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Accessory respiratory is generally accepted to have evolved independently on numerous occasions in adaption to aquatic hypoxia in freshwater habitats. In general, the air-breathing organ in fish is believed to be structurally modified to supplement respiration. In this study, intuitive evidence for elaborate structural modifications of the intestine, an air-breathing organ in mud loach (Misgurnus anguillicaudatus), compared with two other obligate aquatic breathers, grass carp (Ctenopharyngodon idellus) and yellow catfish (Pelteobagrus fulvidraco), were directly provided by histological and morphometric methods. As a result, a sharply decreasing height of mucosal folds and thickness of muscularis were manifested in loach intestine from its anterior to posterior region. Compared with grass carp and yellow catfish, loach had the smallest ratios of mucosal fold height/muscularis thickness to intestinal lumen radius in the posterior intestine. These suggested that the posterior intestine is the air-breathing location for the loach. Furthermore, length density of capillary (0.46±0.05 μm−2) in the posterior intestine of the loach was significantly higher than those of grass carp and yellow catfish. Meanwhile, diffusion distance of air-blood barrier (1.34±0.04 μm) in the posterior intestine of the loach was significantly smaller than those of the other two fish species. In summary, the characteristics of highly vascularized, short diffusion distance of air-blood barrier, thinned and flattened made the posterior intestine a perfect air-breathing location for the loach.