野生黑鸢源芽孢杆菌的分离鉴定及生物学特性研究

doi:10.16431/j.cnki.1671-7236.2025.11.043

摘要/Abstract

摘要： 【目的】从野生黑鸢粪便中分离鉴定耐药更少的芽孢杆菌，并测定其潜在益生性，为动物营养与饲料微生态制剂提供候选菌株。【方法】从新疆野生动物救助站采集黑鸢的新鲜粪便样品，用LB琼脂培养基分离芽孢杆菌，对分离株进行形态学观察、生化鉴定和16S rDNA测序分析，进一步测定分离菌株的耐酸耐胆盐能力、自聚集性、DPPH自由基清除率、药物敏感性、溶血活性和抑菌能力，评估其体外益生能力。【结果】从野生黑鸢粪便中分离得到3株菌，分离菌株在LB琼脂培养基上呈淡黄色褶皱状单菌落，表面粗糙，湿润，为革兰阳性杆状细菌，分别命名为C42a、C45和C5y1。生化鉴定结果显示，3株分离菌V-P、D-木糖、L-阿拉伯糖、D-甘露醇、明胶液化、pH 5.7、7%氯化钠、硝酸盐还原、淀粉水解试验结果呈阳性；柠檬酸盐和丙酸盐试验结果呈阴性。16S rDNA测序结果显示，C42a与地衣芽孢杆菌PP236929.1、C45与地衣芽孢杆菌MF581456.1、C5y1与地衣芽孢杆菌HQ850703.1的相似性均＞97%，确定3株分离菌均为地衣芽孢杆菌。3株菌在pH为2.0时存活率高于32.00%，最高可达75.30%；在0.3%胆盐浓度下均可存活。自聚集能力均在52.22%～88.89%之间，DPPH自由基清除率在63.62%～84.18%之间。药敏试验结果显示，分离菌C42a、C45、C5y1对氨苄西林、亚胺培南、阿米卡星、庆大霉素等11种药物均表现敏感；C42a对红霉素中介，C5y1对克林霉素中介，C45对红霉素和克林霉素均中介。3株菌均不具有溶血活性。C42a和C45对沙门菌和金黄色葡萄球菌有较好的抑制作用。【结论】本研究从野生黑鸢粪便中分离出3株耐药较少的地衣芽孢杆菌C42a、C45、C5y1，均表现出良好的体外益生特性，可作为动物微生态制剂的候选菌株。

关键词: 黑鸢; 地衣芽孢杆菌; 分离鉴定; 生物学特性

Abstract: 【Objective】 This study aimed to isolate and identify Bacillus strains with lower antibiotic resistance from the feces of wild Milvus migrans,and determine the potential beneficial effects,so as to provide candidate strains for animal nutrition and feed microecological preparations. 【Method】 Fresh faecal samples of wild Milvus migrans were collected from Xinjiang wildlife rescue station,and Bacillus was isolated with LB agar medium.Morphological observation,biochemical identification and 16S rDNA sequencing were performed.The ability of acid and bile salt resistance,self-aggregation,DPPH free radical clearance,drug sensitivity,hemolytic activity and antibacterial ability of the isolated strains were further determined to evaluate their in vitro probiotic properties. 【Result】 Three strains of bacteria were isolated from the feces of wild Milvus migrans.On LB agar medium,the isolated strains were light yellow folded single colony with rough and wet surface.They were Gram-positive rod-shaped bacteria,named C42a,C45 and C5y1, respectively.The biochemical identification results showed that the test results of V-P,D-xylose, L-arabinose,D-mannitol,gelatin liquefaction,pH 5.7,7% sodium chloride,nitrate reduction and starch hydrolysis of the three isolated bacteria were positive,and the results of the citrate and propionate tests were negative.16S rDNA sequencing results showed that the similarities between C42a and Bacillus licheniformis PP236929.1,C45 and Bacillus licheniformis MF581456.1,C5y1 and Bacillus licheniformis HQ850703.1 were all ＞97%,and the three isolates were all identified as Bacillus licheniformis.At pH 2.0,the survival rate of the three strains was higher than 32.00% and the highest was 75.30%.It could survive at a bile salt concentration of 0.3%.The self-aggregation capacity was in the range of 52.22% to 88.89%,and the DPPH free radical clearance was in the range of 63.62% to 84.18%.The results of drug sensitivity test showed that C42a,C45 and C5y1 were sensitive to 11 drugs including ampicillin,imipenem,amikacin,gentamicin and so on.C42a was moderately sensitive to erythromycin,C5y1 was moderately sensitive to clindamycin,and C45 was moderately sensitive to erythromycin and clindamycin.The three strains did not have hemolytic activity.C42a and C45 showed good inhibitory effect on Salmonella and Staphylococcus aureus. 【Conclusion】 In this study,three strains of Bacillus licheniformis C42a,C45 and C5y1 with lower antibiotic resistance were isolated from the feces of wild Milvus migrans,which showed good in vitro probiotics and could be used as candidate strains for animal microecological preparations.

Key words: Milvus migrans; Bacillus licheniformis; isolation and identification; biological characteristics

中图分类号:

S831.5

谢怀茹, 潘雨涵, 林峻, 石蕊, 王旭翔, 戴宗茂, 靳茜, 地丽热巴·地里夏提, 阿依乔丽凡·沙汉, 彭啟敏, 肖莹, 麦多, 王金泉. 野生黑鸢源芽孢杆菌的分离鉴定及生物学特性研究[J]. 中国畜牧兽医, 2025, 52(11): 5488-5498.

XIE Huairu, PAN Yuhan, LIN Jun, SHI Rui, WANG Xuxiang, DAI Zongmao, JIN Qian, Dilraba DILIXIATI, Ayiqiaolifan SHAHAN, PENG Qimin, XIAO Ying, MAI Duo, WANG Jinquan. Isolation,Identification and Biological Characteristics of Bacillus Derived from Wild Milvus migrans[J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(11): 5488-5498.

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