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The valorization of lignocellulosic biomass, derived from various bio-waste materials, has received considerable attention as a sustainable approach to improve production chains while reducing environmental impact. Microbial enzymes have emerged as key players in the degradation of polysaccharides, offering versatile applications in biotechnology and industry. Among these enzymes, glycoside hydrolases (GHs) play a central role. Xylanases, in particular, are used in a wide range of applications and are essential for the production of xylose, which can be fermented into bioethanol or find use in many other industries. Currently, fungal secretomes dominate as the main reservoir of lignocellulolytic enzymes, but thermophilic microorganisms offer notable advantages in terms of enzyme stability and production efficiency. Here we present the genomic characterization of Geobacillus stearothermophilus GF16 to identify genes encoding putative enzymes involved in lignocellulose degradation. Thermostable GHs secreted by G. stearothermophilus GF16 were investigated and found to be active on different natural polysaccharides and synthetic substrates, revealing an array of inducible GH activities. In particular, the concentrated secretome possesses significant thermostable xylanase and β-xylosidase activities (5 ×10 3 U/L and 1.7 ×10 5 U/L, respectively), highlighting its potential for application in biomass valorization. We assessed the hemicellulose hydrolysis capabilities of various agri-food wastes using the concentrated secretome of the strain cultivated on xylan. An impressive 300-fold increase in xylose release compared to a commercially available cocktail was obtained with the secretome, underscoring the remarkable efficacy of this approach., Competing Interests: Declaration of Competing Interest All the authors declare no conflict of interests., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

This study investigated the safety characteristics and potential probiotic properties of Enterococcus faecium by using whole genome analysis, and then explored the effect of this strain on the virulence of Listeria monocytogenes in vitro and during the storage of fermented sausages. Results showed that E. faecium B1 presented enterocin A, B, and P, enterolysin A, and UviB, and the exotoxin related genes and exoenzyme related genes were not detected in the genome of E. faecium B1. However, the adherence genes including acm and scm were present in this strain, which also positively correlated with characteristics related to probiotic potential. In addition, E. faecium could adapt to the condition of fermented sausages, and decrease the survival of L. monocytogenes in vitro and in vivo. The expression of the virulence genes (prfA, hly, inlA, and inlB) and sigB-related genes (prli42, rsbT, rsbU, rsbV, rsbW, and sigB) were all inhibited by E. faecium B1 to different extents during the storage of fermented sausages at 4 °C. Moreover, compared with the E. faecium B1 group, the expression level of entA, entB, and entP genes of E. faecium B1 in the co-culture of fermented sausages was increased during the storage, which may be the inhibition mechanism of E. faecium B1 on L. monocytogenes. These results demonstrated that E. faecium B1 could potentially be used as bio-protection to control L. monocytogenes in meat products., Competing Interests: Declaration of competing interest The authors declare no known conflicts of interest (financial, professional, or personal)., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Exopolysaccharides (EPS) are natural macromolecular carbohydrates with good functional activity and physiological activities, which can be utilized as an emulsifier, viscosity enhancer, stabilizer, gelling agent, and water retention agent in a wide range of food products. In this study, the whole genome of Bacillus amyloliquefaciens D189, an EPS-producing bacteria, was sequenced. The result showed that D189 contains a single, circular chromosome of 3,963,356 bp with an average GC content of 45.74% and 3996 coding genes. The gene annotation results showed that D189 is a potentially safe strain and confirmed to be safe associated with hemolytic assay, and antibiotic resistance test. Meanwhile, D189 genome possessed 240 genes related to carbohydrate metabolism. More importantly, D189 could transport 9 sugars and contained a complete biosynthetic pathway for 8 nucleotide sugars. Based on the validation experiments, strain D189 could metabolize 8 sugars (glucose, sucrose, trehalose, fructose, cellobiose, maltose, mannitol, and N-acetyl-D-glucosamine) to produce EPS, with the highest yield of 1.212 g/L when sucrose was the carbon source. Therefore, the whole genome sequencing preliminarily elucidated the physiological mechanism of EPS, providing several pathways for engineering D189 to further enhance the yield of EPS., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Proteus mirabilis is a gram-negative pathogen that caused significant opportunistic infections. In this study we aimed to identify antimicrobial resistance (AMR) genes and virulence determinants in two pan-drug resistant isolate "Bacteria_11" and "Bacteria_27" using whole genome sequencing. Proteus mirabilis "Bacteria_11" and "Bacteria_27" were isolated from two different hospitalized patients in Egypt. Antimicrobial susceptibility determined using Vitek 2 system, then whole genome sequencing (WGS) using MinION nanopore sequencing was done. Antimicrobial resistant genes and virulence determinants were identified using ResFinder, CADR AMR database, Abricate tool and VF analyzer were used respectively. Multiple sequence alignment was performed using MAFFT and FastTree, respectively. All genes were present within bacterial chromosome and no plasmid was detected. "Bacteria_11" and "Bacteria_27" had sizes of approximately 4,128,657 bp and 4,120,646 bp respectively, with GC content of 39.15% and 39.09%. "Bacteria_11" and "Bacteria_27" harbored 43 and 42 antimicrobial resistance genes respectively with different resistance mechanisms, and up to 55 and 59 virulence genes respectively. Different resistance mechanisms were identified: antibiotic inactivation, antibiotic efflux, antibiotic target replacement, and antibiotic target change. We identified several genes associated with aminoglycoside resistance, sulfonamide resistance. trimethoprim resistance tetracycline resistance proteins. Also, those responsible for chloramphenicol resistance. For beta-lactam resistance, only blaVEB and blaCMY-2 genes were detected. Genome analysis revealed several virulence factors contribution in isolates pathogenicity and bacterial adaptation. As well as numerous typical secretion systems (TSSs) were present in the two isolates, including T6SS and T3SS. Whole genome sequencing of both isolates identify their genetic context of antimicrobial resistant genes and virulence determinants. This genomic analysis offers detailed representation of resistant mechanisms. Also, it clarifies P. mirabilis ability to acquire resistance and highlights the emergence of extensive drug resistant (XDR) and pan-drug resistant (PDR) strains. This may help in choosing the most appropriate antibiotic treatment and limiting broad spectrum antibiotic use., (© 2024. The Author(s).)

Carbapenem-resistant Acinetobacter baumannii (CRAB) poses a significant threat to hospitalized patients as effective therapeutic options are scarce. Based on the genomic characteristics of the CRAB strain AB2877 harboring chromosome-borne bla OXA-23 , which was isolated from the bronchoalveolar lavage fluid (BALF) of a patient in a respiratory intensive care unit (RICU), we systematically analyzed antibiotic resistance genes (ARGs) and the genetic context associated with ARGs carried by CRAB strains harboring chromosome-borne bla OXA-23 worldwide. Besides bla OXA-23 , other ARGs were detected on the chromosome of the CRAB strain AB2877 belonging to ST208/1806 (Oxford MLST scheme). Several key genetic contexts associated with the ARGs were identified on the chromosome of the CRAB strain AB2877, including (1) the MDR region associated with bla OXA-23 , tet(B)-tetR(B ), aph(3'')-Ib, and aph(6)-Id (2); the resistance island AbGRI3 harboring armA and mph(E)-msr(E ) (3); the Tn 3 -like composite transposon containing bla TEM-1D and aph(3')-Ia ; and (4) the structure "IS Aba1-bla ADC-25 ." The first two genetic contexts were most common in ST195/1816, followed by ST208/1806. The last two genetic contexts were found most frequently in ST208/1806, followed by ST195/1816.IMPORTANCEThe bla OXA-23 gene can be carried by plasmid or chromosome, facilitating horizontal genetic transfer and increasing carbapenem resistance in healthcare settings. In this study, we focused on the genomic characteristics of CRAB strains harboring the chromosome-borne bla OXA-23 gene, and the important genetic contexts associated with bla OXA-23 and other ARGs were identified, and their prevalent clones worldwide were determined. Notably, although the predominant clonal CRAB lineages worldwide containing the MDR region associated with bla OXA-23 , tet(B)-tetR(B ), aph(3'')-Ib, and aph (6)-Id was ST195/1816, followed by ST208/1806, the CRAB strain AB2877 in our study belonged to ST208/1806. Our findings contribute to the knowledge regarding the dissemination of CRAB strains and the control of nosocomial infection., Competing Interests: The authors declare no conflict of interest.

Objectives: The novel dual-target triazaacenaphthylene, gepotidacin, recently showed promising results in its Phase III randomized controlled trial for the treatment of gonorrhoea. We investigated alterations in the gepotidacin GyrA and ParC targets in gonococci by in silico mining of publicly available global genomes (n = 33 213) and determined gepotidacin MICs in isolates with GyrA A92 alterations combined with other GyrA and/or ParC alterations., Methods: We examined gonococcal gyrA and parC alleles available at the European Nucleotide Archive. MICs were determined using the agar dilution method (gepotidacin) or Etest (four antimicrobials). Models of DNA gyrase and topoisomerase IV were obtained from AlphaFold and used to model gepotidacin in the binding site., Results: GyrA A92 alterations were identified in 0.24% of genomes: GyrA A92P/S/V + S91F + D95Y/A/N (0.208%), A92P + S91F (0.024%) and A92P (0.003%), but no A92T (previously associated with gepotidacin resistance) was found. ParC D86 alterations were found in 10.6% of genomes: ParC D86N/G (10.5%), D86N + S87I (0.051%), D86N + S88P (0.012%) and D86G + E91G (0.003%). One isolate had GyrA A92P + ParC D86N alterations, but remained susceptible to gepotidacin (MIC = 0.125 mg/L). No GyrA plus ParC alterations resulted in a gepotidacin MIC > 4 mg/L. Modelling of gepotidacin binding to GyrA A92/A92T/A92P suggested that gepotidacin resistance due to GyrA A92T might be linked to the formation of a new polar contact with DNA., Conclusions: In silico mining of 33 213 global gonococcal genomes (isolates from 1928 to 2023) showed that A92 is highly conserved in GyrA, while alterations in D86 of ParC are common. No GyrA plus ParC alterations caused gepotidacin resistance. MIC determination and genomic surveillance of potential antimicrobial resistance determinants are imperative., (© The Author(s) 2024. Published by Oxford University Press on behalf of British Society for Antimicrobial Chemotherapy.)

Objectives: Escherichia coli can cause infections in the urinary tract and in normally sterile body sites leading to invasive E. coli disease (IED), including bacteraemia and sepsis, with older populations at increased risk. We aimed to estimate the theoretical coverage rate by the ExPEC4V and 9V vaccine candidates. In addition, we aimed at better understanding the diversity of E. coli isolates, including their genetic and phenotypic antimicrobial resistance (AMR), sequence types (STs), O-serotypes and the bacterial population structure., Methods: Blood and urine culture E. coli isolates (n = 304) were collected from hospitalized patients ≥60 years (n = 238) with IED during a multicentric, observational study across three continents. All isolates were tested for antimicrobial susceptibility, O-serotyped, whole-genome sequenced and bioinformatically analysed., Results: A large diversity of STs and of O-serotypes were identified across all centres, with O25b-ST131, O6-ST73 and O1-ST95 being the most prevalent types. A total of 45.4% and 64.7% of all isolates were found to have an O-serotype covered by the ExPEC4V and ExPEC9V vaccine candidates, respectively. The overall frequency of MDR was 37.4% and ST131 was predominant among MDR isolates. Low in-patient genetic variability was observed in cases where multiple isolates were collected from the same patient., Conclusions: Our results highlight the predominance of MDR O25b-ST131 E. coli isolates across diverse geographic areas. These findings provide further baseline data on the theoretical coverage of novel vaccines targeting E. coli associated with IED in older adults and their associated AMR levels., (© The Author(s) 2024. Published by Oxford University Press on behalf of British Society for Antimicrobial Chemotherapy.)

Objectives: Carbapenem-resistant Klebsiella pneumoniae (CRKP) is a major nosocomial infectious pathogen with rapidly increasing prevalence. The genomic epidemiological characteristics of CRKP nationwide, especially the evolving trends within the predominant clones, should be evaluated clearly., Methods: We collected 3415 K. pneumoniae strains from 28 hospitals across China. Antimicrobial susceptibility testing and WGS were performed. Subsequent genomic analyses, including sequence typing, K-locus (KL) identification, antimicrobial resistance gene screening, and virulence score assessment were performed. The phylogenetic relationship of clonal group 11 was determined based on core-genome analysis, and the presence of the pLVPK-like virulence plasmid in ST11 isolates was confirmed using plasmid core-gene analysis. Additionally, the trends of the ST11 lineage with different KL types on a global scale were investigated using Beast2., Results: Of the K. pneumoniae strains, 708 were identified as CRKP isolates (20.7%), of which 97.7% were MDR. ST11 was the predominant clone, and KPC-2 was the prevalent carbapenemase in China, although the prevalence of specific clones and carbapenemases varied by geographic region. Among ST11 isolates, KL47 and KL64 were the predominant KL types, and KL64 gradually replaced KL47, with a higher percentage of KL64 isolates harbouring the pLVPK-like plasmid. Global genome data showed a significant increase in the effective population size of KL64 over the last 5 years., Conclusions: The prevalence of CRKP was very high in certain regions in China. The increasing convergence of virulence and resistance, particularly in ST11-KL64 isolates, should be given more attention and further investigation., (© The Author(s) 2024. Published by Oxford University Press on behalf of British Society for Antimicrobial Chemotherapy. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Tuberculosis is a global public health concern. Earlier reports suggested the emergence of high rates of drug resistant tuberculosis in Egypt. This study included 102 isolates of Mycobacterium tuberculosis collected from two reference laboratories in Cairo and Alexandria. All clinical isolates were sub-cultured on Löwenstein-Jensen medium and analyzed using both BD BACTEC MGIT 960 SIRE Kit and standard diffusion disk assays to identify the antibiotic sensitivity profile. Extracted genomic DNA was subjected to whole genome sequencing (WGS) using Illumina platform. Isolates that belong to lineage 4 represented > 80%, while lineage 3 represented only 11% of the isolates. The percentage of drug resistance for the streptomycin, isoniazid, rifampicin and ethambutol were 31.0, 17.2, 19.5 and 20.7, respectively. Nearly 47.1% of the isolates were sensitive to the four anti-tuberculous drugs, while only one isolate was resistant to all four drugs. In addition, several new and known mutations were identified by WGS. High rates of drug resistance and new mutations were identified in our isolates. Tuberculosis control measures should focus on the spread of mono (S, I, R, E)- and double (S, E)-drug resistant strains present at higher rates throughout the whole Nile Delta, Egypt., (© 2024. The Author(s).)

Aims: Characterize global genomic features of 86 genomes of Salmonella Gallinarum (SG) and Pullorum (SP), which are important pathogens causing systemic infections in poultry., Methods and Results: All genomes harbored efflux pump encoding gene mdsA and gold tolerance genes golS and golT. Aminoglycoside (aac(6')-Ib, aadA5, aph(6)-Id, aph(3'')-Ib, ant(2'')-Ia), beta-lactam (blaTEM-1, blaTEM-135), efflux pump (mdsB), fosfomycin (fosA3), sulfonamide (sul1, sul2), tetracycline [tet(A)], trimethoprim (dfrA17), acid (asr), and disinfectant (qacEdelta1) resistance genes, gyrA, gyrB, and parC quinolone resistance point mutations, and mercury tolerance genes (mer) were found in different frequencies. Additionally, 310 virulence genes, pathogenicity islands (including SPI-1, 2, 3, 4, 5, 6, 9, 10, 12, 13, and 14), plasmids [IncFII(S), ColpVC, IncX1, IncN, IncX2, and IncC], and prophages (Fels-2, ST104, 500465-1, pro483, Gifsy-2, 103 203_sal5, Fels-1, RE-2010, vB_SenS-Ent2, and L-413C) were detected. MLST showed biovar-specific sequence types, and core genome MLST showed country-specific and global-related clusters., Conclusion: SG and SP global strains carry many virulence factors and important antimicrobial resistance genes. The diverse plasmids and prophages suggest genetic variability. MLST and cgMLST differentiated biovars and showed profiles occurring locally or worldwide., (© The Author(s) 2024. Published by Oxford University Press on behalf of Applied Microbiology International.)

Sponge microbiomes are often highly diverse making it difficult to determine which lineages are important for maintaining host health and homeostasis. Characterising genomic traits associated with symbiosis can improve our knowledge of which lineages have adapted to their host and what functions they might provide. Here we examined five microbial families associated with sponges that have previously shown evidence of cophylogeny, including Endozoicomonadaceae, Nitrosopumilaceae, Spirochaetaceae, Microtrichaceae and Thermoanaerobaculaceae, to better understand the mechanisms behind their symbiosis. We compared sponge-associated genomes to genomes found in other environments and found that sponge-specific clades were enriched in genes encoding many known mechanisms for symbiont survival, such as avoiding phagocytosis and defence against foreign genetic elements. We expand on previous knowledge to show that glycosyl hydrolases with sulfatases and sulfotransferases likely form multienzyme degradation pathways to break and remodel sulfated polysaccharides and reveal an enrichment in superoxide dismutase that may prevent damage from free oxygen radicals produced by the host. Finally, we identified novel traits in sponge-associated symbionts, such as urea metabolism in Spirochaetaceae which was previously shown to be rare in the phylum Spirochaetota. These results identify putative mechanisms by which symbionts have adapted to living in association with sponges., (© 2024 Commonwealth of Australia and The Author(s). Environmental Microbiology published by John Wiley & Sons Ltd.)

Motivation: Mobile genetic elements (MGEs) are as ubiquitous in nature as they are varied in type, ranging from viral insertions to transposons to incorporated plasmids. Horizontal transfer of MGEs across bacterial species may also pose a significant threat to global health due to their capability to harbor antibiotic resistance genes. However, despite cheap and rapid whole-genome sequencing, the varied nature of MGEs makes it difficult to fully characterize them, and existing methods for detecting MGEs often do not agree on what should count. In this manuscript, we first define and argue in favor of a divergence-based characterization of mobile-genetic elements. Results: Using that paradigm, we present skandiver, a tool designed to efficiently detect MGEs from whole-genome assemblies without the need for gene annotation or markers. skandiver determines mobile elements via genome fragmentation, average nucleotide identity (ANI), and divergence time. By building on the scalable skani software for ANI computation, skandiver can query hundreds of complete assemblies against >65 000 representative genomes in a few minutes and 19 GB memory, providing scalable and efficient method for elucidating mobile element profiles in incomplete, uncharacterized genomic sequences. For isolated and integrated large plasmids (>10 kb), skandiver's recall was 48% and 47%, MobileElementFinder was 59% and 17%, and geNomad was 86% and 32%, respectively. For isolated large plasmids, skandiver's recall (48%) is lower than state-of-the-art reference-based methods geNomad (86%) and MobileElementFinder (59%). However, skandiver achieves higher recall on integrated plasmids and, unlike other methods, without comparing against a curated database, making skandiver suitable for discovery of novel MGEs., Availability and Implementation: https://github.com/YoukaiFromAccounting/skandiver., (© The Author(s) 2024. Published by Oxford University Press.)

Plague is a deadly zoonosis that still poses a threat in many regions of the world. We combined epidemiologic, host, and vector surveillance data collected during 1961-1980 from the Araripe Plateau focus in northeastern Brazil with ecologic, geoclimatic, and Yersinia pestis genomic information to elucidate how these factors interplay in plague activity. We identified well-delimited plague hotspots showing elevated plague risk in low-altitude areas near the foothills of the plateau's concave sectors. Those locations exhibited distinct precipitation and vegetation coverage patterns compared with the surrounding areas. We noted a seasonal effect on plague activity, and human cases linearly correlated with precipitation and rodent and flea Y. pestis positivity rates. Genomic characterization of Y. pestis strains revealed a foundational strain capable of evolving into distinct genetic variants, each linked to temporally and spatially constrained plague outbreaks. These data could identify risk areas and improve surveillance in other plague foci within the Caatinga biome.

Bifidobacterium pseudocatenulatum is a prevalent gut microbe in humans of all ages and plays a crucial role in host health. However, its adaptive evolutionary characteristics remain poorly understood. This study analyzed the genome of 247 B. pseudocatenulatum isolates from Chinese, Vietnamese, Japanese and other region populations using population genomics and functional genomics. Our findings revealed high genetic heterogeneity and regional clustering within B. pseudocatenulatum isolates. Significant differences were observed in genome characteristics, phylogeny, and functional genes. Specifically, Chinese and Vietnamese isolates exhibited a higher abundance of genes involved in the metabolism of plant-derived carbohydrates (GH13, GH43, and GH5 enzyme families), aligning with the predominantly vegetable-, wheat- and fruit-based diets of these populations. Additionally, we found widespread transmission of antibiotic resistance genes (tetO and tetW) through mobile genetic elements, such as genomic islands (GIs), resulting in substantial intra-regional differences. Our findings highlight distinct adaptive evolution in B. pseudocatenulatum driven by gene specialization, possibly in response to regional variations in diet and lifestyle. This study sheds light on bifidobacteria colonization mechanisms in the host gut. IMPORTANCE: Gut microbiota, as a key link in the gut-brain axis, helps to maintain the health of the organism, among which, Bifidobacterium pseudocatenulatum (B. pseudocatenulatum) is an important constituent member of the gut microbiota, which plays an important role in maintaining the balance of gut microbiota. The probiotic properties of B. pseudocatenulatum have been widely elaborated, and in order to excavate its evolutionary features at the genomic level, here we focused on the genetic background and evolutionary mechanism of the B. pseudocatenulatum genomes isolated from the intestinal tracts of different populations. Ultimately, based on the phylogenetic tree, we found that B. pseudocatenulatum has high genetic diversity and regional clustering phenomenon, in which plant-derived carbohydrate metabolism genes (GH13, GH43, GH5) showed significant regional differences, and this genetic differentiation drove the adaptive evolution, which likely shaped by diet and lifestyle., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Ltd.)

A novel A. pittii J08 with heterotrophic nitrification and aerobic denitrification (HN-AD) isolated from pond sediments could rapidly degrade inorganic nitrogen (N) and total nitrogen (TN-N) with ammonium (NH 4 + -N) preference. N degradation rate of NH 4 + -N, nitrite (NO 2 - -N) and nitrate (NO 3 - -N) were 3.9 mgL -1 h -1 , 3.0 mgL -1 h -1 and 2.7 mgL -1 h -1 , respectively. In addition, strain J08 could effectively utilize most of detected low-molecular-weight carbon (LMWC) sources to degrade inorganic N with a wide adaptability to various culture conditions. Whole genome sequencing (WGS) analysis revealed that assembled genome of stain J08 possessed the crucial genes involved in dissimilatory/assimilatory NO 3 - -N reduction and NH 4 + -N assimilation. These results indicated that strain J08 could be applied to wastewater treatment in aquaculture., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The rising frequency of multidrug-resistant (MDR) Acinetobacter baumannii infections represents a significant public health challenge in Bangladesh. Genomic analysis of bacterial pathogens enhances surveillance and control efforts by providing insights into genetic diversity, antimicrobial resistance (AMR) profiles, and transmission dynamics. In this study, we conducted a comprehensive bioinformatic analysis of 82 whole-genome sequences (WGS) of A. baumannii from Bangladesh to understand their genomic epidemiological characteristics. WGS of the MDR and biofilm-forming A. baumannii strain S1C revealed the presence of 28 AMR genes, predicting its pathogenicity and classification within sequence type ST2. Multi-locus sequence typing (MLST) genotyping suggested heterogeneity in the distribution of clinical A. baumannii strains in Bangladesh, with a predominance of ST575. The resistome diversity was evident from the detection of 82 different AMR genes, with antibiotic inactivation being the most prevalent resistance mechanism. All strains were predicted to be multidrug-resistant. The observed virulence genes were associated with immune evasion, biofilm formation, adherence, nutrient acquisition, effector delivery, and other mechanisms. Mobile genetic elements carrying AMR genes were predicted in 68.29% (N = 56) of the genomes. The "open" state of the pan-genome and a high proportion of accessory genes highlighted the genome plasticity and diversity of A. baumannii in Bangladesh. Additionally, phylogenomic analysis indicated clustering of A. baumannii strains into three separate clades according to sequence type. In summary, our findings offer detailed insights into the genomic landscape of A. baumannii in Bangladesh, contributing to our understanding of its epidemiology and pathogenicity and informing strategies to combat this pathogen., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Purpose: We aimed to characterise Yersinia enterocolitica from human clinical specimens in Switzerland using epidemiological, microbiological and whole-genome sequencing (WGS) data., Methods: Isolates (n = 149) were collected between January 2019 and December 2023. Epidemiological data was noted and strains were characterized by biochemical and serological typing, antimicrobial susceptibility testing (AST), and WGS-based analysis., Results: Most of the isolates (86%) were from stool specimens and 52% were from male patients. The patients' median age was 28 years (range < 1-94 years). Typing assigned the isolates to bioserotype 4/O:3 (44%), biotype 1A (34%), bioserotype 2/O:9 (21%), and bioserotype 3/O:3 (1%). WGS identified Y. enterocolitica (n = 147), Y. alsatica (n = 1) and Y. proxima (n = 1). Seven isolates were multidrug resistant (MDR) and harboured plasmid pAB829 carrying aph(3″)-Ib, aph(6)-Id, and tet(Y) (n = 1), pAC120 carrying aph(6)-Id and tet(A) (n = 2), or a 12.6 kb Tn2670-like transposon containing catA1, aadA12, sul1, and qacEΔ1 (n = 4). Virulence factors (VFs) included ail (n = 99), invB, (n = 145), ystA (n = 99), ystB (n = 48) and pYV-associated VFs (n = 93). MLST and cgMLST analysis showed that BT 1A strains consisted of several STs and were highly diverse, whereas BT 2/O:9 strains were all ST12 and clustered closely, and BT 4/O:3 strains mostly belonged to ST18 but were more diverse. SNP analysis revealed two highly clonal BT 4/O:3 subpopulations with wide spatio-temporal distribution., Conclusions: Y. enterocolitica BT 1A, BT 2/O:9 and BT 4/O:3 are frequently associated with human yersiniosis in Switzerland. WGS-based subtyping of Y. enterocolitica is a powerful tool to explore the genetic diversity and the pathogenic potential of human isolates., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Probiotics are the health beneficial microorganisms and suitable for food industry if found fit for human consumption. In the present study, Lactiplantibacillus plantarum MCC5231, a probiotic bacterium included in vegetable-based beverages, was evaluated for its safety characteristics and gastrointestinal survival using a combined in silico and in vitro approach. The strain was found to be devoid of hemolytic, lecithinase and gelatinase activities. Additionally, it does not consist any transferable antibiotic resistance genes. Further, whole genome sequence analysis revealed the presence of three intact prophages and 14 virulence-associated genes, however, none of them posed a pathogenic threat. Importantly, MCC5231 do not possess any gene associated with toxin production. The strain harbored a CRISPR system, enhancing defense against prophages. Survival assays under simulated gastric and intestinal fluid conditions demonstrated viability rates of 71.4 % and 83.3 %, respectively. Genetic analysis of the mucin binding protein indicated possession of a type II mucin binding domain, suggesting moderate adhesion to intestinal cells. Furthermore, L. plantarum MCC5231 exhibited the ability to produce exopolysaccharides and form biofilms, which may confer additional protection in the gastrointestinal tract. Based on these findings, L. plantarum MCC5231 appears to be a safe probiotic candidate suitable for commercial use in the food industry., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Prakash M Halami reports financial support was provided by India Ministry of Science & Technology Department of Biotechnology. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Priestia is a genus that was renamed from the genus Bacillus based on the conserved signature indels (CSIs) in protein sequences that separate Priestia species from Bacillus, with the latter only including species closely related to B. subtilis and B. cereus. Diagnosis of anthrax, a zoonotic disease, is implicated by tripartite anthrax virulence genes (lef, pagA, and cya) and poly-γ-D-glutamic acid capsular genes cap-ABCDE of Bacillus anthracis. Due to the amplification of anthrax virulence genes in Priestia isolates, the search for homologous anthrax virulence genes within the Priestia genomes (n = 9) isolated from animal blood smears was embarked upon through whole genome sequencing. In silico taxonomic identification of the isolates was conducted using genome taxonomy database (GTDB), average nucleotide identity (ANI), and multi-locus sequence typing (MLST), which identified the genomes as P. aryabhattai (n = 5), P. endophytica (n = 2) and P. megaterium (n = 2). A pan-genome analysis was further conducted on the Priestia genomes, including the screening of virulence, antibiotic resistance genes and mobile genetic elements on the sequenced genomes. The oligoribonuclease NrnB protein sequences showed that Priestia spp. possess a unique CSI that is absent in other Bacillus species. Furthermore, the CSI in P. endophytica is unique from other Priestia spp. Pan-genomic analysis indicates that P. endophytica clusters separately from P. aryabhattai and P. megaterium. In silico BLASTn genome analysis using the SYBR primers, Taqman probes and primers that target the chromosomal marker (Ba-1), protective antigen (pagA), and lethal factor (lef) on B. anthracis, showed partial binding to Priestia regions encoding for hypothetical proteins, pyridoxine biosynthesis, hydrolase, and inhibitory proteins. The antibiotic resistance genes (ARG) profile of Priestia spp. showed that the genomes contained no more than two ARGs. This included genes conferring resistance to rifamycin and fosfomycin on P. endophytica, as well as clindamycin on P. aryabhattai and P. megaterium. Priestia genomes lacked B. anthracis plasmids and consisted of plasmid replicon types with unknown functions. Furthermore, the amplification of Priestia strains may result in false positives when qPCR is used to detect the virulence genes of B. anthracis in soil, blood smears, and/or environmental samples., Competing Interests: Declaration of competing interest None., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Salmonella is a foodborne pathogen that causes salmonellosis, of which retail chicken meat is a major source. However, the prevalence of Salmonella in different retail chicken supply modes and the threat posed to consumers remains unclear. The prevalence, serotype distribution, antibiotic resistance, and genomic characteristics of Salmonella in three supply modes of retail chicken (live poultry, frozen, and chilled) were investigated using whole-genome sequencing (WGS) and machine learning (ML). In this study, 480 retail chicken samples from live poultry, frozen, and chilled supply modes in Guangzhou from 2020 to 2021, as well as 253 Salmonella isolates (total isolation rate = 53.1 %), were collected. The prevalence of isolates in the live poultry mode (67.5 %, 81/120) was statistically higher than in the frozen (50.0 %, 120/240) and chilled (43.3 %, 52/120) (P < 0.05) modes. Serotype identification showed significant differences in the serotype distribution of Salmonella in different supply modes. S. Enteritis (46.7 %) and S. Indiana (14.2 %) were predominant in the frozen mode. S. Agona (23.5 %) and S. Saintpaul (13.6 %) were predominant in live poultry, while S. Enteritis (40.4 %) and S. Kentucky (17.3 %) were predominant in chilled mode. Antibiotic testing showed that frozen mode isolates were more resistant; the multidrug-resistant (MDR) rate of isolates in the frozen mode reached 91.8 %, significantly higher than in the chilled (86.5 %) and live (74.1 %) (P < 0.05) modes. WGS was performed on 155 top serotypes (S. Enteritidis, S. Kentucky, S. Indiana, and S. Agona). The antibiotic resistance gene analysis showed that the abundance and carrying rate of antibiotic resistance genes of Salmonella in the frozen mode (54 types, 16.1 %) were significantly higher than in other modes (live poultry: 36 types, 9.4 %, P < 0.05; chilled: 31 types, 11.6 %). The bla NDM-1 and bla NDM-9 genes encoding carbapenem resistance were found in frozen mode isolates on a complex transposon consisting of TnAS3-IS26. Virulence factors and plasmid replicons were abundant in the studied frozen mode isolates. In addition, single nucleotide polymorphism (SNP) phylogenetic tree results showed that in the frozen supply mode, the S. Enteritidis clonal clade continued to contaminate retail chicken meat and was homologous to S. Enteritidis strains found in farm chicken embryos, slaughterhouse chicken carcasses, and patients from hospitals in China (SNP 0 = 10). Notably, the pan-genome-based ML model showed that characteristic genes in frozen and live poultry isolates differed. The narZ gene was a key characteristic gene in frozen isolates, encoding nitrate reductase, relating to anaerobic bacterial growth. The ydgJ gene is a key characteristic gene in the live mode and encodes an oxidoreductase related to oxidative function in bacteria. The high prevalence of live poultry mode Salmonella and the transmission of frozen mode MDR Salmonella in this study pose serious risks to food safety and public health, emphasizing the importance of improving disinfection and cold storage measures to reduce Salmonella contamination and transmission. In conclusion, the continued surveillance of Salmonella across different supply models and the development of an epidemiological surveillance system based on WGS is necessary., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

IncQ-type plasmids have become important vectors in the dissemination of bla GES among different bacterial genera and species from different environments around the world, and studies estimating the occurrence of Guiana extended-spectrum (GES)-type β-lactamases are gaining prominence. We analyzed the genetic aspects of two IncQ1 plasmids harboring different bla GES variants from human and environmental sources. The bla GES variants were identified using polymerase chain reaction (PCR) in Aeromonas veronii isolated from hospital effluent and Klebsiella variicola isolated from a rectal swab of a patient admitted to the cardiovascular intensive care unit in a different hospital. Antimicrobial-susceptibility testing and transformation experiments were performed for phenotypic analysis. Whole-genome sequencing was performed using Illumina and Oxford Nanopore platforms. The comparative analysis of plasmids was performed using BLASTn, and the IncQ1 plasmids showed a high identity and similar size. A. veronii harbored bla GES-7 in a class 1 integron (In2061), recently described by our group, and K. variicola carried bla GES-5 in the known class 1 integron. Both integrons showed a fused gene cassette that encodes resistance to aminoglycosides and fluoroquinolones, with an IS6100 truncating the 3'-conserved segment. The fused genes are transcribed together, although the attC site is disrupted. These gene cassettes can no longer be mobilized. This study revealed a mobilome that may contribute to the dissemination of GES-type β-lactamases in Brazil. Class 1 integrons are hot spots for bacterial evolution, and their insertion into small IncQ-like plasmids displayed successful recombination, allowing the spread of bla GES variants in various environments. Therefore, they can become prevalent across clinically relevant pathogens., Competing Interests: Declaration of competing interest None declared., (Copyright © 2024. Published by Elsevier B.V.)

Lactobacillus helveticus exhibits a remarkable proteolytic system. However, the etiology of these protein hydrolysis characteristics, whether caused by extracellular proteinases (EP) or cell envelope proteinases (CEP), has been puzzling researchers. In this study, third-generation Nanopore whole genome sequencing and proteomics analysis were used to unravel the root cause of the aforementioned confusion. The genome of L. helveticus Lh191404 was 2,117,643 bp in length, with 67 secreted proteins were found. Combined with proteomic analysis, it was found that the protein composition of extraction from CEP and EP were indeed the same substance. Bioinformatics analysis indicated that the CEP belonged to the PrtH1 Variant (PrtH1&#95;V) genotype by phylogenetic analysis. The three-dimensional structures of various domains within the PrtH1&#95;V-191404 had been characterized, providing a comprehensive understanding of its structural features. Results of proteinase activity showed that the optimal reaction temperature was 40 °C, with a pH of 6.50. These findings suggested that the origin of EP in L. helveticus Lh191404 may be due to CEP being released into the substrate after detaching from the cell wall. This research is of guiding significance for further understanding the operational mechanism of the protein hydrolysis system in lactic acid bacteria., Competing Interests: Declaration of competing interest The authors confirm that they have no conflicts of interest with respect to the work described in this manuscript., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Nosocomial outbreaks caused by carbapenem-resistant Acinetobacter baumannii (CRAB) strains are rapidly emerging worldwide and are cause for concern. Herein, we aimed to describe the genomic characteristics of CRAB strains isolated from two hospitals in China in 2023. The A. baumannii isolates were mainly collected from the ICU and isolated from the sputum (71.43%, 15/21), followed by urine (14.29%, 3/21). Twenty-one A. baumannii strains possessed a multidrug-resistant (MDR) profile, and whole-genome sequencing showed that they all carried bla OXA-23 . Based on the Pasteur multilocus sequence typing (MLST) scheme, all strains were typed into a sequence type 2 (ST2). Based on the Oxford MLST scheme, six strains belonged to ST540, three of which were ST208, and four strains were assigned to ST784. Kaptive showed most of the strains (38.10%, 8/21) contained KL93. As for the lipoolygosaccharide (OC locus) type, OCL1c and OCL1d were identified, accounting for 33.33% (7/21) and 66.67% (14/21), respectively. Based on the BacWGSTdb server, we found that the strains belonging to ST540 and ST784 were all collected from China. However, the ST938 strains were isolated from Malaysia and Thailand. Comparative genomics analysis showed that the AB10 strain had a closed relationship with SXAB10-SXAB13 strains, suggesting the transmission happened in these two hospitals and other hospital in China. In addition, the 4300STDY7045869 strain, which was collected from Thailand, possessed near genetic relationship with our isolates in this study, suggesting the possible spread among various countries. Additionally, 3-237 single nucleotide polymorphisms were observed among these strains. In conclusion, this study conducted a genome-based study for A. baumannii strains collected from two hospitals in China and revealed their epidemiological and molecular features. Clone spreading occurred in these two hospitals. Hence, there is an urgent need for increased surveillance in hospitals and other clinical settings to prevent and control CRAB spreading., Competing Interests: Declaration of competing interest The authors declare that there is no conflict of interest., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Understanding the prevalence and distribution of CRISPR-Cas systems across different strains can illuminate the ecological and evolutionary dynamics of Clostridium botulinum populations. In this study, we conducted genome mining to characterize the CRISPR-Cas systems of C. botulinum strains. Our analysis involved retrieving complete genome sequences of these strains and assessing the diversity, prevalence, and evolution of their CRISPR-Cas systems. Subsequently, we performed an analysis of homology in spacer sequences from identified CRISPR arrays to investigate and characterize the range of targeted phages and plasmids. Additionally, we investigated the evolutionary trajectory of C. botulinum strains under selective pressures from foreign invasive DNA. Our findings revealed that 306 strains possessed complete CRISPR-Cas structures, comprising 58% of the studied C. botulinum strains. Secondary structure prediction of consensus repeats indicated that subtype II-C, with longer stems compared to subtypes ID and IB, tended to form more stable RNA secondary structures. Moreover, protospacer motif analysis demonstrated that strains with subtype IB CRISPR-Cas systems exhibited 5'-CGG-3', 5'-CC-3', and 5'-CAT-3' motifs in the 3' flanking regions of protospacers. The diversity observed in CRISPR-Cas systems indicated their classification into subtypes IB, ID, II-C, III-B, and III-D. Furthermore, our results showed that systems with subtype ID and III-D frequently harbored similar spacer patterns. Moreover, analysis of spacer sequences homology with phage and prophage genomes highlighted the specific activities exhibited by subtype IB and III-B against phages and plasmids, providing valuable insights into the functional specialization within these systems., Competing Interests: Declaration of competing interest The authors declare that they have no competing interests., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Mobile colistin resistance (mcr) genes are pivotal contributors to last-line of antimicrobial resistance in human infections. Shewanella, historically recognized as a natural environmental bacterium with metal reduction capabilities, recently has been observed in clinical settings. However, limited knowledge has been explored on genetic differences between strains from non-clinical and clinical strains. In this study, we conducted the whole genome sequencing on six Arctic strains, illustrated the phylogenetic relationships on published 393 Shewanella strains that categorized the genus into four lineages (L1 to L4). Over 86.4% of clinical strain group (CG) strains belonged to L1 and L4, carrying mcr-4 genes and a complete metal-reduction pathways gene cluster. Remarkably, a novel Arctic Shewanella strain in L3, exhibits similar genetic characteristics with CG strains that carried both mcr-4 genes and a complete metal reduction pathway gene cluster. It raised concerns about the transmission ability from environment to clinic setting causing in the potential infections, and emphasized the need for monitoring the emerging strains with human infections., Competing Interests: Declaration of competing interest The authors declare no competing interests., (Copyright © 2023. Published by Elsevier B.V.)

Leminorella grimontii strain LG-KP-E1-2-T0 was isolated from Zophobas morio larvae. It showed a susceptibility phenotype compatible with the expression of an inducible extended-spectrum β-lactamase. The presence of a chromosomal bla gene encoding for the class A GRI-1 β-lactamase was revealed by whole-genome sequencing. GRI-1 shared the highest amino acid identity with RIC-1 and OXY-type β-lactamases (76-80%). Analysis of six further publicly-available L. grimontii draft genomes deposited in NCBI revealed that bla GRI-1 was always present. Core-genome analysis indicated that LG-KP-E1-2-T0 was unique from other strains. We provided the first complete genome of L. grimontii and new insights on its chromosomal β-lactamases., (© 2024. The Author(s).)

Background: Chronic leg ulcers are hard to treat and can be a burden, particularly in resource-limited settings where diagnosis is a challenge. Staphylococcus aureus is among the common bacteria isolated from chronic wounds with a great impact on wound healing, particularly in patients with co-morbidities. Antimicrobial resistance genes and virulence factors in Staphylococcus aureus isolates were assessed to support healthcare professionals to make better therapeutic choices, and importantly to curb the development and spread of antibiotic resistance., Methods: A cross-sectional study involved both inpatients and outpatients with chronic leg ulcers was conducted from August 2022 to April 2023 in 2 health facilities in Kilimanjaro region in Tanzania. Antimicrobial susceptibility testing was done using the disk diffusion method. Further, whole genome sequencing was performed to study the genotypic characteristics of the isolates., Results: A total of 92 participants were recruited in which 9 participants were only positive for 10 Staphylococcus aureus isolates upon culture. Five STs among 9 isolates were identified. Most of them belonged to ST8 (44%), with 1 isolate does not belong to any ST. Additionally, 50% of the isolates were methicillin-resistant Staphylococcus aureus (MRSA). All S. aureus isolates had almost similar virulence factors such as hemolysin, proteases and evasions that promote toxin production, protease production and host immune evasion respectively. Moreover, all mecA positive S. aureus isolates were phenotypically susceptible to cefoxitin., Conclusion: Presence of mecA positive S. aureus isolates which are also phenotypically susceptible to cefoxitin implies the possibility of classifying MRSA as MSSA. This may result in the possible emergence of highly cefoxitin - resistant strains in health care and community settings when subsequently exposed to beta-lactam agents. Therefore, combination of whole genome sequencing and conventional methods is important in assessing bacterial resistance and virulence to improve management of patients., Competing Interests: Declaration of competing interest The authors declare that they have no competing interests., (Copyright © 2024. Published by Elsevier B.V.)

Background: Vancomycin-resistant Enterococcus faecium (VREfm) are significant nosocomial pathogens. Sequence type (ST) 80 vanA-encoding VREfm predominate in Irish hospitals, but their transmission is poorly understood., Aims: To investigate transmission and persistence of predominant complex type (CT) VREfm in two wards of an Irish hospital (H1) using whole-genome sequencing, and their intra- and inter-hospital dissemination., Methods: Rectal screening (N = 330, September 2019 to December 2022) and environmental (N = 48, November 2022 to December 2022) E. faecium were investigated. Isolate relatedness was assessed by core-genome multi-locus sequence typing (cgMLST) and core-genome single nucleotide polymorphism (cgSNP) analysis. Likely transmission chains were identified using SeqTrack (https://graphsnp.fordelab.com/graphsnp) using cgSNP data and recovery location. Well-characterized E. faecium (N = 908) from seven Irish hospitals including H1 (June 2017 to July 2022) were also investigated., Findings: Conventional MLST assigned isolates to nine STs (ST80, 82%). cgMLST identified three predominant ST80 CTs (CT2933, CT2932 and CT1916) (55% of isolates) of related isolates (≤20 allelic differences). cgSNP analysis differentiated these CTs into multiple distinct closely related genomic clusters (≤10 cgSNPs). Parisimonious network construction identified 55 likely inter- and intra-ward transmissions with epidemiological support between patients ≤30 days involving 73 isolates (≤10 cgSNPs) from seven genomic clusters. Numerous other likely transmissions over longer time periods without evident epidemiological links were identified, suggesting persistence and unidentified reservoirs contribute to dissemination. The three CTs predominated among E. faecium (N = 1286) in seven hospitals, highlighting inter-hospital spread without known epidemiological links., Conclusion: This study revealed the long-term intra- and inter-hospital dominance of three major CT ST80 VREfm lineages, widespread transmission and persistence, implicating unidentified reservoirs., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Background: Folate is crucial for maintaining health, but humans are unable to synthesize folate and need to obtain it from food. Lactiplantibacillus plantarum can produce the necessary vitamin B for the human body, including folate. Whole genome sequencing technology can clarify the physiological characteristics of folate production in Lactiplantibacillus plantarum. In order to explore new Lactiplantibacillus plantarum that produce folate, the folate production and probiotic characteristics of Lactiplantibacillus plantarum ZFM55 isolated from infant feces were investigated, and whole genome sequencing was performed., Results: The folate synthesis ability of Lactiplantibacillus plantarum ZFM55 were measured, and its total folate production was 299.72 ± 28.81 ng mL -1 . Subsequently, its probiotic properties were explored. The antibacterial test showed that its inhibition zone diameter against Staphylococcus aureus and Salmonella typhimurium was 15.5 ± 0.82 mm and 13.88 ± 0.98 mm, respectively. The tolerance test results indicated that it maintained good activity in simulated gastrointestinal tract and bile salt environments. In vitro intestinal simulation experiments had confirmed that Lactiplantibacillus plantarum ZFM55 can increase the abundance of beneficial bacteria such as Bifidobacteria in the intestine and inhibit the growth of harmful bacteria such as Escherichia&#95;Shigella. Genomic sequencing indicated that the genetic material of Lactiplantibacillus plantarum ZFM55 contains one chromosome and three plasmids, and it has 20 genes related to folate synthesis, which explains its ability to produce folate., Conclusion: This study reports a new potential probiotic that produces folate, and provides ideas for exploring probiotics with specific probiotic characteristics. © 2024 Society of Chemical Industry., (© 2024 Society of Chemical Industry.)

Staphylococcus aureus thrives at animal-human-environment interfaces. A large-scale work from our group indicated that antimicrobial resistance (AMR) in commensal S. aureus strains from wild ungulates is associated with agricultural land cover and livestock farming, raising the hypothesis that AMR genes in wildlife strains may originate from different hosts, namely via exchange of mobile genetic elements (MGE). In this work, we generate the largest available dataset of S. aureus draft genomes from wild ungulates in Portugal and explore their mobilome, which can determine important traits such as AMR, virulence, and host specificity, to understand MGE exchange. Core genome multi-locus sequence typing based on 98 newly generated draft genomes and 101 publicly available genomes from Portugal demonstrated that the genomic relatedness of S. aureus from wild ungulates assigned to livestock-associated sequence types (ST) is greater compared to wild ungulate isolates assigned to human-associated STs. Screening of host specificity determinants disclosed the unexpected presence in wildlife of the immune evasion cluster encoded in φSa3 prophage, described as a human-specific virulence determinant. Additionally, two plasmids, pAVX and pETB, previously associated with avian species and humans, respectively, and the Tn553 transposon were detected. Both pETB and Tn553 encode penicillin resistance through blaZ. Pangenome analysis of wild ungulate isolates shows a core genome fraction of 2133 genes, with isolates assigned to ST72 and ST3224 being distinguished from the remaining by MGEs, although there is no reported role of these in adaptation to wildlife. AMR related gene clusters found in the shell genome are directly linked to resistance against penicillin, macrolides, fosfomycin, and aminoglycosides, and they represent mobile ARGs. Altogether, our findings support epidemiological interactions of human and non-human hosts at interfaces, with MGE exchange, including AMR determinants, associated with putative indirect movements of S. aureus among human and wildlife hosts that might be bridged by livestock., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The genus Pseudochrobactrum encompasses free-living bacteria phylogenetically close to Ochrobactrum opportunistic pathogens and to Brucella, facultative intracellular parasites causing brucellosis, a worldwide-extended and grave zoonosis. Recently, Pseudochrobactrum strains were isolated from Brucella natural hosts on Brucella selective media, potentially causing diagnostic confusions. Strikingly, P. algeriensis was isolated from cattle lymph nodes, organs that are inimical to bacteria. Here, we analyse P. algeriensis potential virulence factors in comparison with Ochrobactrum and Brucella. Consistent with genomic analyses, Western-Blot analyses confirmed that P. algeriensis lacks the ability to synthesize the N-formylperosamine O-polysaccharide characteristic of the lipopolysaccharide (LPS) of smooth Brucella core species. However, unlike other Pseudochrobactrum but similar to some early diverging brucellae, P. algeriensis carries genes potentially synthetizing a rhamnose-based O-polysaccharide LPS. Lipid A analysis by MALDI-TOF demonstrated that P. algeriensis LPS bears a lipid A with a reduced pathogen-associated molecular pattern, a trait shared with Ochrobactrum and Brucella that is essential to generate a highly stable outer membrane and to delay immune activation. Also, although not able to multiply intracellularly in macrophages, the analysis of P. algeriensis cell lipid envelope revealed the presence of large amounts of cationic aminolipids, which may account for the extremely high resistance of P. algeriensis to bactericidal peptides and could favor colonization of mucosae and transient survival in Brucella hosts. However, two traits critical in Brucella pathogenicity are either significantly different (T4SS [VirB]) or absent (erythritol catabolic pathway) in P. algeriensis. This work shows that, while diverging in other characteristics, lipidic envelope features relevant in Brucella pathogenicity are conserved in Brucellaceae. The constant presence of these features strongly suggests that reinforcement of the envelope integrity as an adaptive advantage in soil was maintained in Brucella because of the similarity of some environmental challenges, such as the action of cationic peptide antibiotics and host defense peptides. This information adds knowledge about the evolution of Brucellaceae, and also underlines the taxonomical differences of the three genera compared., Competing Interests: Declaration of Competing Interest The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Glanders, a highly contagious and often fatal disease affecting equids, is caused by Burkholderia mallei. Although sporadic cases of equine glanders have recently been documented in Mongolia, genome sequencing and molecular studies of the bacteria within this region are lacking. This study provided the first molecular characterization of B. mallei isolated from four native Mongolian horses from two different provinces in 2019 and 2022 by applying whole-genome sequencing with two SNP types (previously developed genotyping with 15 SNP markers that provide global coverage of the B. mallei population and the core genome coding SNP typing developed in this study). The Mongolian isolates were located within the L3B1 cluster, which was previously associated with the V-120 strain from Russia. Within the L3B1 cluster shared by neighboring countries, they were in a unique subbranch. In this study, specific SNP markers unique to the Mongolian strains were identified to track these strains using a high-resolution melting analysis (HRMA). This study revealed the unique phylogenetic background of Mongolian strains isolated from the eastern part of Mongolia. HRMA specific to the Mongolian subbranch may contribute to the molecular epidemiological monitoring of glanders in Mongolia and surrounding countries., Competing Interests: Declaration of competing interest The authors declare that there are no conflicts of interest., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Foodborne illnesses pose a substantial health and economic burden, presenting challenges in prevention due to the diverse microbial hazards that can enter and spread within food systems. Various factors, including natural, political and commercial drivers, influence food production and distribution. The risks of foodborne illness will continue to evolve in step with these drivers and with changes to food systems. For example, climate impacts on water availability for agriculture, changes in food sustainability targets and evolving customer preferences can all have an impact on the ecology of foodborne pathogens and the agrifood niches that can carry microorganisms. Whole-genome and metagenome sequencing, combined with microbial surveillance schemes and insights from the food system, can provide authorities and businesses with transformative information to address risks and implement new food safety interventions across the food chain. In this Review, we describe how genome-based approaches have advanced our understanding of the evolution and spread of enduring bacterial foodborne hazards as well as their role in identifying emerging foodborne hazards. Furthermore, foodborne hazards exist in complex microbial communities across the entire food chain, and consideration of these co-existing organisms is essential to understanding the entire ecology supporting pathogen persistence and transmission in an evolving food system., (© 2024. Springer Nature Limited.)

Background: Lacticaseibacillus (formerly Lactobacillus) rhamnosus is widely used in probiotics or food supplements to promote microbiome health and may also be part of the normal microbiota of the human gastrointestinal tract. However, it rarely also causes invasive or severe infections in patients. It has been postulated that these infections may originate from probiotics or from endogenous commensal reservoirs. In this report, we examine the population structure of Lacticaseibacillus rhamnosus and investigate the utility of using bacterial genomics to identify the source of invasive Lacticaseibacillus infections., Methods: Core genome phylogenetic analysis was performed on 602 L. rhamnosus genome sequences from the National Center for Biotechnology public database. This information was then used along with newly generated sequences of L. rhamnosus isolates from yogurt to investigate a fatal case of L. rhamnosus endocarditis., Results: Phylogenetic analysis demonstrated substantial genetic overlap of L. rhamnosus isolates cultured from food, probiotics, infected patients, and colonized individuals. This was applied to a patient who had both consumed yogurt and developed L. rhamnosus endocarditis to attempt to identify the source of his infection. The sequence of the isolate from the patient's bloodstream differed at only one nucleotide position from one of the yogurt isolates. Both isolates belonged to a clade, identified here as clade YC, composed of mostly gastrointestinal isolates from healthy individuals, some of which also differed by only a single nucleotide change from the patient's isolate., Conclusions: As illustrated by this case, whole genome sequencing may be insufficient to reliably determine the source of invasive infections caused by L. rhamnosus., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Santoiemma et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Streptomyces species are experts in the production of bioactive secondary metabolites; however, their taxonomy has fallen victim of the tremendous interest shown by the scientific community, evident in the discovery of numerous synonymous in public repositories. Based on genomic data from NCBI Datasets and nomenclature from the LPSN database, we compiled a dataset of 600 Streptomyces species along with their annotations and metadata. To pinpoint the most suitable taxonomic classification method, we conducted a comprehensive assessment comparing multiple methodologies, including analysis of 16S rRNA, individual housekeeping genes, multilocus sequence analysis (MLSA), and Fast Average Nucleotide Identity (FastANI) on a subset of 409 species with complete data. Due to insufficient resolution of 16S rRNA and inconsistency observed in individual housekeeping genes, we performed a more in-depth analysis, comparing only FastANI and MLSA, which expanded our dataset to include 502 species. With FastANI validated as the preferred method, we conducted pairwise analysis on the entire dataset identifying 59 non-unique species among the 600, and subsequently refined the dataset to 541 unique species. Additionally, we collected data on 724 uncharacterized Streptomyces strains to investigate the uniqueness potential of the unannotated fraction of the Streptomyces genus. Utilizing FastANI, 289 strains could be successfully classified into one of the 541 Streptomyces species. KEY POINTS: • Evaluation of taxonomic classification methods for Streptomyces species. • Whole genome analysis, specifically FastANI, has been chosen as preferred method. • Various reclassifications are proposed within the Streptomyces genus., (© 2024. The Author(s).)

Clarithromycin (CLR) is currently a key antibiotic for Helicobacter pylori infection treatment, however, the data on CLR resistance patterns in Russia are missing. Here, we applied WGS-based approach to H. pylori clinical isolates from Russia to comprehensively investigate sequence variation, identify putative markers of CLR resistance and correlate them with phenotypic susceptibility testing. The phenotypic susceptibility of 44 H. pylori isolates (2014-2022) to CLR was determined by disc diffusion method: 23 isolates were CLR-resistant and 21-CLR-susceptible. All isolates were subjected to WGS and submitted to GenBank. Based on complete sequence analysis, we showed that among all sequence variants, the combination of mutations A2146G/A2147G in the 23S rRNA gene is the most reliable for prediction of phenotypic susceptibility. For the first time, the average number of mutations in 106 virulence-associated genes between resistant and susceptible groups were compared. Moreover, this study presents the first WGS insight into genetic diversity of H. pylori in Russia with a particular focus on the molecular basis of drug resistance: the novel mutations were described as potential markers for the resistance development. Of these, the most prominent was a frameshift deletion (252:CGGGT) in HP0820 coding region, which is a good candidate for further investigation., (© 2024. The Author(s).)

Bacteria can solubilize phosphorus (P) through the secretion of low-molecular-weight organic acids and acidification. However, the genes involved in the production of these organic acids are poorly understood. The objectives of this study were to verify the calcium phosphate solubilization and the production of low-molecular-weight organic acids by diverse genera of phosphate solubilizing bacterial strains (PSBS); to identify the genes related to the synthesis of the organic acids in the genomes of these strains and; to evaluate growth and nutrient accumulation of maize plants inoculated with PSBS and fertilized with Bayóvar rock phosphate. Genomic DNA was extracted for strain identification and annotation of genes related to the organic acids production. A greenhouse experiment was performed with five strains plus 150 mg dm - 3 P 2 O 5 as Bayóvar rock phosphate (BRP) to assess phosphate solubilization contribution to maize growth and nutrition. Paraburkholderia fungorum UFLA 04-21 and Pseudomonas anuradhapurensis UFPI B5-8A solubilized over 60% of Ca phosphate and produced high amounts of citric/maleic and gluconic acids in vitro, respectively. Eleven organic acids were identified in total, although not all strains produced all acids. Besides, enzymes related to the organic acids production were found in all bacterial genomes. Plants inoculated with strains UFPI B5-6 (Enterobacter bugandensis), UFPI B5-8A, and UFLA 03-10 (Paenibacillus peoriae) accumulated more biomass than the plants fertilized with BRP only. Strains UFLA 03-10 and UFPI B5-8A increased the accumulation of most macronutrients, including P. Collectively, the results show that PSBS can increase maize growth and nutrient accumulation based on Bayóvar rock phosphate fertilization., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Background: Streptococcus suis (S. suis) is an important swine and human pathogen. A recent study reported the first isolate of S. suis capable of infecting fish, designated as S. suis strain 3112. The bacterium was isolated from snakeskin gourami (Trichopodus pectoralis), an economically important fish species native to Southeast Asia, and it was previously shown that it can infect and cause lethal streptococcosis in the fish., Results: In this study, we present the complete genome of S. suis 3112. Molecular sequence analysis revealed that it belongs to serotype 6, sequence type 2340. Phylogenetic analysis showed that the bacterium clustered with healthy-pig S. suis isolates, suggestive of an ultimate swine (as opposed to human) origin of the bacterium. Two fluoroquinolone resistance genes are present in the bacterial genome, namely patA and patB. Our results showed that both genes are expressed in our bacterium, and the bacterium is resistant to norfloxacin, but is still sensitive to other fluoroquinolones, including ciprofloxacin, enrofloxacin, and sparfloxacin. Additionally, the bacterium is sensitive to β-lactams, tetracyclines, sulphonamides, and an aminoglycoside., Conclusions: This study reports and describes the complete genome of S. suis 3112, the first isolate of S. suis known to infect fish, and provides further insights into the bacterial isolate, particularly regarding its drug resistance profile. These results will facilitate further investigations of the comparative genomics and pathogenic characteristics of S. suis, as well as the development of control strategies against this newly-identified fish pathogen., (© 2024. The Author(s).)

Cases of diphtheria, even in immunized individuals, are still reported in several parts of the world, including in Brazil. New outbreaks occur in Europe and other continents. In this context, studies on Corynebacterium diphtheriae infections are highly relevant, both for a better understanding of the pathogenesis of the disease and for controlling the circulation of clones and antimicrobial resistance genes. Here we present a case of cutaneous infection by multidrug-resistant Corynebacterium diphtheriae and provide its whole-genome sequencing. Genomic analysis revealed resistance genes, including tet(W), sul1, cmx, rpoB2, rbpA and mutation in rpoB. We performed phylogenetic analyzes and used the BRIG to compare the predicted resistance genes with those found in genomes from other significant isolates, including those associated with some outbreaks. Virulence factors such as spaD, srtBC, spaH, srtDE, surface-anchored pilus proteins (sapD), nonfimbrial adhesins (DIP0733, DIP1281, and DIP1621), embC and mptC (putatively involved in CdiLAM), sigA, dtxR and MdbA (putatively involved) in post-translational modification, were detected. We identified the CRISPR-Cas system in our isolate, which was classified as Type II-U based on the database and contains 15 spacers. This system functions as an adaptive immune mechanism. The strain was attributed to a new sequence type ST-928, and phylogenetic analysis confirmed that it was related to ST-634 of C. diphtheriae strains isolated in French Guiana and Brazil. In addition, since infections are not always reported, studies with the sequence data might be a way to complement and inform C. diphtheriae surveillance., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

A Gram-stain-positive, aerobic, white-coloured, rod-shaped bacteria, designated as a strain dW9 T , was isolated from soil. Strain dW9 T was catalase-positive and oxidase-negative. Strain dW9 T grew at temperature of 20-37°C and at pH of 5.0-7.0. Phylogenetic and 16S rRNA gene analysis indicated that strain dW9 T belonged to the genus Paenibacillus with its closest relative being Paenibacillus filicis S4 T (97.4% sequence similarity). The genome size of dW9 T was 7,787,916 bp with DNA G+C content of 51.3%. The digital DNA-DNA hybridization (dDDH) and average nucleotide identity (ANI) values of dW9 T with its closest relatives were found to be <22.0% and <74.0%, respectively. The only respiratory quinone was MK-7, and the major fatty acids were antiso-C 15:0 and iso-C 16:0 . Overall, the comprehensive taxonomic analysis revealed that strain dW9 T met all the fundamental criteria to be classified as a novel species within the genus Paenibacillus. Accordingly, we propose the name Paenibacillus gyeongsangnamensis sp. nov., with the type strain dW9 T (=KCTC 43431 T =NBRC 116022 T ).

Tatum D. Mortimer works in the field of pathogen population genomics and evolution. In this mSphere of Influence article, she reflects on how "Frequency-dependent selection can forecast evolution in Streptococcus pneumoniae " by Azarian et al. and "Contingency, repeatability, and predictability in the evolution of a prokaryotic pangenome" by Beavan et al. made an impact on her by highlighting the ways in which genomic data can be used to predict pathogen evolution., Competing Interests: The author declares no conflict of interest.

Comprehensive and accurate genome annotation is crucial for inferring the predicted functions of an organism. Numerous tools exist to annotate genes, gene clusters, mobile genetic elements, and other diverse features. However, these tools and pipelines can be difficult to install and run, be specialized for a particular element or feature, or lack annotations for larger elements that provide important genomic context. Integrating results across analyses is also important for understanding gene function. To address these challenges, we present the Beav annotation pipeline. Beav is a command-line tool that automates the annotation of bacterial genome sequences, mobile genetic elements, molecular systems and gene clusters, key regulatory features, and other elements. Beav uses existing tools in addition to custom models, scripts, and databases to annotate diverse elements, systems, and sequence features. Custom databases for plant-associated microbes are incorporated to improve annotation of key virulence and symbiosis genes in agriculturally important pathogens and mutualists. Beav includes an optional Agrobacterium -specific pipeline that identifies and classifies oncogenic plasmids and annotates plasmid-specific features. Following the completion of all analyses, annotations are consolidated to produce a single comprehensive output. Finally, Beav generates publication-quality genome and plasmid maps. Beav is on Bioconda and is available for download at https://github.com/weisberglab/beav., Importance: Annotation of genome features, such as the presence of genes and their predicted function, or larger loci encoding secretion systems or biosynthetic gene clusters, is necessary for understanding the functions encoded by an organism. Genomes can also host diverse mobile genetic elements, such as integrative and conjugative elements and/or phages, that are often not annotated by existing pipelines. These elements can horizontally mobilize genes encoding for virulence, antimicrobial resistance, or other adaptive functions and alter the phenotype of an organism. We developed a software pipeline, called Beav, that combines new and existing tools for the comprehensive annotation of these and other major features. Existing pipelines often misannotate loci important for virulence or mutualism in plant-associated bacteria. Beav includes custom databases and optional workflows for the improved annotation of plant-associated bacteria. Beav is designed to be easy to install and run, making comprehensive genome annotation broadly available to the research community., Competing Interests: The authors declare no conflict of interest.

Klebsiella aerogenes is an understudied and clinically important pathogen. We therefore investigate its population structure by genome analysis aligned with metadata. We sequence 130 non-duplicated K. aerogenes clinical isolates and identify two inter-patient transmission events. We then retrieve all publicly available K. aerogenes genomes (n = 1,026, accessed by January 1, 2023) and analyze them with our 130 genomes. We develop a core-genome multi-locus sequence-typing scheme. We find that K. aerogenes is a species complex comprising four phylogroups undergoing evolutionary divergence, likely forming three species. We delineate remarkable clonal diversity and identify three worldwide-distributed carbapenemase-encoding clonal clusters, representing high-risk lineages. We uncover that K. aerogenes has an open genome equipped by a large arsenal of antimicrobial resistance genes. We identify two genetic regions specific for K. aerogenes, encoding a type VI secretion system and flagella/chemotaxis for motility, respectively, both contributing to the virulence. These results provide much-needed insights into the population structure and pan-genomes of K. aerogenes., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Correct termination of transcription is essential for gene expression. In bacteria, factor-dependent termination relies on the Rho factor, that classically has three conserved domains. Some bacteria also have a functional insertion region. However, the variation in Rho structure among bacteria has not been analyzed in detail. This study determines the distribution, sequence conservation, and predicted features of Rho factors with diverse domain architectures by analyzing 2730 bacterial genomes. About half (49.8%) of the species analyzed have the typical Escherichia coli like Rho while most of the other species (39.8%) have diverse, atypical forms of Rho. Besides conservation of the main domains, we describe a duplicated RNA-binding domain present in specific species and novel variations in the bicyclomycin binding pocket. The additional regions observed in Rho proteins exhibit remarkable diversity. Commonly, however, they have exceptional amino acid compositions and are predicted to be intrinsically disordered, to undergo phase separation, or have prion-like behavior. Phase separation has recently been shown to play roles in Rho function and bacterial fitness during harsh conditions in one species and this study suggests a more widespread role. In conclusion, diverse atypical Rho factors are broadly distributed among bacteria, suggesting additional cellular roles., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Live attenuated vaccines (LAVs) whose virulence would be controlled at the tissue level could be a crucial tool to effectively fight intracellular bacterial pathogens, because they would optimize the induction of protective immune memory while avoiding the long-term persistence of vaccine strains in the host. Rational development of these new LAVs implies developing an exhaustive map of the bacterial virulence genes according to the host organs implicated. We report here the use of transposon sequencing to compare the bacterial genes involved in the multiplication of Brucella melitensis, a major causative agent of brucellosis, in the lungs and spleens of C57BL/6 infected mice. We found 257 and 135 genes predicted to be essential for B. melitensis multiplication in the spleen and lung, respectively, with 87 genes common to both organs. We selected genes whose deletion is predicted to produce moderate or severe attenuation in the spleen, the main known reservoir of Brucella, and compared deletion mutants for these genes for their ability to protect mice against challenge with a virulent strain of B. melitensis. The protective efficacy of a deletion mutant for the plsC gene, implicated in phospholipid biosynthesis, is similar to that of the reference Rev.1 vaccine but with a shorter persistence in the spleen. Our results demonstrate that B. melitensis faces different selective pressures depending on the organ and underscore the effectiveness of functional genome mapping for the design of new safer LAV candidates., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Barbieux et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Corynebacterium pseudotuberculosis (CPS) is an important bacterial animal pathogen. CPS causes chronic, debilitating and currently incurable infectious diseases affecting a wide range of livestock and wild herbivores including camelids worldwide. Belonging to the Corynebacterium diphtheriae complex, this pathogen can also infect humans. The classical characterization of CPS is typically based on the testing of nitrate reductase activity, separating the two biovars Equi and Ovis. However, more refined resolutions are required to unravel routes of infection. This was realized in our study by generating and analyzing whole genome sequencing (WGS) data. Using newly created core genome multilocus sequence typing (cgMLST) profiles we were the first to discover isolates grouping in a cluster adjacent to clusters formed by CPS biovar Equi isolates. This novel cluster includes CPS isolates from alpacas, llamas, camels and dromedaries, which are characterized by a lack of nitrate reductase activity as encountered in biovar Ovis. This is of special interest for molecular epidemiology. Nevertheless, these isolates bear the genes of the nitrate locus, which are characteristic of biovar Equi isolates. However, sequence analysis of the genes narG and narH of the nitrate locus revealed indels leading to frameshifts and inactivity of the enzymes involved in nitrate reduction. Interestingly, one CPS isolate originating from another lama with an insertion in the MFS transporter (narT) is adjacent to a cluster formed by ovine CPS isolates biovar Equi. Based on this knowledge, the combination of biochemical and PCR based molecular biological nitrate reductase detection can be used for a fast and uncomplicated classification of isolates in routine diagnostics in order to check the origin of camelid CPS isolates. Further analysis revealed that partial sequencing of the ABC transporter substrate binding protein (CP258&#95;RS07935) is a powerful tool to assign the biovars and the novel genomovar., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Hiller et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Escherichia coli multi-locus sequence type ST131 is a globally distributed pandemic lineage that causes multidrug-resistant extra-intestinal infections. ST131 E. coli frequently produce extended-spectrum β-lactamases (ESBLs), which confer resistance to many β-lactam antibiotics and make infections difficult to treat. We sequenced the genomes of 154 ESBL-producing E. coli clinical isolates belonging to the ST131 lineage from patients at the University of Pittsburgh Medical Center (UPMC) between 2004 and 2018. Isolates belonged to the well described ST131 clades A (8%), B (3%), and C (89%). Time-dated phylogenetic analysis estimated that the most recent common ancestor (MRCA) for all clade C isolates emerged around 1989, consistent with previous studies. We identified multiple genes potentially under selection in clade C, including the cell wall assembly gene ftsI, the LPS biosynthesis gene arnC, and the yersiniabactin uptake receptor fyuA. Diverse ESBL-encoding genes belonging to the bla CTX-M , bla SHV , and bla TEM families were identified; these genes were found at varying numbers of loci and in variable numbers of copies across isolates. Analysis of ESBL flanking regions revealed diverse mobile elements that varied by ESBL type. Overall, our findings show that ST131 subclade C dominated among patients and uncover possible signals of ongoing adaptation within this ST131 lineage., (© 2024. The Author(s).)

The structure of microbial communities arises from a multitude of factors, including the interactions of microorganisms with each other and with the environment. In this work, we sought to disentangle those drivers by performing a cross-study, cross-biome meta-analysis of microbial occurrence data in more than 5000 samples, applying a novel network clustering algorithm aimed to capture conditional taxa co-occurrences. We then examined the phylogenetic and functional composition of the resulting clusters, and searched for global patterns of assembly both at the community level and in the presence/absence of individual metabolic pathways.Our analysis highlighted the prevalence of functional redundancy in microbial communities, particularly between taxa that co-occur in more than one environment, pointing to a relationship between functional redundancy and environmental adaptation. In spite of this, certain pathways were observed in fewer taxa than expected by chance, suggesting the presence of auxotrophy, and presumably cooperation among community members. This hypothetical cooperation may play a role in genome reduction, since we observed a negative relationship between the size of bacterial genomes and the size of the community they belong to.Overall, our results suggest the microbial community assembly is driven by universal principles that operate consistently across different biomes and taxonomic groups., (© 2024. The Author(s).)

Background: Magnetotactic bacteria (MTB) are a unique group of microorganisms that sense and navigate through the geomagnetic field by biomineralizing magnetic nanoparticles. MTB from the phylum Nitrospirota (previously known as Nitrospirae) thrive in diverse aquatic ecosystems. They are of great interest due to their production of hundreds of magnetite (Fe 3 O 4 ) magnetosome nanoparticles per cell, which far exceeds that of other MTB. The morphological, phylogenetic, and genomic diversity of Nitrospirota MTB have been extensively studied. However, the metabolism and ecophysiology of Nitrospirota MTB are largely unknown due to the lack of cultivation techniques., Methods: Here, we established a method to link the morphological, genomic, and metabolic investigations of an uncultured Nitrospirota MTB population (named LHC-1) at the single-cell level using nanoscale secondary-ion mass spectrometry (NanoSIMS) in combination with rRNA-based in situ hybridization and target-specific mini-metagenomics., Results: We magnetically separated LHC-1 from a freshwater lake and reconstructed the draft genome of LHC-1 using genome-resolved mini-metagenomics. We found that 10 LHC-1 cells were sufficient as a template to obtain a high-quality draft genome. Genomic analysis revealed that LHC-1 has the potential for CO 2 fixation and NO 3 - reduction, which was further characterized at the single-cell level by combining stable-isotope incubations and NanoSIMS analyses over time. Additionally, the NanoSIMS results revealed specific element distributions in LHC-1, and that the heterogeneity of CO 2 and NO 3 - metabolisms among different LHC-1 cells increased with incubation time., Conclusions: To our knowledge, this study provides the first metabolic measurements of individual Nitrospirota MTB cells to decipher their ecophysiological traits. The procedure constructed in this study provides a promising strategy to simultaneously investigate the morphology, genome, and ecophysiology of uncultured microbes in natural environments. Video Abstract., (© 2024. The Author(s).)

The Philippines is a high-incidence country for tuberculosis, with the increasing prevalence of multi- (MDR-TB) and extensively-drug (XDR-TB) resistant Mycobacterium tuberculosis strains posing difficulties to disease control. Understanding the genetic diversity of circulating strains can provide insights into underlying drug resistance mutations and transmission dynamics, thereby assisting the design of diagnostic tools, including those using next generation sequencing (NGS) platforms. By analysing genome sequencing data of 732 isolates from Philippines drug-resistance survey collections spanning from 2011 to 2019, we found that the majority belonged to lineages L1 (531/732; 72.5%) and L4 (European-American; n = 174; 23.8%), with the Manila strain (L1.2.1.2.1) being the most prominent (475/531). Approximately two-thirds of isolates were found to be at least MDR-TB (483/732; 66.0%), and potential XDR-TB genotypic resistance was observed (3/732; 0.4%), highlighting an emerging problem in the country. Genotypic resistance was highly concordant with laboratory drug susceptibility testing. By finding isolates with (near-)identical genomic variation, five major clusters containing a total of 114 isolates were identified: all containing either L1 or L4 isolates with at least MDR-TB resistance and spanning multiple years of collection. Closer inspection of clusters revealed transmission in prisons, some involving isolates with XDR-TB, and mutations linked to third-line drug bedaquiline. We have also identified previously unreported mutations linked to resistance for isoniazid, rifampicin, ethambutol, and fluoroquinolones. Overall, this study provides important insights into the genetic diversity, transmission and circulating drug resistance mutations of M. tuberculosis in the Philippines, thereby informing clinical and surveillance decision-making, which is increasingly using NGS platforms., (© 2024. The Author(s).)