2017–2019年中国南方五省规模化养猪场副猪格拉菌血清型、耐药性及β-内酰胺类耐药基因bla-TEM分子特征分析

Characterization of serotypes, antibiotic resistance, and bla-TEMgene of Glaesserella parasuis in large-scale pig farms in 5 southern provinces of China during 2017–2019

摘要:

【目的】旨在分析当前规模化养殖场副猪格拉菌(Glaesserella parasuis)优势血清型、耐药特性、耐药基因与分子特征。【方法】对源自规模化养猪场21株副猪格拉菌临床分离株，采用PCR鉴定血清型；利用K-B纸片扩散法鉴定其对25种抗生素的耐药表型；采用PCR检测bla-_TEM、bla-_NDM和bla-_CTX等7种耐药基因，并采用Chi-square test和Fisher exact test分析耐药表型和耐药基因型的相关性；耐药基因目的条带测序，并应用CLC Sequence Viewer软件分析β-内酰胺类耐药基因(bla-_TEM)编码蛋白氨基酸关键位点差异与耐药性的关系。【结果】21株副猪格拉菌临床分离株的优势血清型为4和12型；对β-内酰胺类药物苯唑西林的耐药性较强，耐药菌占比达61.9%(13/21)；多重耐药菌株占比高达90.5%(19/21)；β-内酰胺类耐药基因bla-_TEM携带率较高(52.4%，11/21)，且bla-_TEM与β-内酰胺类药物青霉素G、苯唑西林和头孢拉定的耐药性显著相关，部分bla-_TEM编码氨基酸存在可能与副猪格拉菌耐药能力有关的差异位点。【结论】本研究表明，规模化养猪场的副猪格拉菌多重耐药情况仍很严重，并明确了被调查区域β-内酰胺类药物耐药率高的主要原因是携带耐药基因bla-_TEM，为加强对规模化养猪场副猪格拉菌耐药性监测提供理论依据。

关键词:副猪格拉菌;bla-_TEM;分子特征

Abstract:

[Objective] To analyze the serotypes, antibiotic resistance, and resistance genes of 21 isolates of Glaesserella parasuis in large-scale pig farms.[Methods] Serotypes were identified by PCR and the resistance to 25 antibiotics with the K-B disk diffusion method. Seven resistance genes, including bla-_TEM, bla-_NDM, and bla-_CTX, were detected by PCR. Then chi-square test and Fisher's exact test were used to analyze the association between resistance phenotypes and genotypes. The β-lactam resistance gene bla-_TEM was sequenced and the sequences of amino acids encoded by bla-_TEM from different isolates were compared. Finally, the relationship between the differential sites of the amino acids and antibiotic resistance was analyzed by CLC Sequence Viewer. [Results] The isolates were dominated by serotypes 4 and 12. They showed high rate (61.9%, 13/21) of resistance to β-lactam antibiotic oxacillin and 90.5% (19/21) of the isolates demonstrated multi-antibiotic resistance. bla-_TEMwas identified in 52.4% (11/21) of the isolates, which was significantly correlated with the resistance phenotype of penicillin G, oxacillin, and cefradine. There were mutations of the amino acids encoded by bla-_TEM among isolates, which may be related to the antibiotic resistance of the isolates.[Conclusion]Multi-antibiotic resistance of G. parasuis isolates is prevalent in large-scale pig farms, and the high rate of β-lactam resistance is attributed to the inborn resistance gene bla-_TEM. The conclusion lays a theoretical basis for surveillance of antibiotic resistance of G. parasuis in large-scale pig farms.

Key words:Glaesserella parasuis;bla-_TEM;molecular characterization