title: Comparison of multiplex real-time PCR and PCR-reverse blot hybridization assay for the direct and rapid detection of bacteria and antibiotic resistance determinants in positive culture bottles.
abstract: The aim of this study was to evaluate the performance of a commercially available multiplex real-time PCR assay and a PCR-reverse blot hybridization assay (PCR-REBA) for the rapid detection of bacteria and identification of antibiotic resistance genes directly from blood culture bottles and to compare the results of these molecular assays with conventional culture methods. The molecular diagnostic methods were used to evaluate 593 blood culture bottles from patients with bloodstream infections. The detection positivity of multiplex real-time PCR assay for Gram-positive bacteria, Gram-negative bacteria and Candida spp. was equivalent to PCR-REBA as 99.6 %, 99.1 % and 100 %, respectively. Using conventional bacterial cultures as the gold standard, the sensitivity, specificity, positive predictive value and negative predictive value of these two molecular methods were 99.5 % [95 % confidence interval (CI), 0.980-1.000; P<0.001), 100 % (95 % CI, 0.983-1.000; P<0.001), 100 % and 99 %, respectively. However, positivity of the Real-methicillin-resistant Staphylococcusaureus multiplex real-time PCR assay targeting the mecA gene to detect methicillin resistance was lower than that of the PCR-REBA method, detecting an overall positivity of 98.4 % (n=182; 95 % CI, 0.964-1.000; P<0.009) and 99.5 % (n=184; 95 % CI, 0.985-1.000; P<0.0001), respectively. The entire two methods take about 3 h, while results from culture can take up to 48-72 h. Therefore, the use of these two molecular methods was rapid and reliable for the characterization of causative pathogens in bloodstream infections.
authors.affiliation: Optipharm M&D, Inc., Wonju Eco Environmental Technology Center, Wonju, Gangwon, 26493, South Korea.
authors.forename: Hye-Young
authors.initials: HY
authors.lastname: Wang
authors.affiliation: Department of Biomedical Laboratory Science, College of Health Sciences, Yonsei University, Wonju, Gangwon, 26493, South Korea.
authors.forename: Seoyong
authors.initials: S
authors.lastname: Kim
authors.affiliation: Department of Biomedical Laboratory Science, College of Health Sciences, Yonsei University, Wonju, Gangwon, 26493, South Korea.
authors.forename: Jungho
authors.initials: J
authors.lastname: Kim
authors.affiliation: Department of Laboratory Medicine, Yonsei University Wonju College of Medicine, Wonju, Gangwon, 26426, South Korea.
authors.forename: Soon Deok
authors.initials: SD
authors.lastname: Park
authors.affiliation: Department of Internal Medicine, Yonsei University Wonju College of Medicine, Wonju, Gangwon, 26426, South Korea.
authors.forename: Hyo Youl
authors.initials: HY
authors.lastname: Kim
authors.affiliation: Department of Laboratory Medicine, Yonsei University Wonju College of Medicine, Wonju, Gangwon, 26426, South Korea.
authors.forename: Young
authors.initials: Y
authors.lastname: Uh
authors.affiliation: Department of Biomedical Laboratory Science, College of Health Sciences, Yonsei University, Wonju, Gangwon, 26493, South Korea.
authors.forename: Hyeyoung
authors.initials: H
authors.lastname: Lee
mesh_terms: D016470:Bacteremia / Q000175:diagnosis* / Q000382:microbiology; D001419:Bacteria / Q000145:classification / Q000187:drug effects / Q000302:isolation & purification*; D001769:Blood / Q000382:microbiology; D002175:Candida / Q000302:isolation & purification; D058387:Candidemia / Q000175:diagnosis / Q000382:microbiology; D024881:Drug Resistance, Bacterial; D006801:Humans; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D009693:Nucleic Acid Hybridization / Q000379:methods*; D011237:Predictive Value of Tests; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D013997:Time Factors