| Manganese-oxidizing microbes widely exist in the water and soil environment. They are able to oxidize Mn2+ into Mn4+ efficiently and therefore play an important role in the biogeochemical cycling of manganese. Many manganese-oxidizing bacteria can strongly catalyze the oxidation of soluble Mn(II) to generate insoluble manganese oxides, and the production rate of Mn(IV) is much faster than chemical catalysts (up to 100000 times). Therefore, manganese-oxidizing bacteria have promising application prospects in the biological manganese removal research and in treating of high manganese content groundwater.This study mainly focuses on the research of manganese-oxidizing microbes isolated from iron-manganese nodules in soil. Firstly,44 manganese-oxidizing bacteria were isolated from the iron-manganese nodules samples collected from different soil profile throughout the country. Secondly, eight isolates were identified through 16S rDNA sequencing analysis after observing their morphological characteristics. The identification results showed that all these strains belonged to the genus Bacillus, including three strains of B. cereus, one strain of B. thuringiensis, one strain of B. megaterium, one strain of B. pumilus and two other specieses of the genus Bacillus. Because the overall topologies of the MnxG (The gene directly involved in Mn(II) oxidation in Bacillus spp.)-and 16S rDNA-based phylogenetic trees were remarkably similar, the phylogenetic analysis of these strains and the known marine manganese-oxidizing Bacillus spp. was carried out to determine the genetic relationship between the isolates and other known marine manganese-oxidizing Bacillus spp., furtherly study whether the Mn(II) oxidation process between the isolates and the known marine manganese-oxidizing Bacillus spp. has a similar molecular mechanism or not. Meanwhile, the manganese oxidation activity of the 44 isolates and the previous manganese-oxidizing strain of the project was measured. In view of Bacillus sp.WH4 with the highest manganese oxidation activity would be a promising application candidate for the cleaning of water resources polluted by manganese, its fermentation medium was optimized on the basis of determining their growth characteristics.The measuring results of the growth characteristics of Bacillus sp.WH4 indicated that 1 mmol/L of manganese did not effect the growth of Bacillus sp.WH4, and the more suitable temperature and pH was 28℃and 7.0, respectively. The growth curve of the strain showed that it spended 29 h entering logarithmic growth phase. And its more suitable cultivating volume is 50 mL in a 250 mL conical flask.In this study, the fermentation medium for Bacillus sp.WH4 was optimized by response surface methodology (RSM) using the viable count in fermentation broth as an indicator. Firstly, based on the initial fermentation medium, the optimal carbon source and nitrogen sources were selected by the single-factor test, respectively. Secondly, four significant factors, soybean meal, KH2PO4, MgSO4·7H2O and FeSO4·7H2O, were screened out by Plackett-Burman (PB) design. Thirdly, the maximum response region for the Central Composite Design (CCD) was determined through the method of steepest ascent. Finally, the optimal concentration for each significant factor was determined using CCD and RSM analysis. The optimized fermentation medium contained sucrose 2, soybean meal 0.96, K2HPO4 0.8, KH2PO4 0.19, MgSO4·7H2O 0.11, CaCl2 0.02, NaCl 0.5 and FeSO4·7H2O 0.02 (w/%). The results of the fermentation test confirmed that the viable count of Bacillus sp.WH4 in the optimized fermentation medium was 4.2×109 CFU·mL-1, which is nearly four times than that in the basic fermentation medium.
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