以核酸檢測法應用於監測收容所無症狀犬貓小病毒感染情形

Detection of Asymptomatic Parvovirus-Infected Dogs and Cats in Animal Shelters Using a Molecular Assay

陳文德(Wen-Te Chen)

指導教授 ： 萬灼華

國立臺灣大學/獸醫專業學院/分子暨比較病理生物學研究所/碩士(2021年)

https://doi.org/10.6342/NTU202103853

摘要

犬細小病毒（CPV）和貓細小病毒（FPV）是犬和貓最常見的感染性病原，臨床前期感染的動物經常在進入動物收容所後引發致命的疾病爆發。由於商業化的現行檢測試劑可能不能有效和準確地檢測出CPV/FPV感染初期的動物。為了能及早敏感地檢測疾病，本研究針對CPV/FPV之VP基因與顯示檢體樣本DNA品質之housekeeping基因，製備多重引子聚合酶連鎖反應檢測法技術之試劑（CPV/FPV/Actin multiplex PCR assay），此檢測法能同時檢測出CPV和FPV感染動物，同時即使在10個基因拷貝組的CPV/FPV的低感染量也能夠敏感性的被檢測出來。本研究在檢測收容所的狗和貓的糞便時，無論是明顯的臨床症狀表現還是亞臨床狀態下的患病犬貓，多重引子聚合酶連鎖反應檢測試劑成功地鑑定出所有感染的犬貓。然而，五種常用來診斷細小病毒感染方面的商業化試劑套組，卻表現出明顯較低的敏感性，特別是在臨床症狀正常或有輕度/中度腹瀉的動物身上，其敏感度更大幅下降，商業化試劑套組在已感染細小病毒的狗的檢測敏感度為75.0% ，對貓的檢測敏感度則更降低為 37.5% - 50.0% 。因此，為了防止已感染動物，特別是臨床前期無症狀之感染動物進入動物收容所，本研究之多重引子聚合酶連鎖反應檢測試劑可成為一個強有力的工具，並且建議在所有動物進入庇護所之前都必須進行以多重引子聚合酶連鎖反應檢測試劑篩選出已感染犬貓並加以隔離，避免造成更多健康犬貓受到感染，另外再結合實驗動物常見之多樣本池化檢驗策略的情況下，便可達成高成本效益和高度可靠的方法實際應用於動物收容所內大量犬貓健康監測。

關鍵字

犬細小病毒 ； 貓細小病毒 ； 多重引子聚合酶連鎖反應檢測法技術 ； 多樣本池化檢驗策略 ； 亞臨床感染 ； 疾病疫情預防 ； 動物收容所醫學

並列摘要

Canine parvovirus (CPV) and feline parvovirus (FPV), the most prevalent infectious pathogens in dogs and cats, frequently induce lethal outbreaks in shelters through introducing the preclinical-stage infected animals. Commercialized in-house assays may not be able to effectively and accurately identify the canine parvovirus and feline parvovirus infections in early stage. In this study, a CPV/FPV/Actin multiplex PCR assay targeting the VP genes and a housekeeping gene (β-actin) was developed to simultaneously detect the canine parvovirus and feline parvovirus. The multiplex PCR assay was able to specifically detect CPV/FPV as low as 10 copies. In screening the feces of shelter dogs and cats, the multiplex PCR successfully identified all infected animals, in either overtly sick or subclinical condition. However, the three commonly-used commercialized kits exhibited distinctly lower sensitivity in diagnosing the parvoviral infections, especially in animals clinically normal or with mild/moderate diarrhea; their detection sensitivity was 75.0 % for dogs and 37.5-50.0 % for cats. Therefore, to prevent the infected animals, especially those preclinically infected ones, into shelters, the CPV/FPV/Actin multiplex PCR assay should be a powerful tool to screen all animals prior to their entering the shelters. In combining with the sample-pooling strategy, the multiplex PCR testing would be a cost-effective, practical, and highly reliable health monitoring method used in shelters.

並列關鍵字

Canine parvovirus ； feline parvovirus ； fecal multiplex PCR ； the sample-pooling strategy ； subclinical infection ； outbreak prevention ； shelter medicine

參考文獻

1. Abd-Eldaim, M.; Beall, M.J.; Kennedy, M.A. Detection of feline panleukopenia virus using a commercial ELISA for canine parvovirus. Veterinary Therapeutics 2009, 10, E1-6.

2. Addie, D.D.; Toth, S.; Thompson, H.; Greenwood, N.; Jarrett, J.O. Detection of feline parvovirus in dying pedigree kittens. Veterinary Record 1998, 142(14), 353-356.

3. Agbandje, M., McKenna, R., Rossmann, M. G., Strassheim, M. L., Parrish, C. R. Structure determination of feline panleukopenia virus empty particles. Proteins: Structure, Function, and Bioinformatics 1993, 16(2), 155-171.

4. Allison, A.B., Harbison, C.E., Pagan, I., Stucker, K.M., Kaelber, J.T., Brown, J.D., Ruder M. G., Keel M. K., Dubovi E. J., Holmes E. C., Parrish, C. R. Role of multiple hosts in the cross-species transmission and emergence of a pandemic parvovirus. Journal of Virology 2012, 86(2), 865-872.

5. Appel, M. J., Cooper, B. J., Greisen, H., Scott, F., Carmichael, L. E. Canine viral enteritis. I. Status report on corona- and parvo-like viral enteritides. The Cornell veterinarian 1979, 69(3), 123–133.