Global Virulence Research Articles

Transcriptional Profiling of Staphylococcus aureus during the Transition from Asymptomatic Nasal Colonization to Skin Colonization/Infection in Patients with Atopic Dermatitis.

Staphylococcus aureus acts both as a colonizing commensal bacterium and invasive pathogen. Nasal colonization is associated with an increased risk of infection caused by the identical strain. In patients with atopic dermatitis (AD), the degree of S. aureus colonization is associated with the severity of the disease. Here, we comparatively analyzed the in vivo transcriptional profile of S. aureus colonizing the nose and non-diseased skin (non-lesional skin) as opposed to the diseased skin (lesional skin-defined here as infection) of 12 patients with AD. The transcriptional profile during the asymptomatic colonization of the nose closely resembled that of the lesional skin samples for many of the genes studied, with an elevated expression of the genes encoding adhesion-related proteins and proteases. In addition, the genes that modify and remodel the cell wall and encode proteins that facilitate immune evasion showed increased transcriptional activity. Notably, in a subgroup of patients, the global virulence regulator Agr (accessory gene regulator) and downstream target genes were inactive during nasal colonization but upregulated in the lesional and non-lesional skin samples. Taken together, our results demonstrate a colonization-like transcriptional profile on diseased skin and suggest a role for the peptide quorum sensing system Agr during the transition from asymptomatic nasal colonization to skin colonization/infection.

Population genomics uncovers global distribution, antimicrobial resistance, and virulence genes of the opportunistic pathogen Klebsiella aerogenes

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 equippedby 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.

Assessment of the hypothetical protein BB0616 in the murine infection of Borrelia burgdorferi.

bb0616 of Borrelia burgdorferi, the Lyme disease pathogen, encodes a hypothetical protein of unknown function. In this study, we showed that BB0616 was not surface-exposed or associated with the membrane through localization analyses using proteinase K digestion and cell partitioning assays. The expression of bb0616 was influenced by a reduced pH but not by growth phases, elevated temperatures, or carbon sources during in vitro cultivation. A transcriptional start site for bb0616 was identified by using 5' rapid amplification of cDNA ends, which led to the identification of a functional promoter in the 5' regulatory region upstream of bb0616. By analyzing a bb0616-deficient mutant and its isogenic complemented counterparts, we found that the infectivity potential of the mutant was significantly attenuated. The inactivation of bb0616 displayed no effect on borrelial growth in the medium or resistance to oxidative stress, but the mutant was significantly more susceptible to osmotic stress. In addition, the production of global virulence regulators such as BosR and RpoS as well as virulence-associated outer surface lipoproteins OspC and DbpA was reduced in the mutant. These phenotypes were fully restored when gene mutation was complemented with a wild-type copy of bb0616. Based on these findings, we concluded that the hypothetical protein BB0616 is required for the optimal infectivity of B. burgdorferi, potentially by impacting B. burgdorferi virulence gene expression as well as survival of the spirochete under stressful conditions.

The global RNA–RNA interactome of Klebsiella pneumoniae unveils a small RNA regulator of cell division

The ubiquitous RNA chaperone Hfq is involved in the regulation of key biological processes in many species across the bacterial kingdom. In the opportunistic human pathogen Klebsiella pneumoniae, deletion of the hfq gene affects the global transcriptome, virulence, and stress resistance; however, the ligands of the major RNA-binding protein in this species have remained elusive. In this study, we have combined transcriptomic, co-immunoprecipitation, and global RNA interactome analyses to compile an inventory of conserved and species-specific RNAs bound by Hfq and to monitor Hfq-mediated RNA-RNA interactions. In addition to dozens of RNA-RNA pairs, our study revealed an Hfq-dependent small regulatory RNA (sRNA), DinR, which is processed from the 3' terminal portion of dinI mRNA. Transcription of dinI is controlled by the master regulator of the SOS response, LexA. As DinR accumulates in K. pneumoniae in response to DNA damage, the sRNA represses translation of the ftsZ transcript by occupation of the ribosome binding site. Ectopic overexpression of DinR causes depletion of ftsZ mRNA and inhibition of cell division, while deletion of dinR antagonizes cell elongation in the presence of DNA damage. Collectively, our work highlights the important role of RNA-based gene regulation in K. pneumoniae and uncovers the central role of DinR in LexA-controlled division inhibition during the SOS response.

Persistent Salmonella infections in humans are associated with mutations in the BarA/SirA regulatory pathway

Several bacterial pathogens, including Salmonella enterica, can cause persistent infections in humans by mechanisms that are poorly understood. By comparing genomes of isolates longitudinally collected from 256 prolonged salmonellosis patients, we identified repeated mutations in global regulators, including the barA/sirA two-component regulatory system, across multiple patients and Salmonella serovars. Comparative RNA-seq analysis revealed that distinct mutations in barA/sirA led to diminished expression of Salmonella pathogenicity islands 1 and 4 genes, which are required for Salmonella invasion and enteritis. Moreover, barA/sirA mutants were attenuated in an acute salmonellosis mouse model and induced weaker transcription of host immune responses. In contrast, in a persistent infection mouse model, these mutants exhibited long-term colonization and prolonged shedding. Taken together, these findings suggest that selection of mutations in global virulence regulators facilitates persistent Salmonella infection in humans, by attenuating Salmonella virulence and inducing a weaker host inflammatory response.

The Mutation of the DNA-Binding Domain of Fur Protein Enhances the Pathogenicity of Edwardsiella piscicida via Inducing Overpowering Pyroptosis.

Edwardsiella piscicida is an important fish pathogen with a broad host that causes substantial economic losses in the aquaculture industry. Ferric uptake regulator (Fur) is a global transcriptional regulator and contains two typical domains, the DNA-binding domain and dimerization domain. In a previous study, we obtained a mutant strain of full-length fur of E. piscicida, TX01Δfur, which displayed increased siderophore production and stress resistance factors and decreased pathogenicity. To further reveal the regulatory mechanism of Fur, the DNA-binding domain (N-terminal) of Fur was knocked out in this study and the mutant was named TX01Δfur2. We found that TX01Δfur2 displayed increased siderophore production and enhanced adversity tolerance, including a low pH, manganese, and high temperature stress, which was consistent with the phenotype of TX01Δfur. Contrary to TX01Δfur, whose virulence was weakened, TX01Δfur2 displayed an ascended invasion of nonphagocytic cells and enhanced destruction of phagocytes via inducing overpowering or uncontrollable pyroptosis, which was confirmed by the fact that TX01Δfur2 induced higher levels of cytotoxicity, IL-1β, and p10 in macrophages than TX01. More importantly, TX01Δfur2 displayed an increased global virulence to the host, which was confirmed by the result that TX01Δfur2 caused higher lethality outcomes for healthy tilapias than TX01. These results demonstrate that the mutation of the Fur N-terminal domain augments the resistance level against the stress and pathogenicity of E. piscicida, which is not dependent on the bacterial number in host cells or host tissues, although the capabilities of biofilm formation and the motility of TX01Δfur2 decline. These interesting findings provide a new insight into the functional analysis of Fur concerning the regulation of virulence in E. piscicida and prompt us to explore the subtle regulation mechanism of Fur in the future.

PhcX Is a LqsR-family response regulator that contributes to Ralstonia solanacearum virulence and regulates multiple virulence factors.

The bacterial wilt caused by the soil-borne phytopathogen Ralstonia solanacearum is one of the most destructive crop diseases. To achieve a successful infection, R. solanacearum has evolved an intricate regulatory network to orchestrate the expression of an arsenal of virulence factors and fine-tune the allocation of energy. However, despite the wealth of knowledge gained in the past decades, many players and connections are still missing from the network. The importance of our study lies in the identification of PhcX, a novel conserved global regulator with critical roles in modulating the virulence and metabolism of R. solanacearum. PhcX affects many well-characterized regulators and exhibits contrasting modes of regulation from the central regulator PhcA on a variety of virulence-associated traits and genes. Our findings add a valuable piece to the puzzle of how the pathogen regulates its proliferation and infection, which is critical for understanding its pathogenesis and developing disease control strategies.

The role of integration host factor in biofilm and virulence of high-alcohol-producing Klebsiella pneumoniae.

Klebsiella pneumoniae is a well-known human nosocomial pathogen with an arsenal of virulence factors, including capsular polysaccharides (CPS), fimbriae, flagella, and lipopolysaccharides (LPS). Our previous study found that alcohol acted as an essential virulence factor for high-alcohol-producing K. pneumoniae (HiAlc Kpn). Integration host factor (IHF) is a nucleoid-associated protein that functions as a global virulence regulator in Escherichia coli. However, the regulatory role of IHF in K. pneumoniae remains unknown. In the present study, we found that deletion of ihfA or ihfB resulted in a slight defect in bacterial growth, a severe absence of biofilm formation and cytotoxicity, and a significant reduction in alcohol production. RNA sequencing differential gene expression analysis showed that compared with the wild-type control, the expression of many virulence factor genes was downregulated in ΔihfA and ΔihfB strains, such as those related to CPS (rcsA, galF, wzi, and iscR), LPS (rfbABCD), type I and type III fimbriae (fim and mrk operon), cellulose (bcs operon), iron transporter (feoABC, fhuA, fhuF, tonB, exbB, and exbD), quorum sensing (lsr operon and sdiA), type II secretion system (T2SS) and type VI secretion system (T6SS) (tssG, hcp, and gspE). Of these virulence factors, CPS, LPS, fimbriae, and cellulose are involved in biofilm formation. In addition, IHF could affect the alcohol production by regulating genes related to glucose intake (ptsG), pyruvate formate-lyase, alcohol dehydrogenase, and the tricarboxylic acid (TCA) cycle. Our data provided new insights into the importance of IHF in regulating the virulence of HiAlc Kpn. IMPORTANCE Klebsiella pneumoniae is a well-known human nosocomial pathogen that causes various infectious diseases, including urinary tract infections, hospital-acquired pneumonia, bacteremia, and liver abscesses. Our previous studies demonstrated that HiAlc Kpn mediated the development of nonalcoholic fatty liver disease by producing excess endogenous alcohol in vivo. However, the regulators regulating the expression of genes related to metabolism, biofilm formation, and virulence of HiAlc Kpn remain unclear. In this study, the regulator IHF was found to positively regulate biofilm formation and many virulence factors including CPS, LPS, type I and type III fimbriae, cellulose, iron transporter, AI-2 quorum sensing, T2SS, and T6SS in HiAlc Kpn. Furthermore, IHF positively regulated alcohol production in HiAlc Kpn. Our results suggested that IHF could be a potential drug target for treating various infectious diseases caused by K. pneumoniae. Hence, the regulation of different virulence factors by IHF in K. pneumoniae requires further investigation.

Rifampicin-resistant RpoB S522L Vibrio vulnificus exhibits disturbed stress response and hypervirulence traits.

Rifampicin resistance, which is genetically linked to mutations in the RNA polymerase β-subunit gene rpoB, has a global impact on bacterial transcription and cell physiology. Previously, we identified a substitution of serine 522 in RpoB (i.e., RpoBS522L ) conferring rifampicin resistance to Vibrio vulnificus, a human food-borne and wound-infecting pathogen associated with a high mortality rate. Transcriptional and physiological analysis of V. vulnificus expressing RpoBS522L showed increased basal transcription of stress-related genes and global virulence regulators. Phenotypically these transcriptional changes manifest as disturbed osmo-stress responses and toxin-associated hypervirulence as shown by reduced hypoosmotic-stress resistance and enhanced cytotoxicity of the RpoBS522L strain. These results suggest that RpoB-linked rifampicin resistance has a significant impact on V. vulnificus survival in the environment and during infection.

Paired single-cell host profiling with multiplex-tagged bacterial mutants reveals intracellular virulence-immune networks

Encounters between host cells and intracellular bacterial pathogens lead to complex phenotypes that determine the outcome of infection. Single-cell RNA sequencing (scRNA-seq) is increasingly used to study the host factors underlying diverse cellular phenotypes but has limited capacity to analyze the role of bacterial factors. Here, we developed scPAIR-seq, a single-cell approach to analyze infection with a pooled library of multiplex-tagged, barcoded bacterial mutants. Infected host cells and barcodes of intracellular bacterial mutants are both captured by scRNA-seq to functionally analyze mutant-dependent changes in host transcriptomes. We applied scPAIR-seq to macrophages infected with a library of Salmonella Typhimurium secretion system effector mutants. We analyzed redundancy between effectors and mutant-specific unique fingerprints and mapped the global virulence network of each individual effector by its impact on host immune pathways. ScPAIR-seq is a powerful tool to untangle bacterial virulence strategies and their complex interplay with host defense strategies that drive infection outcome.

Deep longitudinal multi-omics analysis of Bordetella pertussis cultivated in bioreactors highlights medium starvations and transitory metabolisms, associated to vaccine antigen biosynthesis variations and global virulence regulation.

Bordetella pertussis is the bacterial causative agent of whooping cough, a serious respiratory illness. An extensive knowledge on its virulence regulation and metabolism is a key factor to ensure pertussis vaccine manufacturing process robustness. The aim of this study was to refine our comprehension of B. pertussis physiology during in vitro cultures in bioreactors. A longitudinal multi-omics analysis was carried out over 26 h small-scale cultures of B. pertussis. Cultures were performed in batch mode and under culture conditions intending to mimic industrial processes. Putative cysteine and proline starvations were, respectively, observed at the beginning of the exponential phase (from 4 to 8 h) and during the exponential phase (18 h 45 min). As revealed by multi-omics analyses, the proline starvation induced major molecular changes, including a transient metabolism with internal stock consumption. In the meantime, growth and specific total PT, PRN, and Fim2 antigen productions were negatively affected. Interestingly, the master virulence-regulating two-component system of B. pertussis (BvgASR) was not evidenced as the sole virulence regulator in this in vitro growth condition. Indeed, novel intermediate regulators were identified as putatively involved in the expression of some virulence-activated genes (vags). Such longitudinal multi-omics analysis applied to B. pertussis culture process emerges as a powerful tool for characterization and incremental optimization of vaccine antigen production.

Auranofin inhibits virulence pathways in Pseudomonas aeruginosa

Pseudomonas aeruginosa is widely attributed as the leading cause of hospital-acquired infections. Due to intrinsic antibiotic resistance mechanisms and the ability to form biofilms, P. aeruginosa infections are challenging to treat. P. aeruginosa employs multiple virulence mechanisms to establish infections, many of which are controlled by the global virulence regulator Vfr. An attractive strategy to combat P. aeruginosa infections is thus the use of anti-virulence compounds. Here, we report the discovery that FDA-approved drug auranofin attenuates virulence pathways in P. aeruginosa, including quorum sensing (QS) and Type IV pili (TFP). We show that auranofin acts via multiple targets, one of which being Vfr. Consistent with inhibition of QS and TFP expression, we show that auranofin attenuates biofilm maturation, and when used in combination with colistin, displays strong synergy in eradicating P. aeruginosa biofilms. Auranofin may have immediate applications as an anti-virulence drug against P. aeruginosa infections.

Structural basis for the inhibitory mechanism of auranofin and gold(I) analogues against Pseudomonas aeruginosa global virulence factor regulator Vfr

Pseudomonas aeruginosa is a leading cause of hospital-acquired infections. Treatment of P. aeruginosa infections is difficult given its multiple virulence mechanisms, intrinsic antibiotic resistance mechanisms, and biofilm-forming ability. Auranofin, an approved oral gold compound for rheumatoid arthritis treatment, was recently reported to inhibit the growth of multiple bacterial species. Here, we identify P. aeruginosa’s global virulence factor regulator Vfr as one target of auranofin. We report the mechanistic insights into the inhibitory mechanism of auranofin and gold(I) analogues to Vfr through structural, biophysical, and phenotypic inhibition studies. This work suggests that auranofin and gold(I) analogues have potential to be developed as anti-virulence drugs against P. aeruginosa.

Immunogenicity of live phoP gene deletion strain of Riemerella anatipestifer serotype 1

Duck infectious serositis is an acute and infectious disease caused by Riemerella anatipestifer (R. anatipestifer) that leads to perihepatitis, pericarditis, meningitis, and airbag inflammation in ducks, which causes serious economic losses to the global duck industry. The phoP/phoR is a novel 2-component signal transduction system first reported in gram-negative bacteria, of which phoP acts as a global regulator and virulence factor. In this study, the phoP gene from the R. anatipestifer YM strain was knocked out using homologous recombination technology and replaced with the spectinomycin resistance gene (Spec). The virulence of the R. anatipestifer YMΔphoP strain was reduced by approximately 47,000 times compared to that of the wild-type R. anatipestifer YM strain. Ducks were immunized with live R. anatipestifer YMΔphoP strain by subcutaneous inoculation at a dose of 106 to 107 CFU (0.2 mL per duck) and challenged with the wild-type R. anatipestifer YM strain 14 days later. The protection rate in the immunized group was 100%. The growth characteristics of ducks in the immunized and negative control groups were normal, and the research demonstrated R. anatipestifer YMΔphoP strain have suitable immunogenicity and protective effects. Thus, the study findings suggest that the novel R. anatipestifer YMΔphoP strain may provide a candidate for the development of a gene deletion activated vaccine against duck infectious serositis.

Pan-Genome Analysis of Campylobacter: Insights on the Genomic Diversity and Virulence Profile.

ABSTRACTThe genus Campylobacter contains pathogens that cause bacterial gastroenteritis in humans and animals. Despite large-scale sequencing efforts to raise clinical awareness of Campylobacter, little is known about the diversity and functions of virulence factors. Here, we constructed the pan-genome of Campylobacter using 39 representative genomes, elucidating their genetic diversity, evolutionary characteristics, and virulence and resistance profiles. The Campylobacter pan-genome was open and showed extensive genome variability, with high levels of gene expansion and contraction as the organism evolved. These Campylobacter members had diverse virulence gene content, and six potential core virulence genes (porA, PEB4, cheY, htrB, Cj1135, and kpsF) have been identified. The conserved mechanisms for Campylobacter pathogenicity were related to adherence, motility, and immune modulation. We emphasized the relative importance of variable virulence genes. Many virulence genes have experienced expansion or contraction in specific lineages, which may be one of the factors causing differences in the content of virulence genes. Additionally, these Campylobacter genomes have a high prevalence of the cmeA and cmeC genes, which are linked to the CmeABC pump and contribute to multidrug resistance. The genomic variations, core and variable virulence factors, and resistance genes of Campylobacter characterized in this study would contribute to a better understanding of the virulence of Campylobacter and more effective use of candidates for drug development and prevention of Campylobacter infections.IMPORTANCE Pathogenic members of the genus Campylobacter are recognized as one of the major causative agents of human bacterial gastroenteritis. This study revealed the pan-genome of 39 Campylobacter species, provided the most updated reconstruction of the global virulence gene pool of 39 Campylobacter species, and identified species-related virulence differences. This study highlighted the basic conserved functionality and specificity of pathogenicity that are crucial to infection, which was critical for improving the diagnosis and prevention of Campylobacter infections.

Effect of Environmental Temperatures on Proteome Composition of Salmonella enterica Serovar Typhimurium.

Salmonella enterica serovar Typhimurium (STM) is a major cause of gastroenteritis and transmitted by consumption of contaminated food. STM is associated to food originating from animals (pork, chicken, eggs) or plants (vegetables, fruits, nuts, and herbs). Infection of warm-blooded mammalian hosts by STM and the underlying complex regulatory network of virulence gene expression depend on various environmental conditions encountered in hosts. However, less is known about the proteome and possible regulatory networks for gene expression of STM outside the preferred host. Nutritional limitations and changes in temperature are the most obvious stresses outside the native host. Thus, we analyzed the proteome profile of STM grown in rich medium (LB medium) or minimal medium (PCN medium) at temperatures ranging from 8 °C to 37 °C. LB medium mimics the nutritional rich environment inside the host, whereas minimal PCN medium represents nutritional limitations outside the host, found during growth of fresh produce (field conditions). Further, the range of temperatures analyzed reflects conditions within natural hosts (37 °C), room temperature (20 °C), during growth under agricultural conditions (16 °C and 12 °C), and during food storage (8 °C). Implications of altered nutrient availability and growth temperature on STM proteomes were analyzed by HPLC/MS-MS and label-free quantification. Our study provides first insights into the complex adaptation of STM to various environmental temperatures, which allows STM not only to infect mammalian hosts but also to enter new infection routes that have been poorly studied so far. With the present dataset, global virulence factors, their impact on infection routes, and potential anti-infective strategies can now be investigated in detail. Especially, we were able to demonstrate functional flagella at 12 °C growth temperature for STM with an altered motility behavior.

The Xanthophyll Carotenoid Lutein Reduces the Invasive Potential of Pseudomonas aeruginosa and Increases Its Susceptibility to Tobramycin.

Recently, the xanthophyll carotenoid lutein has been qualified as a potential quorum sensing (QS) and biofilm inhibitor against Pseudomonas aeruginosa. To address the potential of this xanthophyll compound as a relevant antivirulence agent, we investigated in depth its impact on the invasion capabilities and aggressiveness of P. aeruginosa PAO1, which rely on the bacterial ability to build and maintain protective barriers, use different types of motilities and release myriad virulence factors, leading to host cell and tissue damages. Our data, obtained on the PAO1 strain, indicate that all-trans lutein (Lut; 22 µM) disrupts biofilm formation and disorganizes established biofilm structure without affecting bacterial viability, while improving the bactericidal activity of tobramycin against biofilm-encapsulated PAO1 cells. Furthermore, this xanthophyll affects PAO1 twitching and swarming motilities while reducing the production of the extracellular virulence factors pyocyanin, elastase and rhamnolipids as well as the expression of the QS-regulated lasB and rhlA genes without inhibiting the QS-independent aceA gene. Interestingly, the expression of the QS regulators rhlR/I and lasR/I is significantly reduced as well as that of the global virulence factor regulator vfr, which is suggested to be a major target of Lut. Finally, an oxidative metabolite of Lut, 3′-dehydrolutein, induces a similar inhibition phenotype. Taken together, lutein-type compounds represent potential agents to control the invasive ability and antibiotic resistance of P. aeruginosa.

Air pollution induces Staphylococcus aureus USA300 respiratory tract colonization mediated by specific bacterial genetic responses involving the global virulence gene regulators Agr and Sae.

Exposure to particulate matter (PM), a major component of air pollution, is associated with exacerbation of chronic respiratory disease, and infectious diseases such as community-acquired pneumonia. Although PM can cause adverse health effects through direct damage to host cells, our previous study showed that PM can also impact bacterial behaviour by promoting in vivo colonization. In this study we describe the genetic mechanisms involved in the bacterial response to exposure to black carbon (BC), a constituent of PM found in most sources of air pollution. We show that Staphylococcus aureus strain USA300 LAC grown in BC prior to inoculation showed increased murine respiratory tract colonization and pulmonary invasion in vivo, as well as adhesion and invasion of human epithelial cells in vitro. Global transcriptional analysis showed that BC has a widespread effect on S. aureus transcriptional responses, altering the regulation of the major virulence gene regulators Sae and Agr and causing increased expression of genes encoding toxins, proteases and immune evasion factors. Together these data describe a previously unrecognized causative mechanism of air pollution-associated infection, in that exposure to BC can increase bacterial colonization and virulence factor expression by acting directly on the bacterium rather than via the host.

The Isolation and Characterization of a Broad Host Range Bcep22-like Podovirus JC1.

Bacteriophage JC1 is a Podoviridae phage with a C1 morphotype, isolated on host strain Burkholderia cenocepacia Van1. Phage JC1 is capable of infecting an expansive range of Burkholderia cepacia complex (Bcc) species. The JC1 genome exhibits significant similarity and synteny to Bcep22-like phages and to many Ralstonia phages. The genome of JC1 was determined to be 61,182 bp in length with a 65.4% G + C content and is predicted to encode 76 proteins and 1 tRNA gene. Unlike the other Lessieviruses, JC1 encodes a putative helicase gene in its replication module, and it is in a unique organization not found in previously analyzed phages. The JC1 genome also harbours 3 interesting moron genes, that encode a carbon storage regulator (CsrA), an N-acetyltransferase, and a phosphoadenosine phosphosulfate (PAPS) reductase. JC1 can stably lysogenize its host Van1 and integrates into the 5′ end of the gene rimO. This is the first account of stable integration identified for Bcep22-like phages. JC1 has a higher global virulence index at 37 °C than at 30 °C (0.8 and 0.21, respectively); however, infection efficiency and lysogen stability are not affected by a change in temperature, and no observable temperature-sensitive switch between lytic and lysogenic lifestyle appears to exist. Although JC1 can stably lysogenize its host, it possesses some desirable characteristics for use in phage therapy. Phage JC1 has a broad host range and requires the inner core of the bacterial LPS for infection. Bacteria that mutate to evade infection by JC1 may develop a fitness disadvantage as seen in previously characterized LPS mutants lacking inner core.

The core and accessory Hfq interactomes across Pseudomonas aeruginosa lineages

The major RNA-binding protein Hfq interacts with mRNAs, either alone or together with regulatory small noncoding RNAs (sRNAs), affecting mRNA translation and degradation in bacteria. However, studies tend to focus on single reference strains and assume that the findings may apply to the entire species, despite the important intra-species genetic diversity known to exist. Here, we use RIP-seq to identify Hfq-interacting RNAs in three strains representing the major phylogenetic lineages of Pseudomonas aeruginosa. We find that most interactions are in fact not conserved among the different strains. We identify growth phase-specific and strain-specific Hfq targets, including previously undescribed sRNAs. Strain-specific interactions are due to different accessory gene sets, RNA abundances, or potential context- or sequence- dependent regulatory mechanisms. The accessory Hfq interactome includes most mRNAs encoding Type III Secretion System (T3SS) components and secreted toxins in two strains, as well as a cluster of CRISPR guide RNAs in one strain. Conserved Hfq targets include the global virulence regulator Vfr and metabolic pathways involved in the transition from fast to slow growth. Furthermore, we use rGRIL-seq to show that RhlS, a quorum sensing sRNA, activates Vfr translation, thus revealing a link between quorum sensing and virulence regulation. Overall, our work highlights the important intra-species diversity in post-transcriptional regulatory networks in Pseudomonas aeruginosa.