International Journal of Cytology, Cytosystematics and Cytogenetics

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1 Caryologia International Journal of Cytology, Cytosystematics and Cytogenetics ISSN: (Print) (Online) Journal homepage: Chromosomal variability of Drosophila polymorpha populations from Atlantic Forest remnants of continental and insular environments in the State of Santa Catarina, Brazil Daniela Cristina De Toni, Fabiana De Oliveira Herédia & Vera Lúcia S. Valente To cite this article: Daniela Cristina De Toni, Fabiana De Oliveira Herédia & Vera Lúcia S. Valente (2001) Chromosomal variability of Drosophila polymorpha populations from Atlantic Forest remnants of continental and insular environments in the State of Santa Catarina, Brazil, Caryologia, 54:4, , DOI: / To link to this article: Published online: 28 Jan Submit your article to this journal Article views: 41 View related articles Full Terms & Conditions of access and use can be found at Download by: [ ] Date: 02 August 2016, At: 05:36

2 CARYOLOGIA Vol. 54, no. 4: , 2001 Chromosomal variability of Drosophila polymorpha populations from Atlantic Forest remnants of continental and insular environments in the State of Santa Catarina, Brazil DANIELA CRISTINA DE TONI, FABIANA DE OLIVEIRA HERÉDIA and VERA LÚCIA S. VALENTE* Departamento de Genética, Instituto de Biociências, Universidade Federal do Rio Grande do Sul. Caixa Postal Porto Alegre, RS, Brazil. INTRODUCTION The study of Atlantic Forest insect communities is fundamental for understanding biodiversity, both genetically and ecologically. It is also urgent, due to the rapid rate of destruction of this type of environment, of which only small portions of the original area remain in Brazil. Studies of Drosophila communities from the Atlantic Forest in Southern Brazil were done by VALENTE and ARAÚJO (1986a, b, 1991); SAAVEDRA et al. (1995a, b) who during four years studied both ecological and genetic characteristics of Drosophila populations from places having elements of this type of forest (Atlantic Forest, whether narrowly or * Corresponding author: fax , valente@ufrgs.br Abstract In the present study, we investigated chromosomal polymorphism for paracentric inversions found in Drosophila polymorpha populations common in five fly communities (one continental and four insular) collected in Atlantic Forest remnants in Santa Catarina, Brazil, during a one-year sampling. Our analyses showed that the more polymorphic the populations, the greater the abundance of flies. The insular populations were more polymorphic than the continental one, probably because the greater environmental heterogeneity found in the islands provided more niches for fly colonisation and was more suitable for supporting a community. Stochastic effects, however, need to be taken into account to understand the low polymorphism found at the continental site, since the D. polymorpha sample collected there was small. We also present here the occurrence of six previously undescribed paracentric chromosomal inversions in heterozygosis, increasing to 7 the number of variants identified in the D. polymorpha polytene chromosomes. Key words: Atlantic Forest; chromosomal polymorphism; continental and island populations; Drosophila polymorpha; inversions. broadely defined) in Rio Grande do Sul, the southernmost Brazilian state. In the state of Santa Catarina, a preliminary study was carried out (DE TONI and HOFMANN 1994) in order to identify, taxonomically and systematically, the Drosophila communities inhabiting some Atlantic Forest in the city of Florianópolis, on Santa Catarina Island. In that study, we observed that the D. willistoni subgroup was the most frequently found in such communities, followed by D. polymorpha. The latter species has received the attention of drosophilists because of its polymorphisms: pigmentation of abdominal tergites (DA CUNHA 1949; HEED and BLAKE 1963; MARTINEZ and CORDEIRO 1970) and chromosomal paracentric inversions (DA CUNHA et al. 1953; HEED and RUS- SELL 1971; ROHDE and VALENTE 1996a), both clearly adaptative. The occurrence of heterosis

3 330 DE TONI, HERÉDIA and VALENTE associated with the major esterase enzymatic locus in a natural Brazilian population of this species was found by NAPP and CORDEIRO (1978), who established interspecific relationships among cardini group species by means of protein electrophoresis (NAPP and CORDEIRO 1981). DA CUNHA et al. (1953) in studying chromosomal polymorphism of both D. polymorpha and D. cardinoides (D. cardini group), observed that the former is more polymorphic than the latter, presenting 6 different inversions in analysed samples. They also observed that D. polymorpha is relatively more common in Southeastern Brazil than the other D. cardini group species. ROHDE and VALENTE (1996a), studying urban and wild D. polymorpha and D. cardinoides populations, constructed polytene chromosomal photomaps and analysed qualitatively the chromosomal polymorphism of the two species, describing a new IIR chromosomal arm inversion. Despite its pervasiveness in the Atlantic Forest communities, however, D. polymorpha has been inadequately studied. Santa Catarina State still have several forested areas, conserved in varying degrees, and mainly on Atlantic islands. Furthermore, the location of D. polymorpha on both islands and the continent provides a good opportunity, which this report intends to exploit, to study the role of this type of genetic variant in environments threatened by extinction. MATERIAL AND METHODS Fig. 1 shows the locations where we performed sampling trips. They were: on Santa Catarina Island (27 o 42'S; 48 o 30'W), A) Canto da Lagoa (Atlantic Forest narrowly defined) which constitutes an Atlantic Forest with high relative humidity, four tree straits, and many epiphytes plants. The trees of the main straits have, in average, a diameter of 50 cm, at chestheight. This forest is defined as a secondary forest in advanced state of regeneration; D) Sertão do Peri (Atlantic Forest broadly defined) presenting patches of secondary forest very similar to the one of point A, but with trees of large diameter and economic interest, still not cut, in the more preserved points.; on two Atlantic islands (both Atlantic Forest, and varyingly conserved): B) Ratones Grande Island (27 o 29'30"S; 48 o 36'42"W), located between the island and the continent well preserved and with m 2, site of a conservation unit maintained by the Universidade Federal of Santa Catarina; C) Ratones Pequeno Island (27 o 29'31"S; 48 o 34'04"W), with 96.20m 2 covered by vegetation similar to that of Ratones Grande Island, but partially devastated by anthropic pressure; on the continent: E) State Park of Serra do Tabuleiro, the main protected forest area in Santa Catarina, with 87, m 2, and constituted by at least six different botanical compositions (Atlantic Forest strictly defined) This place is widely conserved, with great numbers of species of great size, economic interest, with immense amount of plant epiphytic plants, that demonstrate the wetness of this place. The humidity is maintained by the conservation of a large intact forest and by a River (Cubatão), that has its spring in this region of collections. Adult Drosophila population samplings were obtained over fermented banana bait. A minimum of three and a maximum of six days passed between visits to each collection site. Field work was performed every two months between July/1997 and June/1998. Isofemale lines of D. polymorpha were established immediately after collection. Chromosomal polymor- Fig. 1 Maps of Brazil and of Santa Catarina State (1), in which are indicated the area corresponding to the five sampling points. A) Canto da Lagoa; B) Ratones Grande Island; C) Ratones Pequeno Island; D) Sertão do Peri; E) Serra do Tabuleiro.

4 INVERSIONS IN DROSOPHILA POLYMORPHA 331 phism of each sample was analysed by observing polytene chromosomes of one female third instar larva per isofemale line. Slides with salivary glands were processed according to the classic technique of ASH- BURNER (1967) and chromosome configurations were photomicrographed under phase contrast. We used the reference photomap of D. polymorpha salivary chromosomes (ROHDE and VALENTE, 1996a) to establish break points of inversions detected in heterozygous state, and describe them for comparison with inversions detected in our samples. The ROFF and BENTZER (1989) test was used to compare chromosomal polymorphism data of D. polymorpha populations. Estimation of inversion frequency similarity between the different populations was made employing the Manhattan distance, represented by dendograms constructed using the UPGMA method. RESULTS AND DISCUSSION In Table 1 we present data on chromosomal polymorphism found in the five Drosophila polymorpha populations studied showing total num- Fig. 2 Photomap of the D. polymorpha (ROHDE and VALENTE 1996a) polytene chromosomes with representation of inversion break points found in the State of Santa Catarina populations. With the exception of IIRA, all remaining inversions (XA, XB, IIRB, IIRC, IIRD and IIIRA) are previously undescribed arrangements.

5 332 DE TONI, HERÉDIA and VALENTE Table 1 Frequencies and absolute numbers of paracentric heterozygous inversions found in Drosophila polymorpha populations sampled in Santa Catarina State. Inversion/ Chromosomal arm Populations Date N total XA XB IIRA IIRB IIRC IIRD IIIRA Het % N / Month/year N % N % N % N % N % N % N % female Aug/ A Oct/ N=111 Nov/ Jan/ Apr/ Total Average Sep/ B Feb/ N=66 May/ Total Average Sep/ C Nov/ N=304 Dec/ Feb/ May/ Total Average Sep/ D Oct/ N=41 Jan/ Total Average Jan/ E Mar/ N=15 Total Average Inv/chroms Legend: A: Canto da Lagoa; B: Ratones Grande Island; C: Ratones Pequeno Island; D: Sertão do Peri; E: Serra do Tabuleiro; N: Total of analyzed isofemales; N / female: mean number of inversions/ female; Het. % : percentage of heterogygotes and Inv/chroms : mean number of inversions per chromosome.

6 INVERSIONS IN DROSOPHILA POLYMORPHA 333 ber of homozygous, total number of heterozygous for each type of inversion detected in the chromosomes of each population and mean number of inversions per female, per chromosomal arm. The most polymorphic chromosomal arm observed was the right arm of the second pair (IIR), followed by the X and the right arm of the third chromosome (IIIR). Noticeably in this Table, the island populations were generally more polymorphic than the continental ones (site E), an unexpected result since findings of several other authors who studied chromosomal polymorphism in insular and continental populations of other Drosophila species have obtained consistently opposite results (DOBZHANSKY 1957; TOWNSEND 1958; AYALA et al. 1971). Certain particularities of our samplings and especially of their environments, however, need to be considered to interpret our data properly. For instance, the number of D. polymorpha specimens captured in Serra do Tabuleiro (continental site E was always fewer than those obtained at the other). Thus, stochastic factors operating in small populations could explain the poverty of chromosomal arrangements (only the IIRA inversion segregates) observed in this population. Great differences exist in the number of isofemale lines established from flies collected at all 5 places: whereas the sample from Ratones Pequeno Island (C) was constituted by 304 isofemale lines, the E sample was made up of only 15 isofemale lines. According to the Island Biogeography Theory (MAC ARTHUR and WILSON 1967, see also HEED 1962), we can consider the area sampled in Serra do Tabuleiro as a small island inside a greater area, in which D. polymorpha was inadequately sampled. In fact, the more polymorphic population samples came from places where the species was more prevalent (see Table 1). Thus, it appears that under more favourable environmental conditions, chromosomal polymorphism tends to increase, as previously observed by DA CUNHA and DOBZHANSKY (1954); DA CUNHA et al. (1950, 1959) and reviewed by BRUSSARD (1984) for other species. Fig. 2 shows the photomap of larval salivary gland polytene chromosomes of D. polymorpha, established by ROHDE and VALENTE (1996a), in which we plotted the apparent new inversion limits detected in our samples. These inversions are: XA, with break points in the proximal region of section 17 and in the medium part of 14; XB, involving sections 15 to 20, with break points in the sub-proximal region of section 20, and in the distal part of 15; IIRA, from the middle of section 48 to the distal portion of 54; IIRB, with break points in the proximal region of section 57 and in the distal portion of 54; inversion IIRC, involving the distal portion of section 47 through the proximal region of section 48; IIRD, with break points in the distal part of 45, and in the middle region of 47 sections, and IIIA, which has break points in the sub-distal part of section 86, and in the middle part of section 83. Heterozygous aspect for each of these inversions is shown in the photomicrographs presented in Fig. 3. Statistical analysis of the comparisons performed using the ROFF and BENTZER (1989) Test among the population sampled, with respect to inversion frequencies, showed that the D. polymorpha populations from A, B, C, and D (insular) do not differ from those of (continental) E. However, insular populations A and D (both from Santa Catarina Island) differ statistically, as do those from the two Ratones Islands (B and C). Comparing the samples obtained on Santa Catarina Island with those of the Ratones, we observed that only the population of Ratones Pequeno (C) resembles that of Sertão do Peri (D), on Santa Catarina Island. All other population comparisons revealed different chromosomal constitutions. According to the model proposed by HEED and RUSSELL (1971), a D. cardinoides strain was a probable remnant of the ancestor from which the remaining species of the cardini group originated, making it older than D. polymorpha. A greater number of inversions are therefore expected in D. cardinoides, and have, in fact, been found in populations of the two species recently studied at urban sites in Rio Grande do Sul State by ROHDE and VALENTE (1996a, b). In the primitive Atlantic Forest environment, however, D. cardinoides is much less abundant than D. polymorpha, and the adjustment of this last one to such environments is perhaps reflected, as happened in our collections, in a high number of inversion types (6), close to that observed for the older species (7) in the city environment by ROHDE and VALENTE (1996a). DA CUNHA et al. (1953), analysing the chromosomal polymorphism of these two species, observed that D. polymorpha presented a higher degree of polymorphism than did D. cardinoides. They mention the occurrence of 3 inversions in D. cardinoides and 6 inversions (one in the sec-

7 DE TONI, HERÉDIA and VALENTE 334 Fig. 3 Heterozygous inversions found in polytene chromosomes of sampled populations of D. polymorpha in Santa Catarina State: XA; XB; IIRA; IIRB; IIRC; IIRA+C; IIRD; IIIRA. Bars = 10 µm.

8 INVERSIONS IN DROSOPHILA POLYMORPHA 335 ond, two in the third and 3 in the X chromosomes), in the same D. polymorpha chromosomes in which we detected polymorphism. These authors, however, do not describe either the chromosomal arms or break points of the inversions observed, rendering comparisons with our data difficult. It is therefore possible that some inversions, here reported as new, had already been observed by those authors. DA CUNHA et al. (1953) proposed that the higher chromosomal polymorphism level in the D. polymorpha populations collected by them in wild environments, could be considered a consequence of their pervasiveness in Southern Brazil. Our data points to the same conclusion. Furthermore, our observations about fly community dynamics in the places studied (data not shown), showed that the most polymorphic D. polymorpha population sampled (Ratones Pequeno) expanded in September, 1997, then strongly contracted. This could mean either that the original population was more polymorphic than we could detect, or that we detected a phenomenon in the field similar to that observed by CARSON and WISOTZKEY (1989) in laboratory populations. These authors observed increased chromosomal variability after the occurrence of a D. silvestris population bottleneck. HEED and RUSSELL (1971), also working with chromosomal polymorphism in the cardini group of Drosophila, observed inversions only in chromosomal arms IIL and X, and concluded that the latter chromosome is the most polymorphic in all species of the D. cardini group. Our data do not support this assumption, since the IIR chromosomal arm presented the highest number of inversions. POWELL et al. (1972) suggested the use of D. pseudoobscura chromosomal polymorphism as a criterion for determining the geographical origin of populations. Doing so, in our study we observed that the small continental population did not present any inversion type different from that of IIRA found by ROHDE and VALENTE (1996a) on the mainland of neighbouring Rio Grande do Sul State, whereas the insular populations presented several other inversions, in the same and other chromosomal arms. Perhaps the greater number of ecological opportunities offered by the island environments contribute to the high genetic variability expressed in our samples as higher D. polymorpha polymorphism. Although the statistical tests indicated homogeneity among both continental and island samples, we believe that the very limited continental area sampled distorted this result, since in that population only one type of inversion was detected, as compared with the great variability found in the others. The similarity found between the samples from Ratones Pequeno (C) and Sertão do Peri, on Santa Catarina Island (D) remains obscure, since they correspond to very different types of environments and the distance separating them is sufficient to hamper population dispersion. When we analysed the dendogram of similarities in inversion frequencies established among all the populations (Fig. 4), we observed that the Ratones Pequeno (C) population is most unlike the others, probably due to the higher chromosomal variant and frequency numbers. This finding supports the classic assumption that greater environmental heterogeneity promotes higher levels of chromosomal polymorphism (DA CUNHA et al. 1950, 1959; DA CUNHA and DOBZHANSKY 1954). All other comparisons made between samplings from Santa Catarina Island, that either dif- Fig. 4 Similarities among the D. polymorpha populations of the State of Santa Catarina shown by chromosomal polymorphism data. Clusters were formed employing Manhattan Distance as a dissimilarity coefficient for UPGMA (unweighted pairgroup method using arithmetic averages) clustering algorithm (SNEATH and SOKAL 1973). A) Canto da Lagoa; B) Ratones Grande Island; C) Ratones Pequeno Island; D) Sertão do Peri; E) Serra do Tabuleiro.

9 336 DE TONI, HERÉDIA and VALENTE fered in environment (A x D) or were similar in flora and climate types (B x A) produced statistically different results. Finally, it should be mentioned that the most polymorphic sample of D. polymorpha in Ratones Pequeno (C), where the species apparently has more ecological opportunities, was taken on September, At that time, climatic factors appeared to be promoting population growth. Different roles of climatic or intrinsic factors affecting separate populations of the same species were reported by VALENTE and ARAÚJO (1986a) for D. willistoni. These authors, working with native populations subject to lesser or greater climatic fluctuations, suggested that unstable climates could diminish the effect of intrinsic factors (such as chromosomal polymorphism) on the regulation of population size. This observation should be considered here. Acknowledgements This research was supported by CAPES (DCDT), and CNPq (FOH and VLSV) fellowships and grants from CNPq, FAPERGS, FINEP, and PROPESQ-UFRGS. Thanks are due to Dr. Paulo R.P.Hofmann and all staff of Universidade Federal de Santa Catarina for the facilities offered for performing field work in the islands and processing of the collected material, to the Ph.D. student Jennifer Brisson of Department of Biology Ian Duncan- St. Louis-MO, for the revision of our English version for this paper, as well as to Dr. Carlos R. Vilela (Universidade de São Paulo) for help in identifying several species of Drosophila collected, and to Miss Nena B. Morale for technical assistance. REFERENCES ASHBURNER M., 1967 Patterns of puffing activity in the salivary gland chromosomes of Drosophila. I. Autosomal puffing patterns in a laboratory stock of Drosophila melanogaster. Chromosoma, 27: AYALA F. J., POWELL J.R. and DOBZHANSKY T., 1971 Polymorphism in continental and island populations of Drosophila willistoni. Proceedings of Natural Academy of Science USA, 68: BRUSSARD P.F., 1984 Geographical patterns and environmental gradients: the central- marginal model in Drosophila revisited. Ann. Rev. Ecol. Syst., 15: CARSON H.L. and WISOTZKEY R.G., 1989 Increase in genetic variance following a population bottleneck. American Naturalist, 134: DA CUNHA A.B., 1949 Genetic analysis of the polymorphism of colour pattern in Drosophila polymorpha. Evolution, 3: DA CUNHA A.B. and DOBZHANSKY T., 1954 A further study of chromosomal polymorphism in Drosophila willistoni in its relation to environment. Evolution, 8: DA CUNHA A.B., BRNCIC D. and SALZANO F.M., 1953 A comparative study of chromosomal polymorphism in certain South American species of Drosophila. Heredity, 7: DA CUNHA A.B., BURLA H. and DOBZHANSKY T., 1950 Adaptive chromosomal polymorphism in Drosophila willistoni. Evolution, 4: DA CUNHA A.B., DOBZHANSKY T., PAVLOVSKY O. and SPASSKY B., 1959 Genetics of natural populations. XXXVIII. Supplementary data on the chromosomal polymorphism in its relations to the environment. Evolution, 13: DE TONI D.C. and HOFMANN P.R.P., 1994 Preliminary taxonomic survey of the genus Drosophila (Diptera: Drosophilidae) at Morro da Lagoa da Conceição; Santa Catarina island, Brazil. Revista Brasileira de Biologia, 55: DOBZHANSKY T., 1957 Genetics of natural populations. XXVI. Chromosomal variability in island and continental populations of Drosophila willistoni from Central America and the West Indies. Evolution, 11: HEED W.B., 1962 Genetic characteristics of island populations. University of Texas Publications, 6205: HEED W.B. and BLAKE P., 1963 A new colour allele at the E locus of Drosophila polymorpha from Northern South America. Genetics, 48: HEED W.B. and RUSSELL J.S., 1971 Phylogeny and population structure in island and continental species of the cardini group of Drosophila. Studies by inversion analysis. University of Texas Publications, 7103: MAC ARTHUR R.H. and WILSON E.O., 1967 The Theory of Island Biogeography. Princeton University Press. Princeton, N. Jersey. MARTINEZ M.N. and CORDEIRO A.R., 1970 Modifiers of colour pattern genes in Drosophila polymorpha. Genetics, 64: NAPP M. and CORDEIRO A.R., 1978 Heterosis in a wild strain of Drosophila polymorpha with a lethal closely linked to the major esterase locus. Biochemical. Genetics, 16: , 1981 Interspecific relationship in the cardini group of Drosophila studied by electrophoresis. Braz. J. Genet., 4: POWELL J.F., LEVENE H. and DOBZHANSKY T., 1972 Chromosomal polymorphism in Drosophila pseudoobscura used for diagnosis of geographic origin. Evolution, 26:

10 INVERSIONS IN DROSOPHILA POLYMORPHA 337 ROFF D.A. and BENTZER P., 1989 The statistical analysis of mitochondrial DNA polymorphisms: c2 and the problem of small samples. Molecular Biology and Evolution, 6: ROHDE C. and VALENTE V.L.S., 1996a Cytological maps and chromosomal polymorphism of Drosophila polymorpha and Drosophila cardinoides. Braz. J. Genet., 19: , 1996b Ecological characteristics of urban populations of Drosophila polymorpha Dobzhansky & Pavan and Drosophila cardinoides Dobzhansky & Pavan (Diptera, Drosophilidae). Rev. Bras. Entomol., 40: SAAVEDRA C.C.R., VALENTE V.L.S. and NAPP M., 1995a An ecological and genetic approach to the study of enzymatic polymorphisms in Drosophila maculifrons. Rev. Bras. Genét., 18: SAAVEDRA C.C.R., CALLEGARI-JACQUES S.M., NAPP M. and VALENTE V.L.S., 1995b A descriptive and analytical study of four Neotropical Drosophilid communities. J. Zool. Syst. Evol. Res., 33: SNEATH P.H. and SOKAL R.R., 1973 Numerical Taxonomy. Freeman, San Francisco. TOWNSEND J.I., 1958 Chromosomal polymorphism in Caribbean island populations of Drosophila willistoni. PNAS USA, 44: VALENTE V.L.S. and ARAÚJO A.M., 1986a Chromosomal polymorphism, climatic factors, and variation in population size of Drosophila willistoni in Southern Brazil. Heredity, 57: , 1986b Comments on breeding sites of Drosophila willistoni Sturtevant (Diptera, Drosophilidae). Rev. Bras. Entomol., 30: , 1991 Ecological aspects of Drosophila species inhabiting wild environments in Southern Brazil (Diptera: Drosophilidae). Rev. Bras. Entomol., 35: Received June 18, 2001; accepted July 5, 2001