Variation in staphyloxanthin production and biofilm formation by Staphylococcus aureus from chronic infections.

AGUSTIN RIVIERE; CRISTIAN DOTTO; MAURICIO SULIGOY; ANDREA LOMBARTE SERRAT; DANIEL SORDELLI; MÓNICA GIACOMODONATO; FERNANDA BUZZOLA

Cordoba

Congreso; XI Congreso Argentino de Microbiología General - SAMIGE 2015; 2015

SAMIGE Sociedad Argentina de Microbiología General

Staphylococcus aureus is one of the most important human pathogens both in the hospital and the community. This bacterium has the ability to cause chronic infections due to the formation of surface-associated aggregates or biofilms. The SigB alternative sigma factor is activated during biofilm formation and triggers the expression of genes associated with pigmentation and biofilm formation. The orange and yellow carotenoid pigments that produce S. aureus are the products of the staphyloxanthin (STX) biosynthetic pathway. The carotenoid pigment biosynthesis genes are organized in the operon crtOPQMN. STX is a virulence factor with antioxidant action and its production is positively regulated by the SigB factor. To further study the contribution of STX in the fitness of S. aureus in chronic infections, we compared the level of pigments produced with the biofilm biomass formed by S. aureus isolated from patients with osteomyelitis. We selected 18 S. aureus isolates from our laboratory collection of strains whose full genome was previously sequenced. The genotypic characterization (clonal complex, sequence typing, agr typing and spa typing) of each strain was performed by specific PCRs and sequence analysis. The pigment extracted was quantified by evaluation of the optical density (OD) at 450 nm and by correlation pigment production with the culture OD. Biofilms formed by S. aureus were measured by static microtiter plate assays. The distribution of the average OD values from the 18 isolates into quartiles permitted to classify the strains as high producers (HP: OD ³ 0.62), producers (P: OD ³ 0.4 and < 0.62), or non-producers (NP: OD < 0.4). The analysis of the levels of pigment extracted revealed that four strains were STX negative, other four isolates produced orange (A450: 0.44 -1.03) pigment and the rest of the strains showed yellow (A450: 0.28 - 0.43) pigment. The NP biofilm isolates expressed only yellow levels of STX. However, almost 30% of the P and HP biofilm strains were STX negative. A couple of genotypically identical (CC5, ST5, t002, agr type II) isolates that showed differences in pigment and biofilm production were selected for bioinformatical analysis. The SigB factor sequences were 100% identical among this pair of isolates whereas a single aminoacidic change at the 108 position of the crtM gene was observed in the STX negative producer strain under comparison. Taken together, these results indicate that STX production (orange pigment) correlates with increased biofilm formation in S. aureus isolates from patients with chronic osteomyelitis. However, a low percentage of the P and HP biofilm isolates were unable to produce orange or yellow pigments suggesting the presence of a genetic defect in the crt operon.