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Enterococcus faecalis HKF7 with lactic acid activity as well as screening culture method and application thereof

An enterococcus faecalis, active technology, applied in the field of bacterial species, can solve the problems of unseen Lactococcus lactis and Enterococcus faecalis in Hainan Island geographical population, unstable purification effect, poor purification effect, etc. Significant effect, strong acid-producing effect

Active Publication Date: 2018-07-06
HAINAN UNIVERSITY
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  • Abstract
  • Description
  • Claims
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AI Technical Summary

Problems solved by technology

The strains of Bacillus licheniformis in different growth environments have a good effect on the purification of aquaculture water in the local environment, but they often show poor purification effect and unstable purification effect in the remote environment.
[0006] At present, the domestic patents on lactic acid bacteria microecological preparations mainly focus on fermented strains for fertilizer production and food. There are few patents on Enterococcus faecalis added to breeding feed, and there are no patents on Lactococcus lactis and Enterococcus faecalis. related patents

Method used

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  • Enterococcus faecalis HKF7 with lactic acid activity as well as screening culture method and application thereof
  • Enterococcus faecalis HKF7 with lactic acid activity as well as screening culture method and application thereof
  • Enterococcus faecalis HKF7 with lactic acid activity as well as screening culture method and application thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] The separation and screening method of embodiment 1 Enterococcus faecalis HKF7, mainly comprises the following steps:

[0051] (1) Sample collection

[0052] Collect seawater samples from the waters of Hainan Wenchang Shrimp Farm, and take a small amount of the collected samples for use. The entire operation process is carried out on a purification ultra-clean bench;

[0053] (2) Expanded culture of sample strains

[0054] Take 0.1 mL of the seawater sample collected in step (1) and inoculate it into a centrifuge tube (10 mL / 50 mL, V / V) containing MRS liquid culture medium, 1 tube for each sample, at 30 ° C, 180 r / min, shaker Medium shaking culture for 24h;

[0055] (3) Preliminary isolation of strains

[0056] For the isolation and purification of the strains, the plate streaking method is combined with the coating method. For the strains cultivated in step (2), 0.1 mL of the bacterial liquid was taken and applied to MRS solid medium (containing 1% light CaCO2). 3 ...

Embodiment 2

[0062] Example 2 Molecular identification of Enterococcus faecalis HKF7.

[0063] In order to identify the strains, scanning electron microscope was used to observe the cell morphology and some physiological and biochemical reactions of the tested strains. figure 2 As shown, it can be preliminarily judged that the strain is Enterococcus faecalis.

[0064] The screened strains were identified by PCR using the universal primers for bacteria: the template DNA was extracted according to the operating instructions of the bacterial DNA extraction kit. PCR amplification of the 16srRNA gene fragment of Enterococcus faecalis HKF7 was performed using the upstream primer 5'-AGAGTTTTGATCCTGGCTCA-3' of the conserved 16S rRNA sequence, see SEQ ID No. 2 and the downstream primer 5'-GGTTACCTTGTTACGACTT-3, see SEQ ID No. 3. Amplified, cloned and sequenced. After sequencing, fragments of 1422 bp were obtained respectively. The accession number in GenBank was MF037702. After sequencing, it was...

Embodiment 3

[0065] Example 3 Acid-producing ability test of Enterococcus faecalis HKF7.

[0066] Acid-producing ability test: firstly use the inoculation loop to pick out the target strain of primary screening, inoculate the strain in MRS liquid medium by aseptic operation, and then incubate in a 30°C incubator with shaking at 180 rpm for 24 hours, for use; prepare MRS solid medium ( plus 1% light CaCO 3 ), pour 20 mL of medium into each petri dish, and use the diluted 10 8 100 μL of bacterial liquid of cfu / mL was spread on the solid medium, the culture dish was placed in a 30°C incubator for 48 hours, and the diameter of the dissolved calcium circle was measured with a vernier caliper. It was finally determined that the diameter of the calcified circle of Enterococcus faecalis HKF7 reached 5.00 to 7.00 mm, such as figure 1 shown, by figure 1 The diameter of the calcification circle shows that E. faecalis HKF7 has strong acid-producing ability.

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Abstract

The invention relates to Enterococcus faecalis HKF7 with lactic acid activity as well as a screening culture method and application thereof. The classified name of the bacterial strain is Enterococcusfaecalis HKF7, the Enterococcus faecalis HKF7 was preserved in China Center for Type Culture Collection (CCTCC), the preservation number is CCTCC NO:M2017297, and the preservation date is May 31, 2017. The bacterial strain is separated and screened from Hainan tropical aquaculture seawater and the intestinal tracts of Hainan tropical living litopenaeus vannamei. The Enterococcus faecalis HKF7 provided by the invention does not have hemolytic activity, has high hydrophobic activity and high self-coagulation rate, high artificial gastroenteric tolerance, high cholate tolerance and high bacteriostatic ability. The litopenaeus vannamei is fed with the Enterococcus faecalis HKF7 serving as a feed additive; and through culture test verification, the Enterococcus faecalis HKF7 can improve the immunity of the organism, promotes the growth of the litopenaeus vannamei, has stable effect and is a bacterial strain with good application prospect.

Description

technical field [0001] The invention belongs to the field of strains, and relates to an Enterococcus faecalis. More specifically, the invention relates to an Enterococcus faecalis HKF7 with lactic acid activity, a screening culture method and application thereof. Background technique [0002] In order to pursue high-efficiency and rapid breeding benefits, the breeding density has been continuously increased and the scale of farming has been expanded, resulting in a decrease in the growth rate of aquatic animals, an increase in the bait coefficient, frequent occurrence of common diseases such as vibriosis, and the abuse of chemical fishery drugs such as antibiotics. This not only increases the cost of farming, but also poses a huge threat to the quality and safety of aquatic products and human health. Therefore, finding an efficient, healthy and safe breeding method has become a research focus of current aquaculture technology. [0003] The application of microecological pre...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12N1/02A23K50/80A23K10/18C12R1/01
CPCA23K10/18A23K50/80C12N1/02C12N1/20C12R2001/46C12N1/205Y02A40/818
Inventor 谢珍玉文金顺章翔龙昊
Owner HAINAN UNIVERSITY
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