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Vibrio alginolyticus bacteriophage microecological preparation as well as preparation method and application thereof

A micro-ecological preparation and technology of Vibrio alginolyticus, applied in the field of microorganisms, can solve problems such as low phage titer, and achieve the effects of increasing titer, prolonging fermentation time, and eliminating content

Inactive Publication Date: 2018-06-01
XIAMEN CANCO BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

The phage fermentation methods disclosed in the above patents all have the problem of low phage titer

Method used

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  • Vibrio alginolyticus bacteriophage microecological preparation as well as preparation method and application thereof
  • Vibrio alginolyticus bacteriophage microecological preparation as well as preparation method and application thereof

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preparation example Construction

[0022] The invention provides a preparation method of Vibrio alginolyticus bacteriophage probiotics, comprising the following steps:

[0023] 1) The Vibrio alginolyticus liquid is inserted into the fermentation medium for cultivation to obtain the host fermentation liquid; the initial cell concentration of the Vibrio alginolyticus host in the fermentation medium is 10 10 ~10 16 CFU / ml;

[0024] 2) Set the concentration to 10 8 ~10 16 Vibrio alginolyticus phage liquid of PFU / ml is inoculated into the host fermentation broth for fermentation; the inoculum amount of Vibrio alginolyticus phage liquid is 1% to 10%;

[0025] 3) When the fermentation culture in the step 2) is carried out for 6-12 hours, add Vibrio alginolyticus liquid to the host fermentation liquid in the step 2), continue the fermentation culture, control the entire fermentation period to 18-24 hours, and obtain the fermentation liquid;

[0026] 4) Inactivating the host of Vibrio alginolyticus with the ferment...

Embodiment 1

[0048] Vibrio alginolyticus was isolated from a culture pond of Penaeus vannamei. Penaeus vannamei culture ponds are located in Shandong, Fujian, Guangdong, Hainan and other places. The collection time is September 2016. Vibrio alginolyticus was isolated from the intestinal tract of Penaeus vannamei, and the phage of Vibrio alginolyticus was isolated from cultured water and seawater of Penaeus vannamei.

[0049] The phage expansion culture method of the isolated 3 strains of Vibrio alginolyticus, the specific steps are as follows: first determine the optimal multiplicity of infection of the phage: the multiplicity of infection (MOI) refers to the number of phages and the number of host bacteria when the phage initially infects the host bacteria. The ratio, also known as the infection multiple. Inoculate 100ul of host bacteria into 200ml NB culture medium, and cultivate to logarithmic phase at 150r / min for 12h. Then take the initial valence as 10 9 PFU / mL phage culture medi...

Embodiment 2

[0051] The method for feeding phage host fermentation to produce phage comprises the following steps:

[0052] 1) Use a 50L fermenter for fermentation. The volume of the fermentation medium in the fermenter is 35L. Insert the host into the fermentation medium so that the number of colonies in the initial medium is 10 10 CFU / ml; the formula of the fermentation medium is as follows: peptone 10g / L; beef extract powder 10gL; sodium chloride 1g / L; pH is 9.0;

[0053] 2) add the 1% 10 of the first strain fermented liquid volume again 8 The Vibrio alginolyticus phage liquid of PFU / ml was cultured in a fermenter, the fermentation temperature was 28° C., the rotation speed was 200 rpm; the pH value was controlled at 9.

[0054] 3) When the fermentation was carried out for 6 hours, the phage host was added to the fermenter, and the concentration was controlled at 10 14 Between CFU / ml, the feeding time is 3h, the flow speed is 2L / h, the feeding volume is 6% of the total fermentation vo...

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Abstract

The invention provides a vibrio alginolyticus bacteriophage microecological preparation as well as a preparation method and application thereof, and belongs to the technical field of microorganisms. The preparation method comprises the following steps: inoculating a vibrio alginolyticus solution and a vibrio alginolyticus bacteriophage solution into a fermental cultivation solution for fermental cultivation; performing the fermental cultivation for 6 to 15 h, then adding the vibrio alginolyticus solution into a host fermentation solution in a flowing manner for continuous fermental cultivation, controlling the whole fermentation period at 18 to 24 h, and inactivating vibrio alginolyticus hosts in the fermented solution, thus obtaining the vibrio alginolyticus bacteriophage microecologicalpreparation. According to the preparation method provided by the invention, fermentation and flowing adding moments are controlled to perform the fermental cultivation for 6 to 15 h, and then the hostsolution is added in the flowing manner, so that the titer of vibrio alginolyticus bacteriophage can be greatly improved without prolonging the fermentation time.

Description

technical field [0001] The invention belongs to the technical field of microbes, and in particular relates to a vibrio alginolyticus bacteriophage micro-ecological preparation and a preparation method and application thereof. Background technique [0002] At present, the main means of controlling and eliminating pathogenic bacteria are still various physical or chemical methods, including the use of various antibiotics. Both physical and chemical methods have obvious disadvantages. First of all, the elimination effect of physical methods on various pathogenic bacteria can only be applied to a certain link in the related field, and cannot be applied to all links in the entire field. Secondly, although the acid-base treatment in the chemical method can achieve a certain control effect, the effect of these methods is extremely unsatisfactory after the harmful bacteria form a biofilm. The research and development of microbial ecological preparations has attracted the attention...

Claims

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Application Information

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IPC IPC(8): C12N7/00C12N1/20C12Q1/04C12R1/92C12R1/63
CPCC12N1/20C12N7/00C12N2795/00051C12Q1/04G01N2333/28
Inventor 姜宗然付汉清郭立
Owner XIAMEN CANCO BIOTECH CO LTD
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