针对常见和新兴猪呼吸道病原体的靶向下一代测序小组的分析和诊断验证
Analytic and Diagnostic Validation of a Targeted Next-Generation Sequencing Panel for Common and Emerging Swine Respiratory Pathogens.
摘要(中文译文)
呼吸道疾病仍然是美国养猪业中代价最高的挑战之一，并且常与多种微生物混合感染相关。常规qPCR检测具有高度敏感性，但多重检测能力有限，且通常无法提供用于病原体表征的测序信息。我们假设，靶向下一代测序（tNGS） panel 能够在保持临床相关敏感性的同时，实现对猪呼吸道病原体的广泛、同步检测。我们开发了一种多重 Ion Torrent tNGS panel，并使用20份系列稀释的qPCR阳性临床样本和合成 gBlock 对照进行了分析验证，随后使用25份qPCR阳性和25份qPCR阴性呼吸道样本进行了诊断验证。大多数靶标在临床相关病原体浓度范围内均可被检出。猪放线杆菌引物显示非特异性扩增，猪链球菌血清分型在临床样本中无法稳定实现，而猪繁殖与呼吸综合征病毒分型仅限于区分北美和欧洲基因型。与常规qPCR的诊断一致性较高（Cohen's κ = 0.84），尽管对低丰度靶标的敏感性有所下降。该检测方法检出了混合感染以及常规qPCR panel 之外的额外病原体。这些发现支持将tNGS作为猪呼吸道疾病的补充诊断和监测工具。
Abstract
Respiratory disease remains one of the mostly costly challenges in the U.S. swine industry and is frequently associated with polymicrobial infections. Routine qPCR assays are highly sensitive but are limited in multiplexing capacity and generally do not provide sequencing information for pathogen characterization. We hypothesized that a target next-generation sequencing (tNGS) panel could provide the broad, simultaneous detection of swine respiratory pathogens while preserving clinically relevant sensitivity. A multiplex Ion Torrent tNGS panel was developed and analytically validated using 20 serially diluted qPCR-positive clinical samples and synthetic gBlock controls, followed by diagnostic validation with 25 qPCR positive and 25 qPCR negative respiratory samples. Most targets were detected across clinically relevant pathogens concentrations. Actinobacillus suis primers showed nonspecific amplification, streptococcus suis serotyping was not consistently achievable in clinical samples, and porcine reproductive and respiratory syndrome virus typing was limited to distinguishing North American and European genotypes. Diagnostic agreement with routine qPCR was high (Cohen's κ = 0.84), although sensitivity decreased for low-abundance targets. The assay detected mixed infections and additional organisms outside routine qPCR panels. These findings support tNGS as a complementary diagnostic and surveillance tool for swine respiratory disease.
如何引用
AMA
Nelly O Elshafie, Rebecca P Wilkes. Analytic and Diagnostic Validation of a Targeted Next-Generation Sequencing Panel for Common and Emerging Swine Respiratory Pathogens.. Microorganisms. 2026; doi:10.3390/microorganisms14051159.
APA
Nelly O Elshafie, Rebecca P Wilkes (2026). Analytic and Diagnostic Validation of a Targeted Next-Generation Sequencing Panel for Common and Emerging Swine Respiratory Pathogens.. Microorganisms. https://doi.org/10.3390/microorganisms14051159
GB/T 7714
Nelly O Elshafie, Rebecca P Wilkes. Analytic and Diagnostic Validation of a Targeted Next-Generation Sequencing Panel for Common and Emerging Swine Respiratory Pathogens.[J]. Microorganisms, 2026 doi:10.3390/microorganisms14051159.
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