Zhang Yunhua, Zhang Zhengyou, Dai Li, Liu Ying, Cheng Maoji, Chen Lijuan
Anhui Province Key Laboratory of Farmland Ecological Conservation and Pollution Prevention, School of Resources and Environment, Anhui Agricultural University, Hefei 230036, China.
College of Animal Science and Technology, Anhui Agricultural University, Hefei 230036, China.
Asian-Australas J Anim Sci. 2018 Jan;31(1):63-70. doi: 10.5713/ajas.17.0018. Epub 2017 Jun 27.
The aim of the study was to isolate gossypol-degrading bacteria and to assess its potential for gossypol degradation.
Rumen liquid was collected from fistulated cows grazing the experimental pasture. Approximately 1 mL of the rumen liquid was spread onto basal medium plates containing 2 g/L gossypol as the only source of carbon and was then cultured at 39°C to isolate gossypol-degrading bacteria. The isolated colonies were cultured for 6 h and then their size and shape observed by microscope and scanning electron microscope. The 16S rRNA gene of isolated colonies was sequenced and aligned using National Center for Biotechnology Information-Basic Local Alignment Search Tool. The various fermentation conditions, initial pH, incubation temperature, inoculum level and fermentationperiod were analyzed in cottonseed meal (CSM). The crude protein (CP), total gossypol (TG), and free gossypol (FG) were determined in CSM after fermentation with isolated strain at 39°C for 72 h.
Screening results showed that a single bacterial isolate, named Rumen Bacillus Subtilis (RBS), could use gossypol as a carbon source. The bacterium was identified by 16S rDNA sequencing as being 98% homologous to the sequence of strain GH38. The optimum fermentation conditions were found to be 72 h, 39°C, pH 6.5, moisture 50%, inoculum level 10 cell/g. In the optimum fermentation conditions, the FG and TG content in fermented CSM decreased 78.86% and 49% relative to the control. The content of CP and the essential amino acids of the fermented CSM increased respectively, compared with the control.
The isolation of a gossypol-degrading bacterium from the cow rumen is of great importance for gossypol biodegradation and may be a valuable potential source for gossypol-degradation of CSM.
本研究旨在分离棉酚降解菌，并评估其降解棉酚的潜力。
从在实验牧场放牧的带瘘管奶牛采集瘤胃液。将约1 mL瘤胃液涂布于以2 g/L棉酚作为唯一碳源的基础培养基平板上，然后在39°C培养以分离棉酚降解菌。将分离出的菌落培养6 h，然后通过显微镜和扫描电子显微镜观察其大小和形状。使用美国国立生物技术信息中心的基本局部比对搜索工具对分离菌落的16S rRNA基因进行测序和比对。在棉籽粕中分析各种发酵条件、初始pH、培养温度、接种量和发酵时间。在39°C用分离菌株发酵72 h后，测定棉籽粕中的粗蛋白（CP）、总棉酚（TG）和游离棉酚（FG）。
筛选结果表明，一种名为瘤胃枯草芽孢杆菌（RBS）的单一细菌分离株能够利用棉酚作为碳源。通过16S rDNA测序鉴定该细菌与菌株GH38的序列同源性为98%。发现最佳发酵条件为72 h、39°C、pH 6.5、水分50%、接种量10个细胞/g。在最佳发酵条件下，发酵棉籽粕中的FG和TG含量相对于对照分别降低了78.86%和49%。与对照相比，发酵棉籽粕的CP含量和必需氨基酸含量分别增加。
从奶牛瘤胃中分离出棉酚降解菌对于棉酚生物降解具有重要意义，可能是棉籽粕棉酚降解的宝贵潜在来源。
