Development of a multiplex PCR assay for detection of Riemerella anatipestifer serotype 1 and serotype 2 strains

IF 2.4 2区农林科学 Q3 MICROBIOLOGY Veterinary microbiology Pub Date : 2025-02-14 DOI:10.1016/j.vetmic.2025.110435

Mengsi Wu , Rong Guo , Meitong Chen , Min Zhu , Zongchao Chen , Chan Ding , Shengqing Yu

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Abstract

Riemerella anatipestifer infection is one of the main infectious diseases that threaten the poultry industry at present. Serotypes 1 and 2 are the main serotypes causing global outbreaks of R. anatipestifer. In this study, we designed the primers specific for R. anatipestifer serotypes 1 and serotype 2 strains, and for R. anatipestifer 16S rRNA (for the species identification) to establish a multiplex PCR assay for rapid detection of the serotype 1 and serotype 2 strains. The specificity test showed that a 505 bp fragment was amplified from serotype 1 strains, and a 1125 bp fragment was amplified from serotype 2 strains. A 843 bp fragment of 16S rRNA was amplified from R. anatipestifer strains with different serotypes. No amplification band was shown for other species of bacterial pathogens, including Escherichia coli, Salmonella, Pasteurella multocida, Mycoplasma gallisepticum and Mycoplasma synoviae. The sensitivity test showed a detection limit of 10² CFU of the multiplex PCR assay. Furthermore, a total of 60 R. anatipestifer clinical isolates were tested for identification of the serotypes, and the results showed that a 843 bp 16S rRNA fragment was amplified from all 60 isolates, confirming they are R. anatipestifer strains. Moreover, a 505 bp serotype 1 fragment was amplified from 9 isolates, and a 1125 bp serotype 2 fragment was amplified from 28 isolates. R. anatipestifer serotype 1 and 2 strains accounted for 61.67 % of the isolates. The results were further validated by slide agglutination test, which showed 100 % consistency with the multiplex PCR.

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来源期刊

Veterinary microbiology 农林科学-兽医学

CiteScore

5.90

自引率

6.10%

发文量

221

审稿时长

52 days

期刊介绍： Veterinary Microbiology is concerned with microbial (bacterial, fungal, viral) diseases of domesticated vertebrate animals (livestock, companion animals, fur-bearing animals, game, poultry, fish) that supply food, other useful products or companionship. In addition, Microbial diseases of wild animals living in captivity, or as members of the feral fauna will also be considered if the infections are of interest because of their interrelation with humans (zoonoses) and/or domestic animals. Studies of antimicrobial resistance are also included, provided that the results represent a substantial advance in knowledge. Authors are strongly encouraged to read - prior to submission - the Editorials (''Scope or cope'' and ''Scope or cope II'') published previously in the journal. The Editors reserve the right to suggest submission to another journal for those papers which they feel would be more appropriate for consideration by that journal. Original research papers of high quality and novelty on aspects of control, host response, molecular biology, pathogenesis, prevention, and treatment of microbial diseases of animals are published. Papers dealing primarily with immunology, epidemiology, molecular biology and antiviral or microbial agents will only be considered if they demonstrate a clear impact on a disease. Papers focusing solely on diagnostic techniques (such as another PCR protocol or ELISA) will not be published - focus should be on a microorganism and not on a particular technique. Papers only reporting microbial sequences, transcriptomics data, or proteomics data will not be considered unless the results represent a substantial advance in knowledge. Drug trial papers will be considered if they have general application or significance. Papers on the identification of microorganisms will also be considered, but detailed taxonomic studies do not fall within the scope of the journal. Case reports will not be published, unless they have general application or contain novel aspects. Papers of geographically limited interest, which repeat what had been established elsewhere will not be considered. The readership of the journal is global.