Title of article :
Comparison of the Detection Limits of the Culture and PCR Methods for the
Detection of Clostridium difficile, Clostridium perfringens, Campylobacter
jejuni, and Yersinia enterocolitica in Human Stool
Author/Authors :
Ganji، Leila نويسنده Department of Pathobiology, School of Public Health,
University of Medical Sciences, Tehran, Iran , , Azimirad، Masoumeh نويسنده Gastroenterology and Liver Diseases Research Center, Shahid Beheshti University Medical Sciences , , Farzi، Nastaran نويسنده Foodborne and Waterborne Disease Research Center, Research Institute for Gastroenterology and Liver Diseases, Shahid Beheshti University of Medical Sciences, Tehran, IR Iran , , Alebouyeh، Masoud نويسنده , , Shirazi، Mohammad Hassan نويسنده Department of Pathobiology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran , , Eshraghi، Seyed Saeed نويسنده Division of Microbiology, Department of Pathobiology, School of Public Health, Tehran University of Medical Sciences , , Mirshafiey، Abbas نويسنده , , Ebrahimi Daryani، Naser نويسنده , , Zali، Mohammad Reza نويسنده Department of Celiac Disease, Gastroenterology and Liver Diseases Research Center, Shahid Beheshti University of Medical Sciences, Tehran ,
Abstract :
Detection of fastidious enteropathogenic bacteria in fecal samples
of patients with gastroenteritis is a challenge in clinical
microbiological laboratories. The aim of this study was to compare the
detection limits of the PCR and culture methods for the diagnosis of
Campylobacter spp., Yersinia spp., Clostridium perfringens, and
Clostridium difficile in human stool samples. Healthy human stool and
sterile phosphate-buffered saline (PBS) samples were separately spiked
with 10-fold dilutions of C. jejuni, C. difficile, Y. enterocolitica,
and C. perfringens reference strains to obtain final concentrations of
101 - 108 colony forming units (CFU) per gram. Dilutions of each
suspension were inoculated onto specific culture media and colony counts
were determined. Polymerase chain reaction (PCR) was carried out on DNA
extracts of each dilution using specific primers. All of the assays were
performed in two separate replicas. In the cases of the culture and PCR
assays, detection limits of 101 and 102 CFU/g for C. difficile, 2 × 104
and 2 × 104 CFU/g for C. perfringens, 104 and 102 CFU/g for C. jejuni,
and 102 and 104 CFU/g for Y. enterocolitica, respectively, were
obtained. In the cases of the spiked PBS samples, a detection limit of
101 for C. jejuni and Y. enterocolitica was obtained using the culture
method. While 102 -fold higher sensitivity was observed for C. jejuni
via PCR compared with the culture assay, equal (C. perfringens) or lower
sensitivity limits (C. difficile and Y. enterocolitica) were detected
for the spiked stool samples with other bacteria. These results showed
differences in the bacterial culture and PCR methods for quantitative
detection of fastidious bacteria in human stool samples. However, a
bacterial load of 104 CFU per gram of stool was measured as a sufficient
amount for detection of the fastidious bacteria by either culture or PCR
assays. More suitable PCR methods could be used for rapid diagnosis of
the slow-growing bacteria in the patients’ stool samples.
