Evaluation of rapid genotype assay for the identification of gram-positive cocci from blood cultures and detection of mecA and van genes]

Rapid and accurate identification of bacterial pathogens grown in blood cultures of patients with sepsis is crucial for prompt initiation of appropriate therapy in order to decrease related morbidity and mortality rates. Although current automated blood culture systems led to a significant improvement in bacterial detection time, more rapid identification systems are still needed to optimise the establishment of treatment. Novel genotype technology which is developed for the rapid diagnosis of sepsis, is a molecular genetic assay based on DNA multiplex amplification with biotinylated primers followed by hybridization to membrane bound probes. The aim of this study was to evaluate the performance of "Genotype (R) BC gram-positive" test for the identification of gram-positive cocci grown in blood cultures and rapid detection of mecA and van genes. This test uses DNA.STRIP (R) technology which includes a panel of probes for identification of 17 gram-positive bacterial species and is able to determinate the methicillin and vancomycin resistance mediating genes (mecA and vanA, vanB, vanC1, vanC2/C3) simultaneously, in a single test run. A total of 55 positive blood cultures from BACTEC (TM) Plus/F (Becton Dickinson, USA) aerobic and pediatric blood culture vials were included in the study. The isolates which exhibit gram-positive coccus morphology by Gram staining were identified by Genotype (R) BC gram-positive test (Hain Life Science, Germany). All of the samples were also identified with the use of Phoenix PMIC/ID Panel (Becton Dickinson, USA) and antibiotic susceptibilities were determined. Of the 55 blood culture isolates, 17 were identified as Staphylococcus epidermidis [all were methicillin-resistant (MR)], 9 were S.aureus (one was MR), 18 were S.hominis (10 were MR), 4 were E.faecalis, 3 were E. faecium (one was vanconycin-resistant), 2 were S.saprophyticus (one was MR), 1 was S.warneri and 1 was S.haemolyticus, by Phoenix automated system. "Genotype (R) BC gram-positive" test results revealed consistency with Phoenix system regarding bacterial identification in 46 (83.6%) of the samples. The two bacteria identified as S.saprophyticus by the Phoenix system could not be identified by the Genotype (R) BC test since this species were not included in the identification panel of the system, however, mecA gene were detected in these two samples by Genotype (R) BC test. Genotype (R) BC test detected mecA gene in five samples which were not detected as methicillin resistant by the Phoenix system. Besides polymicrobial growth was determined in five samples by Genotype (R) BC test, but not by the automated system. One E.faecium isolate with vanA gene was correctly identified by Genotype (R) BC test. In conclusion, Genotype (R) BC gram-positive test is a fast and reliable test for the identification of the most important gram-positive pathogens and mecA and van genes directly from positive blood culture bottles. This test was also found superior than the automated Phoenix system regarding the detection of polymicrobial growth. These data indicated that, routine use of DNA strip technology-based assay would be useful for clinical diagnosis in patients with sepsis.