title: Evaluation of a proposed molecular methodology for the serotyping of Avibacterium paragallinarum.
abstract: A multiplex (m)PCR and a PCR followed by restriction fragment length polymorphism (RFLP) analysis of Avibacterium paragallinarum have been proposed as alternatives to conventional serotyping by the Page scheme. We evaluated both methods, and also sequenced the PCR-RFLP target fragment to reexamine the capacity of molecular serotyping. Eleven reference strains and 27 field isolates were used. Many reference strains and isolates were misidentified as Page serogroup B. The sequence analysis revealed 6 profiles based on the matching rates of the target sequence with the 3 reverse primers of the mPCR. The reference strains and field isolates in profiles 1 and 4 were correctly identified as serogroup A or C by the mPCR. The strains and/or isolates in profiles 2, 3, 5, and 6 could be misidentified as serogroup B or as nontypeable by the mPCR. The homology comparison of the sequences showed that the target sequence of the mPCR, called region 2, was not Page serogroup specific, although some Kume serovars, such as A-1 and C-2, were correctly serotyped. In addition, there was a 9 nucleotide deletion in the sequences of profiles 1, 3, and 5, but not of profiles 2, 4, and 6. Overall, we confirmed that the mPCR and PCR-RFLP molecular assays are not suitable for identifying the serogroups of A. paragallinarum isolates. With further study, analysis of region 2 sequences may have potential as a means of recognizing the Kume serovars of A. paragallinarum isolates.
authors.affiliation: Beijing Key Laboratory for Prevention and Control of Infectious Diseases in Livestock and Poultry, Institute of Animal Husbandry and Veterinary Medicine, Beijing Municipal Academy of Agriculture and Forestry, Beijing, China (Wang, Sun, Zhang, Zhou, Xu, Chen)Queensland Alliance for Agriculture and Food Innovation, The University of Queensland, St. Lucia, Queensland, Australia (Blackall).
authors.forename: Hongjun
authors.initials: H
authors.lastname: Wang
authors.affiliation: Beijing Key Laboratory for Prevention and Control of Infectious Diseases in Livestock and Poultry, Institute of Animal Husbandry and Veterinary Medicine, Beijing Municipal Academy of Agriculture and Forestry, Beijing, China (Wang, Sun, Zhang, Zhou, Xu, Chen)Queensland Alliance for Agriculture and Food Innovation, The University of Queensland, St. Lucia, Queensland, Australia (Blackall) sunhuiling01@163.com.
authors.forename: Huiling
authors.initials: H
authors.lastname: Sun
authors.affiliation: Beijing Key Laboratory for Prevention and Control of Infectious Diseases in Livestock and Poultry, Institute of Animal Husbandry and Veterinary Medicine, Beijing Municipal Academy of Agriculture and Forestry, Beijing, China (Wang, Sun, Zhang, Zhou, Xu, Chen)Queensland Alliance for Agriculture and Food Innovation, The University of Queensland, St. Lucia, Queensland, Australia (Blackall).
authors.forename: Patrick J
authors.initials: PJ
authors.lastname: Blackall
authors.affiliation: Beijing Key Laboratory for Prevention and Control of Infectious Diseases in Livestock and Poultry, Institute of Animal Husbandry and Veterinary Medicine, Beijing Municipal Academy of Agriculture and Forestry, Beijing, China (Wang, Sun, Zhang, Zhou, Xu, Chen)Queensland Alliance for Agriculture and Food Innovation, The University of Queensland, St. Lucia, Queensland, Australia (Blackall).
authors.forename: Zhenhua
authors.initials: Z
authors.lastname: Zhang
authors.affiliation: Beijing Key Laboratory for Prevention and Control of Infectious Diseases in Livestock and Poultry, Institute of Animal Husbandry and Veterinary Medicine, Beijing Municipal Academy of Agriculture and Forestry, Beijing, China (Wang, Sun, Zhang, Zhou, Xu, Chen)Queensland Alliance for Agriculture and Food Innovation, The University of Queensland, St. Lucia, Queensland, Australia (Blackall).
authors.forename: Hongzhuan
authors.initials: H
authors.lastname: Zhou
authors.affiliation: Beijing Key Laboratory for Prevention and Control of Infectious Diseases in Livestock and Poultry, Institute of Animal Husbandry and Veterinary Medicine, Beijing Municipal Academy of Agriculture and Forestry, Beijing, China (Wang, Sun, Zhang, Zhou, Xu, Chen)Queensland Alliance for Agriculture and Food Innovation, The University of Queensland, St. Lucia, Queensland, Australia (Blackall).
authors.forename: Fuzhou
authors.initials: F
authors.lastname: Xu
authors.affiliation: Beijing Key Laboratory for Prevention and Control of Infectious Diseases in Livestock and Poultry, Institute of Animal Husbandry and Veterinary Medicine, Beijing Municipal Academy of Agriculture and Forestry, Beijing, China (Wang, Sun, Zhang, Zhou, Xu, Chen)Queensland Alliance for Agriculture and Food Innovation, The University of Queensland, St. Lucia, Queensland, Australia (Blackall).
authors.forename: Xiaoling
authors.initials: X
authors.lastname: Chen
mesh_terms: D000818:Animals; D002645:Chickens; D006192:Haemophilus Infections / Q000175:diagnosis / Q000382:microbiology / Q000662:veterinary*; D044134:Haemophilus paragallinarum / Q000235:genetics / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction / Q000662:veterinary; D012150:Polymorphism, Restriction Fragment Length; D011201:Poultry Diseases / Q000175:diagnosis* / Q000382:microbiology; D012680:Sensitivity and Specificity; D012703:Serotyping / Q000662:veterinary
keywords: Avibacterium paragallinarum; PCR-RFLP; infectious coryza; multiplex PCR; profile; region 2 sequence; restriction fragment length polymorphism; serotyping