title: Development of a multiplex one step RT-PCR that detects eighteen respiratory viruses in clinical specimens and comparison with real time RT-PCR.
abstract: Rapid and accurate diagnosis of viral respiratory infections is crucial for patient management. Multiplex reverse transcriptase polymerase chain reaction (mRT-PCR) is used increasingly to diagnose respiratory infections and has shown to be more sensitive than viral culture and antigen detection. Objective of the present study was to develop a one-step mRT-PCR that could detect 18 respiratory viruses in three sets. The method was compared with real time RT-PCR (rRT-PCR) for its sensitivity and specificity. Clinical specimens from 843 pediatric patients with respiratory symptoms were used in the study. 503 (59.7%) samples were detected positive by mRT-PCR. Of these 462 (54.8%) exhibited presence of a single pathogen and 41 (4.9%) had multiple pathogens. rRT-PCR detected 439 (52.1%) positive samples, where 419 (49.7%) exhibited one virus and 20 (2.4%) showed co-infections. Concordance between mRT-PCR and rRT-PCR was 91.9% and kappa correlation 0.837. Sensitivity and specificity of mRT-PCR were 99.5% and 83.7% while that of rRT-PCR was 86.9% and 99.4% respectively. Rhinovirus (17.2%) was the most frequently detected virus followed by respiratory syncytial virus B (15.4%), H1N1pdm09 (8.54%), parainfluenza virus-3 (5.8%) and metapneumovirus (5.2%). In conclusion, mRT-PCR is a rapid, cost effective, specific and highly sensitive method for detection of respiratory viruses.
authors: Influenza Division, National Institute of Virology, Pune, India. mlchoudhary@gmail.com
authors: Manohar L
authors: ML
authors: Choudhary
authors: Siddharth P
authors: SP
authors: Anand
authors: Mostafa
authors: M
authors: Heydari
authors: Grishma
authors: G
authors: Rane
authors: Varsha A
authors: VA
authors: Potdar
authors: Mandeep S
authors: MS
authors: Chadha
authors: Akhilesh C
authors: AC
authors: Mishra
mesh_terms: D002648:Child; D002675:Child, Preschool; D006801:Humans; D053118:Influenza A Virus, H1N1 Subtype / Q000235:genetics / Q000302:isolation & purification; D007251:Influenza, Human / Q000175:diagnosis; D029121:Metapneumovirus / Q000235:genetics / Q000302:isolation & purification; D010224:Parainfluenza Virus 3, Human / Q000235:genetics / Q000302:isolation & purification; D010850:Picornaviridae Infections / Q000175:diagnosis; D012367:RNA, Viral / Q000032:analysis*; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D018357:Respiratory Syncytial Virus Infections / Q000175:diagnosis; D012141:Respiratory Tract Infections / Q000175:diagnosis* / Q000821:virology*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012229:Rhinovirus / Q000302:isolation & purification; D012680:Sensitivity and Specificity
chemical_list: D012367:RNA, Viral