Effects Of Heavy Metal Ions On The Growth Characterization Of Methanotrophic Bacteria

In this paper, the effect of heavy metal pollution on the growth of microorganisms in water body has been briefly summarized. Meanwhile, microorganisms as the molecular recognition elements of microbial sensor are exposed into heavy metal pollutants inevitably in the application of the biosensors practical detection, which affect the accuracy of measuring and the service life of microbial sensor. According to this problem, we selected a methanotrophic ME25 as the research target, which is the molecular recognition element of microbial sensor in the methanol detection. The effects of Pb2+, Cd2+, Hg2+ and their combination on the growth of ME25, and the effects of three metal irons on the protein content, MDH activity, soluble protein electrophoresis and MDH isozyme electrophoresis of this strain were systematically studied by spectrophotometry, SDS-PAGE and isozyme electrophoresis. In addition, a common complexant (EDTA) was used to study the effect of the toxicity of heavy metal ions.Three kinds of metal ions above mentioned remarkably inhibited the growth of ME25. The EC50 of ME25 growth inhibition by Pb2+, Cd2+ and Hg2+ was 34.1 mg·L-1,0.0718 mg·L-1,0.0395 mg·L-1 respectively, and their inhibiting action was Hg2+>Cd2+>Pb2+. Strain ME25 was strong resistance to Pb2+ but relatively sensitive to Cd2+ and Hg2+. In addition, for the effect of growth of ME25, the remarkable antagonistic effects of Pb2+-Cd2+ and Pb2+-Hg2+ were presented. However, the remarkable synergistic effects of Cd2+-Hg2+ and Pb2+-Cd2+-Hg2+ were presented. Experiment also showed that Pb2+, Cd2+ and Hg2+ not only inhibited the biomass of ME25, but also existed inhibiting effect on the soluble protein of this strain. And the effect increased with metal ions concentration increasing.With equivoluminal crude protein of bacterial suspension setting to equivoluminal samples, analysis of soluble protein SDS-PAGE and optical density showed that with the increased concentration of three heavy metal ions, the protein bands of electrophoresis spectrum reduced significantly. Except some lower dose treatment of protein bands were raised, the others all presented downtrend. Results indicated that heavy metal ions inhibited the growth and reproduction of ME25 and decreased the biomass. Besides, the protein content of per unit volume bacterial cells were reduced, so the protein content of every protein bands of electrophoresis spectrum were reduced. In addition, the trend of the three metal ions was consistent. Analysis of unit bacterial protein samples showed that the protein bands of electrophoresis spectrum were almost identical, but relative concentration dramatically changed. Some protein bands raised and some reduced. Three kinds of heavy metal ions lead to different Rf of protein bands which presented as raising and reducing. The results showed that heavy metals affect the expression of bacterial cell protein significantly, and the effects of three heavy metal ions on the protein expression were different.Determination of the methanol dehydrogenase activity showed that, three kinds of metal ions remarkably inhibited the enzyme activity of ME25, and the activity decreased with its concentration increasing. Furthermore, with equivoluminal crude protein of bacterial suspension setting to equivoluminal samples, the analysis of methanol dehydrogenase isozyme electrophoresis and optical density showed that, methanol dehydrogenase isozyme spectrum presented four bands, and the brightness of isozyme bands decreased significantly or even disappeared with the concentration of three heavy metal ions increasing. The analysis of isozyme of unit bacterial protein samples showed that, heavy metal ions on methanol dehydrogenase isozyme had different effects on different bands. The first bands were all raised, while the second, third and the fourth bands expressed as downtrend. Besides, the trend of the three metal ions was almost identical, but relative concentration dramatically changed.These results revealed that methanotrophic ME25 as the molecular recognition elements of microbial sensor are exposed into heavy metal pollutants, which seriously influence the growth of ME25 and its metabolism, thus disturb the accuracy of microbial sensor measuring, and then affect the service life of microbial sensor. ME25 biomass was less influenced by EDTA when its concentration was under 5μm·L-1. Adding EDTA to medium could significantly decrease the heavy metal toxicity on the growth of methanotrophic ME25. These results revealed that adding moderate concentration of complexant in the system of methanol microbial sensor could improve its accuracy of measuring. In addition, this research may pave the way for further study of the masking agent of the heavy metal ions.