Azadirachtin and Its Nanoformulation Reshape the Maize Phyllosphere Microbiome While Maintaining Overall Microbial Diversity

The phyllosphere microbiome is an important component of plant-associated ecosystems, and its structure is susceptible to biotic stress and agricultural interventions. However, the non-target effects of plant-derived pesticides and their nanoformulations on the phyllosphere microbial community remain unclear. By using 16S rRNA amplicon sequencing, we investigated the non-target effects of azadirachtin (Aza) and its nanoformulation (O-carboxymethyl chitosan-loaded azadirachtin, O-cmc-aza) on the phyllosphere microbial community of maize, including Spodoptera frugiperda herbivory stress (Attack) as an additional treatment. The results showed that all three treatments significantly altered the phyllosphere microbial community structure, while the overall microbial diversity indices remained stable. Specifically, the Attack treatment significantly enriched bacterial genera such as Akkermansia and Burkholderia-Caballeronia-Paraburkholderia; the Aza treatment mainly increased the abundance of taxa such as Stenotrophomonas and Herbaspirillum, which have been associated in the literature with plant growth promotion; and the O-cmc-aza treatment specifically enriched microbial groups such as Ralstonia and Sphingomonas, which have been reported to include strains involved in pollutant degradation and nitrogen cycling, while reducing the ACE index but maintaining high community evenness. Our results indicated that azadirachtin and its nanoformulations induced compositional changes in the phyllosphere microbiome, without causing marked decline in microbial diversity. This study provides data support for evaluating plant-derived pesticides and nanoformulations with respect to their non-target effect on phyllosphere microbial communities in green agricultural systems.