Supplementary file 1_AMF-mediated rhizospheric interactions by soil microbiota and metabolites in intercropping of tobacco and maize to regulate the soil nutrients.docx

IntroductionColonization of arbuscular mycorrhizal fungi (AMF) can form the root symbiotic network of co-cultured plants roots and hyphae thus promote plant growth. Maize is often intercropped during the harvesting period of tobacco in tobacco-growing areas in China. AMF Colonization has been shown to be an effective approach for regulating the synergistic growth of Nicotiana tabacum and maize.
MethodsIn this study, pot experiments were conducted, and samples were analyzed using 16S rDNA and ITS high-throughput sequencing for bacteria and fungi, respectively,and LC-MS/MS widely targeted metabolomics of soil. Differences in microbiota and metabolites in the rhizosphere soil of tobacco and maize, as well as their correlations with the agrochemical properties of soil at the vegetative stage of crop growth, were investigated under AMF colonization to clarify the feedback regulation of plant growth by AMF inoculation and rhizosphere interactions between Nicotiana tabacum and maize.
ResultsThe results showed that the growth of tobacco and maize inoculated with AMF was better than that of uninoculated plants, and this was related to the enhancement of plant nutrient uptake by AMF and root interactions between the two crops, which resulted in significant increases in the content of alkaline hydrolyzable nitrogen (N), available phosphorus (P), and available potassium (K) in the rhizosphere soil of tobacco. The stem girth of tobacco and the biomass of maize were significantly higher under intercropping than monoculture, as intercropping increased the relative abundances of Penicillium, Trichoderma, Blastomonas, and Sphingomonas in the rhizosphere soil of tobacco and maize; the abundance of Penicillium was higher in rhizosphere soil of AMF inoculated treatments. AMF inoculation and intercropping cultivation respectively led to the down-regulation of differentially expressed metabolites (DEMs) in the rhizosphere soil of tobacco and maize. Additionally, pH and organic matter are key environmental factors influencing soil microbial communities.
DiscussionOverall, intercropping and AMF inoculation mediated rhizospheric interactions by soil microbiota and metabolites in intercropping can regulate plant growth and improving the content of N, P, and K in rhizosphere soil. Our findings provide new insights with implications for AMF application on interactions between the root systems of tobacco with maize or other plants during the tobacco harvesting period.

引言
丛枝菌根真菌（arbuscular mycorrhizal fungi, AMF）定殖可形成共培养植物根系与菌丝的根际共生网络，进而促进植物生长。在中国烟区，烟草收获期常套种玉米。已有研究表明，丛枝菌根真菌定殖是调控烟草（Nicotiana tabacum）与玉米协同生长的有效手段。

方法
本研究开展盆栽试验，分别采用16S rDNA和ITS高通量测序分析土壤细菌与真菌群落，并采用LC-MS/MS广泛靶向代谢组学技术分析土壤样本。本研究探究了AMF定殖条件下，烟草与玉米根际土壤的微生物群与代谢物差异，及其在作物营养生长期与土壤农化性状的相关性，以明确AMF接种对植物生长的反馈调控作用，以及烟草与玉米间的根际互作机制。

结果
研究发现，接种AMF的烟草与玉米生长状况优于未接种对照组，这与AMF增强植物养分吸收以及两作物间的根系互作相关，该效应使得烟草根际土壤的碱解氮（N）、有效磷（P）和速效钾（K）含量显著提升。间作处理下烟草茎粗与玉米生物量均显著高于单作处理，这是由于间作提高了烟草与玉米根际土壤中青霉属（Penicillium）、木霉属（Trichoderma）、芽单胞菌属（Blastomonas）和鞘氨醇单胞菌属（Sphingomonas）的相对丰度；且AMF接种处理的根际土壤中青霉属丰度更高。AMF接种与间作栽培分别导致烟草与玉米根际土壤的差异表达代谢物（differentially expressed metabolites, DEMs）下调表达。此外，pH值与有机质是影响土壤微生物群落的关键环境因子。

讨论
综上，间作与AMF接种通过土壤微生物群与代谢物介导根际互作，可调控植物生长并提升根际土壤的氮、磷、钾含量。本研究结果为烟草收获期AMF在烟草与玉米或其他作物根系互作中的应用提供了新的理论参考。