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Method for quickly concentrating viruses in water body

A water and virus technology, applied in the field of environmental virology and environmental monitoring, can solve the problems of limited promotion and application, high requirements, complex operation, etc., and achieve the effects of low equipment requirements, high recovery efficiency and simple operation.

Inactive Publication Date: 2011-06-15
何雅青 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Most of these methods require high conditions, some require expensive instruments, and some are complicated to operate, so the promotion and application in grassroots units is limited to a certain extent.

Method used

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  • Method for quickly concentrating viruses in water body
  • Method for quickly concentrating viruses in water body
  • Method for quickly concentrating viruses in water body

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] An outbreak of diarrhea occurred in a company. First, norovirus was detected positive in the stool samples of two cases. Epidemiological investigations showed that the attack rate of people who drank the company's direct drinking water was higher than that of people who drank other types of water. Norovirus detection was performed on 9 water samples, and norovirus was detected from 3 samples. The specific method is as follows:

[0022] Carry out virus concentration with the method of the present invention respectively to every part of water sample, the steps are as follows:

[0023] 1. Pretreatment of virus-contaminated water samples

[0024] Take 1 L of water sample, add MgCl2 solution to make the final concentration 0.05 mol / L, then adjust the pH of the solution to 3.5 with 1 mol / L hydrochloric acid, stir and mix evenly on a magnetic stirrer.

[0025] 2. Filter

[0026] Use Millipore's microbial filtration system or Milliflex automatic biological filtration system ...

Embodiment 2

[0037] To monitor enteroviruses in environmental water, river water was randomly selected, a total of 6 water samples, 1000ml / point, and enteroviruses were detected from 1 sample. The specific method is as follows:

[0038] Carry out virus concentration to each part of water sample respectively, the concentration step is the same as embodiment 1.

[0039] Take 200 μl of the resuspended water virus concentrate and extract RNA.

[0040]RT-PCR detection, including: use the extracted viral RNA as a template, apply random primers to obtain viral genome cDNA by reverse transcription under the action of M-MLV reverse transcriptase, the reaction condition is 65°C for 5min, and incubate at 25°C after adding reverse transcriptase 10min, 42°C extension for 1h, 70°C denaturation for 10min. Using viral cDNA as a template, human enterovirus nucleic acid detection universal primer sequence: 5'-TCC GGC CCC TGA ATG CGGCTA ATC C-3'PE1 (downstream): 5'-ACA CGG ACA CCC AAA GTA GTC GGT CC-3 'Am...

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Abstract

The invention discloses a method for quickly concentrating viruses in a water body, which comprises the following steps of: a, pre-treating the virus polluted water body; b, performing suction-filtering on the virus polluted water body pre-treated in the step a by using a microbe filter system under negative pressure; c, fully soaking a water body virus enriched filter membrane into 30ml of beef extract-glycine eluent with pH of 9.5, and then removing the filter membrane by magnetic stirring and eluting to obtain the eluent; and d, adding polyethylene glycol 8000 (PEG8000) into the eluent obtained in the step c till the final concentration of the eluent is 8 percent; and adding NaCl till the final concentration of the eluent is 0.3mol / L, stirring and mixing uniformly, standing for 13 to 16 hours at the temperature of 4 DEG C, centrifuging for 30 minutes at 10,000rpm in a high-speed refrigerated centrifuge, removing the supernate, and re-suspending by using 0.5ml of phosphate buffer solution (PBS) with pH of 7.4 to obtain re-suspended water body virus concentrate. The method is simple to operate, has compact steps, low equipment requirement and high reclamation efficiency, and is convenient for popularization and application in the primary level.

Description

technical field [0001] The invention belongs to the technical fields of environmental virology and environmental monitoring, and specifically relates to a quick and easy membrane filtration method for concentrating viruses in water bodies. It adopts a microbial filtration system and uses a mixed cellulose ester microporous filter membrane to concentrate viruses in natural environment water bodies. In order to improve the sensitivity of virus pollution detection in environmental water. Background technique [0002] As a medium for disease transmission, environmental water bodies often cause outbreaks of water-borne diseases, such as viral hepatitis, viral diarrhea, and other water-borne viral diseases, once they are polluted by viruses. cause serious losses. In recent years, due to the aggravation of water pollution, cases of enterovirus infection caused by water transmission have been reported from time to time, which has attracted widespread attention. With the development...

Claims

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Application Information

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IPC IPC(8): G01N1/40
Inventor 何雅青姚相杰
Owner 何雅青
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