Dietary humic and fulvic acid supplementation improves growth and modulates intestinal morphology and gut microbiome in Rainbow Trout

Abstract Objective Humic substances have been widely used in livestock feeds, but limited research exists on their application in aquafeeds. This study evaluated the effects of dietary humic substances on growth, gut pH, intestinal morphology, and the gut microbiome of juvenile Rainbow Trout Oncorhynchus mykiss. Methods Rainbow Trout averaging 22.9 ± 0.19 g (mean ± SE) in triplicate tanks were fed one of four diets: a control (top-coated with water) or a humic acid–fulvic acid blend (AC Aqua Boost) top-coated at 5, 10, and 15 mL/kg. Fish were fed for 140 d, with biomass recorded biweekly and individual body weight and length measured at trial initiation and completion. Stomach pH and intestinal pH were assessed, villus length and width were measured in proximal and distal intestines, and fecal microbiota were analyzed using 16S ribosomal RNA amplicon sequencing. Results Dietary humic and fulvic acid significantly improved body weight, length, weight gain, thermal growth coefficient, and specific growth rate compared with those in controls. Stomach pH (2.79) differed from proximal and distal intestine pH (7.53), although dietary treatments did not affect gut pH. Villus length in the distal intestine increased in fish that were fed the 5-mL/kg diet. Microbiome analysis showed minor effects at 5 and 10 mL/kg, whereas 15 mL/kg increased Lactobacillus and Clostridium abundance and overall bacterial diversity. Conclusions Overall, the 5-mL/kg treatment improved growth and intestinal morphology, while higher doses modulated the gut microbiome. These findings suggest a threshold concentration influencing trout performance and gut health, supporting the use of supplementation in aquafeeds.