As the main raw material of the tobacco industry, flue-cured tobacco has a long cultivation history all over the world, and its quality is very important in tobacco production (Ren et al., 2015). Continuous cropping refers to the cultivation method of repeatedly planting the same or similar crops in the same field, which is limited by the area of cultivated land and benefits from lower agricultural management costs compared to other cultivation methods, such as rotation, and is widely adopted in many places (Shipton, 1977; Chen et al., 2009). This long-term cropping pattern has inevitably resulted in great impacts on soil quality and field crop production. It usually leads to soil sickness, including deterioration of soil physicochemical properties, increase of pests and diseases, and accumulation of toxic substances. Moreover, long-term cropping can also increase the impact on the soil microbial community. All of these inevitably cause poor growth of crop plants, eventually resulting in yield and quality decline (Garbeva et al., 2004; Tan et al., 2021; Feng et al., 2022; Zhang et al., 2023). A recent study showed that the continuous cropping of sweet potato led to a 20%–30% reduction in yield, as well as a lack of production or even serious plant death (Gao et al., 2019). Continuous potato cropping caused a decline in soil fertility and aggravated the toxic effects of root secretions (Qin et al., 2017). Continuous cropping of chili led to the enrichment of pathogenic microbes (Chen et al., 2021), while continuous cropping of tomatoes significantly reduced soil enzyme activity, microbial biomass, and microbial metabolic activity, leading to changes in microbial community composition and structure, which caused changes in microbial community composition and structure (Fu et al., 2017). To sum up, there are three main factors associated with continuous cropping: imbalance of soil nutrients, shifts in microbial community composition, and autotoxicity of root exudates (Zhu et al., 2018). Tobacco, a type of plant in the Solanaceae family, is particularly intolerant to continuous cropping (Niu et al., 2017). The continuous cropping of Yunnan flue-cured tobacco has led to changes in environmental and biological factors in the soil (Yu et al., 2023). However, due to limited arable land, insufficient human resources, and a lack of scientific and advanced planting concepts, a large portion of tobacco planting still adopts continuous cropping.

The rhizosphere, as the narrow zone of soil that surrounds plant roots, is the habitat of countless microorganisms, and it can interact with plant roots (Philippot et al., 2013). The relationship between plant roots and rhizosphere microorganisms is also mutual: metabolites secreted by plant roots provide a favorable environment for the growth and reproduction of microorganisms (Edwards et al., 2015), and in return, rhizosphere microorganisms have significant effects on root traits and plant nutrition, growth, and development (Li et al., 2017b). As has been well documented, plants, soils, and microorganisms use the rhizosphere as the primary area for signal communication and material and energy exchange (Levy et al., 2017). Therefore, the study of the rhizosphere microbial community will be beneficial in addressing the issue of soil sickness caused by the continuous cropping of tobacco.

Soil microorganisms are critical to many processes including organic matter turnover, mineral nutrition cycling, soil structure maintenance, and toxin degradation (Blagodatskaya and Kuzyakov, 2013). Therefore, the variation of the soil microbial community is essential to the sustainability of soil ecosystem functions. On one hand, in agricultural production, soil microbial structure is determined by various factors, including farmland soil type, cropping pattern, and crop variety (Garbeva et al., 2004). On the other hand, soil productivity and sustainability depend on complex physicochemical and biological characteristics (Avidano et al., 2005). Therefore, the diversity and abundance of soil microbials significantly affect the productivity and sustainability of agricultural soil and thereby affect crop growth and quality (Nayyar et al., 2009). However, the general patterns of classification and functional characteristics of rhizosphere and microbial communities have largely remained unexplored. Understanding and managing the functions of rhizosphere microorganisms is crucial for supporting plant growth in different environmental conditions.

Additionally, because of its rapid response to environmental changes, the characteristics of microbial communities have been proven to be effective bio-indicators of soil conditions and environmental change (Welbaum et al., 2004; Chen et al., 2014). Recent studies have shown that bacterial communities presented huge differences when responding to various treatments, including abrupt flooding (Zhu et al., 2020), varying tillage depth in soil (Anderson et al., 2017), and climate changes (Dacal et al., 2022). The rhizosphere microbial community can also reflect the land use history (Li et al., 2019). Therefore, active management of soil microbial communities to improve soil conditions may be a promising method to improve crop productivity (Glick, 2010). However, specific populations, and the characteristics and relationships between plants and soil microbial communities must be further studied and understood.

In this study, we collected rhizosphere soil samples from seven different tobacco planting sites in vigorous growing stage: a place that had just planted flue-cured tobacco for 1 year (CC1Y), and places that had been continuously cropping flue-cured tobacco for 3 (CC3Y), 5 (CC5Y), 8 (CC8Y), 10 (CC10Y), 15 (CC15Y), and 30 (CC30Y) years, respectively. Soil physicochemical properties were analyzed, and high-throughput 16S rRNA and ITS rRNA gene sequencing was performed. The abundance, diversity, and composition of the microbial communities were analyzed. It was hypothesized that different continuous cropping treatments would lead to differences in soil physicochemical properties and recruit diverse soil microbial communities. There were two main objectives of this study: 1) to explore the effects of continuous tobacco cropping on soil microbial communities and physicochemical properties; and 2) to reveal the correlation between soil properties and soil microbial communities under continuous cropping systems.

In this study, it was found that as the continuous cropping years of tobacco increased, soil physicochemical properties changed, soil quick-available nutrients such as nitrogen, phosphorus, and potassium accumulated, soil organic matter also increased. Soil available nutrients are important indicators for evaluating soil fertility and are the direct source of the three basic elements required by plants, which have a significant impact on crop yield and quality (Oldroyd and Leyser, 2020). In previous studies, the impact of continuous cropping on soil properties of different crops was not the same; it has been shown that long-term continuous cropping of crops such as peanuts and chili leads to an increase in the content of available potassium, available phosphorus, available nitrogen, and soil organic matter (Chen et al., 2021; Li et al., 2022a).

Our sampling method was similar to earlier investigations of continuous tobacco cropping by Yu, involving samples obtained within the same year. Nonetheless, while Yu’s sampling primarily emphasized distinctions between continuous and non-continuous cropping, our sampling method was adopted based on the variations across continuous cropping years (Yu et al., 2023). In Yu’s study, continuous cropping could significantly increase the available nitrogen content and significantly decrease the available potassium of tobacco grown in Yunnan. But in Yan’s study, continuous cropping significantly reduced the soil organic matter, nitrate nitrogen, and available potassium content of Liangshan tobacco, but significantly increased the content of available phosphorus (Yan et al., 2021). The results showed that tobacco cultivation for more than 3 years led to an accumulation of a large amount of available nutrients in the soil, including available potassium and available phosphorus. On one hand, this might be relative to the long-term fertilization during continuous cropping process, and it may also be related to the preferential absorption of plants (Chen et al., 2020). However, in previous studies, soil sickness brought about by continuous cropping could not be prevented by only correcting the imbalanced physicochemical factors in soil (Wilson et al., 2004).

Soil microorganisms have been shown to play important roles, being able to participate in the circulation of elements such as carbon, nitrogen, phosphorus, and sulfur (Yi et al., 2019; Li et al., 2022b). Continuous planting of the same crop is a convenient way to manage farmland, but the cultivation of a single crop affects the soil texture and the structure and function of the root microbial community of the crop (Niu et al., 2016; Wei and Yu, 2018; Li et al., 2020a; Dang et al., 2020). Our study indicated that long-term continuous cropping significantly reduced the diversity and stability of the soil bacterial community, and this is consistent with previous studies (Tan et al., 2021). However, continuous cropping did not show a significant impact on the diversity and stability of fungal community, in agreement with previous studies (Yu et al., 2023). The healthy growth of plants and the formation and maintenance of disease-resistant soils benefit from high diversity in the soil microbiome (Yuan et al., 2014). Therefore, the loss of rhizosphere bacterial richness and diversity may be one of the factors leading to poor growth of tobacco. In this study, the abundance of Chloroflexi and Patescibacteria in soil decreased along with continuous planting years. As reported, the most abundant bacteria belonging to Chloroflexi were involved in various organic transformations and sulfate metabolism, and they could reduce the potential for membrane contamination (Miura et al., 2007; Hug et al., 2013; Narsing Rao et al., 2022). The decrease in Chloroflexi abundance may be one of the reasons for the accumulation of soil organic matter. After years of continuous cropping, the abundance of the Patescibacteria phylum declined. Patescibacteria has been identified as potentially the large autotrophic taxa involved in the nitrogen, sulfur, and iron cycles (Herrmann et al., 2019). A decrease in the abundance of the Patescibacteria phylum indicated the decrease of some part of the microbial communities involved in the circulation of elements. These reductions in the abundance of beneficial microorganisms leading to a huge impact on the rhizosphere environment may be one of the reasons for nutrient imbalances. Ascomycota, Basidiomycota, and Mortierellomycota were dominant fungal phyla, consistent with previous studies (Wang et al., 2020). Ascomycota is a saprophytic soil fungus that plays a major ecological role as a decomposer in nature and is influenced by plant species and planting methods. In this study, there was a significant correlation between Ascomycota and AN, AP, and SOM, and the decrease of Ascomycota abundance may be one of the reasons for the enrichment of various available nutrients. As reported, Mortierella could dissolve mineral phosphorus in soil by synthesizing and secreting oxalic acid (Spaepen et al., 2007), and it can also grow rapidly in iron-rich soil and use its mycelium to facilitate the growth of beneficial bacteria such as Bacillus (Jeewani et al., 2021). Olpidiomycota has also been reported to be significantly enriched in non-continuous cropping sugar beets (Cui et al., 2022). However, an opposite result was shown, where the abundance of Chytridiomycota increased significantly in the continuous cropping of sweet potato (Gao et al., 2019). Together, these comparisons indicated that the microbial community were affected by various factors, among which crop variety played a key role. Additionally, a significant positive correlation between Basidiomycota and continuous cropping year indicated that continuous cropping significantly enriches Basidiomycota. This is in contrast to the results of many previous studies (Beule et al., 2019; Beule et al., 2021; Pang et al., 2021; Xu et al., 2022) but similar in a study of continuous cropping in grape (Liu et al., 2021). Basidiomycota has a wide variety of fungal species, making it one of the most advanced groups of fungi, with extremely wide distribution and complex functions (He et al., 2022). The impact of changes in Basidiomycota abundance requires more detailed research. Above all, the balance of rhizosphere microecology is an important measure for maintaining the balance of soil and plant microbiota, as a good rhizosphere microecological environment can not only promote the absorption and utilization of nutrients by plants but also help plants resist the invasion of diseases (Gonzalez et al., 2018; Zhou et al., 2020; Ling et al., 2022). The disruption of the rhizosphere microecological balance could lead to a decrease in sustainable soil productivity (Berendsen et al., 2012).

Recent studies have suggested that land use history affects crop rhizosphere microbiomes and plant physiology (Li et al., 2019). Soils with specific microbial community structure characteristics can also reflect soil changes and health levels (Shao et al., 2019). In the correlation analysis with soil properties in bacterial samples, Streptomyces, Sphingomonas, Microvirga, and Bryobacter showed a significant positive correlation with continuous cropping years. Streptomyces is one of the most important bacterial communities in soil, with great potential for the bioremediation of toxic compounds (Álvarez et al., 2012), and plays an important role in the decomposition of organic matter that is beneficial for crop production (Qin et al., 2011). The results of this study are consistent with a study of the continuous cropping of sugarcane (Pang et al., 2021) but in contrast to a study of continuous cropping in soybean (Xu et al., 2022), implying the diversity between different crops. Sphingomonas has the function of promoting plant growth and decomposing aromatic compounds (Khan et al., 2014; Zhou et al., 2016). A previous study used it as an inoculum for soil herbicide removal experiments (Li et al., 2017a). Microvirga has the ability to decompose the pyridinecarboxamide compound, and it is used as a bioremediation agent (Zhao et al., 2021). In fungal samples, it was found that Tausonia was the first dominant genus in Basidiomycota taxa, and its abundance increased with continuous cropping years, with significant differences; this partially explained the significant increase in Basidiomycota. Environmental factor correlation analysis at the genus level showed that Tausonia and Solicocozyma under the Basidiomycota phylum were significantly positively correlated with continuous cropping years, consistent with previous studies on continuous cropping of sugar beet (Cui et al., 2022). Previous studies have shown that under the reduced nitrogen fertilizer environment of tobacco, the abundance of Tausonia significantly increased (Shen et al., 2022). Tausonia may participate in regulating tobacco sugar content and may also play a role in plant tolerance to low temperature and fermentation processes (Kim et al., 2020; Wang et al., 2022b). Tausonia pullulans has been reported to be able to delay the mycorrhizal colonization of beneficial fungi in tomato roots (Mestre et al., 2022). As a Solanaceae crop, the tobacco plant may also affect the colonization of beneficial fungi due to the enrichment of the Tausonia genus. Similarly, the Fusarium and Pseudomycohila genera under the Ascomycota phylum also showed a significant positive correlation with the duration of continuous cropping. Fusarium genera have various pathogenic fungi, among which Fusarium oxysporum can directly cause root rot. Therefore, the increase in Fusarium abundance might greatly increase the possibility of soil disease. The changes in these specific genera with different functions during continuous cropping could provide a reference for the exploration of microbial agent products, which could be beneficial in solving soil sickness caused by continuous cropping.

The cycling of nitrogen in soil is influenced by both plant and microbial functional genes (Morse et al., 2018). Previous studies have shown that an increase in nitrogen inhibits microbial activity, such as microbial respiration, and a decrease in microbial biomass carbon (Liu and Greaver, 2010; Hu et al., 2021). In this study, there was a significant decrease in nitrogen metabolism functions and a significant increase in respiratory function in bacterial communities, with significant differences in continuous cropping. This is in contrast to previous studies, indicating that differential microorganisms may not fully explain functional changes. A previous study showed that differential microorganisms can serve as important indicators of environmental change (Berard et al., 2004), but the function of soil microbial communities is not entirely determined by bacteria, and the function of fungi is equally important. In Yu’s research, the correlation between allelochemicals, microbial communities, and tobacco continuous cropping is evident, with allelochemicals leading to a more significant influence than microbial communities. In Yu’s findings, however, soil physicochemical properties did not show a direct correlation with the continuous cropping of tobacco (Yu et al., 2023). Therefore, the exploration of microbial communities should extend beyond surface-level assessments of species richness and diversity. A deeper and more comprehensive examination into their functional roles and interactions should be performed. It should be noted that the bacterial community function in this study was predicted using only the FAPROTAX database, rather than being validated through experiments as other studies have done, which could have led to differences from the actual situation (Tao et al., 2020; Jin et al., 2023; Zhou et al., 2023). Therefore, further experimental verification is needed to assess the impact of microbial community changes on microbial community function. It should also be noted that the sampling method adopted in this study, which involves selecting different plots in the same year, present some limitations to a certain extent. A more accurate method for continuous cropping study would be to sample the same plot for multiple consecutive years. Consecutive sampling would play a crucial role in decreasing the impact of environmental changes on the dynamic shifts within planting soil and microbial communities. It facilitates a clearer comprehension of the change trends in soil and microbial communities. Furthermore, consecutive sampling contributes to evaluating the relative stability of microbial communities, enhancing the reliability of study data. Nevertheless, the consecutive sampling method poses specific challenges for practical implementation.

Of course, considering the complexity of plant microbial and microbial interactions, optimizing soil microbial partnerships to improve soil quality is a challenging task (Brodeur, 2012; Toju et al., 2018; Sessitsch et al., 2019). Sometimes the change of a single microorganism could lead to great changes in the entire community (Tao et al., 2020), indicating the regulatory complexity of microbial community structure. However, the soil shape obtained by changing the rhizosphere microbial community is more durable than changing the fertilization method (Zhou et al., 2023). Therefore, for soil sickness caused by continuous cropping in farmland production, we recommend reducing the application of fertilizers, and instead balancing the microbial community. This is similar to previous research that confirmed that reducing nitrogen inputs could benefit tobacco (Shen et al., 2021). Changing the application of fertilizers to other new types of fertilizers, such as chitin fertilizers (Fan et al., 2022) and organic fertilizers (Chen et al., 2018), is also an effective strategy. Furthermore, changing farming methods would also be an effective way (Liu et al., 2020). These proposed strategies could help to restore the balance of the rhizosphere microecological environment.

This study analyzed the changes in the microbial community structure of tobacco rhizosphere under different continuous cropping years. The relationship between soil microbial community and soil physicochemical properties was revealed. Research has demonstrated that continuous cropping significantly impacted soil physicochemical properties during the long-term continuous cropping process, leading to the enrichment of some soil nutrients. The bacterial and fungal communities in the rhizosphere soil of continuous cropping tobacco underwent significant compositional changes, with some genera significantly associated with continuous cropping. Together, these results could be of benefit for providing a theoretical reference for the proper management and scientific fertilization of tobacco planting soil under continuous cropping conditions.