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Phase variation of poly-N-acetylglucosamine expression in Staphylococcus aureus.

Brooks JL, Jefferson KK - PLoS Pathog. (2014)

Bottom Line: Inactivation of IcaC results in a PIA/PNAG-negative phenotype.There was also a survival advantage for an icaC-negative strain harboring intact icaADB genes relative to an isogenic icaADBC deletion mutant.Together, these results suggest that inactivation of icaC is a mode of phase variation for PIA/PNAG expression, that high-level production of PIA/PNAG carries a fitness cost, and that icaADB may contribute to bacterial fitness, by an unknown mechanism, in the absence of an intact icaC gene and PIA/PNAG production.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Virginia Commonwealth University School of Medicine, Richmond, Virginia, United States of America.

ABSTRACT
Polysaccharide intercellular adhesin (PIA), also known as poly-N-acetyl-β-(1-6)-glucosamine (PIA/PNAG) is an important component of Staphylococcus aureus biofilms and also contributes to resistance to phagocytosis. The proteins IcaA, IcaD, IcaB, and IcaC are encoded within the intercellular adhesin (ica) operon and synthesize PIA/PNAG. We discovered a mechanism of phase variation in PIA/PNAG expression that appears to involve slipped-strand mispairing. The process is reversible and RecA-independent, and involves the expansion and contraction of a simple tetranucleotide tandem repeat within icaC. Inactivation of IcaC results in a PIA/PNAG-negative phenotype. A PIA/PNAG-hyperproducing strain gained a fitness advantage in vitro following the icaC mutation and loss of PIA/PNAG production. The mutation was also detected in two clinical isolates, suggesting that under certain conditions, loss of PIA/PNAG production may be advantageous during infection. There was also a survival advantage for an icaC-negative strain harboring intact icaADB genes relative to an isogenic icaADBC deletion mutant. Together, these results suggest that inactivation of icaC is a mode of phase variation for PIA/PNAG expression, that high-level production of PIA/PNAG carries a fitness cost, and that icaADB may contribute to bacterial fitness, by an unknown mechanism, in the absence of an intact icaC gene and PIA/PNAG production.

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Fitness of MN8m versus JB12.The competitive fitness of MN8m in co-culture with PNAG-negative, non-mucoid JB12 was determined by combining the two strains and enumerating each strain over time. Bars represent the value of each phenotype as percent of total population, determined by enumerating colonies on CRA. Values represent the mean of triplicate samples.
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ppat-1004292-g006: Fitness of MN8m versus JB12.The competitive fitness of MN8m in co-culture with PNAG-negative, non-mucoid JB12 was determined by combining the two strains and enumerating each strain over time. Bars represent the value of each phenotype as percent of total population, determined by enumerating colonies on CRA. Values represent the mean of triplicate samples.

Mentions: The frequency of reversion from non-mucoid (JB12) to mucoid was very low. We hypothesized that the higher frequency with which nonmucoid variants were isolated from MN8m cultures was due to a fitness cost imparted by high-level PIA/PNAG production. To determine if there was a fitness cost associated with constitutive PIA/PNAG synthesis, we inoculated competitive co-cultures with equivalent numbers of MN8m and JB12 and examined shifts in the population over time by assessing colony morphology on CRA. We observed that there does indeed appear to be significant growth advantage in the PIA/PNAG-negative variant JB12, and that by 12 hours, more than 95% of the culture was non-mucoid (Fig. 6). Direct calculation of the fitness cost of PIA/PNAG over-production versus PIA/PNAG loss resulted in fitt (relative bacterial fitness) values of +1.401 at 6 hours, and +1.386 at 12 hours, with a value greater than 1 indicating a significant fitness advantage of the JB12 PIA/PNAG-negative phenotype over the mucoid MN8m.


Phase variation of poly-N-acetylglucosamine expression in Staphylococcus aureus.

Brooks JL, Jefferson KK - PLoS Pathog. (2014)

Fitness of MN8m versus JB12.The competitive fitness of MN8m in co-culture with PNAG-negative, non-mucoid JB12 was determined by combining the two strains and enumerating each strain over time. Bars represent the value of each phenotype as percent of total population, determined by enumerating colonies on CRA. Values represent the mean of triplicate samples.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4117637&req=5

ppat-1004292-g006: Fitness of MN8m versus JB12.The competitive fitness of MN8m in co-culture with PNAG-negative, non-mucoid JB12 was determined by combining the two strains and enumerating each strain over time. Bars represent the value of each phenotype as percent of total population, determined by enumerating colonies on CRA. Values represent the mean of triplicate samples.
Mentions: The frequency of reversion from non-mucoid (JB12) to mucoid was very low. We hypothesized that the higher frequency with which nonmucoid variants were isolated from MN8m cultures was due to a fitness cost imparted by high-level PIA/PNAG production. To determine if there was a fitness cost associated with constitutive PIA/PNAG synthesis, we inoculated competitive co-cultures with equivalent numbers of MN8m and JB12 and examined shifts in the population over time by assessing colony morphology on CRA. We observed that there does indeed appear to be significant growth advantage in the PIA/PNAG-negative variant JB12, and that by 12 hours, more than 95% of the culture was non-mucoid (Fig. 6). Direct calculation of the fitness cost of PIA/PNAG over-production versus PIA/PNAG loss resulted in fitt (relative bacterial fitness) values of +1.401 at 6 hours, and +1.386 at 12 hours, with a value greater than 1 indicating a significant fitness advantage of the JB12 PIA/PNAG-negative phenotype over the mucoid MN8m.

Bottom Line: Inactivation of IcaC results in a PIA/PNAG-negative phenotype.There was also a survival advantage for an icaC-negative strain harboring intact icaADB genes relative to an isogenic icaADBC deletion mutant.Together, these results suggest that inactivation of icaC is a mode of phase variation for PIA/PNAG expression, that high-level production of PIA/PNAG carries a fitness cost, and that icaADB may contribute to bacterial fitness, by an unknown mechanism, in the absence of an intact icaC gene and PIA/PNAG production.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Virginia Commonwealth University School of Medicine, Richmond, Virginia, United States of America.

ABSTRACT
Polysaccharide intercellular adhesin (PIA), also known as poly-N-acetyl-β-(1-6)-glucosamine (PIA/PNAG) is an important component of Staphylococcus aureus biofilms and also contributes to resistance to phagocytosis. The proteins IcaA, IcaD, IcaB, and IcaC are encoded within the intercellular adhesin (ica) operon and synthesize PIA/PNAG. We discovered a mechanism of phase variation in PIA/PNAG expression that appears to involve slipped-strand mispairing. The process is reversible and RecA-independent, and involves the expansion and contraction of a simple tetranucleotide tandem repeat within icaC. Inactivation of IcaC results in a PIA/PNAG-negative phenotype. A PIA/PNAG-hyperproducing strain gained a fitness advantage in vitro following the icaC mutation and loss of PIA/PNAG production. The mutation was also detected in two clinical isolates, suggesting that under certain conditions, loss of PIA/PNAG production may be advantageous during infection. There was also a survival advantage for an icaC-negative strain harboring intact icaADB genes relative to an isogenic icaADBC deletion mutant. Together, these results suggest that inactivation of icaC is a mode of phase variation for PIA/PNAG expression, that high-level production of PIA/PNAG carries a fitness cost, and that icaADB may contribute to bacterial fitness, by an unknown mechanism, in the absence of an intact icaC gene and PIA/PNAG production.

Show MeSH
Related in: MedlinePlus