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Current Microbiology Jun 2024Linezolid resistance in Enterococcus spp. is increasingly considered critically important and a public health threat which mandates the need to understand their genomic...
Linezolid resistance in Enterococcus spp. is increasingly considered critically important and a public health threat which mandates the need to understand their genomic contents and dissemination patterns. Here, we used whole-genome sequencing to characterize the resistome, virulome and mobile genetic elements of nine linezolid-resistant (LZD) enterococci (seven optrA-E. faecalis, one poxtA-E. faecium and one optrA-E. casseliflavus) previously obtained from the nares of healthy dogs, pigs, pig farmers and tracheal samples of nestling storks in Spain. Also, the relatedness of the isolates with publicly available genomes was accessed by core-genome single nucleotide polymorphism (SNP) analysis. The optrA gene of the E. faecalis and E. casseliflavus isolates was located downstream of the fexA gene. The optrA gene in the E. casseliflavus isolate was carried in a plasmid (pURX4962), while those in the seven E. faecalis isolates were chromosomally located. The OptrA proteins were mostly variants of wild type (DP-2: Y176D/T481P; RDK: I104R/Y176D/E256K; DD-3: Y176D/G393D; and EDD: K3E/Y176D/G393D), except two that were wild type (one E. faecalis and one E. casseliflavus). The poxtA gene in the E. faecium isolate was found alone within its contig. The cfrD was upstream of ermB gene in the E. casseliflavus isolate and flanked by ISNCY and IS1216. All the LZD enterococci carried plasmid rep genes (2-3) containing tetracycline, chloramphenicol and aminoglycoside resistance genes. All isolates except E. casseliflavus carried at least one intact prophage, of which E. faecalis-ST330 (X4957) from a pig carried the highest (n = 5). Tn6260 was associated with lnuG in E. faecalis-ST330 while Tn554 was with fexA in E. feaecalis-ST59 isolates. All except E. casseliflavus (n = 0) carried at least two metal resistance genes (MRGs), of which poxtA-carrying E. faecium-ST1739 isolate contained the most (arsA, copA, fief, ziaA, znuA, zosA, zupT, and zur). SNP-based analyses identified closely related optrA-E. faecalis isolates from a pig and a pig farmer on the same farm (SNP = 4). Moreover, optrA- carrying E. faecalis-ST32, -ST59, and -ST474 isolates from pigs were related to those previously described from humans (sick and healthy) and cattle in Spain, Belgium, and Switzerland (SNP range 43-86). These findings strongly suggest the transmission of LZD-E. faecalis between a pig and a pig farmer and potential inter-country dissemination. These highlight the need to strengthen molecular surveillance of LZD enterococci in all ecological niches and body parts to direct appropriate control strategies.
Topics: Animals; Phylogeny; Linezolid; Swine; Drug Resistance, Bacterial; Genome, Bacterial; Dogs; Anti-Bacterial Agents; Enterococcus; Gram-Positive Bacterial Infections; Humans; Whole Genome Sequencing; Spain; Polymorphism, Single Nucleotide; Microbial Sensitivity Tests; Bacterial Proteins; Genomics; Plasmids
PubMed: 38877167
DOI: 10.1007/s00284-024-03737-2 -
Journal of Bacteriology Jun 2024Many bacteria build elaborate molecular machines to import DNA via natural competence, yet this activity is often not identified until strains have been handled and...
UNLABELLED
Many bacteria build elaborate molecular machines to import DNA via natural competence, yet this activity is often not identified until strains have been handled and domesticated in laboratory settings. For example, one of the best studied Gram-positive model organisms, has a poorly transformable ancestor. Transformation in the ancestral strain is inhibited by a transmembrane peptide, ComI, which is encoded on an extrachromosomal plasmid. Although ComI was shown to be necessary and sufficient to inhibit transformation when produced at high levels under an inducible promoter, the mechanism by which ComI inhibits transformation is unknown. Here, we examine the native regulation and mechanism of transformation inhibition by ComI. We find that under native regulation, ComI expression is restricted in the absence of the plasmid. In the presence of the plasmid, we find that ComI is expressed at higher levels in cells that are differentiating into a competent state. The subcellular localization of ComI, however, does not depend on any other competence proteins, and permeabilization activity is concentration-dependent. Time-lapse microscopy reveals that competent cells producing ComI are first permeabilized and then die. Based on these observations, we propose a new model for the mechanism of ComI in which response to competence activation leads to selective elimination of the competent subpopulation.
IMPORTANCE
Natural transformation mechanisms have been studied across several bacterial systems, but few examples of inhibition exist. This work investigates the mechanism of action of a plasmid-encoded transmembrane inhibitor of natural transformation. The data reveal that the peptide can cause cell permeabilization. Permeabilization is synergistic with entry of into the "competent" state, such that cells with the ability to be transformed are preferentially killed. These findings reveal a self-preservation mechanism coupled to the physiological state of the cells that ensures that the population can maintain an unaltered plasmid and its predicted prophage.
PubMed: 38874341
DOI: 10.1128/jb.00413-23 -
Cell Host & Microbe Jun 2024Stress-induced prophages commonly "jump ship" by inducing lysis via the host SOS response. In a recent work, Uppalapati et al. reports an alternate, stress-selective...
Stress-induced prophages commonly "jump ship" by inducing lysis via the host SOS response. In a recent work, Uppalapati et al. reports an alternate, stress-selective strategy. Instead of promoting lysis, the Salmonella Gifsy-1 prophage arrests growth specifically when the SOS response coincides with oxidative stress.
Topics: Oxidative Stress; Prophages; SOS Response, Genetics; Salmonella Phages; Salmonella
PubMed: 38870894
DOI: 10.1016/j.chom.2024.05.010 -
Microbiology Resource Announcements Jun 2024We have revealed the genomic sequence of strain Hakim RU_CBWP isolated from pond surface water. Our assembled genome covers 3.787 Mb with 45.5629× coverage, showcasing...
We have revealed the genomic sequence of strain Hakim RU_CBWP isolated from pond surface water. Our assembled genome covers 3.787 Mb with 45.5629× coverage, showcasing an average GC content of 38.60%. This genome contains two CRISPR arrays, 17 prophages, 22 antibiotic resistance genes, and 20 virulence factor genes.
PubMed: 38864657
DOI: 10.1128/mra.00440-24 -
Environmental Microbiology Jun 2024The environmental bacterium, Pseudomonas putida, possesses a broad spectrum of metabolic pathways. This makes it highly promising for use in biotechnological production...
The environmental bacterium, Pseudomonas putida, possesses a broad spectrum of metabolic pathways. This makes it highly promising for use in biotechnological production as a cell factory, as well as in bioremediation strategies to degrade various aromatic pollutants. For P. putida to flourish in its environment, it must withstand the continuous threats posed by bacteriophages. Interestingly, until now, only a handful of phages have been isolated for the commonly used laboratory strain, P. putida KT2440, and no phage defence mechanisms have been characterized. In this study, we present a new Collection of Environmental P. putida Phages from Estonia, or CEPEST. This collection comprises 67 double-stranded DNA phages, which belong to 22 phage species and 9 phage genera. Our findings reveal that most phages in the CEPEST collection are more infectious at lower temperatures, have a narrow host range, and require an intact lipopolysaccharide for P. putida infection. Furthermore, we show that cryptic prophages present in the P. putida chromosome provide strong protection against the infection of many phages. However, the chromosomal toxin-antitoxin systems do not play a role in the phage defence of P. putida. This research provides valuable insights into the interactions between P. putida and bacteriophages, which could have significant implications for biotechnological and environmental applications.
Topics: Pseudomonas putida; Host Specificity; Prophages; Pseudomonas Phages; Estonia; Bacteriophages
PubMed: 38863081
DOI: 10.1111/1462-2920.16671 -
Journal of Applied Microbiology Jun 2024To investigate alternative resistance mechanisms among seven ceftazidime-avibactam (CZA)-resistant carbapenem-resistant Klebsiella pneumoniae (CRKP) strains lacking...
AIMS
To investigate alternative resistance mechanisms among seven ceftazidime-avibactam (CZA)-resistant carbapenem-resistant Klebsiella pneumoniae (CRKP) strains lacking common antimicrobial resistance genes (ARGs) using whole genome sequencing.
METHODS AND RESULTS
ARG and virulence factors (VFs) were screened using the ARG database CARD and the VF database, respectively, and identified using genomic annotation data with BLAST+. Six strains were ST11 sequence types (STs), and one was ST2123. ST11 strains harbored more ARGs than the ST2123 strains. All seven strains carried multiple ARGs with efflux-mediated antibiotic resistance, including oqxA, oqxB, tet (A), qacEdltal, CRP, H-NS, Kpn-E, F, G, H, acrA, LptD, acrB, acrD, cpxA, mdtB, and mdtC. These efflux-mediated ARGs were identified in most strains and even all strains. Whole genome sequencing revealed that the ST11 strain carried multiple potential prophages, genomic islands, and integrative and conjugative elements, while the ST2123 strain carried an independent potential prophages and a genomic island.
CONCLUSIONS
Whole genome sequencing analysis revealed that these seven CZA-resistant CRKP strains lacking common ARGs exhibited efflux-mediated antibiotic resistance-associated ARGs. The main mechanism by which CRKP resists CZA is antibiotic inactivation. Except for tet (A), no ARGs and validation experiments related to efflux were found. This study's results provide a new possibility for the resistance mechanism of CRKP to CZA, and we will verify this conclusion through experiments in the future.
Topics: Ceftazidime; Klebsiella pneumoniae; Azabicyclo Compounds; Whole Genome Sequencing; Drug Combinations; Anti-Bacterial Agents; Microbial Sensitivity Tests; Genome, Bacterial; Drug Resistance, Multiple, Bacterial; Humans; Carbapenem-Resistant Enterobacteriaceae; Klebsiella Infections; Carbapenems; Virulence Factors
PubMed: 38849309
DOI: 10.1093/jambio/lxae135 -
Antonie Van Leeuwenhoek Jun 2024Yersinia is an important genus comprising foodborne, zoonotic and pathogenic bacteria. On the other hand, species of the so-called group Yersinia enterocolitica-like are...
Yersinia is an important genus comprising foodborne, zoonotic and pathogenic bacteria. On the other hand, species of the so-called group Yersinia enterocolitica-like are understudied and mostly characterized as non-pathogenic, despite of some reports of human infections. The present study aimed to provide genomic insights of Yersinia frederiksenii (YF), Yersinia intermedia (YI) and Yersinia kristensenii (YK) isolated worldwide. A total of 22 YF, 20 YI and 14 YK genomes were searched for antimicrobial resistance genes, plasmids, prophages, and virulence factors. Their phylogenomic relatedness was analyzed by Gegenees and core-genome multi-locus sequence typing. Beta-lactam resistance gene bla and five plasmids replicons (pYE854, ColRNAI, ColE10, Col(pHAD28) and IncN3) were detected in less than five genomes. A total of 59 prophages, 106 virulence markers of the Yersinia genus, associated to adherence, antiphagocytosis, exoenzymes, invasion, iron uptake, proteases, secretion systems and the O-antigen, and virulence factors associated to other 20 bacterial genera were detected. Phylogenomic analysis revealed high inter-species distinction and four highly diverse YF clusters. In conclusion, the results obtained through the analyses of YF, YI and YK genomes suggest the virulence potential of these strains due to the broad diversity and high frequency of prophages and virulence factors found. Phylogenetic analyses were able to correctly distinguish these closely related species and show the presence of different genetic subgroups. These data contributed for a better understanding of YF, YI and YK virulence-associated features and global genetic diversity, and reinforced the need for better characterization of these Y. enterocolitica-like species considered non-pathogenic.
Topics: Yersinia; Phylogeny; Genome, Bacterial; Virulence Factors; Brazil; Yersinia Infections; Humans; Genomics; Prophages; Plasmids; Multilocus Sequence Typing; Virulence
PubMed: 38829455
DOI: 10.1007/s10482-024-01984-8 -
Environmental Pollution (Barking, Essex... May 2024Staphylococcus aureus thrives at animal-human-environment interfaces. A large-scale work from our group indicated that antimicrobial resistance (AMR) in commensal S....
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.
PubMed: 38825220
DOI: 10.1016/j.envpol.2024.124241 -
BMC Genomics Jun 2024Despite Spirochetales being a ubiquitous and medically important order of bacteria infecting both humans and animals, there is extremely limited information regarding...
BACKGROUND
Despite Spirochetales being a ubiquitous and medically important order of bacteria infecting both humans and animals, there is extremely limited information regarding their bacteriophages. Of the genus Treponema, there is just a single reported characterised prophage.
RESULTS
We applied a bioinformatic approach on 24 previously published Treponema genomes to identify and characterise putative treponemal prophages. Thirteen of the genomes did not contain any detectable prophage regions. The remaining eleven contained 38 prophage sequences, with between one and eight putative prophages in each bacterial genome. The prophage regions ranged from 12.4 to 75.1 kb, with between 27 and 171 protein coding sequences. Phylogenetic analysis revealed that 24 of the prophages formed three distinct sequence clusters, identifying putative myoviral and siphoviral morphology. ViPTree analysis demonstrated that the identified sequences were novel when compared to known double stranded DNA bacteriophage genomes.
CONCLUSIONS
In this study, we have started to address the knowledge gap on treponeme bacteriophages by characterising 38 prophage sequences in 24 treponeme genomes. Using bioinformatic approaches, we have been able to identify and compare the prophage-like elements with respect to other bacteriophages, their gene content, and their potential to be a functional and inducible bacteriophage, which in turn can help focus our attention on specific prophages to investigate further.
Topics: Prophages; Phylogeny; Treponema; Genomics; Genome, Bacterial; Computational Biology; Genome, Viral; Bacteriophages
PubMed: 38824509
DOI: 10.1186/s12864-024-10461-5 -
Methods in Molecular Biology (Clifton,... 2024Bacterial viruses (bacteriophages or phages) are the most abundant and diverse biological entities on Earth. There is a renewed worldwide interest in phage-centered... (Comparative Study)
Comparative Study
Bacterial viruses (bacteriophages or phages) are the most abundant and diverse biological entities on Earth. There is a renewed worldwide interest in phage-centered research motivated by their enormous potential as antimicrobials to cope with multidrug-resistant pathogens. An ever-growing number of complete phage genomes are becoming available, derived either from newly isolated phages (cultivated phages) or recovered from metagenomic sequencing data (uncultivated phages). Robust comparative analysis is crucial for a comprehensive understanding of genotypic variations of phages and their related evolutionary processes, and to investigate the interaction mechanisms between phages and their hosts. In this chapter, we present a protocol for phage comparative genomics employing tools selected out of the many currently available, focusing on complete genomes of phages classified in the class Caudoviricetes. This protocol provides accurate identification of similarities, differences, and patterns among new and previously known complete phage genomes as well as phage clustering and taxonomic classification.
Topics: Genome, Viral; Bacteriophages; Genomics; Phylogeny; Computational Biology; Metagenomics
PubMed: 38819567
DOI: 10.1007/978-1-0716-3838-5_14