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The acousto-optic mode coupling in grapefruit microstructured optical fibers (GMOFs) has been investigated. Due to the presence of grape-like large air holes in the fiber cladding, unequal acoustic modulation is generated, and orthogonal acoustic gratings come into being in the GMOF, which is verified by our experimental observation on the mode profile of resonance peaks. The experimental results indicate that two resonance peaks originating from the mode coupling in acoustic-birefringence-induced orthogonal acoustic gratings simultaneously exist in the transmission spectrum, and the wavelength spacing between them reaches as large as 227.4 nm, which indicates that a greatly enhanced acoustic birefringence has been introduced in the GMOF. And moreover, some important grating features, including frequency-dependent grating period, frequency responses of acoustic phase velocity, and acoustic dispersion, have been investigated for the orthogonal acoustic gratings. These studies would be of importance for understanding the acoustic birefringence mechanism in MOF-based acoustic gratings and development of related acousto-optic devices.