Evaluation of nested PCR in detection of Helicobacter pylori targeting a highly conserved gene: HSP60.
OBJECTIVE
To comparatively evaluate a new nested set of primers designed for the detection of Helicobacter pylori targeting a highly conserved heat shock protein gene (Hsp60).


METHODS
A total of 60 subjects having peptic ulcer diseases were tested for the detection of H. pylori using rapid urease test (RUT), histology, culture, and polymerase chain reaction (PCR) in their antral biopsy specimens. A newly designed Hsp60 gene-based primer set was evaluated against commonly used PCR primers for detection of H. pylori.


RESULTS
Forty-six of the 60 study subjects were found positive for culture isolation and all the 46 culture-positive specimens were also positive with Hsp60 gene PCR. Of the 46 culture-positive specimens, 44 were positive for 16S rRNA gene, ureC gene, RUT, and histology whereas only 29 were positive with ureA gene PCR. Of the 14 culture-negative subjects, 10 were positive with 16S rRNA gene, 4 were positive with ureC (glmM) gene PCR, and 2 were positive with RUT and 1 was positive on histology.


CONCLUSIONS
This study shows that nested amplification targeting Hsp60 gene is the most sensitive and specific with LR+ and LR- values of proportional, variant and 0, respectively, when compared with the other three PCR methods. Also, HSP60 gene-specific nested protocol was the most appropriate for detection of H. pylori in clinical specimens. This is particularly valuable because it can be used as a noninvasive method for detecting H. pylori infection in young children and also, in follow-up studies with peptic ulcer patients, on samples like feces and saliva.
Singh
Varsha
V
Department of Biochemistry, Institute of Medical Sciences, Banaras Hindu University, Varanasi, India.
Mishra
Shrutkirti
S
Rao
G R K
GR
Jain
Ashok Kumar
AK
Dixit
V K
VK
Gulati
Anil Kumar
AK
Mahajan
Divya
D
McClelland
Michael
M
Nath
Gopal
G
D001426:Bacterial Proteins / Q000235:genetics*; D001706:Biopsy; D018834:Chaperonin 60 / Q000235:genetics*; D005753:Gastric Mucosa / Q000473:pathology; D016481:Helicobacter Infections / Q000175:diagnosis*; D016480:Helicobacter pylori / Q000235:genetics* / Q000302:isolation & purification*; D006801:Humans; D010437:Peptic Ulcer / Q000382:microbiology; D016133:Polymerase Chain Reaction / Q000379:methods*; D011706:Pyloric Antrum / Q000473:pathology; D012329:RNA, Bacterial / Q000235:genetics; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D012680:Sensitivity and Specificity; D014510:Urease / Q000032:analysis / Q000235:genetics
D001426:Bacterial Proteins; D018834:Chaperonin 60; D012329:RNA, Bacterial; D012336:RNA, Ribosomal, 16S; D014510:Urease