台灣環境水體中水媒腺病毒之調查與分析方法研究

Survey and Analysis of Waterborne Related Adenovirus in Taiwan's Environmental Water

楊怡俊(Yi-Chun Yang)

指導教授 ： 許昺慕

國立中正大學/理學院/地球與環境科學系應用地球物理與環境科學研究所/碩士(2014年)

摘要

腺病毒會藉由人類活動產生的廢水排放而流入環境中，因此有必要針對環境水體中存留之腺病毒研發快速、有效的檢測方法，避免人類接觸或暴露於含有腺病毒的飲用水或娛樂用水。本研究分別採集全台灣水庫和河川環境水體，並進行人類腺病毒(human adenovirus; HAdv)相關議題之研究。環境水體經過GN6濾紙過濾濃縮後，利用靈敏且專一性的巢式PCR檢測水體樣本中的腺病毒，並利用定序分析(Sequence analysis)探討環境水中腺病毒的基因型別。本研究也利用即時定量聚合酶連鎖反應(TaqMan Real-Time PCR)檢測環境水體中腺病毒的濃度。最後，亦使用不同病毒濃縮法，對於環境水體中腺病毒的回收率進行探討。
台灣環境水體中腺病毒之定性分析結果顯示水庫、河川和朴子溪流域檢出率分別為0%、18%和16%。腺病毒的檢出率和總菌落數和導電度具有相關性。檢出型別分為三群：豬腺病毒5型，會造成豬隻的腹瀉症狀；人類腺病毒A群，能引起兒童急性出血性膀胱炎；人類腺病毒41型，能造成人類四歲以下兒童腸胃炎，是環境水體中普遍存在的人類腺病毒型別。人類腺病毒JTVX TaqMan Real-Time PCR定量的濃度濃度範圍在6.10 × 102到2.04 × 109 copies/L之間。本研究12種腺病毒環境水體濃縮法實驗顯示，利用孔徑大小0.2μm的混合纖維濾紙濃縮環境水體方法對於本研究朴子溪水體中之腺病毒有最好的回收率，而硝酸纖維負電濾紙搭配有機凝絮二次濃縮回收效率最差。

關鍵字

正電濾紙 ； 分子篩 ； 硝酸纖維濾紙 ； 混合纖維濾紙 ； 聚合酶連鎖反應 ； 人類腺病毒 ； 環境水體 ； 有機凝絮 ； 回收率

並列摘要

Adenovirus can be introduced into the aquatic environments by human waste discharge. Methods for rapid detection and quantification of adenovirus in water are urgently needed to prevent human exposure to pathogens through drinking and recreational waters. In this study, reservoir and river water samples were collected over Taiwan, and analyzed for human adenovirus (HAdvs). Adenoviruses were recovered from the water samples using GN6 membrane filter. The sensitivity and specificity of nested PCR primer sets were used for detection of adenovirus in the water samples. Then, the genotypes of adenovirus were identified by sequence analysis. This study also use TaqMan Real-Time PCR for quantification of adenovirus in the water samples. Finally, recovered rate of different concentration methods were evaluated for human adenovirus in the water samples. The results showed that 0%, 18% and 16% of HAdvs present in reservoir, river and puzih river, respectively. The significant differences (Mann-Whitney U Test, p < 0.05) were found between the incidence of HAdvs and HPC as well as Conductivity. Several types of adenovirus were detected in Taiwan suface water samples: Porcine adenovirus 5, which can cause gastroenteritis in pigs; Human adenovirus A (Including 12 and 31), which can cause Haemorrhagic cystitis in children; Human adenovirus 41, which are unique in being responsible for most cases of Ad-associated gastroenteritis in children. The HAdvs concentration ranging from 6.10×102 to 2.04×109 copies/L. Twelve concentrated method was compared for adenovirus recovery efficiencies in puzih river water. Optimal recovery was obtained at 0.2μm Mixed Cellulosed Ester. Worst recovery was obtained at nitrocellulose filter membrane with organic flocculation.

並列關鍵字

NanoCeram ； 1MDS ； Nitrocellulose filter membrane ； Mixed Cellulosed Ester ； PCR ； Human adenovirus ； Environmental water ； Molecular filtration ； Organic flocculation ； recovery

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