Isolation And Identification Of A Bacillus Pumilus And Optimization Of Its Fermentation Conditions

With the development of the industrialization, lots of synthetic compounds havebeen discharged into the environment. Some of the synthetic compounds with highlytoxic, carcinogenic effects are hard to been degraded naturally. With the diffusion ofgarbage penetrants, these harmful materials could cause serious pollution togroundwater and soil around. Environment pollution control has been the the problemthat various countries in the world tried to overcome and the most popularmeasurement to deal with the contaminant is chemical, physical treatments combinedwith biological treatments. In recent years, according to different kinds of pollutants,scientists have developed different treatment methods, microbial method of which hasobvious advantage. But as barely garbage classification in the people daily life, it hasthe high concentration of metal ions in the garbage penetrant so that it does need todevelop a strain which has high expression and strong resistance to degrade thegarbage penetrant. Some studies have reported that Bacillus pumilus has the verygood effect on the degradation of waste. As the advantages of high speed ofpropagation, strong resilience and the ability of producing steady spores, Bacilluspumilus has been one of the more and more notable bio-control materials.The external environment has important effect in the growth of Bacillus pumilus.When the external environment is not suitable for growth, the strain would becomeinto spore from the vegetative cell. Moreover, carbon source, nitrogen source andinorganic salts can all affect the growth of bacteria, so it is need to filter the optimumcarbon source, nitrogen source, inorganic salts and best fermentation conditions.This research isolated strain from municipal landfill leachate and their cells showedthe characteristic of rod-shaped and Gram-positive. The16S rRNA gene sequences ishighly conserved. So according to the16S rRNA gene sequences and gyrA gene、rpoAgene of Bacillus pumilus, we design six pairs of specific primers and run the specificPCR reactions. The strain was identified as Bacillus pumilus by PCR products. Thismethod could be easily and quickly identify Bacillus pumilus, avoiding the16S rRNA gene sequences analysis and database comparison.Optimum ingredient of medium for the growth of the strain was determined bysingle factor and orthogonal experiments. The optimized culture medium consists of0.1%of Corn starch,1.0%of Yeast Extract,0.3%of Beef Extract,0.5%of NaCl and0.49%of MgSO4. The best fermentation conditions is37℃,pH7.2and5%ofinoculation quantity. At this condition, the OD320of strains at10h rose from1.44to6.93.It was found that spores were easier to store than vegetative cells and resistant tothe bad environment strongly. In the environment where is suitable for the growth,they could turn to the vegetative cell. The sporulation efficiency was a key factor ofaffecting the microbial agents used. We tried to find a condition with high sporulationefficiency.We examined the effect of Mn2+ion concentration on sporulation efficiency of thestrain. The results showed that0.007%of Mn2+ion had the highest effect onsporulation efficiency. This result provided a foundation for industrial production.