Abstract: Sthenoteuthis oualaniensis constitutes a significant fishery resource in the South China Sea. Previous studies have indicated that intestinal microbiota varies with sex and sexual maturity and is closely associated with growth traits and feeding patterns. Among them, the intestinal microbiota in female sexual maturity of medium-form populations of S. oualaniensis differed significantly from those of the micro-form population and individuals of different sexes. To investigate the intestinal microbial community structure and metabolic functions of the cultivable microbiota in female sexual maturity of the medium-form populations of S. oualaniensis, we collected the specimens from the offshore northern South China Sea and the northern Nansha fishing ground, and comparatively analyzed the intestinal microbial community structure, and the metabolic profiles of culturable isolates on carbon and nitrogen substrates. Mycoplasma dominated the intestinal microbiota in both regions, accounting for 82.95% and 93.37% of relative abundance, respectively. There were no significant differences in the α-diversity indices (Shannon, Simpson, Ace and Chao1) (p>0.05). However, six non-dominant bacterial genera, including Enhydrobacter of the phylum Proteobacteria, exhibited significantly higher abundances in the offshore northern South China Sea than in the northern Nansha fishing ground (p<0.05). NMDS ordination revealed a clear spatial separation between the two regions, indicating that environmental heterogeneity drove differentiation among non-dominant taxa. Functional predictions indicated highly similar between regions, particularly in major COG categories such as translation and ribosomal structure and amino-acid transport and metabolism. The average well color development (AWCD) values of the culturable microbial communities peaked at 168^th hour (1.48) and 120^th hour (1.46), respectively. Carbohydrate utilization was highest at 24^th hour, with absorbance values of 10.97 and 11.84. The culturable gut microbiota showed high utilization (Rs>1.0) for 13 and 12 carbon and nitrogen sources in the two regions, with no significant difference in diversity indices (p>0.05). Overall, the intestinal microbiota of sexually mature, medium-form female S. oualaniensis exhibited conserved core taxa, α-diversity, functional repertoire, and metabolic characteristics across the two regions, whereas divergence occurred only among non-dominant taxa diverged in response to environmental heterogeneity.