Humalite shapes the wheat rhizosphere soil microbiome by altering microbial community structure, diversity, and network stability

The application of humic substances to enhance soil health and crop yield has gained considerable interest in recent years, mainly due to their organic origin and capacity to improve the physicochemical and biological properties of the soil. Humalite, a rich source of humic substances found in southern Alberta, Canada, is particularly valuable due to its low ash and heavy metal levels. Despite its agricultural potential, its effects on the soil microbiome have yet to be evaluated. This study utilized 16S rRNA gene and ITS2 region amplicon sequencing to examine bacterial and fungal communities in rhizosphere soil collected from wheat plants cultivated at five Humalite application rates (0, 200, 400, 800, and 1600 kg/ha) in combination with nitrogen, phosphorus, and potassium (NPK) at recommended levels based on soil test under controlled greenhouse conditions. Results indicated that Humalite application influenced microbial community composition by increasing the abundance of beneficial bacterial ( Flavisolibacter , Gaiella , Geomonas and Sphingomonas ) and fungal ( Solicoccozyma , Clonostachys, Trichoderma ) genera while reducing pathogenic and harmful taxa (Bedellovibrionota and Fusarium ). The Humalite application reduced bacterial diversity while increasing fungal diversity specifically at 800 and 1600 kg/ha, and increased the co-occurrence network stability. Notably, the abundance of various taxa involved in reducing N 2 O emissions (Methylomirabilota, Gemmatimonadota, Terrimonas ) was higher in Humalite-treated soils. Overall, Humalite application modulated rhizosphere microbial communities, enhancing beneficial taxa and network connectivity while suppressing pathogenic and harmful taxa. These changes suggest that Humalite creates a more balanced, resilient, and health-promoting soil microbiome. • This study investigated the effects of Humalite on the wheat rhizosphere soil microbiome. • Humalite treatment altered microbial community composition and alpha and beta diversities. • The abundance of beneficial bacterial and fungal taxa increased while harmful taxa decreased following Humalite application. • Humalite application increased the stability of the bacterial and fungal co-occurrence network. • The abundance of various bacterial taxa associated with nitrogen loss pathways was reduced under Humalite treatments.