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Piglet diarrhea coliphage and application thereof

A technology of Escherichia coli and phage, applied in the direction of phage, virus/phage, application, etc., can solve the problems that antibiotics have poor effect on pathogenic Escherichia coli, cannot target multiple Escherichia coli at the same time, and are expensive, etc., to achieve non-toxicity Side effects, good pH and temperature tolerance, and high safety effects

Active Publication Date: 2021-08-03
QINGDAO PHAGEPHARM BIO TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Bile salts, brilliant green, etc. have inhibitory effects on it; they are sensitive to sulfonamides, streptomycin, chloramphenicol, etc., but with the extensive use of antibiotics, drug resistance has emerged, and antibiotics are resistant to pathogenic Escherichia coli effect is getting worse
There are many vaccines for preventing pathogenic Escherichia coli at home and abroad, the price is expensive, and there are too many serotypes of Escherichia coli, and one vaccine cannot target multiple Escherichia coli serotypes at the same time, so it is sought to develop an effective treatment and prevention of piglet diarrhea of drugs or products are imminent

Method used

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  • Piglet diarrhea coliphage and application thereof
  • Piglet diarrhea coliphage and application thereof
  • Piglet diarrhea coliphage and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Example 1 Isolation of phage and its biological characteristics

[0029] Recovery of bacteria and preparation of bacteria solution

[0030] Pick the Escherichia coli cryopreservation solution and draw lines on the SS medium, put it in a 37°C incubator and cultivate it for 16-24 hours. After overnight, pick a single colony with a sterile white tip and inoculate it into 5ml of nutrient broth. Place in a shaker at 37°C and shake at 170rpm for 12h to obtain the bacterial liquid.

[0031] Phage Isolation and Purification

[0032] Put the feces samples and sewage brought back from the pig farm into the triangular flasks filled with nutrient broth, add 1% Escherichia coli in proportion to each triangular flask, stir evenly, and put them in a shaker at 37°C, 170rpm Shake overnight. Centrifuge the overnight mixed sample liquid at 11000rpm for 5min, and filter it with a 0.22μm filter to form a phage stock solution; mix the phage stock solution with the host bacteria, incubate ...

Embodiment 2

[0048] Whole Genome Sequencing of Example 2 Phage vB-EcoP-ZPD342

[0049] The whole genome of the phage was sequenced using an Illumina Miseq (San Diego, CA, USA) sequencer. Using the Miseq Reagent Kit v2 kit to construct a 600bp sequencing library, the main process is as follows: Genomic DNA is ultrasonically fragmented, the ends are blunted, and after adding specific adapters, the DNA is amplified, purified and screened to obtain the constructed sequencing library. The original phage sequencing data were assembled and spliced ​​using the software Newbler2.9. Finally, the size of the phage genome is 39014bp, which is double-stranded DNA, the G+C content is 48.76%, and contains 51 predicted open reading frames.

[0050] The 51 ORFs protein sequences of phage vB-EcoP-ZPD342 were compared using the online tool BLASTp and the Conserved Domain Database (CDD). Among them, phage vB-EcoP-ZPD342 contained 45 known encoded functional proteins, and the remaining ORFs were hypothetical ...

Embodiment 3

[0066] The mensuration of embodiment 3 phage lysis spectrum and in vitro lysis test

[0067] Determination of phage lysis profiles

[0068] The lysis spectrum of the phage was measured by the double-layer plate method, and the steps were as follows: prepare the bacterial suspension of the host bacteria according to the method in Example 1, and 100 strains of pathogenic E. Bacillus standard strains K88, K99, 987P, K88ac. Use the double-layer plate method to detect whether it is lysed. After the upper layer of agar is solidified, place it in an incubator at 37°C and incubate it upside down for 6-8 hours, then observe the results. See Table 3.

[0069] Table 3 Escherichia coli phage lysis profile

[0070]

[0071]

[0072]

[0073] Phage vB-EcoP-ZPD342 has a wide cleavage spectrum. For 100 pathogenic Escherichia coli epidemic strains, phage vB-EcoP-ZPD342 can lyse 92 of them, with a cleavage rate of 92%. The phage can lyse K88, K99, 987P, Escherichia coli of F41, LT1,...

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Abstract

The invention discloses and relates to a piglet diarrhea coliphage and application thereof, the piglet diarrhea coliphage is named as vB-EcoP-ZPD342, and the preservation number is CGMCC NO.18863. The coliphage preparation can effectively prevent and treat piglet diarrhea caused by escherichia coli, can be used for disinfecting escherichia coli in a farm environment and breeding equipment, and can also be used as a preservative of meat products. The coliphage preparation provided by the invention is colorless, non-toxic and odorless, does not pollute the environment when being sprayed in the environment, and has broad-spectrum bactericidal performance. The coliphage preparation disclosed by the invention provides a new idea for preventing and treating piglet diarrhea, killing escherichia coli in a farm environment and preventing and treating meat products.

Description

technical field [0001] The invention belongs to the field of biology, and relates to an E. coli phage, which can prevent and treat diarrhea of ​​piglets, and sterilize and disinfect the farm environment and breeding equipment, and is used as a preservative for meat products. Background technique [0002] Piglet diarrhea is a common disease in pig farms. Pathogenic Escherichia coli infection causes piglet yellow scour, piglet pullorum, and piglet edema, with high morbidity and mortality. This kind of disease is more likely to occur in piglets during the period from birth to weaning. After the piglets are infected, the symptoms often manifest as vomiting, diarrhea, dehydration and rapid weight loss, which has a great impact on the growth of piglets and also causes huge economic losses to the farm. . At the same time, meat products are easily contaminated by microorganisms and Escherichia coli during storage and transportation, and pathogenic Escherichia coli contamination can...

Claims

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Application Information

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IPC IPC(8): C12N7/00A61K35/76A61P31/04A61P1/12A23K10/18A23B4/22A01N63/40A01P3/00A01P1/00C12R1/92
CPCC12N7/00A61K35/76A61P31/04A61P1/12A23K10/18A23B4/22A01N63/40C12N2795/10221C12N2795/10231C12N2795/10232A23V2002/00A23V2200/10Y02A50/30
Inventor 潘强任慧英孙虎芝闫艳新袁嘉婧张灿
Owner QINGDAO PHAGEPHARM BIO TECH CO LTD
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