Toxoplasma gondii: isolation, biological and molecular characterisation of samples from free-range Gallus gallus domesticus from countryside Southeast Brazil

Abstract Toxoplasma gondii presents a high prevalence worldwide, infecting several animals. Felines are considered the definitive hosts and among the intermediate hosts we highlight mammals and birds. The man can become infected by ingesting tissue cysts present in birds and mammals. Biological and molecular aspects of T. gondii allows a better understanding of the epidemiology of toxoplasmosis. This work is a serologic screening of 58 chickens grown (Gallus gallus domesticus) for human consumption in Espírito Santo State, by means of indirect haemagglutination assay (IHA). Thirteen chickens tested positive for anti-T. gondii antibodies. The heart and brain of five positive chickens were harvested, treated with pepsin and inoculated separately, in two Swiss mice, intraperitoneally. Tachyzoites were observed in the peritoneum of all the animals, between seven and 10 days after the inoculum. Ten isolates were obtained and biologically characterised in BALB/c mice inoculated with 101 to 104 tachyzoites. All isolates were classified as virulent or intermediately virulent. Isolates were genotyped by means of polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis, revealing three different genotypes. None of the isolates exhibited the clonal type I, II or III genotype. No genotypic differences were observed between the isolates from the brain or heart from the same bird.

摘要 刚地弓形虫（Toxoplasma gondii）在全球范围内具有较高的流行率，可感染多种动物。猫科动物被认为是其终末宿主，中间宿主则以哺乳类和鸟类为主。人类可通过摄入携带于鸟类和哺乳类动物体内的组织包囊而感染该寄生虫。对刚地弓形虫生物学与分子层面的研究，有助于更深入地理解弓形虫病的流行病学特征。本研究对巴西圣埃斯皮里图州用于人类食用的58只家鸡（Gallus gallus domesticus）开展血清学筛查，检测方法为间接血凝试验（indirect haemagglutination assay, IHA）。其中13只家鸡的抗刚地弓形虫抗体检测呈阳性。研究人员采集其中5只抗体阳性家鸡的心脏与脑组织，经胃蛋白酶处理后，分别通过腹腔注射途径接种于两只瑞士小鼠体内。接种后7至10天，可在所有受试小鼠的腹膜中观察到速殖子（tachyzoites）。共获得10个弓形虫分离株，并在接种了10¹至10⁴个速殖子的BALB/c小鼠体内完成生物学特性鉴定。所有分离株均被归类为强毒株或中等毒力株。通过聚合酶链反应-限制性片段长度多态性（polymerase chain reaction-restriction fragment length polymorphism, PCR-RFLP）分析对分离株进行基因分型，结果显示存在3种不同的基因型。未发现任何分离株呈现经典的I型、II型或III型克隆基因型。同一受试家鸡的脑组织与心脏组织分离株之间未观察到基因型差异。