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A lytic Vibrio harvelii phage rdp-vp-19012 and its application

A technology of RDP-VP-19012 and Vibrio harveylius, which is applied in the field of microorganisms and can solve problems such as threats to consumer health

Active Publication Date: 2021-11-26
RECOM QINGDAO BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Antibiotics have always been used to control the disease. However, the emergence of drug-resistant strains and the residue of antibiotics in farmed animals have seriously threatened the health of consumers. Countries such as the European Union have banned the use of antibiotics to prevent Harvey vibriosis in aquaculture animals. Therefore, Urgent search for new eco-friendly antimicrobials

Method used

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  • A lytic Vibrio harvelii phage rdp-vp-19012 and its application
  • A lytic Vibrio harvelii phage rdp-vp-19012 and its application
  • A lytic Vibrio harvelii phage rdp-vp-19012 and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] Example 1 Isolation and Identification of Vibrio Harvey

[0032] Sampling from the hepatopancreas of diseased prawns, using aseptic operation method, streaked on TCBS medium, cultured at 37°C for 18-24 hours, and showed round yellow colonies with neat edges on TCBS agar plate, with a diameter of about 2-3 mm; then pick a typical colony and streak it on the Chromagar Vibrio chromogenic medium. After culturing at 37°C for 18-24 hours, white colonies appear on the Chromagar Vibrio chromogenic medium. Pick typical colonies and continue to streak Line purification was performed 3 times, and then a single colony was picked and inoculated in 5 mL of 2216E liquid medium, and cultured with shaking at 200 rpm at 37°C for 8 hours to obtain a uniform turbid bacterial suspension. Through 16sRNA molecular identification, it was determined to be Vibrio harveylius, and one of them was named BV-19013.

Embodiment 2

[0033] Example 2 Drug resistance experiment of Vibrio harveylius

[0034] The isolated Vibrio harveii was tested for drug susceptibility to 12 common antibiotics by disk diffusion method. Drug sensitive disks included cefixime (CEF, 5 μg / tablet), gentamicin (GEN, 10 μg / tablet), tetracycline (TET, 30 μg / tablet), ciprofloxacin (CIP, 5 μg / tablet), norfloxacin (NOR, 10 μg / tablet), chloramphenicol (CHL, 30 μg / tablet tablets), co-trimoxazole (T / S, 1.25 / 23.75 μg / tablet), rifampicin (RIF, 5 μg / tablet), furazolidone (FUR, 300 μg / tablet), amoxicillin (AMO, 10 μg / tablet), erythromycin (ERY, 15 μg / tablet), enrofloxacin (ENR, 5 μg / tablet). Take 100 μL concentration of 10 8 The cell / mL bacterial suspension was coated on the MH plate containing 2% NaCl, and the antibiotic paper was pasted on it. After overnight incubation at 37°C, the diameter of the inhibition zone was recorded. Escherichia coli ATCC25922 was used as the quality control bacteria, and the drug resistance was judged by r...

Embodiment 3

[0037] Example 3 Isolation and Identification of Vibrio Harvey Bacteriophage

[0038] The sewage sample used in the experiment of the present invention was collected from the surrounding sewage of farmers in Jimo, Qingdao in 2019, and was used as a water sample for bacteriophage isolation.

[0039] (1) Take 10 ml of sewage, centrifuge at 10,000rpm for 5 minutes, remove larger impurities and most of the bacteria, and then use a 0.22 micron microporous membrane to filter and sterilize. Take 3 milliliters of the filtrate and 3 milliliters of the host bacterial suspension, add them together into 20 milliliters of autoclaved 2216E liquid medium, and then place them in a 37°C incubator for overnight culture. After culturing, take 5 ml, centrifuge at 10,000 rpm for 5 minutes, and then filter and sterilize with a 0.22 micron microporous filter. The filtrate is the stock solution intended to contain phage. Then, use the double-plate method to identify whether there are phages, and if ...

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Abstract

The invention discloses a lytic Vibrio harveylius phage RDP-VP-19012 and its application. Its host is Vibrio harveylius. The head of the phage is regular hexagonal and the diameter of the head is about 74nm when observed through an electron microscope. , the length of the tail is 95nm, it is a long tail phage, the full length of the genome is 125581bp, and the preservation number is CGMCC No.19985. The Vibrio harvelii phage RDP‑VP‑19012 has a strong lysing effect on Vibrio harvelii in water, has high temperature tolerance and a wide range of acid and alkali tolerance, and can effectively treat the pathogenic bacteria in aquaculture. The Vibrio harvelii disease caused by it can be used as a disinfectant, which provides a source of phage for the industrialized, large-scale and programmed production of phages and the treatment of bacterial diseases in aquaculture animals caused by Vibrio harveylia.

Description

technical field [0001] The invention relates to the field of microorganisms, in particular to a strain of lytic Vibrio harveii phage RDP-VP-19012 and its application. Background technique [0002] Vibrio harvei is a marine pathogen that can infect many aquatic animals such as crustaceans, shellfish, and fish, causing acute sepsis, exophthalmos, and chronic skin ulcers in sick animals. Antibiotics have always been used to control the disease. However, the emergence of drug-resistant strains and the residue of antibiotics in farmed animals have seriously threatened the health of consumers. Countries such as the European Union have banned the use of antibiotics to prevent Harvey vibriosis in aquaculture animals. Therefore, There is an urgent need to find new eco-friendly antimicrobial agents. [0003] Phage is a bacterial virus with high abundance in nature, strong host specificity, and will not damage the micro-ecological environment. After phages infect the host bacteria, n...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/00C02F3/34C12R1/92C02F103/20
CPCC02F3/34C02F2103/20C02F2303/04C12N7/00C12N2795/10321C12N2795/10331
Inventor 杜新永江守杰李霞马如霞李先胜赵丹丹罗成盛张庆刘玉庆盖春云刘爽
Owner RECOM QINGDAO BIOTECH CO LTD
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