First cytogenetic information on four checkered beetles (Coleoptera, Cleridae).

The karyotypes of four species of Cleridae (Coleoptera): Trichodes favarius (Illiger, 1802), Trichodes quadriguttatus Adams, 1817, Trichodes reichei (Mulsant et Rey, 1863), and Tilloidea transversalis (Charpentier, 1825) were reported for the first time with this study. The chromosome numbers of these four species were determined as 2n = 18, sex chromosome system Xyp, and all chromosomes were metacentric (the except y chromosome). Together with this study, the chromosome data of only 17 species are available in this family. It is remarkable that all of them display the same chromosome number and similar karyotypes. This may make the effect of karyotypical features important in interpreting the evolutionary process of Cleridae. Atılay Yağmur Okutaner.

Introduction

The containing 16 families and including approximately 10,000 taxonomically defined species is an important superfamily of (Gimmel et al. 2019). After , is the second largest Cleroid family with almost 3700 species and 350 genera in 13 subfamilies described so far (Opitz 2010; Bulak et al. 2012; Gunter et al. 2013; Gerstmeier 2018). are widespread in all continents (except for the Antarctic) and has the highest diversity in the tropics (Gunter et al. 2013). Former analyses of phylogenetic and taxonomic relationships of were especially based on morphology (Gerstmeier and Eberle 2011; Opitz 2012; Gunter et al. 2013). Therefore, these relationships were generally determined according to morphological characters with traditional classification systems. The molecular phylogeny of the family is extensively discussed in Gunter et al. (2013).

The data given by chromosomal characters may help to understand the evolutionary relationships of species or higher taxa. Karyological data from the studies in recent years present important findings of genetic structure, life cycle, ecological characteristics, evolution, taxonomy, and phylogeny of insects (Shaarawi and Angus 1991; Gokhman and Kuznetsova 2006). For those reasons, karyotypic features may be referable as a taxonomic character in solving taxonomic problems, assessing relationships, and phylogenetic classification. (Dobigny et al. 2004; Gokhman and Kuznetsova 2006; Miao and Hua 2017).

Although the have a large representative and wide distribution area, only 18 species (13 , 5 ) of the superfamily have been cytogenetically studied so far. The 13 species of in five genera ( Gahan, 1910, Kirby, 1818, Latreille, 1806, Herbst, 1792, and Olivier, 1795) display monotypic chromosome number as “2n = 18”, the basic sex chromosome system for as Xyp, and metacentric/submetacentric morphology for all chromosomes (Smith and Virkki 1978; Schneider et al. 2007; Mendes-Neto et al. 2010).

This study was carried out to support cytogenetic data of the family . The chromosomal first data belonging to four species, (Illiger, 1802), Adams, 1817, (Mulsant et Rey, 1863), and (Charpentier, 1825) were given in this study.

Material and methods

The localities of collected adult specimens are as follows: 16 (Illiger, 1802): Hıdırbey village of Samandağ county in Hatay province, ; 13 Adams, 1817: Göksun county in Kahramanmaraş province ; 8 : Sıddıklı town in Kırşehir province and 14 (Charpentier, 1825): Kesikköprü town in Kırşehir province (Leg: A.Y. Okutaner). The specimens were identified by Hüseyin Ozdikmen and were stored in Zoology Lab of Kırsehir Ahi Evran University.

Living beetles were transferred to the laboratory. The gonads and midguts were dissected and isolated from abdominal contents with the aid of a stereomicroscope microscope. The chromosomal preparation procedure was performed according to the method described by Rozek (1994) with partial modifications. The chromosomal preparation procedure in this study was based on the method described by Rozek (1994) with some modifications. The tissues were treated 15–30 min at room temperature with a hypotonic solution containing 1% sodium citrate and 0.005% w/v colchicine. Tissue samples were transferred to cryotubes including 3:1 ethanol: acetic acid solution and stored in the freezer. Each treated sample was placed on a clean slide and disintegrated lightly. With the subsequent addition of the acetic acid: distilled water (1:1) solution, another slide was firmly covered over this slide. These slides were immediately frozen in liquid nitrogen and uncoupled to be stained in 4% Giemsa solution.

The chromosomes of females were obtained only from . Meiotic chromosome sets of all species were obtained from testis tissues. The chromosome sets fixed on the slides were photographed at 100X magnification with Olympus BX53F microscope equipped with a camera. Chromosome measurements were calculated in terms of µm using the “ImageJ” program with the “levan” plug-in. The chromosome measurements were made from different meiosis metaphase plates of each species and the ideograms were formed with the average for these measurements.

Results and discussion

The number of the diploid chromosome complement was determined as 2n = 18 and the sex chromosome system as Xyp for each species: , , , and . The males of these four species display n = 8 + Xyp meioformula. Their chromosome sets (autosomes and X chromosomes) consist of metacentric chromosomes except for subtelocentric y chromosome. Sex chromosome system (association of Xyp) in meiosis I, and the presence of y chromosome in meiosis II were clearly demonstrated (Figs 1, 2).

Figure 1.

A Female Mitotic metaphase of B, C male meiotic metaphases of (B meiosis II; C meiosis I) D, E male meiotic metaphases of (D, E meiosis II) F male mitotic metaphase of G, H male meiotic metaphases of (G meiosis I; H meiosis II) I male mitotic metaphase of J, K male meiotic metaphases of (J, K meiosis II) L male mitotic metaphase of .

Figure 2.

Ideograms of the haploid chromosomes.

The idiogram shows that the first two chromosome pairs of the species belonging to the genus are larger than others and a gradual decrease in size in the karyotype of (Fig. 2).

In the previous literature, there is cytogenetic information of only 13 checkered beetles (2 subfamilies, 5 genera). Additionally, cytogenetic data of 4 different species were presented for the first time in this study. After all given data, the diploid chromosome numbers have been presented as 2n = 18 and the sex chromosome system as Xyp of all these 17 species. However, four species of have observed different chromosome numbers and two different sex chromosome systems XO and Xyp, the chromosome morphologies of these four species are metacentric except for the y chromosome as similar to the (Table 1).

Table 1.

The chromosome data of the and .

Taxa	Haploid Formula	Diploid Number/Formula	Citations
Cleridae
Thanasimus dubius (Fabricius, 1777) (Clerinae)	8+Xyp		Smith (1950)
Trichodes nutalli (Kirby, 1818) (Clerinae)	8+Xyp		Smith (1953)
Enoclerus nigripes rujiventris (Spinola, 1844) (Clerinae)	8+Xyp	18
Enoclerus sp. (Clerinae)	8+Xyp	18	Smith (1960)
Trichodes ornatus (Linsley et MacSwain, 1943) (Clerinae)	8+Xyp	18
Thanasimus formicarius (Linnaeus, 1758) (Clerinae)	8+Xyp	18	Virkki (1960)
Trichodes apiarius (Linnaeus, 1758) (Clerinae)	8+Xyp	18
Enoclerus sp. (Clerinae)	8+Xyp		Virkki (1963)
Priocera spinosa (Fabricius, 1801) (Clerinae)	8+Xyp
Enoclerus moestus (Klug, 1842) (Clerinae)	8+Xyp	18	Smith and Virkki (1978)
Thanasimus undatulus (Say, 1835) (Clerinae)	8+Xyp
Necrobia ruficollis (Fabricius, 1775) (Corynetinae)	8+Xyp	18	Yadav and Dange (1989)
Necrobia rujipes (De Geer, 1775) (Corynetinae)	8+Xyp	18
Trichodes favarius (Illiger, 1802) (Clerinae)	8+Xyp	18	This Study
Trichodes quadriguttatus Adams, 1817 (Clerinae)	8+Xyp	18
Trichodes reichei (Mulsant et Rey, 1863) (Clerinae)	8+Xyp	18
Tilloidea transversalis (Charpentier, 1825) (Tillinae)	8+Xyp	18
Melyridae
Endeodes collaris LeConte, 1853 (Malachiinae)		18+X0	Smith and Virkki (1978)
Collops sp. (Malachiinae)		16+X0
Hoppingiana hudsonica LeConte 1866 (Dasytinae)		12+Xyp
Astylus variegatus (Germar, 1824) (Melyrinae)		16+Xyp	Schneider at all (2007)
Astylus antis (Perty, 1830) (Melyrinae)	8+Xp or yp	16+Xyp	de Oliveira Mendes-Neto et al. (2010)

Diploid chromosome number 20 and sex chromosome system Xyp are considered ancestral cytogenetic features of , especially the (Smith and Wirkki 1978). According to the limited number of previous studies, it can be said that 2n = 18 chromosome numbers formed by decreasing the ancestral chromosome set (2n = 20) and Xyp sex chromosome system belonging to family are quite conservative.

Although it shows variation in the family , the numerical changes of chromosomes may not have an important role in the karyotypic evolution of the family . Except for the Y chromosome, the metacentric/submetacentric form of all chromosomes may have created a balance for the karyotype of the species. The absence of acrocentric and telocentric chromosomes can reduce the possibility of new centric fusions such as Robertsonian Translocation (Schubert 2007; Chmátal et al. 2014). On the other hand, being resistant to mechanism of chromosome aberration such as chromosome breaks and euploidy may also have created chromosome number stability in the evolutionary process of the family.

In all these respects, the stability of the chromosome set of the family is quite remarkable. If these results can be supported by expanding further studies, the cytogenetic features of would be very useful taxonomic and evolutionary characters.