Influenza A virus (IAV) subtypes H1N1, H1N2, and H3N2 are endemic in swine herds in most pork producing countries; however, the viruses circulating in different geographic regions are antigenically and genetically distinct. In this sense, the availability of a rapid diagnostic assay to detect locally adapted IAVs and discriminate the virus subtype in clinical samples from swine is extremely important for monitoring and control of the disease. This study describes the development and validation of a multiplex RT-PCR assay for detection and subtyping of IAV from pigs. The analytical and diagnostic specificity of the assays was 100% (94.3–100.0, CI 95%), and the limit of detection was 10 −3 TCID 50 /mL. A total of 100 samples (IAV isolates and clinical specimens) were tested, and the virus subtype was determined for 80 samples (80%; 71.1–86.7, CI 95%). From these, 50% were H1N1, 22.5% were H1N2, and 7.5% were H3N2. Partial subtyping was determined for 8.75% samples (H1pdmNx and HxN2). Additionally, mixed infections with two virus subtypes (H1N2 + H3N2 and H1N1pdm + H1pdmN2; 2.5%) and reassortant viruses (H1pdmN2, 6.25%; and H1N1hu, 2.5%) were detected by the assay. A rapid detection of the most prevalent IAV subtypes and lineages in swine is provided by the assays developed here, improving the IAV diagnosis in Brazilian laboratories, and contributing to the IAV monitoring.

中文翻译：

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巴西猪流感病毒检测和亚型区分的两种多重 RT-PCR 检测方法的评价

甲型流感病毒 (IAV) 亚型 H1N1、H1N2 和 H3N2 在大多数猪肉生产国的猪群中流行；然而，在不同地理区域传播的病毒在抗原和基因上是不同的。从这个意义上说，可用于检测局部适应的 IAV 并区分猪临床样本中的病毒亚型的快速诊断分析的可用性对于监测和控制疾病极为重要。本研究描述了用于检测和分型猪 IAV 的多重 RT-PCR 检测方法的开发和验证。测定的分析和诊断特异性为100%（94.3-100.0，CI 95%），检测限为10 -3 TCID 50 /mL。共检测了 100 个样本（IAV 分离株和临床标本），确定了 80 个样本的病毒亚型（80%；71.1-86.7，置信区间 95%）。其中，50% 是 H1N1，22.5% 是 H1N2，7.5% 是 H3N2。确定了 8.75% 样品（H1pdmNx 和 HxN2）的部分亚型。此外，该测定还检测到两种病毒亚型（H1N2 + H3N2 和 H1N1pdm + H1pdmN2；2.5%）和重组病毒（H1pdmN2，6.25%；和 H1N1hu，2.5%）的混合感染。这里开发的检测方法可以快速检测猪中最普遍的 IAV 亚型和谱系，改善巴西实验室的 IAV 诊断，并有助于 IAV 监测。