Mycobacterium avium subspecies paratuberculosis (MAP), the cause of Johne disease, is a slow growing mycobacterium. Viable MAP detection is difficult, inconstant and time-consuming. The purpose of this study was to compare a rapid phage/qPCR assay performed on peripheral blood mononuclear cells (PBMCs) with three standard methods of MAP detection: fecal MAP PCR; plasma antigen-specific IFN-γ & serum MAP ELISA hypothesizing that, if sensitive and specific, Johne animals would be positive and Control animals negative. We studied a well characterized herd of Holstein cattle that were naturally infected with MAP and their Controls. With phage/qPCR 72% (23/32) of Johne and 35% (6/17) of Controls were MAP positive. With fecal PCR 75% (24/32) of Johne and 0% (0/17) of Controls were MAP positive. With plasma antigen-specific IFN-γ 69% (22/32) of Johne and 12% (2/17) of Controls were MAP positive. With serum MAP ELISA, 31% (10/32) of Johne and 0% (0/17) of Controls were MAP positive. When phage / qPCR and fecal PCR results were combined, 100% (32/32) Johne and 35% (6/17) of Control animals were MAP positive. Younger Control animals (1–3 years) had significantly fewer plaques (25 ± 17 SEM) than older Controls (4–12 years) (309 ± 134 p = 0.04). The same trend was not observed in the Johne animals (p = 0.19). In contrast to our hypothesis, using the phage/qPCR assay we find that viable circulating MAP can rapidly be detected from the blood of animals infected with, as well as those in the Control group evidently colonized by MAP. These data indicate that the presence of viable MAP in blood does not necessarily signify that an animal must of necessity be demonstrably ill or be MAP positive by standard diagnostic methods.

中文翻译：

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牛分枝杆菌噬菌体检测法与标准诊断方式在自然感染约翰内氏病牛中检测活鸟分枝杆菌副结核亚种及标准诊断方法的比较

禽分枝杆菌亚种副结核病（MAP）是约翰内氏病的病因，是一种生长缓慢的分枝杆菌。可行的MAP检测非常困难，持续且耗时。这项研究的目的是将对外周血单核细胞（PBMC）进行的快速噬菌体/ qPCR分析与MAP检测的三种标准方法进行比较：粪便MAP PCR；粪便MAP PCR；粪便MAP PCR；粪便MAP PCR。血浆抗原特异性IFN-γ和血清MAP ELISA假设，如果敏感和特异，则Johne动物将为阳性，而对照动物为阴性。我们研究了特征鲜明的荷斯坦牛群，它们自然被MAP及其对照感染。使用噬菌体/ qPCR，72％（23/32）的约翰和35％（6/17）的对照为MAP阳性。通过粪便PCR，75％（24/32）的Johne和0％（0/17）的对照为MAP阳性。使用血浆抗原特异性IFN-γ时，有69％（22/32）的Johne和12％（2/17）的对照为MAP阳性。使用血清MAP ELISA，有31％（10/32）的Johne和0％（0/17）的对照为MAP阳性。当将噬菌体/ qPCR和粪便PCR结果相结合时，100％（32/32）的Johne和35％（6/17）的对照动物的MAP阳性。较年轻的对照动物（1-3年）的斑块（25±17 SEM）明显少于较早的对照动物（4-12岁）（309±134 p = 0.04）。在Johne动物中未观察到相同的趋势（p = 0.19）。与我们的假设相反，使用噬菌体/ qPCR分析，我们发现可以从感染了动物的动物的血液以及被MAP明显克隆的对照组的血液中快速检测到可行的循环MAP。