Method for extracting and enriching phage from activated sludge

A technology of activated sludge and bacteriophage, which is applied in the direction of viruses/phages, biochemical equipment and methods, microorganisms, etc., can solve problems such as complex action mechanisms, and achieve the effects of short operation time, mild reaction conditions, and reduced dosage

Active Publication Date: 2019-05-28
ZHEJIANG UNIV
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  • Claims
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Problems solved by technology

However, activated sludge is composed of biological aggregates composed of complex flora and extracellular secretions, which exist in various forms in the sludge system...

Method used

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  • Method for extracting and enriching phage from activated sludge
  • Method for extracting and enriching phage from activated sludge
  • Method for extracting and enriching phage from activated sludge

Examples

Experimental program
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Embodiment

[0039] The purpose of this embodiment is to efficiently extract phage particles from aerobic granular sludge, specifically according to the following steps:

[0040] Step 1: Activated Sludge Grinding

[0041] Take 40ml of activated sludge and grind it in a 15ml cell grinder to disperse the sludge sample. Grind 10ml each time, four times in total. Finally, wash with 5ml PBS buffer solution, and collect all the sludge grinding solution and buffer solution in a 45ml centrifuge tube for the next step of enzymatic hydrolysis.

[0042] Before the sludge sample is ground, the sludge sample can be taken to measure its MLVSS, which can be used as a basic indicator of biomass.

[0043] Step 2: Sludge EPS hydrolysis

[0044] The above sludge samples were hydrolyzed by amylase to remove EPS in the sludge samples and reduce the impact of EPS on phage release. The amount of enzyme added to the sludge sample was twice the amount recommended by the kit. In this experiment, starch glucoam...

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Abstract

The invention discloses a method for extracting and enriching a phage from aerobic granular sludge. The method comprises the following steps: (1) sludge grinding; (2) extracellular polymeric substancehydrolysis; (3) ultrasonic/vortex oscillation; (5) supernatant extraction; (5) secondary ultrafiltration concentration; and (6) sample preservation and fixation. The method is simple and convenient to operate, the reaction condition is mild, no toxic reagent is used, the problems that a granule sludge system is low in extraction efficiency due to EPS (extracellular polymeric substance) wrapping,and baterial cell lysis causes host DNA pollution are solved, and the method is an efficient and feasible phage extracting method. Compared with a traditional method for purifying a phage through combination of PEG and salting out, medicaments such as PEG and NaCl is not required to be used, the purifying time is shortened to about 6 h from orginal 24 h; and the obtained sample is low in salt concentration, further dialysis and desalting are not needed, and the obtained concentrated solution can be directly used for related operations such as a transmission electron microscope, DNA extraction,CsCl2 gradient centrifugation.

Description

technical field [0001] The invention relates to a method for extracting bacteriophage from activated sludge, belonging to the field of wastewater biological treatment. Background technique [0002] Bacteriophage is a type of bacterial virus and is currently known to be the most abundant organism in the natural environment. Vigorous phages prey on host bacteria through five steps: adsorption, injection, integration, assembly, and lysis. In recent years, phages have been gradually applied to pathogen detection, bacterial disease control, and pipeline biofilm suppression, showing great potential. The abundance of phages in activated sludge can reach 100-1000 times the abundance of phages in natural water, which has huge resource potential. However, activated sludge is composed of biological aggregates composed of complex flora and extracellular secretions, which exist in various forms in the sludge system, and the mechanism of interaction with host bacteria is extremely compl...

Claims

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Application Information

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IPC IPC(8): C12N7/00
Inventor 俞卓栋张智明朱亮徐向阳
Owner ZHEJIANG UNIV
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