Heredity 56 (1986) 365—372 The Genetical Society of Great Britain Received 18 September 1985

Cytogenetic studies in gomphocerine . I. Comparative analysis of chromosome C-banding pattern

J. Cabrero and J. P. M. Camacho Departamento de Genética, Facultad de Ciencias, Universidad de Granada, 18071 Granada, Spain.

Chromosome C-banding pattern have been studied in 20 species of gomphocerine grasshoppers. Paracentromeric C- bands were present in the chromosomes of all species analysed, which favours the equilocal model of heterochromatin distribution. Interstitial or distal C-bands, on the other hand, appeared, at most, in two different chromosome pairs within the same chromosome complement, which is not expected on the equilocal model. Interspecific comparisons of C-banding patterns reveal the existence of differences between species of either the same or of different genera, which emphasises the dynamic nature of heterochromatin. The shows more similarities for C-banding between the species of the genera Stenobothrus, Euchorthippus, and Omocestus than between these species and those of , which seem to indicate the existence of a close relationship between these three genera.

INTRODUCTION caught in the South of the Iberian Peninsula, and belong to the subfamily (Orthop- Acrididgrasshoppers are very uniform in chromo- tera, ). They include representatives of some form and number (John and Hewitt, 1966). 20 species: specifically Truxalis nasuta (L.), For this reason this family has long been regarded Dociostaurus genei (Ocsk.), Ramburiella hispanica as chromosomally conservative since the majority (Ramb.), Euchorthippus chopardi Desc., E. pul- have 2n =22+XOc/XXtelo-subtelocentric vinatus (F.-W.), Stenobothrus festivus Bol., S. chromosomes. The Gomphocerinae is the most grammicus (Caz.), Omocestus bolivari Chop., 0. distinctive subfamily within the Acrididae since panteli (Bol.), 0. raymondi (Yers.), 0. !lorenteae about half of the species analysed to date have Stauroderus scalaris (F.-W.), Chorthippus 2n = Pasc., 16+XOd/XX9with three long metacentric parallelus (Zett.), Ch. dorsatus (Zett.), Ch. jucun- chromosome pairs which are presumed to have dus (Fisch.), Ch. apicalis (H.-S.), Ch. nevadensis arisen by centric fusions. Despite this apparent Pasc., Ch. binotatus (Charp.), Ch. vagans karyotypic conservatism, the Acrididae show (Eversm.) and Ch. brunneus (Thunb.). In all a extensive intraspecific variation for supernumerary total of 771 male and 334 female adults were ana- chromosome segments (John and King, 1977; lysed together with 697 embryos. 1982; 1983; Hewitt, 1979; Camacho et a!., 1984), Adult individuals were analysed cytologically as well as interspecific variation for C-banding by the methods described in Camacho and Cabrero (King and John, 1980; Santos et a!., 1983) and (1982). The embryos were studied cytologically nuclear DNA content (John and Hewitt, 1966; following the technique described by Webb et al. Kiknadze and Vysotskaya, 1969; Rees et a!., 1978; (1978), and C-banding was carried out as Gosálvez et a!., 1980). described in Camacho et a!. (1984). In this paper we analyse C-banding pattern in In all species analysed the autosomes have been 20 species of gomphocerine grasshoppers in order classified into three size groups: L (long), M to determine the interspecific differences which (medium) and S (short). exist between them. RESULTS MATERIALSAND METHODS Inthe majority of the 20 species analysed there Allthe individuals analysed in this report were were variations in the C-banding pattern involving 366 J. CABRERO AND J. P. M. CAMACHO the presence of supernumerary chromosome seg- extremes, though they may vary even between ments. These variations will be reported separately, different chromosomes of the same species. For and here we will consider the standard C-banding example, in Ramburiella hispanica (fig. 3) while pattern found in a majority of individuals within the four larger chromosome pairs have large para- the species analysed. centromeric C-bans the remainder have medium C-heterochromatin content and distribution in or small bands. In Omocestus panteli (fig. 9) para- the 20 species are shown in figs. 1-20 and table 1. centromeric C-bands are medium sized, with C-bands may show three different locations: para- exception of that in the S8 which is small. Chorthip- centromeric, interstitial and distal. Striking inter- pus apicalis (fig. 16) provides an extreme example specific differences exist for C-banding pattern in of the heterogeneous distribution of paracen- respect of the presence or absence of interstitial tromeric C-heterochromatin since it shows large and distal C-bands but more especially for the size C-bands in the L1, L2, L3, M6 and S8 chromosomes, of the paracentromeric bands. medium C-bands in the M7 and small C-bands in the X, M4 and M5. Other species show paracen- ParacentromericC-bands tromeric C-bands of medium size in all chromo- somes; this is so in Euchorthippus chopardi (fig. 4), Thechromosomes of all species analysed, like E. pulvinatus (fig. 5), Stenobothrusfestivus (fig. 6), those studied by King and John (1980) and Santos S. grammicus (fig. 7), Omocestus bolivari (fig. 8), et a!. (1983), have C-bands in the vicinity of the 0. llorenteae (fig. 11), Chorthippus parallelus (fig. centromeric regions. However, these paracen- 13), Ch.jucundus (fig. 15), Ch. nevadensis (fig. 17) tromeric C-bands may be small, representing and Ch. vagans (fig. 19). strictly centromeric heterochromatin, or large, in which case they are composed of both centric and interstitialC-bands proximal bands. The smallest centromeric C-bands occur in Omocestus raymondi (fig. 10) and Whileinterstitial bands are frequent in some (fig. 14). Large compound species of Australian acridoids (King and John, paracentromeric C-bands are clearly exemplified 1980; John et aL, 1985) they are invariably scarce in Stauroderus scalaris (fig. 12), Chorthippus bino- in Spanish species (Santos et a!., 1983). In the tatus (fig. 18) and Ch. brunneus (fig. 20). In a present study we have observed them in 11 species majority of the species, however, paracentromeric but even here they never occur in more than two C-bands are intermediate between these two size chromosome pairs in any one species. A majority

Table 1C-banding pattern in 20 species of gomphocerine grasshoppers Paracentromenc C-bands Interstitial Distal Species 2n Fig. Large Medium Small C-bands C-bands

Dociostaurus genei 23 1 — L2-M4, M8 L1, M5-M7, X, M9-S11 L1 — Truxalis nasuta 23 2 — M9, 510 L1-M8, S11, X M9 M9 Ramburiella hispanica 23 3 L1M4 M5-S11, X — — — Euchorthippus chopardi 17 4 — All — X L3 Euchorthippus pulvinatus 17 5 — All — M6 — Stenobothrusfestivus 17 6 — All — — M6 Stenobothrus grammicus 17 7 — All — — — Omocestus bolivari 17 8 — All — — X Omocestus panteli 17 9 — All except S8 S8 L3 — Omocestus raymondi 17 10 — — All X L2, L3 Oinocestus llorenteae 17 11 — All — X — Stauroderus scalaris 17 12 All — — — — Chorthippus parallelus 17 13 — All — — L3 Chorthippus dorsatus 17 14 — — All — — Chorthippusjucundus 17 15 — All — L2, L3 — Chorthippus apicalis 17 16 L1-L3, M6, S0 M7 M4, M5, X — M Chorthippus nevadensis 17 17 — All — L2 M6, M7 Chorthippus binotatus 17 18 All — — — — 17 19 — All — L2, L3 M4 17 20 All — — L2, L3 M7 CYTOGENETIC STUDIES IN GOMPHOCERINE GRASSHOPPERS. I 367 C -l 0 0 — al -& r -4 0 0 • •2 4.. ee1 ¼ r - 0 S 4Sr (4 r p.) UI 0 1 1:: S S • a r r S W a°"e

Figures1—5 C-banding pattern in the haploid chromosome complements of gomphocerine grasshoppers. (1) Dociostaurus genei, (2) Truxalis nasuta, (3) Ramburiella hispanica, (4) Euchorthippus chopardi, (5) E. pulvinatus. of interstitial C-bands are located in the long (L) (fig. 20). The megameric M9 chromosome of and the X chromosomes. This includes the X of Truxalis nasuta (fig. 2) has an interstitial C-band Euchorthippus chopardi (fig. 4), Omocestus ray- and, finally, the megameric M6 chromosome of mondi (fig. 10) and 0. llorenteae (fig. 11), the L2 Euchorthippus pulvinatus (fig. 5) possesses an of Chorthippus nevadensis (fig. 17), the L3 of 0. apparently distal C-band which, in less condensed panteli (fig. 9) and the L2 and the L3 of Ch.jucundus chromosomes, is clearly interstitial (see figs. 3 and (fig. 15), Ch. vagans (fig. 19) and Ch. brunneus 4 in Santos and Giráldez, 1982). 368 J. CABRERO AND J. P. M. CAMACHO 'C 'C I,) r I;. C — p S I S 0 a a St -4 S S 0 9 3 a - 4: -t 0 4 1.

Figures6—16 C-banding pattern in the haploid chromosome complements of gomphocerine grasshoppers. (6) Stenobothrusfestivus, (7) S. grammicus, (8) Omocestus bolivari, (9) 0. panteli, (10) 0. raymondi. CYTOGENETIC STUDIES IN GOMPHOCERINE GRASSHOPPERS. I 369 r r '4 "I IC -F r • • • a a tie -& S p a S S 4 • jS n. - - • • S -s a 'I), art

Figures 11—15C-banding pattern in the haploid chromosome complements of gomphocerine grasshoppers. (11) Omocestus llorenteae, (12)Stauroderusscalaris, (13) Chorthippus parallelus, (14) Ch. dorsatus, (15) Ch. jucundus. 370 J. CABRERO AND J. P. M. CAMACI-10 r 1 1 p__ 3' 'S S a —. - p. a a -J S e a a 'a t 0 Sb • - .t S a e a ect fl -Se a, jfr9

Figures16—20 C-banding pattern in the haploid chromosome complements of gomphocerine grasshoppers. (16) Chorthippus apicalis, (17) Ch.nevadensis,(18) Ch. binotatus, (19) Ch. vagans, (20) Ch. brunneus. CYTOGENETIC STUDIES IN GOMPHOCERINE GRASSHOPPERS. I 371

Distal C-bands limited by the fact that one cannot be sure that chromosomes of similar relative length are Theseare as infrequent as the interstistial C-bands and, like them, appear at most in two chromosome necessarily homologous in all genomes (King and John, 1980). The only possible exceptions are the pairs within the same species. In the majority of X and the megameric chromosomes, two elements cases the distal C-bands are small in size. This is which, presumably, each have a common origin so in the M7 chromosomes of Chorthippus brunneus within the Acrididae. However, in spite of the (fig. 20), the M6 and M7 of Ch. nevadensis (fig. heteropycnosis of both chromosomes during male 17), the M4 of Ch. apicalis (fig. 16), and the X of meiosis, their responses to C-banding are not Omocestus bolivari (fig. 8). In other species, distal especially distinctive. The only recognisable mega- C-bands are medium or large, as is the case in the meric chromosomes which show an unusual C- L3 chromosome of Euchorthippus chopardi (fig. 4), banding pattern are the M9 chromosome of the L2 and L3 of Omocestus raymondi (fig. 10), the Truxalis nasuta (fig. 2) and the M6 of Euchorthippus M6 of Stenobothrus festivus (fig. 6), the M9 of pulvinatus (fig. 5), Stenobothrus festivus (fig. 6), Truxalis nasuta (fig. 2) and the C-band in the short Chorthippus apicalis (fig. 16) and Ch. nevadensis arm of the M4 chromosome of Chorthippus vagans (fig. 17). On the other hand, some X chromosomes (fig. 19). show a C-band located proximally as in Stenoboth- rusfestivus (fig. 6) and S. grammicus (fig. 7), inter- DISCUSSION stitially as in Euchorthippus chopardi (fig. 4), Omocestus raymondi (fig. 10) and 0. llorenteae (fig. TheC-banding patterns of the 20 gomphocerine 11) or distally as in 0. bolivari (fig. 8). 'While the species analysed in this paper, like those of other megameric chromosomes of these species do not acridoid grasshoppers (King and John, 1980; San- seem to have much in common, the X chromo- tos et a!., 1983), fit the equilocal model for hetero- somes show more similarities between the species chromatin distribution proposed by Heitz (1933; of the genera Stenobothrus, Euchorthippus and 1935). Thus the 'constitutive heterochromatin of Omocestus than between these and the species of non homologous chromosomes of a same diploid Chorthippus, which suggests a close relationship set tends to be located in similar sites". Without between these three genera. However, neither the doubt, paracentromeric C-bands provide the similarities nor the differences between these gen- strongest support for the equilocal model, given era are absolute, which implies that the evolution- that their presence is constant in all chromosomes ary relationship of these taxa is too complex to be of all species, and that their size is usually uniform deduced from C-banding patterns. In other words, for a majority of the chromosomes within a given C-banding patterns may change so quickly that species. Exceptionally, as in Dociostaurus genei they do not provide sensible evolutionary markers. and Truxalis nasuta, paracentromeric C-bands are The pronounced tendency for C-heterochromatin medium sized in some chromosomes but small in to vary in grasshoppers has been evidenced by others, while in Ramburiella hispanica they may many reports (John and King, 1977; 1982; 1983; be large or medium, and in Chorthippus apicalis Hewitt, 1979; Santos et al., 1983; Camacho et a!., there are large, medium and small paracentromeric 1984) but especially by those of Shaw et a!. (1976). C-bands. Recent studies of acridoid C-hetero- A further fact that merits comment is the origin chromatin by fluorochromes (John et a!., 1985) of the short arms of subtelocentric chromosomes have demonstrated that C-heterochromatic blocks in some species. While in Stauroderus scalaris (fig. which occupy similarpositions on non 12), Chorthippus binotatus (fig. 18) and Ch. brun- homologous chromosomes in a species do not neus (fig. 20) they appear deeply stained following necessarily coincide either in size or in composi- C-banding, in Ch. vagans (fig. 19), with the excep- tion. This heterochromatin heterogeneity, shown tion of the M4 chromosome, the short arms of the by differential fluorescence, implies that equilocal- M, S and X chromosomes do not C-band. Con- ity as defined by position does not apply invariably sequently, while darkly C-banded short arms to structure. Interspecific comparisons of C- would have arisen by amplification, those of Ch. banding patterns reveal differences between vagans have presumably arisen by pericentric species of either the same or of different genera inversion. These two types of chromosomal rear- (Santos et a!., 1983; this paper), which undoub- rangements have also been held to explain the tedly is also due to the dynamic nature of hetero- origin of different kinds of short arms in the chromatin (Morescaichi, 1977). It is important to chromosomes of different species of Australian recognise that interspecific comparisons are morabine grasshoppers (White, 1975). 372 J. CABRERO AND J. P. M. CAMACHO

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