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1. Phage Infection Restores PQS Signaling and Enhances Growth of a Pseudomonas aeruginosa lasI Quorum-Sensing Mutant

   https://doi.org/10.1128/jb.00557-21  

   Høyland-Kroghsbo, Nina Molin; Bassler, Bonnie L. (January 2022, Journal of Bacteriology)

   Bondy-Denomy, Joseph (Ed.)

   ABSTRACT Chemical communication between bacteria and between bacteria and the bacteriophage (phage) viruses that prey on them can shape the outcomes of phage-bacterial encounters. Quorum sensing (QS) is a bacterial cell-to-cell communication process that promotes collective undertaking of group behaviors. QS relies on the production, release, accumulation, and detection of signal molecules called autoinducers. Phages can exploit QS-mediated communication to manipulate their hosts and maximize their own survival. In the opportunistic pathogen Pseudomonas aeruginosa , the LasI/R QS system induces the RhlI/R QS system, and in opposing manners, these two systems control the QS system that relies on the autoinducer called PQS. A P. aeruginosa Δ lasI mutant is impaired in PQS synthesis, leading to accumulation of the precursor molecule HHQ, and HHQ suppresses growth of the P. aeruginosa Δ lasI strain. We show that, in response to a phage infection, the P. aeruginosa Δ lasI mutant reactivates QS, which, in turn, restores pqsH expression, enabling conversion of HHQ into PQS. Moreover, downstream QS target genes encoding virulence factors are induced. Additionally, phage-infected P. aeruginosa Δ lasI cells transiently exhibit superior growth compared to uninfected cells. IMPORTANCE Clinical isolates of P. aeruginosa frequently harbor mutations in particular QS genes. Here, we show that infection by select temperate phages restores QS, a cell-to-cell communication mechanism in a P. aeruginosa QS mutant. Restoration of QS increases expression of genes encoding virulence factors. Thus, phage infection of select P. aeruginosa strains may increase bacterial pathogenicity, underscoring the importance of characterizing phage-host interactions in the context of bacterial mutants that are relevant in clinical settings. 

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2. Quorum sensing controls Vibrio cholerae multicellular aggregate formation

   https://doi.org/10.7554/eLife.42057  

   Jemielita, Matthew; Wingreen, Ned S; Bassler, Bonnie L (December 2018, eLife)

   Bacteria communicate and collectively regulate gene expression using a process called quorum sensing (QS). QS relies on group-wide responses to signal molecules called autoinducers. Here, we show that QS activates a new program of multicellularity in Vibrio cholerae. This program, which we term aggregation, is distinct from the canonical surface-biofilm formation program, which QS represses. Aggregation is induced by autoinducers, occurs rapidly in cell suspensions, and does not require cell-division, features strikingly dissimilar from those characteristic of V. cholerae biofilm formation. Extracellular DNA limits aggregate size, but is not sufficient to drive aggregation. A mutagenesis screen identifies genes required for aggregate formation, revealing proteins involved in V. cholerae intestinal colonization, stress response, and a protein that distinguishes the current V. cholerae pandemic strain from earlier pandemic strains. We suggest that QS-controlled aggregate formation is important for V. cholerae to successfully transit between the marine niche and the human host. 

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3. Transcriptome profiling of type VI secretion system core gene tssM mutant of Xanthomonas perforans highlights regulators controlling diverse functions ranging from virulence to metabolism

   https://doi.org/10.1128/spectrum.02852-23  

   Ramamoorthy, Sivakumar; Pena, Michelle; Ghosh, Palash; Liao, Ying-Yu; Paret, Mathews; Jones, Jeffrey B.; Potnis, Neha (January 2024, Microbiology Spectrum)

   Burbank, Lindsey Price (Ed.)

   ABSTRACT Type VI secretion system (T6SS) is a versatile, contact-dependent contractile nano-weapon in Gram-negative bacteria that fires proteinaceous effector molecules directly into prokaryotic and eukaryotic cells aiding in manipulation of the host and killing of competitors in complex niches. In plant pathogenic xanthomonads, T6SS has been demonstrated to play these diverse roles in individual pathosystems. However, the molecular network underlying the regulation of T6SS is still elusive inXanthomonasspp. To bridge this knowledge gap, we conducted anin vitrotranscriptome screen using plant apoplast mimicking minimal medium, XVM2 medium, to decipher the effect oftssMdeletion, a core gene belonging to T6SS-cluster i3*, on the regulation of gene expression inXanthomonas perforansstrain AL65. Transcriptomic data revealed that a total of 277 and 525 genes were upregulated, while 307 and 392 genes were downregulated in the mutant strain after 8 and 16 hours of growth in XVM2 medium. The transcript abundance of several genes associated with flagellum and pilus biogenesis as well as type III secretion system was downregulated in the mutant strain. Deletion oftssMof cluster-i3* resulted in upregulation of several T6SS genes belonging to cluster-i3*** and genes involved in biofilm and cell wall biogenesis. Similarly, transcription regulators likerpoN, Pho regulon,rpoE, andcsrAwere identified to be upregulated in the mutant strain. Our results suggest that T6SS modulates the expression of global regulators likecsrA,rpoN, andphoregulons, triggering a signaling cascade, and co-ordinates the expression of suite of virulence factors, stress response genes, and metabolic genes. IMPORTANCET6SS has received attention due to its significance in mediating interorganismal competition through contact-dependent release of effector molecules into prokaryotic and eukaryotic cells. Reverse-genetic studies have indicated the role of T6SS in virulence in a variety of plant pathogenic bacteria, including the one studied here,Xanthomonas. However, it is not clear whether such effect on virulence is merely due to a shift in the microbiome-mediated protection or if T6SS is involved in a complex virulence regulatory network. In this study, we conducted in vitro transcriptome profiling in minimal medium to decipher the signaling pathways regulated by tssM-i3* inX. perforansAL65. We show that TssM-i3* regulates the expression of a suite of genes associated with virulence and metabolism either directly or indirectly by altering the transcription of several regulators. These findings further expand our knowledge on the intricate molecular circuits regulated by T6SS in phytopathogenic bacteria. 

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4. Regulatory small RNA, Qrr2 is expressed independently of sigma factor-54 and can function as the sole Qrr sRNA to control quorum sensing in Vibrio parahaemolyticus

   https://doi.org/10.1128/JB.00350-21  

   Tague, J.G.; Hong, J.; Kalburge, S.S.; Boyd, E.F. (October 2021, Journal of Bacteriology)

   null (Ed.)

   Bacterial cells alter gene expression in response to changes in population density in a process called quorum sensing (QS). In Vibrio harveyi, LuxO, a low cell density activator of sigma factor-54 (RpoN), is required for transcription of five non-coding regulatory sRNAs, Qrr1-Qrr5, which each repress translation of the master QS regulator LuxR. Vibrio parahaemolyticus, the leading cause of bacterial seafood-borne gastroenteritis, also contains five Qrr sRNAs that control OpaR (the LuxR homolog), controlling capsule polysaccharide (CPS), motility, and metabolism. We show that in a Δ luxO deletion mutant, opaR was de-repressed and CPS and biofilm were produced. However, in a Δ rpoN mutant, opaR was repressed, no CPS was produced, and less biofilm production was observed compared to wild type. To determine why opaR was repressed, expression analysis in Δ luxO showed all five qrr genes were repressed, while in Δ rpoN the qrr2 gene was significantly de-repressed. Reporter assays and mutant analysis showed Qrr2 sRNA can act alone to control OpaR. Bioinformatics analysis identified a sigma-70 (RpoD) -35 -10 promoter overlapping the canonical sigma-54 (RpoN) -24 -12 promoter in the qrr2 regulatory region. The qrr2 sigma-70 promoter element was also present in additional Vibrio species indicating it is widespread. Mutagenesis of the sigma-70 -10 promoter site in the Δ rpoN mutant background, resulted in repression of qrr2. Analysis of qrr quadruple deletion mutants, in which only a single qrr gene is present, showed that only Qrr2 sRNA can act independently to regulate opaR . Mutant and expression data also demonstrated that RpoN and the global regulator, Fis, act additively to repress qrr2 . Our data has uncovered a new mechanism of qrr expression and shows that Qrr2 sRNA is sufficient for OpaR regulation. Importance The quorum sensing non-coding sRNAs are present in all Vibrio species but vary in number and regulatory roles among species. In the Harveyi clade, all species contain five qrr genes, and in V. harveyi these are transcribed by sigma-54 and are additive in function. In the Cholerae clade, four qrr genes are present, and in V. cholerae the qrr genes are redundant in function. In V. parahaemolyticus , qrr2 is controlled by two overlapping promoters. In an rpoN mutant, qrr2 is transcribed from a sigma-70 promoter that is present in all V. parahaemolyticus strains and in other species of the Harveyi clade suggesting a conserved mechanism of regulation. Qrr2 sRNA can function as the sole Qrr sRNA to control OpaR. 

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5. Natural silencing of quorum-sensing activity protects Vibrio parahaemolyticus from lysis by an autoinducer-detecting phage

   https://doi.org/10.1371/journal.pgen.1010809  

   Duddy, Olivia P.; Silpe, Justin E.; Fei, Chenyi; Bassler, Bonnie L. (July 2023, PLOS Genetics)

   Matic, Ivan (Ed.)

   Quorum sensing (QS) is a chemical communication process that bacteria use to track population density and orchestrate collective behaviors. QS relies on the production, accumulation, and group-wide detection of extracellular signal molecules called autoinducers. Vibriophage 882 (phage VP882), a bacterial virus, encodes a homolog of theVibrioQS receptor-transcription factor, called VqmA, that monitors theVibrioQS autoinducer DPO. Phage VqmA binds DPO at high host-cell density and activates transcription of the phage geneqtip. Qtip, an antirepressor, launches the phage lysis program. Phage-encoded VqmA when bound to DPO also manipulates host QS by activating transcription of the host genevqmR. VqmR is a small RNA that controls downstream QS target genes. Here, we sequenceVibrio parahaemolyticusstrain O3:K6 882, the strain from which phage VP882 was initially isolated. The chromosomal region normally encodingvqmRandvqmAharbors a deletion encompassingvqmRand a portion of thevqmApromoter, inactivating that QS system. We discover thatV.parahaemolyticusstrain O3:K6 882 is also defective in its other QS systems, due to a mutation inluxO, encoding the central QS transcriptional regulator LuxO. Both thevqmR-vqmAandluxOmutations lockV.parahaemolyticusstrain O3:K6 882 into the low-cell density QS state. Reparation of the QS defects inV.parahaemolyticusstrain O3:K6 882 promotes activation of phage VP882 lytic gene expression and LuxO is primarily responsible for this effect. Phage VP882-infected QS-competentV.parahaemolyticusstrain O3:K6 882 cells lyse more rapidly and produce more viral particles than the QS-deficient parent strain. We propose that, inV.parahaemolyticusstrain O3:K6 882, constitutive maintenance of the low-cell density QS state suppresses the launch of the phage VP882 lytic cascade, thereby protecting the bacterial host from phage-mediated lysis. 

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