对虾养殖水体中一株降解氨氮芽孢杆菌的分离鉴定及其发酵配方的优化

Isolation and identification of a strain of probiotic Bacillus from a breeding outdoor pond of shrimp (Litopenaeus vannamei) and optimization of its fermentation formula

本实验在(28±1)℃下从对虾养殖池塘中分离筛选具有氨氮(包括游离氨NH 3 、铵盐NH4^+)降解效果的芽孢杆菌( Bacillus ),并采用单因素实验对筛选出的芽孢杆菌进行发酵配方优化,以此挑选出低廉高产的发酵培养基配方。菌株分离筛选及氨氮降解实验结果显示:经初步分离,得到5株芽孢杆菌(DS1、DS2、DS3、DS4、DS5),再通过氨氮降解实验得到一株能够显著降解水体氨氮的芽孢杆菌DS5,5 d内菌株DS5对水体氨氮降解迅速,在第4 天达到最大降解率(36.41±0.07)%,且最后3 d水体氨氮持续稳定在36%左右;通过菌株16S rDNA序列分析及生理生化鉴定,将菌株DS5(GenBank登录号:MK629979)初步鉴定为枯草芽孢杆菌( Bacillus subtilis )。菌株DS5发酵配方实验结果表明:在28℃下,当碳源为5‰葡萄糖、氮源为10‰豆粕、无机盐为5‰氯化钠时,菌株生长良好,培养基含菌数较高,综合考虑成本、菌含量等因素,选择5‰葡萄糖-10‰豆粕-5‰氯化钠作为该菌的最优发酵培养基配方。

In this experiment, Bacillus with the degradation effect of ammonia nitrogen,including free ammonia (NH 3 -N) and ammonium salt (NH4^+),was isolated and screened from a breeding outdoor pond of shrimp ( Litopenaeus vannamei ) under(28±1)℃.The single factor experiment was used to optimize the fermentation formula of the screened Bacillus and select the low cost and high yield fermentation medium formula.The results of strain isolation and screening,and ammonia nitrogen degradation experiment showed that:after preliminary isolation,five strains of Bacillus (DS1,DS2,DS3,DS4 and DS5) were obtained,and then one strain of Bacillus DS5 that could significantly degrade ammonia nitrogen in water was obtained through ammonia nitrogen degradation experiment.Within five days,the DS5 strain rapidly degraded the ammonia nitrogen in water,reaching the maximum degradation rate (36.41±0.07)% on the 4th day,and the ammonia nitrogen in the last 3 days water remained stable at about 36%.The strain DS5 was preliminarily identified as Bacillus subtilis by 16S rDNA sequence analysis as well as physiological and biochemical identification.Fermentation formula experimental results of strain DS5 showed that under 28℃,when carbon source adding 5‰ glucose,nitrogen source adding 10‰ soybean meal and inorganic salt adding 5‰ sodium chloride,the strain grew well,and the number of bacteria in culture medium was higher than others.To consider the cost and the amount of bacteria,5‰ glucose-10‰ soybean meal-5‰ sodium chloride was selected as the optimum fermentation medium for this strain.