Multiplex PCR for direct identification of Campylobacter spp. in human and chicken stools.
Differentiation between Campylobacter jejuni and Campylobacter coli is problematic in clinical specimens due to fastidious growth requirements and limited biochemical tests. This study describes a rapid, multiplex PCR protocol for the direct detection and differentiation of C. jejuni and C. coli in stools. An evaluation was carried out of this multiplex protocol based on the detection of cadF (genus specific), and hipO (C. jejuni) and asp (C. coli) genes, using stool from patients with Campylobacter enteritis and chicken. Protocol sensitivity was assessed and specificity determined using a panel of enteric bacteria, and evaluation of 30 diarrhoeic stool specimens culture negative for Campylobacter. Of the 114 specimens (54 human and 60 chicken) evaluated by the protocol, 70 (61.4 %) were identified as C. jejuni, 35 (30.7 %) as C. coli and 9 (7.9 %) as a mixed infection/colonization with both species. All mixed infections were identified as C. jejuni by culture. Among the stool specimens that were culture negative for Campylobacter, two (6.7 %) were C. jejuni positive by multiplex PCR. The protocol sensitivity limit was 0.015-0.016 ng C. jejuni and C. coli DNA mul(-1) in the specimen. There was no cross-reaction with the reference strains assessed. Comparison of hippurate test and multiplex PCR demonstrated 17 isolates with false-positive hippurate enzymic activity and 7 with false-negative activity. This rapid protocol (turnaround time 6 h) is highly sensitive and specific for direct evaluation of stool for these pathogens. It has significant application for routine clinical diagnostic and epidemiological purposes.
Al Amri
Aisha
A
Department of Microbiology, Immunology and Infectious Diseases, College of Medicine and Medical Sciences, Arabian Gulf University, PO Box 22979, Manama, Kingdom of Bahrain.
Senok
Abiola C
AC
Department of Clinical Sciences, College of Medicine, University of Sharjah, United Arab Emirates.
Ismaeel
Abdulrahman Yusuf
AY
Department of Microbiology, Immunology and Infectious Diseases, College of Medicine and Medical Sciences, Arabian Gulf University, PO Box 22979, Manama, Kingdom of Bahrain.
Al-Mahmeed
Ali E
AE
Department of Microbiology, Immunology and Infectious Diseases, College of Medicine and Medical Sciences, Arabian Gulf University, PO Box 22979, Manama, Kingdom of Bahrain.
Botta
Giuseppe A
GA
Department of Medical and Morphological Sciences, Institute of Microbiology, Udine Medical School, Udine, Italy.|Department of Microbiology, Immunology and Infectious Diseases, College of Medicine and Medical Sciences, Arabian Gulf University, PO Box 22979, Manama, Kingdom of Bahrain.
D000818:Animals; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics; D001715:Bird Diseases / Q000175:diagnosis / Q000382:microbiology; D002169:Campylobacter Infections / Q000175:diagnosis* / Q000382:microbiology / Q000662:veterinary; D017000:Campylobacter coli / Q000145:classification* / Q000302:isolation & purification*; D016123:Campylobacter jejuni / Q000145:classification* / Q000302:isolation & purification*; D002352:Carrier Proteins / Q000235:genetics; D002645:Chickens; D003429:Cross Reactions; D004751:Enteritis / Q000382:microbiology; D005243:Feces / Q000382:microbiology*; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; C107872:CadF protein, Campylobacter jejuni; D002352:Carrier Proteins