Research Article
Research Article
The complete mitochondrial genomes of five Agrilinae (Coleoptera, Buprestidae) species and phylogenetic implications
expand article infoZhonghua Wei
‡ China West Normal University, Nanchong, China
Open Access


Five complete mitochondrial genomes of five species from the subfamily Agrilinae were sequenced and annotated, including Coraebus diminutus Gebhardt, 1928 (15,499 bp), Coraebus cloueti Théry, 1893 (15,514 bp), Meliboeus sinae Obenberger, 1935 (16,108 bp), Agrilus sichuanus Jendek, 2011 (16,521 bp), and Sambus femoralis Kerremans, 1892 (15,367 bp). These mitogenomes ranged from 15,367 to 16,521 bp in length and encoded 37 typical mitochondrial genes: 13 protein-coding genes (13 PCGs), 2 ribosomal RNA genes (2 rRNAs), 22 transfer RNA genes (22 tRNAs), and a control region (CR). Most of PCGs had typical ATN start codons and terminated with TAR or an incomplete stop codon T–. Among these five mitogenomes, Leu2, Ile, Phe, Ser2, Gly, Met, and Val were the seven most frequently encoded amino acids. Interestingly, in A. sichuanus, a 774 bp insertion was present at trnW and trnC junction, which is unusual in Buprestidae. Additionally, phylogenetic analyses were performed based on three kinds of nucleotide matrixes (13 PCGs, 2 rRNAs, and 13 PCGs + 2 rRNAs) using Bayesian inference and maximum-likelihood methods. The results showed that the clade of Buprestidae was well separated from outgroups and all Agrilinae species formed to a single highly supported clade. The tribe Coraebini was polyphyletic, as the genus Meliboeus (Coraebini) clustered with the genus Trachys (Tracheini). The rRNA genes had important impact for the tree topology of Agrilinae. Compared to the tribes Tracheini and Agrilini, the tribe Coraebini is a younger group.


Comparative analysis, mitogenome, phylogenetic analysis


The superfamily Buprestoidea, which contains the families Buprestidae and Schizopodidae, differs from other groups of the Elateriformia by their serrate antennae, hypognathous head, transverse suture of metaventrite present, and two connate basal abdominal ventrites (Bellamy and Volkovitsh 2016). The buprestid beetles are a large group containing six subfamilies, 521 genera, and more than 15,000 species widely distributed in the world (Bellamy 2008; Kubáň et al. 2016). The adults exhibit a broad range of host utilization in leaves, flowers, and stems, whereas larvae are mostly internal feeders on roots and stems, or feed on foliage of woody or herbaceous plants (Bellamy and Volkovitsh 2016). Only adults of the Australian Xyroscelis crocata were reported to feed on the sap of the host plant Macrozamia communis (Bellamy 1997).

Although taxonomists have made important contributions to the buprestid classification of subfamilies and tribes based on several morphological characteristics (Cobos 1980, 1986; Tôyama 1987; Hołyński 1988, 1993, 2009; Bellamy 2003), the problems of the overall classification in Buprestoidea remain unsettled.

In the past two decades, molecular systematic approaches have been used to resolve unsettled classification and phylogenetic relationships in Insecta (Short and Fikáček 2013; Cline et al. 2014; Robertson et al. 2015; Kundrata et al. 2017; Gimmel et al. 2019; Lee et al. 2020). As to Buprestidae, Bernhard et al. (2005) first used molecular phylogenetic methods based on three mitochondrial markers (nad1, 12S, and 16S) and confirmed that the Agrilus viridis complex, which is widely distributed across Eurasia, is monophyletic. Pentinsaari et al. (2014) and Pellegrino et al. (2017) used mitochondrial markers to evaluate the diversity of A. viridis complex, their results suggest that different feeding forms of A. viridis represent distinct species. Subsequently, Evans et al. (2015) performed the first large-scale phylogenetic trees combing nuclear and mitochondrial data from 141 species to understand the higher-level relationships in Buprestidae. In that study, the monophyly of the family Schizopodidae and subfamilies Agrilinae, Julodinae, and Galbellinae were strongly supported, while the interrelationships of Chrysochroinae and Buprestinae remained uncertain. Hansen et al. (2016) used molecular systematic methods based on nuclear and mitochondrial data (coi and ak) to investigate the relationships within Chrysobothris femorata species group, and their results showed that some morphological species were not well separated. Kelnarova et al. (2019) provided a molecular phylogeny of Agrilus species from the Northern Hemisphere and their results suggest that DNA barcoding is a powerful species identification to Agrilus.

During this time, the mitogenome emerged as a valuable source for higher-level phylogenetic analyses, evolutionary strategies, and genetic diversity analyses (Saccone et al. 1999; Krzywinski et al. 2011; Cameron 2014; Qin et al. 2015; Song et al. 2019; Wang et al. 2019). Several buprestid mitogenomes have been sequenced and reported, such as the mitogenome of Chrysochroa fulgidissima (Schönherr, 1817) by Hong et al. (2009); the mitogenome of Agrilus planipennis Fairmaire, 1888 by Duan et al. (2017), who also performed phylogenetic analyses based on 13 PCGs of 45 mitogenomes of coleopterans; the mitogenome of Trachys variolaris Saunders, 1873 by Cao and Wang (2019a); and the mitogenome of Coraebus cavifrons Descarpentries & Villiers, 1967 by Cao and Wang (2019b). More detailed information of buprestid mitogenomes is presented in Table 1.

Table 1.

Information on the mitogenomes of Buprestidae and two outgroups used in this study.

No. Taxa Accession no. Genome size (bp) A% A+T% AT skew GC skew References
1 Coraebus diminutus OK189521 15,499 38.34 68.42 0.12 −0.25 This study
2 Coraebus cloueti OK189520 15,514 38.53 69.27 0.11 −0.25 This study
3 Meliboeus sinae OK189522 16,108 40.18 72.42 0.11 −0.22 This study
4 Sambus femoralis OK349489 15,367 40.98 73.23 0.12 −0.18 This study
5 Agrilus sichuanus OK189519 16,521 40.19 71.73 0.12 −0.21 This study
6 Agrilus planipennis KT363854 15,942 40.25 71.90 0.12 −0.24 Duan et al. 2017
7 Agrilus mali MN894890 16,204 40.34 74.46 0.08 −0.18 Sun et al. 2020
8 Coraebus cavifrons MK913589 15,686 38.94 69.79 0.12 −0.18 Cao and Wang 2019b
9 Trachys auricollis MH638286 16,429 38.94 71.05 0.10 −0.20 Xiao et al. 2019
10 Trachys troglodytiformis KX087357 16,316 41.03 74.62 0.10 −0.19 Unpublished
11 Trachys variolaris MN178497 16,771 39.92 72.11 0.11 −0.21 Cao and Wang 2019a
12 Melanophila acuminata MW287594 15,853 38.74 75.66 0.02 −0.25 Peng et al. 2021
13 Anthaxia chinensis MW929326 15,881 40.12 73.61 0.09 −0.29 Chen et al. 2021
14 Chrysochroa fulgidissima EU826485 15,592 40.31 69.92 0.15 −0.24 Hong et al. 2009
15 Acmaeodera sp. FJ613420 16,217 38.11 68.41 0.11 −0.25 Sheffield et al. 2009
16 Heterocerus parallelus (outgroup) KX087297 15,845 41.90 74.03 0.13 −0.24 Unpublished
17 Dryops ernesti (outgroup) KX035147 15,672 39.04 72.98 0.07 −0.23 Unpublished

Currently, the subfamily Agrilinae contains four tribes (Agrilini, Coraebini, Aphanisticini, and Tracheini); however, the phylogenetic placement of several genera of this subfamily remains unstable. The genera in the tribes Coraebini and Agrilini were revised by Kubáň et al. (2000). In that study, the genus Sambus in the tribe Coraebini was transferred to Agrilini based on the female behavior of ovipositing on rather smooth surfaces of living plants. Later, Kubáň (2016) placed the genera Sambus, Parasambus, and Pseudagrilus in incertae sedis. In order to solve these problems, we contribute mitogenomic data of five species of buprestids, Coraebus diminutus Gebhardt, 1928, Coraebus cloueti Théry, 1893, Meliboeus sinae Obenberger, 1935, Agrilus sichuanus Jendek, 2011, and Sambus femoralis Kerremans, 1892, and perform a molecular phylogenetic analysis in this study. The phylogenetic trees of 15 species from nine genera belonging to four subfamilies of Buprestidae were constructed based on the newly sequenced and previously reported mitogenomes (Table 1).

Material and methods

Sampling and DNA extraction

Specimens of five species were collected using an entomological net. Among them, C. diminutus, C. cloueti, M. sinae, and A. sichuanus were collected in the Dayaoshan Mountains in Guangxi Zhuang Autonomous Region, and S. femoralis was collected at Yingjiang County in Yunnan Province, China. Specimens were immediately preserved in 95% ethanol in the field after collected and then stored at –24 °C in the laboratory. The specimens were identified based on morphological characteristics under a Leica M205 FA stereomicroscope. Total DNA was extracted from muscle tissues using the Ezup Column Animal Genomic DNA Purification Kit (Shanghai, China) following the manufacturer’s instructions.

Sequencing, sequence assembly, annotation, and heterogeneity

DNA sequencing and de novo assembly of each mitogenome were performed by Beijing Aoweisen Gene Technology Co. Ltd (Beijing, China). 22 tRNA genes were identified using the MITOS webserver, with the parameters of the Invertebrate Mito genetic code (Bernt et al. 2013). Their secondary structures were plotted manually from the MITOS predictions using Adobe Illustrator. Every sequence of tRNA genes was manually checked separately. The PCGs were identified as open reading frames corresponding to the 13 PCGs. The rRNAs and control regions were identified by the boundaries of the tRNA genes. The tRNA secondary structures were identified using tRNAscan-SE (Lowe and Chan 2016). Mitogenome maps (Suppl. material 1: Fig. S1) were produced using Organellar Genome DRAW (OGDRAW) (Greiner et al. 2019). The Base composition and relative synonymous codon usage values were determined using MEGA 6.0 (Kumar et al.2016). Strand asymmetry was calculated using the formulae AT-skew = (A – T) / (A + T), and GC-skew = (G – C) / (G + C) (Perna and Kocher 1995). In the control region (CR), tandem repeat elements were detected by Tandem Repeats Finder (Benson 1999). The heterogeneous analysis of the 13 PCGs and two rRNAs datasets were performed using AliGROOVE 1.06 (Kück et al. 2014), and the nucleotide diversity (Pi) and the ratio of Ka/Ks of PCGS were calculated with DnaSP v. 5 (Librado and Rozas 2009).

Phylogenetic analyses

Phylogenetic trees for A. sichuanus, C. diminutus, C. cloueti, M. sinae, S. femoralis, and 10 other buprestid species belonging to four subfamilies were reconstructed by three separate datasets (13 PCGs, 2 rRNAs, and 13 PCGs + 2 rRNAs) using different best-fit models (Table 4). The mitogenomes of Heterocerus parallelus (Heteroceridae) and Dryops ernesti (Dryopidae) were used as outgroups, as they are phylogenetically distant from Buprestidae in the suborder Polyphaga (Xiao et al. 2019). The phylogenetic analyses were performed using PhyloSuite v. 1.2.2 (Zhang et al. 2020). Nucleotide sequences of the 13 PCGs and 2 rRNAs of all 17 mitogenomes were aligned using ClustalW (Thompson et al. 1994) and trimmed using trimAl v. 1.2 (Capella-Gutiérrez et al. 2009). The best-fit model for three datasets was determined by ModelFinder based on Bayesian information criterion. The maximum-likelihood (ML) and Bayesian inference (BI) methods were used to reconstruct the phylogenetic trees by IQ-tree v. 1.6.8 (Guindon et al. 2010) and MrBayes v. 3.2.6 program respectively (Ronquist et al. 2012). Bayesian analyses were run with two independent chains spanning 2,000,000 generations, four Markov chains, sampling at every 100 generations, and a burn-in period of 0.25 for each chain. The phylogenetic trees were edited and visualized by Figtree v. 1.4.3.

Results and discussion

Genome organization and base composition

The complete mitogenomes of the buprestids A. sichuanus, C. diminutus, C. cloueti, M. sinae, and S. femoralis have the following GenBank accession numbers attributed to them: OK189519, OK189521, OK189520, OK189522, OK349489. The mitogenomes of these five species contained the 37 typical mitochondrial genes (13 PCGs, 22 tRNAs, and 2 rRNAs) and a control region (CR) (Table 2). The composition and arrangement of the mitochondrial genes in these species (Table 2) were highly similar as those in most other buprestid species (Duan et al. 2017; Cao and Wang 2019a, 2019b; Xiao et al. 2019; Chen et al. 2021; Peng et al. 2021).

Table 2.

The five newly annotated Buprestidae mitogenomes. The order of these five species in the table is as follows: Agrilus sichuanus, Coraebus diminutus, Coraebus cloueti, Meliboeus sinae, and Sambus femoralis. – not determined.

Gene Strand Position From To Start codons Stop condons Anticodon Intergenic nucleotides
trnI J 1/1/1/1/1 65/63/63/64/65 GAT -3/-3/-3/5-3
trnQ N 63/61/61/70/63 131/129/129/138/131 AAG -1/0/0/0/-1
trnM J 131/129/129/138/131 199/196/196/205/196 CAA 0/0/0/0/0
nad2 J 200/197/197/206/197 1222/1219/1219/1231/1210 ATC/ATT/ATT/ATC/ATT TAA/TAG/TAA/TAA/TAA 1/1/-2/0/-2
trnW J 1224/1221/1218/1232/1209 1293/1286/1283/1303/1273 ACA 774/-8/-13/13/-8
trnC N 2068/1279/1276/1296/1266 2130/1339/1336/1356/1326 GCA 0/2/2/0/0
trnY N 2131/1342/1339/1357/1327 2195/1404/1401/1419/1387 GAA 9/1/1/1/1
cox1 J 2205/1406/1403/1421/1389 3735/2936/2933/2951/2919 –/–/–/–/– TAA/TAA/TAA/TAA/TAA 0/0/0/0/0
trnL2 J 3736/2937/2934/2952/2920 3802/3003/3001/3016/2984 AAG 0/0/0/0/0
cox2 J 3803/3004/3002/3017/2985 4484/3670/3668/3698/3666 ATT/ATA/ATA/ATC/ATT TAA/TAA/TAA/TAA/TAA 0/0/0/0/0
trnK J 4485/3671/3669/3699/3667 4553/3740/3738/3768/3736 CAA 0/0/0/0/0
trnD J 4554/3741/3739/3769/3737 4618/3803/3802/3830/3798 GAC 0/0/0/0/0
atp8 J 4619/3804/3803/3831/3799 4777/3962/3961/3989/3954 ATT/ATA/ATC/ATT/ATA TAG/TAA/TAA/TAA/TAG 0/-7/-7/-7-7
atp6 J 4771/3956/3955/3983/3948 5445/4630/4629/4657/4622 ATG/ATG/ATG/ATG/ATG TAA/TAA/TAA/TAA/TAA -1/-1/-1/-1/-1
cox3 J 5445/4630/4629/4657/4622 6233/5416/5415/5443/5405 ATG/ATG/ATG/ATG/ATG TAG/TAA/TAA/TAA/TAA 8/0/0/0/0
trnG J 6242/5417/5416/5444/5406 6306/5477/5476/5509/5469 ACC 0/0/0/0/0
nad3 J 6307/5478/5477/5510/5470 6660/5831/5830/5863/5823 ATT/ATT/ATT/ATT/ATT TAG/TAG/TAG/TAG/TAG -2/-2/-2/-2/-2
trnA J 6659/5830/5829/5862/5822 6721/5890/5889/5924/5884 AGC 0/-1/-1/-1/0
trnR J 6722/5890/5889/5924/5885 6781/5952/5951/5988/5947 ACG 1/-1/-1/-1/1
trnN J 6783/5952/5951/5988/5949 6849/6017/6016/6051/6013 GAA 0/0/0/0/0
trnS1 J 6850/6018/6017/6052/6014 6916/6075/6074/6117/6080 ACA 1/0/7/-1/0
trnE J 6918/6076/6082/6117/6081 6982/6139/6143/6179/6143 AAC -1/-4/-4/-1/-1
trnF N 6982/6136/6140/6179/6143 7045/6198/6202/6240/6207 GAA 0/0/0/0/0
nad5 N 7046/6199/6203/6241/6208 8768/7915/7919/7960/7915 ATA/ATT/ATT/ATT/ATA TAA/TAA/TAA/TAA/TAA 0/0/0/0/0
trnH N 8769/7916/7920/7961/7916 8830/7977/7981/8026/7978 GAG 0/0/0/0/0
nad4 N 8831/7978/7982/8027/7979 10,166/9295/9299/9362/9308 ATG/ATG/ATG/ATG/ATG TAA/TAA/TAA/TAA/TAA -7/-7/-7/-7/-7
nad4L N 10,160/9289/9293/9356/9302 10,444/9576/9580/9640/9589 ATG/ATG/ATG/ATG/ATA TAA/TAA/TAA/TAA/TAA 4/3/3/2/1
trnT J 10,449/9580/9584/9643/9591 10,511/9642/9646/9704/9654 AGA -1/-1/-1/-1/-1
trnP N 10,511/9642/9646/9704/9654 10,574/9704/9708/9769/9717 AGG 1/1/1/1/1
nad6 J 10,576/9706/9710/9771/9719 11,079/10,185/10,189/10,259/10,192 ATT/ATA/ATA/ATG/ATT TAA/TAA/TAA/TAA/TAA -1/-1/-1/-1/-1
cytb J 11,079/10,185/10,189/10,259/10,192 12,224/11,327/11,331/11,401/11,334 ATG/ATG/ATG/ATG/ATG TAA/TAG/TAG/TAG/TAG 8/-2/-2/-2/-2
trnS2 J 12,233/11,326/11,330/11,400/11,333 12,298/11,391/11,395/11,465/11,400 ACA 17/9/9/19/14
nad1 N 12,316/11,411/11,415/11,485/11,415 13,266/12,361/12,365/12,432/12,365 TTG/TTG/TTG/TTG/TTG TAA/TAA/TAA/TAG/TAA 1/1/1/0/1
trnL1 N 13,268/12,363/12,367/12,433/12,367 13,334/12,427/12,431/12,495/12,434 AAG 0/0/0/0/0
rrnL N 13,335/12,428/12,432/12,496/12,435 14,605/13,693/13,697/13,757/13,692 0/0/0/0/0
trnV N 14,606/13,694/13,698/13,758/13,693 14,674/13,762/13,766/13,826/13,761 AAC 0/0/0/0/0
rrnS N 14,675/13,763/13,767/13,827/13,762 15,379/14,480/14,483/14,531/14,457 0/0/0/0/0
A + T rich region 15,380/14,481/14,484/14,532/14,458 16,521/15,499/15,514/16,108/15,367 0/0/0/0/0

Four of the 13 PCGs (nad1, nad4L, nad4, and nad5), eight tRNAs (trnQ, trnV, trnL1, trnP, trnH, trnF, trnY, and trnC), and two rRNAS (rrnL and rrnS) are encoded on the N-strand, whereas the other 23 genes (9 PCGs and 14 tRNAs) are encoded on the J-strand. The mitogenome sequence of these five buprestid species ranged in size from 15,367 to 16,521 bp.

The mean A + T nucleotide contents of five complete mitogenomes were similar: 68.42% in C. diminutus, 69.27% in C. cloueti, 72.42% in M. sinae, 71.73% in A. sichuanus, and 73.23% in S. femoralis. The entire mitogenomes had a higher A + T contents of 68.42–73.23% (66.05–72.50% for PCGs, 70.95–74.03% for tRNA genes, 75.20–77.33% for rRNA genes, and 74.17–78.38% for the CR) than G + C contents, which is consistent with the typical base of buprestid mitogenomes. The overall AT skews in these five complete mitogenomes were 0.12, 0.11, 0.11, 0.12, and 0.12, respectively. These five species showed a positive TA skew, suggesting that a slight AT bias which are similar to those observed in other buprestid species (Duan et al. 2017; Cao and Wang 2019a, 2019b; Xiao et al. 2019; Chen et al. 2021; Peng et al. 2021).

Protein-coding regions, codon usage, and nucleotide diversity

The total lengths of PCGs in these five buprestid species ranged from 11,090 to 11,158 bp, accounting for 67.54–72.17% of the entire mitogenomes. Similar to the other buprestid mitogenomes, nad5 and atp8 were found to be the largest (1708–1723 bp) and smallest (156–159 bp) genes, respectively. The majority of PCGs strictly started with an ATN (ATA/ATT/ATC/ATG) start codon, except for the nad1 starting with TTG. All PCGs strictly terminated with TAR (TAG/TAA) or an incomplete stop codon T–. Similar to most previously sequenced members of Buprestidae, the AT skew (0.11–0.12) of these five PCGs (Table 3) were similar among the 15 buprestid species. Summaries of the numbers of amino acids in the annotated PCGs and relative synonymous codon usage are presented in Figs 1 and 2. Overall codon usage among the sequenced buprestid mitogenomes was found to be similar, with Leu2, Ile, Phe, Ser2, Gly, Met, and Val being the seven most frequently coded amino acids.

Table 3.

Summarized mitogenomic characteristics of the five buprestid species in this study.

Species PCGs rRNAs tRNA CR
Size (bp) A+T content AT skew Size (bp) A+T content AT skew Size (bp) A+T content AT skew Size (bp) A+T content AT skew
A. sichuanus 11,158 70.08 −0.15 1976 75.96 −0.13 1444 74.03 −0.0009 1142 74.17 0.06
C. diminutus 11,093 66.05 −0.14 1984 75.20 −0.11 1477 70.95 0.03 1019 77.72 0.02
C. cloueti 11,093 67.09 −0.15 1983 75.39 −0.11 1414 71.22 0.019 1031 78.27 0.02
M. sinae 11,135 70.70 −0.15 1967 77.33 −0.11 1435 72.13 0.007 1577 78.38 0.13
S. femoralis 11,090 72.50 −0.16 1954 75.69 −0.13 1430 73.85 0.03 910 75.82 0.18
Figure 1. 

Numbers of different amino acids in the mitogenomes of the five buprestid species; the stop codon is not included. AS: Agrilus sichuanus, CC: Coraebus cloueti, CD: Coraebus diminutus, MS: Meliboeus sinae, and SF: Sambus femoralis.

Figure 2. 

RSCU (relative synonymous codon usage) of the mitogenomes of the five buprestid species; the stop codons are not included.

The nucleotide diversity (Pi) of the 13 PCGs among five species of Agrilinae is provided (Fig. 3), which ranged from 0.202 to 0.375. In these genes, nad2 (Pi = 0.375) presented the highest variability, followed by nad6 (Pi = 0.346), nad4 (Pi = 0.300), and nad5 (Pi = 0.290); cox1 (Pi = 0.20) exhibited the lowest variability. The ratio of Ka/Ks (Fig. 4) for each gene of the 13 PCGs was calculated. The values of nad4 and nad4L are distinctly higher than others, which suggests that the genes nad4 and nad4L have a relatively higher evolutionary rate.

Figure 3. 

Nucleotide diversity (Pi) of 13 PCGs among five newly sequenced Agrilinae mitogenomes.

Figure 4. 

The ratio of Ka/Ks of 13 PCGs among the 15 reported Buprestidae mitogenomes.

tRNA, rRNA genes, and heterogeneity

The length of rrnL genes ranged from 1258 bp (S. femoralis) to 1271 bp (A. sichuanus), whereas rrnS ranged from 696 bp (S. femoralis) to 718 bp (C. diminutus). The A + T content of the rRNA genes ranged from 75.20% (C. diminutus) to 77.33% (M. sinae) (Table 3). Compared with those in other sequenced buprestid mitogenomes, the rRNA genes in these five newly sequenced buprestid mitogenomes are highly conserved (Hong et al. 2009; Duan et al. 2017; Cao and Wang 2019a, 2019b; Xiao et al. 2019; Sun et al. 2020; Chen et al. 2021; Peng et al. 2021). These rRNAs were located between the CR and trnL1, and separated by trnV. The total lengths of the 22 tRNA genes ranged from 1414 bp (C. cloueti) to 1444 bp (C. diminutus), whereas individual tRNA genes typically ranged in size from 58 to 70 bp, among which, eight tRNAs were encoded on the N-strand and the remaining 14 encoded on the J-strand. The secondary structures of tRNAs showed a standard clover-leaf structure (Suppl. material 1: Figs S2–S6), except for tRNA-Ser (Fig. 5) which lacks or has an unusual dihydrouridine arm, which forms a loop commonly found in other insects (Xiao et al. 2011; Park et al. 2012; Yu et al. 2016; Yan et al. 2017; Yu and Liang 2018; Li et al. 2019). In A. sichuanus, the longest intergenic nucleotide (774 bp) was located between trnW and trnC, which is an interesting and specific phenomenon in Buprestidae. The degree of heterogeneity of the 13 PCGs dataset was higher than that of the two rRNAs dataset (Suppl. material 1: Fig. S7). Additionally, the heterogeneity in sequence divergences was slightly stronger for Coraebus than for other buprestid genera (Suppl. material 1: Fig. S7).

Figure 5. 

The predicted secondary structures of the tRNA-Ser in the mitogenomes of the five buprestid species.

Control region

The CR, also known as the A + T-rich region (Wolstenholme 1992), was the largest non-coding region and located between trnI and rrnS. The length of CR ranged from 910 bp (S. femoralis) to 1577 bp (M. sinae). The A + T content (74.17–78.38%) of the CR of these five species was found to be higher than that of the whole genome (68.42–73.23%), PCGs (66.05–72.50%), rRNAs (75.20–77.33%), and tRNAs (70.95–73.85%) (Table 3). Moreover, the compositional analysis revealed that the mitogenomes of the five buprestid species had a positive AT skew (0.02–0.18) in the CR. In these five species, only C. cloueti and C. diminutus had no tandem repeat element detected; however, those of A. sichuanus (20 and 40 bp), M. sinae (53 bp), and S. femoralis (265 bp) had different lengths.

Table 4.

Best-fit models of three datasets used for phylogeny.

ML method BI method
13 PCGs +2 rRNAs GTR+F+I+G4 GTR+F+I+G4

Phylogenetic analyses

Both ML and BI trees using three datasets produced identical topologies (Figs 68), (Buprestinae + ((Chrysochroniae + Polycestinae) + Agrilinae)), in terms of subfamily-level relationship. The monophyly of Buprestidae is corroborated again, as all the buprestid species converged together as an independent clade, and two outgroup taxa obviously separated from the buprestid clade. The target species C. diminutus, C. cloueti, Meliboeus sinae, Agrilus sichuanus, and Sambus femoralis, as well as other species of Agrilinae, converged together as an independent clade. And the target species, M. sinae, was most closely related to the genus Trachys with high value support (Figs 68) which is inconsistent with the previous studies (Kubáň et al. 2000; Evans et al. 2015). The relationship of Agrilinae clades obtained from 2 rRNAs and 13 PCGs + 2 rRNAs datasets are identical but with different topology from the 13 PCGs dataset. In the topology generated from the 13 PCGs dataset, S. femoralis and Agrilus were clustered into a single branch with high support value (Fig. 6, ML: 77, BI: 1) whereas, in the topology generated from the 2 rRNAs and 13 PCGs + 2 rRNAs datasets, S. femoralis split from base of the Agrilinae clades (Figs 7, 8). Based on these results the position of the genus Sambus in the tribe Agrilini was not suitable and suspect. The different tree topologies suggested that the rRNA genes were extremely valuable for the phylogenetic analysis of Agrilinae. Coraebini is the most diverse tribe in Agrilinae, and 10 subtribes are defined (Kubáň et al. 2000). The genus Meliboeus (Meliboeina) and Coraebus (Coraebina) in different clades suggested that the tribe Coraebini was polyphyletic, which is consistent with the previous study of Evans et al. (2015). The samples used in this study might be too limited for a comprehensive phylogeny of Buprestidae which still needs a deep study in the future.

Figure 6. 

Phylogenetic relationships of 15 selected buprestid species using both BI and ML analyses based on 13 PCGs of mitogenomes. The numbers on the branches show posterior probability (BI tree), whereas the values under branches are bootstrap (ML tree).

Figure 7. 

Phylogenetic relationships of 15 selected buprestid species using both BI and ML analyses based on 2 rRNAs of mitogenomes. The numbers on the branches show posterior probability (BI tree), whereas the values under branches are bootstrap (ML tree).


In this study, five mitogenomes (15,367–16,521 bp) were newly sequenced and annotated, including representatives from the tribes Coraebini and Agrilini in subfamily Agriinae. The mitogenomes of the genera Sambus and Meliboeus are reported for the first time. These five sequences showed a positive AT skew, and the amino acids Leu, Ile, Phe, Ser2, Gly, Met, and Val were most frequently used. The secondary structures of tRNA-Ser are absent the D-arm, which is similar to other orders of Insecta. The rRNA genes are valuable for phylogenetic analyses of Agrilinae as they could affect the tree topologies. The results show that Coraebini is polyphyletic, and the genus Sambus belongs to neither Coraebini nor Agrilini. However, more mitogenome samplings are needed to resolve the phylogeny of the Buprestidae in the future to better understand the phylogenetics of jewel beetles.

Figure 8. 

Phylogenetic relationships of 15 selected buprestid species using both BI and ML analyses based on 13 PCGs + 2 rRNAs of mitogenomes. The numbers on the branches show posterior probability (BI tree), whereas the values under branches are bootstrap (ML tree).


I am sincerely grateful to Lanrui Wang (Qingyang, Gansu, China) and Yingqi Liu (China Agricultural University, Beijing, China) for their guidance in using the software. I also thank Dr Hui-Feng Zhao (Langfang Normal University, Hebei, China) and Menglin Wang (China West Normal University, Sichuan, China) for revising the manuscript. This work was supported by the Doctoral Scientific Research Foundation of China West Normal University (20E054).


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Supplementary material

Supplementary material 1 

Figures S1–S7

Zhonghua Wei

Data type: Images (pdf file)

Explanation note: Figure S1. The mitogenome maps of Agrilus sichuanus, Coraebus cloueti, Coraebus diminutus, Meliboeus sinae, and Sambus femoralis. Figure S2. The secondary cloverleaf structure for the tRNAs of Agrilus sichuanus. Figure S3. The secondary cloverleaf structure for the tRNAs of Coraebus cloueti. Figure S4. The secondary cloverleaf structure for the tRNAs of Coraebus diminutus. Figure S5. The secondary cloverleaf structure for the tRNAs of Meliboeus sinae. Figure S6. The secondary cloverleaf structure for the tRNAs of Sambus femoralis. Figure S7. Heterogeneous sequence divergence within datasets 13 PCGs and 2 rRNAs of Buprestidae species.

This dataset is made available under the Open Database License ( The Open Database License (ODbL) is a license agreement intended to allow users to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited.
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