Examining gene transcription alternative splicing can distinguish gene-splicing events associated with stress and identify biomarkers that can be used to monitor stress conditions and decrease sampling invasiveness. Previously, we in silico predicted that a gene called prolyl 4-hydroxylase subunit alpha-2 (P4HA2) with retained intron is expressed under stress conditions. In this study, we aimed to validate the intron's retention and hypothesized that it would continue to show differential expression in fecal and mucus samples. The in vitro expression of the intron verifies the in silico data, where the intron of interest was upregulated during stress conditions. Separately, a microbiome study using the same samples identified bacterial taxa predictive of stress. Higher levels of 3 bacterial biomarker taxa species belonging to the genera Romboutsia and Cetobacterium were more abundant in the stressed fish. However, a pathogenic class of bacteria (Gammaproteobacteria) was significantly higher in the control group.