmonella isolates that are genetically diverse and to evaluate their ability to form biofilm under different conditions relevant to a processing environment.Twenty Salmonella isolates representative of 10 serovars were subtyped using Clustered regularly interspaced short palindromic repeats (CRISPR)-typing to assess the genetic diversity between isolates of each serovar. Biofilm formation of the isolates on both plastic and stainless-steel surfaces at 25 and 15°C was assessed. At 25°C, 8/20 isolates each produced strong and moderate biofilm on plastic surface compared to stainless-steel (3/20 and 13/20 respectively). At 15°C, 5/20 produced strong biofilm on plastic surface and none on stainless-steel. Several isolates produced weak biofilm on plastic (11/20) and stainless-steel (16/20) surfaces. Serovar Schwarzengrund consistently produced strong biofilm while serovars Heidelberg and Newport produced weak biofilm.These results suggest that Salmonellae differ in their attachment depending on the surface and temperature conditions encountered, which may influence persistence in the processing environment.These differences in biofilm formation could provide useful information for mitigation of Salmonella in processing environments.