Folia biologica (Kraków), vol. 52 (2004), No 1-2 Notes on Chromosome Numbers and C-banding Patterns in Karyotypes of Some Weevils from Central Europe (Coleoptera, Curculionoidea: Apionidae, Nanophyidae, Curculionidae) Dorota LACHOWSKA, Milada HOLECOVÁ and Maria RO¯EK Accepted January 27, 2004 LACHOWSKA D., HOLECOVÁ M., RO¯EK M. 2004. Notes on chromosome numbers and C-banding patterns in karyotypes of some weevils from Central Europe (Coleoptera, Curculionoidea: Apionidae, Nanophyidae, Curculionidae). Folia biol. (Kraków) 52: 61-66. Chromosome numbers and C-banding patterns of sixteen weevil species are presented. The obtained results confirm the existence of two groups of species with either a small or large amount of heterochromatin in the karyotype. The first group comprises twelve species (Apionidae: Oxystoma cerdo, Eutrichapion melancholicum, Ceratapion penetrans, Ceratapion austriacum, Squamapion flavimanum, Rhopalapion longirostre; Nanophyidae: Nanophyes marmoratus; Curculionidae: Centricnemus (=Peritelus) leucogrammus, Sitona humeralis, Sitona lineatus, Sitona macularis, Sitona suturalis). In weevils with a small amount of heterochromatin, tiny grains on the nucleus during interphase are visible, afterwards appearing as dark dots during mitotic and meiotic prophase. The second group comprises four species from the curculionid subfamily Cryptorhynchinae (Acalles camelus, Acalles commutatus, Acalles echinatus, Ruteria hypocrita) which possess much larger heteropycnotic chromosome parts visible during all nuclear divisions. The species examined have pericentromeric C-bands on autosomes and on the X chromosome. Key words: Curculionoidea, Apionidae, Nanophyidae, Curculionidae, C-bands, chromosome number. Dorota LACHOWSKA and Maria RO¯EK, Department of Experimental Zoology, Institute of Systematics and Evolution of Animals, Polish Academy of Sciences, S³awkowska 17, 31-016 Kraków, Poland. E-mail: [email protected] [email protected] Milada HOLECOVÁ, Department of Zoology, Comenius University, Mlynská dolina B-1, 842-15 Bratislava, Slovakia. E-mail: [email protected] The cytogenetic distribution of heterochromatin Material and Methods can be visualized by the C-banding technique. Chromosome banding analysis is useful in estab- lishing the nature of chromosomal differences, but For the cytogenetic study, adult males were col- unfortunately most karyotypic data have been ob- lected in forest and grassland habitats of Slovakia tained by standard analysis, only a minor fraction and Poland from June to October 2003. Their go- of papers have analysed banded karyotypes of weevils nads were dissected and used as material for (HOLECOVÁ et al. 1997, 2002; HSIAO &HSIAO squashes. The gonads were fixed according to the 1984; RO¯EK &HOLECOVÁ 2000; RO¯EK et al. 2004). method described by RO¯EK (1994) with minor modification (RO¯EK &HOLECOVÁ 2000; RO¯EK The present paper is a continuation of investiga- &LACHOWSKA 2001). C-bands were determined tions on the C-banding patterns of Palaearctic wee- using modifications of the procedure described by vils. The aim of our study is (1) to describe chromo- RO¯EK et al. (2004). The squashed slides were some numbers for 8 weevil species examined for treated with a 0.3 N HCl at 20-23oC for 1min and the first time, (2) to analyse the C-banding patterns with a freshly prepared solution of 5% barium hy- on chromosomes of 16 species, and (3) to investi- droxide at 20-23oC for 2-3 min. Next, they were gate whether the small amount of heterochromatin rinsed with distilled water and incubated in 2xSSC is characteristic for genus, tribe or family. at 50oC for 1h. Dry slides were stained with 4% The systematics of Curculionoidea are based on Giemsa for 10 to 20 min. Observations of chromo- STREJÈEK (1993) and ALONSO-ZARAZAGA and somes and photomicrographs were made using a LYAL (1999). Jenaval light microscope (C. Zeiss, Jena). The ma- 62 D. LACHOWSKA et al. terial is deposited in the Institute of Systematics Eight Apionidae species: Eutrichapion melan- and Evolution of Animals (Kraków). cholicum, Ceratapion penetrans, Ceratapion aus- triacum, Squamapion flavimanum; Nanophyidae: Nanophyes marmoratus; Curculionidae: Acalles Results and Discussion commutatus, Sitona macularis, Sitona suturalis were investigated for the first time. Chromosome numbers were analysed in sixteen In Apionidae meiotic stages were exclusively weevil species from the families Apionidae, Nano- observed in the preparations. All species examined phyidae and Curculionidae (Table 1). The chro- possess the same chromosome number: n%=10+Xyp, mosome numbers of eight species – Rhopalapion 2n=22 (Figs 3-5, 7). The sex chromosomes were longirostre, Oxystoma cerdo, Acalles camelus, connected achiasmatically (Figs 3-5, 7). In Oxys- Acalles echinatus, Ruteria hypocrita, Centricne- toma cerdo, metaphase II was observed (Fig. 2). In mus (=Peritelus) leucogrammus, Sitona lineatus, this species the chromosomes were metacentric, Sitona humeralis, were described earlier (TAKE- with slight differences in length. The X chromo- NOUCHI 1974; PETRYSZAK 1977; HOLECOVÁ et some was the longest, whereas the y chromosome al. 1999a, b; LACHOWSKA et al. 1999, 2001). was the smallest element in the set. These results Table 1 Species of weevils in which chromosomes were examined Family, subfamily, tribe Geographic source Chromosome Species and date of collection number References Family: Apionidae Oxystoma cerdo S Poland, Zawoja, 2n=22 HOLECOVÁ et al. (Gerstaecker, 1854) June 13, 2003 n%=10+Xyp 1999b Eutrichapion melancholicum SW Slovakia, Borská níina lowland, Devínska Nová 2n=22 Present study (Wencker, 1864) Ves, August 21, 2003 n%=10+Xyp Ceratapion penetrans SW Slovakia, Borská níina lowland, Devínske Jazero, 2n=22 Present study (Germar, 1817) August 21, 2003 n%=10+Xyp Ceratapion austriacum SW Slovakia, Malé Karpaty Mts., Devínska Kobyla, 2n=22 Present study (Wagner, 1904) August 18, 2003 n%=10+Xyp Squamapion flavimanum SW Slovakia, Borská níina lowland, Devínske Jazero, 2n=22 Present study (Gyllenhal, 1833) August 21, 2003 n%=10+Xyp Rhopalapion longirostre SW Slovakia, Bratislava env., 2n=22 HOLECOVÁ et al. (Oliver, 1807) June 9, 2003 n%=10+Xyp 1999a Family: Nanophyidae Nanophyes marmoratus SW Slovakia, Borská níina lowland, Devínske Jazero, 2n=22 Present study (Goeze, 1777) Juli 14, 2003 n%=10+Xyp Family: Curculionidae Subfamily: Cryprorhynchinae Tribe: Cryptorhynchini Acalles camelus SW Slovakia, Malé Karpaty Mts., Devínska Kobyla, 2n=30 LACHOWSKA et (Fabricius, 1792) September 28, 2003 n%=14+Xyp al. 2001 Acalles commutatus SW Slovakia, Malé Karpaty Mts., Lozorno, 2n=28 Present study Dieckmann, 1982 October 19, 2003 n%=13+Xyp Acalles echinatus SW Slovakia, Malé Karpaty Mts., Devínska Kobyla, 2n=30 LACHOWSKA et (Germar, 1824) October 12, 2003 n%=14+Xyp al. 2001 Ruteria hypocrita SW Slovakia, Malé Karpaty Mts., Devínska Kobyla, 2n=30 LACHOWSKA et (Boheman, 1837) October 4, 2003 n%=14+Xyp al. 2001 Subfamily: Entiminae Tribe: Peritelini Centricnemus (=Peritelus) SW Slovakia, Malé Karpaty Mts., Devínska Kobyla, 2n=22 PETRYSZAK 1977, Leucogrammus n%=10+Xy LACHOWSKA et Germar, 1824 August 15, 2003 p al. 1999 Sitonini Sitona humeralis SW Slovakia, Borská níina lowland, Devínske Jazero, 2n=22 HOLECOVÁ et al. Stephens, 1831 August 21, 2003, n%=10+Xyp 1999a Sitona lineatus SW Slovakia, Borská níina lowland, Závod-Abrod, 2n=22 TAKENOUCHI (Linnaeus, 1758) August 15, 2003 n%=10+Xyp 1974 Sitona macularis SW Slovakia, Borská níina lowland, Závod-Abrod, 2n=22 Present study (Marsham, 1802) August 15, 2003 n%=10+Xyp Sitona suturalis SW Slovakia, Malé Karpaty Mts., Devínska Kobyla, 2n=22 Present study Stephens, 1831 August 18, 2003 n%=10+Xyp C-banding Patterns in Weevils 63 confirmed an earlier observation (HOLECOVÁ et Acalles. The present karyological observations al. 1999b). In Ceratapion austriacum metaphase confirm the hypothesis that A. commutatus and A. II was constituted by meta-, submeta- and subtelo- echinatus are sibling species differing in morphol- centric chromosomes. The X chromosome was ogy of male genitalia, habitat (microhabitat) pref- submetacentric and the longest, while the y chro- erence and also in karyology. mosome was subtelocentric (Fig. 6). From the genus Centricnemus only one species – The C-banding results revealed that species C. leucogrammus, was analysed and the results from the family Apionidae possess a small amount confirmed earlier observations (PETRYSZAK 1977; of heterochromatin. Short, heterochromatic bands LACHOWSKA et al. 1999). This species has a small were visible in prophase stages (Figs1&8). C-bands in pachytene (Fig. 21). In Nanophyidae only prophase and metaphase Four examined species from the genus Sitona: S. I stages with meioformula n%=10+Xyp were ob- humeralis, S. lineatus, S. macularis, and S. sutu- served in Nanophyes marmoratus. A small amount ralis, possessed the same chromosome number of heterochromatin was visible as in Apionidae. and meioformula n%=10+Xyp, a small amount of In Curculionidae three species from the genus heterochromatin was observed (Figs 21-24). Acalles, one from Ruteria, one from Centricnemus According to the authors’ observations, twelve (=Peritelus) and four from Sitona were analysed. weevils surveyed here share the diploid comple- In Acalles and Ruteria, 30 chromosomes with ment of 22 chromosomes, and the meioformula n%=10+Xyp (Table 1). This is the most character- meioformula n%=14+Xyp were observed. The karyo- types were asymmetric with meta-, submeta- and istic chromosome number for