Trade-Offs in Growth Performance, Serum Biochemistry, and Fillet Amino Acid Profiles of Red Tilapia Fed Graded Levels of Azolla microphylla Meal in a Fishmeal Replacement Blend, with Exploratory Observations on Intestinal Microbiota

This study evaluated the replacement of commercial fishmeal with a combination of Azolla microphylla meal (AMM) and soybean meal in red tilapia (Oreochromis spp.) diets at 0%, 25%, 50%, and 75% fishmeal replacement over 60 days, followed by a sublethal Aeromonas hydrophila exposure. Soybean meal was increased alongside AMM to maintain isonitrogenous and isoenergetic diets; therefore, the observed responses reflect the combined replacement of fishmeal rather than the effect of AMM alone. Growth performance (WG, ADG, SGR) progressively declined and FCR increased with higher fishmeal replacement levels (p < 0.05). Serum aspartate transaminase (AST) rose and alkaline phosphatase (ALP) fell at elevated replacement levels. Fillet total essential amino acids were significantly reduced only at 75% fishmeal replacement, indicating that moderate substitution (up to 50%) preserves nutritional quality. As an exploratory, descriptive endpoint, the intestinal microbiota was profiled using pooled samples (n = 1 composite per treatment per time point), precluding statistical inference. Within the pooled post-challenge profiles, Cetobacterium represented a high relative proportion in T1–T3, whereas the pooled T4 profile showed a more heterogeneous taxonomic composition. Because no independent microbiota replicates were available, these observations are descriptive and hypothesis-generating rather than evidence of dietary or challenge effects. Overall, fishmeal replacement above 25% impaired growth, whereas fillet total essential amino acids were maintained through 50% replacement. These effects should be interpreted as the combined outcome of replacing fishmeal with AMM and soybean meal. The pooled microbiota profiles provide preliminary observations for future replicated studies.