Integrative analysis reveals casing layer dynamics during Agaricus bisporus cultivation and the growth promoting effect of Pseudomonas putida AT130

Abstract

Agaricus bisporus is the most widely consumed edible mushroom and an important source of dietary protein and bioactive compounds. The casing layer plays a critical role in its cultivation, where microbial communities and metabolic activities strongly affect yield and quality, however, the stage-resolved research gap remains that it is still unclear which microbial and metabolic shifts in the casing layer are associated with primordia initiation and subsequent quality formation. In this study, the ecological and metabolic basis of mushroom quality formation was investigated by the combination of microbiome sequencing, non-targeted metabolomics, and functional genomics. Microbial communities exhibited stage-specific dynamics, with significant restructuring during primordia formation, when community cohesion and niche breadth reached their highest levels (0.68 ± 0.06 and 4.48 ± 1.05, respectively). A total of 1108 non-volatile metabolites were identified from metabolomic profiling. The adenosine and tryptophan exhibited significant changes and were enriched in energy and amino acid metabolism pathways. A representative strain, Pseudomonas putida AT130, was isolated from the genus Pseudomonas. The gene clusters related to phosphate solubilization, potassium mobilization, lignin degradation, and indole-3-acetic acid biosynthesis were revealed by genome analysis with the multifunctional activities being confirmed through in vitro assays. Pot experiments further showed that AT130 inoculation improved mushroom performance, increasing fruiting body yield by 123.55% and enhancing nutritional traits (protein and soluble sugars increased, whereas ash decreased) relative to the control. These findings linked casing-layer microbiota with mushroom quality and identified AT130 as a promising food-grade bioinoculant to enhance A. bisporus nutritional value and productivity.