Effects Of Waterflooding And Organic Manure Addition On Soil Autotrophic Microorganism Communities

As an active group living in pedosphere,autotrophic microbes play important roles in counter-regulate the aromospheric CO2 to soil carbon pool and affected by factors both natural and artificial.The research based on three types of farmland soil(clay,sand and loam)and set different treatment through pigeon droppings addition and waterflooding to simulate different land utilization types and management measures under agroec.ological systems.The incubated soil DNA is extracted and through which,we amplified cbbLR and cbbM gene(large subunit of RubisCO I,II)and then using techniques such as Q-PCR,PCR-RFLP and Illumina MiSeq sequencing to analyze autotrophic bacterial communities composition,diversity and abundance,and further explore the potential carbon fixation bacteria as well as their response to different habitat and soil physicochemical variables.Based on the quantitative fluorescence analysis,we found that the abundance of cbbLR gene distributed from 0.01 to 2.3×107 copy g-1 but cbbM gene copy generally lower than cbbLR gene for 1 to 2 orders of magnitude.Two genes abundance in soil texture follow the same trend:clay ? loam>sandy.While waterflooding and pigeon drooping addition contribute to significant differences in the two genes abundance:for cbbLR gene abundance,pigeon droppings,and waterflooding didn't bring about the significant changes among treatments.For cbbM gene,addition of pigeon droppings significantly reduced the cbbMgene abundance,but waterflooding appared remarkable effect only when pigeon droppings added.In addition,organic matter and total nitrogen,total potassium and available phosphorus were proved to be significantly influenced cbbLR and cbbM genes abundance.PCR-RFLP method was used to construct cbbLR gene libraries.The results showed that the microorganism diversity in clay resemble to those in loam,but are significantly greater than in sandy.Generally speaking,antijamming capability against pigeon droppings and waterflooding are as follows:sandy>loam>clay.Among positive clones sequencing,we found that ?-and ?-proteobacteria accounted for absolute advantage in cbbLR gene libraries,and several genura of Actinobacteria and Verrucomicrobia are also detected.All sequences belonged to 18 genera,apart from 4 dominant genera Rubrivivax,Rhodopseudomonas Rhodobacter and Burkholderia,we found two newly named genera in recent years:Methylacidimicrobium(methane bacteria)and Polymorphum(the newly found genera under Rhodobacteraceae).We use Illumina MiSeq sequencing technology to build cbbM gene libraries.Through multivariate statistical analysis,we found that waterflooding and pigeon dropping addition less affected microbial community in clay and loam,but significantly influenced the diversity of the community in sand:waterflooding only,pigeon dropping addition only,and waterflooding plus pigeon dropping were significantly increased cbbM genetic diversity in the sandy soil.In addition,the main bacterias which participate in CO2 fixation including purple non-sulfur bacteria(PNSB):Rhodopseudomonas,Rhodovulum,Phaeospirillum and Rhodobacter;Iron oxidizing bacteria(FeOB):Sideroxydans,Leptothrix;Sulfur oxidizing bacteria(SOB):Sulfuricella,Acidithiobacillus,Acidihalobacter,Thiomonas,Thiodictyon,Thiobacillus,Thioalkalicoccus,Thiocystis,hydroxide bacteria(HOB anywhere):Cupriavidus;Azotobacter(NFB):Bradyrhizobium;Hasten magnetic bacteria(MTB):Magnetospira,Magnetospirillum,Magnetovibrio and other bacteria.Among them,the diverse sulfur oxidizing bacteria are the major flora which responsible for CO2 fixation.The relative influence of different habitats on the cbbLR and cbbM gene are the same:soil types>pigeon dropping addition>waterflooding.Physicochemical variables such as organic matter,pH and potassium content is the main factor affecting the solid carbon cbbLR flora,and the soil pH,available nutrients such as VK and AN affect cbbM flora significantely.