Cunha M M, Carneiro A C A V, Costa J G L, Vitor R W A
Departamento de Parasitologia, Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, 31270-901, Belo Horizonte, MG, Brazil.
J Parasitol. 2016 Feb;102(1):157-60. doi: 10.1645/15-813. Epub 2015 Sep 29.
Genotyping of Toxoplasma gondii is traditionally performed using DNA obtained from tachyzoites after isolation by bioassay in mice. In this study, genotyping of T. gondii was performed by multiplex nested polymerase chain reaction restriction fragment length polymorphism (Mn-PCR-RFLP) in DNA obtained from the lungs of experimentally infected mice, the hearts of naturally infected free-range chickens, and human blood samples of newborns with congenital toxoplasmosis. The efficiency of Mn-PCR varied according to the marker. We obtained complete genotypes of all of the mice lung samples. In chickens, total or partial genotyping was performed on all of the 15 samples. Two complete genotypes were obtained, including one identified for the first time, and another previously described in different hosts including dogs, cats, and humans. In blood from infants, partial genotypes were obtained in 8 of the 12 samples. Mouse bioassay is the most efficient method to obtain DNA from T. gondii , but direct tissue genotyping enhances the likelihood of obtaining molecular information on T. gondii and is an effective tool as a complement to isolation in mice. In this study, we genotyped Toxoplasma gondii directly from human (blood samples of newborns with congenital toxoplasmosis) and free-range chickens (hearts) by Mn-PCR-RFLP. We present partial and complete genotypes and provide technical and scientific information about T. gondii genotyping methods.
传统上，通过在小鼠体内进行生物测定分离速殖子后，利用从速殖子中获得的DNA对弓形虫进行基因分型。在本研究中，通过多重巢式聚合酶链反应-限制性片段长度多态性（Mn-PCR-RFLP）对从实验感染小鼠的肺、自然感染的散养鸡的心脏以及先天性弓形虫病新生儿的人类血液样本中获得的DNA进行弓形虫基因分型。Mn-PCR的效率因标记物而异。我们获得了所有小鼠肺样本的完整基因型。在鸡中，对所有15个样本进行了全部或部分基因分型。获得了两种完整基因型，其中一种是首次鉴定出来的，另一种是先前在包括狗、猫和人类在内的不同宿主中描述过的。在婴儿血液中，12个样本中有8个获得了部分基因型。小鼠生物测定是从弓形虫获取DNA的最有效方法，但直接对组织进行基因分型增加了获得弓形虫分子信息的可能性，并且是作为小鼠分离补充的有效工具。在本研究中，我们通过Mn-PCR-RFLP直接对人类（先天性弓形虫病新生儿的血液样本）和散养鸡（心脏）中的弓形虫进行基因分型。我们展示了部分和完整基因型，并提供了有关弓形虫基因分型方法的技术和科学信息。
