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 thecommunity. This bacterium has the ability to cause chronic infections due to the formation ofsurface-associated aggregates or biofilms. The SigB alternative sigma factor is activated during biofilmformation 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 thestaphyloxanthin (STX) biosynthetic pathway. The carotenoid pigment biosynthesis genes areorganized in the operon crtOPQMN. STX is a virulence factor with antioxidant action and its productionis 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 biomassformed by S. aureus isolated from patients with osteomyelitis. We selected 18 S. aureus isolates fromour laboratory collection of strains whose full genome was previously sequenced. The genotypiccharacterization (clonal complex, sequence typing, agr typing and spa typing) of each strain wasperformed by specific PCRs and sequence analysis. The pigment extracted was quantified byevaluation of the optical density (OD) at 450 nm and by correlation pigment production with the cultureOD. Biofilms formed by S. aureus were measured by static microtiter plate assays. The distribution ofthe average OD values from the 18 isolates into quartiles permitted to classify the strains as highproducers (HP: OD ≥ 0.62), producers (P: OD ≥ 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, otherfour 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 wereselected for bioinformatical analysis. The SigB factor sequences were 100% identical among this pairof isolates whereas a single aminoacidic change at the 108 position of the crtM gene was observed inthe STX negative producer strain under comparison.Taken together, these results indicate that STX production (orange pigment) correlates with increasedbiofilm formation in S. aureus isolates from patients with chronic osteomyelitis. However, a lowpercentage of the P and HP biofilm isolates were unable to produce orange or yellow pigmentssuggesting the presence of a genetic defect in the crt operon.