Linking cuticle chemistry and function: long-chain alkanes drive variation in cuticular water loss in Themeda triandra, a widespread C4 perennial grass

Cuticular conductance (gmin) plays a key role in limiting non-stomatal water loss, yet the structural and biochemical determinants of this trait remain poorly understood in grasses. We investigated drought responses in six accessions of Themeda triandra, spanning a wide climatic range, to understand how cuticle traits regulate variation in gmin. We measured cuticle thickness (light microscopy), % epicuticular wax cover (scanning electron microscopy), alkane chemistry (gas chromatography) and leaf biomechanical properties (via fracture tests). gmin was measured as part of a previous study. Alkane composition - particularly a higher abundance of C31 - was consistently associated with lower gmin, pointing to a biochemical basis for reduced cuticular water loss. In contrast, % epicuticular wax cover, cuticle thickness, and leaf toughness varied among accessions and treatments but showed no relationship with gmin. These results indicate that cuticular water loss in T. triandra is regulated primarily by biochemical traits rather than by structural or mechanical traits. They emphasize the importance of considering cuticle chemistry, rather than surface coverage or thickness, when studying drought tolerance in grasses, highlighting the biochemical basis of adaptive diversity in this widespread native species.