China Animal Husbandry and Veterinary Medicine ›› 2025, Vol. 52 ›› Issue (2): 901-911.doi: 10.16431/j.cnki.1671-7236.2025.02.039

• Basic Veterinary Medicine • Previous Articles    

Isolation,Identification and Pathogenicity Analysis of a Strain of Haemophilus parasuis Serotype 2

LI Qianwen1,2,3, HUA Tao1,2,3, CHANG Chen1,2,3, YAN Xingyu1,2,3, LI Junxing4, TANG Bo1,2,3   

  1. 1. Institute of Veterinary Immunology & Engineering, Jiangsu Academy of Agricultural Sciences, Nanjing 210014, China;
    2. National Center for Engineering and Technology Research of Veterinary Bioproducts, Nanjing 210014, China;
    3. GuoTai (Taizhou) Center of Technology Innovation for Veterinary Biologicals, Taizhou 225300, China;
    4. Institute of Animal Husbandry and Veterinary Research, Zhejiang Academy of Agricultural Sciences, Hangzhou 310021, China
  • Received:2024-05-20 Published:2025-01-24

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Abstract: 【Objective】 The purpose of this experiment was to clarify the infection of Haemophilus parasuis (Hps) in a pig farm in Zhejiang province,and clarify the serotype,biological characteristics and pathogenicity of the isolate,so as to provide theoretical reference for clinical control of Hps. 【Method】 The strain was isolated and purified from lung,heart blood,joint fluid and other disease materials of infected pigs.Gram staining microscopy,16S rRNA gene PCR amplification and sequencing were performed to determine the serotype of the isolate.The virulence genes,susceptibility to common antibiotics and pathogenicity of the isolate were detected.The secreted protein was extracted by saturated ammonium sulfate method,and the main enrichment regions of the protein were analyzed by electrophoresis. 【Result】 The isolate grew round,smooth and moist,pale in color,and pinpoint-sized colonies with neat edges on TSA medium.After inoculation into TSB medium,the isolate grew slowly at 1-3 h,entered the logarithmic phase at 3-4 h,and entered the stable phase at 4-12 h.Gram staining microscopy showed red short rod shape,long rod shape,varying in length,suspected Hps.PCR amplification of 16S rRNA gene showed a specific band of 680 bp,which was consistent with expectations.The results of serotype identification showed that specific bands of Hps serotype 2 were detected.BLAST comparison showed that the isolate was Hps serotype 2.Virulence gene test results showed that the isolate carried cdtA,cdtB,cdtC,vtaA1, rfaD,rfaE, gnd,cpaD,espP2,vacJ,rfaF,vtaA2,htrA and ompP2 genes.When the isolate infected BALB/c mice at a dose of 1.0×109 CFU/mL,the mice died within 5-10 h，and caused nucleus concentration and necrosis in the liver,a large amount of serous exudate in the alveolar cavity,mixed with red blood cells and bacteria,and inflammatory cell infiltration in the spleen.The results of drug sensitivity test showed that the isolate was resistant to cefoperazone,cefuroxime sodium,piperacillin and penicillin,intermediate to chloramphenicol,and sensitive to ciprofloxacin,levofloxacin and azithromycin.The size of secreted protein mainly concentrated in 25-180 ku. 【Conclusion】 In this study,a strain of Hps serotype 2 was isolated and identified.The isolate encoded a variety of virulence genes,was pathogenic,had obvious resistance to a variety of commonly used antibacterial drugs,and secreted a variety of proteins.The results indicated that quinolones,macrolides and aminoglycosides could be used for prevention and control in this farm.

Key words: Haemophilus parasuis; serotype; isolation and identification; pathogenicity

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