title: Nested PCR Assay for Eight Pathogens: A Rapid Tool for Diagnosis of Bacterial Meningitis.
abstract: OBJECTIVE
  Bacterial meningitis is a dreadful infectious disease with a high mortality and morbidity if remained undiagnosed. Traditional diagnostic methods for bacterial meningitis pose a challenge in accurate identification of pathogen, making prognosis difficult. The present study is therefore aimed to design and evaluate a specific and sensitive nested 16S rDNA genus-based polymerase chain reaction (PCR) assay using clinical cerebrospinal fluid (CSF) for rapid diagnosis of eight pathogens causing the disease.
  
  
  METHODS
  The present work was dedicated to development of an in-house genus specific 16S rDNA nested PCR covering pathogens of eight genera responsible for causing bacterial meningitis using newly designed as well as literature based primers for respective genus. A total 150 suspected meningitis CSF obtained from the patients admitted to Central India Institute of Medical Sciences (CIIMS), India during the period from August 2011 to May 2014, were used to evaluate clinical sensitivity and clinical specificity of optimized PCR assays.
  
  
  RESULTS
  The analytical sensitivity and specificity of our newly designed genus-specific 16S rDNA PCR were found to be ≥92%. With such a high sensitivity and specificity, our in-house nested PCR was able to give 100% sensitivity in clinically confirmed positive cases and 100% specificity in clinically confirmed negative cases indicating its applicability in clinical diagnosis.
  
  
  CONCLUSIONS
  Our in-house nested PCR system therefore can diagnose the accurate pathogen causing bacterial meningitis and therefore be useful in selecting a specific treatment line to minimize morbidity. Results are obtained within 24 h and high sensitivity makes this nested PCR assay a rapid and accurate diagnostic tool compared to traditional culture-based methods.
authors.affiliation: Research Laboratory, Central India Institute of Medical Sciences, Nagpur, Maharashtra, India.
authors.forename: Sharda P
authors.initials: SP
authors.lastname: Bhagchandani
authors.affiliation: Research Laboratory, Central India Institute of Medical Sciences, Nagpur, Maharashtra, India.
authors.forename: Sushant
authors.initials: S
authors.lastname: Kubade
authors.affiliation: Research Laboratory, Central India Institute of Medical Sciences, Nagpur, Maharashtra, India.
authors.forename: Priyanka P
authors.initials: PP
authors.lastname: Nikhare
authors.affiliation: Research Laboratory, Central India Institute of Medical Sciences, Nagpur, Maharashtra, India.
authors.forename: Sonali
authors.initials: S
authors.lastname: Manke
authors.affiliation: Research Laboratory, Central India Institute of Medical Sciences, Nagpur, Maharashtra, India.
authors.forename: Nitin H
authors.initials: NH
authors.lastname: Chandak
authors.affiliation: Research Laboratory, Central India Institute of Medical Sciences, Nagpur, Maharashtra, India.
authors.forename: Dinesh
authors.initials: D
authors.lastname: Kabra
authors.affiliation: Research Laboratory, Central India Institute of Medical Sciences, Nagpur, Maharashtra, India.
authors.forename: Neeraj N
authors.initials: NN
authors.lastname: Baheti
authors.affiliation: Research Laboratory, Central India Institute of Medical Sciences, Nagpur, Maharashtra, India.
authors.forename: Vijay S
authors.initials: VS
authors.lastname: Agrawal
authors.affiliation: Research Laboratory, Central India Institute of Medical Sciences, Nagpur, Maharashtra, India.
authors.forename: Pankaj
authors.initials: P
authors.lastname: Sarda
authors.affiliation: Research Laboratory, Central India Institute of Medical Sciences, Nagpur, Maharashtra, India.
authors.forename: Parikshit
authors.initials: P
authors.lastname: Mahajan
authors.affiliation: Research Laboratory, Central India Institute of Medical Sciences, Nagpur, Maharashtra, India.
authors.forename: Ashish
authors.initials: A
authors.lastname: Ganjre
authors.affiliation: Environmental Genomics Unit, National Environmental Engineering Research Institute, Nagpur, Maharashtra, India.
authors.forename: Hemant J
authors.initials: HJ
authors.lastname: Purohit
authors.affiliation: Research Laboratory, Central India Institute of Medical Sciences, Nagpur, Maharashtra, India.
authors.forename: Lokendra
authors.initials: L
authors.lastname: Singh
authors.affiliation: Research Laboratory, Central India Institute of Medical Sciences, Nagpur, Maharashtra, India.
authors.forename: Girdhar M
authors.initials: GM
authors.lastname: Taori
authors.affiliation: Research Laboratory, Central India Institute of Medical Sciences, Nagpur, Maharashtra, India.
authors.forename: Hatim F
authors.initials: HF
authors.lastname: Daginawala
authors.affiliation: Research Laboratory, Central India Institute of Medical Sciences, Nagpur, Maharashtra, India. raj_ciims@rediffmail.com.
authors.forename: Rajpal S
authors.initials: RS
authors.lastname: Kashyap
mesh_terms: D000150:Acinetobacter / Q000302:isolation & purification; D000293:Adolescent; D000328:Adult; D000368:Aged; D002648:Child; D002675:Child, Preschool; D017931:DNA Primers; D004269:DNA, Bacterial / Q000235:genetics; D004755:Enterobacteriaceae / Q000302:isolation & purification; D005260:Female; D006190:Haemophilus / Q000302:isolation & purification; D006801:Humans; D007194:India; D008297:Male; D016920:Meningitis, Bacterial / Q000134:cerebrospinal fluid / Q000175:diagnosis*; D008837:Micrococcus / Q000302:isolation & purification; D008875:Middle Aged; D009343:Neisseria / Q000302:isolation & purification; D016133:Polymerase Chain Reaction / Q000379:methods*; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA; D013210:Staphylococcus / Q000302:isolation & purification; D013291:Streptococcus / Q000302:isolation & purification; D055815:Young Adult
chemical_list: D017931:DNA Primers; D004269:DNA, Bacterial; D012336:RNA, Ribosomal, 16S