scholarly journals Roles of the Structural Symbiosis Polysaccharide (syp) Genes in Host Colonization, Biofilm Formation, and Polysaccharide Biosynthesis in Vibrio fischeri

2012 ◽  
Vol 194 (24) ◽  
pp. 6736-6747 ◽  
Author(s):  
S. Shibata ◽  
E. S. Yip ◽  
K. P. Quirke ◽  
J. M. Ondrey ◽  
K. L. Visick
Keyword(s):  
Biofilm Formation ◽  
Vibrio Fischeri ◽  
Host Colonization ◽  
Polysaccharide Biosynthesis ◽  
Symbiosis Polysaccharide

scholarly journals RscS Functions Upstream of SypG To Control the syp Locus and Biofilm Formation in Vibrio fischeri

2008 ◽  
Vol 190 (13) ◽  
pp. 4576-4583 ◽  
Author(s):  
Elizabeth A. Hussa ◽  
Cynthia L. Darnell ◽  
Karen L. Visick
Keyword(s):  
Biofilm Formation ◽  
Large Scale ◽  
Vibrio Fischeri ◽  
Response Regulator ◽  
Bacterial Cells ◽  
Euprymna Scolopes ◽  
Regulatory Pathways ◽  
Two Component ◽  
Two Component Signal Transduction ◽  
Symbiosis Polysaccharide

ABSTRACTTwo-component signal transduction systems, composed of sensor kinase (SK) and response regulator (RR) proteins, allow bacterial cells to adapt to changes such as environmental flux or the presence of a host. RscS is an SK required forVibrio fischerito initiate a symbiotic partnership with the Hawaiian squidEuprymna scolopes, likely due to its role in controlling the symbiosis polysaccharide (syp) genes and thus biofilm formation. To determine which RR(s) functions downstream of RscS, we performed epistasis experiments with a library of 35 RR mutants. We found that several RRs contributed to RscS-mediated biofilm formation inV. fischeri. However, only thesyp-encoded symbiosis regulator SypG was required for both biofilm phenotypes andsyptranscription induced by RscS. These data support the hypothesis that RscS functions upstream of SypG to induce biofilm formation. In addition, this work also revealed a role for thesyp-encoded RR SypE in biofilm formation. To our knowledge, no other study has used a large-scale epistasis approach to elucidate two-component signaling pathways. Therefore, this work both contributes to our understanding of regulatory pathways important for symbiotic colonization byV. fischeriand establishes a paradigm for evaluating two-component pathways in the genomics era.

scholarly journals Hybrid histidine kinase BinK represses Vibrio fischeri biofilm signaling at multiple developmental stages

2021 ◽  
Author(s):  
Denise A. Ludvik ◽  
Katherine M. Bultman ◽  
Mark J. Mandel
Keyword(s):  
Gene Expression ◽  
Gene Regulation ◽  
Histidine Kinase ◽  
Biofilm Formation ◽  
Vibrio Fischeri ◽  
Bacterial Biofilm ◽  
Negative Regulator ◽  
Light Organ ◽  
Host Colonization ◽  
Hybrid Histidine Kinase

ABSTRACTThe symbiosis between the Hawaiian bobtail squid, Euprymna scolopes, and its exclusive light-organ symbiont, Vibrio fischeri, provides a natural system in which to study host-microbe specificity and gene regulation during the establishment of a mutually-beneficial symbiosis. Colonization of the host relies on bacterial biofilm-like aggregation in the squid mucus field. Symbiotic biofilm formation is controlled by a two-component signaling (TCS) system consisting of regulators RscS-SypF-SypG, which together direct transcription of the Syp symbiotic polysaccharide. TCS systems are broadly important for bacteria to sense environmental cues and then direct changes in behavior. Previously, we identified hybrid histidine kinase BinK as a strong negative regulator of V. fischeri biofilm regulation, and here we further explore the function of BinK. To inhibit biofilm formation, BinK requires the predicted phosphorylation sites in both the histidine kinase (H362) and receiver (D794) domains. Furthermore, we show that a strain lacking BinK yields RscS non-essential for host colonization, and imaging of aggregate size revealed no benefit to the presence of RscS in a background lacking BinK. Strains lacking RscS still suffered in competition, suggesting another function for the protein. Finally, we show that BinK functions to inhibit biofilm gene expression in the light organ crypts, providing evidence for biofilm gene regulation at later stages of host colonization. Overall, this study provides direct evidence for opposing activities of RscS and BinK and yields novel insights into biofilm regulation during the maturation of a beneficial symbiosis.IMPORTANCEBacteria are often in a biofilm state, and transitions between planktonic and biofilm lifestyles are important for pathogenic, beneficial, and environmental microbes. The critical nature of biofilm formation during Vibrio fischeri colonization of the Hawaiian bobtail squid light organ provides an opportunity to study development of this process in vivo using a combination of genetic and imaging approaches. The current work refines the signaling circuitry of the biofilm pathway in V. fischeri, provides evidence that biofilm regulatory changes occur in the host, and identifies BinK as one of the regulators of that process. This study provides information about how bacteria regulate biofilm gene expression in an intact animal host.

scholarly journals Hybrid histidine kinase BinK represses Vibrio fischeri biofilm signaling at multiple developmental stages

2021 ◽  
Author(s):  
Denise A. Ludvik ◽  
Katherine M. Bultman ◽  
Mark J. Mandel
Keyword(s):  
Gene Expression ◽  
Gene Regulation ◽  
Histidine Kinase ◽  
Biofilm Formation ◽  
Vibrio Fischeri ◽  
Bacterial Biofilm ◽  
Negative Regulator ◽  
Light Organ ◽  
Host Colonization ◽  
Hybrid Histidine Kinase

The symbiosis between the Hawaiian bobtail squid, Euprymna scolopes, and its exclusive light-organ symbiont, Vibrio fischeri, provides a natural system in which to study host-microbe specificity and gene regulation during the establishment of a mutually-beneficial symbiosis. Colonization of the host relies on bacterial biofilm-like aggregation in the squid mucus field. Symbiotic biofilm formation is controlled by a two-component signaling (TCS) system consisting of regulators RscS-SypF-SypG, which together direct transcription of the Syp symbiotic polysaccharide. TCS systems are broadly important for bacteria to sense environmental cues and then direct changes in behavior. Previously, we identified hybrid histidine kinase BinK as a strong negative regulator of V. fischeri biofilm regulation, and here we further explore the function of BinK. To inhibit biofilm formation, BinK requires the predicted phosphorylation sites in both the histidine kinase (H362) and receiver (D794) domains. Furthermore, we show that RscS is not essential for host colonization when binK is deleted from strain ES114, and imaging of aggregate size revealed no benefit to the presence of RscS in a background lacking BinK. Strains lacking RscS still suffered in competition. Finally, we show that BinK functions to inhibit biofilm gene expression in the light organ crypts, providing evidence for biofilm gene regulation at later stages of host colonization. Overall, this study provides direct evidence for opposing activities of RscS and BinK and yields novel insights into biofilm regulation during the maturation of a beneficial symbiosis. IMPORTANCE Bacteria are often in a biofilm state, and transitions between planktonic and biofilm lifestyles are important for pathogenic, beneficial, and environmental microbes. The critical nature of biofilm formation during Vibrio fischeri colonization of the Hawaiian bobtail squid light organ provides an opportunity to study development of this process in vivo using a combination of genetic and imaging approaches. The current work refines the signaling circuitry of the biofilm pathway in V. fischeri, provides evidence that biofilm regulatory changes occur in the host, and identifies BinK as one of the regulators of that process. This study provides information about how bacteria regulate biofilm gene expression in an intact animal host.

scholarly journals The Cyclic-di-GMP Phosphodiesterase BinA Negatively Regulates Cellulose-Containing Biofilms in Vibrio fischeri

2010 ◽  
Vol 192 (5) ◽  
pp. 1269-1278 ◽  
Author(s):  
Christine M. Bassis ◽  
Karen L. Visick
Keyword(s):  
Amino Acids ◽  
Intracellular Signaling ◽  
Biofilm Formation ◽  
Vibrio Fischeri ◽  
Cellulose Biosynthesis ◽  
Euprymna Scolopes ◽  
Cyclic Diguanylate ◽  
Cellulose Production ◽  
Different Types ◽  
Symbiosis Polysaccharide

ABSTRACT Bacteria produce different types of biofilms under distinct environmental conditions. Vibrio fischeri has the capacity to produce at least two distinct types of biofilms, one that relies on the symbiosis polysaccharide Syp and another that depends upon cellulose. A key regulator of biofilm formation in bacteria is the intracellular signaling molecule cyclic diguanylate (c-di-GMP). In this study, we focused on a predicted c-di-GMP phosphodiesterase encoded by the gene binA, located directly downstream of syp, a cluster of 18 genes critical for biofilm formation and the initiation of symbiotic colonization of the squid Euprymna scolopes. Disruption or deletion of binA increased biofilm formation in culture and led to increased binding of Congo red and calcofluor, which are indicators of cellulose production. Using random transposon mutagenesis, we determined that the phenotypes of the ΔbinA mutant strain could be disrupted by insertions in genes in the bacterial cellulose biosynthesis cluster (bcs), suggesting that cellulose production is negatively regulated by BinA. Replacement of critical amino acids within the conserved EAL residues of the EAL domain disrupted BinA activity, and deletion of binA increased c-di-GMP levels in the cell. Together, these data support the hypotheses that BinA functions as a phosphodiesterase and that c-di-GMP activates cellulose biosynthesis. Finally, overexpression of the syp regulator sypG induced binA expression. Thus, this work reveals a mechanism by which V. fischeri inhibits cellulose-dependent biofilm formation and suggests that the production of two different polysaccharides may be coordinated through the action of the cellulose inhibitor BinA.

scholarly journals Control of competence in Vibrio fischeri

2021 ◽  
Author(s):  
Joshua J. Cohen ◽  
Steven J. Eichinger ◽  
Danae A. Witte ◽  
Connor J. Cook ◽  
Pat M. Fidopiastis ◽  
...  
Keyword(s):  
Biofilm Formation ◽  
Vibrio Fischeri ◽  
Genetic Manipulation ◽  
Model Organism ◽  
Dna Uptake ◽  
Host Colonization ◽  
Host Interactions ◽  
Light Production ◽  
Competence Factor ◽  
Control Transformation

Vibrio species, including the squid symbiont Vibrio fischeri, become competent to take up DNA under specific conditions. For example, V. fischeri becomes competent when grown in the presence of chitin oligosaccharides or upon overproduction of the competence regulatory factor TfoX. While little is known about the regulatory pathway(s) that control V. fischeri competence, this microbe encodes homologs of factors that control competence in the well-studied V. cholerae. To further develop V. fischeri as a genetically-tractable organism, we evaluated the roles of some of these competence homologs. Using TfoX-overproducing cells, we found that competence depends upon LitR, the homolog of V. cholerae master quorum sensing and competence regulator HapR, and on homologs of putative pili genes that in V. cholerae facilitate DNA uptake. Disruption of genes for negative regulators upstream of LitR, namely the LuxO protein and the sRNA Qrr1, resulted in increased transformation frequencies. Unlike LitR-controlled light production, however, competence did not vary with cell density under tfoX-overexpressing conditions. Analogous to V. cholerae, the requirement for LitR could be suppressed by loss of the Dns nuclease. We also found a role for the putative competence regulator CytR. Finally, we determined that transformation frequencies varied depending on the TfoX-encoding plasmid, and developed a new dual tfoX- and litR-overexpression construct that substantially increased the transformation frequency of a less genetically-tractable strain. By advancing the ease of genetic manipulation of V. fischeri, these findings will facilitate the rapid discovery of genes involved in physiologically-relevant processes, such as biofilm formation and host colonization. Importance The ability of bacteria to take up DNA (competence) and incorporate foreign DNA into their genomes (transformation) permits them to rapidly evolve and gain new traits and/or acquire antibiotic resistances. It also facilitates laboratory-based investigations into mechanisms of specific phenotypes, such as those involved in host colonization. Vibrio fischeri has long been a model for symbiotic bacteria-host interactions as well as for other aspects of its physiology such as bioluminescence and biofilm formation. Competence of V. fischeri can be readily induced upon overexpression of the competence factor TfoX. Relatively little is known about the V. fischeri competence pathway, although homologs of factors known to be important in V. cholerae competence exist. By probing the importance of putative competence factors that control transformation of V. fischeri, this work deepens our understanding of the competence process and advances our ability to genetically manipulate this important model organism.

scholarly journals Multiple Vibrio fischeri genes are involved in biofilm formation and host colonization

2012 ◽  
Vol 81 (3) ◽  
pp. 562-573 ◽  
Author(s):  
Alba Chavez-Dozal ◽  
David Hogan ◽  
Clayton Gorman ◽  
Alvaro Quintanal-Villalonga ◽  
Michele K. Nishiguchi
Keyword(s):  
Biofilm Formation ◽  
Vibrio Fischeri ◽  
Host Colonization

scholarly journals Vibrio fischeri Biofilm Formation Prevented by a Trio of Regulators

2018 ◽  
Vol 84 (19) ◽  
Author(s):  
Cecilia M. Thompson ◽  
Anne E. Marsden ◽  
Alice H. Tischler ◽  
Jovanka Koo ◽  
Karen L. Visick
Keyword(s):  
Biofilm Formation ◽  
Vibrio Fischeri ◽  
Negative Control ◽  
Negative Regulators ◽  
Content Type ◽  
Genetically Manipulated ◽  
Biofilm Development ◽  
Matrix Control ◽  
Symbiosis Polysaccharide

ABSTRACT Biofilms, complex communities of microorganisms surrounded by a self-produced matrix, facilitate attachment and provide protection to bacteria. A natural model used to study biofilm formation is the symbiosis between Vibrio fischeri and its host, the Hawaiian bobtail squid, Euprymna scolopes. Host-relevant biofilm formation is a tightly regulated process and is observed in vitro only with strains that have been genetically manipulated to overexpress or disrupt specific regulators, primarily two-component signaling (TCS) regulators. These regulators control biofilm formation by dictating the production of the symbiosis polysaccharide (Syp-PS), the major component of the biofilm matrix. Control occurs both at and below the level of transcription of the syp genes, which are responsible for Syp-PS production. Here, we probed the roles of the two known negative regulators of biofilm formation, BinK and SypE, by generating double mutants. We also mapped and evaluated a point mutation using natural transformation and linkage analysis. We examined traditional biofilm formation phenotypes and established a new assay for evaluating the start of biofilm formation in the form of microscopic aggregates in shaking liquid cultures, in the absence of the known biofilm-inducing signal calcium. We found that wrinkled colony formation is negatively controlled not only by BinK and SypE but also by SypF. SypF is both required for and inhibitory to biofilm formation. Together, these data reveal that these three regulators are sufficient to prevent wild-type V. fischeri from forming biofilms under these conditions. IMPORTANCE Bacterial biofilms promote attachment to a variety of surfaces and protect the constituent bacteria from environmental stresses, including antimicrobials. Understanding the mechanisms by which biofilms form will promote our ability to resolve them when they occur in the context of an infection. In this study, we found that Vibrio fischeri tightly controls biofilm formation using three negative regulators; the presence of a single one of these regulators was sufficient to prevent full biofilm development, while disruption of all three permitted robust biofilm formation. This work increases our understanding of the functions of specific regulators and demonstrates the substantial negative control that one benign microbe exerts over biofilm formation, potentially to ensure that it occurs only under the appropriate conditions.

scholarly journals Discovery of Calcium as a Biofilm-Promoting Signal for Vibrio fischeri Reveals New Phenotypes and Underlying Regulatory Complexity

2018 ◽  
Vol 200 (15) ◽  
Author(s):  
Alice H. Tischler ◽  
Louise Lie ◽  
Cecilia M. Thompson ◽  
Karen L. Visick
Keyword(s):  
Biofilm Formation ◽  
Vibrio Fischeri ◽  
Response Regulator ◽  
Negative Regulator ◽  
Regulatory Control ◽  
Cellulose Synthesis ◽  
Sensor Kinase ◽  
Content Type ◽  
First Time ◽  
Symbiosis Polysaccharide

ABSTRACT Vibrio fischeri uses biofilm formation to promote symbiotic colonization of its squid host, Euprymna scolopes. Control over biofilm formation is exerted at the level of transcription of the symbiosis polysaccharide (syp) locus by a complex set of two-component regulators. Biofilm formation can be induced by overproduction of the sensor kinase RscS, which requires the activities of the hybrid sensor kinase SypF and the response regulator SypG and is negatively regulated by the sensor kinase BinK. Here, we identify calcium as a signal that promotes biofilm formation by biofilm-competent strains under conditions in which biofilms are not typically observed (growth with shaking). This was true for RscS-overproducing cells as well as for strains in which only the negative regulator binK was deleted. The latter results provided, for the first time, an opportunity to induce and evaluate biofilm formation without regulator overexpression. Using these conditions, we determined that calcium induces both syp-dependent and bacterial cellulose synthesis (bcs)-dependent biofilms at the level of transcription of these loci. The calcium-induced biofilms were dependent on SypF, but SypF's Hpt domain was sufficient for biofilm formation. These data suggested the involvement of another sensor kinase(s) and led to the discovery that both RscS and a previously uncharacterized sensor kinase, HahK, functioned in this pathway. Together, the data presented here reveal both a new signal and biofilm phenotype produced by V. fischeri cells, the coordinate production of two polysaccharides involved in distinct biofilm behaviors, and a new regulator that contributes to control over these processes. IMPORTANCE Biofilms, or communities of surface-attached microorganisms adherent via a matrix that typically includes polysaccharides, are highly resistant to environmental stresses and are thus problematic in the clinic and important to study. Vibrio fischeri forms biofilms to colonize its symbiotic host, making this organism useful for studying biofilms. Biofilm formation depends on the syp polysaccharide locus and its regulators. Here, we identify a signal, calcium, that induces both SYP-PS and cellulose-dependent biofilms. We also identify a new syp regulator, the sensor kinase HahK, and discover a mutant phenotype for the sensor kinase RscS. This work thus reveals a specific biofilm-inducing signal that coordinately controls two polysaccharides, identifies a new regulator, and clarifies the regulatory control over biofilm formation by V. fischeri.

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