Reverse osmosis-based enrichment of virus in water samples: an experiment study
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摘要:
目的 建立基于反渗透技术的水体病毒富集体系。 方法 以2种灭活病毒疫苗及2种病毒核酸质粒为样本,使用反渗透技术对水中病毒进行富集,并通过实时荧光定量(qRT-PCR)实验对不同材质的反渗透膜、不同孔径及过滤次数的富集效果进行测定与优化。 结果 富集效率随反渗透膜孔径的降低而提高,0.10 μm孔径的反渗透膜对2种待测病毒样本的平均富集效率达到(22.86 ± 0.84)%;尼龙、醋酸纤维素、聚醚砜等3种不同材质的反渗透膜对2种待测病毒样本的富集效率接近;富集效率随水样滤次数的增加而提高,但病毒样品在过滤2次与3次比较富集效率提高不明显。 结论 0.10 μm孔径的醋酸纤维素材质滤膜,在2次过滤的条件下,脊髓灰质炎病毒达到32.64 %的病毒富集效率。 Abstract:Objective To establish a reverse osmosis technology-based method for enrichment of virus in water samples. Methods Two inactivated virus vaccines and two viral nucleic acid plasmids were added into simulated water samples and detected with real-time quantitative reverse transcriptase PCR (qRT-PCR) after reverse osmosis-based enrichment. Reverse osmosis membrane material, membrane pore size, and filtration time were optimized in the enrichments. Results The enrichment efficiency increased with the decrease of pore size of the reverse osmosis membrane. The average virus enrichment efficiency of reverse osmosis membrane with the pore size of 0.10 μm was 22.86 ± 0.84% for 2 simulated water samples. The virus enrichment efficiency of nylon, cellulose acetate and polyethersulfone membranes were similar between each other. The enrichment efficiency goes up with the increase of filtration times, but the difference in the efficiency was not significant between two and three times of filtration. Conclusion The efficiency can be up to 32.64% for the enrichment of polio virus in water samples using cellulose acetate filter membrane with 0.10 μm aperture and two times of filtration. -
Key words:
- environment health /
- virus pollution of water /
- reverse osmosis enrichment
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表 1 不同孔径醋酸纤维素滤膜反渗透膜富集效率(%)比较
待测样本 0.45 μm 0.22 μm 0.10 μm F 值 ΔCT 富集效率(%) ΔCT 富集效率(%) ΔCT 富集效率(%) 诺如病毒核酸质粒 1.92 ± 0.69 6.05 ± 1.71 2.30 ± 0.31 7.87 ± 2.14 a 2.34 ± 0.36 8.02 ± 1.23 a 41.532 乙肝病毒核酸质粒 2.96 ± 0.05 12.33 ± 1.44 3.20 ± 0.20 14.77 ± 1.12 a 3.23 ± 0.09 15.04 ± 0.74 a 88.922 手足口灭活病毒 3.10 ± 0.59 13.64 ± 1.84 3.34 ± 0.14 16.12 ± 1.03 a 3.38 ± 0.11 16.54 ± 0.64 a 113.369 脊髓灰质炎灭活病毒 3.96 ± 0.58 25.21 ± 1.13 4.15 ± 0.13 28.48 ± 1.87 a 4.20 ± 0.04 29.17 ± 1.03 a 257.744 注:a 与0.45 μm孔距富集效率比较,P < 0.05。 表 2 0.10 μm孔径不同材质反渗透膜富集效率(%)比较
待测样本 聚醚砜滤膜 醋酸纤维素 尼龙滤膜 F 值 ΔCT 富集效率(%) ΔCT 富集效率(%) ΔCT 富集效率(%) 诺如病毒核酸质粒 2.27 ± 0.29 7.71 ± 1.61 2.34 ± 0.36 8.02 ± 1.23 2.21 ± 0.36 7.37 ± 3.14 47.354 乙肝病毒核酸质粒 3.14 ± 0.41 14.05 ± 1.15 3.23 ± 0.09 15.04 ± 0.74 3.11 ± 0.09 13.77 ± 2.82 0.733 手足口灭活病毒 3.37 ± 0.11 16.46 ± 1.31 3.38 ± 0.11 16.54 ± 0.64 3.41 ± 0.11 17.12 ± 3.14 10.254 脊髓灰质炎灭活病毒 4.14 ± 0.12 27.82 ± 1.51 4.20 ± 0.04 29.17 ± 1.03 4.15 ± 0.04 28.13 ± 1.69 11.697 表 3 0.10 μm孔径的醋酸纤维素滤膜滤过3次富集效率(%)比较
待测样本 1次 2次 3次 F 值 ΔCT 富集效率(%) ΔCT 富集效率(%) ΔCT 富集效率(%) 诺如病毒核酸质粒 2.34 ± 0.36 8.02 ± 1.23 2.62 ± 0.31 9.79 ± 1.14 a 2.74 ± 0.06 10.82 ± 1.04 b 29.475 乙肝病毒核酸质粒 3.23 ± 0.09 15.04 ± 0.74 3.35 ± 0.30 16.37 ± 0.92 a 3.45 ± 0.22 17.61 ± 0.52 b 98.418 手足口灭活病毒 3.38 ± 0.11 16.54 ± 0.64 3.44 ± 0.04 17.37 ± 1.31 a 3.47 ± 0.04 17.72 ± 0.89 76.784 脊髓灰质炎灭活病毒 4.20 ± 0.04 29.17 ± 1.03 4.35 ± 0.27 32.64 ± 1.87 a 4.38 ± 0.60 33.14 ± 1.11 187.784 注: a 与第1次富集效率比较,P < 0.05;b 与第2次富集效率比较,P < 0.05。 -
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