4.3 Community shift of CO2 and N2 fixation bacteria and driving factors during 20 year of IM
The CO2 and N2 fixation bacterial communities responded differently to IM. The CO2fixation bacterial community changed gradually with IM duration, with the structures associated with CK and IM20 differing the most. However, a relatively similar structure for soil diazotrophs was observed between CK and IM20, indicating that community of N2 -fixing bacteria responded to the disturbance at earlier stage of IM but returned to resemble the original structure at IM20. These results suggest that the community structure of soil diazotrophic bacteria is more resilient than that of CO2 bacteria, despite declines in abundance. Soils from all treatments had six predominant species of CO2 fixation bacteria but these varied in their relative abundances. The relative abundance of the most predominant T-RF 177 bp was lowest (P < 0.05) in CK, indicating IM improved the species represented by T-RF. RDA revealed that numbers of this T-RF in IM topsoil samples were positively related to AP and δ 13C, indicating soil AP might favor the associated species. In addition, soil P content has also been shown to be a major regulator for cbbL -containing algae composition (Yuan et al., 2015). The most dominant group was composed of several groups of α, β ,γ -Proteobacteria − Bradyrhizobium sp. , Rhodospirillum centenum , Thioflavicoccus mobilis ,Stappia meyerae , Mesorhizobium ciceri , and Starkeya novella. In the contrast, the relative abundance of T-RFs 44bp and 360bp in topsoil decreased sharply after 6 or 10 years of IM, suggesting that the associated species may have been inhibited by rich soil nutrition. Proteobacteria are fast-growing copiotrophs that thrive in environments with high carbon availability (Fierer, Bradford, & Jackson, 2007). Soils in this studied contained five dominant types of diazotrophic bacteria, with the highest relative abundance observed (22.1-54.4% for T-RF 180bp) being reached in the middle stages of IM. These higher abundances of T-RFs exhibited an interesting relationship of ‘growth and decline’ between groups of CK and IM20 vs the rest group of IM treatments. This phenomenon suggested that different species of diazotrophic bacteria have unique environmental preferences. It was difficult to recognize taxa were favored under IM because several T-RFs identified to same species. The number of factors driving changes within the diazotropic community greater than for CO2 fixation bacteria and included AK, SOC, C : N and AN content in topsoils, and AP and C:N in the subsoils. Two taxa of diazotrophic bacteria, Rhizobium sp. and Azorhizobium doebereinerae , were the most dominant of those detected in the soils sampled.
4.4 Alteration of diversity of CO2 and N2 fixation bacterialcommunity and driving factors during 20 year of IM
Just as the abundances of CO2- and N2-fixing bacteria responded differently to IM, so too did the corresponding measures of diversity used in this study. A dip in Shannon and Evenness index values detected for CO2-fixing bacteria at IM10 in both topsoils and subsoils was opposed to the opposite trend observed for the Simpson index. Lower values for Shannon and Evenness indices after 10 years of IM may reflect a collective effect of large applications of manure. The Shannon and Evenness indices, which reflect the total numbers and evenness of microbial species, was decreased by manure input, which may have stimulated certain groups of CO2-fixing bacteria bacteria. Then, as the effect of manure declined with further duration of IM, the indices recovered gradually. On the contrary, the Simpson index reflects the primarily the dominant species and so may have increased significantly at IM10 due to the dominance of TRF 177 bp. The recovery of Shannon and Evenness index after 10 years of IM may be attributed to an increase in the number of species, but these were quite different from those in the CK and IM6. Overall, these results indicate that the communities of CO2 fixation bacteria were altered by IM practice. Conversely, both the Shannon and Evenness indices for the diazotrophic communities indicated a return to the original state (CK) suggesting that their communities was not impacted significantly by IM practice. The above results implied that CO2 -fixing bacteria was more sensitive to IM practice with application of MCM than diazotrophic bacteria.