scholarly journals Influence ofoxyRon Growth, Biofilm Formation, and Mobility of Vibrio parahaemolyticus

2015 ◽  
Vol 82 (3) ◽  
pp. 788-796 ◽  
Author(s):  
Chun-Hui Chung ◽  
Shin-yuan Fen ◽  
Shu-Chuan Yu ◽  
Hin-chung Wong
Keyword(s):  
Biofilm Formation ◽  
Vibrio Parahaemolyticus ◽  
Deletion Mutant ◽  
Cumene Hydroperoxide ◽  
Content Type ◽  
Antioxidative Function ◽  
Cell Mobility ◽  
Inhibition Of Growth ◽  
Dipotassium Hydrogen Phosphate ◽  
In The Wild

ABSTRACTVibrio parahaemolyticusis a common marine food-borne enteropathogen. In this study, we examined the antioxidative activity, growth, biofilm formation, and cell mobility of anoxyRdeletion mutant and its genetically complementary strain ofV. parahaemolyticus.oxyRis the regulator of catalase andahpCgenes. Protection against extrinsic H2O2and against the organic peroxides cumene hydroperoxide andtert-butyl hydroperoxide was weaker in the deletion mutant than in its parent strain. Expression of the major functional antioxidative genes,ahpC1and VPA1418, was markedly decreased in theoxyRmutant. Growth of this mutant on agar medium was significantly inhibited by autoclaved 0.25% glucose and by 0.25% dipotassium hydrogen phosphate, 0.5% monosaccharides (glucose, galactose, xylose, and arabinose), or 114.8 mM phosphates. The inhibition of the growth of thisoxyRmutant by extrinsic peroxides, autoclaved sugars, and phosphates was eliminated by the complementaryoxyRgene or by the addition of catalase to the autoclaved medium, while no inhibition of growth was observed when filter-sterilized sugars were used. The formation of biofilm and swimming mobility were significantly inhibited in theoxyRmutant relative to that in the wild-type strain. This investigation demonstrates the antioxidative function ofoxyRinV. parahaemolyticusand its possible roles in biofilm formation, cell mobility, and the protection of growth in heated rich medium.

scholarly journals Functions of VPA1418 and VPA0305 Catalase Genes in Growth of Vibrio parahaemolyticus under Oxidative Stress

2016 ◽  
Vol 82 (6) ◽  
pp. 1859-1867 ◽  
Author(s):  
Ching-Lian Chen ◽  
Shin-yuan Fen ◽  
Chun-Hui Chung ◽  
Shu-Chuan Yu ◽  
Cheng-Lun Chien ◽  
...  
Keyword(s):  
Vibrio Parahaemolyticus ◽  
Culture Medium ◽  
Incubation Temperature ◽  
Content Type ◽  
Inhibition Of Growth ◽  
Homologous Genes ◽  
Mutant Strains ◽  
In The Wild ◽  
Incubation Temperatures ◽  
Logarithmic Growth

ABSTRACTThe marine foodborne enteropathogenVibrio parahaemolyticushas four putative catalase genes. The functions of twokatE-homologous genes,katE1(VPA1418) andkatE2(VPA0305), in the growth of this bacterium were examined using gene deletion mutants with or without complementary genes. The growth of the mutant strains in static or shaken cultures in a rich medium at 37°C or at low temperatures (12 and 4°C), with or without competition fromEscherichia coli, did not differ from that of the parent strain. When 175 μM extrinsic H2O2was added to the culture medium, bacterial growth of the ΔkatE1strain was delayed and growth of the ΔkatE1ΔkatE2and ΔkatE1ΔahpC1double mutant strains was completely inhibited at 37°C for 8 h. The sensitivity of the ΔkatE1strain to the inhibition of growth by H2O2was higher at low incubation temperatures (12 and 22°C) than at 37°C. The determined gene expression of these catalase andahpCgenes revealed thatkatE1was highly expressed in the wild-type strain at 22°C under H2O2stress, while thekatE2andahpCgenes may play an alternate or compensatory role in the ΔkatE1strain. This study demonstrated thatkatE1encodes the chief functional catalase for detoxifying extrinsic H2O2during logarithmic growth and that the function of these genes was influenced by incubation temperature.

scholarly journals Homologous c-di-GMP-Binding Scr Transcription Factors Orchestrate Biofilm Development in Vibrio parahaemolyticus

2020 ◽  
Vol 202 (6) ◽  
Author(s):  
John H. Kimbrough ◽  
J. Thomas Cribbs ◽  
Linda L. McCarter
Keyword(s):  
Transcription Factors ◽  
Biofilm Formation ◽  
Vibrio Parahaemolyticus ◽  
Matrix Protein ◽  
Second Messenger ◽  
Minor Role ◽  
Binding Motif ◽  
Swarming Motility ◽  
Content Type ◽  
Biofilm Development

ABSTRACT The marine bacterium and human pathogen Vibrio parahaemolyticus rapidly colonizes surfaces by using swarming motility and forming robust biofilms. Entering one of the two colonization programs, swarming motility or sessility, involves differential regulation of many genes, resulting in a dramatic shift in physiology and behavior. V. parahaemolyticus has evolved complex regulation to control these two processes that have opposing outcomes. One mechanism relies on the balance of the second messenger c-di-GMP, where high c-di-GMP favors biofilm formation. V. parahaemolyticus possesses four homologous regulators, the Scr transcription factors, that belong in a Vibrio-specific family of W[F/L/M][T/S]R motif transcriptional regulators, some members of which have been demonstrated to bind c-di-GMP. In this work, we explore the role of these Scr regulators in biofilm development. We show that each protein binds c-di-GMP, that this binding requires a critical R in the binding motif, and that the biofilm-relevant activities of CpsQ, CpsS, and ScrO but not ScrP are dependent upon second messenger binding. ScrO and CpsQ are the primary drivers of biofilm formation, as biofilms are eliminated when both of these regulators are absent. ScrO is most important for capsule expression. CpsQ is most important for RTX-matrix protein expression, although it contributes to capsule expression when c-di-GMP levels are high. Both regulators contribute to O-antigen ligase expression. ScrP works oppositely in a minor role to repress the ligase gene. CpsS plays a regulatory checkpointing role by negatively modulating expression of these biofilm-pertinent genes under fluctuating c-di-GMP conditions. Our work further elucidates the multifactorial network that contributes to biofilm development in V. parahaemolyticus. IMPORTANCE Vibrio parahaemolyticus can inhabit open ocean, chitinous shells, and the human gut. Such varied habitats and the transitions between them require adaptable regulatory networks controlling energetically expensive behaviors, including swarming motility and biofilm formation, which are promoted by low and high concentrations of the signaling molecule c-di-GMP, respectively. Here, we describe four homologous c-di-GMP-binding Scr transcription factors in V. parahaemolyticus. Members of this family of regulators are present in many vibrios, yet their numbers and the natures of their activities differ across species. Our work highlights the distinctive roles that these transcription factors play in dynamically controlling biofilm formation and architecture in V. parahaemolyticus and serves as a powerful example of regulatory network evolution and diversification.

scholarly journals Spermidine Inversely Influences Surface Interactions and Planktonic Growth in Agrobacterium tumefaciens

2016 ◽  
Vol 198 (19) ◽  
pp. 2682-2691 ◽  
Author(s):  
Yi Wang ◽  
Sok Ho Kim ◽  
Ramya Natarajan ◽  
Jason E. Heindl ◽  
Eric L. Bruger ◽  
...  
Keyword(s):  
Agrobacterium Tumefaciens ◽  
Biofilm Formation ◽  
Direct Synthesis ◽  
Wild Type ◽  
Cyclic Diguanylate ◽  
Content Type ◽  
Exogenous Addition ◽  
In The Wild ◽  
Growth Requirement ◽  
Transposon Insertions

ABSTRACTIn bacteria, the functions of polyamines, small linear polycations, are poorly defined, but these metabolites can influence biofilm formation in several systems. Transposon insertions in an ornithine decarboxylase (odc) gene inAgrobacterium tumefaciens, predicted to direct synthesis of the polyamine putrescine from ornithine, resulted in elevated cellulose. Null mutants forodcgrew somewhat slowly in a polyamine-free medium but exhibited increased biofilm formation that was dependent on cellulose production. Spermidine is an essential metabolite inA. tumefaciensand is synthesized from putrescine inA. tumefaciensvia the stepwise actions of carboxyspermidine dehydrogenase (CASDH) and carboxyspermidine decarboxylase (CASDC). Exogenous addition of either putrescine or spermidine to theodcmutant returned biofilm formation to wild-type levels. Low levels of exogenous spermidine restored growth to CASDH and CASDC mutants, facilitating weak biofilm formation, but this was dampened with increasing concentrations. Norspermidine rescued growth for theodc, CASDH, and CASDC mutants but did not significantly affect their biofilm phenotypes, whereas in the wild type, it stimulated biofilm formation and depressed spermidine levels. Theodcmutant produced elevated levels of cyclic diguanylate monophosphate (c-di-GMP), exogenous polyamines modulated these levels, and expression of a c-di-GMP phosphodiesterase reversed the enhanced biofilm formation. Prior work revealed accumulation of the precursors putrescine and carboxyspermidine in the CASDH and CASDC mutants, respectively, but unexpectedly, both mutants accumulated homospermidine; here, we show that this requires a homospermidine synthase (hss) homologue.IMPORTANCEPolyamines are small, positively charged metabolites that are nearly ubiquitous in cellular life. They are often essential in eukaryotes and more variably in bacteria. Polyamines have been reported to influence the surface-attached biofilm formation of several bacteria. InAgrobacterium tumefaciens, mutants with diminished levels of the polyamine spermidine are stimulated for biofilm formation, and exogenous provision of spermidine decreases biofilm formation. Spermidine is also essential forA. tumefaciensgrowth, but the related polyamine norspermidine exogenously rescues growth and does not diminish biofilm formation, revealing that the growth requirement and biofilm control are separable. Polyamine control of biofilm formation appears to function via effects on the cellular second messenger cyclic diguanylate monophosphate, regulating the transition from a free-living to a surface-attached lifestyle.

scholarly journals Complete Genome Sequence of a Deletion Mutant of Vibrio parahaemolyticus from Pacific White Shrimp (Penaeus vannamei)

2018 ◽  
Vol 6 (25) ◽  
Author(s):  
Siddhartha Kanrar ◽  
Arun K. Dhar
Keyword(s):  
Genome Sequence ◽  
Vibrio Parahaemolyticus ◽  
Deletion Mutant ◽  
Pacific White Shrimp ◽  
White Shrimp ◽  
Penaeus Vannamei ◽  
Content Type ◽  
A Genome ◽  
Acute Hepatopancreatic Necrosis Disease ◽  
Experimental Challenge

ABSTRACT Vibrio parahaemolyticus carrying the toxin genes pirA and pirB causes acute hepatopancreatic necrosis disease in shrimp. A genome sequence of V. parahaemolyticus strain R13 was determined that showed deletions of the entire pirA gene and the 5ʹ end of the pirB gene and does not cause the disease in experimental challenge.

scholarly journals Deletion Mutant Library for Investigation of Functional Outputs of Cyclic Diguanylate Metabolism in Pseudomonas aeruginosa PA14

2014 ◽  
Vol 80 (11) ◽  
pp. 3384-3393 ◽  
Author(s):  
Dae-Gon Ha ◽  
Megan E. Richman ◽  
George A. O'Toole
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Biofilm Formation ◽  
Deletion Mutant ◽  
Deletion Mutants ◽  
Mutant Library ◽  
Potential Utility ◽  
Cyclic Diguanylate ◽  
Swarming Motility ◽  
Content Type ◽  
Swimming Motility

ABSTRACTWe constructed a library of in-frame deletion mutants targeting each gene inPseudomonas aeruginosaPA14 predicted to participate in cyclic di-GMP (c-di-GMP) metabolism (biosynthesis or degradation) to provide a toolkit to assist investigators studying c-di-GMP-mediated regulation by this microbe. We present phenotypic assessments of each mutant, including biofilm formation, exopolysaccharide (EPS) production, swimming motility, swarming motility, and twitch motility, as a means to initially characterize these mutants and to demonstrate the potential utility of this library.

scholarly journals Involvement of Stress-Related GenespolBandPA14_46880in Biofilm Formation of Pseudomonas aeruginosa

2014 ◽  
Vol 82 (11) ◽  
pp. 4746-4757 ◽  
Author(s):  
Sahar A. Alshalchi ◽  
Gregory G. Anderson
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Biofilm Formation ◽  
Acute Infection ◽  
Regulatory Gene ◽  
Chronic Infections ◽  
Content Type ◽  
Abiotic Surfaces ◽  
Mutant Strains ◽  
In The Wild ◽  
Airway Cells

ABSTRACTChronic infections ofPseudomonas aeruginosaare generally established through production of biofilm. During biofilm formation, production of an extracellular matrix and establishment of a distinct bacterial phenotype make these infections difficult to eradicate. However, biofilm studies have been hampered by the fact that most assays utilize nonliving surfaces as biofilm attachment substrates. In an attempt to better understand the mechanisms behindP. aeruginosabiofilm formation, we performed a genetic screen to identify novel factors involved in biofilm formation on biotic and abiotic surfaces. We found that deletion of genespolBandPA14_46880reduced biofilm formation significantly compared to that in the wild-type strain PA14 in an abiotic biofilm system. In a biotic biofilm model, wherein biofilms form on cultured airway cells, the ΔpolBand ΔPA14_46880strains showed increased cytotoxic killing of the airway cells independent of the total number of bacteria bound. Notably, deletion mutant strains were more resistant to ciprofloxacin treatment. This phenotype was linked to decreased expression ofalgR, an alginate transcriptional regulatory gene, under ciprofloxacin pressure. Moreover, we found that pyocyanin production was increased in planktonic cells of mutant strains. These results indicate that inactivation ofpolBandPA14_46880may inhibit transition ofP. aeruginosafrom a more acute infection lifestyle to the biofilm phenotype. Future investigation of these genes may lead to a better understanding ofP. aeruginosabiofilm formation and chronic biofilm infections.

scholarly journals AcheZ-Like Gene inAzorhizobium caulinodansIs a Key Gene in the Control of Chemotaxis and Colonization of the Host Plant

2017 ◽  
Vol 84 (3) ◽  
Author(s):  
Xiaolin Liu ◽  
Wei Liu ◽  
Yu Sun ◽  
Chunlei Xia ◽  
Claudine Elmerich ◽  
...  
Keyword(s):  
Host Plant ◽  
Active Site ◽  
Deletion Mutant ◽  
Sinorhizobium Meliloti ◽  
Response Regulator ◽  
General Situation ◽  
Symbiotic Association ◽  
Azorhizobium Caulinodans ◽  
Content Type ◽  
In The Wild

ABSTRACTChemotaxis can provide bacteria with competitive advantages for survival in complex environments. The CheZ chemotaxis protein is a phosphatase, affecting the flagellar motor inEscherichia coliby dephosphorylating the response regulator phosphorylated CheY protein (CheY∼P) responsible for clockwise rotation. AcheZgene has been found inAzorhizobium caulinodansORS571, in contrast to other rhizobial species studied so far. The CheZ protein in strain ORS571 has a conserved motif similar to that corresponding to the phosphatase active site inE. coli. The construction of acheZdeletion mutant strain and ofcheZmutant strains carrying a mutation in residues of the putative phosphatase active site showed that strain ORS571 participates in chemotaxis and motility, causing a hyperreversal behavior. In addition, the properties of thecheZdeletion mutant revealed that ORS571 CheZ is involved in other physiological processes, since it displayed increased flocculation, biofilm formation, exopolysaccharide (EPS) production, and host root colonization. In particular, it was observed that the expression of severalexpgenes, involved in EPS synthesis, was upregulated in thecheZmutant compared to that in the wild type, suggesting that CheZ negatively controlsexpgene expression through an unknown mechanism. It is proposed that CheZ influences theAzorhizobium-plant association by negatively regulating early colonization via the regulation of EPS production. This report established that CheZ inA. caulinodansplays roles in chemotaxis and the symbiotic association with the host plant.IMPORTANCEChemotaxis allows bacteria to swim toward plant roots and is beneficial to the establishment of various plant-microbe associations. The level of CheY phosphorylation (CheY∼P) is central to the chemotaxis signal transduction. The mechanism of the signal termination of CheY∼P remains poorly characterized amongAlphaproteobacteria, except forSinorhizobium meliloti, which does not contain CheZ but which controls CheY∼P dephosphorylation through a phosphate sink mechanism.Azorhizobium caulinodansORS571, a microsymbiont ofSesbania rostrata, has an orphancheZgene besides twocheYgenes similar to those inS. meliloti. In addition to controlling the chemotaxis response, the CheZ-like protein in strain ORS571 is playing a role by decreasing bacterial adhesion to the host plant, in contrast to the general situation where chemotaxis-associated proteins promote adhesion. In this study, we identified a CheZ-like protein amongAlphaproteobacteriafunctioning in chemotaxis and theA. caulinodans-S. rostratasymbiosis.

scholarly journals Activities of Alkyl Hydroperoxide Reductase Subunits C1 and C2 of Vibrio parahaemolyticus against Different Peroxides

2014 ◽  
Vol 80 (23) ◽  
pp. 7398-7404 ◽  
Author(s):  
Chun-Hui Chung ◽  
Tsung-yong Ma ◽  
Shin-yuan Fen ◽  
Hin-chung Wong
Keyword(s):  
Liquid Medium ◽  
Vibrio Parahaemolyticus ◽  
Vibrio Vulnificus ◽  
Incubation Temperature ◽  
Cumene Hydroperoxide ◽  
Organic Peroxides ◽  
Homologous Proteins ◽  
Alkyl Hydroperoxide Reductase ◽  
Content Type ◽  
Alkyl Hydroperoxide

ABSTRACTAlkyl hydroperoxide reductase subunit C gene (ahpC) functions were characterized inVibrio parahaemolyticus, a commonly occurring marine food-borne enteropathogenic bacterium. TwoahpCgenes,ahpC1(VPA1683) andahpC2(VP0580), encoded putative two-cysteine peroxiredoxins, which are highly similar to the homologous proteins ofVibrio vulnificus. The responses of deletion mutants ofahpCgenes to various peroxides were compared with and without gene complementation and at different incubation temperatures. The growth of theahpC1mutant andahpC1 ahpC2double mutant in liquid medium was significantly inhibited by organic peroxides, cumene hydroperoxide andtert-butyl hydroperoxide. However, inhibition was higher at 12°C and 22°C than at 37°C. Inhibiting effects were prevented by the complementaryahpC1gene. Inconsistent detoxification of H2O2byahpCgenes was demonstrated in an agar medium but not in a liquid medium. Complementation with anahpC2gene partially restored the peroxidase effect in the doubleahpC1 ahpC2mutant at 22°C. This investigation reveals thatahpC1is the chief peroxidase gene that acts against organic peroxides inV. parahaemolyticusand that the function of theahpCgenes is influenced by incubation temperature.

scholarly journals Uncovering Roles of Streptococcus gordonii SrtA-Processed Proteins in the Biofilm Lifestyle

2020 ◽  
Vol 203 (2) ◽  
pp. e00544-20
Author(s):  
Brittany L. Nairn ◽  
Grace T. Lee ◽  
Ashwani K. Chumber ◽  
Patrick R. Steck ◽  
Mahmoud O. Mire ◽  
...  
Keyword(s):  
Cell Wall ◽  
Biofilm Formation ◽  
Deletion Mutant ◽  
Ex Vivo ◽  
Sortase A ◽  
Cell Wall Proteins ◽  
Mutant Library ◽  
Rich Medium ◽  
Content Type ◽  
Markerless Deletion

ABSTRACTStreptococcus gordonii is a commensal oral organism. Harmless in the oral cavity, S. gordonii is an opportunistic pathogen. S. gordonii adheres to body surfaces using surface adhesive proteins (adhesins), which are critical to subsequent formation of biofilm communities. As in most Gram-positive bacteria, S. gordonii surface proteins containing the C-terminal LPXTG motif cleavage sequence are processed by sortase A (SrtA) to become covalently attached to the cell wall. To characterize the functional diversity and redundancy in the family of SrtA-processed proteins, an S. gordonii DL1 markerless deletion mutant library was constructed of each of the 26 putative SrtA-processed proteins. Each library member was evaluated for growth in rich medium, biofilm formation on plastic, saliva and salivary fractions, cell surface hydrophobicity (CSH), hemagglutination, and integration into an ex vivo plaque biofilm community. Library members were compared to the non-SrtA-processed adhesins AbpA and AbpB. While no major growth differences in rich medium were observed, many S. gordonii LPXTG/A proteins impacted biofilm formation on one or more of the substrates. Several mutants showed significant differences in hemagglutination, hydrophobicity, or fitness in the ex vivo plaque model. From the identification of redundant and unique functions in these in vitro and ex vivo systems, functional stratification among the LPXTG/A proteins is apparent.IMPORTANCES. gordonii interactions with its environment depend on the complement of cell wall proteins. A subset of these cell wall proteins requires processing by the enzyme sortase A (SrtA). The identification of SrtA-processed proteins and their functional characterization will help the community to better understand how S. gordonii engages with its surroundings, including other microbes, integrates into the plaque community, adheres to the tooth surface, and hematogenously disseminates to cause blood-borne infections. This study identified 26 putative SrtA-processed proteins through creation of a markerless deletion mutant library. The library was subject to functional screens that were chosen to better understand key aspects of S. gordonii physiology and pathogenesis.

scholarly journals Inactivation ofcysLInhibits Biofilm Formation by Activating the Disulfide Stress Regulator Spx inBacillus subtilis

2019 ◽  
Vol 201 (8) ◽  
Author(s):  
Kazuo Kobayashi
Keyword(s):  
Bacillus Subtilis ◽  
Biofilm Formation ◽  
Deletion Mutant ◽  
Reduced Form ◽  
Sulfite Reductase ◽  
Dependent Manner ◽  
Cysteine Biosynthesis ◽  
Content Type ◽  
Phosphoadenosine Phosphosulfate ◽  
Biofilm Matrix

ABSTRACTBacillus subtilisforms biofilms in response to internal and external stimuli. I previously showed that thecysLdeletion mutant was defective in biofilm formation, but the reason for this remains unidentified. CysL is a transcriptional activator of thecysJIoperon, which encodes sulfite reductase, an enzyme involved in cysteine biosynthesis. Decreased production of sulfite reductase led to biofilm formation defects in the ΔcysLmutant. The ΔcysLmutation was suppressed by disruptingcysHoperon genes, whose products function upstream of sulfite reductase in the cysteine biosynthesis pathway, indicating that defects in cysteine biosynthesis were not a direct cause for the defective biofilm formation observed in the ΔcysLmutant. ThecysHgene encodes phosphoadenosine phosphosulfate reductase, which requires a reduced form of thioredoxin (TrxA) as an electron donor. High expression oftrxAinhibited biofilm formation in the ΔcysLmutant but not in the wild-type strain. Northern blot analysis showed thattrxAtranscription was induced in the ΔcysLmutant in a disulfide stress-induced regulator Spx-dependent manner. On the basis of these results, I propose that the ΔcysLmutation causes phosphoadenosine phosphosulfate reductase to consume large amounts of reduced thioredoxin, inducing disulfide stress and activating Spx. Thespxmutation restored biofilm formation to the ΔcysLmutant. The ΔcysLmutation reduced expression of theepsoperon, which is required for exopolysaccharide production. Moreover, overexpression of theepsoperon restored biofilm formation to the ΔcysLmutant. Taken together, these results suggest that the ΔcysLmutation activates Spx, which then inhibits biofilm formation through repression of theepsoperon.IMPORTANCEBacillus subtilishas been studied as a model organism for biofilm formation. In this study, I explored why thecysLdeletion mutant was defective in biofilm formation. I demonstrated that the ΔcysLmutation activated the disulfide stress response regulator Spx, which inhibits biofilm formation by repressing biofilm matrix genes. Homologs of Spx are highly conserved among Gram-positive bacteria with low G+C contents. In some pathogens, Spx is also reported to inhibit biofilm formation by repressing biofilm matrix genes, even though these genes and their regulation are quite different from those ofB. subtilis. Thus, the negative regulation of biofilm formation by Spx is likely to be well conserved across species and may be an appropriate target for control of biofilm formation.

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