4 DISCUSSION
Recent studies indicated that changes of soil microbial communities as a result of various agricultural management practices contribute to plant performance (Zhou et al., 2014; Gao et al., 2019). Organic farming (Reganold & Wachter, 2016), no-tillage (Ashworth et al., 2017), and rotation (Liu et al., 2019) have attracted the most scientific attention in the field of microbial research, while natural fallow practice has not been investigated in detail to this regard. Importantly, natural fallow practice is widely used worldwide because of labor shortage and areas unsuitable for mechanic agriculture. Therefore, the present study investigated the microbial mechanisms underlying the different response of N. tabacum plants to changes of soil microbiomes induced by natural fallow and continuous N. tabacum cropping practices. Clearly, different environmental conditions impose different selection forces on plants (Anderson, 2016; Jansson & Hofmockel, 2020), while biomass accumulation and allocation strategies may vary over time and across environments (Poorter et al., 2012). First, the effects of microorganisms on N. tabacum fitness were investigated by reassembling soil microbial communities. The results indicated that changes of soil microbiomes significantly decreased the total biomass accumulation, changed biomass allocation patterns, and altered the investment strategies in N. tabacum leaves grown in FS. However, no similar changes were observed in plants grown in CCS (Figures 1a and b). This suggests that soil microorganisms influence plant performance related to the abiotic soil context (Lozano et al., 2017; Mapelli et al., 2018). The LMA level is a key trait to assess plant growth and an important indicator for plant strategies (Poorter et al., 2009). In the present study, LMA levels showed a plastic response to environmental differences and plants grown in FS preferred to invest more in leaves and acquired a high leaf density during the process of microbial community reassembly (Figure 1c). In general, herbaceous or woody plants with a slower relative growth ratio (RGR) have a higher LMA level (Poorter et al., 2009). Thus, this implies that N. tabacum growth may suffer more from negative feedback of the reassembling soil microbial communities in response to natural fallow practice than in response to continuous cropping practice. In summary, these results suggested that soil microorganisms of FS play a more important role in determining N. tabacumfitness than those of CCS. Considering that, in practice, continuousN. tabacum cropping does not exceed three years, the damage from the changes of soil microbiomes to N. tabacum growth may be exaggerated at least within the short length of continuous cropping.
Soil microorganisms regulate many ecosystem processes and play key roles in nutrient cycling (Bender et al., 2016). However, the community structure and function also appear to be readily influenced by agricultural management practices. For example, bacterial network properties under continuous N. tabacum cropping are more sensitive to soil variables (Chen et al., 2018), while the abundances of several beneficial fungal species increased in a continuous soybean cropping practice (Liu et al., 2019). The present study identified no significant difference of microbial community richness under both management practices. However, bacterial and fungal community richness was higher for bulk soil in F_NS than in F_S, while no significant differences were found between C_NS and C_S. This suggests that the abiotic context of FS exerts stronger effects on the reassembly of soil microbiomes (de Vries et al., 2012; Erlandson et al., 2018). Furthermore, the taxonomy of microbial diversity indicated differences related to the management practice (Figures 2 and S3). For example, although Proteobacteria, Actinobacteria, and Bacteroidetes dominated both sample types, Firmicutes showed a relatively higher abundance in CCS than in FS. Previous studies indicated that Firmicutes was closely related to organic management (Hartman et al., 2018) and functioned in response to receiving manure fertilizer (Hartmann et al., 2014). Hence, this implies that relatively lower abundance of Firmicutes OTUs in FS may result in a low nutrient utilization efficiency on the bacterial level. This indirectly influenced N. tabacum growth, and finally decreased biomass accumulation. Moreover, PCoA showed marked differences among fungal communities of bulk soil; however, there was no significant difference in the rhizosphere of fungal communities (Figure 3). This indicates that the rhizosphere serves as a carbon-rich niche for the establishment of microbial communities while bulk soil is rapidly depleted of carbon and other nutrients by heterotrophic microbes (Sasse et al., 2018). Thus, this further suggests that similar colonization and coexistence conditions for fungi may exist in rhizosphere of N. tabacum plants both under continuous cropping and natural fallow practice. The results of the present study are consistent with a previous study, showing that species richness was less variable in the responses to different cropping systems than species composition (Hartman et al., 2018).
In addition, this study highlighted the differences of context-specific microbial clades between CCS and FS (Figures 4a and b; Tables 1 and 2). Several bacterial groups, such as Firmicutes and Actinobacteria, only showed significant enrichment in the bulk soil and rhizosphere of C_NS. This suggested that these bacterial groups were associated with receiving both manure fertilizer (Hartmann et al., 2014 and 2018) and soil fertility (Mapelli et al., 2018). In parallel, several bacterial groups were also found to be exclusively enriched in F_NS. For example, previous studies implied that Sphingomonadales (including family Sphingomonadaceae) has the ability to degrade a wide range of aromatic compounds (Balkwill et al., 2006), cope with stress conditions, and adapt to new habitats (Vaz-Moreira et al., 2011). Therefore, it has been suggested that the relatively high abundance of Sphingomonadales found in FS may be useful to improve N. tabacum growth environments. Several key microbes involved in Xanthobacteraceae contribute to nitrogen cycling (Zhu et al., 2018). The present study showed that Xanthobacteraceae were significantly enriched in rhizosphere in F_NS, suggesting that these microorganisms in the rhizosphere may play a key role in helping N. tabacum plants to acquire nitrogen nutrition when they grow in FS. In addition to bacterial clades, several fungal clades, such as Rozellomycota, Eurotiomycetes, and Sordariomycetes, were exclusively detected in the bulk soil in F_NS. This suggests that these play distinct roles in the plant-soil feedback when N. tabacumplants are grown in FS. The fungal class Leotiomycetes (family Myxotrichaceae and genus Oidiodendron ) was significantly enriched in rhizosphere in C_NS. Myxotrichaceae is cellulolytic, and was implied to play a significant role in the decomposition dynamics of litter (Rice et al., 2006). Tremellomycetes and Dothideomycetes dominated the rhizosphere in F_NS and may result in the difference between the structural and functional differences from C_NS. Moreover, this study also documented specific microbial clades in the reassembling soil microbial communities (Figure 4c, Table S3). For example, the bacterial family Rhodanobacteraceae, which belongs to the class of Gammaproteobacteria, dominated the rhizosphere in F_S, and was implied to have the capacity to degrade polycyclic aromatic hydrocarbons (Cazals et al., 2019). Importantly, no fungal clade was significantly enriched among the reassembled fungal communities. This again suggests that similar colonization and coexistence conditions may exist for fungi in the rhizosphere of N. tabacum . In conclusion, these differences in composition suggest that not only plant performance (Sasse et al., 2018), but also the specific differences in the abiotic context exert distinct effects on the reassembly of the soil microbiome.
As far as we know, several common factors, including abiotic and biotic factors, likely lead to the assembly of a core microbiome (Naylor et al., 2017; Erlandson et al., 2018; Lundberg & Teixeira, 2018; Perez-Jaramillo et al., 2019). In the present study, only 24.1% bacterial OTUs and 13.2% fungal OTUs were identified as the core microbiomes (Table S4), suggesting that most OTUs were specific and thatN. tabacum plants may have a strong strength of rhizosphere effect. Previous studies suggested that the strength of the rhizosphere effect is likely associated with the developmental stage of a plant (Chaparro et al., 2013), root exudation (Zhalnina et al., 2018), host genotype (Bulgarelli et al., 2015), and domestication (Edwards et al., 2019). This study further compared the compositional differences between bulk soil and rhizosphere and identified several context-specific microbial species, especially those exclusively enriched in the rhizosphere (Tables S5 and S6). This suggested that those species likely elucidate the differences of rhizosphere bacterial and fungal microbiomes between FS and CCS.
5 CONCLUSIONS
The findings of the present study imply that specific microbial community assembly and especially a number of context-specific microbial clades under different management practices (continuous cropping and natural fallow) may lead to different growth and development inN. tabacum . In addition, a set of positive changes of soil microbiomes induced by the natural fallow practice could not completely enhance N. tabacum fitness, while additional agricultural management practices such as targeted microbial fertilizer application or rotation may compensate for the shortcomings arising from a single measure.