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. 2017 Jul 31;2(2):475–476. doi: 10.1080/23802359.2017.1361344

The complete mitochondrial genome of the cavity-nesting honeybee, Apis cerana (Insecta: Hymenoptera: Apidae) from Borneo

Hisashi Okuyama a, Salim Tingek b, Jun-ichi Takahashi a,✉
PMCID: PMC7800181  PMID: 33473869

Abstract

The complete mitochondrial genome of the cavity-nesting honeybee Apis cerana from Sabah on Borneo Island was analyzed using next-generation sequencing. The mitochondrial genome of A. cerana was a circular molecule of 15,884 bp and was similar to that of the other cavity-nesting honeybee species. The average AT content in the A. cerana mitochondrial genome was 84.4%. It was predicted to contain 13 protein-coding, 22 tRNA, and two rRNA genes, along with one A + T-rich control region.

Keywords: Asian honeybee, Illumina sequencing, genetic distance, Apis cerana, Borneo

The Asian cavity-nesting honeybee, Apis cerana, is widely distributed in the Asian continent and surrounding islands. Multivariate morphometric analysis of A. cerana suggested the presence of four or more subspecies (Ruttner 1988). Mitochondrial DNA analyses of the partial DNA sequences indicated that A. cerana from Borneo has higher genetic diversity compared to that in Asian continent populations (Smith and Hagen 1996; Smith et al. 2000; Takahashi et al. 2002; Tanaka et al. 2003; Arias and Sheppard 2005), but its phylogenetic position remains uncertain (Tanaka et al. 2001; Lo et al. 2010). To our knowledge, this is the first study to analyze the complete mitochondrial genome of A. cerana from Borneo.

Adult workers of A. cerana in Sabah, Malaysia, were collected in March 2000 (the specimen was stored in the National Museum of Nature and Science, Japan, accession number: NSMT-I-HYM74241). Genomic DNA isolated from worker was sequenced using Illumina’s HiSeq platform. The resultant reads were assembled and annotated using the MITOS web server (Germany; Bernt et al. 2013) and Geneious R9 (Biomatters, Auckland, New Zealand). A phylogenetic tree was constructed using MEGA6 (Tamura et al. 2013) and TREEFINDER v.2011 (Jobb et al. 2004) by using the nucleotide sequences of the 13 protein-coding genes.

The A. cerana mitochondrial genome was found to form a closed loop that is 15,884 bp long (AP018149). The A. cerana mitochondrial genome represented a typical hymenopteran pattern and was similar to the common A. cerana mitochondrial genome organization, comprising 13 protein-coding genes, 22 putative tRNA genes, two rRNA genes, and an A + T-rich control region. The average AT content of the A. cerana japonica mitochondrial genome was 84.36%. Similar to honeybee mitochondrial genomes, the heavy strand encoded nine protein-coding genes and 14 tRNA genes, and the light strand encoded four protein-coding genes, eight tRNAs, and two rRNA genes. The ATP6 and ATP8 genes shared 19 nucleotides. Nine protein-coding genes of the A. cerena mitochondrial genome started with ATT; ATP6, COIII, and Cytb genes, with ATG; and ATP8 gene, with ATC, all of which have been commonly found in the A. cerana mitochondrial genome (Tan et al. 2011; Takahashi et al. 2016; Okuyama et al. 2017). The stop codon of each of these protein-coding genes was either TAA, similar to the case in other honeybees. All of the tRNA genes typically possessed cloverleaf secondary structures, except for tRNA-Ser, which lacked the dihydrouridine arm.

Phylogenetic analysis was conducted using 13 mitochondrial protein-coding genes with 19 closely related taxa (Figure 1). The genetic distance and mutation site of 13 mitochondrial protein-coding genes in the A. cerana subspecies mitochondrial genome were 0.0195 and 206 on average, respectively. The Bornean A. cerana was closer to A. cerana cerana than to A. cerana japonica, consistent with the findings of previous multivariate morphometric studies (Radloff et al. 2010). The complete mitochondrial DNA sequence of A. cerana has only been identified for seven populations, including that from Borneo. However, the complete mitochondrial DNA sequence provides additional valid information for assessing the phylogeny of this group.

Figure 1.

Figure 1.

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Phylogenetic relationships (maximum likelihood) among the species of the genus Apis (Hymenoptera) determined using the mitochondrial DNA nucleotide sequences of the 13 protein-coding genes. Numbers beside the nodes are percentages of 1000 bootstrap values. Apis florea and Apis andreniformis were used as an out-group. Alphanumeric terms in parentheses indicate the GenBank accession numbers.

Acknowledgements

We are grateful to Prof. Tadaharu Yoshida and Prof. Masami Sasaki (Tamagawa University) for kindly providing honeybee sample.

Disclosure statement

The authors report no conflicts of interest. The authors alone are responsible for the content and writing of this article.

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