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首页> 外文期刊>Journal of Parasitology >SPECIFIC DETECTION OF NEOSPORA CANINUM OOCYSTS IN FECAL SAMPLES FROM EXPERIMENTALLY-INFECTED DOGS USING THE POLYMERASE CHAIN REACTION
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SPECIFIC DETECTION OF NEOSPORA CANINUM OOCYSTS IN FECAL SAMPLES FROM EXPERIMENTALLY-INFECTED DOGS USING THE POLYMERASE CHAIN REACTION

机译：聚合酶链反应从实验感染犬中特异性检测粪便中新孢子虫卵囊

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Neospora caninum oocysts, passed in the feces of a definitive host (dog), were isolated, and genomic DNA was extracted. A polymerase cahin reaction (PCR) targeting the N. caninum-specific Nc 5 genomic sequence was performed using the isolated DNA. A synthesized competitor molecule containing part of the Nc 5 sequence was included in the assay as a check against false-negative PCR results and to quantify N. caninum oocyst DNA in fecal samples. A standard curve of the ratio of fluorescence intensity of PCR-amplified competitor to that of oocyst DNA was constructed to compare oocyst equivalents from fecal samples containing unknown numbers of N. caninum oocysts and to assess the sensitivity of the assay. The specificity of the assay was determined using the Nc 5–specific primers in PCR assays against other parasites likely to be found in canine feces. Genomic DNA sequences from the canine coccidians Hammondia heydorni, Cryptosporidium parvum, Sarcocystis cruzi, S. tenella, and Isospora ohioensis and the canine helminth parasites Strongyloides stercoralis, Toxocara canis, Dipylidium caninum, and Ancylostoma caninum were not amplified. In addition, genomic DNA sequences from oocysts of coccidian parasites that might contaminate dog feces, such as Hammondia hammondi, Toxoplasma gondii, or Eimeria tenella, were not amplified in the PCR assay. The assay should be useful in epidemiological surveys of both domestic and wild canine hosts and in investigations of oocyst biology in experimental infections.
机译：分离到确定宿主（狗）粪便中的新孢子虫卵囊，并提取基因组DNA。使用分离的DNA进行针对犬新孢子虫特异性Nc 5基因组序列的聚合酶cahin反应（PCR）。包含部分Nc 5序列的合成竞争分子被包括在测定中，以检查假阴性PCR结果并定量粪便样品中的犬新孢子虫卵囊DNA。构建PCR扩增的竞争者与卵囊DNA的荧光强度之比的标准曲线，以比较来自粪便样本的卵囊当量，其中粪便样本中含有未知数量的犬新孢子虫卵囊，并评估测定的灵敏度。使用Nc 5特异性引物在PCR分析中针对犬粪便中可能发现的其他寄生虫，确定了分析的特异性。来自犬球虫Hammondia heydorni，小隐孢子虫，Sarcocystis cruzi，S。tenella和OH的Isospora寄生虫的基因组DNA序列以及犬蠕虫的寄生虫，如丝状线虫，Toxocara canis，Dipylidium caninum和Ancylostoma caninum。此外，PCR检验未扩增可能污染狗粪便的球虫寄生虫卵囊的基因组DNA序列，例如Hammondia hammondi，Toxoplasma gondii或Eimeria tenella。该测定法应可用于家犬和野生犬宿主的流行病学调查以及实验性感染中卵囊生物学的调查。

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