柔嫩艾美耳球虫野外抗药性虫株的RAPD分析

RAPD Analyses of the Drug-resistance Strains of Eimeria Tenella Isolated From Fields

用RAPD方法对柔嫩艾美耳球虫 (Eimeriatenella)的 8个分别来源于黑龙江、北京、四川、甘肃和广东的中山、新会、东莞以及广州近郊的江村镇的野外分离虫株和 2个药物敏感性虫株进行了遗传多态性分析。同时用 9种抗球虫药物即马杜霉素 (5mg/kg)、盐霉素 (6 0mg/kg)、莫能霉素 (12 0mg/kg)、拉沙菌素 (90mg/kg)、克球多(15 0mg/kg)、常山酮 (3mg/kg)、氯氢苯乙氰 (1mg/kg)、尼卡巴嗪 (12 5mg/kg)和球痢灵 (12 5mg/kg)分别对 8个野外虫株进行抗药性试验 ,8个虫株对上述药物均有不同程度的抗药性。RAPD分析表明 :10个样品都有相似而清晰的主带 ,每个泳道有 5～ 13条带不等 ,大小为 0 18～ 2 10kb。它们之间的相似率 (SI)大于 4 0 5 8% ,最大的为90 72 % ,这种相似率属种内变异水平。根据SI值可把 10个样品划分为 3个类群 :广东虫株类群 ,群内比较 ,平均SI值为 76 6 0 % ;广东省外虫株类群 ,群内比较的平均SI值为 72 14 % ;敏感株类群 ,其间的SI值为 89 2 7%。在另一方面 ,广东株与广东省外株、敏感株之间比较 ,其SI的平均值分别仅为 6 3 0 3%和 4 7 2 5 % 。

The genetic relatedness of 8 field isolates derived from Heilongjiang,Beijing,Sichuan,Gansu province and Zhongshan,Xinhui,Dongguan,Jiangchun(Guangzhou suburb)of Guangdong province,and 2 drug sensitive strains of Eimeria tenella was assayed by means of random amplified polymorphic DNA(RAPD).The 8 field isolates were determined to be resistant with different extent to 9 anticoccials:maduramacin(5 mg/kg),salinomycin(60 mg/kg),monencin (120 mg/kg),lasanosidum(90 mg/kg),clopidol(150 mg/kg),halofugenone(3mg/kg),diclazuril(1 mg/kg),nicarbazin(125 mg/kg)and zoalene(125 mg/kg).5 different olignucleotides were used to generate similarity coefficients for the drug resistant and sensitive strains of E.tenella. 5 13 DNA segments,depending on the strain primer combination,were amplified with the various primers.Amplification products ranged in size from 0 18 2 10 kb.Band match analyses were used to calculate similarity coefficient(SI).A similarity analysis of E.tenella different strains yielded similarity coefficient of 40 58% 90 72%,belonging to infra species level.SI was very high between 2 drug sensitive strains(89 2%),and among strains(76 6%)derived from Guangdong province,as well as isolates(72 14%)derived from Heilongjiang,Beijing,Sichuan, Gansu province.On the other hand,SI was 63 03% between groups of Guangdong isolates and strains outside Guangdong;47 25% between drug resistant strain(field isolates)and sensitive strains,indicating that genetic difference existed among strains of E.tenella with various drugsensitivity.