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首页> 外文期刊>Aquaculture Reports >Evaluation of a reliable non-invasive molecular test for the diagnosis of the causative agent of acute hepatopancreatic necrosis disease of shrimp
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Evaluation of a reliable non-invasive molecular test for the diagnosis of the causative agent of acute hepatopancreatic necrosis disease of shrimp

机译：可靠的非侵入性分子检测对虾急性肝胰腺坏死病病原诊断的评估

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Acute hepatopancreatic necrosis disease (AHPND, also known as early mortality syndrome, EMS) has caused substantial mortality, up to 100%, in populations of penaeid shrimp cultured in SE Asia and in Latin America. The disease is caused by the bacterium Vibrio parahaemolyticus, which secretes binary toxins (PirA"v"p and PirB"v"p) resulting in the deterioration of the hepatopancreas tissue of infected shrimp. Diagnosis, screening, and monitoring of AHPND in shrimp populations involve sacrificing individuals to obtain tissue samples. This sampling method is undesirable when applied to valuable populations of broodstock. Here, we evaluated a non-invasive diagnostic method based on shrimp fecal samples that are analyzed by PCR. Small groups of Pacific white shrimp Penaeus vannamei were exposed to low levels of AHPND-bacteria and their feces were collected prior to any mortality observed (in the bioassays #1 and #2). Two protocols were evaluated. In one, DNA extracted from the fecal samples was directly analyzed by PCR. In the other, the fecal samples were cultured in TSB+ for 6h to enrich the bacterial populations, then the enriched bacterial broth was used for PCR analyses. Our results showed that the presence of V. parahaemolyticus could be detected both in fecal DNA samples and in the enriched bacterial broth, but the bands from the bacterial broth showed stronger amplification than the DNA; 12 strong positive in the enriched bacterial broth, but only 7 strong positive in the fecal DNA samples. Also, the AHPND bacteria present in the feces is infectious, determined by a bioassay of feeding specific pathogen free indicator shrimp with AHPND-feces (in the bioassay #3), and this proves that the AHPND can be transmitted through a fecal-oral route.
机译：在东南亚和拉丁美洲养殖的对虾虾种群中，急性肝胰腺坏死病（AHPND，也称为早期死亡率综合症，EMS）已导致相当大的死亡率，高达100％。该疾病是由副溶血性弧菌引起的，该细菌分泌二元毒素（PirA“ v” p和PirB“ v” p），导致被感染虾的肝胰脏组织变质。对虾种群中AHPND的诊断，筛选和监测涉及牺牲个体以获得组织样本。当将其应用于有价值的亲鱼种群时，这种采样方法是不可取的。在这里，我们评估了基于虾粪便样本的非侵入性诊断方法，该样本通过PCR分析。太平洋白虾南美白对虾的小群暴露于低水平的AHPND细菌，在观察到任何死亡率之前收集了它们的粪便（在生物测定1和2中）。评价了两种方案。一种是从粪便样品中提取的DNA通过PCR直接分析。另一方面，将粪便样品在TSB +中培养6小时以富集细菌种群，然后将富集的细菌肉汤用于PCR分析。我们的结果表明，在粪便DNA样品和富集的细菌肉汤中均可检测到副溶血性弧菌，但细菌肉汤中的条带显示出比DNA更强的扩增。在富集的细菌肉汤中有12个强阳性，但在粪便DNA样品中只有7个强阳性。此外，粪便中存在的AHPND细菌具有传染性，这是通过对特定的无病原体指示虾饲喂AHPND粪便的生物测定法确定的（生物测定法3），这证明AHPND可以通过粪便-口腔途径传播。

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