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Escherichia coli bacteriophage composition and application thereof

A technology of Escherichia coli and phage, which is applied in the field of microorganisms, can solve the problems of bacteria and fungi proliferation, good disinfection effect of bacteria, and reduced production performance, etc., and achieve fast sterilization, host specificity, and safe killing effect

Active Publication Date: 2020-06-30
QINGDAO PHAGEPHARM BIO TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Nowadays, large-scale chicken farms mostly use nipple-type automatic drinking water system as drinking water equipment, and the nipple-type automatic drinking water system pipes are completely closed, and it is difficult to clean the interior, and the chicken farm is to improve the feed conversion rate, egg production rate and enhance the immunity of chickens. , anti-stress ability, etc., usually add multidimensional products through the drinking water system, which accelerates the pollution of the drinking water system, excessive bacteria, etc., and even forms biofilms, which leads to disease in chickens, thereby reducing production performance and increasing costs
[0005] At present, the cleaning of the waterline mainly uses acidifiers, descaling agents and disinfectants, etc. However, the long-term use of acidifiers will cause a large number of bacteria and fungi in the waterline to proliferate; When other medicines or vaccines are used in drinking water, the disinfectant in the water line will reduce the efficacy of the medicine or inactivate the vaccine
Therefore, there is no waterline cleaner that has a good disinfection effect on bacteria in the waterline and has no side effects.

Method used

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  • Escherichia coli bacteriophage composition and application thereof
  • Escherichia coli bacteriophage composition and application thereof
  • Escherichia coli bacteriophage composition and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] Example 1 Isolation and preparation of phage

[0045] 1. Sample collection

[0046] The chicken manure sample in the present invention was collected from a chicken farm in Panjin City, Liaoning Province.

[0047] 2. Resuscitation culture of host bacteria and preparation of propagation fluid

[0048] The host bacteria are E. coli isolated and stored in this laboratory, and diseased chickens from a chicken farm in Jiangsu Province are isolated. Use a sterile inoculating loop to dip the host bacteria liquid, streak inoculation on MacConkey agar medium, and cultivate in a 37°C incubator for 16-24 hours to obtain a single colony. Pick a single colony, inoculate it in a 5mL NB broth test tube, and culture it with shaking at 170 rpm / min for 12-16 hours at 37°C to obtain a fresh host bacterial growth solution for use.

[0049] 3. Treatment of fecal samples, sewage and other samples

[0050] Put an appropriate amount of fecal water into the bubble sample bottle, add 200μL of host bacteri...

Embodiment 2

[0054] Example 2 Proliferation and purification of phage

[0055] 1. Purification of phage

[0056] Using the double-layer plate method, pick a single, smooth and transparent plaque from the double-layer plate that forms the plaque, and inoculate it in a sterile EP tube containing 1 ml of NB broth, and shake at 37°C and 170 rpm. 30min, get the phage extract; centrifuge at 11000rpm for 5min, take 100μL of the centrifuged phage extract and 100μL of the host bacteria proliferation solution, mix well, incubate at 37℃ for 5min, add to the preheated melted upper agar, mix, and quickly pour it into the lower agar On the medium plate, after the upper agar is solidified, invert it in a 37°C incubator for 6-8 hours. Repeat the above steps 3-5 times to repeatedly purify the phage until a plaque with uniform size, shape and distribution is obtained.

[0057] 2. Preparation of phage proliferation solution

[0058] From the purified plaque double-layer plate, deduct a single plaque and inoculate ...

Embodiment 3

[0059] Example 3 Observation and identification of phage morphology

[0060] The method of morphological observation is: take 20μL of phage sample and drop it on a copper mesh with carbon coating film, let it settle naturally for 15min, blot dry with filter paper, and stain with 2%(W / V) phosphotungstic acid (PTA) After 1-2 min, the filter paper is slightly blotted dry, and after drying, observe and take pictures with a transmission electron microscope.

[0061] Observation result: phage PD06 as image 3 As shown, the head is icosahedron, the head is about 100nm wide, about 100nm long, and the tail is about 110nm long. According to the phage classification, the phage morphology in this study conforms to the characteristics of the Mycophage family and belongs to the Mycophage family.

[0062] Phage PD114 as Figure 4 As shown, the head has a polyhedral structure with a head length of about 60nm, a width of about 63nm, and a tail length of about 150nm. According to the classification of...

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Abstract

The invention discloses a bacteriophage composition which is formed by compounding an escherichia coli bacteriophage PD06 and an escherichia coli bacteriophage PD114 and is constructed on the basis ofa cocktail prevention and control method, and an application of the bacteriophage composition in a medicament for preventing and treating chicken colibacillosis. The invention also discloses a biological bacteriostatic agent, disinfectant and food additive which take the bacteriophage composition as an active component. The bacteriophage composition not only can quickly, efficiently and safely kill pathogenic escherichia coli and even drug-resistant escherichia coli, but also can effectively reduce the death rate of sick chicks. As a waterline cleaning agent, the bacteriophage composition caneffectively degrade biofilms formed by enrichment of escherichia coli in a waterline, and can purify drinking water. In addition, the bacteriophage composition also has the advantages of safety, no residue and no side effect.

Description

Technical field [0001] The present invention relates to the technical field of microorganisms, in particular to a coliphage composition and its application. Background technique [0002] Avian colibacillosis is a common poultry disease caused by certain pathogenic Escherichia coli. The onset of colibacillosis is not seasonal. Chickens at all stages may be infected, especially young chickens; The excrement of feces and the contamination of eggshells by feces can lead to increased mortality of chicken embryos and an increase in the proportion of weak chicks. The disease can also be infected through the digestive tract, respiratory tract, umbilical cord, etc. through infectious agents such as drinking water, feed, air, and cages contaminated by Escherichia coli, which can cause catarrhal cavity and fallopian tube inflammation. Depending on the type of disease, it can cause a variety of symptoms, including Escherichia coli sepsis, air sacculitis, granuloma, perhepatitis, peritonitis...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/00C12Q1/10C12Q1/06A23K10/18A23B4/22A23B5/16A61K35/76A61P31/04A01N63/40A01P1/00C12R1/19
CPCA23B4/22A23B5/16A23V2002/00A61K35/76A23K10/18A61P31/04C12N7/00C12N2795/10121C12Q1/06C12Q1/10A23V2200/10A23V2250/5026A23V2250/5114A23V2250/61A23V2250/628
Inventor 潘强任慧英孙虎芝刘广芹王翠
Owner QINGDAO PHAGEPHARM BIO TECH CO LTD
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