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Cyclic di-GMP-dependent signaling pathways in the pathogenic Firmicute Listeria monocytogenes.

Chen LH, Köseoğlu VK, Güvener ZT, Myers-Morales T, Reed JM, D'Orazio SE, Miller KW, Gomelsky M - PLoS Pathog. (2014)

Bottom Line: The last gene of the cluster encodes the fourth listerial GGDEF domain protein, PssE, that functions as an I-site c-di-GMP receptor essential for exopolysaccharide synthesis.The c-di-GMP-inducible exopolysaccharide causes cell aggregation in minimal medium and impairs bacterial migration in semi-solid agar, however, it does not promote biofilm formation on abiotic surfaces.The exopolysaccharide and another, as yet unknown c-di-GMP-dependent target, drastically decrease listerial invasiveness in enterocytes in vitro, and lower pathogen load in the liver and gallbladder of mice infected via an oral route, which suggests that elevated c-di-GMP levels play an overall negative role in listerial virulence.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, University of Wyoming, Laramie, Wyoming, United States of America.

ABSTRACT
We characterized key components and major targets of the c-di-GMP signaling pathways in the foodborne pathogen Listeria monocytogenes, identified a new c-di-GMP-inducible exopolysaccharide responsible for motility inhibition, cell aggregation, and enhanced tolerance to disinfectants and desiccation, and provided first insights into the role of c-di-GMP signaling in listerial virulence. Genome-wide genetic and biochemical analyses of c-di-GMP signaling pathways revealed that L. monocytogenes has three GGDEF domain proteins, DgcA (Lmo1911), DgcB (Lmo1912) and DgcC (Lmo2174), that possess diguanylate cyclase activity, and three EAL domain proteins, PdeB (Lmo0131), PdeC (Lmo1914) and PdeD (Lmo0111), that possess c-di-GMP phosphodiesterase activity. Deletion of all phosphodiesterase genes (ΔpdeB/C/D) or expression of a heterologous diguanylate cyclase stimulated production of a previously unknown exopolysaccharide. The synthesis of this exopolysaccharide was attributed to the pssA-E (lmo0527-0531) gene cluster. The last gene of the cluster encodes the fourth listerial GGDEF domain protein, PssE, that functions as an I-site c-di-GMP receptor essential for exopolysaccharide synthesis. The c-di-GMP-inducible exopolysaccharide causes cell aggregation in minimal medium and impairs bacterial migration in semi-solid agar, however, it does not promote biofilm formation on abiotic surfaces. The exopolysaccharide also greatly enhances bacterial tolerance to commonly used disinfectants as well as desiccation, which may contribute to survival of L. monocytogenes on contaminated food products and in food-processing facilities. The exopolysaccharide and another, as yet unknown c-di-GMP-dependent target, drastically decrease listerial invasiveness in enterocytes in vitro, and lower pathogen load in the liver and gallbladder of mice infected via an oral route, which suggests that elevated c-di-GMP levels play an overall negative role in listerial virulence.

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Impaired spreading of the L. monocytogenes ΔpdeB/C/D mutant to the liver and gallbladder in a foodborne model of infection.Groups of BALB/c/By/J mice were fed 5.9–7.5×108 CFU of the indicated L. monocytogenes strains and bacterial loads were assessed 60 h post-infection. Dashed lines indicate the limit of detection for each tissue. Bars denote mean values for pooled data from three separate experiments. **, significantly different (p<0.05). Prism 5 for Mac (GraphPad) was used to perform unpaired Student's t-tests.
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ppat-1004301-g008: Impaired spreading of the L. monocytogenes ΔpdeB/C/D mutant to the liver and gallbladder in a foodborne model of infection.Groups of BALB/c/By/J mice were fed 5.9–7.5×108 CFU of the indicated L. monocytogenes strains and bacterial loads were assessed 60 h post-infection. Dashed lines indicate the limit of detection for each tissue. Bars denote mean values for pooled data from three separate experiments. **, significantly different (p<0.05). Prism 5 for Mac (GraphPad) was used to perform unpaired Student's t-tests.

Mentions: To assess the role of c-di-GMP signaling in vivo, we used a newly developed mouse model of foodborne listeriosis [67]. Groups of BALB/c/By/J mice were fed either wild-type EGD-e or the ΔpdeB/C/D mutant, and the bacterial load in various tissues was assessed 60 h post infection. There was no significant difference in colonization of the ileum, colon or spleen at this time point (Fig. 8). However, the ΔpdeB/C/D triple mutant was significantly impaired in colonizing both the liver and the gallbladder. The decreased bacterial load in the liver appears to be linked to EPS, since the ΔpssC mutation in the ΔpdeB/C/D background restored the bacterial load to the wild-type level (Fig. 8). In fact, no significant differences in bacterial loads in the liver were observed when the same L. monocytogenes strains were injected intravenously (Fig. S2), suggesting that increased levels of c-di-GMP may alter the ability of the bacteria to disseminate from the gut.


Cyclic di-GMP-dependent signaling pathways in the pathogenic Firmicute Listeria monocytogenes.

Chen LH, Köseoğlu VK, Güvener ZT, Myers-Morales T, Reed JM, D'Orazio SE, Miller KW, Gomelsky M - PLoS Pathog. (2014)

Impaired spreading of the L. monocytogenes ΔpdeB/C/D mutant to the liver and gallbladder in a foodborne model of infection.Groups of BALB/c/By/J mice were fed 5.9–7.5×108 CFU of the indicated L. monocytogenes strains and bacterial loads were assessed 60 h post-infection. Dashed lines indicate the limit of detection for each tissue. Bars denote mean values for pooled data from three separate experiments. **, significantly different (p<0.05). Prism 5 for Mac (GraphPad) was used to perform unpaired Student's t-tests.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4125290&req=5

ppat-1004301-g008: Impaired spreading of the L. monocytogenes ΔpdeB/C/D mutant to the liver and gallbladder in a foodborne model of infection.Groups of BALB/c/By/J mice were fed 5.9–7.5×108 CFU of the indicated L. monocytogenes strains and bacterial loads were assessed 60 h post-infection. Dashed lines indicate the limit of detection for each tissue. Bars denote mean values for pooled data from three separate experiments. **, significantly different (p<0.05). Prism 5 for Mac (GraphPad) was used to perform unpaired Student's t-tests.
Mentions: To assess the role of c-di-GMP signaling in vivo, we used a newly developed mouse model of foodborne listeriosis [67]. Groups of BALB/c/By/J mice were fed either wild-type EGD-e or the ΔpdeB/C/D mutant, and the bacterial load in various tissues was assessed 60 h post infection. There was no significant difference in colonization of the ileum, colon or spleen at this time point (Fig. 8). However, the ΔpdeB/C/D triple mutant was significantly impaired in colonizing both the liver and the gallbladder. The decreased bacterial load in the liver appears to be linked to EPS, since the ΔpssC mutation in the ΔpdeB/C/D background restored the bacterial load to the wild-type level (Fig. 8). In fact, no significant differences in bacterial loads in the liver were observed when the same L. monocytogenes strains were injected intravenously (Fig. S2), suggesting that increased levels of c-di-GMP may alter the ability of the bacteria to disseminate from the gut.

Bottom Line: The last gene of the cluster encodes the fourth listerial GGDEF domain protein, PssE, that functions as an I-site c-di-GMP receptor essential for exopolysaccharide synthesis.The c-di-GMP-inducible exopolysaccharide causes cell aggregation in minimal medium and impairs bacterial migration in semi-solid agar, however, it does not promote biofilm formation on abiotic surfaces.The exopolysaccharide and another, as yet unknown c-di-GMP-dependent target, drastically decrease listerial invasiveness in enterocytes in vitro, and lower pathogen load in the liver and gallbladder of mice infected via an oral route, which suggests that elevated c-di-GMP levels play an overall negative role in listerial virulence.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, University of Wyoming, Laramie, Wyoming, United States of America.

ABSTRACT
We characterized key components and major targets of the c-di-GMP signaling pathways in the foodborne pathogen Listeria monocytogenes, identified a new c-di-GMP-inducible exopolysaccharide responsible for motility inhibition, cell aggregation, and enhanced tolerance to disinfectants and desiccation, and provided first insights into the role of c-di-GMP signaling in listerial virulence. Genome-wide genetic and biochemical analyses of c-di-GMP signaling pathways revealed that L. monocytogenes has three GGDEF domain proteins, DgcA (Lmo1911), DgcB (Lmo1912) and DgcC (Lmo2174), that possess diguanylate cyclase activity, and three EAL domain proteins, PdeB (Lmo0131), PdeC (Lmo1914) and PdeD (Lmo0111), that possess c-di-GMP phosphodiesterase activity. Deletion of all phosphodiesterase genes (ΔpdeB/C/D) or expression of a heterologous diguanylate cyclase stimulated production of a previously unknown exopolysaccharide. The synthesis of this exopolysaccharide was attributed to the pssA-E (lmo0527-0531) gene cluster. The last gene of the cluster encodes the fourth listerial GGDEF domain protein, PssE, that functions as an I-site c-di-GMP receptor essential for exopolysaccharide synthesis. The c-di-GMP-inducible exopolysaccharide causes cell aggregation in minimal medium and impairs bacterial migration in semi-solid agar, however, it does not promote biofilm formation on abiotic surfaces. The exopolysaccharide also greatly enhances bacterial tolerance to commonly used disinfectants as well as desiccation, which may contribute to survival of L. monocytogenes on contaminated food products and in food-processing facilities. The exopolysaccharide and another, as yet unknown c-di-GMP-dependent target, drastically decrease listerial invasiveness in enterocytes in vitro, and lower pathogen load in the liver and gallbladder of mice infected via an oral route, which suggests that elevated c-di-GMP levels play an overall negative role in listerial virulence.

Show MeSH
Related in: MedlinePlus