一种使用锁核酸（LNA）TaqMan 探针实时 RT-PCR 检测 1 型和 2 型 H5N1 流感病毒的方法。
A real-time RT-PCR for detection of clade 1 and 2 H5N1 influenza A virus using locked nucleic acid (LNA) TaqMan probes.
机构信息
Oxford University Clinical Research Unit, Hospital for Tropical Diseases, 190 Ben Ham Tu, Dist 05, Ho Chi Minh City, Viet Nam.
出版信息
Virol J. 2010 Feb 22;7:46. doi: 10.1186/1743-422X-7-46.
BACKGROUND
The emergence and co-circulation of two different clades (clade 1 and 2) of H5N1 influenza viruses in Vietnam necessitates the availability of a diagnostic assay that can detect both variants.
RESULTS
We developed a single real-time RT-PCR assay for detection of both clades of H5N1 viruses, directly from clinical specimens, using locked nucleic acid TaqMan probes. Primers and probe used in this assay were designed based on a highly conserved region in the HA gene of H5N1 viruses. The analytical sensitivity of the assay was < 0.5 PFU and 10-100 ssDNA plasmid copies. A total of 106 clinical samples (58 from patients infected with clade 1, 2.1 or 2.3 H5N1 viruses and 48 from uninfected or seasonal influenza A virus-infected individuals) were tested by the assay. The assay showed 97% concordance with initial diagnostics for H5 influenza virus infection with a specificity of 100%.
CONCLUSIONS
This assay is a useful tool for diagnosis of H5N1 virus infections in regions where different genetic clades are co-circulating.
背景
两种不同的 H5N1 流感病毒（1 型和 2 型）在越南同时出现并共同传播，这就需要有一种诊断检测方法，能够同时检测这两种变体。
结果
我们开发了一种单重实时 RT-PCR 检测方法，使用锁定核酸 TaqMan 探针，直接从临床标本中检测两种 H5N1 病毒。该检测方法中使用的引物和探针是根据 H5N1 病毒的 HA 基因中的高度保守区域设计的。该检测方法的分析灵敏度<0.5 PFU 和 10-100 ssDNA 质粒拷贝。该检测方法共检测了 106 份临床样本（58 份来自感染了 1 型、2.1 或 2.3 型 H5N1 病毒的患者，48 份来自未感染或感染季节性甲型流感病毒的个体）。该检测方法与 H5 流感病毒感染的初始诊断具有 97%的一致性，特异性为 100%。
结论
该检测方法是在不同遗传分支同时流行的地区诊断 H5N1 病毒感染的有用工具。
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