Clinical evaluation of NucliSENS magnetic extraction and NucliSENS analyte-specific reagents for real-time detection of human metapneumovirus in pediatric respiratory specimens.
In this study, we evaluated the NucliSENS miniMAG (MM) and easyMAG (EM) nucleic acid extraction platforms (bioMérieux, Durham, NC) in combination with the NucliSENS EasyQ basic kit and analyte-specific reagents (ASRs) (bioMérieux) for the detection of human metapneumovirus (hMPV) in respiratory samples. Total nucleic acids from pediatric clinical samples (n = 653) and an hMPV-specific inhibition control (h-IC) were coextracted using the MM and/or the EM. Nucleic acid sequence-based amplification and real-time molecular beacon detection of hMPV were performed using a NucliSENS EasyQ analyzer (bioMérieux). Positive results were confirmed using an in-house-validated reverse transcriptase PCR ASR-based assay. The inclusion of the h-IC monitored the entire process, including the efficiency of nucleic acid extraction, amplification, and detection. The percentages of samples with inhibited amplification of the h-IC after initial NA extraction by EM and MM were 1.88% and 3.17%, respectively. After reprocessing of a new aliquot, the final h-IC inhibition rates were 0% (EM) and 1.06% (MM). The limit of detection of the assay was between 2 (EM extraction) and 10 (MM extraction) RNA copies/reaction, and specificity was 100% when testing viral respiratory isolates and clinical samples. hMPV was detected in 5.6% of pediatric samples tested and was also detected in three coinfections with respiratory syncytial virus (RSV). hMPV was the second most frequently detected respiratory virus in children of 0 to 2 years of age, after RSV. In summary, NucliSENS extraction and ASRs provided a sensitive and specific method for the detection of hMPV in respiratory samples.
Ginocchio
Christine C
CC
North Shore-Long Island Jewish Health System Laboratories, Molecular Diagnostics, Department of Laboratory Medicine, 10 Nevada Drive, Lake Success, NY 11042, USA. cginocch@nshs.edu
Manji
Ryhana
R
Lotlikar
Madhavi
M
Zhang
Fan
F
D002675:Child, Preschool; D006801:Humans; D007223:Infant; D007231:Infant, Newborn; D029121:Metapneumovirus / Q000235:genetics / Q000302:isolation & purification*; D015335:Molecular Probes; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D018184:Paramyxoviridae Infections / Q000821:virology; D012367:RNA, Viral / Q000032:analysis / Q000302:isolation & purification; D011933:Reagent Kits, Diagnostic; D012137:Respiratory System / Q000821:virology*; D012680:Sensitivity and Specificity
D015335:Molecular Probes; D012367:RNA, Viral; D011933:Reagent Kits, Diagnostic