A Novel Technique For Rapid Selection Of MCs-degrading Bacteria And The Study On The Degradation Efficiency Of Isolated Strains

Currently, biodegradation of microcystin by bacteria has being one of the most promising methods for removing MCs from water. The process of isolation and identification are complicated and it will spend a lot of time and money. In our study, we found the conserved sequence of microcystin degrading genes mlrA by comparing and analyzing all mlrA genes on NCBI. Then we designed a pairs of specific primers to select the degrading bacteria by polymerase chain reaction( PCR) methods.Based on rapid detection of the conserved sequence, the microcystin-degrading bacteria would be selected. We isolated and purified eight strains from rotten cyanobacterias of Taihu Lake, then the mlrA gene of each strain were detected by PCR. The results of PCR showed that five strains named A1, A2, A6, Q1 and Q3 in our lab contained the target gene of mlrA. In order to verify the reliability of this method, the primers designed by other scholars were used to amplify mlrA gene in these eight strains. It was found that the five strains possess the mlr A gene encoding the most important enzyme for MCs degradation, while it couldn’t be detected in the other three strains. In addition, the five strains were capable of degrading MCs at the rate of 25.4%, 23.6%, 27.1%, 20.7% and 22.1% separately, and other three strains almost had no degradation. The results of biodegradation and mlrA gene amplification indicated that the method of PCR can be used to select the MCs-degrading bacteria rapidly and simply.Based on morphological, physiological characteristics and analysis of the 16 S rRNA gene sequence, the five MCs-degrading strains were identified as Kurthia gibsonii, Catellibacterium terrae, Acinetobacter lwoffii, Bacillus cereus and Brevundimonas diminuta.The five MCs-degrading strains were used to investigate their degradation ability of standard MC-LR. It was found that these five strains can use MC-LR as the sole carbon and energy sources. All of them were grow fast in two days after inoculation. The MC-LR was degraded by five strains at the rate of 59.9%, 62.5%, 57.8%, 64% and 61.5% separately within seven days. This is the first study to report that Kurthia gibsonii and Brevundimonas diminuta could degrade MCs. In our study, the five isolated strains shown to possess a homologous mlrA gene responsible for degrading MCs, this finding suggest that mlrA gene may also exist in Non-Sphingomonadaceae.We researched the effect of intracellular and extracellular extracts on the degradation of MC-LR in five strains. The result showed that both intracellular and extracellular extracts were capable of degrading MCs. However, the degradation rates of intracellular extracts were better than extracellular extracts in each bacterium. The degradation rates of intracellular extracts were 48.1%, 32.5%, 11.6%, 40.3% and 32.5% respectively. To date, only one biodegradation pathway for microcystin has been reported, and three intracellular enzymes play a crucial role in this pathway. Accordingly, the previous results suggest that these five strains might degrade MCs through a similar pathway which researched by Dr. Bourne.