Application of a dual-locus metabarcoding approach for a more comprehensive account of cattle dietary items in a semi-arid African savanna with special reference to forbs

Increasing livestock densities and more severe drought events challenge sustainable management in South Africa’s semi-arid savannas. Effective mitigation strategies require accurate assessments of livestock foraging behaviour. By utilising high-throughput sequencing technology, this study evaluated the use of a dual-locus metabarcoding approach ( trn L and rbc L) together with study-area-specific reference libraries, to analyse cattle diets in two bioregions of the eastern semi-arid South African savanna. Both markers demonstrated the ability to identify various plant families, but trn L exhibited a higher diversity in terms of family and genus identification at both sampling sites. Forbs, although comprising a diverse component of savanna plant communities, have relatively small above-ground biomass, but can still serve as crucial forage items, especially during dry periods. Our study underscores the significant role of forbs in cattle diets, demonstrating a shift in cattle foraging preferences from grass-based diets to higher inclusions of forbs and woody taxa during the drier season. Although grasses, such as Setaria , were still prevalent, forbs, belonging to the genera Malvastrum , Asparagus , Pollichia and Ipomoea were also important food items for cattle as well as woody taxa belonging to Fabaceae, Combretaceae, Ebenaceae, and Malvaceae with a selection of food items from trees and shrubs from genera Albizia , Combretum , Euclea and Vachellia. Furthermore, our study highlights the value of a dual-locus metabarcoding approach for understanding herbivorous diets. Using trn L and rbc L markers, with study-area-specific reference libraries, improves taxonomic resolution for accurately reconstructing cattle diets in semi-arid savannas. This study may improve biodiversity estimates and inform sustainable rangeland management strategies in semi-arid African savanna ecosystems.