Humalite restructures microbial community composition, functions, and interaction networks in the canola rhizosphere

Humic substances (HSs) are increasingly recognized for their role in enhancing soil biological functions and promoting sustainable crop production. Humalite, an organic soil amendment derived from deposits in Southern Alberta, is a valuable source of HSs due to its high humic content and low heavy metal contaminations. While Humalite has demonstrated agronomic potential, its effects on crop-associated microbiomes remain understudied. This study investigated the effects of varying Humalite application rates, under recommended NPK fertilization, on bacterial and fungal communities in the canola ( Brassica napus ) rhizosphere. Rhizosphere soils collected at the flowering stage were analyzed using high-throughput sequencing of the 16S rRNA gene and the ITS2 region. Results indicated that, Humalite reduced bacterial and fungal alpha diversity, especially at 800 and 1600 kg ha −1 . Intra and inter-kingdom networks analyses demonstrated increased bacterial connectivity and stronger inter-kingdom interactions at 400, 800 and 1600 kg ha −1 Humalite, whereas fungal networks became more modular and less connected. Functional prediction analysis suggested enrichment of microbial groups potentially associated with aerobic chemoheterotrophy and organic matter decomposition. Humalite also increased the abundance of several bacterial genera associated with plant growth promotion and nutrient cycling, including Pseudomonas , Flavobacterium , Agrobacterium , Chryseobacterium , and Massilia . Fungal responses were more variable, with enrichment of taxa such as Hyphopichia occurring alongside declines in diversity, network connectivity and the abundance of genera associated with nutrient cycling and plant growth. Overall, this research demonstrates that Humalite alters the structure, functional potential, and interaction networks of canola rhizosphere microbial communities.