Logan T M, Linthicum K J, Moulton J R, Ksiazek T G
Disease Assessment Division, U.S. Army Medical Research Institute of Infectious Diseases, Fort Detrick, Frederick, MD 21702-5011.
J Virol Methods. 1993 Apr;42(1):33-44. doi: 10.1016/0166-0934(93)90174-p.
A viral antigen-capture ELISA was compared to a viral plaque-assay on human cell monolayers for detection and quantification of Crimean-Congo hemorrhagic fever virus in triturated experimentally infected Hyalomma truncatum ticks. In suspensions of ticks exposed as larvae to viremic mice, the ELISA detected 13% positive as compared to 3% (n = 721) positive by plaque-assay. Adult ticks inoculated with virus and sampled up to 102 days later were 84% positive by ELISA compared to 36% (n = 273) positive by plaque-assay. The two tests detected similar proportions of positive ticks in the two weeks immediately after viral inoculation; however, the ELISA was positive in 100% of inoculated adult ticks from 18-102 days post-inoculation while the plaque-assay was positive in 33% (n = 135) of the same specimens. CCHF viral antigen was detected in 10% (n = 101) of first-generation progeny tested as unfed larva pools by ELISA, yet no virus was detected by plaque-assay, indicating that either non-infective viral fragments or very low levels of live virus were detected by ELISA in these tick progeny. As detected by plaque assay, virus inoculated unfed adult ticks were virtually all infected by day 8 post-inoculation; by day 21 post-inoculation only 33% were detected as positive. A cohort of these ticks were allowed to blood feed from day 21-31 post-inoculation. When assayed after feeding all female ticks and nearly 50% of male ticks were detected as virus-positive. This indicates that the virus likely persisted in the unfed ticks below the level of detectability of the plaque-assay and increased in the blood fed ticks up to a detectable level. The ELISA however, detected 100% of ticks as virus-positive from day 14 post-inoculation throughout the remainder of the study, regardless of feeding status (day 102 post-inoculation). These results indicate that antigen-detection ELISA is more sensitive in detecting CCHF virus in ticks than plaque-assay. Since an infected tick remains antigen-positive by ELISA for possibly the remainder of its life, this assay will be a major improvement in field surveys and vector competency studies of ticks for CCHF virus.
将一种病毒抗原捕获酶联免疫吸附测定（ELISA）与在人细胞单层上进行的病毒蚀斑测定相比较，以检测和定量研磨后的实验感染的残缘璃眼蜱中的克里米亚-刚果出血热病毒。在幼虫期暴露于病毒血症小鼠的蜱悬液中，ELISA检测出13%为阳性，而蚀斑测定法检测出3%为阳性（n = 721）。接种病毒并在长达102天后取样的成年蜱，ELISA检测出84%为阳性，而蚀斑测定法检测出36%为阳性（n = 273）。在病毒接种后的两周内，两种检测方法检测到的阳性蜱比例相似；然而，在接种后18 - 102天，ELISA检测接种的成年蜱100%为阳性，而同一标本的蚀斑测定法检测出33%为阳性（n = 135）。通过ELISA检测，在作为未进食幼虫池检测的第一代后代中，10%（n = 101）检测到克里米亚-刚果出血热病毒抗原，但蚀斑测定法未检测到病毒，这表明在这些蜱后代中，ELISA检测到的可能是非感染性病毒片段或极低水平的活病毒。通过蚀斑测定法检测，接种病毒的未进食成年蜱在接种后第8天几乎全部被感染；接种后第21天，只有33%被检测为阳性。让一组这些蜱在接种后第21 - 31天吸血。吸血后检测发现，所有雌性蜱和近50%的雄性蜱为病毒阳性。这表明病毒可能在未进食的蜱中以低于蚀斑测定法可检测水平持续存在，并在吸血蜱中增加到可检测水平。然而，ELISA在接种后第14天至研究剩余时间内检测到100%的蜱为病毒阳性，无论其进食状态如何（接种后第102天）。这些结果表明，抗原检测ELISA在检测蜱中的克里米亚-刚果出血热病毒方面比蚀斑测定法更敏感。由于受感染的蜱通过ELISA可能在其余生中保持抗原阳性，该检测方法将在蜱的克里米亚-刚果出血热病毒现场调查和媒介能力研究中带来重大改进。
