DISCUSSION
4.1 | The relative abundance of rhizosphere soil microbial community changed
At the phylum level, the difference in the carbon sequestration level of Masson pine families did not change the dominant phylum of soil bacteria, but the relative abundance of the bacterial phyla changed significantly. The dominant phyla mainly included Acidobacteria, Proteobacteria and Chloroflexi (Figure 2); this result is consistent with the results of previous studies (Bei, Moser, Wu, Müller, & Liesack, 2019). The relative abundance of Acidobacteria increased with increasing carbon sequestration in the different families, and the differences among families reached significant and extremely significant levels (Figure 2). Acidobacteria are important soil microorganisms that play a very important role in the soil carbon cycle and in structuring ecosystems (Wang, et al., 2016). In this experiment, the total organic carbon content in the samples from the three families gradually increased with increasing carbon sequestration (Table 2), showing the same change trend as that of Acidobacteria. This result suggests that some members of Acidobacteria use soil carbon as the substrate for their metabolic activities and that the carbon content in the soil promotes this process. Previous research has also confirmed this view (Pankratov, et al., 2008). On the other hand, in our study, the relative abundance of Proteobacteria decreased with increasing carbon sequestration in the three families, although previous studies have shown that the relative abundance of Proteobacteria was associated with a higher carbon utilization rate (Fierer, Bradford, & Jackson, 2007). Zheng (2014) explored the impact of ecological restoration on the structure of the soil bacterial community of mangroves (Rhizophora apiculata ) and found that the relative abundance of Proteobacteria was negatively correlated with the content of soil organic carbon. The phenomenon observed in our study may be due to the decisive influence of carbon sequestration in the Masson pine on Proteobacteria; there is evidence that the abovementioned vegetation had a greater impact on the bacterial taxa involved in soil carbon sequestration than other factors, such as soil environmental factors (Zhao, 2019). However, the specific mechanism of this influence and its process of action are still unclear and need to be further explored. Notably, Actinobacteria have been demonstrated to use mycelia to break down recalcitrant organic carbon (Dang, Yu, Le, Liu, & Zhao, 2017), and Chloroflexi showed strong carbon sequestration (Li, 2019). However, the results of this study showed that the relative abundance of these two phyla did not change significantly with the increase in carbon sequestration in the different families (Figure 2 A). We speculated that the cause of this phenomenon might be the low proportion of the two bacteria in the total rhizosphere community.
In terms of fungi, the dominant phyla mainly included Basidiomycota, Ascomycota, Mucoromycota and an unclassified phylum (Unclassified_k_functional). The relative abundance of Basidiomycota increased with the increase in the carbon sequestration level of the Masson pine families. The relative abundance of Ascomycota gradually decreased with increasing carbon sequestration, and the dominant rhizosphere fungi changed from Ascomycota to Basidiomycota (Figure 2, Figure S1). Wang et al. (2019) investigated the influence of grassland restoration in a Chinese pine (Pinus tabuliformis ) forest on fungi and found that when the carbon content in the soil decreased, the relative abundance of Ascomycota decreased significantly, and Basidiomycota replaced Ascomycota as the dominant fungal phylum; these results are consistent with the results of this experiment. Although the corresponding molecular mechanism is not fully understood, there is evidence that up to 30% of plant photosynthetic products can be transferred from plant roots to fungi (Gonzalez, et al., 2018; Nehls, Grunze, Willmann, Reich, & Kuester, 2007). Moreover, some families of Basidiomycota have been confirmed to have enzymes that degrade complex macromolecular organic matter in the soil (Floudas, et al., 2012). When the soil carbon content increased, the abundance of Agaricomycetes significantly increased (Li, et al., 2020). Therefore, this might be the reason why Basidiomycota became the dominant fungus after the soil carbon content increased.
4.2 | Most members of the core microorganic community are involved in soil carbon metabolism
Among the three Masson pine families with different carbon sequestration abilities, the core microorganisms in the rhizosphere soil were abundant. For bacteria, the core genera mainly includedBryobacter , Acidibacter , Acidothermus andBradyrhizobium (Figure 3). Among them, Acidothermus andBryobacter can directly utilize carbon in soil as a primary energy source and participate in the soil carbon cycle ( Du, et al., 2017; Pankratov, et al., 2008; Kalyuzhnaya, Lidstrom, & Chistoserdova, 2008). Therefore, with the increase in carbon sequestration in the Masson pine families, these genera both became core rhizosphere bacteria. On the other hand, both Acidibacter andBradyrhizobium are affiliated with Proteobacteria; their relative abundance declined with increasing carbon sequestration in the Masson pine families because these two genera are mainly involved in the nitrogen rather than the carbon metabolism process in soil ( Wang, Lin, Huang, Yang, & Qu, 2019; Zhong, 2019; Liu, 2019). However, the carbon sequestration process in Masson pine had no significant impact on the nitrogen content of rhizosphere soil (Table 2). Therefore, both of these genera may have become core microbial genera simply because of their high overall abundance in soil, since Proteobacteria are common in almost all soil types (Zhang, & Xu, 2008).
In terms of fungi, the core genera of the microorganism community were fewer, only Russula , Geminibasidium , Saitozyma andPenicillium (Figure S2). Because the structure of the fungal community was less stable than that of the bacterial community, the soil environment had a more obvious influence on the fungal community (Chen, et al., 2018; Xiao, Zhao, Yan, & Guan, 2018). In this experiment, the core fungal genera were related to the carbon sequestration process in the Masson pine families to a certain extent. Studies have shown that carbon sequestration by plants stimulates the growth of Russulaand Geminibasidium and that these fungi regulate the uptake and accumulation of carbon by plants in a feedback system (Zhou, et al., 2011; Qiao, Zhou, Chai, Jia, & Li, 2017). The relative abundance ofPenicillium is mainly influenced by the soil carbon content (Hu, 2015; He, 2016). The carbon content of the rhizosphere soil increased due to carbon sequestration by Masson pine (Table 2), which led toPenicillium becoming one of the core fungi in the microbial community.
4.3 | The soil properties affect the microorganic community
Previous studies have shown that the composition of the soil microbial community is significantly correlated with soil nutrient levels (Tian, et al., 2017). Nitrogen limitation is common in most terrestrial ecosystems, and it often leads to intense competition between microorganisms and plants (Liu et al., 2016) as well as to changes in the classification and functional characteristics of the microbial community (Leff et al., 2015). The most typical changes observed in this study were decreases in the relative abundance of some rhizosphere fungal communities, such as Acidibacter and Bradyrhizobium(Figure S2). On the other hand, soil moisture is another important limiting factor in forests and has a strong influence on the soil bacterial community (Zhang, Liu, Xue, & Wang, 2016), especially Proteobacteria (Figure 2, Table 2). Liu et al. (2020) found that the relative abundance of Proteobacteria in rhizosphere and nonrhizosphere soil under safflower (Carthamus tinctorius ) was significantly negatively correlated with soil moisture, which was similar to the results of this study. However, other studies have noted that the relative abundance of Proteobacteria in the nonrhizosphere soil of grassland vegetation (including Stipa bungeana and Artemisia vestita ) was positively correlated with soil moisture (Liu, Huang, & Zeng, 2016). This suggests that the effects of soil moisture on Proteobacteria might also be influenced by vegetation types and rhizosphere effects. However, the experimental results showed that soil moisture had little influence on fungi (Figure 4 B). Zhao et al. (2016) demonstrated that in semiarid grasslands, water restriction reduced the relative abundance of bacteria but did not change that of fungi, which increased the ratio of fungi to bacteria. Notably, pine forest soil has a higher carbon content than soils of other forest types, making it possible to identify fungal taxa that degrade low-quality substrates (Macdonald, et al., 2009). For instance, due to the lower substrate mass in coniferous forests, the diversity of Basidiomycetes and other lignin-degrading fungi was higher than that in prairie soils with lower carbon content (Allison, Hanson, & Treseder, 2007). In this study, with the increase in the carbon content in soil, the relative abundance of Basidiomycetes also increased (Figure S1 B). Soil pH has been reported to play a key role in controlling microbial community composition (Cao et al., 2017). However, in this study, the effects of differences in soil pH among the different families were not significant; it is speculated that pH has little influence on the bacterial and fungal communities in Masson pine rhizosphere soil, except on genera that are relatively sensitive to pH, such as Saitozyma (Figure 4 B, D).
Notably, previous studies have shown that there is a strong relationship between rhizosphere microbes and plant root exudates (He, 2017). The changes in rhizosphere microbial communities among Masson pine families with different carbon sequestration capacities were discussed in this study, but the specific effects of rhizosphere exudates on soil microorganisms under these conditions were not analyzed. With the increase in carbon sequestration, was there a dominant substance that influenced the relative abundance of different rhizosphere bacteria or fungi? What were the patterns and mechanisms of the influence of these exudates? Were there any interactions between these secretions? These topics should be addressed in further research.