The complete mitochondrial genome of the firefly, Diaphanes citrinus (Olivier), (coleoptera: Lampyridae).

We report the complete mitochondrial genome of firefly, Diaphanes citrinus (Olivier). The circular genome of 18,594 bp and has a base composition of A (41.26%), C (11.64%), G (10.18%), T (36.92%). Similar to other Metazoa, our sequence contains 13 protein-coding genes, 21 transfer RNA genes, two ribosomal RNA genes, and a non-coding AT-rich region. We sequenced the mitochondrial genome of fireflies to analyze phylogenetic relationships and deduce the evolution of their flashing signals.

Introduction

Firefly flashing has fascinated humankind for millennia, and its mystery has been gradually solved (Trimmer et al. 2001; Ghiradella and Schmidt 2004; Aprille et al. 2004). The genus Diaphanes, was established by Motschulsky (1853), has over 90 species recorded from the Oriental and the Ethiopian realms (Olivier 1907, 1910; Mcdermott 1964, 1966). Among this spieces, Diaphanes citrinus is uniqe by its diaphanous area, alongate body, and its special orange coloration (Jeng et al. 2001; Fu 2014).

Mitochondrial genome sequences are essential to a deeper understanding of the evolution of Lampyridae and other luminescent beetles (Ermakov et al. 2006). Here, we elucidate the mtDNA genome of D. citrinus.

Specimens used in this study were collected from Huanjiangmulun attractions, Hechi City, Guangxi Province(25°05′N, 107°93′E), and were stored in Natural History Museum, Huazhong Agricultural University, Wuhan, Hubei, China (its accession no. DC2014092405).

Specific primers were designed based on these conserved regions sequences. The PCR reaction was carried out with LA Taq polymerase for 35 cycles at 94 °C for 30 s, and annealedat 50 °C for 30 s, followed by extension at 72 °C for 1 min per 1 kb. Sequences were assembled using the software DNAstar v7.1 (DNAstar, Madison, WI) and adjusted manually to generate the complete sequence of mitochondrial DNA.

The complete mitochondrial genome sequence of D.citrinus (GenBank:MH651351) has 18594 bp and has a base composition of A (41.26%), C (11.64%), G (10.18%), T (36.92%). Similar to other Metazoa, our sequence contains 13 protein-coding genes, 21 transfer RNA genes, two ribosomal RNA genes, a non-coding AT-rich region and one tandem repeats unit(TRU), which represents a typical insect mitochondrial genome (Wolstenholme 1992). The open frames of the 13 protein-coding genes were inferred from three other fireflies:Aquatica leii, Luciola substriata, and Pyrocoelia rufa (Lee et al. 2003; Jiao et al. 2015; Mu et al. 2016). All 13 PCGs initiated with ATN (ATT, ATA, and ATG) codon. Among those genes, four PCGs (ATP6, ND4, ND4L, CTYB) initiated from ATG, and five PCGs (COI, ND2, ND5, ND6, ND1) initiated from ATT, and four PCGs (COII, COIII, ND3, ATP8) initiated from ATA. In addition, an incomplete terminal codon namely single T was found in five PGGS (COII, COIII, ND5, ND4, ND6), In case of other eight PGGS, TAA (ND2, COI, ATP8, ATP6, ND4L) or TAG (ND3, CTYB, ND1) was used.

The phylogenetic tree among the 12 species based on mitochondrial genome sequences were aligned in MEGA 5(Phoenix, AZ) (with 1000 bootstrap replicates) to construct a Neighbour-Joining tree (Figure 1). The result shows D.citrinus is most closely related to Pyrocoelia rufa, which belongs to an entirely different genus in the Lampyridae.

Figure 1.

Molecular phylogeny of D.citrinus and 11 other firefly species based on the complete mitochondrial genome. The complete mitochondrial genome was downloaded from GenBank and the phylogenic tree was constructed by neighbour-joining method with 1000 bootstrap replicates. MtDNA accession numbers used for tree construction are as follows: Pteroptyx maipo (MF686051), Aquatica ficta (KX758085), Pyrocoelia rufa (AF452048), Aquatica leii (KF667531), Luciola curtithorax (MG770613), Aquatica wuhana (KX758086), Luciola cruciata (AB849456), Asymmetricata circumdata (KX229747), Aquatica lateralis (LC306678), Abscondita anceyi (MH020192) and Luciola substriata (recently identified as Sclerotia flavida by Ballantyne et al. 2016)(KP313820).

In conclusion, the complete mitochondrial genome sequence of D.citrinus provides an important molecular framework for further phylogenetic analyses of fireflies.