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Nature Mar 2022Colibactin is a chemically unstable small-molecule genotoxin that is produced by several different bacteria, including members of the human gut microbiome. Although the...
Colibactin is a chemically unstable small-molecule genotoxin that is produced by several different bacteria, including members of the human gut microbiome. Although the biological activity of colibactin has been extensively investigated in mammalian systems, little is known about its effects on other microorganisms. Here we show that colibactin targets bacteria that contain prophages, and induces lytic development through the bacterial SOS response. DNA, added exogenously, protects bacteria from colibactin, as does expressing a colibactin resistance protein (ClbS) in non-colibactin-producing cells. The prophage-inducing effects that we observe apply broadly across different phage-bacteria systems and in complex communities. Finally, we identify bacteria that have colibactin resistance genes but lack colibactin biosynthetic genes. Many of these bacteria are infected with predicted prophages, and we show that the expression of their ClbS homologues provides immunity from colibactin-triggered induction. Our study reveals a mechanism by which colibactin production could affect microbiomes and highlights a role for microbial natural products in influencing population-level events such as phage outbreaks.
Topics: Bacteria; Bacterial Toxins; Bacteriolysis; Microbial Interactions; Peptides; Polyketides; Prophages; SOS Response, Genetics; Virus Activation
PubMed: 35197633
DOI: 10.1038/s41586-022-04444-3 -
Microbial Genomics Jan 2022The rapid emergence of multidrug-resistant is being driven largely by the spread of specific clonal groups (CGs). Of these, CG147 includes 7-gene multilocus sequence...
The rapid emergence of multidrug-resistant is being driven largely by the spread of specific clonal groups (CGs). Of these, CG147 includes 7-gene multilocus sequence typing (MLST) sequence types (STs) ST147, ST273 and ST392. CG147 has caused nosocomial outbreaks across the world, but its global population dynamics remain unknown. Here, we report a pandrug-resistant ST147 clinical isolate from India (strain DJ) and define the evolution and global emergence of CG147. Antimicrobial-susceptibility testing following European Committee on Antimicrobial Susceptibility Testing (EUCAST) guidelines and genome sequencing (Illumina and Oxford Nanopore Technologies, Unicycler assembly) were performed on strain DJ. Additionally, we collated 217 publicly available CG147 genomes [National Center for Biotechnology Information (NCBI), May 2019]. CG147 evolution was inferred within a temporal phylogenetic framework (beast) based on a recombination-free sequence alignment (Roary/Gubbins). Comparative genomic analyses focused on resistance and virulence genes and other genetic elements (BIGSdb, Kleborate, PlasmidFinder, phaster, ICEfinder and CRISPRCasFinder). Strain DJ had a pandrug-resistance phenotype. Its genome comprised the chromosome, seven plasmids and one linear phage-plasmid. Four carbapenemase genes were detected: and two copies of in the chromosome, and a second copy of on an 84 kb IncFII plasmid. CG147 genomes carried a mean of 13 acquired resistance genes or mutations; 63 % carried a carbapenemase gene and 83 % harboured . All CG147 genomes presented GyrA and ParC mutations and a common subtype I-E CRISPR-Cas system. ST392 and ST273 emerged in 2005 and 1995, respectively. ST147, the most represented phylogenetic branch, was itself divided into two main clades with distinct capsular loci: KL64 (74 %, DJ included, emerged in 1994 and disseminated worldwide, with carbapenemases varying among world regions) and KL10 (20 %, emerged in 2002, predominantly found in Asian countries, associated with carbapenemases NDM and OXA-48-like). Furthermore, subclades within ST147-KL64 differed at the yersiniabactin locus, OmpK35/K36 mutations, plasmid replicons and prophages. The absence of IncF plasmids in some subclades was associated with a possible activity of a CRISPR-Cas system. CG147 comprises pandrug-resistant or extensively resistant isolates, and carries multiple and diverse resistance genes and mobile genetic elements, including chromosomal . Its emergence is being driven by the spread of several phylogenetic clades marked by their own genomic features and specific temporo-spatial dynamics. These findings highlight the need for precision surveillance strategies to limit the spread of particularly concerning CG147 subsets.
Topics: Drug Resistance, Multiple, Bacterial; Evolution, Molecular; Genome, Bacterial; Genomics; India; Interspersed Repetitive Sequences; Klebsiella pneumoniae; Microbial Sensitivity Tests; Multigene Family; Phylogeny; Plasmids; Prophages; Virulence Factors; Whole Genome Sequencing
PubMed: 35019836
DOI: 10.1099/mgen.0.000737 -
MBio Feb 2024Bacteriophages are large and diverse components of the biosphere, and many phages are temperate. Upon infection, temperate phages can establish lysogeny in which a...
Bacteriophages are large and diverse components of the biosphere, and many phages are temperate. Upon infection, temperate phages can establish lysogeny in which a prophage is typically integrated into the bacterial chromosome. Here, we describe the phenomenon of tRNA-dependent lysogeny, a previously unrecognized behavior of some temperate phages. tRNA-dependent lysogeny is characterized by two unusual features. First, a phage-encoded tyrosine family integrase mediates site-specific recombination between a phage site and a bacterial site overlapping a host tRNA gene. However, and share only a short (~10 bp) common core such that a functional tRNA is not reconstructed upon integration. Second, the phage encodes a tRNA of the same isotype as the disrupted but essential host tRNA, complementing its loss, and consequently is required for the survival of lysogenic progeny. As expected, an integrase-defective phage mutant forms turbid plaques, and bacterial progeny are immune to superinfection, but they lack stability, and the prophage is rapidly lost. In contrast, a tRNA-defective phage mutant forms clear plaques and more closely resembles a repressor mutant, and lysogens are recovered only at very low frequency through the use of secondary attachment sites elsewhere in the host genome. Integration-proficient plasmids derived from these phages must also carry a cognate phage tRNA gene for efficient integration, and these may be useful tools for mycobacterial genetics. We show that tRNA-dependent lysogeny is used by phages within multiple different groups of related viruses and may be prevalent elsewhere in the broader phage community.IMPORTANCEBacteriophages are the most numerous biological entities in the biosphere, and a substantial proportion of phages are temperate, forming stable lysogens in which a prophage copy of the genome integrates into the bacterial chromosome. Many phages encode a variety of tRNA genes whose roles are poorly understood, although it has been proposed that they enhance translational efficiencies in lytic growth or that they counteract host defenses that degrade host tRNAs. Here, we show that phage-encoded tRNAs play key roles in the establishment of lysogeny of some temperate phages. They do so by compensating for the loss of tRNA function when phages integrate at an site overlapping a tRNA gene but fail to reconstruct the tRNA at the attachment junction. In this system of tRNA-dependent lysogeny, the phage-encoded tRNA is required for lysogeny, and deletion of the phage tRNA gives rise to a clear plaque phenotype and obligate lytic growth.
Topics: Lysogeny; Bacteriophages; Prophages; Integrases; Plasmids
PubMed: 38236026
DOI: 10.1128/mbio.03260-23 -
MBio Oct 2022The formation of membrane vesicles (MVs) by Gram-positive bacteria has gained increasing attention over the last decade. Recently, models of vesicle formation have been...
The formation of membrane vesicles (MVs) by Gram-positive bacteria has gained increasing attention over the last decade. Recently, models of vesicle formation have been proposed and involve the digestion of the cell wall by prophage-encoded or stress-induced peptidoglycan (PG) hydrolases and the inhibition of PG synthesis by β-lactam antibiotics. The impact of these mechanisms on vesicle formation is largely dependent on the strain and growth conditions. To date, no information on the production of vesicles by the lactobacilli family has been reported. Here, we aimed to characterize the MVs released by the Gram-positive bacteria BL23 and also investigated the mechanisms involved in vesicle formation. Using electron microscopy, we established that the size of the majority of BL23 vesicles ranged from 50 to 100 nm. Furthermore, we showed that the vesicles were released consistently throughout the growth of the bacteria in standard culture conditions. The protein composition of the vesicles released in the supernatant was identified and a significant number of prophage proteins was detected. Moreover, using a mutant strain harboring a defective PLE2 prophage, we were able to show that the spontaneous and mitomycin-triggered induction of the prophage PLE2 contribute to the production of MVs by BL23. Finally, we also demonstrated the influence of prophages on the membrane integrity of bacteria. Overall, our results suggest a key role of the prophage PLE2 in the production of MVs by BL23 in the absence or presence of genotoxic stress. The last few decades have demonstrated that membrane vesicles (MVs) produced by microorganisms can have a wide variety of functions. This diversity places MVs at the crossroads of major research topics in current microbiology such as antibiotic resistance, horizontal gene transfer, cell communication, biofilm development, bacteriophage resistance, and pathogenesis. In particular, vesicles produced by probiotic strains have been shown to play a significant role in their beneficial effects. Thus, the study of vesicle biogenesis is a key element for promoting and improving their release. Overall, our results suggest a key role of spontaneous and mitomycin-triggered prophage induction in MV production by the Gram-positive bacteria BL23. This phenomenon is of great interest as prophage-induced MVs could potentially influence bacterial behavior, stress resistance, and vesicle functions.
Topics: Peptidoglycan; Virus Activation; Lacticaseibacillus casei; Prophages; N-Acetylmuramoyl-L-alanine Amidase; Anti-Bacterial Agents; Mitomycins; beta-Lactams
PubMed: 36200778
DOI: 10.1128/mbio.02375-22 -
International Journal of Molecular... May 2023is a common human commensal and opportunistic pathogen. In recent years, the clinical isolation and resistance rates of have shown a yearly increase, leading to a...
is a common human commensal and opportunistic pathogen. In recent years, the clinical isolation and resistance rates of have shown a yearly increase, leading to a special interest in mobile genetic elements. Prophages are a representative class of mobile genetic elements that can carry host-friendly genes, transfer horizontally between strains, and coevolve with the host's genome. In this study, we identified 15,946 prophages from the genomes of 1437 fully assembled deposited in the NCBI database, with 9755 prophages on chromosomes and 6191 prophages on plasmids. We found prophages to be notably diverse and widely disseminated in the genomes. The prophages encoded multiple putative virulence factors and antibiotic resistance genes. The comparison of strain types with prophage types suggests that the two may be related. The differences in GC content between the same type of prophages and the genomic region in which they were located indicates the alien properties of the prophages. The overall distribution of GC content suggests that prophages integrated on chromosomes and plasmids may have different evolutionary characteristics. These results suggest a high prevalence of prophages in the genome and highlight the effect of prophages on strain characterization.
Topics: Humans; Prophages; Klebsiella pneumoniae; Plasmids; Genomics; Virulence Factors; Anti-Bacterial Agents; Genome, Bacterial
PubMed: 37298067
DOI: 10.3390/ijms24119116 -
Nature Communications Oct 2023Bacteriophages (phages) are the most abundant biological entities on Earth, exerting a significant influence on the dissemination of bacterial virulence, pathogenicity,...
Bacteriophages (phages) are the most abundant biological entities on Earth, exerting a significant influence on the dissemination of bacterial virulence, pathogenicity, and antimicrobial resistance. Temperate phages integrate into the bacterial chromosome in a dormant state through intricate regulatory mechanisms. These mechanisms repress lytic genes while facilitating the expression of integrase and the CI master repressor. Upon bacterial SOS response activation, the CI repressor undergoes auto-cleavage, producing two fragments with the N-terminal domain (NTD) retaining significant DNA-binding ability. The process of relieving CI NTD repression, essential for prophage induction, remains unknown. Here we show a specific interaction between the ClpX protease and CI NTD repressor fragment of phages Ф11 and 80α in Staphylococcus aureus. This interaction is necessary and sufficient for prophage activation after SOS-mediated CI auto-cleavage, defining the final stage in the prophage induction cascade. Our findings unveil unexpected roles of bacterial protease ClpX in phage biology.
Topics: Virus Activation; Staphylococcus aureus; Peptide Hydrolases; Bacteriophages; Prophages
PubMed: 37852980
DOI: 10.1038/s41467-023-42413-0 -
Viruses Nov 2021Prophage 919TP is widely distributed among and is induced to produce free φ919TP phage particles. However, the interactions between prophage φ919TP, the induced phage...
Prophage 919TP is widely distributed among and is induced to produce free φ919TP phage particles. However, the interactions between prophage φ919TP, the induced phage particle, and its host remain unknown. In particular, phage resistance mechanisms and potential fitness trade-offs, resulting from phage resistance, are unresolved. In this study, we examined a prophage 919TP-deleted variant of and its interaction with a modified lytic variant of the induced prophage (φ919TP ). Specifically, the phage-resistant mutant was isolated by challenging a prophage-deleted variant with lytic phage φ919TP . Further, the comparative genomic analysis of wild-type and φ919TP -resistant mutant predicted that phage φ919TP selects for phage-resistant mutants harboring a mutation in key steps of lipopolysaccharide (LPS) O-antigen biosynthesis, causing a single-base-pair deletion in gene . Our study showed that the -mediated O-antigen defect can cause pleiotropic phenotypes, e.g., cell autoaggregation and reduced swarming motility, emphasizing the role of phage-driven diversification in . The developed approach assists in the identification of genetic determinants of host specificity and is used to explore the molecular mechanism underlying phage-host interactions. Our findings contribute to the understanding of prophage-facilitated horizontal gene transfer and emphasize the potential for developing new strategies to optimize the use of phages in bacterial pathogen control.
Topics: Bacterial Proteins; Cholera; Host Microbial Interactions; Host Specificity; Lysogeny; O Antigens; Prophages; Vibrio cholerae O1; Virus Activation
PubMed: 34960610
DOI: 10.3390/v13122342 -
MSphere Aug 2021Prophages are often involved in host survival strategies and contribute toward increasing the genetic diversity of the host genome. Prophages also drive horizontal...
Prophages are often involved in host survival strategies and contribute toward increasing the genetic diversity of the host genome. Prophages also drive horizontal propagation of various genes as vehicles. However, there are few retrospective studies contributing to the propagation of antimicrobial resistance (AMR) and virulence factor (VF) genes by prophage. We extracted the complete genome sequences of seven pathogens, including ESKAPE bacteria and Escherichia coli from a public database, and examined the distribution of both the AMR and VF genes in prophage-like regions. We found that the ratios of AMR and VF genes greatly varied among the seven species. More than 70% of Enterobacter cloacae strains had VF genes, but only 1.2% of Klebsiella pneumoniae strains had VF genes from prophages. AMR and VF genes are unlikely to exist together in the same prophage region except in and , and the distribution patterns of prophage types containing AMR genes are distinct from those of VF gene-carrying prophage types. AMR genes in the prophage were located near transposase and/or integrase. The prophage containing class 1 integrase possessed a significantly greater number of AMR genes than did prophages with no class 1 integrase. The results of this study present a comprehensive picture of AMR and VF genes present within, or close to, prophage-like elements and different prophage patterns between AMR- or VF-encoding prophage-like elements. Although we believe phages play an important role in horizontal gene transfer in exchanging genetic material, we do not know the distribution of the antimicrobial resistance (AMR) and/or virulence factor (VF) genes in prophages. We collected different prophage elements from the complete genome sequences of seven species-Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, Enterobacter cloacae, and Escherichia coli-and characterized the distribution of antimicrobial resistance and virulence genes located in the prophage region. While virulence genes in prophage were species specific, antimicrobial resistance genes in prophages were highly conserved in various species. An integron structure was detected within specific prophage regions such as P1-like prophage element. Maximum of 10 antimicrobial resistance genes were found in a single prophage region, suggesting that prophages act as a reservoir for antimicrobial resistance genes. The results of this study show the different characteristic structures between AMR- or VF-encoding prophages.
Topics: Anti-Bacterial Agents; Bacteria; Cross Infection; Drug Resistance, Bacterial; Enterobacter cloacae; Escherichia coli; Genome, Bacterial; Humans; Klebsiella pneumoniae; Prophages; Virulence; Virulence Factors
PubMed: 34232073
DOI: 10.1128/mSphere.00452-21 -
Infection, Genetics and Evolution :... Aug 2021The Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) is a bacterial defense mechanism against bacteriophages composed of two different parts: the...
The Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) is a bacterial defense mechanism against bacteriophages composed of two different parts: the CRISPR array and the Cas genes. The spacer acquisition is done by the adaptation module consisting of the hallmark Cas1 Cas2 proteins, which inserts new spacers into the CRISPR array. Here we aimed to describe the CRISPR-Cas system in Proteus mirabilis (P. mirabilis) isolates. CRISPR loci was observed in 30 genomic contents of 109 P. mirabilis isolates that each locus was consisted of two CRISPR arrays and each array had a different preserved leader sequences. Only the type I-E CRISPR-Cas system was common in these isolates. The source of the spacers was identified, including phages and prophages. CRISPR spacer origin analysis also identified a conserved PAM sequence of 5'-AAG-3' nucleotide stretch. Through collecting spacers, CRISPR arrays of P. mirabilis isolates were expanded mostly by integration of bacteriophageal source of spacers. This study shows novel findings in the area of the P-mirabilis CRISPR-Cas system. In this regard, among analyzed genome of P. mirabilis isolates, Class I CRISR-Cas systems were dominant, and all belonged to type I-E. In the flanks of the CRISPR, some other elements with regulatory role were also found. A motif of 11 nt size was found to be preserved among the analyzed genome. We believe that it might has a CRISPR-Cas system transcription facilitator by targeting the Rho element.
Topics: Bacteriophages; CRISPR-Cas Systems; Prophages; Proteus mirabilis
PubMed: 33905883
DOI: 10.1016/j.meegid.2021.104881 -
Nucleic Acids Research Dec 2021The life cycle of temperate phages includes a lysogenic cycle stage when the phage integrates into the host genome and becomes a prophage. However, the identification of...
The life cycle of temperate phages includes a lysogenic cycle stage when the phage integrates into the host genome and becomes a prophage. However, the identification of prophages that are highly divergent from known phages remains challenging. In this study, by taking advantage of the lysis-lysogeny switch of temperate phages, we designed Prophage Tracer, a tool for recognizing active prophages in prokaryotic genomes using short-read sequencing data, independent of phage gene similarity searching. Prophage Tracer uses the criterion of overlapping split-read alignment to recognize discriminative reads that contain bacterial (attB) and phage (attP) att sites representing prophage excision signals. Performance testing showed that Prophage Tracer could predict known prophages with precise boundaries, as well as novel prophages. Two novel prophages, dsDNA and ssDNA, encoding highly divergent major capsid proteins, were identified in coral-associated bacteria. Prophage Tracer is a reliable data mining tool for the identification of novel temperate phages and mobile genetic elements. The code for the Prophage Tracer is publicly available at https://github.com/WangLab-SCSIO/Prophage_Tracer.
Topics: Animals; Anthozoa; Bacteria; Genome, Archaeal; Genome, Bacterial; Interspersed Repetitive Sequences; Prophages; Sequence Alignment; Sequence Analysis, DNA; Software
PubMed: 34551431
DOI: 10.1093/nar/gkab824