瘤胃源粪臭素降解菌的分离鉴定及其降解特性研究

doi:10.13560/j.cnki.biotech.bull.1985.2023-0934

摘要/Abstract

摘要：

【目的】 旨在从绵羊瘤胃中分离粪臭素降解菌，并评估其粪臭素降解能力和生长性能，以期开发适用于反刍动物降臭的直接饲喂微生物。【方法】 以绵羊瘤胃液为分离来源，使用含有粪臭素的MSM培养基进行富集和分离，通过细菌菌落形态观察进行初步分类；应用16S rRNA基因扩增、测序及系统发育分析进行物种鉴定；绘制菌株生长曲线，通过HPLC技术测定粪臭素降解曲线。【结果】 从绵羊瘤胃液中分离出25株粪臭素降解菌，经菌落形态鉴定选出11株代表菌株进行后续研究。物种鉴定结果显示，MSML2和MSML6属于枯草芽孢杆菌（Bacillus subtilis），MSML4、MSML5、MSML7和MSML10属于阿氏普里斯特氏菌（Priestia aryabhattai），MSML3和MSML11属于污染伯克霍尔德氏菌（Burkholderia contaminans），MSML1、MSML8和MSML9属于成都假单胞菌（Pseudomonas chengduensis）。其中，MSML5生长速度最快，在约12 h后进入稳定期，稳定期菌体浓度最高；MSML3和MSML8在前16 h生长缓慢，32 h后进入稳定期。在粪臭素降解方面，MSML2粪臭素降解效率最高，48 h内的降解率达到23.03%，其次是MSML7、MSML8和MSML10，降解率均超过20%。【结论】 成功从绵羊瘤胃中分离获得25株粪臭素降解菌，涉及4个物种，首次报道了具备粪臭素降解能力的枯草芽孢杆菌、阿氏普里斯特氏菌和成都假单胞菌，为直接饲喂反刍动物的菌剂开发提供了宝贵的菌株资源。

关键词: 瘤胃, 粪臭素降解率, 生物降解, 分离鉴定, 枯草芽孢杆菌, 直接饲喂微生物

Abstract:

【Objective】 The aim of this study is to isolate skatole-degrading bacteria from the rumen of sheep and estimate its skatole-degrading ability and growth performance, to develop suitable direct-fed microbials for ruminants to reduce odor.【Method】 The MSM medium containing skatole was used for enrichment and isolation from the rumen fluid of sheep. The bacterial colony morphology was used for primary classification. Amplification, sequencing and phylogenetic analysis of 16S rRNA gene were carried out for further identification. The growth curve was drawn, and degradation curve of skatole was determined using HPLC.【Result】 A total of 25 skatole-degrading strains were obtained, and 11 representative strains（MSML1-11）of them were selected based on colony morphology for further study. MSML2 and MSML6 belonged to Bacillus subtilis, MSML4, MSML5, MSML7 and MSML10 belonged to Priestia aryabhattai, MSML3 and MSML11 belonged to Burkholderia contaminans. MSML1, MSML8 and MSML9 belonged to Pseudomonas chengduensis. Among these strains, MSML5 grew at the fastest rate. Its stable phase appeared after 12 h, and the bacterial concentration in the stable phase was the highest. MSML3 and MSML8 grew slowly in the first 16 h and reached a stable phase after about 32 h. In terms of skatole degradation, MSML2 showed the highest skatole degradation efficiency of 23.03% within 48 h, followed by MSML7, MSML8 and MSML10, with a degradation rate higher than 20%.【Conclusion】 This research successfully isolated skatole-degrading bacteria from the rumen of sheep, including four different species. Bacillus subtilis, Priestia aryabhattai and Pseudomonas chengduensis with skatole-degrading ability are reported for the first time, which provides valuable resources for the development of microbial agents directly fed to ruminants.

Key words: rumen, skatole degrading efficiency, biodegradation isolation and identification, Bacillus subtilis, direct-fed microbials（DFM）

王璐, 刘梦雨, 张富源, 纪守坤, 王云, 张英杰, 段春辉, 刘月琴, 严慧. 瘤胃源粪臭素降解菌的分离鉴定及其降解特性研究[J]. 生物技术通报, 2024, 40(3): 305-311.

WANG Lu, LIU Meng-yu, ZHANG Fu-yuan, JI Shou-kun, WANG Yun, ZHANG Ying-jie, DUAN Chun-hui, LIU Yue-qin, YAN Hui. Isolation and Identification of Rumen Skatole-degrading Bacteria and Analysis on Their Degradation Characteristics[J]. Biotechnology Bulletin, 2024, 40(3): 305-311.

图/表 4

图1 代表性瘤胃源粪臭素降解菌株菌落形态

Fig. 1 Colony morphology of representative rumen skatole-degrading bacteria

图2 瘤胃源粪臭素降解菌的系统进化树

Fig. 2 Phylogenetic tree of rumen skatole-degrading bacteria

图3 粪臭素降解菌生长和降解曲线

Fig. 3 Growth and skatole degradation curves of skatole-degrading isolates

图4 不同菌株48 h粪臭素降解率

Fig. 4 Skatole degradation rate by different isolates at 48 h

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