Mosquito-borne diseases pose ongoing global health concerns, demanding more cost-efficient methods to detect pathogens to support enhanced surveillance efforts. This study introduces an adapted TRIzol-based high-throughput RNA extraction protocol, tailored for the detection of California serogroup viruses in pooled mosquito samples in a rapid and cost-effective manner. This approach provided consistent RNA yields and sensitive viral detection relative to two commercial extraction kits (QIAGEN RNeasy Mini Kit and MACHEREY–NAGEL NucleoSpin RNA Plus Kit). The incorporation of a user-friendly and non-spiking-based RT-qPCR internal control designed for the 18S rRNA gene in mosquitoes minimizes potential false positives/negatives, improving the fidelity of viral detection outcomes. Effective RNA yields, purity, and successful target amplification across 25 mosquito species and varied pool sizes (1–50 mosquitoes per tube) affirm the reliability of our approach. The extraction method is cost-effective, with an incurred cost of $0.58 CAD per sample, in contrast to the $5.25 CAD cost per sample of the two kits, rendering it promising for mosquito-borne disease surveillance initiatives.

蚊媒疾病引起了持续的全球健康问题，需要更具成本效益的方法来检测病原体，以支持加强监测工作。本研究引入了一种基于 TRIzol 的高通量 RNA 提取方案，专为快速且经济高效地检测混合蚊子样本中的加州血清群病毒而定制。相对于两种商业提取试剂盒（QIAGEN RNeasy Mini Kit 和 MACHEREY-NAGEL NucleoSpin RNA Plus Kit），这种方法提供了一致的 RNA 产量和灵敏的病毒检测。结合专为蚊子 18S rRNA 基因设计的用户友好且基于非尖峰的 RT-qPCR 内部对照，最大限度地减少潜在的假阳性/阴性，提高病毒检测结果的保真度。 25 种蚊子和不同池大小（每管 1-50 只蚊子）的有效 RNA 产量、纯度和成功的目标扩增证实了我们方法的可靠性。该提取方法具有成本效益，每个样本的成本为 0.58 加元，而两个试剂盒每个样本的成本为 5.25 加元，这使其在蚊媒疾病监测方面很有前景。