台灣南部地區牛隻隱孢子蟲感染之盛行率、基因型別及病原性角色

Prevalence, Genotyping and Pathogenic Role of Cattle Cryptosporidium Infection in Southern Taiwan.

曾巨羽(Chu-Yu Tseng)

指導教授 ： 吳永惠

國立屏東科技大學/農學院/獸醫學系所/碩士(2008年)

https://doi.org/10.6346/NPUST.2008.00244

摘要

為評估台灣地區乳牛場隱孢子蟲感染之發生情形，於2006年8月至2007年1月，首先對台灣南部3縣24場228頭乳牛採其糞便進行修飾式抗酸染色法 (modified acid-fast stain of Ziehl-Neelsen method, MZN法)及巢氏聚合酶鏈鎖反應 (nested polymerase chain reaction, nested PCR法)，檢查有無隱孢子蟲感染，並對7場MZN陽性場之7株nested PCR陽性產物進行定序來與GenBank所發表之隱孢子蟲基因型別比對，結果隱孢子蟲感染之檢出率在nested PCR法為28.9 % (66/228)，顯著 (P<0.05)高於MZN法之7.0 % (16/228)；場陽性率為62.5 % (15/24)，牛隻陽性率在nested PCR和MZN法分別為28.9 % (66/228)和7.0 % (16/228)；感染場數比率和感染牛隻頭數比率在三縣間均無顯著差異 (P>0.05)；然而小於1月齡犢牛組之感染率顯著高於 (P<0.05)其他較大年齡組別之牛隻；下痢牛之隱孢子蟲陽性率顯著 (P<0.05)高於無下痢牛，反之糞中出現卵囊之牛隻，其下痢比率亦顯著高於 (P<0.05)糞中無出現卵囊之牛隻；7株nested PCR陽性產物之核酸定序比對，其相似性與GenBaank 之Cryptosporidium parvum bovine type達96.0-99.6 %。其次，為進一步釐清隱孢子蟲感染在犢牛下痢之病原性角色，對6場MZN陽性場每場8頭共48頭犢牛，於
出生1個月內每隔3天以MZN法測定每公克糞便隱孢子蟲卵囊數 (oocyst per gram, OPG)和記錄其糞便性狀，並以市售偵測抗原之ELISA套組抽檢下痢便中有無Cryptosporidium parvum、Rotavirus、Coronavirus和E. coli F5 等之併發感染，結果C. parvum混合Rotavirus、Coronavirus和/或E. coli F5感染者占 87.9 % (58/66)，僅有7.6 % (5/66)是C. parvum單獨感染；但是隨著糞中OPG急遽上升犢牛總是都會出現水樣下痢，糞便性狀總是隨著OPG之量而變化，且隨著卵囊消失糞便都會轉為正常，故由以上結果我們認為隱孢子蟲感染在台灣地區犢牛下痢佔有很重要之病原性角色。

關鍵字

微小隱孢子蟲感染 ； 新生犢牛下痢症候群 ； 台灣

並列摘要

To evaluate the incidence of Cryptosporidium infection in dairy herds in Taiwan, feces of 228 cattles from 24 farms in three counties of southern Taiwan were examined by modified Ziehel-Neelsen stain (MZN) and nested polymerase chain reaction (nested PCR) for Cryptosporidium oocyst from August 2006 to January 2007. Any of the seven nested PCR positive samples collected from seven MZN positive farms were sequenced and compared with the genotypes of Cryptosporidium reported in Genbank. The results show that the detection rate of Cryptosporidium infection was 28.9 % (66/228) by nested PCR, which is significantly (P<0.05) higher than MZN (7.0%, 16/228). Farm positive rates were 62.5 % (15/24) by nested PCR and 37.5 % (9/24) by MZN. Cattle positive rates were 28.9 % (66/228) by nested PCR and 7.0 % (16/228) by MZN, respectively. No significant differences of farm positive rate and cattle positive rate ware found among three counties of southern Taiwan, but infection rate in the group of under-one-month old calves was significantly (P<0.05) higher than other elderly groups. The positive rate of Cryptosporidium infection in the diarrheic cattles was significantly (P<0.05) higher than that in the non-diarrheic cattles. The diarrheic rate of Cryptosporidium oocyst positive cattles was also significantly (P<0.05) higher than that of Cryptosporidium oocyst negative cattles. The similarity of nucleotide sequences between seven nested PCR positive products and Cryptosporidium parvum bovine genotype in GenBank is 96.0-99.6 %. Next, to evaluate the pathogenic role of Cryptosporidium in calf diarrhea, a total of 48 neonatal calves from six MZN positive farms with 8 calves per farm were continuously calculated the oocyst per gram (OPG) by MZN three days interval during the first month age after calving. The consistency of feces was recorded, and the commercial Ag-detection-ELISA diagnostic kit for Cryptosporidium parvum, Rotavirus, Coronavirus and E. coli F5 was used to examine the diarrheic feces. The results show that the C. parvum infection mixed with Rotavirus, Coronavirus and/or E. coli F5 was 87.9 % (58/66), and only 7.6 % (5/66) was purely C. parvum infection. Watery diarrheic feces were always seen following with the rapid increase of the OPG. Fecal consistency always changed with the level of OPG. The feces always convert to normal consistency following the disappearance of oocyst. Then we concluded that Cryptosporidium infection plays a very important pathogenic role in calf diarrhea in Taiwan.

並列關鍵字

Cryptosporidium parvum infection ； neonatal calf diarrhea ； Taiwan

參考文獻

渡邊裕子。台灣地區隱孢子蟲症之疫學研究。博士論文，國立中興大學獸醫學研究所。1-108，2006。

許昺慕、吳惠琳、曾凱熙、季德玉、葉珍羽。台灣地區豢養動物糞便檢體中隱孢子蟲存在情形之檢測。健康管理學刊1卷1期：76-86，2003。

Abe N, Iseki M. Identification of genotypes of Cryptosporidium parvum isolates from ferrets in Japan. Parasitol Res 89: 400-424, 2003.

Acres SD. Enterotoxigenic Escherichia coli infections in newborn calves: a review. J dairy Sci 68: 229-256, 1985.

Alvarez-Pellitero P, Sitjà-Bobadilla A. Cryptosporidium molnari n. sp. (Apicomplexa: Cryptosporidiidae) infecting two marine fish species, Sparus aurata L. and Dicentrarchus labrax L. Int J Parasitol 32: 1007-1021, 2002.