Laboratório de Micoplasmas, Departamento de Microbiologia, Instituto de Ciências Biomédicas II, Universidade de São Paulo, Brazil; Laboratório de Microbiologia e Imunologia, Nucleo de Tecnologia em Saúde, Instituto Multidisciplinar em Saúde, Universidade Federal da Bahia, Brazil.
Vet Microbiol. 2013 Dec 27;167(3-4):670-4. doi: 10.1016/j.vetmic.2013.07.031. Epub 2013 Aug 9.
Ureaplasma diversum in veterinary studies is an undesirable microbe, which may cause infection in bulls and may result in seminal vesiculitis, balanopostitis, and alterations in spermatozoids, whereas in cows, it may cause placentitis, fetal alveolitis, abortion, and birth of weak calves. U. diversum is released through organic secretions, especially semen, preputial and vaginal mucus, conjunctival secretion, and milk. The aim of the present study was to develop a TaqMan probe, highly sensitive and specific quantitative PCR (qPCR) assay for the detection and quantification of U. diversum from genital swabs of bovines. Primers and probes specific to U. diversum 16S rRNA gene were designed. The specificity, detection limit, intra- and inter-assay variability of qPCR to detect this ureaplasma was compared with the results of the conventional PCR assay (cPCR). Swabs of vaginal mucus from 169 cows were tested. The qPCR assay detected as few as 10 copies of U. diversum and was 100-fold more sensitive than the cPCR. No cross-reactivity with other Mollicutes or eubacteria was observed. U. diversum was detected in 79 swabs (46.42%) by qPCR, while using cPCR it was detected in 42 (25%) samples. The difference in cPCR and qPCR ureaplasma detection between healthy and sick animals was not statistically significant. But the U. diversum load in samples from animals with genital disorders was higher than in healthy animals. The qPCR assay developed herein is highly sensitive and specific for the detection and quantification of U. diversum in vaginal bovine samples.
在兽医研究中，解脲支原体是一种不理想的微生物，它可能会引起公牛感染，导致精囊炎、包皮龟头炎和精子改变，而在奶牛中，它可能会引起胎盘炎、胎儿肺炎、流产和弱仔出生。解脲支原体通过有机分泌物释放，特别是精液、包皮和阴道黏液、结膜分泌物和牛奶。本研究旨在开发一种 TaqMan 探针，用于检测和定量牛生殖器拭子中的解脲支原体，该探针具有高度敏感和特异性的定量 PCR（qPCR）检测方法。设计了针对解脲支原体 16S rRNA 基因的引物和探针。比较了 qPCR 检测这种脲原体的特异性、检测限、内和间试验变异性与常规 PCR 检测（cPCR）的结果。检测了 169 头奶牛的阴道黏液拭子。qPCR 检测的下限低至 10 个拷贝的解脲支原体，比 cPCR 灵敏 100 倍。未观察到与其他柔膜体或真细菌的交叉反应。qPCR 在 79 个拭子（46.42%）中检测到解脲支原体，而 cPCR 在 42 个（25%）样本中检测到。cPCR 和 qPCR 检测健康和患病动物的脲原体差异无统计学意义。但在患有生殖器官疾病的动物样本中，解脲支原体的负荷量高于健康动物。本文开发的 qPCR 检测方法高度敏感和特异性，可用于检测和定量牛阴道样本中的解脲支原体。
