scholarly journals Investigation of a new mechanism of desiccation-stress tolerance in Salmonella

2013 ◽  
Author(s):  
Shlomo Sela ◽  
Michael McClelland
Keyword(s):  
Desiccation Tolerance ◽  
Biofilm Formation ◽  
Function Analysis ◽  
Swarming Motility ◽  
Comparable Amount ◽  
Dehydration Tolerance ◽  
Regulatory Pathways ◽  
Pathogen Survival ◽  
Surfactant Activity

Low-moisture foods (LMF) are increasingly involved in foodborne illness. While bacteria cannot grow in LMF due to the low water content, pathogens such as Salmonella can still survive in dry foods and pose health risks to consumer. We recently found that Salmonella secretes a proteinaceous compound during desiccation, which we identified as OsmY, an osmotic stress response protein of 177 amino acids. To elucidate the role of OsmY in conferring tolerance against desiccation and other stresses in Salmonella entericaserovarTyphimurium (STm), our specific objectives were: (1) Characterize the involvement of OsmY in desiccation tolerance; (2) Perform structure-function analysis of OsmY; (3) Study OsmY expression under various growth- and environmental conditions of relevance to agriculture; (4) Examine the involvement of OsmY in response to other stresses of relevance to agriculture; and (5) Elucidate regulatory pathways involved in controlling osmY expression. We demonstrated that an osmY-mutant strain is impaired in both desiccation tolerance (DT) and in long-term persistence during cold storage (LTP). Genetic complementation and addition of a recombinantOsmY (rOsmY) restored the mutant survival back to that of the wild type (wt). To analyze the function of specific domains we have generated a recombinantOsmY (rOsmY) protein. A dose-response DT study showed that rOsmY has the highest protection at a concentration of 0.5 nM. This effect was protein- specific as a comparable amount of bovine serum albumin, an unrelated protein, had a three-time lower protection level. Further characterization of OsmY revealed that the protein has a surfactant activity and is involved in swarming motility. OsmY was shown to facilitate biofilm formation during dehydration but not during bacterial growth under optimal growth conditions. This finding suggests that expression and secretion of OsmY under stress conditions was potentially associated with facilitating biofilm production. OsmY contains two conserved BON domains. To better understand the role of the BON sites in OsmY-mediated dehydration tolerance, we have generated two additional rOsmY constructs, lacking either BON1 or BON2 sites. BON1-minus (but not BON2) protein has decreased dehydration tolerance compared to intact rOsmY, suggesting that BON1 is required for maximal OsmY-mediated activity. Addition of BON1-peptide at concentration below 0.4 µM did not affect STm survival. Interestingly, a toxic effect of BON1 peptide was observed in concentration as low as 0.4 µM. Higher concentrations resulted in complete abrogation of the rOsmY effect, supporting the notion that BON-mediated interaction is essential for rOsmY activity. We performed extensive analysis of RNA expression of STm undergoing desiccation after exponential and stationary growth, identifying all categories of genes that are differentially expressed during this process. We also performed massively in-parallel screening of all genes in which mutation caused changes in fitness during drying, identifying over 400 such genes, which are now undergoing confirmation. As expected OsmY is one of these genes. In conclusion, this is the first study to identify that OsmY protein secreted during dehydration contributes to desiccation tolerance in Salmonella by facilitating dehydration- mediated biofilm formation. Expression of OsmY also enhances swarming motility, apparently through its surfactant activity. The BON1 domain is required for full OsmY activity, demonstrating a potential intervention to reduce pathogen survival in food processing. Expression and fitness screens have begun to elucidate the processes of desiccation, with the potential to uncover additional specific targets for efforts to mitigate pathogen survival in desiccation. 

scholarly journals Role of Intracellular Proteases in the Antibiotic Resistance, Motility, and Biofilm Formation of Pseudomonas aeruginosa

2011 ◽  
Vol 56 (2) ◽  
pp. 1128-1132 ◽  
Author(s):  
Lucía Fernández ◽  
Elena B. M. Breidenstein ◽  
Diana Song ◽  
Robert E. W. Hancock
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Antibiotic Resistance ◽  
Biofilm Formation ◽  
Regulatory Networks ◽  
Swarming Motility ◽  
Content Type ◽  
Adverse Conditions ◽  
Related Proteins ◽  
Intracellular Proteases

ABSTRACTPseudomonas aeruginosapossesses complex regulatory networks controlling virulence and survival under adverse conditions, including antibiotic pressure, which are interconnected and share common regulatory proteins. Here, we screen a panel of 13 mutants defective in intracellular proteases and demonstrate that, in addition to the known alterations in Lon and AsrA mutants, mutation of three protease-related proteins PfpI, ClpS, and ClpP differentially affected antibiotic resistance, swarming motility, and biofilm formation.

scholarly journals Analysis of the Pseudomonas aeruginosa Aminoglycoside Differential Resistomes Allows Defining Genes Simultaneously Involved in Intrinsic Antibiotic Resistance and Virulence

2019 ◽  
Vol 63 (5) ◽  
Author(s):  
Fernando Sanz-García ◽  
Carolina Alvarez-Ortega ◽  
Jorge Olivares-Pacheco ◽  
Paula Blanco ◽  
José Luis Martínez ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Biofilm Formation ◽  
High Throughput Screening ◽  
Elastase Activity ◽  
Swarming Motility ◽  
Content Type ◽  
Structural Family ◽  
Transposon Insertion ◽  
Bacterial Genes

ABSTRACT High-throughput screening of transposon insertion libraries is a useful strategy for unveiling bacterial genes whose inactivation results in an altered susceptibility to antibiotics. A potential drawback of these studies is they are usually based on just one model antibiotic for each structural family, under the assumption that the results can be extrapolated to all members of said family. To determine if this simplification is appropriate, we have analyzed the susceptibility of mutants of Pseudomonas aeruginosa to four aminoglycosides. Our results indicate that each mutation produces different effects on susceptibility to the tested aminoglycosides, with only two mutants showing similar changes in the susceptibility to all studied aminoglycosides. This indicates that the role of a particular gene in the resistome of a given antibiotic should not be generalized to other members of the same structural family. Five aminoglycoside-hypersusceptible mutants inactivating glnD, hflK, PA2798, PA3016, and hpf were chosen for further analysis in order to elucidate if lower aminoglycoside susceptibility correlates with cross-hypersusceptibility to other antibiotics and with impaired virulence. Our results indicate that glnD inactivation leads to increased cross-susceptibility to different antibiotics. The mutant in this gene is strongly impaired in virulence traits such as pyocyanin production, biofilm formation, elastase activity, and swarming motility and the ability to kill Caenorhabditis elegans. Thus, GlnD might be an interesting target for developing antibiotic coadjuvants with antiresistance and antivirulence properties against P. aeruginosa.

scholarly journals Putative RNA Ligase RtcB Affects the Switch between T6SS and T3SS in Pseudomonas aeruginosa

2021 ◽  
Vol 22 (22) ◽  
pp. 12561
Author(s):  
Maryam Dadashi ◽  
Lin Chen ◽  
Ahmad Nasimian ◽  
Saeid Ghavami ◽  
Kangmin Duan
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Biofilm Formation ◽  
Rna Binding ◽  
Opportunistic Pathogen ◽  
Virulence Determinants ◽  
Sequencing Data ◽  
Secretion Systems ◽  
Regulatory Pathways ◽  
Burn Victims

The opportunistic pathogen Pseudomonas aeruginosa is a significant cause of infection in immunocompromised individuals, cystic fibrosis patients, and burn victims. To benefit its survival, the bacterium adapt to either a motile or sessile lifestyle when infecting the host. The motile bacterium has an often activated type III secretion system (T3SS), which is virulent to the host, whereas the sessile bacterium harbors an active T6SS and lives in biofilms. Regulatory pathways involving Gac-Rsm or secondary messengers such as c-di-GMP determine which lifestyle is favorable for P. aeruginosa. Here, we introduce the RNA binding protein RtcB as a modulator of the switch between motile and sessile bacterial lifestyles. Using the wild-type P. aeruginosa PAO1, and a retS mutant PAO1(∆retS) in which T3SS is repressed and T6SS active, we show that deleting rtcB led to simultaneous expression of T3SS and T6SS in both PAO1(∆rtcB) and PAO1(∆rtcB∆retS). The deletion of rtcB also increased biofilm formation in PAO1(∆rtcB) and restored the motility of PAO1(∆rtcB∆retS). RNA-sequencing data suggested RtcB as a global modulator affecting multiple virulence factors, including bacterial secretion systems. Competitive killing and infection assays showed that the three T6SS systems (H1, H2, and H3) in PAO1(∆rtcB) were activated into a functional syringe, and could compete with Escherichia coli and effectively infect lettuce. Western blotting and RT-PCR results showed that RtcB probably exerted its function through RsmA in PAO1(∆rtcB∆retS). Quantification of c-di-GMP showed an elevated intracellular levels in PAO1(∆rtcB), which likely drove the switch between T6SS and T3SS, and contributed to the altered phenotypes and characteristics observed. Our data demonstrate a pivotal role of RtcB in the virulence of P. aeruginosa by controlling multiple virulence determinants, such as biofilm formation, motility, pyocyanin production, T3SS, and T6SS secretion systems towards eukaryotic and prokaryotic cells. These findings suggest RtcB as a potential target for controlling P. aeruginosa colonization, establishment, and pathogenicity.

scholarly journals Role of PvdQ in Pseudomonas aeruginosa virulence under iron-limiting conditions

Microbiology ◽  
2010 ◽  
Vol 156 (1) ◽  
pp. 49-59 ◽  
Author(s):  
Pol Nadal Jimenez ◽  
Gudrun Koch ◽  
Evelina Papaioannou ◽  
Mariana Wahjudi ◽  
Joanna Krzeslak ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Biofilm Formation ◽  
Iron Homeostasis ◽  
Gene Deletion ◽  
Infection Model ◽  
Swarming Motility ◽  
Low Concentrations ◽  
Biofilm Development ◽  
Long Chain

PvdQ, an acylase from Pseudomonas aeruginosa PAO1, has been shown to have at least two functions. It can act as a quorum quencher due to its ability to degrade long-chain N-acylhomoserine lactones (AHLs), e.g. 3-oxo-C12-HSL, leading to a decrease in virulence factors. In addition, PvdQ is involved in iron homeostasis by playing a role in the biosynthesis of pyoverdine, the major siderophore of P. aeruginosa. In accordance with earlier studies on RNA level, we could show at the protein level that PvdQ is only expressed when iron is present at very low concentrations. We therefore set out to investigate the two functions of PvdQ under iron-limiting conditions. Gene deletion of pvdQ does not affect growth of P. aeruginosa but abrogates pyoverdine production, and results in an accumulation of 3-oxo-C12-HSL. Phenotypic analyses of our ΔpvdQ mutant at low iron concentrations revealed that this mutant is impaired in swarming motility and biofilm formation. Additionally, a plant and a Caenorhabditis elegans infection model demonstrated that the deletion of pvdQ resulted in reduced virulence. None of the phenotypes in the present study could be linked to the presence or absence of AHLs. These results clearly indicate that under iron-limiting conditions PvdQ plays a major role in swarming motility, in biofilm development and in infection that is more likely to be linked to the pyoverdine pathway rather than the LasI/LasR/3-oxo-C12-HSL quorum-sensing circuit.

scholarly journals Sensor Kinase PA4398 Modulates Swarming Motility and Biofilm Formation in Pseudomonas aeruginosa PA14

2014 ◽  
Vol 81 (4) ◽  
pp. 1274-1285 ◽  
Author(s):  
Janine Strehmel ◽  
Anke Neidig ◽  
Michael Nusser ◽  
Robert Geffers ◽  
Gerald Brenner-Weiss ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Biofilm Formation ◽  
Sensor Kinase ◽  
Wild Type ◽  
Swarming Motility ◽  
Content Type ◽  
Regulatory Systems ◽  
Opportunistic Human Pathogen ◽  
Swarming Behavior

ABSTRACTPseudomonas aeruginosais an opportunistic human pathogen that is able to sense and adapt to numerous environmental stimuli by the use of transcriptional regulators, including two-component regulatory systems. In this study, we demonstrate that the sensor kinase PA4398 is involved in the regulation of swarming motility and biofilm formation inP. aeruginosaPA14. A PA4398−mutant strain was considerably impaired in swarming motility, while biofilm formation was increased by approximately 2-fold. The PA4398−mutant showed no changes in growth rate, rhamnolipid synthesis, or the production of the Pel exopolysaccharide but exhibited levels of the intracellular second messenger cyclic dimeric GMP (c-di-GMP) 50% higher than those in wild-type cells. The role of PA4398 in gene regulation was investigated by comparing the PA4398−mutant to the wild-type strain by using microarray analysis, which demonstrated that 64 genes were up- or downregulated more than 1.5-fold (P< 0.05) under swarming conditions. In addition, more-sensitive real-time PCR studies were performed on genes known to be involved in c-di-GMP metabolism. Among the dysregulated genes were several involved in the synthesis and degradation of c-di-GMP or in the biosynthesis, transport, or function of the iron-scavenging siderophores pyoverdine and pyochelin, in agreement with the swarming phenotype observed. By analyzing additional mutants of selected pyoverdine- and pyochelin-related genes, we were able to show that not onlypvdQbut alsopvdR,fptA,pchA,pchD, andpchHare essential for the normal swarming behavior ofP. aeruginosaPA14 and may also contribute to the swarming-deficient phenotype of the PA4398−mutant in addition to elevated c-di-GMP levels.

Defensive Role of Plant-Derived Secondary Metabolites: Indole and Its’ Derivatives

2020 ◽  
Vol 9 (2) ◽  
pp. 78-88
Author(s):  
Mulugeta Mulat ◽  
Raksha Anand ◽  
Fazlurrahman Khan
Keyword(s):  
Biofilm Formation ◽  
Growth And Development ◽  
Functional Role ◽  
Plant Pathogens ◽  
Bacterial Species ◽  
Indole Derivatives ◽  
Resistance Level ◽  
Defensive Role ◽  
Insect Attack

The diversity of indole concerning its production and functional role has increased in both prokaryotic and eukaryotic systems. The bacterial species produce indole and use it as a signaling molecule at interspecies, intraspecies, and even at an interkingdom level for controlling the capability of drug resistance, level of virulence, and biofilm formation. Numerous indole derivatives have been found to play an important role in the different systems and are reported to occur in various bacteria, plants, human, and plant pathogens. Indole and its derivatives have been recognized for a defensive role against pests and insects in the plant kingdom. These indole derivatives are produced as a result of the breakdown of glucosinolate products at the time of insect attack or physical damages. Apart from the defensive role of these products, in plants, they also exhibit several other secondary responses that may contribute directly or indirectly to the growth and development. The present review summarized recent signs of progress on the functional properties of indole and its derivatives in different plant systems. The molecular mechanism involved in the defensive role played by indole as well as its’ derivative in the plants has also been explained. Furthermore, the perspectives of indole and its derivatives (natural or synthetic) in understanding the involvement of these compounds in diverse plants have also been discussed.

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