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Vulnerable patients in an intensive care unit (ICU) setting are at high risk of infection from bacteria including gut-colonising Escherichia coli and Klebsiella species. Complex ICU procedures often depend on successful antimicrobial treatment, underscoring the importance of understanding the extent of patient colonisation by multi-drug-resistant organisms (MDROs) in large UK ICUs. Previous work on ICUs globally uncovered high rates of colonisation by transmission of MDROs, but the situation in UK ICUs is less understood. Here, we investigated the diversity and antibiotic resistance gene (ARG) carriage of bacteria present in one of the largest UK ICUs at the Queen Elizabeth Hospital Birmingham (QEHB), focusing primarily on E. coli as both a widespread commensal and a globally disseminated multi-drug-resistant pathogen. Samples were taken during highly restrictive coronavirus disease 2019 (COVID-19) control measures from May to December 2021. Whole-genome and metagenomic sequencing were used to detect and report strain-level colonisation of patients, focusing on E. coli sequence types (STs), their colonisation dynamics and antimicrobial resistance gene carriage. We found a lack of multi-drug resistance (MDR) in the QEHB. Only one carbapenemase-producing organism was isolated, a Citrobacter carrying bla KPC-2 . There was no evidence supporting the spread of this strain, and there was little evidence overall of nosocomial acquisition or circulation of colonising E. coli . Whilst 22 different E. coli STs were identified, only 1 strain of the pandemic ST131 lineage was isolated. This ST131 strain was non-MDR and was found to be a clade A strain, associated with low levels of antibiotic resistance. Overall, the QEHB ICU had very low levels of pandemic or MDR strains, a result that may be influenced in part by the strict COVID-19 control measures in place at the time. Employing some of these infection prevention and control measures where reasonable in all ICUs might therefore assist in maintaining low levels of nosocomial MDR.

Bacteria are fascinating research objects in many disciplines for countless reasons, and whole-genome sequencing (WGS) has become the paramount methodology to advance our microbiological understanding. Meanwhile, access to cost-effective sequencing platforms has accelerated bacterial WGS to unprecedented levels, introducing new challenges in terms of data accessibility, computational demands, heterogeneity of analysis workflows and, thus, ultimately its scientific usability. To this end, a previous study released a uniformly processed set of 661 405 bacterial genome assemblies obtained from the European Nucleotide Archive as of November 2018. Building on these accomplishments, we conducted further genome-based analyses like taxonomic classification, multilocus sequence typing and annotation of all genomes. Here, we present BakRep, a searchable large-scale web repository of these genomes enriched with consistent genome characterizations and original metadata. The platform provides a flexible search engine combining taxonomic, genomic and metadata information, as well as interactive elements to visualize genomic features. Furthermore, all results can be downloaded for offline analyses via an accompanying command line tool. The web repository is accessible via https://bakrep.computational.bio.

Introduction. Acinetobacter baumannii is a common cause of multidrug-resistant (MDR) nosocomial infections worldwide, including Vietnam. Hypothesis. Analysis of crucial genetic factors may link to epidemiological characteristics and antibiotic resistance of A. baumannii clinical strains in Vietnamese hospitals. Methodology. Fifty-one A . baumannii clinical strains from six different tertiary hospitals in Vietnam were analysed using whole genome sequencing (WGS), between 2017 and 2019. Results. Eleven sequence types (STs) were identified, including four STs reported for the first time in Vietnam based on the PubMLST database and three new STs not previously documented. ST1336, ST1260 and ST575 were found exclusively in Vietnam. These STs were widely distributed in all hospitals in Vietnam, with ST2 and ST571 being the most dominant. Resistant rates to eight antibiotics, belonging to four antibiotic groups, were very high (72.5-94.1 %) with high MIC values, while resistance to colistin was 29.4%. Fifty-one isolates were identified as MDR, with 100% (51/51) isolates carrying antimicrobial-resistant (AMR) genes, and 52 antibiotic-resistant genes were detected among these strains, including β-lactam (22 genes), chloramphenicol (5 genes), lincosamide (2 genes), aminoglycoside (11 genes), rifampicin (1 gene), quinolone (2 genes), sulfonamide and trimethoprim (4 genes) and tetracycline (5 genes) resistance. The most commonly found mobile structures carried partial or complete transposons: ISaba24/ISEc29/ISEc35 contains a series of antibiotic-resistant genes. Conclusion. The WGS results of the 51 strains of A. baumannii provided important information regarding the distribution of STs and associated antibiotic-resistant genes among A. baumannii strains.

The Actinomycetota (formerly Actinobacteria) genus Clavibacter includes phytopathogens with devasting effects in several crops. Clavibacter michiganensis , the causal agent of tomato bacterial canker, is the most notorious species of the genus. Yet, its origin and natural reservoirs remain elusive, and its populations show pathogenicity profiles with unpredictable plant disease outcomes. Here, we generate and analyse a decade-long genomic dataset of Clavibacter from wild and commercial tomato cultivars, providing evolutionary insights that directed phenotypic characterization. Our phylogeny situates the last common ancestor of C. michiganensis next to Clavibacter isolates from grasses rather than to the sole strain we could isolate from wild tomatoes. Pathogenicity profiling of C. michiganensis isolates, together with C. phaseoli and C. californiensis as sister taxa and the wild tomato strain, was found to be congruent with the proposed phylogenetic relationships. We then identified gene enrichment after the evolutionary event, leading to the appearance of the C. michiganesis clade, including known pathogenicity factors but also hitherto unnoticed genes with the ability to encode adaptive traits for a pathogenic lifestyle. The holistic perspective provided by our evolutionary analyses hints towards a host shift event as the origin of C. michiganensis as a tomato pathogen and the existence of pathogenic genes that remain to be characterized.

An endosymbiotic bacterium of the genus Sodalis , designated as strain HZ T , was cultured from the parasitoid wasp Spalangia cameroni , which develops on the pupae of various host flies. The bacterium was detected in S. cameroni developed on houseflies, Musca domestica , in a poultry facility in Hazon, northern Israel. After culturing, this bacterium displayed no surface motility on Luria-Bertani agar and was rod-shaped and irregular in size, ~10-30 nm in diameter and 5-20 µm in length. Phylogenetic analyses revealed that strain HZ T is closely related to Sodalis praecaptivus strain HS T , a free-living species of the genus Sodalis that includes many insect endosymbionts. Although these bacteria maintain >98% sequence identity in shared genes, genomic characterization revealed that strain HZ T has undergone substantial reductive evolution, such that it lacks many gene functions that are maintained in S. praecaptivus strain HS T . Based on the results of phylogenetic, genomic and chemotaxonomic analyses, we propose that this endosymbiont should be classified in a new subspecies as S. praecaptivus subsp. s palangiae subsp. nov. The type strain for this new subspecies is HZ T (=ATCC TSD-398 T =NCIMB 15482 T ). The subspecies Sodalis praecaptivus subsp. praecaptivus strain HS T is created automatically with the type strain ATCC BAA-2554 T (=DSMZ 27494 T ).

A novel Gram-stain-positive, facultatively anaerobic, non-motile, catalase-positive, oxidase-negative and ovoid cocci, designated as A1S7 T , was isolated from the gut of a marine sandworm ( Alitta virens ). Strain A1S7 T exhibited optimal growth at temperatures of 20-30 ℃, pH 6-8 and in the presence of 2-4% (w/v) NaCl. Phylogenetic analysis based on the 16S rRNA gene sequence indicated that strain A1S7 T belonged to the genus Janibacter , exhibiting a similarity of 99.0% to Janibacter cremeus KCTC 49873 T , followed by Janibacter anophelis KCTC 19282 T (98.8%), Janibacter hoylei KCTC 49872 T (98.4%), Janibacter limosus KACC 20518 T (98.2%) and Janibacter corallicola KACC 21120 T (97.2%). The complete genome sequence of strain A1S7 T revealed a genome size of 3360920 bp with a genomic G+C content of 70.1 mol%. The orthologous average nucleotide identity and the digital DNA-DNA hybridization values between strain A1S7 T and Janibacter cremeus KCTC 49873 T were determined to be 89.5 and 37.2%, respectively. The major respiratory quinone was MK-8(H 4 ). The predominant fatty acids (>10%) included iso -C 16:0 , C 17:1  ω 8 c , C 18:1  ω 9 c and C 17:0 . Polar lipids comprised diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol, one unknown phosphoglycolipid and three unknown polar lipids. The cell-wall peptidoglycan type was A1 γ . The major whole-cell sugars were ribose, mannose and glucose. Based on phenotypic, phylogenetic, genotypic and chemotaxonomic properties, strain A1S7 T represents a novel species in the genus Janibacter , for which the name Janibacter alittae sp. nov. is proposed. The type strain is A1S7 T (=KCTC 49714 T = JCM 36706 T ).

The genome of the methanotrophic bacterium Methylohalobius crimeensis strain 10Ki contains a gene cluster that encodes a putative coenzyme-M (CoM)-dependent pathway for oxidation of epoxyethane, based on homology to genes in bacteria that grow on ethylene and propylene as sole substrates. An alkene monooxygenase was not detected in the M. crimeensis genome, so epoxyethane is likely produced from co-oxidation of ethylene by the methane monooxygenase enzyme. Similar gene clusters were detected in about 10% of available genomes from aerobic methanotrophic bacteria, primarily strains grown from rice paddies and other wetlands. The sparse occurrence of the gene cluster across distant phylogenetic groups suggests that multiple lateral gene transfer events have occurred in methanotrophs. In support of this, the gene cluster in M. crimeensis was detected within a large genomic island predicted using multiple methods. Growth studies, reverse transcription-quantitative PCR (RT-qPCR) and proteomics were performed to examine the expression of these genes in M. crimeensis . Growth and methane oxidation activity were completely inhibited by the addition of >0.5% (v/v) ethylene to the headspace of cultures, but at 0.125% and below, the inhibition was only partial, and ethylene was gradually oxidized. The etnE gene encoding epoxyalkane:CoM transferase was strongly upregulated in ethylene-exposed cells based on RT-qPCR. Proteomics analysis confirmed that EtnE and nine other proteins encoded in the same gene cluster became much more predominant after cells were exposed to ethylene. The results suggest that ethylene is strongly inhibitory to M. crimeensis , but the bacterium responds to ethylene exposure by expressing an epoxide oxidation system similar to that used by bacteria that grow on alkenes. In the obligate methanotroph M. crimeensis , this system does not facilitate growth on ethylene but likely alleviates toxicity of epoxyethane formed through ethylene co-oxidation by particulate methane monooxygenase. The presence of predicted epoxide detoxification systems in several other wetland methanotrophs suggests that co-oxidation of ambient ethylene presents a stress for methanotrophic bacteria in these environments and that epoxyethane removal has adaptive value.

Four strains, representing two novel Bifidobacterium species, were isolated from water kefir, a fermented beverage. 16S rRNA gene analysis suggested that the novel species share high identities (98.82-98.89%) with Bifidobacterium aquikefiri LMG 28769 T . Complete genomes were assembled with a short- and long-read hybrid sequencing approach. In agreement with the 16S rRNA gene analysis, phylogenetics with 117 marker genes places the novel species closest to B. aquikefiri LMG 28769 T as well. The isolates have average nucleotide identity (ANI) scores ranging from 81.46 to 84.84% and digital DNA-DNA hybridization (dDDH) scores from 23.9 to 38.5% with the closest related species, as well as ANI scores between the proposed new species of 80.50%, indicating that the isolates represent two novel species. Matrix-assisted laser desorption/ionization-time of flight chemotaxonomic analysis supported the gene-based taxonomic placement. We propose the names Bifidobacterium fermentum sp. nov. and Bifidobacterium aquikefiricola sp. nov. for these novel species within the Bifidobacterium genus. The proposed type strain B. fermentum WK012_4_13 T (= LMG 33104 T = DSM 116073 T ; GenBank accession number GCF_041080835.1) has a genome size of 2.43 Mbp, with a G+C content of 56.00 mol%. The proposed type strain for B. aquikefiricola WK041_4_12 T (= LMG 33105 T = DSM 116074 T ; GenBank accession number GCF_041080795.1) has a genome size of 2.36 Mbp and a G+C content of 53.94 mol%. B. fermentum cells are Gram-positive staining, non-motile, non-spore-forming, fructose-6-phosphate phosphoketolase (F6PPK)-positive, catalase- and oxidase-negative and bacillary club shaped. B. aquikefiricola cells are Gram-positive staining, non-motile, non-spore-forming, F6PPK-positive, catalase- and oxidase-negative and square rod shaped.

Two separate bacterial strains, PMTSA4 T and PMR2A8, were isolated from the rhizospheric soils of bell pepper plants grown in a plant nursery. These strains are Gram-negative, non-motile and rod-shaped and grow in aerobic conditions. They exhibit a positive reaction for catalase activity but negative results for oxidase activity. Phylogenetic analysis of the 16S rRNA gene sequences revealed that the strains PMTSA4 T and PMR2A8 are closely related to Flavobacterium piscinae ICH-30 T (95.6%, respectively), Flavobacterium ahnfeltiae 10Alg 130 T (95.5%) and Flavobacterium maris KMM 9535 T (95.3%), aligning them within the genus Flavobacterium . Digital DNA-DNA hybridization (dDDH) values and average nucleotide identities (ANIs) of the whole-genome sequences for the two strains and related Flavobacterium species were significantly below the established thresholds for prokaryotic species delineation (<70% for dDDH and <95% for ANI). The observed values were as follows: Flavobacterium aquatile LMG 4008 T (dDDH: 19.8% and ANI: 75.5%), F. piscinae ICH-30 T (dDDH: 18.6% and ANI: 73.3%) and F. stagni WWJ 16 T (dDDH: 18.5% and ANI: 72.0%). The strains have genome sizes of 3 068 185 bp and 3 068 330 bp, with a G+C content of 32.5 mol%. In phenotypic characterization, the new strains grew at 10-35 °C and tolerated up to 4% NaCl at pH 5-9 (optimum pH 8). The predominant cellular fatty acids were observed to be iso-C 15:0 , iso-C 17:0 3-OH and iso-C 15:0 3-OH. Menaquinone-6 was the predominant quinone. Considering the results from phenotypic, chemotaxonomic, phylogenetic and genomic analyses, it is proposed that the strains PMTSA4 T and PMR2A8 represent a novel species within the genus Flavobacterium .

Antimicrobial resistance (AMR) poses a significant threat to global public health, with the potential to cause millions of deaths annually by 2050. Effective surveillance of AMR pathogens is crucial for monitoring and predicting their behaviour in response to antibiotics. However, many public health professionals lack the necessary bioinformatics skills and resources to analyse pathogen genomes effectively. To address this challenge, we developed AMRColab, an open-access bioinformatics analysis suite hosted on Google Colaboratory. AMRColab enables users with limited or no bioinformatics training to detect and visualize AMR determinants in pathogen genomes using a 'plug-and-play' approach. The platform integrates established bioinformatics tools such as AMRFinderPlus and hAMRonization, allowing users to analyse, compare and visualize trends in AMR pathogens easily. A trial run using methicillin-resistant Staphylococcus aureus (MRSA) strains demonstrated AMRColab's effectiveness in identifying AMR determinants and facilitating comparative analysis across strains. A workshop was conducted and feedback from participants indicated high confidence in using AMRColab and a willingness to incorporate it into their research. AMRColab's user-friendly interface and modular design make it accessible to a diverse audience, including medical laboratory technologists, medical doctors and public health scientists, regardless of their bioinformatics expertise. Future improvements to AMRColab will include enhanced visualization tools, multilingual support and the establishment of an online community platform. AMRColab represents a significant step towards democratizing AMR surveillance and empowering public health professionals to combat AMR effectively.

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.

Acetic acid bacteria - belonging to the Acetobacteraceae family - are found in the gut of many sugar-feeding insects. In this study, six strains have been isolated from the hemipteran leafhopper Euscelidius variegatus . While they exhibit high 16S rRNA gene sequence similarities to uncultured members of the Acetobacteraceae family, they could not be unequivocally assigned to any particular type species. Considering the clonality of the six isolates, the EV16P T strain was used as a representative of this group of isolates. The genome sequence of EV16P T is composed of a 2.388 Mbp chromosome, with a DNA G+C content of 57 mol%. Phylogenetic analyses based on the 16S rRNA gene sequence and whole-genome multilocus sequence analysis indicate that EV16P T forms a monophyletic clade with the uncultivated endosymbiont of Diaphorina citri , the Candidatus Kirkpatrickella diaphorinae. Such a phylogenetic clade is positioned between those of Asaia-Swaminathania and Kozakia . The genomic distance metrics based on gene and protein sequences support the proposal that EV16P T is a new species belonging to a yet-undescribed genus. It is a rod-shaped Gram-stain-negative bacterium, strictly aerobic, non-motile, non-spore-forming, showing optimal growth without salt (NaCl) at 30 °C and pH of 6-7. The major quinone is Q10, and the dominant cellular fatty acids (>10%) are C 18:l ω 7c, C 19 : 0 cyclo ω 6c, C 16 : 0 and C 19 : 1 2OH. The polar lipid profile comprises diphosphatidylglycerol, phosphatidylethanolamine and phosphatidylcholine, along with unidentified aminophospholipids, glycophospholipids, aminolipids and lipids. Based on a polyphasic approach, including phylogenetic, phylogenomic, genome relatedness, phenotypic and chemotaxonomic characterisations, EV16P T (= KCTC 8296 T , = DSM 117028 T ) is proposed as a representative of a novel species in a novel genus with the proposed name Sorlinia euscelidii gen. nov., sp. nov., in honour of Prof. Claudia Sorlini, an Italian environmental microbiologist at the University of Milan who inspired the research on microbial diversity, including symbiosis in plants and animals.

The novel strain CBA3628 T was isolated from kimchi, a Korean fermented vegetable. CBA3628 T is a cocci-shaped, Gram-stain-positive, catalase- and oxidase-negative and facultatively anaerobic bacterium. The results of phylogenetic analysis based on 16S rRNA gene sequencing indicated that CBA3628 T represented a member of the genus Leuconostoc of the family Leuconostocaceae . CBA3628 T has a circular chromosomal genome and three plasmids of 1 864 558 bp (37% DNA G+C content), containing 1,887 genes, 1,762 predicted protein-coding genes, 4 complete rRNA loci and 70 tRNA genes. The cells were non-haemolytic, non-motile and non-spore forming. The optimal growth of CBA3628 T occurred at 30 °C, pH 6.0 and with 0-2% (w/v) NaCl. The major polar lipids of CBA3628 T were diphosphatidylglycerol and phosphatidylglycerol. The major fatty acids (>10%) of CBA3628 T were C 16  :  0 , C 20  :  0 and C 19 : 0 cyclo ω8 c . CBA3628 T contained A3α-type peptidoglycans. CBA3628 T was most closely related to Leuconostoc mesenteroides subsp. mesenteroides ATCC 8293 T , L. mesenteroides subsp. dextranicum DSM 20484 T and L. suionicum DSM 20241 T with 99.52% 16S rRNA gene sequence similarity. However, the average nucleotide identities of 91.9%, 91.7% and 91.1% and the digital DNA-DNA hybridisation values of 45.6%, 45.4% and 45.4% indicated that the novel isolate represented a distinct species. Phylogenetic analyses of both the 16S rRNA gene and genome sequences revealed that CBA3628 T formed a distinct phylogenetic lineage within the genus Leuconostoc and was most closely related to Leuconostoc litchii MB7 T . The ANI and dDDH values between CBA3628 T and L. litchii MB7 T were 84.9 and 22.8%, respectively. Functional genes belonging to COG categories E, J and K were enriched in the genome of CBA3628 T (>7.9%). On the basis of its physiological, chemotaxonomic, phylogenetic and genomic properties, strain CBA3628 T represents a novel species from the genus Leuconostoc , for which we propose the name Leuconostoc koreense sp. nov., with the type strain CBA3628 T (= KACC 23049 T = DSM 116836 T ).

Group B Streptococcus (GBS) causes severe infections in neonates and adults with comorbidities. Prophages have been reported to contribute to GBS evolution and pathogenicity. However, no studies are available to date on the presence and diversity of prophages in GBS isolates from humans in South America. This study provides insights into the prophage content of 365 GBS isolates collected from clinical samples in the context of an Argentinean multicentric study. Using whole-genome sequence data, we implemented two previously proposed methods for prophage typing: a PCR approach (carried out in silico ) coupled with a blastx-based method to classify prophages based on their prophage group and integrase type, respectively. We manually searched the genomes and identified 325 prophages. However, only 80% of prophages could be accurately categorized with the previous approaches. Integration of phylogenetic analysis, prophage group and integrase type allowed for all to be classified into 19 prophage types, which correlated with GBS clonal complex grouping. The revised prophage typing approach was additionally improved by using a blastn search after enriching the database with ten new genes for prophage group classification combined with the existing integrase typing method. This modified and integrated typing system was applied to the analysis of 615 GBS genomes (365 GBS from Argentina and 250 from public databases), which revealed 29 prophage types, including two novel integrase subtypes. Their characterization and comparative analysis revealed major differences in the lysogeny and replication modules. Genes related to bacterial fitness, virulence or adaptation to stressful environments were detected in all prophage types. Considering prophage prevalence, distribution and their association with bacterial virulence, it is important to study their role in GBS epidemiology. In this context, we propose the use of an improved and integrated prophage typing system suitable for rapid phage detection and classification with little computational processing.

Introduction. Streptococcus uberis is a common cause of mastitis in cattle, leading to significant economic losses. The widespread use of antimicrobials has contributed to the emergence of resistance, which poses a severe challenge in controlling S. uberis infection. Aim. The objective of this study was to gain insights into the antimicrobial resistance (AMR) and epidemiological typing of S. uberis isolated from milk collected from bovine mastitis on dairy farms in Thuringia. Methodology. In this study, 84 S. uberis isolates were obtained from cattle with clinical mastitis in Thuringia, their phenotypic and genotypic AMR were analyzed and their phylogenetic relationship was explored using whole-genome sequencing. Results. Genetically heterogeneous strains were found on the farms, but clusters of highly similar strains also circulated within the same farms. All isolates were sensitive to ampicillin, penicillin, ceftiofur, and vancomycin. However, 42.9%, 42.9%, 22.6%, 19.0%, and 13.0% were resistant to tetracycline, doxycycline, clindamycin, pirlimycin, and erythromycin, respectively. Thirty-nine strains were phenotypically resistant to two or more tested antibiotics. We identified a plasmid associated with macrolide and lincosamide resistance in 12% of the strains. Conclusion. The emergence of S. uberis strains resistant to multiple antibiotics highlights the importance of S. uberis surveillance and the prudent use of antimicrobials.

Vibrio cholerae is a Gram-negative bacterium found in aquatic environments and is the aetiological agent of cholera, characterized by acute watery diarrhoea and severe dehydration. Cholera presents a significant global health burden of an estimated 1.3-5 million annual cases, with the current pandemic caused by a toxigenic lineage of the O1 El Tor biotype called seventh pandemic El Tor (7PET) that is still ongoing. Whilst it is known that non-7PET lineages can cause sporadic disease, little is known about the transmission of these non-epidemic lineages. Thirty-four V. cholerae isolates were obtained from travellers returning from Indonesia to Australia between 2005 and 2017. These were whole genome sequenced, placed into a global phylogenetic context with 883 isolates, and screened for known genes associated with antimicrobial resistance and virulence. This analysis revealed that 30 isolates fell within non-7PET lineages and four within the 7PET lineage. Both 7PET and non-7PET isolates carried genes for resistance to antibiotics that are commonly used in cholera treatment such as tetracyclines and fluoroquinolones. Diverse virulence factors were also present in non-7PET isolates, with two isolates notably carrying toxin-coregulated pilus genes, which are primarily responsible for intestinal colonization in 7PET V. cholerae . This study demonstrates the role of travel in long-range carriage of epidemic and non-epidemic lineages of V. cholerae, and how sentinel travel surveillance can enrich our knowledge of V. cholerae diversity, reveal new biology about the spread of diverse lineages with differing disease potential and illuminate disease presence in endemic regions with limited surveillance data.

Acinetobacter baumannii is a globally distributed opportunistic pathogen in human health settings, including in intensive care units (ICUs). We investigated the contamination of a French small animal ICU with A. baumannii . We discovered repeated animal contamination by A. baumannii , and phylogenetic analysis traced contamination back to a potential foreign animal origin. Genomic analysis combined with antibiotic susceptibility testing revealed heteroresistance to penicillin and aminoglycoside mediated by insertion sequence dynamics and also suggest a potential cross-resistance to human-restricted piperacillin-tazobactam combination. The A. baumannii isolates of the animal ICU belong to the International Clone 2 commonly found in human health settings. Our results suggest a high adaptation of this lineage to healthcare settings and provide questions on the requirements for genetic determinants enabling adaptation to host and abiotic conditions.

Introduction. Reports of β-lactamase-producing Haemophilus influenzae are increasing worldwide. Aim . This study aimed to elucidate the molecular characteristics and evolution of β-lactamase-producing H. influenzae . Methodology. A total of 159 clinical isolates were characterized using multi-locus sequence typing. Antimicrobial resistance genes and integrative and conjugative element (ICE) types were identified through PCR and DNA sequencing. The genetic structure of ICE was further investigated using whole-genome sequencing. Results. Out of 159 clinical isolates, 20.8% ( n =33) were β-lactamase producers. Thirteen sequence types (STs) were identified. ST 103, 155, 165 and 388 have been identified in previous studies, suggesting that strains with these STs tend to acquire the β-lactamase gene bla TEM-1 . Among β-lactamase producers, 66.7% ( n =22) of bla TEM-1 were located on ICE. The ICEs could be classified into two groups based on their sequence (types I and II). Among these strains, 2017-Y3 harboured a macrolide resistance gene, mef (A/E) , in ICE. A comparative analysis of the ICE region of this strain and those from other countries suggested that each isolate was derived from ICE type I or II. These regions, including mef (A/E ), were similar to those of Tn 6822 , which is commonly found in Streptococcus . Conclusions. This study revealed several STs associated with the acquisition of β-lactamase genes on ICEs. Additionally, ICE evolution involved the acquisition of exogenous genes. The accumulation of resistance genes in ICE raises concerns regarding the emergence of multidrug-resistant H. influenzae .

Carbapenem-resistant Klebsiella pneumoniae (CRKP) has recently emerged as a notable public health concern, while the underlying drivers of CRKP transmission among patients across different healthcare facilities have not been fully elucidated. To explore the transmission dynamics of CRKP, 45 isolates were collected from both the intensive care unit (ICU) and non-ICU facilities in a teaching hospital in Guangdong, China, from March 2020 to August 2023. The collection of clinical data and antimicrobial resistance phenotypes was conducted, followed by genomic data analysis for these isolates. The mean age of the patients was 75.2 years, with 18 patients (40.0%) admitted to the ICU. The predominant strain in hospital-acquired CRKP was sequence type 11 (ST11), with k-locus type 64 and serotype O1/O2v1 (KL64:O1/O2v1), accounting for 95.6% (43/45) of the cases. The CRKP ST11 isolates from the ICU exhibited a low single nucleotide polymorphism (SNP) distance when compared to isolates from other departments. Genome-wide association studies identified 17 genes strongly associated with SNPs that distinguish CRKP ST11 isolates from those in the ICU and other departments. Temporal transmission analysis revealed that all CRKP isolates from other departments were genetically very close to those from the ICU, with fewer than 16 SNP differences. To further elucidate the transmission routes among departments within the hospital, we reconstructed detailed patient-to-patient transmission pathways using hybrid methods that combine TransPhylo with an SNP-based algorithm. A clear transmission route, along with mutations in potential key genes, was deduced from genomic data coupled with clinical information in this study, providing insights into CRKP transmission dynamics in healthcare settings.

Pseudonocardia species comprise a genus of filamentous, sporulating bacteria belonging to the phylum Actinomycetota, formerly Actinobacteria. They are found in marine and freshwater sediments and soils and associated with marine animals, insects, and plants. To date, they have mostly been studied because of their mutually beneficial symbiosis with fungus-growing ants in the tribe Attini . They have also attracted interest due to their biosynthetic capabilities, including the production of variably glycosylated polyenes and other novel antifungal compounds, and for their capacity to grow on a variety of hydrocarbons. The majority of clinically used antibiotics are derived from the specialised metabolites of filamentous actinomycete bacteria and most of these come from the genus Streptomyces . However, in the quest for novel chemistry there is increasing interest in studying other filamentous actinomycete genera, including Pseudonocardia . Here we outline the biological properties, genome size and structure and key features of the genus Pseudonocardia , namely their specialised metabolites and ecological roles.

One bacterial strain, designated as C22-A2 T , was isolated from Lake LungmuCo in Tibet. Cells of strain C22-A2 T were long rod-shaped, Gram-stain-negative, non-spore-forming, with positive catalase and oxidase activity. Optimal growth occurred at 20-25 °C, pH 8.0 and with 3.0-7.0% (w/v) NaCl. Phylogenetic analysis of 16S rRNA gene and whole genome sequences revealed that strain C22-A2 T belonged to the genus Virgibacillus , showing the highest 16S rRNA gene similarity to Virgibacillus halodenitrificans DSM 10037 T (97.6%). The average nucleotide identity values between strain C22-A2 T and the type strains of related species in the genus Virgibacillus were less than 74.4% and the digital DNA-DNA hybridization values were less than 20.2%, both below the species delineation thresholds of 95 and 70% respectively. The genome analysis revealed that strain C22-A2 T harboured genes responsible for osmotic and oxidative stress, enabling it to adapt to its surrounding environment. In terms of biochemical and physiological characteristics, strain C22-A2 T shared similar characteristics with the genus Virgibacillus, including the predominant cellular fatty acid anteiso-C 15 : 0 , the major respiratory quinone MK-7, as well as the polar lipids phosphatidylglycerol and diphosphatidylglycerol. Based on the comprehensive analysis of phylogenetic, phylogenomic, morphological, physiological and biochemical characteristics, strain C22-A2 T is proposed to represent a novel species of the genus Virgibacillus , named as Virgibacillus tibetensis sp. nov. (=CGMCC 1.19202 T =KCTC 43426 T ).

Next-generation sequencing methods have become essential for studying bacterial biology and pathogenesis, often depending on high-quality, closed genomes. In this study, we utilized a hybrid sequencing approach to assemble the genome of C6706, a widely used Vibrio cholerae model strain. We present a manually curated annotation of the genome, enhancing user accessibility by linking each coding sequence to its counterpart in N16961, the first sequenced V. cholerae isolate and a commonly used reference genome. Comparative genomic analysis between V. cholerae C6706 and N16961 uncovered multiple genetic differences in genes associated with key biological functions. To determine whether these genetic variations result in phenotypic differences, we compared several phenotypes relevant to V. cholerae pathogenicity like genetic stability, acid sensitivity, biofilm formation and motility. Notably, V. cholerae N16961 exhibited greater motility and reduced biofilm formation compared to V. cholerae C6706. These phenotypic differences appear to be mediated by variations in quorum sensing and cyclic di-GMP signalling pathways between the strains. This study provides valuable insights into the regulation of biofilm formation and motility in V. cholerae .

Klebsiella pneumoniae is a Gram-negative bacterium that causes both community- and healthcare-associated infections. Although various virulence factors and highly pathogenic phenotypes have been reported, the pathogenicity of K. pneumoniae is still not fully understood. In this study, we utilized whole-genome sequencing data of 168 clinical K. pneumoniae strains to assess pathogenicity. This work was based on the concept that the genetic composition of individual genomes (referred to as holistic gene content) of the strains may contribute to their pathogenicity. Holistic gene content analysis revealed two distinct groups of K. pneumoniae strains ('major group' and 'minor group'). The minor group included strains with known highly pathogenic clones (ST23, ST375, ST65 and ST86). The minor group had higher rates of capsular genotype K1 and presence of nine specific virulence genes ( rmpA , iucA , iutA , irp2 , fyuA , ybtS , iroN , allS and clbA ) compared to the major group. Pathogenicity was assessed using Galleria mellonella larvae. Infection experiments revealed lower survival rates of larvae infected with strains from the minor group, indicating higher virulence. In addition, the minor group had a higher string test positivity rate than the major group. Holistic gene content analysis predicted possession of virulence genes, string test positivity and pathogenicity as observed in the G. mellonella infection model. Moreover, the findings suggested the presence of as yet unrecognized genomic elements that are either involved in the acquisition of virulence genes or associated with pathogenicity.

The pathovar-based taxonomy of the Xanthomonas translucens group is very confusing due to an overlap of plant host ranges and level of host specificity. Here, whole-genome sequence-based parameters (digital DNA-DNA hybridization and blast-based average nucleotide identity), phylogenomic, biochemical and phenotypical data were used to taxonomically analyse the 11 known pathovars of the X. translucens complex. This polyphasic approach taxonomically assigned the 11 pathovars of X. translucens complex into three distinct species, two of which are new: X. translucens , X. cerealis sp. nov. and X. graminis sp. nov. X. translucens consists of three pathovars: pv. translucens (=pv. hordei ), pv. pistaciae strain A ICMP 16316 PT and pv. undulosa (=pv. secalis ). X. cerealis sp. nov. encompasses the pv. cerealis strain LMG 679 PT and pv. pistaciae strain B ICMP 16317 PT with genome similarity of 92.7% (dDDH) and 99.0% (ANIb) suggesting taxonomically similar genotypes. The other new species, X. graminis sp. nov., consists of the remaining five designated pathovars (pv. graminis , pv. arrhenatheri, pv. poae , pv. phleipratensis and pv. phlei ) with highly variable dDDH and ANIb values ranging from 74.5 to 93.0% and from 96.7 to 99.2%, respectively, an indication of a very divergent taxonomic group. Only strains of pvs. phlei and phleipratensis showed the highest genomic similarities of 93.0% (dDDH) and 99.2% (ANIb), suggesting synonymic pathovars as both infect the same plant hosts. The dDDH and ANI data were corroborated by phylogenomics clustering. The fatty acid contents were similar but the type strain of X. graminis sp. nov. exhibited 20% less C 15 : 0 iso and 40% more C 17 : 0 iso fatty acids than the other species. Based on phenotypic, biochemical and whole-genome sequence data, we propose two new species, Xanthomonas cerealis sp. nov. and Xanthomonas graminis sp. nov. with type strains LMG 679 T (=NCPPB 1944 T ) and LMG 726 T (=NCPPB 2700 T ), respectively.

The family Anoxybacillaceae was recently proposed encompassing the genera Anoxybacillus , Geobacillus , Parageobacillus , Saccharococcus and Thermolongibacillus . Of these genera, Anoxybacillus contains >50% of the Anoxybacillaceae species. However, Anoxybacillus species form multiple unrelated clades in phylogenetic trees and their evolutionary relationships are unclear. To clarify the evolutionary relationships of Anoxybacillus and other Anoxybacillaceae species, detailed phylogenomic and comparative analyses were conducted on 38 Anoxybacillaceae species with available genomes. In a phylogenomic tree based on 1148 core proteins, all Anoxybacillus , Geobacillus , Parageobacillus , Saccharococcus and Thermolongibacillus species, excepting Anoxybacillus sediminis , formed a strongly supported clade representing the family Anoxybacillaceae . Five conserved signature indels (CSIs) reported here are also uniquely found in these species, providing robust means for the demarcation of family Anoxybacillaceae in molecular terms. In our phylogenomic tree and in the Genomic Taxonomy Database, Anoxybacillus species formed four distinct clades designated as Anoxybacillus sensu stricto (containing the type species A. pushchinoensis ), Anoxybacillus &#95;A, Anoxybacillus &#95;B and Anoxybacillus &#95;C. Our analyses have identified 17 novel CSIs which offer means to reliably distinguish species from these clades based upon multiple uniquely shared molecular characteristics. Additionally, we have identified three and seven CSIs specific for the genera Geobacillus and Brevibacillus , respectively. All seven Brevibacillus -specific CSIs are also shared by Anoxybacillus sediminis , which branches reliably with this genus. Based on the strong phylogenetic and molecular evidence presented here, we are proposing that the genus Anoxybacillus should be restricted to only the species from Anoxybacillus sensu stricto clade, whereas the species from Anoxybacillus &#95;A, Anoxybacillus &#95;B, and Anoxybacillus &#95;C clades should be transferred into three novel genera Anoxybacteroides gen. nov., Paranoxybacillus gen. nov. and Thermaerobacillus gen. nov., respectively. Additionally, we are also proposing the transfer of Anoxybacillus sediminis to the genus Brevibacillus . The proposed changes, which reliably depict the evolutionary relationships among Anoxybacillaceae species, should be helpful in the studies of these organisms.

Six bacterial strains, Mut1 T , Mut2, Alt1, Alt2, Alt3 T , and Alt4, were isolated from soil samples collected in parks in Gothenburg, Sweden, based on their ability to utilize the insoluble polysaccharides α-1,3-glucan (mutan; Mut strains) or the mixed-linkage α-1,3/α-1,6-glucan (alternan; Alt strains). Analysis of 16S rRNA gene sequences identified all strains as members of the genus Streptomyces . The genomes of the strains were sequenced and subsequent phylogenetic analyses identified Mut2 as a strain of Streptomyces laculatispora and Alt1, Alt2 and Alt4 as strains of Streptomyces poriferorum, while Mut1 T and Alt3 T were most closely related to the type strains Streptomyces drozdowiczii NBRC 101007 T and Streptomyces atroolivaceus NRRL ISP-5137 T , respectively. Comprehensive genomic and biochemical characterizations were conducted, highlighting typical features of Streptomyces , such as large genomes (8.0-9.6 Mb) with high G+C content (70.5-72.0%). All six strains also encode a wide repertoire of putative carbohydrate-active enzymes, indicating a capability to utilize various complex polysaccharides as carbon sources such as starch, mutan, and cellulose, which was confirmed experimentally. Based on phylogenetic and phenotypic characterization, our study suggests that strains Mut1 T and Alt3 T represent novel species in the genus Streptomyces for which the names Streptomyces castrisilvae sp. nov. and Streptomyces glycanivorans sp. nov. are proposed, with strains Mut1 T (=DSM 117248 T =CCUG 77596 T ) and Alt3 T (=DSM 117252 T =CCUG 77600 T ) representing the respective type strains.

Two-novel filamentous actinobacteria designated strains 2-2 T and 2-15 T were isolated from soil of a coal mining site in Mongolia, and their taxonomic positions were determined using a polyphasic approach. Phylogenetic analyses based on 16S rRNA gene sequences showed that each of the strains formed a distinct clade within the genus Amycolatopsis . The 16S rRNA gene sequence similarity analysis showed that both strains were mostly related to Amycolatopsis rhabdoformis NCIMB 14900 T with 99.0 and 99.4% sequence similarity, respectively. The genome-based comparison indicated that strain 2-2 T shared the highest digital DNA-DNA hybridization value of 35.6% and average nucleotide identity value of 86.9% with Amycolatopsis pretoriensis DSM 44654 T , and strain 2-15 T shared the corresponding values of 36.5 and 87.9% with A. rhabdoformis NCIMB 14900 T , all of which being well below the thresholds for species delineation. The chemotaxonomic properties of both strains were typical of the genus Amycolatopsis . In silico prediction of chemotaxonomic markers was also carried out, and the results were consistent with the chemotaxonomic profiles of the genus. Genome mining for secondary metabolite production in strains 2-2 T and 2-15 T revealed the presence of 29 and 24 biosynthetic gene clusters involved in the production of polyketide synthase, non-ribosomal peptide synthetase, ribosomally synthesized and post-translationally modified peptides, lanthipeptide, terpenes, siderophore, and a number of other unknown type compounds. Both strains showed broad antifungal activity against several filamentous fungi and also antibacterial activity against methicillin-resistant Staphylococcus aureus and Acinetobacter baumannii . The phenotypic, biochemical, and chemotaxonomic properties indicated that both strains could be clearly distinguished from other species of Amycolatopsis , and thus the names Amycolatopsis nalaikhensis sp. nov. (type strain, 2-2 T =KCTC 29695 T =JCM 30462 T ) and Amycolatopsis carbonis (type strain, 2-15 T =KCTC 39525 T =JCM 30563 T ) are proposed accordingly.

Two novel actinomycetal strains, designated CC-R113 T and CC-R104 T , were isolated from the tissues of two macroalgae collected on the northern Portuguese coast. Phylogenetic analyses based on the 16S rRNA gene showed that strain CT-R113 T belongs to the genus Nocardiopsis , being closely related to Nocardiopsis umidischolae 66/93 T and Nocardiopsis tropica VKM Ac-1457 T , with 98.65 and 98.39 % sequence similarity, respectively. The clade formed between the three type strains was confirmed by phylogenomic analysis. The genome of strain CT-R113 T was 7.27 Mb in size with a G+C content of 71.3 mol %, with average nucleotide identity (ANI) values of 89.59 and 90.14 % with strains 66/93 T and VKM Ac-1457 T , respectively. The major cellular fatty acids were identified as C 18 : 1  ω 9 c , iso-C 16 : 0 and anteiso-C 17 : 0 . Menaquinone 10 (MK-10) was the major respiratory quinone. Comparative analysis of 16S rRNA gene sequences showed that strain CC-R104 T belongs to the genus Rhodococcus and is most closely related to Rhodococcus pyridinivorans DSM 44555 T , with 98.24 % sequence similarity. However, phylogenomic analysis revealed that strain CC-R104 T establishes a clade with Rhodococcus artemisae DSM 45380 T , being more distant from Rhodococcus pyridinivorans DSM 44555 T . The genome of strain CC-R104 T was 5.34 Mb in size with a G+C content of 67.01 mol%. The ANI value between strains CC-R104 T and DSM 45380 T was 81.2 % and between strains CC-R104 T and DSM 44555 T was 81.5 %. The major cellular fatty acids were identified as C 18 : 1  ω 9 c , C 16 : 0 and summed feature 3. Menaquinone 8 (MK-8) was the only respiratory quinone. For both CC-R113 T and CC-R104 T , optimum growth was observed at pH 7.0, 28 °C and 0-5 % NaCl and whole-cell hydrolysates contained meso -diaminopimelic acid as the cell-wall diamino acid. On the basis of phenotypic, molecular and chemotaxonomic characteristics, strains CT-R113 T and CC-R104 T are considered to represent novel species, for which the names Nocardiopsis codii sp. nov. (type strain CT-R113 T =LMG33234 T =UCCCB172 T ) and Rhodococcus chondri sp. nov. (type strain CC-R104 T =LMG33233 T =UCCCB171 T ) are proposed.

Streptomyces is among the most extensively studied genera of bacteria but its complex taxonomy remains contested and is suspected to contain significant species-level misclassification. Resolving the classification of Streptomyces would benefit many areas of applied microbiology that rely on an accurate ground truth for grouping of related organisms, including comparative genomics-based searches for novel antimicrobials. We survey taxonomic conflicts between 16S rRNA and whole genome-based Streptomyces classifications using 2276 publicly available Streptomyces genome assemblies and 48 981 publicly available full-length 16S rRNA Streptomyces sequences from silva, Greengenes, Ribosomal Database Project (RDP), and NCBI (National Centre for Biotechnology Information) databases. We construct a full-length 16S gene tree for 14 239 distinct Streptomyces sequences that resolves three major lineages of Streptomyces , but whose topology is not consistent with existing taxonomic assignments. We use these sequence data to delineate 16S and whole genome landscapes for Streptomyces , demonstrating that 16S and whole-genome classifications are frequently in disagreement, and that 16S zero-radius Operational Taxonomic Units (zOTUs) are often inconsistent with Average Nucleotide Identity (ANI)-based taxonomy. Our results strongly imply that 16S rRNA sequence data does not map to taxonomy sufficiently well to delineate Streptomyces species routinely. We propose that alternative marker sequences should be adopted by the community for classification and metabarcoding. Insofar as Streptomyces taxonomy has been determined or supported by 16S sequence data and may in parts be in error, we also propose that reclassification of the genus by alternative approaches may benefit the Streptomyces community.

The 16S rRNA gene of Thermobacterium salinum TK19130 T had the highest sequence similarity to that of Luteirhabdus pelagi A3-108 T (99.7%). Phylogeny of 16S rRNA gene and whole genome sequences indicated that T. salinum TK19130 T and L. pelagi A3-108 T are closely related, and represented an independent clade. Whole genome comparisons showed that T. salinum TK19130 T and L. pelagi A3-108 T shared average amino acid identity of 95.3%, indicating they could be merged into the same genus. The digital DNA-DNA hybridization and average nucleotide identity values between T. salinum TK19130 T and L. pelagi A3-108 T were 52.5 and 93.3%, respectively. These values were below the recommended threshold values of prokaryotic species delineation. Thus, based on the principle of priority, we proposed the transfer of Thermobacterium salinum Chen et al . 2023 to the genus Luteirhabdus Ren et al . 2022 as Luteirhabdus salina comb. nov.

Campylobacter is the leading bacterial cause of infectious intestinal disease, but the pathogen typically accounts for a very small proportion of the overall stool microbiome in each patient. Diagnosis is even more difficult due to the fastidious nature of Campylobacter in the laboratory setting. This has, in part, driven a change in recent years, from culture-based to rapid PCR-based diagnostic assays which have improved diagnostic detection, whilst creating a knowledge gap in our clinical and epidemiological understanding of Campylobacter genotypes - no isolates to sequence. In this study, direct metagenomic sequencing approaches were used to assess the possibility of replacing genome sequences with metagenome sequences; metagenomic sequencing outputs were used to describe clinically relevant attributes of Campylobacter genotypes. A total of 37 diarrhoeal stool samples with Campylobacter and five samples with an unknown pathogen result were collected and processed with and without filtration, DNA was extracted, and metagenomes were sequenced by short-read sequencing. Culture-based methods were used to validate Campylobacter metagenome-derived genome (MDG) results. Sequence output metrics were assessed for Campylobacter genome quality and accuracy of characterization. Of the 42 samples passing quality checks for analysis, identification of Campylobacter to the genus and species level was dependent on Campylobacter genome read count, coverage and genome completeness. A total of 65% (24/37) of samples were reliably identified to the genus level through Campylobacter MDG, 73% (27/37) by culture and 97% (36/37) by qPCR. The Campylobacter genomes with a genome completeness of over 60% ( n =21) were all accurately identified at the species level (100%). Of those, 72% (15/21) were identified to sequence types (STs), and 95% (20/21) accurately identified antimicrobial resistance (AMR) gene determinants. Filtration of stool samples enhanced Campylobacter MDG recovery and genome quality metrics compared to the corresponding unfiltered samples, which improved the identification of STs and AMR profiles. The phylogenetic analysis in this study demonstrated the clustering of the metagenome-derived with culture-derived genomes and revealed the reliability of genomes from direct stool sequencing. Furthermore, Campylobacter genome spiking percentages ranging from 0 to 2% total metagenome abundance in the ONT MinION sequencer, configured to adaptive sequencing, exhibited better assembly quality and accurate identification of STs, particularly in the analysis of metagenomes containing 2 and 1% of Campylobacter jejuni genomes. Direct sequencing of Campylobacter from stool samples provides clinically relevant and epidemiologically important genomic information without the reliance on cultured genomes.

We aimed to elucidate the relationship between Kineococcus terrestris and Kineococcus aureolus through whole-genome-based analysis. The genome-derived 16S rRNA gene sequences of K. terrestris KCTC 39738 T and K. aureolus KCTC 39739 T shared a 100% similarity. Phylogenetic trees based on 16S rRNA gene and whole-genome sequences revealed that K. terrestris KCTC 39738 T and K. aureolus KCTC 39739 T formed a robust clade, indicating a close relationship between them. Genomic comparison showed that the two strains shared 99.1% average nucleotide identity, 92.0% digital DNA-DNA hybridization and 98.9% average amino acid identity values, all of which exceeded the recommended threshold values for species classification. Most phenotypic characteristics between the two species were almost identical. Based on the above evidence, we propose the reclassification of Kineococcus aureolus Xu et al . 2017 as a later heterotypic synonym of Kineococcus terrestris Xu et al . 2017. Since these two species were proposed in the same article, the principle of priority does not apply. Our proposal is supported by the fact that the nomenclatural authorities first described Kineococcus terrestris .

In this opinion piece, we consider the meaning of the term 'wild type' in the context of microbiology. This is especially pertinent in the post-genomic era, where we have a greater awareness of species diversity than ever before. Genomic heterogeneity, in vitro evolution/selection pressures, definition of 'the wild', the size and importance of the pan-genome, gene-gene interactions (epistasis), and the nature of the 'wild-type gene' are all discussed. We conclude that wild type is an outdated and even misleading phrase that should be gradually phased out.

Mycoplasma ovipneumoniae is associated with respiratory disease in wild and domestic Caprinae globally, with wide variation in disease outcomes within and between host species. To gain insight into phylogenetic structure and mechanisms of pathogenicity for this bacterial species, we compared M. ovipneumoniae genomes for 99 samples from 6 countries (Australia, Bosnia and Herzegovina, Brazil, China, France and USA) and 4 host species (domestic sheep, domestic goats, bighorn sheep and caribou). Core genome sequences of M. ovipneumoniae assemblies from domestic sheep and goats fell into two well-supported phylogenetic clades that are divergent enough to be considered different bacterial species, consistent with each of these two clades having an evolutionary origin in separate host species. Genome assemblies from bighorn sheep and caribou also fell within these two clades, indicating multiple spillover events, most commonly from domestic sheep. Pangenome analysis indicated a high percentage (91.4 %) of accessory genes (i.e. genes found only in a subset of assemblies) compared to core genes (i.e. genes found in all assemblies), potentially indicating a propensity for this pathogen to adapt to within-host conditions. In addition, many genes related to carbon metabolism, which is a virulence factor for Mycoplasmas, showed evidence for homologous recombination, a potential signature of adaptation. The presence or absence of annotated genes was very similar between sheep and goat clades, with only two annotated genes significantly clade-associated. However, three M. ovipneumoniae genome assemblies from asymptomatic caribou in Alaska formed a highly divergent subclade within the sheep clade that lacked 23 annotated genes compared to other assemblies, and many of these genes had functions related to carbon metabolism. Overall, our results suggest that adaptation of M. ovipneumoniae has involved evolution of carbon metabolism pathways and virulence mechanisms related to those pathways. The genes involved in these pathways, along with other genes identified as potentially involved in virulence in this study, are potential targets for future investigation into a possible genomic basis for the high variation observed in disease outcomes within and between wild and domestic host species.

Introduction. Commensal Neisseria spp. are highly prevalent in the oropharynx as part of the healthy microbiome. N. meningitidis can colonise the oropharynx too from where it can cause invasive meningococcal disease. To identify N. meningitidis , clinical microbiology laboratories often rely on Matrix Assisted Laser Desorption/Ionisation Time of Flight Mass Spectrometry (MALDI-TOF MS). Hypothesis/Gap statement. N. meningitidis may be misidentified by MALDI-TOF MS. Aim. To conduct genomic surveillance of oropharyngeal Neisseria spp. in order to: (i) verify MALDI-TOF MS species identification, and (ii) characterize commensal Neisseria spp. genomes. Methodology. We analysed whole genome sequence (WGS) data from 119 Neisseria spp. isolates from a surveillance programme for oropharyngeal Neisseria spp. in Belgium. Different species identification methods were compared: (i) MALDI-TOF MS, (ii) Ribosomal Multilocus Sequence Typing (rMLST) and (iii) rplF gene species identification. WGS data were used to further characterize Neisseria species found with supplementary analyses of Neisseria cinerea genomes. Results. Based on genomic species identification, isolates from the oropharyngeal Neisseria surveilence study were composed of the following species: N. meningitidis ( n =23) , N. subflava ( n =61), N. mucosa ( n =15), N. oralis ( n =8), N. cinerea ( n =5), N. elongata ( n =3), N. lactamica ( n =2), N. bacilliformis ( n =1) and N. polysaccharea ( n =1). Of these 119 isolates, four isolates identified as N. meningitidis ( n =3) and N. subflava ( n =1) by MALDI-TOF MS , were determined to be N. polysaccharea ( n =1) , N. cinerea ( n =2) and N. mucosa ( n =1) by rMLST. Phylogenetic analyses revealed that N. cinerea isolates from the general population ( n =3, cluster one) were distinct from those obtained from men who have sex with men (MSM, n =2, cluster two). The latter contained genomes misidentified as N. meningitidis using MALDI-TOF MS. These two N. cinerea clusters persisted after the inclusion of published N. cinerea WGS ( n =42). Both N. cinerea clusters were further defined through pangenome and Average Nucleotide Identity (ANI) analyses. Conclusion. This study provides insights into the importance of genomic genus-wide Neisseria surveillance studies to improve the characterization and identification of the Neisseria genus.

Gene- and genome-based approaches were used to determine whether Vigna little leaf (ViLL) phytoplasma, which occurs in northern Australia, is a distinct ' Candidatus Phytoplasma' species. The ViLL 16S rRNA gene sequences exhibited the highest known similarity to species in the 16SrXXIX-A and 16SrIX-D subgroups, namely ' Candidatus Phytoplasma omanense' (98.03-98.10%) and ' Candidatus Phytoplasma phoenicium' (96.87-97.20%), respectively. A total of 48 single-copy orthologue genes were identified to be shared among the two draft ViLL phytoplasma genomes, 30 publicly available phytoplasma genomes, and one Acholeplasma laidlawii genome as the outgroup taxon. Phylogenomic assessments using the 48 shared single-copy orthologue genes supported that ViLL and ' Ca . Phytoplasma phoenicium' were closely related yet distinct species. The 16S rRNA gene sequence analysis and phylogenomic assessment indicate that ViLL phytoplasmas are a distinct taxon. As such, a novel species, ' Candidatus Phytoplasma vignae', is proposed. Strain BAWM-336 (genome accession number JAUZLI000000000) detected in Momordica charantia (bitter melon) serves as the reference strain of this species, with infected plant material deposited in the Victorian Plant Pathology Herbarium (VPRI) as VPRI 44369.

Four strains (MSK211, MSK294 T , MSK312, MSK433) of a novel Dolosigranulum species were cultured from nasopharyngeal swabs collected from mother-infant dyads in southern Botswana. These strains grew optimally on tryptic soy agar with 5% sheep blood solid medium and in fastidious bacteria broth. Colonies on tryptic soy agar with 5% sheep blood agar appeared grey or white with a flat, smooth surface and variable alpha haemolysis. Cells were Gram-positive, non-spore-forming, non-motile cocci that lacked catalase or oxidase activity. Major fatty acids were C 16 : 0 (palmitic acid), C 18 : 1  ω9 c (oleic acid), and C 18 : 0 (stearic acid). Analyses of 16S rRNA gene sequences identified these strains as belonging to the genus Dolosigranulum (family Carnobacteriaceae ), which currently contains only a single validly published species ( Dolosigranulum pigrum ). Whole-genome sequencing revealed that the genomes of these strains are 1.98-2.07 Mbp in size and have a G+C content of 39.6-39.9 mol%. Comparisons of these genomes to publicly available genomes of D. pigrum yielded average nucleotide identities and in silico DNA-DNA hybridization values of 92.3-92.9% and 49.1-51.4%, respectively. These results indicate that these strains represent a novel species of Dolosigranulum , for which we propose the name Dolosigranulum savutiense sp. nov., with the type strain MSK294 T (=DSM 117171 T =JCM 36673 T ).

The present study used whole-genome data to clarify the taxonomic assignment of two closely related Williamsia species. Genomic information for 10 type strains was available at the time of conducting this analysis. One group of type strains was found to be conspecific, namely Williamsia muralis Kämpfer et al . 1999 and Williamsia marianensis Pathom-aree et al . 2006. The 16S rRNA gene sequences showed 99 % similarity between these type strains. Whole-genome-based comparisons showed that Williamsia muralis DSM 44343 T and Williamsia marianensis DSM 44944 T shared 98.07 % average nucleotide identity, 98.29 % average amino acid identity and 84.80 % digital DNA-DNA hybridization values. These values exceeded the threshold values for bacterial species delineation. Further, phylogenomic analysis based on the core genomes of the strains under study confirmed that Williamsia muralis DSM 44343 T and Williamsia marianensis DSM 44944 T formed a monophyletic clade. Based on this evidence, we propose the reclassification of Williamsia marianensis Pathom-aree et al . 2006 as a later heterotypic synonym of Williamsia muralis Kämpfer et al . 1999.

A novel, Gram-positive, facultatively anaerobic, and non-motile bacterial strain, designated B2T-5 T , was isolated from jeotgal, a traditional Korean fermented seafood. Colonies grown on gifu anaerobic medium agar plates were cream-coloured, irregular, and umbonate with curled margins. Optimal growth of strain B2T-5 T occurred at 20 °C, pH 8.0, and in the presence of 1% (w/v) NaCl. Strain B2T-5 T was negative for oxidase and catalase activity. Hippurate was not hydrolysed and acetoin was not produced. The major cellular fatty acids were C 18 : 1  ω 9 c and C 16 : 0 . The cell-wall peptidoglycan was of the A4 α type containing l-Lys-d-Asp. The predominant respiratory quinone was menaquinone 7. The major polar lipids were diphosphatidylglycerol, phosphatidylglycerol, and phosphatidylcholine. According to the phylogenetic analysis based on 16S rRNA gene sequences, strain B2T-5 T was most closely related to Vagococcus teuberi DSM 21459 T , showing 98.2% sequence similarity. Genome sequencing of strain B2T-5 T revealed a genome size of 2.0 Mbp and a G+C content of 33.8 mol%. The average nucleotide identities of strain B2T-5 T with Vagococcus teuberi DSM 21459 T , Vagococcus bubulae SS1994 T , and Vagococcus martis D7T301 T were 75.0, 74.7, and 75.1%, respectively. Based on the phenotypic, chemotaxonomic, and genotypic data, strain B2T-5 T represents a novel species of the genus Vagococcus , for which the name Vagococcus jeotgali sp. nov. is proposed. The type strain is B2T-5 T (=KCTC 21223 T =JCM 35937 T ).

The genus Segeticoccus includes only one species with validly published name, Segeticoccus rhizosphaerae . The type strain Segeticoccus rhizosphaerae YJ01 T was isolated from soil in Korea, while Ornithinicoccus soli XNB-1 T was isolated from soil in China. Both strains share similar phenotypic and chemotaxonomic characteristics, including predominant menaquinone MK-8(H 4 ) and major polar lipids diphosphatidylglycerol and phosphatidylglycerol. Whole genome sequences revealed a DNA G+C content of 70.1 mol% for both strains and 100% similarity in their 16S rRNA gene sequences. Phylogenetic analysis showed they form a distinct cluster separate from other genera. Genomic comparisons showed average nucleotide identity and digital DNA-DNA hybridization values of 99.16 and 94.2%, respectively, indicating they represent a single species. Based on this genomic evidence, Ornithinicoccus soli Jiang et al . 2020 is proposed to be a later heterotypic synonym of Segeticoccus rhizosphaerae Lee and Whang 2020.

Defining the population structure of a pathogen is a key part of epidemiology, as genomically related isolates are likely to share key clinical features such as antimicrobial resistance profiles and invasiveness. Multiple different methods are currently used to cluster together closely related genomes, potentially leading to inconsistency between studies. Here, we use a global dataset of 26 306  Streptococcus pneumoniae genomes to compare four clustering methods: gene-by-gene seven-locus MLST, core genome MLST (cgMLST)-based hierarchical clustering (HierCC) assignments, life identification number (LIN) barcoding and k-mer-based PopPUNK clustering (known as GPSCs in this species). We compare the clustering results with phylogenetic and pan-genome analyses to assess their relationship with genome diversity and evolution, as we would expect a good clustering method to form a single monophyletic cluster that has high within-cluster similarity of genomic content. We show that the four methods are generally able to accurately reflect the population structure based on these metrics and that the methods were broadly consistent with each other. We investigated further to study the discrepancies in clusters. The greatest concordance was seen between LIN barcoding and HierCC (adjusted mutual information score=0.950), which was expected given that both methods utilize cgMLST, but have different methods for defining an individual cluster and different core genome schema. However, the existence of differences between the two methods shows that the selection of a core genome schema can introduce inconsistencies between studies. GPSC and HierCC assignments were also highly concordant (AMI=0.946), showing that k-mer-based methods which use the whole genome and do not require the careful selection of a core genome schema are just as effective at representing the population structure. Additionally, where there were differences in clustering between these methods, this could be explained by differences in the accessory genome that were not identified in cgMLST. We conclude that for S. pneumoniae , standardized and stable nomenclature is important as the number of genomes available expands. Furthermore, the research community should transition away from seven-locus MLST, whilst cgMLST, GPSC and LIN assignments should be used more widely. However, to allow for easy comparison between studies and to make previous literature relevant, the reporting of multiple clustering names should be standardized within the research.

Investigating the genomic epidemiology of major bacterial pathogens is integral to understanding transmission, evolution, colonization, disease, antimicrobial resistance and vaccine impact. Furthermore, the recent accumulation of large numbers of whole genome sequences for many bacterial species enhances the development of robust genome-wide typing schemes to define the overall bacterial population structure and lineages within it. Using the previously published data, we developed the Pneumococcal Genome Library (PGL), a curated dataset of 30 976 genomes and contextual data for carriage and disease pneumococci recovered between 1916 and 2018 in 82 countries. We leveraged the size and diversity of the PGL to develop a core genome multilocus sequence typing (cgMLST) scheme comprised of 1222 loci. Finally, using multilevel single-linkage clustering, we stratified pneumococci into hierarchical clusters based on allelic similarity thresholds and defined these with a taxonomic life identification number (LIN) barcoding system. The PGL, cgMLST scheme and LIN barcodes represent a high-quality genomic resource and fine-scale clustering approaches for the analysis of pneumococcal populations, which support the genomic epidemiology and surveillance of this leading global pathogen.

Two bacterial strains, SP1S1-4 T and SP2S1-2 T , were isolated from sediment samples collected in the Stockholm archipelago in November 2021. Following whole-genome sequencing, these strains were identified as tentatively belonging to two novel Shewanella genospecies, based on digital DNA-DNA hybridization, as implemented in the Type Strain Genome Server. Shewanella septentrionalis , Shewanella baltica and Shewanella hafniensis were, in this order and within a narrow genomic relatedness range, their closest genotypic relatives. Additional sampling and sequencing efforts led to the retrieval of distinct isolates that were monophyletic with SP1S1-4 T and SP2S1-2 T , respectively, based on phylogenomic analysis of whole-genome sequences. Comparative analyses of genome sequence data, which included blast-based average nucleotide identity, core genome-based and core proteome-based phylogenomics, in addition to MALDI-TOF MS-based protein profiling, confirmed the distinctness of the putative novel genospecies with respect to their closest genotypic relatives. A comprehensive phenotypic characterisation of SP1S1-4 T and SP2S1-2 T revealed only minor differences with respect to the type strains of S. septentrionalis , S. baltica and S. hafniensis . Based on the collective phylogenomic, proteomic, and phenotypic evidence presented here, we describe two novel genospecies within the genus Shewanella , for which the names Shewanella scandinavica sp. nov. and Shewanella vaxholmensis sp. nov. are proposed. The type strains are, respectively, SP2S1-2 T (=CCUG 76457 T =CECT 30688 T ), with a draft genome sequence of 5 041 805 bp and a G+C content of 46.3 mol%, and SP1S1-4 T (=CCUG 76453 T =CECT 30684 T ), with a draft genome sequence of 4 920147 bp and a G+C content of 46.0 mol%. Our findings suggest the existence of a species complex formed by the species S. baltica , S. septentrionalis , S. scandinavica sp. nov., and S. vaxholmensis sp. nov., with S. hafniensis falling in the periphery, where distinct genomic species clusters could be identified. However, this does not exclude the possibility of a continuum of genomic diversity within this sedimental ecosystem, as discussed herein with additional sequenced isolates.

Introduction. Klebsiella spp. are important bacteria that colonize the human intestine, especially in preterm infants; they can induce local and systemic disease under specific circumstances, including inflammatory bowel disease, necrotizing enterocolitis and colorectal cancer. Hypothesis. Klebsiella spp. colonized in the intestine of the neonates in the neonatal intensive care unit (NICU) may be associated with disease and antibiotic resistance, which will be hazardous to the children. Aim. Our aim was to know about the prevalence, antimicrobial resistance and genome characteristics of Klebsiella spp. in neonate carriers. Methodology. Genome sequencing and analysis, and antimicrobial susceptibility testing were mainly performed in this study. Results. The isolation rates of Klebsiella spp. strains were 3.7% (16/436) in 2014 and 4.3% (18/420) in 2021. Cases with intestinal-colonized Klebsiella spp. were mainly infants with low birth weights or those with pneumonia or hyperbilirubinemia. According to the core-pan genomic analysis, 34 stains showed gene polymorphism and a sequence type (ST) of an emerging high-risk clone (ST11). Eight strains (23.5%) were found to be resistant to 2 or more antibiotics, and 46 genes/gene families along with nine plasmids were identified that conferred resistance to antibiotics. In particular, the two strains were multidrug-resistant. Strain A1256 that is related to Klebsiella quasipneumoniae subsp. similipneumoniae was uncommon, carrying two plasmids similar to IncFII and IncX3 that included five antibiotic resistance genes. Conclusion. The prevention and control of neonatal Klebsiella spp. colonization in the NICU should be strengthened by paying increased attention to preventing antimicrobial resistance in neonates.

Genome sequencing of Clostridium clostridioforme strain LM41 revealed the presence of an atypically high proportion of mobile genetic elements for this species, with a particularly high abundance of prophages. Bioinformatic analysis of prophage sequences sought to characterize these elements and identify prophage-linked genes contributing to enhanced fitness of the host bacteria in the dysbiotic gut. Using PHASTER, PhageScope and manual curation, this work has identified 15 prophages: 4 predicted to be intact, 2 predicted to be defective and 9 which are unclassified. Quantitative PCR (qPCR) analysis revealed spontaneous release of four of the LM41 prophages (φ1, φ2, φ4 and φ10) into the culture supernatant, with virion-like particles visualized using transmission electron microscopy. The majority (12/14) of these particles had morphology akin to podoviruses, which is consistent with morphology predictions for φ1 and φ4. We observed diversity in the lysogeny mechanisms utilized by the prophages, with examples of the classical λ-like CI/Cro system, the ICE Bs 1 ImmR/ImmA-like system and the Mu-like C/Ner system. Classical morons, such as toxins or immune evasion factors, were not observed. We did, however, identify a variety of genes with roles in mediating restriction modification and genetic diversity, as well as some candidate genes with potential roles in host adaptation. Despite being the most abundant entities in the intestine, there is a dearth of information about phages associated with members of the microbiome. This work begins to shed light on the contribution of these elements to the lifestyle of C. clostridioforme LM41.

Two Gram-stain-negative, aerobic, rod-shaped, orange-coloured bacterial strains, designated strain C216 T and strain M2295, were isolated from mature mushroom compost from composting facilities in Victoria and South Australia, Australia, respectively. External structures such as flagella or pili were not observed on the cells under scanning electron microscopy. Optimal growth was found to occur at 45 °C, at pH 7.25 and in the absence of NaCl on Emerson's 350 YpSs medium. The genome sequence of strain C216 T was 3 342 126 bp long with a G+C content of 40.5 mol%. Functional analysis of the genome of strain C216 T revealed genes encoding chitinolytic and hemi-cellulolytic functions, with 166 predicted genes associated with carbohydrate metabolism (8.9% of the predicted genes). These functions are important for survival in the mushroom compost environment, which is rich in hemicelluloses. No antibiotic resistance genes were found in the genome sequence. The major fatty acids of strain C216 T were iso-C 15 : 0 (56.7%), iso-C 17 : 0 3-OH (15.6%), C 16 : 1  ω 7 c /iso-C 15 : 0 2-OH (7.3%) and iso-C 15 : 1 G (6.1%). The only respiratory quinone was MK-7. The major polar lipid of strain C216 T was phosphatidylethanolamine, but three unidentified phospholipids, four unidentified aminophospholipids/aminolipids and one unidentified glycolipid were also detected. Phylogenetic analysis based on proteins encoded by the core genome (bac120, 120 conserved bacterial genes) showed that strain C216 T forms a distinct lineage in the family Chitinophagaceae and that the closest identified relative is Niabella soli (69.69% ANI). These data demonstrate that strain C216 T represents a novel genus and novel species within the family Chitinophagaceae , for which we propose the name Mycovorax composti . The type strain is C216 T (=DSM 114558 T =LMG 32998 T ).

Haemophilus influenzae is part of the human nasopharyngeal microbiota and a pathogen causing invasive disease. The extensive genetic diversity observed in H. influenzae necessitates discriminatory analytical approaches to evaluate its population structure. This study developed a core genome multilocus sequence typing (cgMLST) scheme for H. influenzae using pangenome analysis tools and validated the cgMLST scheme using datasets consisting of complete reference genomes ( N = 14) and high-quality draft H. influenzae genomes ( N = 2297). The draft genome dataset was divided into a development dataset ( N = 921) and a validation dataset ( N = 1376). The development dataset was used to identify potential core genes, and the validation dataset was used to refine the final core gene list to ensure the reliability of the proposed cgMLST scheme. Functional classifications were made for all the resulting core genes. Phylogenetic analyses were performed using both allelic profiles and nucleotide sequence alignments of the core genome to test congruence, as assessed by Spearman's correlation and ordinary least square linear regression tests. Preliminary analyses using the development dataset identified 1067 core genes, which were refined to 1037 with the validation dataset. More than 70% of core genes were predicted to encode proteins essential for metabolism or genetic information processing. Phylogenetic and statistical analyses indicated that the core genome allelic profile accurately represented phylogenetic relatedness among the isolates ( R 2 = 0.945). We used this cgMLST scheme to define a high-resolution population structure for H. influenzae , which enhances the genomic analysis of this clinically relevant human pathogen.

Changing climates are allowing the geographic expansion of ticks and their animal hosts, increasing the risk of Borrelia -caused zoonoses in Canada. However, little is known about the genomic diversity of Borrelia from the west of the Canadian Rockies and from the tick vectors Ixodes pacificus , Ixodes auritulus and Ixodes angustus . Here, we report the whole-genome shotgun sequences of 51 Borrelia isolates from multiple tick species collected on a range of animal hosts between 1993 and 2016, located primarily in coastal British Columbia. The bacterial isolates represented three different species from the Lyme disease-causing Borrelia burgdorferi sensu lato genospecies complex [ Borrelia burgdorferi sensu stricto ( n =47), Borrelia americana ( n =3) and Borrelia bissettiae ( n =1)]. The traditional eight-gene multi-locus sequence typing (MLST) strategy was applied to facilitate comparisons across studies. This identified 13 known Borrelia sequence types (STs), established 6 new STs, and assigned 5 novel types to the nearest sequence types. B. burgdorferi s. s. isolates were further differentiated into ten ospC types, plus one novel ospC with less than 92 % nucleotide identity to all previously defined ospC types. The MLST types resampled over extended time periods belonged to previously described STs that are distributed across North America. The most geographically widespread ST, ST.12, was isolated from all three tick species. Conversely, new B. burgdorferi s. s. STs from Vancouver Island and the Vancouver region were only detected for short periods, revealing a surprising transience in space, time and host tick species, possibly due to displacement by longer-lived genotypes that expanded across North America.This article contains data hosted by Microreact.

Sphingopyxis lutea DHUNG17 T was compared with Sphingopyxis jiangsuensis XHP0097 T to examine the taxonomic relationship between the two type strains. The 16S rRNA gene sequence of S. lutea DHUNG17 T shared high similarity (99.9 %) to that of S. jiangsuensis XHP0097 T . The results of phylogenetic analyses based on 16S rRNA gene sequences indicated that the two strains formed a tight cluster within the genus Sphingopyxis . The average amino acid identity, average nucleotide identity and digital DNA-DNA hybridization values between S. lutea DHUNG17 T and S. jiangsuensis XHP0097 T were below 99.0, 99.1 and 92.2±1.7 %, respectively, all of which were greater than the species delineation threshold for AAI (95.5 %), ANI (95-96 %) and dDDH (70 %), strongly indicating that the two strains represented a single species. Based on the combined phylogenetic, genomic and phenotypic characterization presented here, we propose Sphingopyxis lutea as a later heterotypic synonym of Sphingopyxis jiangsuensis .

Introduction. Maternal screening tests and prophylactic antibiotics are important to prevent neonatal and infant group B streptococcal (GBS) infections. Hypothesis/Gap Statement. The performance of enrichment broth media for GBS screening that are available in Japan is unclear. Whole-genome data of GBS isolates from pregnant women in Japan is lacking. Aim. The aim of this study was to compare the protocol performance of six enrichment broths and two subculture agar plates, which were all available in Japan, for GBS detection. In addition, we showed whole-genome data of GBS isolates from pregnant women in Japan. Methodology. We collected 133 vaginal-rectal swabs from pregnant women visiting clinics and hospitals in Nagasaki Prefecture, Japan, and compared the protocol performance of 6 enrichment broths and 2 subculture agar plates. All GBS isolates collected in this study were subjected to whole-genome sequencing analysis. Results. We obtained 133 vaginal-rectal swabs from pregnant women at 35-37 weeks of gestation from 8 private clinics and 2 local municipal hospitals within Nagasaki Prefecture, Japan. The detection rate of the protocol involving the six enrichment broths and subsequent subcultures varied between 95.5 and 100 %, depending on the specific choice of enrichment broth. The GBS carriage rate among pregnant women in this region was 18.8 %. All 25 isolates derived from the swabs were susceptible to penicillin, whereas 48 and 36 % of the isolates demonstrated resistance to erythromycin and clindamycin, respectively. The distribution of serotypes was highly diverse, encompassing seven distinct serotypes among the isolates, with the predominant serotype being serotype V ( n = 8). Serotype V isolates displayed a tendency towards increased resistance to erythromycin and clindamycin, with all resistant isolates containing the ermB gene. Conclusion. There was no difference in performance among the culture protocols evaluated in this study. GBS strains isolated from pregnant women appeared to have greater genomic diversity than GBS strains detected in neonates/infants with invasive GBS infections. To confirm this result, further studies with larger sample sizes are needed.