Studies On Microbial Molecular Ecology In Rhizosphere Soil Of Tolerant Plants Under The Stress Of Landfill Gas

Sanitary landfill technique of Municipal Solid Waste (MSW) disposal plays an important role, but faces more and more challenges. Retardation of revegetation, ever escalating water and air pollution caused by landfill and the degradation of land resource and landscape Scenary are becoming the major issues to be addressed in conserving the city ecology. According to the investigations on vegetation of Tianziling landfill, several tolerant plants under the stress of landfill gases were the frist time to be selected by us. Based on these findings, a novel technique for landfill revegetation was designed using municipal sewage sludge to enhance the landfill cover soil. The variation of the composition of microbial community in rhizosphere soil of tolerant plants: Chenopodium album L. (C. album) and Festuca ovina under the stress of landfill gas (LFG) was studied at laboratory level. Microbial molecular ecology of the methanotrophic bacteria community in rhizosphere soil of tolerant plants, enhanced by the addition of sewage sludge and compost to the landfill cover was investigated under the stress of LFG.The survey carried out in and around the Tianziling landfill of Hangzhou city, to understand the composition of vegetation, revealed that there are 86 species of plants belonging to 35 families, inside the landfill. The results show that it needs 7-8 years to cover the whole landfill by natural vegetation, and the dominant plants are herbs mainly composed of wild weeds, while plants belong to arbor and frutex are very rare in the landfill. The evolving trend is fit to the trends of general disused sites. However, the natural wild weeds play a destructive role in landfill revegetation. The effective strategy to enhance landfill revegetation is therefore first, to control the unlimited extension of wild weeds in the landfill and then, to select tolerant plants to the landfill amended with the landfill cover soil with biosolid and other nutrients.Based on the investigation of vegetation and the principles of landscape reclamation in landfill, 20 species of arbor and frutex plants and 9 species of herbs were used for the vegetation study in the Tianziling landfill site. It is found that Pinus elliottii, Ligustrum lucidum, Cercis chinensis, Festuca ovina, Eragrostis pilosa and Chenopdium album L (C. album) ect, are more tolerant to the landfill conditions. Celtis julianae,Zelkova schneideriana,Pistacia Chinese,Photinia serrulata,Cercis chinensis and Chenopdium album L. were selected for the first time as the tolerant plants used for landfill revegetation. These tolerant plants were seclected to be planted in the landfill according to the "Arbor + Frutex + Herb" principle, and a successful field demonstration was done to provide the novel landfill revegetation techniques of using municipal sewage sludge to enrich the landfill cover soil. The revegetation time of a landfill is expected to be shortening at least by two years with this technique.The effect of a LFG tolerant plant: C. album on methane oxidation activity (MOA) and bacterial community composition in landfill cover soil was investigated at field level. Soil samples from four simulated lysimeters with and without LFG and plant vegetation were taken at 4 stages during the development cycle of the plant. Results showed that the total number of culturable bacteria in soil could significantly be increased (P < 0.05) by the growth of C. album. The total number of methanotrophs and MOA in soils with LFG was significantly higher (P < 0.05) than in soils without LFG on the sampling days of 90, 150 and 210. Further, the total number of methanotrophs and MOA in lysimeters with LFG, increased in the presence of C. album when the plant entered the seed setting stage. Polymerase chain reaction and denaturing gradient gel electrophoresis (PCR-DGGE) gel patterns of 16S rDNA gene fragment and band sequencing analyses apparently differed among soil bacterial communities in the presence of LFG and plant vegetation. Members of the genus Methylosarcina were found to be the most active and dominant methanotrophs in rhizosphere soil of C. album with LFG, while Methylococcus, Methylocystis, and Methylosinus were the primary methanotroph genera in LFG soil without C. album. Thus, C. album appears to be methanotrophic bacteria specific, in the presence of LFG. Soil MOA and microbial diversity can also be affected significantly by the presence of this plant. The inclusion of plants that are capable of tolerating LFG and specific on active methanotrophs may be critical in controling CH₄ emissions from landfills.Traditional cultivation and the rolling tube method combined with modern molecular microbiological techniques including the Terminal Fragment Length Polymorphism (T-RFLP) Analysis, PCR-DGGE and Clone techques were used to detect the Festuca ovina rhizosphere soil bacterial community compostion, the soil culturable methanotrphic bacteria and the soil methanotrphic bacteria community compostion under the landfill cover soil amended with sewage suldge and sewage sludge compost. The results showed that the biomass, chlorophyll content and photo-physiological characteristics of Festuca ovina were significantly increased by the LFG under the landfill cover soil amended with sewage suldge and sewage sludge compost(P<0.05). The T-RFLP anaysis showed that, the addition of sewage sludge or sewage sludge compost to the landfill cover plays an important role in improving the soil bacterial community compositon. The total number of methanotrophs and MOA in soils stressed by LFG could significantly be increased (P < 0.05) by using sewage sludge and sewage sludge compost. The results of PCR-DGGE combined 16S rDNA phylogenetic analysis showed that the methanotrophic bacterial community and composition of rhizosphere are obviously influenced by the stress of LFG and the addition of sewage sludge or sewage sludge compost to landfill cover. Type II methanotrophic bacteria also became one of the dominant microbes. The dominant methanotrophic bacteria in landfill cover soil, before the stress of LFG and with out the plantation of Festuca ovina were Methylococcus and Methylobactor which are type I methanotrophic bacteria. However, the dominant methanotrophic bacteria in rhizosphere soil of Festuca ovina amended with sewage sludge compost changed to Methylococcus, Methylocaldum and Methylocystis under the stress of LFG. Type I methanotrophic bacteria were more dominant than Type II methanotrophic bacteria under this situation. In contrast, when sewage sludge was used to amend the landfill cover soil, the dominant methanotrophic bacteria in rhizosphere soil of Festuca ovina turned to Methylocaldum, Methylocystis and Methylosinus under the stress of LFG, and Type II methanotrophic bacteria were more dominant under this situation. Consequently, the type of landfill cover has a significant influence on shaping the methanotrophic bacteria community composition and MOA in the rhizosphere soil of the tolerant plants, and the mechanisms may probably be different. The mechanisms attributable to this finding are: the improved availability of nutrients to the tolerant plants and the increased soil porosity, assuring increased supply of oxygen to the plant roots which can create niches for the colonization of methanotrophic bacteria. The variation of methanotrophic bacteria community composition in rhizosphere soil of tolerant plants among the different types of landfill cover is one of the key factors bringing about the significant differences of MOA in different kinds of landfill covers.