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Investigating Extracellular DNA Release in Staphylococcus xylosus Biofilm In Vitro.

ABSTRACT: Staphylococcus xylosus forms biofilm embedded in an extracellular polymeric matrix. As extracellular DNA (eDNA) resulting from cell lysis has been found in several staphylococcal biofilms, we investigated S. xylosus biofilm in vitro by a microscopic approach and identified the mechanisms involved in cell lysis by a transcriptomic approach. Confocal laser scanning microscopy (CLSM) analyses of the biofilms, together with DNA staining and DNase treatment, revealed that eDNA constituted an important component of the matrix. This eDNA resulted from cell lysis by two mechanisms, overexpression of phage-related genes and of cidABC encoding a holin protein that is an effector of murein hydrolase activity. This lysis might furnish nutrients for the remaining cells as highlighted by genes overexpressed in nucleotide salvage, in amino sugar catabolism and in inorganic ion transports. Several genes involved in DNA/RNA repair and genes encoding proteases and chaperones involved in protein turnover were up-regulated. Furthermore, S. xylosus perceived osmotic and oxidative stresses and responded by up-regulating genes involved in osmoprotectant synthesis and in detoxification. This study provides new insight into the physiology of S. xylosus in biofilm.

SUBMITTER: Leroy S

PROVIDER: S-EPMC8617998 | biostudies-literature | 2021 Oct

REPOSITORIES: biostudies-literature

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Investigating Extracellular DNA Release in Staphylococcus xylosus Biofilm In Vitro.

Leroy Sabine S Lebert Isabelle I Andant Carine C Micheau Pierre P Talon Régine R

Microorganisms 20211021 11

Staphylococcus xylosus forms biofilm embedded in an extracellular polymeric matrix. As extracellular DNA (eDNA) resulting from cell lysis has been found in several staphylococcal biofilms, we investigated S. xylosus biofilm in vitro by a microscopic approach and identified the mechanisms involved in cell lysis by a transcriptomic approach. Confocal laser scanning microscopy (CLSM) analyses of the biofilms, together with DNA staining and DNase treatment, revealed that eDNA constitut ...[more]

PMID: 34835318

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Project description:Staphylococcus xylosus, a coagulase-negative, non-pathogenic bacterium, responsible for opportunistic infections in humans and bovine mastitis, has the ability to form biofilms, which are responsible for persistent infections and antibiotic resistance. In our study, azithromycin significantly inhibited biofilm formation by altering protein expression. Of the 1764 proteins measured by the isobaric Tag for Relative and Absolute Quantification (iTRAQ) technique, only 148 proteins showed significantly different expression between the azithromycin-treated and untreated cells. Most ribosomal proteins were markedly up-regulated, and the expression of the proteins involved in histidine biosynthesis, which, in turn, influence biofilm formation, was down-regulated, particularly imidazole glycerophosphate dehydratase (IGPD). Previously, we had observed that IGPD plays an important role in biofilm formation by S. xylosus. Therefore, hisB expression was studied by real-time PCR, and the interactions between azithromycin and IGPD were predicted by molecular docking analysis. hisB was found to be significantly down-regulated, and six bond interactions were observed between azithromycin and IGPD. Many active atoms of azithromycin did not interact with the biologically active site of IGPD. Surface plasmon resonance analysis used to further study the relationship between IGPD and azithromycin showed minimum interaction between them. Histidine content in the azithromycin-treated and untreated groups was determined. We noted a slight difference, which was not consistent with the expression of the proteins involved in histidine biosynthesis. Therefore, histidine degradation into glutamate was also studied, and we found that all proteins were down-regulated. This could be the reason why histidine content showed little change between the treated and untreated groups. In summary, we found that azithromycin is a potential inhibitor of S. xylosus biofilm formation, and the underlying mechanism was preliminarily elucidated in this study.

Project description:BackgroundPolymicrobial infections are responsible for significant mortality and morbidity in adults and children. Staphylococcus epidermidis and Candida albicans are the most frequent combination of organisms isolated from polymicrobial infections. Vascular indwelling catheters are sites for mixed species biofilm formation and pose a significant risk for polymicrobial infections. We hypothesized that enhancement of biofilms in a mixed species environment increases patient mortality and morbidity.ResultsMixed species biofilms of S. epidermidis and C. albicans were evaluated in vitro and in a subcutaneous catheter infection model in vivo. Mixed species biofilms were enhanced compared to single species biofilms of either S. epidermidis or C. albicans. A mixed species environment increased catheter infection and increased dissemination of S. epidermidis in mice. Microarrays were used to explore differential gene expression of S. epidermidis in the mixed species biofilms. In mixed species biofilms, compared to single species S. epidermidis biofilms, 2.7% of S. epidermidis genes were upregulated and 6% were down regulated. Staphylococcal autolysis repressors lrgA and lrgB were down regulated 36-fold and 27-fold respectively. The role of biofilm extracellular DNA was investigated by quantitation and by evaluating the effects of DNAse in a concentration and time dependent manner. S. epidermidis specific eDNA was increased in mixed species biofilms and further confirmed by degradation with DNAse.ConclusionsMixed-species biofilms are enhanced and associated with increased S. epidermidis-specific eDNA in vitro and greater systemic dissemination of S. epidermidis in vivo. Down regulation of the lrg operon, a repressor of autolysis, associated with increased eDNA suggests a possible role for bacterial autolysis in mixed species biofilms. Enhancement and systemic dissemination of S. epidermidis may explain adverse outcomes after clinical polymicrobial infections of S. epidermidis and C. albicans.

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Investigating Extracellular DNA Release in Staphylococcus xylosus Biofilm In Vitro.

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Investigating Extracellular DNA Release in <i>Staphylococcus xylosus</i> Biofilm In Vitro.

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