Investigating the Phylogenetic Relationships of the Iranian Honey Bee with Other Bee Breeds Worldwide Using the Cytochrome Oxidase I Region
Authors: Sarhaddi Avval Mansoureh , Fazeli-Nasab Bahman , Pahlavan Anahita , Mirzaei Ali Reza 
Published: December 2024

Background: Honey bees, classified within the insect class and widely distributed globally, consist of 26 identified subspecies across five evolutionary lines. Among these, the Iranian honey bee (Apis mellifera meda) is notable for its presence in Iran and neighboring regions. This subspecies exhibits unique biological traits, including a high reproductive capacity and effective propolis collection, distinguishing it from other local bee populations. Understanding genetic diversity within these populations is crucial for conservation and breeding efforts. Objectives: The aim of this study is to determine the phylogenetic relationships and natural variations of the Iranian honey bee in comparison to other bee breeds worldwide using the cytochrome oxidase I (COI) region. Methods: The study focuses on the identification of COI gene sequences in honeybees. The mitochondrial COI gene sequences related to the Iranian honey bee and eight populations of other honey bee breeds from around the world (24 ecotypes) were extracted and aligned from the genome database (NCBI). BLAST was used for sequence retrieval, and Clustal W was employed for alignment among species. Phylogenetic analysis was performed using MEGA software, applying the Neighbor Joining method to construct a phylogenetic tree based on the edited sequences. The percentage of nucleotide substitutions and replacements was calculated using the maximum likelihood method. Additionally, molecular diversity indices were computed using DnaSP software to assess genetic variation and conservation parameters among populations. Results: The bioinformatics analysis of biological data revealed that transfer substitution occurred more frequently than crossover substitution. Additionally, the percentage of substitution in pyrimidine bases was higher than in purine bases. The dN/dS substitution ratio in the gene locus sequence across the examined ecotypes was estimated at 0.08, indicating purifying selection during the evolution of this gene locus. Furthermore, examination of the COI region of the mitochondrial genome identified a total of 4 haplotypes, 20 mutations, and 195 polymorphic loci within the entire studied population. The neutrality values, based on Tajima's test, indicated effects related to genetic drift, genetic bottlenecks, or balancing selection during the evolutionary history of the studied population. Phylogenetic analysis revealed nine distinct categories of evolutionary paths for the COI gene in the studied ecotypes. In the phylogenetic tree, the Iranian honey bee was placed in a separate cluster, exhibiting the highest and lowest phylogenetic distances relative to Apis koschevnikovi and Apis mellifera, respectively. Conclusions: This research indicates that the conserved region of the mitochondrial cytochrome oxidase gene among various bee species is minimal, highlighting significant polymorphism and susceptibility to mutations. A total of 20 mutations and 195 polymorphic sites were identified, suggesting purifying selection processes that contribute to the gene's biological functions. The study also demonstrates that the COI region can effectively differentiate Iranian honey bee populations from others globally. Overall, these findings enhance our understanding of genetic diversity and evolutionary relationships within bee species, underscoring the importance of continued research for conservation efforts.