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Prophages are prevalent among bacterial species, including strains carrying antibiotic resistance genes (ARGs). Prophage induction can be triggered by the SOS response to stressors, leading to cell lysis. In environments polluted by chemical stressors, ARGs and prophage co-harboring strains might pose an unknown risk of spreading ARGs through chemical pollutant-mediated prophage induction and subsequent cell lysis. In this study, we investigated the effects of common non-antibiotic water pollutants, triclosan and silver nanoparticles, on triggering prophage induction in clinical isolates carrying ARGs and the subsequent uptake of released ARGs by the naturally competent bacterium Acinetobacter baylyi. Our results demonstrate that both triclosan and silver nanoparticles, at environmentally relevant concentrations and those found in commercial products, significantly enhance prophage induction among various clinical isolates. Transmission electron microscopy imaging and plaque assays confirmed the production of infectious phage particles under non-antibiotic pollutants-mediated prophage induction. In addition, the rate of ARG transformation to A. baylyi significantly increased after the release of extracellular ARGs from prophage induction-mediated cell lysis. The mechanism of non-antibiotic pollutants-mediated prophage induction is primarily associated with excessive oxidative stress, which provokes the SOS response. Our findings offer insights into the role of non-antibiotic pollutants in promoting the dissemination of ARGs by triggering prophage induction.

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Natural and chemically modified polysaccharides are extensively employed across a wide array of industries, leading to their prevalence in the waste streams of industrialized societies. With projected increasing demand, a pressing challenge is to swiftly assess and predict their biodegradability to inform the development of new sustainable materials. In this study, we developed a scalable method to evaluate polysaccharide breakdown by measuring microbial growth and analyzing microbial genomes. Our approach, applied to polysaccharides with various structures, correlates strongly with well-established regulatory methods based on oxygen demand. We show that modifications to the polysaccharide structure decreased degradability and favored the growth of microbes adapted to break down chemically modified sugars. More broadly, we discovered two main types of microbial communities associated with different polysaccharide structures─one dominated by fast-growing microbes and another by specialized degraders. Surprisingly, we were able to predict biodegradation rates based only on two genomic features that define these communities: the abundance of genes related to rRNA (indicating fast growth) and the abundance of glycoside hydrolases (enzymes that break down polysaccharides), which together predict nearly 70% of the variation in polysaccharide breakdown. This suggests a trade-off, whereby microbes are either adapted for fast growth or for degrading complex polysaccharide chains, but not both. Finally, we observe that viral elements (prophages) encoded in the genomes of degrading microbes are induced in easily degradable polysaccharides, leading to complex dynamics in biomass accumulation during degradation. In summary, our work provides a practical approach for efficiently assessing polymer degradability and offers genomic insights into how microbes break down polysaccharides.

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Bioelectrochemical systems (BESs) exploit electroactive biofilms (EABs) for promising applications in biosensing, wastewater treatment, energy production, and chemical biosynthesis. However, during the operation of BESs, EABs inevitably decay. Seeking approaches to rejuvenate decayed EABs is critical for the sustainability and practical application of BESs. Prophage induction has been recognized as the primary reason for EAB decay. Herein, we report that introducing a competitive species of Geobacter uraniireducens suspended prophage induction in Geobacter sulfurreducens and thereby rejuvenated the decayed G. sulfurreducens EAB. The transcriptomic profile of G. sulfurreducens demonstrated that the addition of G. uraniireducens significantly affected the expression of metabolism- and stress response system-related genes and in particular suppressed the induction of phage-related genes. Mechanistic analyses revealed that interspecies ecological competition exerted by G. uraniireducens suppressed prophage induction. Our findings not only reveal a novel strategy to rejuvenate decayed EABs, which is significant for the sustainability of BESs, but also provide new knowledge for understanding phage-host interactions from an ecological perspective, with implications for developing therapies to defend against phage attack.

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Infections caused by carbapenem-resistant Acinetobacter baumannii (CRAB) present significant healthcare challenges due to limited treatment options. Bacteriophage (phage) therapy offers potential as an alternative treatment. However, the high host specificity of phages poses challenges for their therapeutic application. To broaden the phage spectrum, laboratory-based phage training using the Appelmans protocol was employed in this study. As a result, the protocol successfully expanded the host range of a phage cocktail targeting CRAB. Further analysis revealed that the expanded host range phages isolated from the output cocktail were identified as recombinant derivatives originating from prophages induced from encountered bacterial strains. These findings provide valuable genetic insights into the protocol's mechanism when applied to phages infecting A. baumannii strains that have never been investigated before. However, it is noteworthy that the expanded host range phages obtained from this protocol exhibited limited stability, raising concerns about their suitability for therapeutic purposes.

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Shiga toxin-producing Escherichia coli (STEC) are foodborne pathogens implicated in diseases including hemolytic uremic syndrome (HUS) and hemorrhagic colitis (HC). The main virulence factor are Shiga toxins; their production and secretion are by-products of the expression of late genes of prophages upon sub-lethal environmental stimuli exposure. Hence, the lysogenic prophage after a stress switch to lytic cycle spreading the Stx phages. In the present study, 35 STEC were screened for the presence and the ability to release Shiga toxin-encoding bacteriophages. Three bacterial strains showed signals of prophage presence both in plate and in PCR. Subsequently, these bacterial strains were subjected to stressors that simulate cheese manufacturing conditions: NaCl (1, 1.5 and 2% w/v), lactic acid (0.5, 1.5 and 3% v/v), anaerobic growth, pasteurization (72°C for 15 s), UV irradiation. The ability to release prophage was evaluated by Real Time qPCR. Induction of the prophages showed that the addition of NaCl at 1.5 and 2% significantly increased viral release compared to control. Conversely, the addition of lactic acid had a significant repressive effect. The other applied stressors had no significant effect in phage release according to the experimental conditions adopted.

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Achromobacter species colonization of Cystic Fibrosis respiratory airways is an increasing concern. Two adult patients with Cystic Fibrosis colonized by Achromobacter xylosoxidans CF418 or Achromobacter ruhlandii CF116 experienced fatal exacerbations. Achromobacter spp. are naturally resistant to several antibiotics. Therefore, phages could be valuable as therapeutics for the control of Achromobacter. In this study, thirteen lytic phages were isolated and characterized at the morphological and genomic levels for potential future use in phage therapy. They are presented here as the Achromobacter Kumeyaay phage collection. Six distinct Achromobacter phage genome clusters were identified based on a comprehensive phylogenetic analysis of the Kumeyaay collection as well as the publicly available Achromobacter phages. The infectivity of all phages in the Kumeyaay collection was tested in 23 Achromobacter clinical isolates; 78% of these isolates were lysed by at least one phage. A cryptic prophage was induced in Achromobacter xylosoxidans CF418 when infected with some of the lytic phages. This prophage genome was characterized and is presented as Achromobacter phage CF418-P1. Prophage induction during lytic phage preparation for therapy interventions require further exploration. Large-scale production of phages and removal of endotoxins using an octanol-based procedure resulted in a phage concentrate of 1 × 109 plaque-forming units per milliliter with an endotoxin concentration of 65 endotoxin units per milliliter, which is below the Food and Drugs Administration recommended maximum threshold for human administration. This study provides a comprehensive framework for the isolation, bioinformatic characterization, and safe production of phages to kill Achromobacter spp. in order to potentially manage Cystic Fibrosis (CF) pulmonary infections.

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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 ß-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 Lacticaseibacillus casei BL23 and also investigated the mechanisms involved in vesicle formation. Using electron microscopy, we established that the size of the majority of L. casei 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 L. casei 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 L. casei BL23 in the absence or presence of genotoxic stress. IMPORTANCE 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 Lacticaseibacillus casei BL23. This phenomenon is of great interest as prophage-induced MVs could potentially influence bacterial behavior, stress resistance, and vesicle functions.

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Phages are viruses that infect bacteria. They affect various microbe-mediated processes that drive biogeochemical cycling on a global scale. Their influence depends on whether the infection is lysogenic or lytic. Temperate phages have the potential to execute both infection types and thus frequently switch their infection modes in nature, potentially causing substantial impacts on the host-phage community and relevant biogeochemical cycling. Understanding the regulating factors and outcomes of temperate phage life cycle transition is thus fundamental for evaluating their ecological impacts. This review thus systematically summarizes the effects of various factors affecting temperate phage life cycle decisions in both culturable phage-host systems and natural environments. The review further elucidates the ecological implications of the life cycle transition of temperate phages with an emphasis on phage/host fitness, host-phage dynamics, microbe diversity and evolution, and biogeochemical cycles.

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Antibiotic resistance is rapidly spreading via the horizontal transfer of resistance genes in mobile genetic elements. While plasmids are key drivers of this process, few integrative phages encode antibiotic resistance genes. Here, we find that phage-plasmids, elements that are both phages and plasmids, often carry antibiotic resistance genes. We found 60 phage-plasmids with 184 antibiotic resistance genes, providing resistance for broad-spectrum-cephalosporins, carbapenems, aminoglycosides, fluoroquinolones, and colistin. These genes are in a few hot spots, seem to have been cotranslocated with transposable elements, and are often in class I integrons, which had not been previously found in phages. We tried to induce six phage-plasmids with resistance genes (including four with resistance integrons) and succeeded in five cases. Other phage-plasmids and integrative prophages were coinduced in these experiments. As a proof of concept, we focused on a P1-like element encoding an extended spectrum ß-lactamase, blaCTX-M-55. After induction, we confirmed that it is capable of infecting and converting four other E. coli strains. Its reinduction led to the further conversion of a sensitive strain, confirming that it is a fully functional phage. This study shows that phage-plasmids carry a large diversity of clinically relevant antibiotic resistance genes that they can transfer across bacteria. As plasmids, these elements seem plastic and capable of acquiring genes from other plasmids. As phages, they may provide novel paths of transfer for resistance genes because they can infect bacteria that are distant in time and space from the original host. As a matter of alarm, they may also mediate transfer to other types of phages. IMPORTANCE The dissemination of antimicrobial resistance is a major threat to global health. Here, we show that a group of temperate bacterial viruses (phages), termed phage-plasmids, commonly encode different and multiple types of resistance genes of high clinical importance, often in integrons. This is unexpected, as phages typically do not carry resistance genes and, hence, do not confer upon their hosts resistance via infection and genome integration. Our experiments with phage-plasmids isolated from clinical settings confirmed that they infect sensitive strains and render them antibiotic resistant. The spread of antibiotic resistance genes by phage-plasmids is worrisome because it dispenses cell-to-cell contact, which is necessary for canonical plasmid transfer (conjugation). Furthermore, their integrons become genetic platforms for the acquisition of novel resistance genes.

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Viruses are key actors of ecosystems and have major impacts on global biogeochemical cycles. Prophages deserve particular attention as they are ubiquitous in bacterial genomes and can enter a lytic cycle when triggered by environmental conditions. We explored how temperature affects the interactions between prophages and other biological levels using an opportunistic pathogen, the bacterium Serratia marcescens, which harbours several prophages and that had undergone an evolution experiment under several temperature regimes. We found that the release of one of the prophages was temperature-sensitive and malleable to evolutionary changes. We further discovered that the virulence of the bacterium in an insect model also evolved and was positively correlated with phage release rates. We determined through analysis of genetic and epigenetic data that changes in the bacterial outer cell wall structure possibly explain this phenomenon. We hypothezise that the temperature-dependent phage release rate acted as a selection pressure on S. marcescens and that it resulted in modified bacterial virulence in the insect host. Our study system illustrates how viruses can mediate the influence of abiotic environmental changes to other biological levels and thus be involved in ecosystem feedback loops.

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Viruses are ubiquitously distributed in the marine environment, influencing microbial population dynamics and biogeochemical cycles on a large scale. Due to their small size, they fall into the oceanographic size-class definition of dissolved organic matter (DOM; <0.7 µm). The purpose of our study was to investigate if there is a detectable imprint of virus particles in natural DOM following standard sample preparation and molecular analysis routines using ultrahigh-resolution mass spectrometry (FT-ICR-MS). Therefore, we tested if a molecular signature deriving from virus particles can be detected in the DOM fingerprint of a bacterial culture upon prophage induction and of seawater containing the natural microbial community. Interestingly, the virus-mediated lysate of the infected bacterial culture differed from the cell material of a physically disrupted control culture in its molecular composition. Overall, a small subset of DOM compounds correlated significantly with virus abundances in the bacterial culture setup, accounting for <1% of the detected molecular formulae and <2% of the total signal intensity of the DOM dataset. These were phosphorus- and nitrogen-containing compounds and they were partially also detected in DOM samples from other studies that included high virus abundances. While some of these formulae matched with typical biomolecules that are constituents of viruses, others matched with bacterial cell wall components. Thus, the identified DOM molecular formulae were probably not solely derived from virus particles but were partially also derived from processes such as the virus-mediated bacterial cell lysis. Our results indicate that a virus-derived DOM signature is part of the natural DOM and barely detectable within the analytical window of ultrahigh-resolution mass spectrometry when a high natural background is present.

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Mobile genetic elements (MGEs) drive bacterial evolution, alter gene availability within microbial communities, and facilitate adaptation to ecological niches. In natural systems, bacteria simultaneously possess or encounter multiple MGEs, yet their combined influences on microbial communities are poorly understood. Here, we investigate interactions among MGEs in the marine bacterium Sulfitobacter pontiacus. Two related strains, CB-D and CB-A, each harbor a single prophage. These prophages share high sequence identity with one another and an integration site within the host genome, yet these strains exhibit differences in "spontaneous" prophage induction (SPI) and consequent fitness. To better understand mechanisms underlying variation in SPI between these lysogens, we closed their genomes, which revealed that in addition to harboring different prophage genotypes, CB-A lacks two of the four large, low-copy-number plasmids possessed by CB-D. To assess the relative roles of plasmid content versus prophage genotype on host physiology, a panel of derivative strains varying in MGE content were generated. Characterization of these derivatives revealed a robust link between plasmid content and SPI, regardless of prophage genotype. Strains possessing all four plasmids had undetectable phage in cell-free lysates, while strains lacking either one plasmid (pSpoCB-1) or a combination of two plasmids (pSpoCB-2 and pSpoCB-4) produced high (>105 PFU/mL) phage titers. Homologous plasmid sequences were identified in related bacteria, and plasmid and phage genes were found to be widespread in Tara Oceans metagenomic data sets. This suggests that plasmid-dependent stabilization of prophages may be commonplace throughout the oceans. IMPORTANCE The consequences of prophage induction on the physiology of microbial populations are varied and include enhanced biofilm formation, conferral of virulence, and increased opportunity for horizontal gene transfer. These traits lead to competitive advantages for lysogenized bacteria and influence bacterial lifestyles in a variety of niches. However, biological controls of "spontaneous" prophage induction, the initiation of phage replication and phage-mediated cell lysis without an overt stressor, are not well understood. In this study, we observed a novel interaction between plasmids and prophages in the marine bacterium Sulfitobacter pontiacus. We found that loss of one or more distinct plasmids-which we show carry genes ubiquitous in the world's oceans-resulted in a marked increase in prophage induction within lysogenized strains. These results demonstrate cross talk between different mobile genetic elements and have implications for our understanding of the lysogenic-lytic switches of prophages found not only in marine environments, but throughout all ecosystems.

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Leuconostoc has been used as a principal starter in natural kimchi fermentation, but limited research has been conducted on its phages. In this study, prophage distribution and characterization in kimchi-derived Leuconostoc strains were investigated, and phage induction was performed. Except for one strain, 16 Leuconostoc strains had at least one prophage region with questionable and incomplete regions, which comprised 0.5-6.0% of the bacterial genome. Based on major capsid protein analysis, ten intact prophages and an induced incomplete prophage of Leu. lactis CBA3626 belonged to the Siphoviridae family and were similar to Lc-Nu-like, sha1-like, phiMH1-like, and TPA_asm groups. Bacterial immunology genes, such as superinfection exclusion proteins and methylase, were found on several prophages. One prophage of Leu. lactis CBA3626 was induced using mitomycin C and was confirmed as belonging to the Siphoviridae family. Homology of the induced prophage with 21 reported prophages was not high (< 4%), and 47% identity was confirmed only with TPA_asm from Siphoviridae sp. isolate ct3pk4. Therefore, it is suggested that Leuconostoc from kimchi had diverse prophages with less than 6% genome proportion and some immunological genes. Interestingly, the induced prophage was very different from the reported prophages of other Leuconostoc species.

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The toxin-antitoxin (TA) genetic modules control various bacterial events, such as plasmid maintenance, persister cell formation, and phage defense. They also exist in mobile genetic elements, including prophages; however, their physiological roles remain poorly understood. Here, we demonstrate that hokW-sokW, a putative TA locus encoded in Sakai prophage 5 (Sp5) in enterohemorrhagic Escherichia coli O157: H7 Sakai strain, functions as a type I TA system. Bacterial growth assays showed that the antitoxic activity of sokW RNA against HokW toxin partially requires an endoribonuclease, RNase III, and an RNA chaperone, Hfq. We also demonstrated that hokW-sokW assists Sp5-mediated lysis of E. coli cells when prophage induction is promoted by the DNA-damaging agent mitomycin C (MMC). We found that MMC treatment diminished sokW RNA and increased both the expression level and inner membrane localization of HokW in a RecA-dependent manner. Remarkably, the number of released Sp5 phages decreased by half in the absence of hokW-sokW. These results suggest that hokW-sokW plays a novel role as a TA system that facilitates the release of Sp5 phage progeny through E. coli lysis.

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By targeting key regulatory hubs of their host, bacteriophages represent a powerful source for the identification of novel antimicrobial proteins. Here, a screening of small cytoplasmic proteins encoded by the CGP3 prophage of Corynebacterium glutamicum resulted in the identification of the gyrase-inhibiting protein Cg1978, termed Gip. Pull-down assays and surface plasmon resonance revealed a direct interaction of Gip with the gyrase subunit A (GyrA). The inhibitory activity of Gip was shown to be specific to the DNA gyrase of its bacterial host C. glutamicum. Overproduction of Gip in C. glutamicum resulted in a severe growth defect as well as an induction of the SOS response. Furthermore, reporter assays revealed an RecA-independent induction of the cryptic CGP3 prophage, most likely caused by topological alterations. Overexpression of gip was counteracted by an increased expression of gyrAB and a reduction of topA expression at the same time, reflecting the homeostatic control of DNA topology. We postulate that the prophage-encoded Gip protein plays a role in modulating gyrase activity to enable efficient phage DNA replication. A detailed elucidation of the mechanism of action will provide novel directions for the design of drugs targeting DNA gyrase.

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BACKGROUND: Temperate phages influence the density, diversity and function of bacterial populations. Historically, they have been described as carriers of toxins. More recently, they have also been recognised as direct modulators of the gut microbiome, and indirectly of host health and disease. Despite recent advances in studying prophages using non-targeted sequencing approaches, methodological challenges in identifying inducible prophages in bacterial genomes and quantifying their activity have limited our understanding of prophage-host interactions. RESULTS: We present methods for using high-throughput sequencing data to locate inducible prophages, including those previously undiscovered, to quantify prophage activity and to investigate their replication. We first used the well-established Salmonella enterica serovar Typhimurium/p22 system to validate our methods for (i) quantifying phage-to-host ratios and (ii) accurately locating inducible prophages in the reference genome based on phage-to-host ratio differences and read alignment alterations between induced and non-induced prophages. Investigating prophages in bacterial strains from a murine gut model microbiota known as Oligo-MM12 or sDMDMm2, we located five novel inducible prophages in three strains, quantified their activity and showed signatures of lateral transduction potential for two of them. Furthermore, we show that the methods were also applicable to metagenomes of induced faecal samples from Oligo-MM12 mice, including for strains with a relative abundance below 1%, illustrating its potential for the discovery of inducible prophages also in more complex metagenomes. Finally, we show that predictions of prophage locations in reference genomes of the strains we studied were variable and inconsistent for four bioinformatic tools we tested, which highlights the importance of their experimental validation. CONCLUSIONS: This study demonstrates that the integration of experimental induction and bioinformatic analysis presented here is a powerful approach to accurately locate inducible prophages using high-throughput sequencing data and to quantify their activity. The ability to generate such quantitative information will be critical in helping us to gain better insights into the factors that determine phage activity and how prophage-bacteria interactions influence our microbiome and impact human health. Video abstract.

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A predator-prey model is used to investigate the interactions between phages and bacteria by considering the lytic and lysogenic life cycles of phages and the prophage induction. We provide answers to the following conflictual research questions: (1) what are conditions under which the presence of phages can purify a bacterial infected environment? (2) Can the presence of phages triggers virulent bacterial outbreaks? We derive the basic offspring number $\mathcal N_0$ that serves as a threshold and the bifurcation parameter to study the dynamics and bifurcation of the system. The model exhibits three equilibria: an unstable environment-free equilibrium, a globally asymptotically stable (GAS) phage-free equilibrium (PFE) whenever $\mathcal N_0<1$, and a locally asymptotically stable environment-persistent equilibrium (EPE) when $\mathcal N_0>1$. The Lyapunov-LaSalle techniques are used to prove the GAS of the PFE and estimate the EPE basin of attraction. Through the center manifold approximation, topological types of the PFE are precised. Existence of transcritical and Hopf bifurcations are established. Precisely, when $\mathcal N_0>1$, the EPE loses its stability and periodic solutions arise. Furthermore, increasing $\mathcal N_0$ can purify an environment where bacteriophages are introduced. Purposely, we prove that for large values of $\mathcal N_0$, the overall bacterial population asymptotically approaches zero, while the phage population sustains. Ecologically, our results show that for small values of $\mathcal N_0$, the existence of periodic solutions could explain the occurrence of repetitive bacteria-borne disease outbreaks, while large value of $\mathcal N_0$ clears bacteria from the environment. Numerical simulations support our theoretical results.

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Shiga toxin-producing Escherichia coli (STEC) O145 is a major serotype associated with severe human disease. Production of Shiga toxins (Stxs), especially Stx2a, is thought to be correlated with STEC virulence. Since stx genes are located in prophages genomes, induction of prophages is required for effective Stxs production. Here, we investigated the production of Stxs in 12 environmental STEC O145:H28 strains under stresses STEC encounter in natural habitats and performed comparative analysis with two O145:H28 clinical strains, one linked to a 2010 U.S. lettuce-associated outbreak (RM13514) and the other linked to a 2007 Belgium ice cream-associated outbreak (RM13516). Similar to the outbreak strains, all environmental strains belong to Sequence Type (ST)-78 using the EcMLST typing scheme. Although all Stx1a-prophages were grouped together, variations in Stx1a production were observed prior to or following the inductions. Among all stx2a positive environmental strains, only the Stx2a-prophage in cattle isolate RM9154-C1 was clustered with the Stx2a-prophages in RM13514, the Stx2a-phage induced from a STEC O104:H4 strain linked to the 2011 outbreak of enterohemorrhagic infection in Germany, and the Stx2a-prophage in STEC O157:H7 strain EDL933, a prototype of enterohemorrhagic E. coli. Furthermore, the Stx2a-prophage in RM9154-C1 shared the same chromosomal insertion site and carried the same antiterminator Q gene and the late promoter PR' as the Stx2a-prophage in RM13514. Following mitomycin C or enrofloxacin treatment, the production of Stx2a in RM9154-C1 was the highest among all environmental strains tested. In contrast, following acid challenge and recovery, the production of Stx2a in RM9154-C1 was the lowest among all the environmental strains tested, at a level comparable to the clinical strains. A significant increase in Stx2a production was detected in all strains when exposed to H2O2, although the induction fold was much lower than those by other inducers. This low-efficiency induction of Stx-prophages by H2O2, a natural inducer of Stx-prophages, supports the hypothesis of bacterial altruism in controlling Stxs production, a strategy that assures the survival of the STEC population as a whole by sacrificing a small fraction of cells for Stxs production and release. Differential induction of Stxs among strains carrying nearly identical Stx-prophages suggests a role of host bacteria in regulating Stxs production. Our study revealed diverse Stx-prophages in STEC O145:H28 strains that were genotypically indistinguishable. Identification of a cattle isolate harboring a Stx2a-prophage associated with high virulence supports the premise that cattle, a natural reservoir of STEC, serve as a source of hypervirulent STEC strains.

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Bacteriophages (phages) are the most abundant biological entity in the human body, but until recently the role that phages play in human health was not well characterized. Although phages do not cause infections in human cells, phages can alter the severity of bacterial infections by the dissemination of virulence factors amongst bacterial hosts. Recent studies, made possible with advances in genome engineering and microscopy, have uncovered a novel role for phages in the human body - the ability to modulate the physiology of the mammalian cells that can harbor intracellular bacteria. In this review, we synthesize key results on how phages traverse through mammalian cells - including uptake, distribution, and interaction with intracellular receptors - highlighting how these steps in turn influence host cell killing of bacteria. We discuss the implications of the growing field of phage-mammalian cell interactions for phage therapy.

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Pseudomonas aeruginosa is responsible for long-term infections and is particularly resistant to treatments when hiding inside the extracellular matrix or biofilms. Phage therapy might represent an alternative to antibiotic treatment, but up to 10% of clinical strains appear to resist multiple phages. We investigated the characteristics of P. aeruginosa clinical strains naturally resistant to phages and compared them to highly susceptible strains. The phage-resistant strains were defective in lipopolysaccharide (LPS) biosynthesis, were nonmotile and displayed an important degree of autolysis, releasing phages and pyocins. Complete genome sequencing of three resistant strains showed the existence of a large accessory genome made of multiple insertion elements, genomic islands, pyocins and prophages, including two phages performing lateral transduction. Mutations were found in genes responsible for the synthesis of LPS and/or type IV pilus, the major receptors for most phages. CRISPR-Cas systems appeared to be absent or inactive in phage-resistant strains, confirming that they do not play a role in the resistance to lytic phages but control the insertion of exogenous sequences. We show that, despite their apparent weakness, the multiphage-resistant strains described in this study displayed selective advantages through the possession of various functions, including weapons to eliminate other strains of the same or closely related species.