A microRNA Catalogue for Red Abalone Larvae Reveals Developmental and Acidification-Associated Abundance Patterns

Larval development in marine mollusks requires coordinated gene regulation and is particularly sensitive to ocean acidification, yet post-transcriptional regulation remains poorly characterized in non-model gastropods. We characterized the microRNA (miRNA) repertoire and respiratory response of red abalone (Haliotis rufescens) larvae reared at pH 8.0 and 7.6. Small-RNA sequencing was performed at the trochophore (24 h post-fertilization, hpf) and competent veliger (110 hpf) stages, with three biological replicates per stage and pH treatment, while oxygen consumption was measured at 24, 48, 60, and 110 hpf. Genome-anchored annotation identified 147 precursor loci encoding 117 distinct mature miRNA sequences, including 70 matching reference databases and 47 candidate novel miRNAs. The developmental stage was the predominant source of variation in miRNA abundance, with the first principal component explaining 87.3% of the variation, whereas acidification affected a comparatively small subset of miRNAs. Cumulative oxygen consumption from 24 to 110 hpf was 57% higher at pH 7.6 than at pH 8.0. Both intergenic and intragenic miRNAs contributed to the acidification-associated response, with no significant preferential representation of either genomic-origin class. Computational predictions link pH-responsive miRNAs to genes associated with aerobic metabolism, morphogenesis, biomineralization, chromatin regulation, and stress responses. Because these miRNA-target relationships are computational predictions and the physiological and molecular measurements were not biologically paired, they do not establish a direct regulatory relationship. These results provide the first larval miRNA catalogue for H. rufescens and identify developmental- and acidification-associated miRNA abundance patterns and candidate miRNA-target relationships for future functional validation.