Lactobacillus salivarius NHE-LsE33 and application thereof

Abstract

The invention provides Lactobacillus salivarius NHE-LsE33. The Lactobacillus salivarius NHE-LsE33 is used for separating intestinal tracts of piglets and is preserved in China General Microbiological Culture Collection Center (CGMCC for short, the address is Institute of Microbiology, Chinese Academy of Sciences, No.3, No.1 Yard, West Beichen Road, Chaoyang District, Beijing, the postal code is 100101), the preservation number is CGMCC NO.22510, and the preservation date is May 12, 2021. The lactobacillus salivarius NHE-LsE33 has remarkable prebiotics, and can be used for effectively inhibiting growth and reproduction of pathogenic bacteria such as intestinal pathogenic escherichia coli, staphylococcus aureus, salmonella typhimurium, salmonella choleraesuis, salmonella, shigella, bacillus perfringens, proteus penneri, aeromonas hydrophila and vibrio parahaemolyticus. The lactobacillus salivarius NHE-LsE33 can be prepared into a micro-ecological preparation to be applied to animal breeding, and the strain of the micro-ecological preparation for feed is widened.

Description

technical field

[0001] The invention belongs to the field of probiotics for feeding, and in particular relates to a probiotic Lactobacillus salivarius NHE-LsE33 and its application. Background technique

[0002] Feed mold, mildew and toxins produced by mildew fungi directly affect the quality and safety of feed storage and processing, and seriously endanger the health of livestock, poultry and human beings. After the feed is moldy or polluted by molds, most of the molds are non-toxic molds, which reduce the palatability of the feed and the nutritional value of the feed; a small number of molds are toxin-producing molds, which can cause acute or chronic poisoning of livestock and poultry , Some mycotoxins also have toxic effects such as carcinogenic, teratogenic and mutagenic.

[0003] At present, the main ways to prevent mildew in feed and feed raw materials are to reduce the water content of raw materials, strengthen the management of production and storage, and add single...

Examples

Embodiment 1

[0065] Example 1 Isolation, screening and identification of Lactobacillus salivarius NHE-LsE33

[0066] 1. Isolation and purification of lactic acid bacteria:

[0067] After the healthy piglets were slaughtered and the intestines were taken, 2.5 grams of intestinal contents were quickly collected under aseptic conditions, placed in a triangular flask filled with 22.5mL of sterilized saline, and shaken at a constant temperature at 37°C for 1 hour, then used 10 times The specific dilution method is to dilute to 100,000 times in turn, choose three dilutions of 1,000 times, 10,000 times and 100,000 times, draw 0.1mL and spread it on the improved MRS agar plate, and incubate the plate upside down at 37°C for 48 hours after coating. Use an inoculation loop to pick out the colonies of suspected lactic acid bacteria with obvious calcium-dissolving circles and larger than 5mm, and carry out streak isolation and culture on the MRS agar plate. After 48 hours of cultivation, pick the colo...

Embodiment 2

[0091] The preparation of embodiment 2 Lactobacillus salivarius NHE-LsE33 fermented liquid

[0092] Get Lactobacillus salivarius NHE-LsE33 (preservation number is CGMCC No.22510) seed liquid (the live bacteria concentration is 10 9 CFU / mL) 2mL, inoculated in 100mL shake flask fermentation medium for shake flask fermentation culture; after shake flask fermentation, carry out 10L seed fermentation tank culture, take 100mL shake flask fermentation seed liquid and inoculate into 10L seed fermentation tank for seed culture , 10L fermentation tank liquid volume is 5L seed culture medium. After the cultivation, 5L of seed liquid was inoculated into the fermentation medium in the 100L fermenter to carry out fermentation culture, and the liquid volume of the 100L fermenter was 70L of fermentation culture medium. After the fermentation, the number of viable bacteria in the fermentation broth was detected to be 1.2×10 10 CFU / mL, the fermentation broth was stored in the refrigerator at ...

Embodiment 3

[0101] Example 3 Probiotic verification of Lactobacillus salivarius NHE-LsE33

[0102] On the aseptic operating table, the concentration is 10 9CFU / mL of pathogenic bacteria (enteropathogenic Escherichia coli, Staphylococcus aureus, Salmonella typhimurium, Salmonella choleraesuis, Salmonella, Shigella, Clostridium perfringens, Proteus penninii, Aeromonas hydrophila bacteria, Vibrio parahaemolyticus) bacterial suspension was added to nutrient agar cooled to 45 ℃ (for Clostridium perfringens, the medium was replaced with tryptone-sulfite-cycloserine agar, and the culture conditions required strict anaerobic conditions ) culture medium (after sterilization), and prepare a pathogenic bacteria agar plate of about 4mm, the concentration of pathogenic bacteria in the agar plate is 10 9 CFU / mL. Place the sterilized Oxford cup on the culture medium, gently pressurize it so that it is in contact with the culture medium without gaps, and after 10 minutes, add 200 μL of the preserved fe...

technical field

[0001] The invention belongs to the field of probiotics for feeding, and in particular relates to a probiotic Lactobacillus salivarius NHE-LsE33 and its application. Background technique

[0002] Feed mold, mildew and toxins produced by mildew fungi directly affect the quality and safety of feed storage and processing, and seriously endanger the health of livestock, poultry and human beings. After the feed is moldy or polluted by molds, most of the molds are non-toxic molds, which reduce the palatability of the feed and the nutritional value of the feed; a small number of molds are toxin-producing molds, which can cause acute or chronic poisoning of livestock and poultry , Some mycotoxins also have toxic effects such as carcinogenic, teratogenic and mutagenic.

[0003] At present, the main ways to prevent mildew in feed and feed raw materials are to reduce the water content of raw materials, strengthen the management of production and storage, and add single...

Examples

Embodiment 1

[0065] Example 1 Isolation, screening and identification of Lactobacillus salivarius NHE-LsE33

[0066] 1. Isolation and purification of lactic acid bacteria:

[0067] After the healthy piglets were slaughtered and the intestines were taken, 2.5 grams of intestinal contents were quickly collected under aseptic conditions, placed in a triangular flask filled with 22.5mL of sterilized saline, and shaken at a constant temperature at 37°C for 1 hour, then used 10 times The specific dilution method is to dilute to 100,000 times in turn, choose three dilutions of 1,000 times, 10,000 times and 100,000 times, draw 0.1mL and spread it on the improved MRS agar plate, and incubate the plate upside down at 37°C for 48 hours after coating. Use an inoculation loop to pick out the colonies of suspected lactic acid bacteria with obvious calcium-dissolving circles and larger than 5mm, and carry out streak isolation and culture on the MRS agar plate. After 48 hours of cultivation, pick the colo...

Embodiment 2

[0091] The preparation of embodiment 2 Lactobacillus salivarius NHE-LsE33 fermented liquid

[0092] Get Lactobacillus salivarius NHE-LsE33 (preservation number is CGMCC No.22510) seed liquid (the live bacteria concentration is 10 9 CFU / mL) 2mL, inoculated in 100mL shake flask fermentation medium for shake flask fermentation culture; after shake flask fermentation, carry out 10L seed fermentation tank culture, take 100mL shake flask fermentation seed liquid and inoculate into 10L seed fermentation tank for seed culture , 10L fermentation tank liquid volume is 5L seed culture medium. After the cultivation, 5L of seed liquid was inoculated into the fermentation medium in the 100L fermenter to carry out fermentation culture, and the liquid volume of the 100L fermenter was 70L of fermentation culture medium. After the fermentation, the number of viable bacteria in the fermentation broth was detected to be 1.2×10 10 CFU / mL, the fermentation broth was stored in the refrigerator at ...

Embodiment 3

[0101] Example 3 Probiotic verification of Lactobacillus salivarius NHE-LsE33

[0102] On the aseptic operating table, the concentration is 10 9CFU / mL of pathogenic bacteria (enteropathogenic Escherichia coli, Staphylococcus aureus, Salmonella typhimurium, Salmonella choleraesuis, Salmonella, Shigella, Clostridium perfringens, Proteus penninii, Aeromonas hydrophila bacteria, Vibrio parahaemolyticus) bacterial suspension was added to nutrient agar cooled to 45 ℃ (for Clostridium perfringens, the medium was replaced with tryptone-sulfite-cycloserine agar, and the culture conditions required strict anaerobic conditions ) culture medium (after sterilization), and prepare a pathogenic bacteria agar plate of about 4mm, the concentration of pathogenic bacteria in the agar plate is 10 9 CFU / mL. Place the sterilized Oxford cup on the culture medium, gently pressurize it so that it is in contact with the culture medium without gaps, and after 10 minutes, add 200 μL of the preserved fe...

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