Xue Yuan Chao, Bertsch Jennifer, Monacy Kaylin, Haynes Carter, Williams-Bouyer Natalie, Judy Barbara M, Newman Patrick C, Ksiazek Thomas G, Marushchak Lyudmyla V, Gray Gregory C, Ren Ping
Department of Pathology, University of Texas Medical Branch, Galveston, Texas, USA.
Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, Texas, USA.
J Clin Microbiol. 2025 Aug 13;63(8):e0068125. doi: 10.1128/jcm.00681-25. Epub 2025 Jul 18.
The increasing transmission of highly pathogenic avian influenza (HPAI) H5N1 from animals to humans underscores the urgent need for enhanced diagnostic capabilities in clinical microbiology laboratories. Although analysis suggests that commercial multiplex respiratory panels can detect H5N1, these assays lack subtyping H5N1 capabilities, and their real-world performance remains largely unverified. In this study, we evaluated the limit of detection (LoD) for H5N1 using three commercial molecular diagnostic assays routinely employed at our institution: bioMérieux BioFire Respiratory 2.1 Panel, Cepheid Xpert Xpress CoV-2/Flu/RSV Plus, and Hologic Panther Fusion SARS-CoV-2/Flu A/B/RSV assays. All three reliably detected H5N1 were at low viral concentrations. To enable H5N1 detection, we also developed a real-time RT-PCR H5 single-plex assay on the Hologic Panther Fusion Open Access platform for reflex testing of influenza A-positive specimens. In conclusion, although current commercial assays lack influenza A H5 subtype differentiation, our validation data provide critical performance information. When integrated with a targeted H5 assay, these tools can enhance clinical decision-making and public health surveillance by reducing the risk of missed H5N1 infection cases.IMPORTANCEThis study addresses a growing public health concern: the spread of bird flu (H5N1) from animals to humans. Most hospital laboratories use commercial tests to detect respiratory viruses like the flu, but these tests cannot tell if someone has the specific and more dangerous H5N1 strain. To help solve this, we tested three commonly used diagnostic tools and found that they can detect H5N1 even at low levels. However, since they cannot identify the specific H5 subtype, we also developed and validated a follow-up test that runs on one of the existing laboratory high-throughput equipment. This test can confirm whether a patient infected with the flu has the H5N1 strain. By combining these tools, hospital laboratories can improve early detection of H5N1, support better patient care, and help public health officials respond more effectively to outbreaks.
高致病性禽流感（HPAI）H5N1从动物向人类的传播日益增加，凸显了临床微生物学实验室增强诊断能力的迫切需求。尽管分析表明，商用多重呼吸道检测板可检测出H5N1，但这些检测方法缺乏对H5N1进行亚型分型的能力，其在实际应用中的性能仍 largely unverified。在本研究中，我们使用了本机构常规使用的三种商用分子诊断检测方法，评估了H5N1的检测限（LoD）：bioMérieux BioFire Respiratory 2.1检测板、Cepheid Xpert Xpress CoV-2/Flu/RSV Plus以及Hologic Panther Fusion SARS-CoV-2/Flu A/B/RSV检测方法。所有这三种方法均能在低病毒浓度下可靠地检测出H5N1。为实现H5N1的检测，我们还在Hologic Panther Fusion开放获取平台上开发了一种实时逆转录聚合酶链反应（RT-PCR）H5单重检测方法，用于对甲型流感阳性标本进行复检。总之，尽管目前的商用检测方法缺乏对甲型流感H5亚型的区分能力，但我们的验证数据提供了关键的性能信息。当与靶向H5检测方法相结合时，这些工具可通过降低漏诊H5N1感染病例的风险，加强临床决策和公共卫生监测。
重要性
禽流感（H5N1）从动物向人类的传播。大多数医院实验室使用商用检测方法来检测流感等呼吸道病毒，但这些检测方法无法判断某人是否感染了特定且更危险的H5N1毒株。为解决这一问题，我们测试了三种常用的诊断工具，发现它们即使在低水平下也能检测出H5N1。然而，由于它们无法识别特定的H5亚型，我们还开发并验证了一种后续检测方法，该方法可在现有的实验室高通量设备之一上运行。此检测方法可确认感染流感的患者是否感染了H5N1毒株。通过结合这些工具，医院实验室可改善对H5N1的早期检测，支持更好的患者护理，并帮助公共卫生官员更有效地应对疫情爆发。
