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RecA mediated spontaneous deletions of the icaADBC operon of clinical Staphylococcus epidermidis isolates: a new mechanism of phenotypic variations.

ABSTRACT: Phenotypic variation of Staphylococcus epidermidis involving the slime related ica operon results in heterogeneity in surface characteristics of individual bacteria in axenic cultures. Five clinical S. epidermidis isolates demonstrated phenotypic variation, i.e. both black and red colonies on Congo Red agar. Black colonies displayed bi-modal electrophoretic mobility distributions at pH 2, but such phenotypic variation was absent in red colonies of the same strain as well as in control strains without phenotypic variation. All red colonies had lost ica and the ability to form biofilms, in contrast to black colonies of the same strain. Real time PCR targeting icaA indicated a reduction in gene copy number within cultures exhibiting phenotypic variation, which correlated with phenotypic variations in biofilm formation and electrophoretic mobility distribution of cells within a culture. Loss of ica was irreversible and independent of the mobile element IS256. Instead, in high frequency switching strains, spontaneous mutations in lexA were found which resulted in deregulation of recA expression, as shown by real time PCR. RecA is involved in genetic deletions and rearrangements and we postulate a model representing a new mechanism of phenotypic variation in clinical isolates of S. epidermidis. This is the first report of S. epidermidis strains irreversibly switching from biofilm-positive to biofilm-negative phenotype by spontaneous deletion of icaADBC.

SUBMITTER: Nuryastuti T

PROVIDER: S-EPMC2480603 | biostudies-literature | 2008 Aug

REPOSITORIES: biostudies-literature

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recA mediated spontaneous deletions of the icaADBC operon of clinical Staphylococcus epidermidis isolates: a new mechanism of phenotypic variations.

Nuryastuti Titik T van der Mei Henny C HC Busscher Henk J HJ Kuijer Roel R Aman Abu T AT Krom Bastiaan P BP

Antonie van Leeuwenhoek 20080504 2

Phenotypic variation of Staphylococcus epidermidis involving the slime related ica operon results in heterogeneity in surface characteristics of individual bacteria in axenic cultures. Five clinical S. epidermidis isolates demonstrated phenotypic variation, i.e. both black and red colonies on Congo Red agar. Black colonies displayed bi-modal electrophoretic mobility distributions at pH 2, but such phenotypic variation was absent in red colonies of the same strain as well as in control strains wi ...[more]

PMID: 18454346

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Project description:S. epidermidis is a primary cause of biofilm-mediated infections in humans due to adherence to foreign bodies. A major staphylococcal biofilm accumulation molecule is polysaccharide intracellular adhesin (PIA), which is synthesized by enzymes encoded by the icaADBC operon. Expression of PIA is highly variable among clinical isolates, suggesting that PIA expression levels are selected in certain niches of the host. However, the mechanisms that govern enhanced icaADBC transcription and PIA synthesis in these isolates are not known. We hypothesized that enhanced PIA synthesis in these isolates was due to function of IcaR and/or TcaR. Thus, two S. epidermidis isolates (1457 and CSF41498) with different icaADBC transcription and PIA expression levels were studied. Constitutive expression of both icaR and tcaR demonstrated that both repressors are functional and can completely repress icaADBC transcription in both 1457 and CSF41498. However, it was found that IcaR was the primary repressor for CSF41498 and TcaR was the primary repressor for 1457. Further analysis demonstrated that icaR transcription was repressed in 1457 in comparison to CSF41498, suggesting that TcaR functions as a repressor only in the absence of IcaR. Indeed, DNase I footprinting suggests IcaR and TcaR may bind to the same site within the icaR-icaA intergenic region. Lastly, we found mutants expressing variable amounts of PIA could rapidly be selected from both 1457 and CSF41498. Collectively, we propose that strains producing enhanced PIA synthesis are selected within certain niches of the host through several genetic mechanisms that function to repress icaR transcription, thus increasing PIA synthesis.IMPORTANCEStaphylococcus epidermidis is a commensal bacterium that resides on our skin. As a commensal, it protects humans from bacterial pathogens through a variety of mechanisms. However, it is also a significant cause of biofilm infections due to its ability to bind to plastic. Polysaccharide intercellular adhesin is a significant component of biofilm, and we propose that the expression of this polysaccharide is beneficial in certain host niches, such as providing extra strength when the bacterium is colonizing the lumen of a catheter, and detrimental in others, such as colonization of the skin surface. We show here that fine-tuning of icaADBC transcription, and thus PIA synthesis, is mediated via two transcriptional repressors, IcaR and TcaR.

Project description:Staphylococcus epidermidis is a common commensal of healthy conjunctiva and it can cause endophthalmitis, however its presence in conjunctivitis, keratitis and blepharitis is unknown. Molecular genotyping of S. epidermidis from healthy conjunctiva could provide information about the origin of the strains that infect the eye. In this paper two collections of S. epidermidis were used: one from ocular infection (n = 62), and another from healthy conjunctiva (n = 45). All isolates were genotyped by pulsed field gel electrophoresis (PFGE), multilocus sequence typing (MLST), staphylococcal cassette chromosome mec (SCCmec), detection of the genes icaA, icaD, IS256 and polymorphism type of agr locus. The phenotypic data included biofilm production and antibiotic resistance. The results displayed 61 PFGE types from 107 isolates and they were highly discriminatory. MLST analysis generated a total of 25 STs, of which 11 STs were distributed among the ocular infection isolates and lineage ST2 was the most frequent (48.4%), while 14 STs were present in the healthy conjunctiva isolates and lineage ST5 was the most abundant (24.4%). By means of a principal coordinates analysis (PCoA) and a discriminant analysis (DA) it was found that ocular infection isolates had as discriminant markers agr III or agr II, SCCmec V or SCCmec I, mecA gene, resistance to tobramycin, positive biofilm, and IS256+. In contrast to the healthy conjunctiva isolates, the discriminating markers were agr I, and resistance to chloramphenicol, ciprofloxacin, gatifloxacin and oxacillin. The discriminant biomarkers of ocular infection were examined in healthy conjunctiva isolates, and it was found that 3 healthy conjunctiva isolates [two with ST2 and another with ST9] (3/45, 6.66%) had similar genotypic and phenotypic characteristics to ocular infection isolates, therefore a small population from healthy conjunctiva could cause an ocular infection. These data suggest that the healthy conjunctiva isolates do not, in almost all cases, infect the eye due to their large genotypic and phenotypic difference with the ocular infection isolates.

Dataset Information

RecA mediated spontaneous deletions of the icaADBC operon of clinical Staphylococcus epidermidis isolates: a new mechanism of phenotypic variations.

Publications

recA mediated spontaneous deletions of the icaADBC operon of clinical Staphylococcus epidermidis isolates: a new mechanism of phenotypic variations.

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