Biofilm formation and morphology of group A streptococci isolates of patients with sepsis or necrotising fasciitis

Abstract number: 1134_03_427

Presterl E., Eder M., Reichmann S., Gattringer R., Hirschl A.M.

Objectives:  

Group A streptococci may cause severe sepsis associated with multiorgan failure and high mortality. Additionally, they are the primary cause of necrotizing fasciitis. The objective of this study was to evaluate the ability to form biofilms and the morphology of these biofilms produced by GAS isolated from blood of patients with sepsis and from the tissue of patients with necrotizing fasciitis.

Methods:  

Twenty-six GAS isolates from blood cultures of patients with sepsis and bacteraemia, and 26 isolates from tissue of patients with necrotizing cellulitis and fasciitis were collected. They were isolated and identified using routine methods, and frozen at –73°C in BHI broth with 3% glycerol. To test their ability to form biofilms, the isolates were grown in microtiter plates in tryptone soya broth for 24 or 48 hours. To examine the morphology of the biofilm formation the isolates were either examined unfixed or fixed with 2% glutaraldehyd using electrone microscope scanning (Philips XL30 ESEM). For quantification they were fixed with 2% glutaraldehyd and dyed with 1% crystal violet to measure the mean optical density (OD using a routine microtiter-plate-reader at 550 nm wavelength. To calculate differences in the frequency in the groups the Fisher's exact test and to calculated differences in the OD the Mann–WhitneyU-Test were used.

Results:  

Seventeen out of 26 isolates from patients with sepsis and 14 out of 26 isolates fom patients with necrotizing fasciitis formed biofilms (difference not significant). Scanning electron studies confirmed biofilm formation however it looked somewhat scarce in isolates from necrotizing fasciitis. For the biofilm forming isolates, the OD of isolates from sepsis were significantly higher than the OD of isolates from necrotizing fasciitis (0.996 versus 0.578, p < 0.001).

Conclusions:  

GAS isolates from both, bloodstream and tissue, had ability to form biofilms. Biofilms of blood stream isolates were significantly denser than biofilms of the tissue isolates. This surplus opacity of blood stream isolates was confirmed by the scanning electron microscopy studies which revealed a more scanty looking morphology of the tissue GAS isolates. The lesser density of the biofilms of GAS isolated from the tissue might be important for the pathogenesis of necrotizing fascitiis probably facilitating to the rapid spread of the bacteria within the tissue.