A cytolethal distending toxin gene-based multiplex PCR assay for detection of Campylobacter spp. in stool specimens and comparison with culture method.
In this study, we evaluated the applicability of cytolethal distending toxin (cdt) gene-based species-specific multiplex PCR for the direct detection and identification of Campylobacter jejuni, C. coli and C. fetus from stool specimens of patients with gastroenteritis in comparison to culture methods. A total of 711 stool specimens were examined for the isolation or detection of campylobacters by using Skirrow's selective agar culture plates, a filtration method and the multiplex PCR assay. Forty-one and 36 C. jejuni strains were isolated by culture and filtration methods, respectively. In addition, 2 and 3 C. coli strains were isolated by Skirrow and the filtration methods, respectively. However, when the multiplex PCR was employed, the cdtB genes of C. jejuni and C. coli were detected in 45 and 4 stool samples, respectively, and 9 C. jejuni PCR-positive samples by multiplex PCR were negative by culture method. Sequence analysis of the PCR products obtained from 8 stool specimens from which campylobacters were not isolated by culture method but the sequences exactly matched with that of the cdtB gene of C. jejuni strain 81-176. None of the remaining stool samples which were culture negative for campylobacters produced any amplicon. Stool samples were defined as Campylobacter-positive if detected by any method. The sensitivity of the multiplex PCR was 83%, which was higher than Skirrow (74%) and filtration method (66%). These data indicate that cdtB gene-based multiplex PCR is a rapid and more sensitive method to identify the most important species of Campylobacter for human diseases. (248).
Graduate School of Life and Environmental Sciences, Osaka Prefecture University, 1-1 Rinku Ourai-Kita, Izumisano, Osaka 598-8531, Japan. shinji@vet.osakafu-u.ac.jp
Sachi
S
Shiramaru
Masahiro
M
Asakura
Haruna
H
Inoue
Akira
A
Nagita
Akio
A
Matsuhisa
Shinji
S
Yamasaki
D001427:Bacterial Toxins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D002167:Campylobacter / Q000235:genetics* / Q000302:isolation & purification; D018929:Cell Culture Techniques / Q000379:methods; D005243:Feces / Q000382:microbiology*; D005374:Filtration / Q000379:methods; D005759:Gastroenteritis / Q000175:diagnosis* / Q000382:microbiology*; D006801:Humans; D008969:Molecular Sequence Data; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D017422:Sequence Analysis, DNA; D013045:Species Specificity
D001427:Bacterial Toxins; C067044:cytolethal distending toxin