鸡源凝结芽孢杆菌的分离鉴定及耐受性分析

doi:10.16431/j.cnki.1671-7236.2022.11.009

中国畜牧兽医 ›› 2022, Vol. 49 ›› Issue (11): 4187-4196.doi: 10.16431/j.cnki.1671-7236.2022.11.009

鸡源凝结芽孢杆菌的分离鉴定及耐受性分析

孙彦婷¹, 康宇¹, 詹妍¹, 张冬萱¹, 刘畅², 边传周¹, 张晓静², 白静², 乔宏兴¹

1. 河南牧业经济学院动物医药学院, 郑州 450046;
2. 河南省微生物生物转化中药工程实验室, 郑州 450046

收稿日期:2022-04-15 出版日期:2022-11-05 发布日期:2022-11-04
通讯作者: 乔宏兴 E-mail:zzmzqhx@163.com
作者简介:孙彦婷,E-mail:yxytsun2101@126.com。
基金资助:
河南省重点研发项目（221111111300）；河南省重点研发与推广专项项目（202102110247、192102110075）；河南省教育厅高等学校重点科研计划项目（16A230007）；河南牧业经济学院博士启动基金项目（53000212、2021HNUAHEDF004）；河南牧业经济学院教育教学改革研究与实践项目（2021-XJLX-139）；河南牧业经济学院预防兽医学科团队建设项目（MXK2016102）；河南牧业经济学院科技创新团队项目（2018KYTD13、2018KYTD16）；河南省现代农业产业技术体系团队项目（HARS-22-12-G2）；河南省高等学校教学名师项目（教高[2020]434号）

Isolation,Identification and Stress Resistance Analysis of Bacillus coagulans from Chickens

SUN Yanting¹, KANG Yu¹, ZHAN Yan¹, ZHANG Dongxuan¹, LIU Chang², BIAN Chuanzhou¹, ZHANG Xiaojing², BAI Jing², QIAO Hongxing¹

1. College of Veterinary, Henan University of Animal Husbandry and Economics, Zhengzhou 450046, China;
2. Henan Province Microbial Biotransformation Traditional Chinese Medicine Engineering Laboratory, Zhengzhou 450046, China

Received:2022-04-15 Online:2022-11-05 Published:2022-11-04

摘要/Abstract

摘要： 【目的】试验旨在寻找1株性能稳定、环境耐受性强的凝结芽孢杆菌菌株，以在实际生产中发挥其替抗功能。【方法】本研究采集160日龄健康海兰褐蛋鸡的粪便样本，进行细菌分离纯化，并对分离菌进行形态学特征观察、生理生化鉴定、16S rDNA测序分析。通过绘制该菌的生长动力曲线确定其生长规律；利用耐酸、耐胆盐试验分析该菌株的耐受性；通过单因素试验对分离菌株的培养条件进行优化。【结果】分离菌在普通培养基上长出表面粗糙、不规则的灰白色菌落，镜检为杆状、芽孢端生的革兰氏阳性菌。结合生化试验及16S rDNA基因序列分析综合鉴定该菌株为凝结芽孢杆菌，命名为NJ001。生长动力曲线表明，7~24 h为该菌株的对数生长期，24 h活菌数达到顶峰，凝结芽孢杆菌NJ001的最佳培养时间为24 h；耐受性试验结果显示，该菌株pH 3.0温育2 h存活率为59.5%，pH为4.0时存活率达到87.6%以上，对酸有较强耐受力；0.3%胆盐处理2 h存活率为54.0%，胆盐浓度升至0.5%时存活率仍有50%以上，该菌对胆盐有较强的耐受性。单因素试验结果发现，该菌的最佳培养温度为40 ℃，接种量2%，初始pH 7.0，摇床转速220 r/min。综合以上最佳培养条件，菌株活菌数可达最大值，为1.93×10⁸ CFU/mL。【结论】所分离的菌株为具有较强耐酸耐胆盐特性的凝结芽孢杆菌，其最适生长条件易于实现；且与动物体内胃肠道环境基本吻合，具备作为微生态制剂的应用潜力。

关键词: 鸡; 凝结芽孢杆菌; 分离鉴定; 耐受性

Abstract: 【Objective】 The aim of this experiment was to find a Bacillus coagulans strain with stable performance and strong environmental resistance, so as to play its role instead of antibiotics in actual production.【Method】 The fecal samples from 160-day old healthy Hyline Brown laying hens were collected for isolation and purification of bacteria, and morphological characteristics observation, physiological and biochemical identification, 16S rDNA sequencing analysis and identification of the isolated bacteria were carried out.The growth rule was determined by plotting the growth dynamic curve of the bacterium.Acid and bile salt resistance tests were used to analyze the resistance of the strain.The culture conditions of isolated strains were optimized by single factor experiment.【Result】 The isolated bacteria grew gray white colonies with rough and irregular surface on the ordinary culture medium, and were rod-shaped and spore terminal Gram-positive bacteria under microscope.Combined with biochemical experiments and 16S rDNA sequence analysis, the strain was identified as Bacillus coagulans, named NJ001.The growth dynamic curve showed that 7-24 h was the logarithmic growth period of the strain, and the number of viable number reached the peak at 24 h.The optimal culture time of Bacillus coagulans NJ001 was 24 h.The results of tolerance test showed that the survival rate was 59.5% at pH 3.0 for 2 h, and 87.6% at pH 4.0, showing strong resistance to acid.The survival rate was 54.0% after treatment with 0.3% bile salt for 2 h, and the survival rate was still more than 50% when the concentration of bile salt increased to 0.5%.The results of single factor experiment showed that the optimum culture temperature was 40 ℃, inoculation amount 2%, initial pH 7.0 and shaking speed was 220 r/min.Based on the above optimum cultivation conditions, the maximum viable count of the strain was 1.93×10⁸ CFU/mL.【Conclusion】 The isolated strain was Bacillus coagulans with strong acid and bile salt resistance, and its optimal growth conditions which was basically consistent with the gastrointestinal environment of animals were easy to achieve, so it had the potential of application as a microecological agent.

Key words: chicken; Bacillus coagulans; isolation and identification; stress reistance

中图分类号:

S852.61⁺6

孙彦婷, 康宇, 詹妍, 张冬萱, 刘畅, 边传周, 张晓静, 白静, 乔宏兴. 鸡源凝结芽孢杆菌的分离鉴定及耐受性分析[J]. 中国畜牧兽医, 2022, 49(11): 4187-4196.

SUN Yanting, KANG Yu, ZHAN Yan, ZHANG Dongxuan, LIU Chang, BIAN Chuanzhou, ZHANG Xiaojing, BAI Jing, QIAO Hongxing. Isolation,Identification and Stress Resistance Analysis of Bacillus coagulans from Chickens[J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(11): 4187-4196.

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鸡源凝结芽孢杆菌的分离鉴定及耐受性分析

Isolation,Identification and Stress Resistance Analysis of Bacillus coagulans from Chickens

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