Comparative study on isolation techniques and characterization of Helicobacter pullorum in broiler chickens (Gallus sp.) and their farm environment in Selangor, Malaysia

Helicobacter pullorum is becoming important as an emerging zoonotic pathogen. It has been isolated from poultry in many countries but reports are lacking in developing countries, including Malaysia. Helicobacter pullorum is a fastidious organism, and not readily culturable. They are generally misdiagnosed as Campylobacter and currently are identified by molecular method. This study was conducted to determine the appropriate isolation method, the prevalence of H. pullorum in broiler chickens and farm environment, antibiotic resistance and plasmid profiles of H. pullorum isolated and to characterize the H. pullorum isolates. The first part of the study showed 14 of 57 (24.6%) chickens from five farms were positive for H. pullorum of which 8 (14%) were co-colonized with Campylobacter species. In this study, three methods reported by other researchers were used (Method I, II & III) that is, based on Ceelen et al. (2006c) as method I, Zanoni et al. (2007) as method II and Miller et al. (2006) as method III. In two methods (method I & II), discrete colonies of H. pullorum were not obtained but were mixed with Campylobacter species, while (one) method III gave both discrete colonies of H. pullorum and mixed colonies. In terms of better isolation, ease of preparation and recovery of pure isolates, this (Method III) third method was the method of choice. To improve the H. pullorum isolation percentage, a further modification was done to this Method III by adding an enrichment step and modifying the incubation temperature. Using this modified method (Method IV), 24 of 30 (80.0%) chickens from three farms were positive for H. pullorum while only 17 of 30 (56.7%) were positive using Method III. Method IV gave a better recovery of H. pullorum from chicken caecal contents. The overall prevalence in the second part of the study showed H. pullorum was present in 51 % broilers using method IV alone in chickens from another ten farms, eight open-house farms and two close-house farms.