Diversity Distribution Patterns And Mechanisms Of Phyllosphere Microbial Community Of Rubber Tree

The phyllosphere is the largest microbial habitat on Earth,and phyllosphere microbiota is critical to their host health,growth,reproduction,stress tolerance,etc.As one of the main sources of natural rubber,rubber trees are mainly distributed in Hainan and Yunnan in China,and the rubber forest ecosystem is one of the most widely distributed forest vegetation in tropical areas in China.However,the pattern of microbial diversity in the phyllosphere of rubber trees and the mechanisms of the community assembly remains unclear.In this paper,we investigated the distribution patterns of phyllosphere microbial diversity in rubber trees in two major rubber-growing regions of China:Hainan Island and Xishuangbanna,and explored the community assembly and driving factors of phyllosphere microbial communities;The effects of leaf aging on the composition and diversity of phyllosphere microbial communities were studied;and explored the potential sources of phyllosphere microbes of rubber trees.The results are as follows:1.We examined the phyllosphere bacterial and fungal composition,diversity,and main drivers of these microbes using Illumina sequencing and assembly.We found that the main dominant phyla of bacteria were Proteobacteria,Firmicutes,and Actinomycetes.The main dominant phyla of fungi are Ascomycota and Basidiomycetes.In addition,PCo A and PERMANOVA showed that the seasonal variation of phyllosphere microbial communities was small,and the fungal community was mainly affected by geographical location(R~2=0.17,P=0.001),and showed a significant biogeographic pattern.Among environmental factors,climatic factors(i.e.MAT and MAP)are the main drivers of the fungal community.The bacterial community was mainly affected by the plant compartment(R~2=0.26,P=0.001),and the factors influencing the bacterial community at different compartments were also different,the main factor influencing epiphytic bacteria were external factors(i.e.climatic factors),and the internal factors(i.e.total nitrogen,total phosphorus,and water content)were the main driving force for epiphytic bacteria community assembly.This suggests that phyllosphere bacterial and fungal communities are driven by different factors.2.Through the study of microorganism species coexistence and construction processes in the phyllosphere,we found that there was a host filtering effect from the leaf surface to the leaf interior and that bacteria responded more strongly to the host filtering effect than fungi.Using the neutral model and normalized stochasticity ratio to quantify the relative contribution of deterministic and stochastic processes in the assembly of phyllosphere microbiomes,we found that the host filtering effect account for the enhancement of the deterministic process of community assembly of bacterial(epiphytic:MST=38.7%,endophytic:MST=27.1%)and fungal community(epiphytic:MST=33.6%,epiphytic:MST=28.4%).The host filtering effect decreased the bacterial diversity,abundance,and network complexity,while the fungal diversity was not significant,and the complexity of the network complexity increased.Co-occurrence networks revealed that the fungi,rather than bacteria were often taking a central ecological position in endophytic.3.Through the analysis of the effects of leaf senescence on phyllosphere microbial diversity and community composition,we found that theαdiversity andβdiversity of bacterial communities changed significantly(P<0.05),while fungal changes were not significant,and leaf senescence had a much stronger influence on the bacterial phylum and OTU abundance than fungal communities,indicating that leaf senescence had a greater influence on bacterial communities than fungal.The number of core bacterial and fungal OTUs that were enriched or decreased in the endophytic community during leaf senescence was significantly higher than that on the epiphytic,yellow leaves retained only 7.49%of the bacteria and 4.25%of the fungi found in green leaves in the endophytic samples,while 32.17%of the bacteria and 17.95%of the fungi were retained in the epiphytic samples,indicating that the effects of leaf senescence were greater in the endophytic microbial community than in the epiphytic community.These results show that the endophytic bacterial community dominates the process of the leaf senescence.During the leaf senescence,α-Proteobacteria and Actinobacteria significantly increase in the endophytic samples,enhancing chemoheterotrophic functions.The drastic changes in the endophytic environment also leads to the re-establishment of the bacterial community,dominated by stochastic processes.The ratio of endophytic to epiphytic OTUs of the bacterial community in yellow samples is higher than in green samples(yellow:40.88%;green:20.77%),and the selectively filtration effect decreased significantly with leaf senescence(P<0.05),indicating a decrease in host filtering effect during leaf senescence,which increases the diversity and complexity of the community.4.By comparing the microbial communities in different microhabitats of rubber trees,it was found that the diversity and abundance of soil-associated microorganisms(bulk soil,rhizosphere soil,and rhizoplane soil)were higher than those of plant-associated microorganisms(root-endosphere,phyllosphere,leaf-endosphere),and the bacterial diversity in the phyllosphere was much lower than that in the belowground parts.Both bacterial(R~2=0.38,P=0.001)and fungal(R~2=0.17,P=0.001)community structures were clearly separated into aboveground and belowground parts,indicating ecological niche-dominated community assembly.Using the FEAST method to analyze the potential sources of leaf-associated microorganisms,we found that 66.9%and 62.9%of leaf endosphere bacteria and fungi,respectively,originated from the leaf surface.However,only 8.18%and4.38%of the epiphytic and endophytic bacteria originated from the belowground part,and the epiphytic and endophytic fungal community had approximately 0.72%and 0.61%of the community sourced from below-ground.Additionally,only 1.3%and 0.9%of the fungi in the belowground part originate from the aboveground part.It was also found that soil is a major potential source of root microbiota.In summary,this study reveals the patterns and drivers of phyllosphere microbial diversity in rubber trees,with bacterial and fungal communities driven by geographic location and ecological niche,respectively;The significant effect of host selection effect on the assembly of phyllosphere microbial community of rubber trees was confirmed,but bacteria and fungi responded differently to host filtration effect;The study also revealed that the response of leaf-associated microbial communities to leaf senescence varied depending on microbial groups and ecological niches,with the bacterial community in the leaf interior dominating the process of leaf senescence.The study provided a preliminary understanding of the main sources of leaf-associated microbes,with limited microbial exchange occurring between the aboveground and underground parts.Our study provides new insights into the mechanisms of phyllosphere microbial diversity and assembly in rubber trees,extending our understanding of microbial community dynamics and the stages of leaf development in tropical terrestrial ecosystems.