Abad F X, Pintó R M, Villena C, Gajardo R, Bosch A
Department of Microbiology, University of Barcelona, Spain.
Appl Environ Microbiol. 1997 Aug;63(8):3119-22. doi: 10.1128/aem.63.8.3119-3122.1997.
A method based on infection of CaCo-2 cultured cell monolayers (CC) and reverse transcription-PCR (RT-PCR) was developed for the specific detection of infectious astrovirus. The procedure was validated by titrating poliovirus stocks in parallel in CaCo-2 cells by determining the most probable number of cytopathogenic units and by cell culture and subsequent RT-PCR (CC-RT-PCR). CC-RT-PCR was then employed to measure the persistence of astrovirus suspended in dechlorinated tap water. After 60 days, the decay of astrovirus infectivity was 2 log units at 4 +/- 1 degrees C and 3.2 log units at 20 +/- 1 degrees C, while after 90 days, the titer reduction was 3.3 and 5 log units at 4 +/- 1 degrees C and 20 +/- 1 degrees C, respectively. Astrovirus decay in the presence of free chlorine (FC) was monitored by CC-RT-PCR. Residual infectivity was found after 2 h in the presence of 1 mg of FC/liter. Under these conditions, astrovirus shows a log titer reduction (LTR) or 4, while 0.5 mg of FC/liter induced an LTR of 2.4. The possibility of acquiring data on the survival of fastidious viruses in the environment opens new perspectives on the epidemiology of some significant infections transmitted by the fecal-oral route.
开发了一种基于感染培养的CaCo-2细胞单层(CC)和逆转录聚合酶链反应(RT-PCR)的方法,用于特异性检测传染性星状病毒。通过在CaCo-2细胞中平行滴定脊髓灰质炎病毒储备液,通过确定细胞病变单位的最可能数量以及通过细胞培养和随后的RT-PCR(CC-RT-PCR)来验证该程序。然后使用CC-RT-PCR来测量悬浮在去氯自来水中的星状病毒的持久性。60天后,星状病毒感染力在4±1℃时下降2个对数单位,在20±1℃时下降3.2个对数单位,而90天后,在4±1℃和20±1℃时滴度分别降低3.3和5个对数单位。通过CC-RT-PCR监测游离氯(FC)存在下星状病毒的衰减。在存在1mg FC/升的情况下2小时后发现有残余感染力。在这些条件下,星状病毒的对数滴度降低(LTR)为4,而0.5mg FC/升诱导的LTR为2.4。获取有关挑剔病毒在环境中存活数据的可能性为一些通过粪-口途径传播的重要感染的流行病学开辟了新的前景。