1株柯乐猪源贝莱斯芽孢杆菌的分离鉴定及生物学特性分析

doi:10.16431/j.cnki.1671-7236.2025.03.044

摘要/Abstract

摘要： 【目的】从柯乐猪粪便中筛选具有优良益生特性的益生菌，以期为筛选畜禽微生态制剂候选菌株提供参考。【方法】采集新鲜柯乐猪粪便，用0.3%胆盐初筛耐胆盐菌株，分离纯化后通过菌落形态和16S rDNA测序进行鉴定，并对分离株的生长特性、耐受特性、产酶特性、药物敏感性以及动物安全性进行分析。【结果】分离筛选获得1株耐胆盐贝莱斯芽孢杆菌，将其命名为W1，该分离株在LB固体培养基上呈圆形、乳白色、不透明，表面皱褶干燥、边缘不整齐；革兰染色镜检可见阳性短杆菌，扫描电镜观察菌体呈短杆状。分离株在LB液体培养基中培养4 h后进入对数生长期，28 h后进入稳定期。将2%接种量的分离株菌液置于人工胃液和人工肠液中处理1 h后，其存活率分别为78.44%和86.62%。分离株在纤维素酶、淀粉酶和蛋白酶筛选培养基中形成的透明圈直径分别为(15.44±2.27)、(25.11±0.69)和(29.69±1.76) mm。药敏试验结果显示，分离株对四环素、卡那霉素、庆大霉素、阿奇霉素、诺氟沙星和环丙沙星表现敏感，对青霉素、阿莫西林和红霉素表现中介。动物安全性试验结果显示，连续20 d灌胃0.1 mL 1×10⁹CFU/mL分离菌液，对照组和试验组小鼠生长状况良好，解剖观察脏器无肉眼可见病变，试验组小鼠平均日增重显著高于对照组(P＜0.05)，两组间血常规指标均无显著差异(P＞0.05)。【结论】分离筛选得到1株生物学特性良好的贝莱斯芽孢杆菌，其对人工胃液和人工肠液具有一定耐受能力，产纤维素酶、淀粉酶和蛋白酶，对小鼠无毒害作用，有望作为畜禽微生态制剂的候选菌株。

关键词: 柯乐猪; 贝莱斯芽孢杆菌; 生物学特性; 微生态制剂

Abstract: 【Objective】 The aim of this study was to screen probiotics with excellent probiotic properties from Kele pigs feces,and to provide reference for screening candidate strains of microecological preparations for livestock and poultry.【Method】 Fresh Kele pigs feces were collected,and bile salt-tolerant strains were initially screened with 0.3% bile salt.After isolation and purification,colony morphology and 16S rDNA sequencing were used to identify the isolates.The biological characteristics of its growth,tolerance characteristics,enzyme production characteristics,drug sensitivity characteristics and animal safety were analyzed.【Result】 A strain of bile salt-tolerant Bacillus velezensis was isolated and screened,named W1.The isolate was round,milky white and opaque on LB solid medium,with a wrinkled and dry surface and untidy edges.Gram-stained microscopic examination showed positive brevibacterium,and the bacteria were short rod shaped by scanning electron microscope.The isolates entered the logarithmic phase after 4 h in LB liquid medium,and entered the stationary phase after 28 h.The survival rate was 78.44% and 86.62%,respectively,after 2% of the inoculated liquid was placed in artificial gastric fluid and artificial intestinal fluid for 1 h.The diameter of transparent circles formed by isolate in cellulase,amylase and protease screening media was (15.44±2.27),(25.11±0.69) and (29.69±1.76) mm,respectively.The results of drug sensitivity test showed that the isolate was sensitive to tetracycline,kanamycin,gentamicin,azithromycin,norfloxacin and ciprofloxacin,and was intermediate to penicillin,amoxicillin and erythromycin.The results of animal safety test showed that the mice in control and the experimental groups were in good growth condition by gavage of 0.1 mL 1×10⁹ CFU/mL isolated bacterial solution for 20 d,and no visible lesions were observed in the organs by anatomy.The average daily gain of mice in experimental group was significantly higher than that of control group (P＜0.05),and there was no significant difference in blood routine indexes between the two groups (P ＞ 0.05).【Conclusion】 A strain of Bacillus velezensis with good biological characteristics was isolated and screened.It had certain tolerance to artificial gastric and intestinal fluids,produced cellulase,amylase and protease,and was not toxic to mice.It was expected to be a candidate strain for livestock and poultry microecological prearations.

Key words: Kele pigs; Bacillus velezensis; biological characteristics; microecological preparation

中图分类号:

S816.73

吴诗卉, 周桂仙, 王敏乐, 廖义潇, 文明, 杨颖. 1株柯乐猪源贝莱斯芽孢杆菌的分离鉴定及生物学特性分析[J]. 中国畜牧兽医, 2025, 52(3): 1437-1446.

WU Shihui, ZHOU Guixian, WANG Minle, LIAO Yixiao, WEN Ming, YANG Ying. Isolation,Identification and Biological Characteristics of a Strain of Bacillus velezensis from Kele Pigs[J]. China Animal Husbandry and Veterinary Medicine, 2025, 52(3): 1437-1446.

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