Lu Xiaoyan, Erdman Dean D
Gastroenteritis and Respiratory Viruses Laboratory Branch, Division of Viral Diseases, Centers for Disease Control and Prevention, Atlanta, GA, USA.
Gastroenteritis and Respiratory Viruses Laboratory Branch, Division of Viral Diseases, Centers for Disease Control and Prevention, Atlanta, GA, USA.
J Virol Methods. 2016 Nov;237:174-178. doi: 10.1016/j.jviromet.2016.05.020. Epub 2016 Jun 27.
Human adenoviruses (HAdVs) are medically important respiratory pathogens. Among the 7 recognized species (A-G), species C HAdVs (serotypes 1, 2, 5 and 6) are globally endemic and infect most people early in life. Species C HAdV infections are most often subclinical or mild and can lead to persistent shedding from the gastrointestinal and upper respiratory tracts. They can also cause severe disseminated disease in newborn and immunocompromised persons, where rapid and quantitative detection and identification of the virus would help guide therapeutic intervention. To this end, we developed quantitative type-specific real-time PCR (qPCR) assays for HAdV-1, -2, -5 and -6 targeting the HAdV hexon gene. All type-specific qPCR assays reproducibly detected as few as 5 copies/reaction of quantified hexon recombinant plasmids with a linear dynamic range of 8 log units (5-5×10 copies). No non-specific amplifications were observed with concentrated nucleic acid from other HAdV types or other common respiratory pathogens. Of 199 previously typed HAdV field isolates and positive clinical specimens, all were detected and correctly identified to type by the qPCR assays; 10 samples had 2 HAdV types and 1 sample had 3 types identified which were confirmed by amplicon sequencing. The species C HAdV qPCR assays permit rapid, sensitive, specific and quantitative detection and identification of four recognized endemic HAdVs. Together with our previously developed qPCR assays for the epidemic respiratory HAdVs, these assays provide a convenient alternative to classical typing methods.
人腺病毒(HAdVs)是具有重要医学意义的呼吸道病原体。在7种已确认的病毒种类(A - G)中,C种人腺病毒(血清型1、2、5和6)在全球流行,大多数人在幼年时就会感染。C种人腺病毒感染大多为亚临床感染或症状轻微,可导致胃肠道和上呼吸道持续排毒。它们还可在新生儿和免疫功能低下者中引起严重的播散性疾病,快速、定量地检测和鉴定病毒有助于指导治疗干预。为此,我们针对人腺病毒1、2、5和6,开发了靶向人腺病毒六邻体基因的定量型特异性实时荧光定量聚合酶链反应(qPCR)检测方法。所有型特异性qPCR检测方法均能重复检测到低至5拷贝/反应的定量六邻体重组质粒,线性动态范围为8个对数单位(5 - 5×10拷贝)。未观察到来自其他型人腺病毒或其他常见呼吸道病原体的浓缩核酸产生非特异性扩增。在199份先前已分型的人腺病毒临床分离株和阳性临床标本中,所有标本均通过qPCR检测方法被检测到并正确分型;10份样本检测到2种人腺病毒型别,1份样本检测到3种型别,经扩增子测序确认。C种人腺病毒qPCR检测方法可对四种公认的流行人腺病毒进行快速、灵敏、特异和定量的检测与鉴定。连同我们先前开发的针对流行性呼吸道人腺病毒的qPCR检测方法,这些检测方法为传统分型方法提供了一种便捷的替代方法。