Identification and detection of mecA and van genes of gram positive cocci by rapid assay among positive blood cultures

Detection of bacterial isolates in blood cultures of septicemic patients plays an important role in diagnostic microbiology, due to high rates of patient morbidity and mortality. The positive impact of rapid reporting of bacterial pathogens on patient outcomes and hospital costs have been previously statedCurrent automated continuous monitoring blood culture systems marked a significant advance in reducing bacteria detection time. However, rapid identification systems are also required to optimise treatment orientationThe Genotype technology, a rapid molecular genetic assay based on DNA multiplex amplification with biotinylated primers followed by hybridization to membrane bound probes, complies with the requirements for a fast diagnosis of sepsis. We evaluated the new Genotype®BC Gram-positive test kits (Hain Life Science, Germany). The test includes a panel of probes for identification of 17 Gram-positive (GP) bacteria species and simultaneous determination of methicillin and vancomycin resistance (mecA and van genes). The vancomycin resistance mediating genes vanA, vanB, vanC1, and vanC2/C3 as well as the methicillin resistance mediating mecA gene can be detected in the same stepThe study was performed on 55 positive blood cultures from BACTEC blood culture vials (aerobic and pediatric vials). First, a Gram stain was carried out to select Genotype®BC GP, then identification were performed by the Genotype®BC GP test, and by PHOENIX PMIC/ID Panel (Becton Dickinson Diagnostic Instrument Systems, Spark, Md, USA ) after subculture and phenotypic susceptibility determination. The operating procedure was very easy to carry out and required a small amount of starting material (15 ?l of blood culture). The results were available 4.5 hours after the results of microscopic analysis. Results are generally founds correct as compared with PHOENIX automatize system. Two izolates were CoNS (Staphylococcus saprophyticus) not identifiable by the lack of specific probes on the strip. Also succesfully identified of polimicrobial infections by this test in 5 blood culture specimens that not determined automatize system. Detection of polimicrobial infections were excellent by this test. One Enterococcus faecium isolate with a positive vanA gene isolated was correctly differentiated by the assayThis study provides the first evaluation of genotypeBC tests directly from BACTEC blood cultures vials. In conclusion the evaluated GenoType blood culture assay proved fast and reliable results in detecting the most important Gram positive sepsis pathogens and the mecA and van genes directly from positive blood culture bottles. According to our results, this DNA strip technology based assay can easily be incorporated into routine diagnosis.