毒害艾美耳球虫山西株的分离和鉴定

Isolation and Identification of Eimeria necatrix Shanxi Strain

为了鉴定造成太谷区某鸡场57日龄蛋鸡死亡的鸡球虫种类,明确其生物学特性,本试验采用饱和食盐水漂浮法和单卵囊分离技术从病死鸡粪便中分离纯化得到1株球虫;将分离球虫口服接种SPF鸡,观察并测定卵囊最短孢子化时间、裂殖体寄生部位和大小、卵囊寄生部位和大小、潜隐期、死亡率、增重情况、肠道病变记分等指标,了解该虫株的生物学特点并初步鉴定其类型,随后采用序列特征性扩增区域(SCAR)标记技术进一步确定虫株类型。结果显示,分离虫株的裂殖体寄生于空肠,平均大小为(63.17±7.66)μm×(47.12±6.91)μm,卵囊仅盲肠可见,平均大小为(19.92±0.20)μm×(19.27±0.11)μm,虫株潜隐期为152 h,最短孢子化时间为18 h,接种5.0×104个卵囊/羽和1.1×105个卵囊/羽的分离虫株可分别造成40%和50%的实验鸡死亡,相对增重率下降至31.61%和20.87%,病变记分分别达到3.9和4.0;SCAR标记扩增显示,分离虫株与毒害艾美耳球虫(E.necatrix)的同源性达97.1%,命名为E.necatrix-CN-SX-2018。结果表明,本试验分离得到1株致病性较强的E.necatrix,不仅丰富了山西省E.necatrix流行毒株的生物信息,也为鸡球虫疫苗评价提供了良好的种子资源。

This study aimed to identify the species of chicken coccidia that caused the death of 57-day-old layers in a chicken farm in Taigu District, and to determine its biological characteristics. A strain of coccidia was isolated and purified from the feces of dead chicken by saturated saline water floating method and single oocyst separation technique. After oral inoculation of the coccidia to SPF chickens, the shortest sporulation time, the parasitic position and size of the schizont, the parasitic position and size of the oocyst, the latent period, mortality, weight gain and the score of intestinal lesions were measured to assess the biological characteristics of the coccidia strain, and to preliminarily identify the type of the coccidia. Sequence characterized amplified region(SCAR) marker technology was used to determine its strain type. The results showed the schizonts parasitized the jejunum, with an average size of(63.17±7.66) μm×(47.12±6.91) μm, the oocysts were only visible in the cecum, with an average size of(19.92±0.20) μm×(19.27±0.11) μm, the latent period of the coccidia was 152 h, and the shortest sporulation time was 18 h. Inoculation of experimental chickens with 5.0×10~4 oocysts/feather and 1.1×10~5 oocysts/feather of the coccidia strain caused 40% and 50% mortality, the relative weight gain rate dropped to 31.61% and 20.87% compared to control chickens, the lesion score reached 3.9 and 4.0,respectively. SCAR marker amplification showed that the homology between the isolated coccidia and Eimeria necatrix(E. necatrix) was 97.1%,and the isolated coccidia strain was thus named E. necatrix-CN-SX-2018. The isolation of a highly pathogenic strain of E. necatrix not only enriches the collection of the epidemic strains of chicken coccidiosis in Shanxi Province, but also provides a valuable strain for the evaluation of chicken coccidiosis vaccines.