Genetic Characterization of <scp><i>Apis mellifera jemenitica</i></scp> (Hymenoptera: Apidae) Based on Mitochondrially Encoded NADH Dehydrogenase 4 (ND4)

ABSTRACT In this study, sequences of the mitochondrial NADH dehydrogenase 4 (ND4) gene for 137 native Apis mellifera jemenitica colonies from Saudi Arabia were analyzed. Results revealed eight new ND4 nucleotide haplotypes (1311 bp) and four amino acid haplotypes. Most mt ND4 gene sequences (&gt; 92%) of A. m. jemenitica were identical. However, four samples (3%) showed unusually highly polymorphic sequences. Within‐population evolutionary diversity mean in ND4 nucleotide sequences was very low (0.0004). ND4 sequence variations among A . m. jemenitica and the most closely related subspecies A. m. syriaca and A. m. lamarckii were minimal by an average genetic distance of less than 0.005. The lowest evolutionary divergence mean was reported between A. m. jemenitica and A. m. syriaca . Results also showed that most A. mellifera subspecies included in this study could be separated from most A. m. jemenitica samples using one or more variations in amino acid sequences. Distinct amino acid sequence variations, particularly at codon 316, and differences in the restriction map of SspI can separate most A. m. jemenitica samples from the closely related subspecies, A. m. lamarckii and A. m. syriaca . Phylogenetic analyses showed that 19% of the DN4 nucleotide sequences of all samples and 100% of the amino acid sequences from Tabuk and Madinah clustered with the A. m. syriaca reference sequence, indicating a closer evolutionary relationship with the neighboring subspecies, A. m. syriaca . Results also demonstrated the likelihood of a small population with high genetic variability. This might indicate the presence of intensively hybridized colonies within the entire A. m. jemenitica population native to Saudi Arabia. Results also revealed that mt ND4 sequence analyses could be used successfully to distinguish A. m. jemenitica of Saudi Arabia from other A. mellifera subspecies belonging to different evolutionary lineages such as A. m. mellifera , A. m. carnica , A. m. simensis , and A. m. intermissa . The close evolutionary relationship between A. m. jemenitica and A. m. syriaca or A. m. lamarckii and the approaches to separate them should be considered in any conservation efforts of A. m. jemenitica in Saudi Arabia.