C 1996 The Japan Mendel Society Cytologia 61: 33-39, 1996 Karyotype Evolution of Drymadusini (Decticinae, Orthoptera) Elzbieta Warchalowska-Sliwa1 and Alexander G. Bugrov2 'Department of Experimental Zoology , Institute of Systematics and Evolution of Animals, Polish Academy of Sciences, Slawkowska 17, 31-016 Krakow, Poland 2Institute of Systematics and Ecology of the Siberian Branch , Russian Academy of Sciences, Frunze 11, 630091 Novosybirsk and Department of Natural Sciences, Novosibirsk State University, 630090 Novosibirsk, Russia Accepted October 30, 1995 The karyotype variability, especially differences in chromosome numbers and morphology permit the investigation of the manner, rate, and form of karyotype evolution in closely related groups. The chromosome systems of the Decticinae have been described by many authors, e.g., by John and Hewitt (1968), White (1973), Hewitt (1979), Ueshima and Rentz (1979), Camacho et al. (1981a), Warchalowska-Sliwa (1984, 1988), Warchalowska-Sliwa et al. (1992, 1994), Warchalowska-Sliwa and Michailova (1993). However, karyological studies of species of the tribe Drymadusini are so far scare and the chromosome numbers of only three species having been reported (Bugrov 1990). The number of the genera in the Drymadusa group was established by Ramme (1939). This group includes the following genera Ceraecercus Uv., Drymadusa Stein., Afrodrymadusa Rme., Paradrymadusa Herm., Drymadusella Rme., Anadolua Rme., Phytodrymadusa Rme., Scotodrymadusa Rme., Ferganusa Uv., Paratlanticus Rme., Atlanticus Scudd., and Bergiola Stschelk. with question mark. Most of the species belonging to Drymadusini show small restricted ranges. Some of them are endemic in various montain ranges. On the other hand, many species are associated with arid or nemoral areas of the Far East (Sergeev 1993). In this study the karyological analysis of eight species of the Drymadusini group was undertaken. The karyotype analysis includes the number and morphology of chromosomes, C-banding and NOR location patterns. Materials and methods Eight species of Drymadusini belonging to the fauna of the maritime region of Russia, Dagestan, Tadzhikistan, Turkmenistan, and North Korea were studied (Table 1). Males were injected with 0.1% colchicine for 1.5-2.0 hr. The testes were fixed in glacial acetic-alcohol (1 : 3), and the fixed materials washed and kept in 70% ethanol or Carnoy solution. Air-dried preparations were made by squashing tissues in 60% acetic acid and then freezing in dry ice. C-bands were obtained by the barium hydroxide denaturation technique (Sumner 1972) with minor original modifications. The silver staining method for NOR regions was performed as previously reported (Warchalowska-Sliwa et al. 1992). Chromosomes were classified according to Levan et al. (1964). Results Chromosome number and shape In Table 2 the number and shape of chromosomes in the species analysed are shown. The chromosome numbers in this group range from 2n•‰ =25 to 31. The karyotype of Drymadu- 34 Elibieta Warchalowska-liwa and Alexander G. Bugrov Cytologia 61 Table 1 . List of taxa and collection localities 1 2 3 4 5 6 Figs. 1-6. Karyotypes of: 1. Drymadusella hissarica-2n,Z -=31, NF=31; 2. Atlanticus brun- neri-2n=29, NF=32, the pair M2 with secondary construction; 3. Anadrymadusa robusta-2n= 27, NF=29, with C-bands, small distal C-bands in M2; 4. Bergiola montana-2n=27, NF=30, 5. Tadzhikia pavlovskii-2n = 27, NF =31 with two metacentric pairs (L1, L2); 6. Anatlanticus koreanus-2n = 25, NF = 29 with two metacentric pairs (L1, L2) and secondary constructions in the X chromosome. The X chromosomes are the last elements of the karyotypes. ( X 800). sella hissarica has a basic complement of 2ncll = 31 acrocentrics and NF (Fundamental Number) = 31 (Fig. 1), whereas in Atlanticus brunneri the complement is reduced to 29, NF = 32. Only one pair of autosomes and the X are metacentric (Fig. 2). Four species with 2no' = 27 chromosomes: Anadrymadusa picta, A. robusta, and Paratlanticus ussuriensis possess NF = 29 with one metacentric pair (Fig. 3), while Bergiola monatana possesses NF = 30 with one metacentric pair and metacentric X chromosomes (Fig. 4). The fifth species with the same chromosome number, Tadzhikia pavlovskii (NF = 31), has two metacentric pairs (Fig. 5). The lowest chromosome number in this group was found in Anatlanticus koreanus which possesses 2n♂=25 and NF=29 and is complemented with two metacentric pairs (Fig. 6). Only in 1996 Karyotype Evolution of Drymadusini 35 Fig . 7 . Karyotype evolution scheme in the Drymadusini. Table 2. General karyotypical features and C-heterochromatin location in the species of Drymadusini analysed (m-metacentric, a-acrocentric, * variation of C-heterochromatin) Drymadusella hissarica and Tadzhikia pavlovskii are the X chromosome the largest element, in the remaining species analysed the X chromosome is the second element in size (Figs. 1-6). At mitotic metaphase with Giemsa staining in the X chromosome of Anatlanticus koreanus and the M2 autosome of Atlanticus brunneri secondary constructions were observed (Figs. 2, 6). The scheme of karyotype evolution in Drymadusini is proposed in Fig. 7. Chromosome C-banding patterns Table 2 shows the C-banding patterns of 7 species analysed, and Figs. 1, 3, 5, 8 and 9 give 36 Elibieta Warchalowska-Sliwa and Alexander G. Bugrov Cytologia 61 8 9 10 11 12 Figs. 8-12. 8, Mitotic metaphase of Tadzhikia pavlovskii with C-bands; thick C-bands in the four small-sized bivalent (arrows). 9, Metaphase I of Anatlanticus koreanus with C-bands, the heteromorphic small bivalent (arrow). 10, Diplotene/diakinesis of Atlanticus brunneri with NOR (arrow). 11, Diplotene of Paratlanticus ussuriensis preserves NORs of the L2 and S bivalents (arrows). 12, Diakinesis of Tadzhikia pavlovskii preserves NORs of the L2 or L3 and S bivalents (arrows). ( X 1,000). some examples of the results. C-bands may show three different locations: paracentromeric, interstitial and distal. Most of the species analysed have paracentromeric C-bands in the vicinity of the centromeric regions. However, in Tadzhikia pavlovskii in two metacentric autosomes (L1, L3) C-bands are not visible (Fig. 5). In most cases, the paracentromeric C-bands are restricted to the centromeric region (thin C-bands), e.g. in the whole autosomes complement of Drymadusella hissarica (Fig. 1), Atlanticus brunneri, Paratlanticus ussuriensis, and Anatlanticus koreanus. In other cases, they occupy the region next to the centromere (thick C-bands) as in the X chromosome of Drymadusella hissarica, Anadrymadusa picta, A. robusta (Figs. 1, 3) and in the four small-sized pairs of Tadzhikia pavlovskii (Fig. 8), whereas the remaining pairs possess thin C-bands. Four of the seven species shown in Table 2 have interstitial C-bands on one or more chromosomes. Interstitial C-bands are located near the paracentromeric region (the X in Anatlaticus koreanus) or the interstitial region (pair M3 in Anadrymadusa picta and A. robusta Fig. 3, pairs M4-M6 in Drymadusella hissarica, and M2 in Atlaticus brunneri). When distal C-bands are present they are located in chromosomes of different size, and are thin in size (Table 2, Figs. 1, 3). It is also worth mentioning that sometimes in one small pair of Anatlanticus koreanus there occurs a variation between the distal C-bands of two homolo- gous chromosomes (Fig. 9). One small pair of Tadzhikia pavlovskii is heteromorphic, but it is very difficult to decide whether this is connected with the occurrence of a supernumerary segment or submetacentric chromosome. 1996 Karyotype Evolution of Drymadusini 37 NOR location The chromosome NOR location is known only in five of the eight species analysed in this work. For Drymadusella hissarica, Anatlanticus koreanus and Bergiola monatna the NOR analysis was not undertaken owing to the absence of diplotene-diakinesis in the studied cells. Three species, namely Atlanticus brunneri, Anadrymadusa picta, and A. robusta, showed a single active NOR located on the M2 bivalent in an interstitial position (Fig. 10). Paratlanticus ussuriensis and Tadzhikia pavlivskii showed two active NORs. However, only one of them showed primary NOR (this activity in the majority of cells of each individual) located on M2 in Paratlanticus ussuriensis and on L2 or L3 in Tadzhikia pavlovskii. The secondary NORs in these species are detected in one of the small bivalents (Figs. 11, 12). Discussion Most of the Old World Decticinae have been recorded as having 2n c71= 31 (NF = 31) (John and Hewitt 1968, White 1973, Hewitt 1979, Warchaiowska-Sliwa et al. 1992, 1994). However, the New World decticine genera, with one or more pairs of metacentrics, diverge from this standard karyotype, thus the chromosome numbers of these species range from 2n = 22 to 31 in the male (Ueshima and Rentz 1979). The results obtained in the cytological analysis of the eight species of Drymadusini described in this paper, indicate the presence of a more intensive karyotype evolution than that in other genera of the Old World Decticinae. The basic karyotype 2n = 31 is present in Drymadusella hissarica, which coincides with the earlier described species Uvarovina daurica and Eulithoxenus mongolicus (Bugrov 1990). On the other hand, the seven species possess karyotypes different from the basic one. This variation is mainly caused by chromosomal rearrangement such as centric fusions and pericentric inversions. In species with other karyotypes, may be distinguished two groups: 1. when NF is similar to the basic number (NF = 31), and 2. when NF is different from it, and some explanation of this change should be sought. A hypothesis of the karyotype evolution of Drymadusini is shown in Fig. 7. Atlanticus brunneri and Tadzhikia pavlovskii belong to the first group, while Atlanticus brunneri shows 2n = 29, with one pair of the metacentric autosomes caused by centric fusion of two pairs of acrocentrics. In such a way, the autosomes are reduced in Metrioptera saussureana (John and Hewitt 1968), Pholidoptera aptera (Warchatowska-liwa 1988), Montana deghestanica (Warchalowska-liwa et al. 1994), and some of the North American Decticinae (Ueshima and Rentz 1979).