Limits...
Pleiotropic phenotypes of a Yersinia enterocolitica flhD mutant include reduced lethality in a chicken embryo model.

Townsend MK, Carr NJ, Iyer JG, Horne SM, Gibbs PS, Prüss BM - BMC Microbiol. (2008)

Bottom Line: Compared to the wild-type strain, isogenic flhD and fliA mutants exhibited increased growth on purines and reduced growth on N-acetyl-D-glucosamine and D-mannose, when used as a sole carbon source.The flhD mutant caused reduced chicken embryo lethality when compared to wild-type bacteria.Phenotypes of flhD and fliA mutants are related to central metabolism and virulence and correlate with gene regulation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Veterinary and Microbiological Sciences, North Dakota State University, Fargo, USA. megan.townsend@ndsu.edu

ABSTRACT

Background: The Yersinia enterocolitica flagellar master regulator FlhD/FlhC affects the expression levels of non-flagellar genes, including 21 genes that are involved in central metabolism. The sigma factor of the flagellar system, FliA, has a negative effect on the expression levels of seven plasmid-encoded virulence genes in addition to its positive effect on the expression levels of eight of the flagellar operons. This study investigates the phenotypes of flhD and fliA mutants that result from the complex gene regulation.

Results: Phenotypes relating to central metabolism were investigated with Phenotype MicroArrays. Compared to the wild-type strain, isogenic flhD and fliA mutants exhibited increased growth on purines and reduced growth on N-acetyl-D-glucosamine and D-mannose, when used as a sole carbon source. Both mutants grew more poorly on pyrimidines and L-histidine as sole nitrogen source. Several intermediates of the tricarboxylic acid and the urea cycle, as well as several dipeptides, provided differential growth conditions for the two mutants. Gene expression was determined for selected genes and correlated with the observed phenotypes. Phenotypes relating to virulence were determined with the chicken embryo lethality assay. The assay that was previously established for Escherichia coli strains was modified for Y. enterocolitica. The flhD mutant caused reduced chicken embryo lethality when compared to wild-type bacteria. In contrast, the fliA mutant caused wild-type lethality. This indicates that the virulence phenotype of the flhD mutant might be due to genes that are regulated by FlhD/FlhC but not FliA, such as those that encode the flagellar type III secretion system.

Conclusion: Phenotypes of flhD and fliA mutants are related to central metabolism and virulence and correlate with gene regulation.

Show MeSH

Related in: MedlinePlus

Phenotype MicroArrays, nutrients. Growth on sole carbon and nitrogen sources was determined with Phenotype MicroArrays (Biolog, Hayward, CA) for wild-type 8081c cells and their isogenic flhD and fliA mutants. Bacterial solutions in IF-0 were pipetted onto PM1 (carbon sources, Panels A and C) and PM3 (nitrogen sources, Panels B and D) plates. Plates were incubated at 25°C (Panels A and B) or 4°C (Panels C and D). Growth on all plates was determined as OD630. The background OD630 (obtained from well A1) was subtracted from the OD630 for each nutrient on each plate. Growth ratios are expressed as OD630 of the wild-type cells divided by OD630 of each of the mutant strains (darker bars, flhD mutant; lighter bars, fliA mutant) for each nutrient. Nutrients that provided better growth conditions for the wild-type cells are printed in green, nutrients that provided better growth conditions for the mutants are printed in red. Dipeptides are printed in blue. The experiment was performed three to four times and the mean of the population is presented. Error bars indicate one standard deviation (1 s).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC2262085&req=5

Figure 1: Phenotype MicroArrays, nutrients. Growth on sole carbon and nitrogen sources was determined with Phenotype MicroArrays (Biolog, Hayward, CA) for wild-type 8081c cells and their isogenic flhD and fliA mutants. Bacterial solutions in IF-0 were pipetted onto PM1 (carbon sources, Panels A and C) and PM3 (nitrogen sources, Panels B and D) plates. Plates were incubated at 25°C (Panels A and B) or 4°C (Panels C and D). Growth on all plates was determined as OD630. The background OD630 (obtained from well A1) was subtracted from the OD630 for each nutrient on each plate. Growth ratios are expressed as OD630 of the wild-type cells divided by OD630 of each of the mutant strains (darker bars, flhD mutant; lighter bars, fliA mutant) for each nutrient. Nutrients that provided better growth conditions for the wild-type cells are printed in green, nutrients that provided better growth conditions for the mutants are printed in red. Dipeptides are printed in blue. The experiment was performed three to four times and the mean of the population is presented. Error bars indicate one standard deviation (1 s).

Mentions: Fig. 1 shows the determined phenotypes of the strains on carbon and nitrogen sources. At 25°C (Panel A), flhD mutants grew better on the purines adenosine, inosine, and 2-deoxyadenosine as carbon sources, as well as acetic acid, L-asparagine, L-glutamate, and D-alanine. Wild-type cells grew better on N-acetyl-D-glucosamine and D-mannose. Wild-type cells also grew better on L-histidine, the pyrimidines cytodine, cytosine, uridine, and thymidine, in addition to D-glucosamine, and N-acetyl-D-glucosamine as nitrogen sources (Panel B). The mutants grew better on L-asparagine and L-proline. In addition, several dipeptides provided differential growth conditions for wild-type cells and flhD mutants. No difference was observed between flhD and fliA mutants.


Pleiotropic phenotypes of a Yersinia enterocolitica flhD mutant include reduced lethality in a chicken embryo model.

Townsend MK, Carr NJ, Iyer JG, Horne SM, Gibbs PS, Prüss BM - BMC Microbiol. (2008)

Phenotype MicroArrays, nutrients. Growth on sole carbon and nitrogen sources was determined with Phenotype MicroArrays (Biolog, Hayward, CA) for wild-type 8081c cells and their isogenic flhD and fliA mutants. Bacterial solutions in IF-0 were pipetted onto PM1 (carbon sources, Panels A and C) and PM3 (nitrogen sources, Panels B and D) plates. Plates were incubated at 25°C (Panels A and B) or 4°C (Panels C and D). Growth on all plates was determined as OD630. The background OD630 (obtained from well A1) was subtracted from the OD630 for each nutrient on each plate. Growth ratios are expressed as OD630 of the wild-type cells divided by OD630 of each of the mutant strains (darker bars, flhD mutant; lighter bars, fliA mutant) for each nutrient. Nutrients that provided better growth conditions for the wild-type cells are printed in green, nutrients that provided better growth conditions for the mutants are printed in red. Dipeptides are printed in blue. The experiment was performed three to four times and the mean of the population is presented. Error bars indicate one standard deviation (1 s).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2262085&req=5

Figure 1: Phenotype MicroArrays, nutrients. Growth on sole carbon and nitrogen sources was determined with Phenotype MicroArrays (Biolog, Hayward, CA) for wild-type 8081c cells and their isogenic flhD and fliA mutants. Bacterial solutions in IF-0 were pipetted onto PM1 (carbon sources, Panels A and C) and PM3 (nitrogen sources, Panels B and D) plates. Plates were incubated at 25°C (Panels A and B) or 4°C (Panels C and D). Growth on all plates was determined as OD630. The background OD630 (obtained from well A1) was subtracted from the OD630 for each nutrient on each plate. Growth ratios are expressed as OD630 of the wild-type cells divided by OD630 of each of the mutant strains (darker bars, flhD mutant; lighter bars, fliA mutant) for each nutrient. Nutrients that provided better growth conditions for the wild-type cells are printed in green, nutrients that provided better growth conditions for the mutants are printed in red. Dipeptides are printed in blue. The experiment was performed three to four times and the mean of the population is presented. Error bars indicate one standard deviation (1 s).
Mentions: Fig. 1 shows the determined phenotypes of the strains on carbon and nitrogen sources. At 25°C (Panel A), flhD mutants grew better on the purines adenosine, inosine, and 2-deoxyadenosine as carbon sources, as well as acetic acid, L-asparagine, L-glutamate, and D-alanine. Wild-type cells grew better on N-acetyl-D-glucosamine and D-mannose. Wild-type cells also grew better on L-histidine, the pyrimidines cytodine, cytosine, uridine, and thymidine, in addition to D-glucosamine, and N-acetyl-D-glucosamine as nitrogen sources (Panel B). The mutants grew better on L-asparagine and L-proline. In addition, several dipeptides provided differential growth conditions for wild-type cells and flhD mutants. No difference was observed between flhD and fliA mutants.

Bottom Line: Compared to the wild-type strain, isogenic flhD and fliA mutants exhibited increased growth on purines and reduced growth on N-acetyl-D-glucosamine and D-mannose, when used as a sole carbon source.The flhD mutant caused reduced chicken embryo lethality when compared to wild-type bacteria.Phenotypes of flhD and fliA mutants are related to central metabolism and virulence and correlate with gene regulation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Veterinary and Microbiological Sciences, North Dakota State University, Fargo, USA. megan.townsend@ndsu.edu

ABSTRACT

Background: The Yersinia enterocolitica flagellar master regulator FlhD/FlhC affects the expression levels of non-flagellar genes, including 21 genes that are involved in central metabolism. The sigma factor of the flagellar system, FliA, has a negative effect on the expression levels of seven plasmid-encoded virulence genes in addition to its positive effect on the expression levels of eight of the flagellar operons. This study investigates the phenotypes of flhD and fliA mutants that result from the complex gene regulation.

Results: Phenotypes relating to central metabolism were investigated with Phenotype MicroArrays. Compared to the wild-type strain, isogenic flhD and fliA mutants exhibited increased growth on purines and reduced growth on N-acetyl-D-glucosamine and D-mannose, when used as a sole carbon source. Both mutants grew more poorly on pyrimidines and L-histidine as sole nitrogen source. Several intermediates of the tricarboxylic acid and the urea cycle, as well as several dipeptides, provided differential growth conditions for the two mutants. Gene expression was determined for selected genes and correlated with the observed phenotypes. Phenotypes relating to virulence were determined with the chicken embryo lethality assay. The assay that was previously established for Escherichia coli strains was modified for Y. enterocolitica. The flhD mutant caused reduced chicken embryo lethality when compared to wild-type bacteria. In contrast, the fliA mutant caused wild-type lethality. This indicates that the virulence phenotype of the flhD mutant might be due to genes that are regulated by FlhD/FlhC but not FliA, such as those that encode the flagellar type III secretion system.

Conclusion: Phenotypes of flhD and fliA mutants are related to central metabolism and virulence and correlate with gene regulation.

Show MeSH
Related in: MedlinePlus