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Quantification of Malassezia pachydermatis by real-time PCR in swabs from the external ear canal of dogs

机译:实时PCR定量狗外耳道拭子中的马拉色霉菌

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摘要

is part of the normal microbiota of canine skin and external ear canal, and is also associated with otitis externa in dogs. Laboratory detection of otitis relies on the presence of elevated numbers of the yeast on cytologic examination of otic exudate. Although cytology has high specificity, it has low sensitivity, resulting in false-negatives and posing a challenge for clinicians to accurately diagnose otitis. We developed a quantitative PCR (qPCR) to detect and quantify yeasts and validate the method with swabs from external ear canals of dogs. Our qPCR uses the β-tubulin gene, a single-copy gene, as a target. The limit of quantification was established as 0.18 ng/reaction, equivalent to 2.0 × 10 genome equivalents (gEq). Swabs from healthy dogs yielded quantification values of ≤2.7 × 10  gEq in the qPCR, whereas swabs from dogs with otitis yielded quantification values of ≥2.5 × 10  gEq. Our qPCR assay provides accurate quantification of yeasts from swab samples from dogs, is more sensitive than cytology, and could be used to monitor response to treatment. Our assay could also be valuable in a research setting to better understand the pathogenesis of .
机译:是犬皮肤和外耳道正常微生物群的一部分,并且还与狗的中耳炎有关。耳炎的实验室检测取决于对耳分泌液进行细胞学检查时酵母菌数量增加。尽管细胞学具有高特异性,但敏感性低,导致假阴性,并且对临床医生准确诊断中耳炎构成了挑战。我们开发了定量PCR(qPCR)来检测和定量酵母,并用狗外耳道的拭子验证该方法。我们的qPCR使用单拷贝基因β-微管蛋白基因作为靶标。定量限为0.18μng/反应,相当于2.0×10基因组当量(gEq)。健康犬的拭子在qPCR中的定量值为≤2.7×10 gEq,而中耳炎犬的拭子的定量值为≥2.5×10 gEq。我们的qPCR分析可准确定量狗拭子样本中的酵母,比细胞学更敏感,可用于监测对治疗的反应。我们的测定方法在更好地理解肝炎的发病机制中可能也有价值。

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