一株降解牛血液蛋白菌株的分离鉴定及其降解条件优化

Isolation and Identification of a Bovine Blood Protein Degradation Strain and Preliminary Optimization of Its Degradation Conditions

为了解决屠宰场废弃血液造成的环境污染及其有效利用问题,取样自日喀则地区屠宰场的废弃血液堆积土壤,梯度稀释涂布于血琼脂平板上进行分离,挑选有较大溶血环的菌落纯化,继而保藏菌种。对保藏菌株进行形态学、生理生化、16S rRNA基因序列鉴定并进行药敏试验、菌株生长条件的探究。用福林酚法测定保藏菌株的蛋白酶活力,初步优化血液培养基条件,提升保藏菌株血液降解效率。结果表明,得到了一株高效降解血液蛋白的菌株,编号为NWMCC0047,经过形态学鉴定、生理生化鉴定和16S rRNA基因序列鉴定,表明该菌株为乙酰微小杆菌(Exiguobacterium acetylicum)。其蛋白酶活力可达19.00 U/mL。最适生长条件:碳源为葡萄糖、氮源为牛肉膏、无机盐为磷酸氢二钠、pH值为7、温度为20℃。通过菌株对牛血液蛋白降解条件的初步优化,当血液添加量为10%(体积分数)、麦麸添加20 g/L、不添加无机盐、37℃培养85 h时,其降解率可达21.97%。实验为降解废弃血污提供了简单可参考的操作方法且为生产氨基酸肥料提供候选菌株。

In order to solve the problem of environmental pollution and effective utilization of waste blood in slaughterhouses,the samples were collected from waste blood accumulation soil of slaughterhouses in Shigatse area,and the samples were diluted and coated on blood agar plate for separation.The colonies with large hemolytic rings were selected for purification,and then the strains were preserved.Morphology,physiology and biochemistry,16 S rRNA gene sequence identification,drug sensitivity test and growth conditions of the preserved strains were studied.The protease activity of preserved strains was determined by Folin phenol method,and the blood culture medium conditions were preliminarily optimized to improve the blood degradation efficiency of the preserved strains.The results showed that a strain with high efficiency in degrading blood protein was obtained,and numbered as NWMCC0047.After morphological identification,physiological and biochemical identification,and 16 S rRNA gene sequence identification,the strain was identified as Exiguobacterium acetylicum.Its protease activity reached up to 19.00 U/mL.The optimum growth conditions were as follows:glucose as carbon source,beef extract as nitrogen source,disodium hydrogen phosphate as inorganic salt,pH 7,and temperature 20℃.Through the preliminary optimization of the degradation conditions of bovine blood protein,the degradation rate reached 21.97% when the amount of blood was 10%,wheat bran was 20 g/L,inorganic salt was not added,and cultured at 37℃ for 85 h.The experiment provides a simple and referable operation method for the degradation of waste blood pollution and provides candidate strains for the production of organic amino acid fertilizer.