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Cultivation process of serum-free anaerobic high-density fermentation of Streptococcus equi subspecies zooepidemicus

A technology of high-density fermentation and Streptococcus equi, which is applied in the field of agricultural microorganisms, can solve the problems of vaccine safety hazards and other problems, and achieve the effects of improving immunogenicity, fast growth speed, and improving protection effect

Active Publication Date: 2019-10-18
山东滨州沃华生物工程有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Streptococcus equi subspecies zooepidemicus is a facultative anaerobic microorganism that requires serum and glucose as nutrients during the growth process, but the residue of these heterologous substances will pose a safety hazard to the vaccine

Method used

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  • Cultivation process of serum-free anaerobic high-density fermentation of Streptococcus equi subspecies zooepidemicus
  • Cultivation process of serum-free anaerobic high-density fermentation of Streptococcus equi subspecies zooepidemicus

Examples

Experimental program
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Effect test

Embodiment 1

[0024] A serum-free anaerobic high-density fermentation process for Streptococcus equi subspecies zooepidemicus, comprising the steps of:

[0025] 1) Take the freeze-dried Streptococcus equi subsp. zooepidemicus strain ST171 fermented seed bacteria and activate them in TSA medium supplemented with 5% adult bovine serum for 12-16 hours until a single colony grows;

[0026] 2) Pick a single colony to the slant of TSA agar supplemented with 5% adult bovine serum, and cultivate it for 12-16 hours to obtain first-class seeds;

[0027] 3) Wash the slant of TSA agar supplemented with 5% adult bovine serum with 2 mL of TSB liquid, inoculate the primary seeds on the TSB medium supplemented with 5% adult bovine serum, and shake on a shaker at 37°C and 100rpm Cultivate for 3-4h until OD 600 When the value reaches 0.4-0.6, the secondary seed is obtained;

[0028] 4) Transfer the secondary seed solution to the fermentation medium (enriched casein medium) in a 10L fermenter with an inocul...

Embodiment 2

[0030] A serum-free anaerobic high-density fermentation process for Streptococcus equi subspecies zooepidemicus, comprising the steps of:

[0031] 1) Take the freeze-dried Streptococcus equi subsp. zooepidemicus strain ST171 fermented seed bacteria and activate them in TSA medium supplemented with 7% adult bovine serum for 12-16 hours until a single colony grows;

[0032] 2) Pick a single colony to the slant of TSA agar supplemented with 7% adult bovine serum, and cultivate it for 12-16 hours to obtain first-class seeds;

[0033] 3) Wash the slant of TSA agar supplemented with 7% adult bovine serum with 3 mL of TSB liquid, inoculate the primary seeds on the TSB medium supplemented with 7% adult bovine serum, and shake on a shaker at 38°C and 100rpm Cultivate for 3-4h until OD 600 When the value reaches 0.4-0.6, the secondary seed is obtained;

[0034] 4) Transfer the secondary seed solution to the fermentation medium (enriched casein medium) in a 15L fermenter with an inocul...

Embodiment 3

[0036] A serum-free anaerobic high-density fermentation process for Streptococcus equi subspecies zooepidemicus, comprising the steps of:

[0037] 1) Take the freeze-dried Streptococcus equi subsp. zooepidemicus strain ST171 fermented seed bacteria and activate them in TSA medium supplemented with 5% adult bovine serum for 12-16 hours until a single colony grows;

[0038] 2) Pick a single colony to the slant of TSA agar supplemented with 5% adult bovine serum, and cultivate it for 12-16 hours to obtain first-class seeds;

[0039] 3) Wash the slant of TSA agar supplemented with 5% adult bovine serum with 2 mL of TSB liquid, inoculate the primary seeds on the TSB medium supplemented with 5% adult bovine serum, and shake on a shaker at 37°C and 100rpm Cultivate for 3-4h until OD 600 When the value reaches 0.4-0.6, the secondary seed is obtained;

[0040] 4) Transfer the secondary seed solution to the fermentation medium (enriched casein medium) in a 10L fermenter with an inocul...

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Abstract

The invention discloses serum-free anaerobic high-density fermentation culture process for Streptococcus equi subsp. Zooepidemicu and belongs to the field of agricultural microbiology. Streptococcus equi subsp. Zooepidemicu fermented high-density antigens are obtained through the sequential steps of resuscitating lyophilized seeds with TSA, carrying out primary culture on a slant of TS agar, carrying out secondary seed culture on a TSB medium, and anaerobically fermenting in enriched trypsin-casein medium free of serum and glucose. Study on industrial enlarged production is carried out through the method of anaerobic fermentation culture without adding any serum or glucose, the content of fermented antigens is not reduced, three is no need for ventilation regulation and dissolved oxygen control during fermentation, the antigen content is high, process parameters are easy to control, and supplements are reduced; the process is stable and high in growth speed compared with the existing fermentation process and is easy for implementing large-scale high-density culture of Streptococcus equinus antigens; immunogenicity of a strain is improved, and protection for vaccines is improved.

Description

technical field [0001] The invention relates to the technical field of agricultural microorganisms, in particular to a serum-free anaerobic high-density fermentation culture process of Streptococcus equi subspecies zooepidemicus. Background technique [0002] Streptococcus equi subspecies zooepidemicus belongs to Group C Streptococcus according to Langer's classification. This bacterium can cause a variety of animal infections. The clinical symptoms are sepsis, meningitis, arthritis, etc., and it is generally endemic. With the development of intensive pig farming, streptococcal suis occurs frequently in China. Epidemiological surveys showed that, in addition to Streptococcus suis type 2, Streptococcus equi subspecies zooepidemicus was still the main pathogen of Streptococcus suis in various parts of China. At present, the main means of controlling the disease is still vaccines, mainly attenuated vaccines and inactivated vaccines. [0003] Streptococcus equi attenuated stra...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/20C12R1/46
Inventor 朱杰王楠牛成明董新荣
Owner 山东滨州沃华生物工程有限公司
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