Arachidonic acid synthetic pathways of the oyster protozoan parasite, Perkinsus marinus: evidence for usage of a delta-8 pathway

The meront stage of the oyster protozoan parasite, Perkinsus marinus, is capable of synthesizing saturated and unsaturated fatty acids including the essential fatty acid, arachidonic acid [20:4( n−6)]. Eukaryotes employ either delta-6 (Δ-6) or delta-8 (Δ-8) desaturase pathway or both to synthesize arachidonic acid. To elucidate the arachidonic acid synthetic pathways in P. marinus, meronts were incubated with deuterium-labeled precursors [18:1( n−9)-d6, 18:2( n−6)-d4, 18:3( n−3)-d4, and 20:3( n−3)-d8]. The lipids were extracted, converted to fatty acid methyl esters, and analyzed using gas chromatography/mass spectrometry and gas chromatography/flame ionization detection. Deuterium-labeled 18:2( n−6), 20:2( n−6), 20:3( n−6), and 20:4( n−6) were detected in meront lipids after 1-, 3-, 5-, and 10-day incubation with 18:1( n−9)-d6. Deuterium-labeled 20:2( n−6), 20:3( n−6) and 20:4( n−6) were found in lipids from meronts after incubation with 18:2( n−6)-d4 methyl ester. No labeled 18:3( n−6) was detected in either incubation. Apparently, when incubated with 18:1( n−9)-d6, the parasite first desaturated 18:1( n−9)-d6 to 18:2( n−6)-d6 by Δ-12 desaturase, then to 20:2( n−6)-d6 by elongation, and ultimately desaturated to 20:3( n−6)-d6 and 20:4( n−6)-d6 using the sequential Δ-8 and Δ-5 desaturation. Similarly, when incubated with 18:2( n−6)-d4, P. marinus converted the 18:2( n−6)-d4 to 20:2( n−6)-d4 by elongation and 20:2( n−6)-d4 to 20:3( n−6)-d4 by Δ-8 desaturase then by Δ-5 desaturase to 20:4( n−6)-d4. These results provide evidence that P. marinus employed the Δ-8 rather Δ-6 pathway for arachidonic acid synthesis. Additional support for the presence of a Δ-8 pathway was the demonstrated ability of the parasite to metabolize 18:3( n−3)-d4 to 20:3( n−3)-d4 and 20:4( n−3)-d4, and 20:3( n−3)-d8 to 20:4( n−3)-d6 and 20:5( n−3)-d6 using the sequential position-specific Δ-8 and Δ-5 desaturases.