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A lipid analysis of the tissues of a cold-seep mytilid mussel collected from the Louisiana slope of the Gulf of Mexico was used in conjunction with a compound-specific isotope analysis to demonstrate the presence of methanotrophic symbionts in the mussel gill tissue and to demonstrate the host's dependence on bacterially synthesized metabolic intermediates. The gill tissue contained large amounts of group-specific methanotrophic biomarkers, bacteriohopanoids, 4-methylsterols, lipopolysaccharide-associated hydroxy fatty acids, and type I-specific 16:1 fatty acid isomers with bond positions at ⌬8, ⌬10, and ⌬11. Only small amounts of these compounds were detected in the mantle or other tissues of the host animal. A variety of cholesterol and 4-methylsterol isomers were identified as both free and steryl esters, and the sterol double bond positions suggested that the major bacterially derived gill sterol [11.0% 4␣-methyl-cholesta-8(14),24-dien-3␤-ol] was converted to host cholesterol (64.2% of the gill sterol was cholest-5-en-3␤-ol). The stable carbon isotope values for gill and mantle preparations were, respectively, ؊59.0 and ؊60.4‰ for total tissue, ؊60.6 and ؊62.4‰ for total lipids, ؊60.2 and ؊63.9‰ for phospholipid fatty acids, and ؊71.8 and ؊73.8‰ for sterols. These stable carbon isotope values revealed that the relative fractionation pattern was similar to the patterns obtained in pure culture experiments with methanotrophic bacteria (R. E. Summons, L. L. Jahnke, and Z. Roksandic, Geochim. Cosmochim. Acta 58:2853-2863, 1994) further supporting the conversion of the bacterial methylsterol pool.

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