The aim of this Regional Research Communication was to validate a panel of 30 microsatellite markers recommended by FAO/ISAG for studies of biodiversity in cattle to improve the characterisation of Cuban buffalo populations. The water buffalo (Bubalus bubalis) is an economically important livestock species. Therefore, research focused on the study of the genetic relationships among water buffalo populations is useful to support conservation decisions and to design breeding schemes. Twenty-eight of the 30 tested regions were amplified, one of which (ETH10) turned out to be monomorphic. A total of 143 alleles were observed in the Cuban water buffalo population. The average number of alleles per locus was 5·04. The number of alleles per polymorphic locus ranged from two (INRA 63 and MM12) to nine (ETH185). The observed and expected heterozygosity ranged from 0·108 (HAUT24) to 0·851 (CSSM66) and 0·104 (MM12) to 0·829(INRA32), respectively. The polymorphic information content (PIC) ranged from 0·097 (MM12) to 0·806 (INRA32), and the overall value for these markers was 0·482. Within the population, inbreeding estimates (FIS) was positive in 14 of the 30 loci analysed. This study thus highlights the usefulness of heterologous bovine microsatellite markers to assess the genetic variability in Cuban water buffalo breeds. Furthermore, the results can be utilised for future breeding strategies and conservation.