| This thesis contained two research aspects. The first part is the polyphasic taxonomy identification of strain TS3 which was isolated from heavy metal contaminated soil. The second part is the isolation of soil manganese resistant bacterial strains and identification of their manganese oxidation abilities.A bacterial isolate, designated strain TS3, was isolated from soil samples collected from a metal mine in Tieshan District, Daye City, Hubei Province. It showed highly resistance levels to multi metal(loid)s, particularly for Cu. Cells of this strain were Gram-negative, rod-shaped (0.3-0.6×1.2-2.0μm) and motile with flagella. Aerobic growth can be observed at 10-40℃, with the optimum temperature at 28℃. The pH range for growth is 5.0-9.0, with the optimum pH at 7.0-7.5. The strain had ubiquinone Q-8 as the predominant respiratory quinone, phosphatidylethanolamine (PE), phosphatidylglycerol (PG) and diphosphatidylglycerol (DPG) as the major polar lipids and summed feature 3 (comprising C16:1ω7c and/or iso-C15:0 2-OH,40.3%), C16:0 (21.1%) and C18:1ω7c (15.2%) as the major fatty acids. The G+C content was 65.9 mol%. Analysis of the 16S rRNA gene sequence indicated that strain TS3T showed high sequence similarities with Massilia niastensis 5516S-1T (KACC 12599T) (98.5%), Massilia consociata CCUG 58010T (97.6%), Massilia aerilata 5516S-111 (KACC 12505T) (97.4%) and Massilia varians CCUG 35299T (97.2%). The DNA-DNA hybridization values revealed low relatedness between strains TS3T, and M. niastensis KACC 12599T (36.5%), and M. consociata CCUG 58010T (27.1%), and M. aerilata KACC 12505T (22.7%), and M. varians CCUG 35299T (46.5%). On the basis of phenotypic and phylogenetic characteristics, strain TS3T is affiliated to genus Massilia, but is clearly differentiated from other species and represents a novel member, for which the name Massilia tieshanensis sp. nov. is proposed. The type strain is TS3T (=CCTCC AB2010202T=KACC 14940T). The main innovation of this section is the discovery and indetification of a novel bacterial species. Since this strain was isolated from heavy metal contaminated areas, this finding is significant for taxonomic research of metal-resistant bacteria.Manganese resistant bacteria (18 strains) were isolated from the soil collected from Queyu, Shandong Province. A total of 11 different bacterial strains were gotten after PCR-RFLP analysis. We extracted the DNA of these bacteria, performed PCR amplification of 16S rRNA genes, then compared the sequence similarities in the EzTaxon Server 2.1 database. We found that these strains belonged to Bacillus (5 strains), Pseudomonas (1 strain), Paracoccus (2 strains), Cupriavidus (1 strain), Ensifer (1 strain) and Gordonia (1 strain). The Bacillus genus was the dominant manganese resistant bacterial genus. The detection of resistant levels of these bacteria to Mn2+(MICs, Minimal Inhibitory Concentrations) showed that they all had the highr Mn2+ resistances with MICs ranged from 20 mM to 70 mM. Strains DB5(Pseudomonas sp.), DC7 (Gordonia sp.) and DB1(Bacillus sp.) showed obvious manganese oxidizing than others by preliminary test using LBB (Leukobethelin blue) reagent. Scanning electron microscope showed the cells of strain DB5 had obvious changes after cultivating in K medium with Mn2+. The study of this aspect enriched the resources of manganese-resistant and manganese-oxidizing bacteria and provided a foundation for the future study of manganese bioremediation. |
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