Study On Mercury Resistance And Enrichment Behavior Mechanism Of A Marine Organism, Pseudomonas Aeruginosa

Pseudomonas pseudoalcaligenes A1 is a marine bacterium with high resistance to mercury. Strain A1 is able to grow in the presence of 120mg/L Hg2+, and its maximum Hg2+ accumulation capacity comes up to 179.533 mg/g. Transcriptome analysis of A1 with/without Hg2+ stress(A11/A10) was carried out in order to explore it's mechanism of high mercury resistance. After the functional annotation of the genes, the GO and KEGG Pathway enrichment analysis was performed. The results showed that 2695 Unigenes can be annotated to the NR, NT, Swiss-Prot, KEGG, COG and GO database. In addition, 21.5% of the annotated genes are annotated to genes of other species and most of them can be annotated to Pseudomonas. The number of differentially expressed genes induced by Hg2+ is 472, including 262 up-regulated genes and 165 down-regulated genes.Unigene97-All and CL78.Contig1-All are greatly-differentially expressed genes correlating with the resistance to heavy metals. The blast comparison revealed that CL78.Contig1-All and Unigene97-All are the genes coding heavy metal efflux pump Czc A and mercury reductase Mer A, respectively. CzcA is a heavy metal efflux pump that can pump heavy metals out of the cells, thus reducing the toxicity of heavy metals. The protein coded by Unigene 97-All showed a potential functioning domain for Hg2+ transport and reduction. Blast alignment revealed that Unigene 97-All coding protein is quite similar to Mer A, a well-known mercury reductase existing in most of the Hg-resistant microorganisms. This finding showed that protein coded by Unigene 97-All might be mercury reductase.Genetic engineering technology was applied to study whether Unigene97-All and CL78.Contig1-All are heavy metal resistance genes. After the two genes were respectively cloned into p ET-28 a and then transformed into E.coli BL21(DE3),the recombinant strains U97 and CL78 were constructed successfully.Then the mercury resistance and quantity of mercury equilibrium accumulation of each recombinant strain were evaluated. The results showed that both U97 and CL78 were able to grow in the presence of 6mg/L and 2mg/L Hg2+, respectively, inferring resistance to mercury.There was no obvious difference in Hg2+ accumulation among the recombinant strains and the negative control group. The maximum ccumulation capacities of U97, CL78 and negative control group E.coli BL21 reached 85.47mg/g, 89.28mg/g and 90.09mg/g, respectively, suggesting these two resistance genes did not improve Hg2+ accumulation capacities of the recombinant strains. Therefore we can draw a conclusion that there is no direct relation between heavy metal resistance and accumulation capacity for bacteria.