不同地区鸡毒支原体的分离鉴定及药物敏感性研究

doi:10.16431/j.cnki.1671-7236.2023.01.037

摘要/Abstract

摘要： 【目的】了解中国不同地区鸡毒支原体（Mycoplasma gallisepticum，MG）的流行和耐药情况，为禽支原体病的监测与防控提供科学依据。【方法】在广东、福建和安徽3个省份疑似MG感染的养殖场采集的43份气囊样品中分离培养和纯化MG流行菌株，并对其进行培养特性观察、生化及染色鉴定、血清学特异性鉴定、PCR测序分析、致病性试验和对常见抗菌药物敏感性试验。【结果】分离到3株疑似MG，均可使培养基颜色变黄，在固体培养基上呈典型"煎蛋"样菌落，可发酵葡萄糖，不水解精氨酸，不能利用尿素，符合MG培养特性。姬姆萨染色观察菌体形态、血清学特异性鉴定结果进一步证实分离株为MG。mgc2基因测序结果显示，3株分离株与强毒代表株亲缘关系更近。致病性试验结果显示，3株分离株与MG感染临床症状一致，发病率为70%~90%，致病力较强。药物敏感性试验结果显示，3株分离株均对恩诺沙星、替米考星、土霉素、氟苯尼考耐药，对泰万菌素和沃尼妙林均敏感，对大观霉素、金霉素、泰乐菌素和泰妙菌素的敏感性具有地域差异：福建株对大观霉素表现明显耐药，安徽株对泰妙菌素表现耐药，而广东株对大观霉素、金霉素和泰妙菌素均较敏感。【结论】本试验成功分离到3株MG，致病力较强，且均存在一定程度耐药，对药物的敏感程度有地区性差异，今后仍需加强各地区MG的药物敏感性监测，减少耐药性的产生与扩散。

关键词: 鸡毒支原体; 分离鉴定; 药物敏感性试验

Abstract: 【Objective】 The aim of this experiment was to investigate the prevalence and drug resistance of Mycoplasma gallisepticum (MG) in different areas of China, so as to provide a scientific basis for the surveillance, prevention and control of MG disease.【Method】 The prevalent strains were isolated and purified from the 43 infected air sacs from suspected MG infected chickens in poultry farms of Guangdong, Fujian and Anhui provinces.The isolates were identified by culture characteristics test, biochemical characteristics test, Giemsa staining, serological methods and then PCR sequencing analysis, pathogenicity and sensitivity to common antibiotics were tested.【Result】 3 isolates were obtained successfully, and could change the broth medium from red to yellow.They showed typical "fried egg"-like colony in the solid medium and could ferment glucose, not hydrolyze arginine, and not decompose urea, which were consistent with the culture characteristics of MG.Results of Giemsa staining and serological specific identification further confirmed the isolates to be MG. The sequence analysis of the mgc2 gene revealed that the 3 isolates were more closely related to the typical virulent strains.Animal challenge tests showed that the clinical symptoms of the 3 isolates were consistent with those of MG infection, and the incidence was 70%-90%, the 3 isolates had high pathogenicity.The results of antimicrobial susceptibility test showed that the 3 isolates were resistant to enrofloxacin, temicoxin, oxytetracycline and flufenicol, and were sensitive to tylvalosin and warnimilin.The sensitivity of the 3 isolates to spectinomycin, aureomycin, tyloxacin and tiamulin was different in different regions.The Fujian strain showed obvious resistance to spectinomycin, the Anhui strain showed resistance to tiamulin, and the Guangdong strain was sensitive to spectinomycin, aureomycin and tiamulin.【Conclusion】 3 MG isolates with high pathogenicity were isolated, and they were drug-resistant, their sensitivities to drugs were regional.In the future, it was still necessary to strengthen the drug sensitivity monitoring of MG in various regions to reduce the emergence and spread of drug resistance.

Key words: Mycoplasma gallisepticum; isolation and identification; drug sensitive test

中图分类号:

S852.62

魏津, 王甲, 赵浩然, 刘博, 刘元杰, 姚文生, 刘燕, 马欣. 不同地区鸡毒支原体的分离鉴定及药物敏感性研究[J]. 中国畜牧兽医, 2023, 50(1): 368-376.

WEI Jin, WANG Jia, ZHAO Haoran, LIU Bo, LIU Yuanjie, YAO Wensheng, LIU Yan, MA Xin. Isolation, Identification and Antimicrobial Susceptibility of Mycoplasma gallisepticum from Different Areas[J]. China Animal Husbandry and Veterinary Medicine, 2023, 50(1): 368-376.

参考文献

[1] HU W Y, ZHANG W, SYED W A S, et al.Mycoplasma gallisepticum infection triggered histopathological changes, oxidative stress and apoptosis in chicken thymus and spleen[J].Developmental and Comparative Immunology, 2021, 114(1):103832.
[2] 王育伟, 岳华, 张晓晖, 等.鸡毒支原体研究概况[J].动物医学进展, 2016, 37(7):106-110. WANG Y W, YUE H, ZHANG X J, et al.The situation of research on Mycoplasma galliscepticum[J].Progress in Veterinary Medicine, 2016, 37(7):106-110.(in Chinese)
[3] 牛家强, 徐业芬, 胡思顺, 等.藏鸡鸡毒支原体的分离鉴定及对常用抗菌药物的敏感性试验[J].中国兽医学报, 2018, 38(11):2084-2093. NIU J Q, XU Y F, HU S S, et al.Isolation and identification of Mycoplasma galliscepticum from Tibetan chickens in Tibet and its sensitivity to common antimicrobial agents[J].Chinese Journal of Veterinary Science, 2018, 38(11):2084-2093.(in Chinese)
[4] 刘英慧, 陈亦杏, 李仕楷, 等.鹅源鸡毒支原体分离鉴定与致病性研究[J].中国家禽, 2022, 44(2):106-111. LIU Y H, CHEN Y X, LI S K, et al.Isolation and identification on one strain of Mycoplasma gallisepticum from geese and study of its pathogenicity[J].China Poultry, 2022, 44(2):106-111.(in Chinese)
[5] MUHAMMAD I, HU W Y, MOHAMMAD Z K, et al.Current status of vaccine research, development, and challenges of vaccines for Mycoplasma gallisepticum[J].Poultry Science, 2020, 99(6):4195-4202.
[6] 陈建.鸡毒支原体病的流行病学、临床表现、诊断与防控措施[J].现代畜牧科技, 2021, 83(11):87-88. CHEN J.Epidemiology, clinical manifestations and diagnosis and prevention and control measures of Mycoplasma galliscepticum[J].Modern Animal Husbandry Technology, 2021, 83(11):87-88.(in Chinese)
[7] 谢昊晋.鸡毒支原体流行病学调查及病例分析[D].扬州:扬州大学, 2020. XIE H J.Epidemiological investigation and case analysis of Mycoplasma gallisepticum[D].Yangzhou:Yangzhou University, 2020.(in Chinese)
[8] 孟林春.蛋鸡场鸡毒支原体(MG)感染的监测和两种检测MG抗体的血清学方法建立[D].扬州:扬州大学, 2022. MENG L C.The surveillance of Mycoplasma gallisepticum (MG) infection in layer flocks and the establishment of two serological methods for detection of MG antibody[D].Yangzhou:Yangzhou University, 2020.(in Chinese)
[9] XIAO X, SUN J, YANG T, et al.Pharmacokinetic/pharmacodynamic profiles of tiamulin in an experimental intratracheal infection model of Mycoplasma gallisepticum[J].Frontiers in Veterinary Science, 2020, 3(9):75.
[10] HOFACRE C L.Antimicrobial Drug Use in Poultry[M].5th ed.Hoboken:John Wiley & Sons Inc., 2013.
[11] GERCHMAN I, LEVISOHN S, MIKULA I, et al.Characterization of in vivo-acquired resistance to macrolides of Mycoplasma gallisepticum strains isolated from poultry[J].Veterinary Research, 2011, 42(1):90.
[12] 中国兽药典委员会.中华人民共和国兽药典(2020年版)三部[S].2020. COMMITTEE OF CHINA VETERINARY PHARMACOPOEIA.Veterinary Pharmacopoeia of People's Republic of China, (2020 edition) Ⅲ[S].2020.(in Chinese)
[13] OFFICEINTER NATIONAL DES EPIZOOTIES.OIE Terrestrial Manual[M].Paris:World Organization for Animal Health, 2018.
[14] GARCIA M, IKUTA N, LEVISOHN C, et al.Evaluation and comparison of various PCR methods for detection of Mycoplasma gallisepticum infection in chickens[J]. Avian Diseases, 2005, 49(1):125-132.
[15] TANNER A C, WU C.Adaptation of the Sensititre^® broth microdilution technique to antimicrobial susceptibility testing of Mycoplasma gallisepticum[J].Avian Diseases, 1992, 36(3):714-717.
[16] 吴春琳, 黄宝钦, 蓝天韵, 等.福建白羽肉鸡鸡毒支原体的分离鉴定及最小抑菌浓度测定[J].中国畜牧兽医, 2021, 48(4):1489-1497. WU C L, HUANG B Q, LAN T Y, et al.Isolation, identification, and minimum inhibitory concentration determination of Mycoplasma gallisepticum from white-feathered broilers in Fujian[J].China Animal Husbandry & Veterinary Medicine, 2021, 48(4):1489-1497.(in Chinese)
[17] 张楠.五种抗菌药物对鸡毒支原体的药动学和药效学研究[D].广州:华南农业大学, 2017. ZHANG N.Pharmacokinetic/pharmacodynamic (PK-PD) profiles of antibacterial agents against Mycoplasma gallisepticum[D].Guangzhou:South China Agricultural University, 2017.(in Chinese)
[18] HNATOW L L, KEELER C L JR, TESSMER L L, et al.Characterization of MGC2, a Mycoplasma gallisepticum cytadhesin with homology to the Mycoplasma pneumoniae 30-kilodalton protein P30 and Mycoplasma genitalium P32[J].Infection and Immunity, 1998, 66(7):3436-3442.
[19] ALEXANDER V S, NIKOLAY P E, ALEXANDER N K, et al.Biological characterization of Russian Mycoplasma gallisepticum field isolates[J].Avian Pathology, 2011, 40(4):213-219.
[20] 郭伟娜, 毕丁仁, 郭锐, 等.6种常用抗菌药物对鸡毒支原体的药物敏感性试验[J].中国兽医杂志, 2008, 44(10):64-65. GUO W N, BI D R, GUO R, et al.Drug sensitivity tests of six commonly used antibiotics to Mycoplasma gallisepticum[J].Chinese Journal of Veterinary Medicine, 2008, 44(10):64-65.(in Chinese)
[21] 孔意端, 林居纯, 陈继荣, 等, 鸡毒支原体不同地区分离株对常用抗菌药物的敏感性试验[J].动物医学进展, 2008, 29(6):35-38. KONG Y D, LIN J C, CHEN J R, et al.Sensitivity of Mycoplasma gallisepticum isolates from different geographical location to common antimicrobial agents[J].Progress in Veterinary Medicine, 2008, 29(6):35-38.(in Chinese)
[22] HUANG Z L, WU Y Z, ZHOU Z C, et al.Pharmacokinetic and pharmacodynamic integration and resistance analysis of tilmicosin against Mycoplasma gallisepticum in an in vitro dynamic model[J].Frontiers in Pharmacology, 2019, 10(6):670.
[23] 郭杨丽, 宋翔, 刘乃芝, 等.1株鸡毒支原体的分离鉴定及体外敏感中药的筛选[J].中国畜牧兽医, 2021, 48(3):1064-1071. GUO Y L, SONG X, LIU N Z, et al.Isolation and identification of one strain of Mycoplasma gallisepticum and screening of sensitive traditional Chinese medicine in vitro[J].China Animal Husbandry & Veterinary Medicine, 2021, 48(3):1064-1071.(in Chinese)
[24] GHARAIBEH S, AL-RASHDAN M.Change in antimicrobial susceptibility of Mycoplasma gallisepticum field isolates[J].Veterinary Microbiology, 2011, 150(4):379-383.
[25] MORROW C J, KREIZINGER Z, ACHARI R R, et al.Antimicrobial susceptibility of pathogenic Mycoplasmas in chickens in Asia[J].Veterinary Microbiology, 2020, 250:108840.