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Objectives

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  1. Combination of a microtiter plate method with the amplification of the icaA/aap genes is aneffective tool to determine biofilm formation in Staphylococcus epidermidis isolates from Latvian patients I. Liduma1,2, U. Bers1,2, A.Gorbatjuka1, A. Zilevica1, T. Tracevska1,21- Faculty of Medicine, University of Latvia, 2- Laboratory of Bioanalyticaland Biodosimetry Methods, Faculty of Biology, University of Latvia Objectives Methods Coagulase-negative staphylococci (CoNS) are major nosocomial pathogens known by their ability to form biofilm on the surfaces of foreign bodies introduced (implanted) into the organism. The accumulative phase of biofilm formation is linked to the production of polysaccharide intercellular adhesin (PIA), which is synthesized by icaADBC-encoded proteins and the accumulation associated protein (AAP) encoded by aap. In this study show the reliability of microtiter plate method combined with the icaA/aap genes to determine biofilm forming potentialin patients from four Latvian hospitals. . The study involved S.epidermidis cultures from clinical and control groups isolated in 2009-2010. 81 cultures were isolated from blood (n=45) and intravenous catheters (n=36) in a case of laboratory confirmed bacteraemia from patients of four hospitals in Riga, Latvia.  Fifty-five S.epidermidis isolates from normal nose epithelium flora were included in the control group of 20-22 years old healthy volunteers. The fragments of aap (399bp) and icaA (502bp) genes were detected by amplification and visualised by gel electrophoresis. Biofilm formation using microtitre plate was carried out as described by Christensen et al. Methicillin resistance was determined by the amplification of the mecA gene 310-bp fragment. Results Results on combination of microtiter plate method with amplification of the ica/aap genes are shown in Figure 1. The statistical analysis by Chi-square test showed that results were statistically significant for BF+/aap+/icaA+ (p=0.03) and BF−/aap−/icaA− (p=0.009). Amplification of mecA gene determined methicillin resistance in 79 (97.5%) of clinical group and in 3 (5.4%) of control group samples . Biofilm formation in 35 (43,2%) of S.epidermidis isolates of clinical group and 12 (21,8%) of control group isolates were detected . . . . Fig. 1. Frequency of biofilm forming clinical and non clinical S.epidermidis strains in the presence or absence of icaA and aap genes. Conclusions Our data indicates that the microtiter plate method only in combination with icaA/aap genes amplification is a realiable tool for predicting invasiveness of S.epidermidis strains. Methicillin resistance in nosocomial S.epidermidis strains is growing in Latvia (97.5% compared to 60-80% in 2004) and remains at low level among healthy people, however, this tendence will be analysed in an additional, large scale study. Project “Capacity building for interdisciplinary biosafety research” No 2009/0224/1DP/1.1.1.2.0/09/ APIA/VIAA/055 coofunded by European Social Fund

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