title: Diagnostic significance of nested polymerase chain reaction for sensitive detection of Pneumocystis jirovecii in respiratory clinical specimens.
abstract: A total of 327 clinical specimens, including both invasive and noninvasive samples, obtained from 275 patients with various types of underlying immunocompromised conditions and a clinical suspicion of Pneumocystis pneumonia (PCP) were subjected to 2 different nested polymerase chain reaction (PCR) assays. The target genes used for nested PCR were mitochondrial large subunit ribosomal RNA (mtLSU rRNA) and internal transcribed spacer (ITS) region. The results were compared with a single-round PCR targeting major surface glycoprotein (MSG) gene. Amplification was successful in 16% of cases by mtLSU rRNA nested PCR, in 14.5% by ITS nested PCR, and in 10.9% by MSG PCR. The nested mtLSU rRNA PCR was found to be more sensitive (100% sensitive and 98.7% specific) and useful in detecting PCP for its use in routine diagnosis in our settings. Thus, this assay may be quite useful in the identification of patients who are in the early stage of Pneumocystis jirovecii infection with an organism load that could not be easily detected by the single-step PCR.
authors.affiliation: Department of Microbiology, All Indian Institute of Medical Sciences, Ansari Nagar, New Delhi 110029, India.
authors.forename: Rashmi
authors.initials: R
authors.lastname: Gupta
authors.forename: Bijay Ranjan
authors.initials: BR
authors.lastname: Mirdha
authors.forename: Randeep
authors.initials: R
authors.lastname: Guleria
authors.forename: Lalit
authors.initials: L
authors.lastname: Kumar
authors.forename: Jyotish Chandra
authors.initials: JC
authors.lastname: Samantaray
authors.forename: Sanjay Kumar
authors.initials: SK
authors.lastname: Agarwal
authors.forename: Sushil Kumar
authors.initials: SK
authors.lastname: Kabra
authors.forename: Kalpana
authors.initials: K
authors.lastname: Luthra
mesh_terms: D017931:DNA Primers / Q000235:genetics; D004271:DNA, Fungal / Q000235:genetics; D021903:DNA, Ribosomal Spacer / Q000235:genetics; D006801:Humans; D008969:Molecular Sequence Data; D016720:Pneumocystis Infections / Q000175:diagnosis* / Q000382:microbiology; D045363:Pneumocystis carinii / Q000235:genetics / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012313:RNA / Q000235:genetics; D000077278:RNA, Mitochondrial; D012335:RNA, Ribosomal / Q000235:genetics; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA; D013183:Sputum / Q000382:microbiology*
chemical_list: D017931:DNA Primers; D004271:DNA, Fungal; D021903:DNA, Ribosomal Spacer; D000077278:RNA, Mitochondrial; D012335:RNA, Ribosomal; D012313:RNA