Intra-Populational and Intra-Individual Mosaicisms of Uranoscopus Scaber 1

l-leradity6l {1991) 325—330 Received llJanuar, 7991 OThe Genetical Society of Great Britain

Intra-populational and intra-individual mosaicisms of Uranoscopus scaber 1. (Perciformes, Uranoscopidae)

R. VITTURI, F. CATALANO, M. R. 10 CONTE, A. M. ALESS1, F. P. AMICO & D. C0LOMBERA* Institute of Zoology, University of Pa/errno, V/a Archirafi 18, 90723 Palermo and *oepa,.tment of Biology, University of Padue, Via Trieste 75, 35127 Padue, Italy

Karyotypicanalysis on 25 specimens of Uranoscopus scaber from the Gulf of Palermo has revealed the occurrence of an intra-populational mosaicism of Robertsonian type with three distinct diploid numbers: 2n =30,2n 29 and 2n =28.Intra-individual mosaicism in the chromosome number (2n =30 and 31 in two specimens and 2n= 28 and 29 in one specimen) is also reported. One large metacentric pair is characterized by secondary constrictions, variable in length in different fish. Polymorphisms of these areas seem to have a genetic, rather than transcriptional, basis.

Keywords:karyology,mosaicism, pisces, Uranoscopus scaber.

organizer regions (NORs) were characterized following Introduction the method of Howell & Black (1980); hetero- Althoughmore than 200 species of teleostean Perci- chromatic regions were revealed using the C-banding formes have been analysed cytogenetically (Sola a at, method of Sumner (1972); G-banding was performed 1981), many families of this order remain completely according to Seabright (1971). uncharacteri.zed in this respect. One of these is the Observations and photomicrographs were made family Uranoseopidae, within which the genus using a Jenamed 2 light microscope. Chromosomes Uranoscopus is represented by only one species in the were classified on the basis of the arm-length ratio Mediterranean (Whitehead eta!., 1986). (Levan eta!., 1964). Thispaper analyses the chromosome constitutions of Uranoscopus scaber from the Gulf of Palermo and Results describes both intra-popalational and intra-individual mosaicism in the chromosome numbers as well as Ofthe 64 specimens analysed in this study, only 25 structural polymorphism involving nucleolus organizer produced preparations suitable for chromosome regions (NORs). analysis. Counts of 20 spreads per specimen gave three distinct karyomorphs: with 2n 30 (three females, two males and nine sexually immature), 2n =29 (one Materialsand methods female, one male and three sexually immature) and Sixty-fourspecimens of Uranoscopus scaber caught by 2n =28 (two females, two males and two sexually seine in the Gulf of Palermo and transported live to the immature). These karyomorphs, designated as A-, B- Tnstitute of Zoology, Palermo, Italy, were examined. All and C-type complements, had arm numbers (FN) of individuals were classified according. to Whitehead a 56, 54 and 52, respectively, at. (1986) and sexed by examination of the gonads. The A-type complement (Fig. 1 a) could be arranged Five specimens are now preserved in the Museum of in 15 pairs. The first nine were metacentrics; pairs 10 the Institute of Zoology, University of Palermo. and 11 were submetacentrics, 12 and 13 were meta- Chromosome preparations were obtained by the centrics and pairs 14 and 15 were telocentrics. Homo- standard air-drying technique applied to kidney and logous chromosomes of pair 9 were characterized by spleen tissues. Slides were stained with 5 per cent secondary constrictions which differed in size. Two Giemsa pH 6.8, for conventional analysis. Nucleolus specimens, both possessing the modal diploid number 325 326 R. VITTURI ETAL. M a — == as -w pp —t -S -S -' — a S-s a' a 0 NJ 11 'C at * 'V 4 a S * St

Fig.1 (a) Giemsa non-differentially stained A-type. (b) Aneuploid spread (2n 31)with chromosomes arranged in groups according to morphology and size. 2n29 ('g) flh21IuU KUnx UA

Al(2) ______(a) (b)

Fig. 2 (a) Mitotic metaphase spread with a modal diploid number of 29 and (b) chromosomes arranged in groups according to morphology and size (B-type). of 30 chromosomes, had three and four (out of 20) chromosomes, displayed three (out of 20) spreads with cells respectively with 2n 31 (Fig. 2b); in both indivi- 2n =29(Fig. 3b); in this case an extra chromosome duals an extra chromosome of the group of the belonged to the large metacentric group. medium-sized metacentrics (pairs 12 and 13) was In silver-stained chromosome preparations, secon- observed. dary constrictions of pair 9 were consistently involved The B-type (Fig. 2) consisted of 19 large meta- in active nucleolus organization (Fig. 4a) and a correla- centrics, two medium submetacentrics, four medium tion in dimensions between the NORs and these areas metacentrics and four telocentrics. was observed (Fig. 4b). Secondary constrictions of The C-type complement (Fig. 3a) differed from the chromosomes 9 appeared to be large in size in both A-type with respect to the presence of two large bi- homologues of six fish (Fig. 4a, sec i), small in both armed chromosomes derived from pairs 10 and 11. homologues of four fish (Fig. 4b, see iii) and of differ- Homologous chromosomes of pair 9 had secondary ent dimensions in 15 fish (Fig. 4c, see ii); in addition constrictions which were both large in size. In addition, they showed a uniform length in all spreads of each one individual, with the modal diploid number of 28 individual. (Fig. Saandb,seearrows). WhentheG-banding arranged ingroupsaccordingtomorphologyandsize Fig. Of 15karyotypedspreads, 12 sornes, particularlyinteresting resultswereobserved. which possessedcellswith both28and29chromo- procedure wasperformed onthesinglespecimen, constrictions ofpair9 all chromosomepairs(Fig.Snandb);alsosecondary in staining,wereobservedthecentromericregionsof in size.(b)Pair9ofdifferentfish:g= Fig. 4 diploid number of28chromosomes,while threehad After G-banding,largezones,whichappearedlight 3 (a)Gientsanon-differentiallystainedC-type.(b)0-bandedaneuploidmetaphasewith2n=

(a) S.- • Silver-stained chromosomeswiththeNORs:(i)largeinsize; (ii)differentinsize,onelargeandsmall;(iii)small as -c as

were aS — t C- lightly stainedthroughout 4 as c) giemsa S.______showed

stained andn= 3 (0

the modal a air, S a tI_ S. NOR a -s pa a __ "I

stained. 0 9, C-types (Fig.3dande).Secondary constrictionsofpair regions ofallchromosome pairsofbothA-and sions wereobservedand located inthecentromeric chromatic bandsofapproximately thesamedimen- Sc). correlation intheir0-bandingpatterns(Figs3band showed averylargesecondaryconstrictionandgood 29 chromosomes(Fig.3h).Inthelatter,threeelements we which From C-bandinganalysis, conspicuoushetero- s. C-tm a -' appeared moderately C-positive,weredistin- a S MOSAICISMS OFU.3C48ER321

at; 29; NJ chromosomes are N) a $ :3 II Ni NJ 328 R. VITTLJRI ETAL. 2n30 S = 28 9e I. It S 'S — Al', IEg • 5 to

45,S S 'a. r b,'1, .6 CM (b) (c) (a) ______

2n30 2n=28

S I S 1% S 0, S a

Fig. 5(a) 0-banded spread with 2n =30.(b) 0-banded spread with 2n 28. (c) Partial 0-banded aneuploid spread (2n 29) (arrows indicate three elements each with a large secondary constriction). (d) C-banded spread with 2n =30.{e) C-banded spread with 2,i =28. guishable from the corresponding centromeric derived from the fusion of one homologue of pair 10 C-bands which were more darkly stained (Fig. 3d and with one of pair 11. e, see arrows). Based on the consideration that, in U. scalier, the Roberisonian fusion between two bi-armed chromo- Discussion somes gives rise to a morphologically non-differentiated metacentric, it is clear that the short arms of the Karyotypicvariability in the chromosome number original chromosomes involved in the translocation ranging from Zn =28 to 2n =30reveals the occur- have been lost. As abnormal specimens were not rence of an inter-individual mosaicism of Robertsonian encountered in the population examined, we are origin in Uranoscopus scalier from the Gulf of Palermo. inclined to accept the hypothesis of Bertollo a a!. This mosaicism is clearly due to the fusion of one (1979), that the missing chromosomal portions element of pair 10 with one element of pair 11, B- and possibly contain gene loci not responsible for functions C-type complements being the heterozygous and of vital importance. homozygous conditions for the fused elements respec- Forty-eight mono-armed chromosomes are con- tively. sidered to be the ancestral karyotype in fish (Ohno, Hsu et a!. (1975) proposed that such a fusion could 1974). This may suggest that the A-type is primitive in produce a monocentric or dicentric bi-armed chromo- the population under study. Moreover, the presence of some. However, the latter hypothesis must be the fundamental numbers 56, 54 and 52, which are discarded due to the absence, in the fused metacen- close to the ancestral number of 48, and the presence trics, of a heterochromatin size corresponding to that of a high number of large metacentric chromosomes in MOSAICISMS OF U. SCA HER 329 the actual U. .cca her specimens, seems to support the Secondary constrictions of pair 9 stain like most of derivation of the A-type complement through the homogeneously staining regions (HSRs) (Trent ci numerous Robertsonian fusions occurring during the at, 1984), i.e. light after non-differentially staining and evolution of this species. G-banding, moderately stained with C-banding and Another type of variation was observed in three darkly stained with silver nitrate when composed of specimens; however, because a comparison of the total ribosomal DNA. It is widely held that HSRs are cyto- number of cells analysed per specimen reveals that it logical manifestations of DNA sequence amplification was confined to only a few spreads, we have classified and that they are characterized by displaying extreme it as a low level intra-individual mosaicism. In particu- fragility (Cowell, 1982). If this is true for the secondary lar, when the 0-banding procedure was conducted on a constrictions of pair 9, wide heteromorphism of these single mosaic specimen with 2n =28and 29, it was areas would find a convincing explanation. observed that all three mutant cells encountered showed three elements with large secondary constrictions. This has allowed us to conclude that, in this case, Acknowledgements trisomy involving pair 9 could be responsible for the Thisresearch was supported by a 60 per cent MPI intra-individual numerical variability. In contrast, as grant during 1988—1989. both mosaic specimens with 2n =30 and 31 mutant cells contain an extra chromosome of the group of the References medium-sized metacentrics (Fig. ib), trisomy involving pairs 12 or 13 could have occurred. BEIITOLLO,L A. C., TAXAHASHI, c. 5. AND MORE1RA FILHO, o. 1979. Note that intra-individual mosaicism has been Karyotypic studies of the genus Hop!ias (Pisces, reported only in the genus Salvelinus (Davisson ci at, Erythrinidae). Rev. Brash. Genet. ii. 1, 17—37. 1973; Hartley & Borne, 19Mb) and in Gobiusfallax BLACK, D. A. AND HOWELL, w. M. 1978. A distinctive chromo- (Thode a at, 1988), while intra-populational somal race of the cyprinodontid fish, Fundulus notatus, mosaicisms in the form of Robertsonian translocations from the upper Tombigbee river system of Alabama and are widespread in teleostean fish. The latter were Mississippi. Copela, 2, 280—288. CHEN, r It 1971. A comparative chromosome study of twenty described in species of the same genus (Chen, 1971), in kihifish species of the genus Fundulus (Teleostei, different populations of the same species (Black & Cyprinodontidae). Chromosoma, 32, 436—455. Howell, 1978; Ojima & Kashiwagi, 1981) and in COWELL, J. a. 1982. Double minutes and homogeneously specimens of the same population (Ojima & Kashiwagi, staining regions: Gene amplification in mammalian cells. 1981; Hartley & Home, 1984a and b; Vitturi ci at, Ann. Rev. Genee., 16, 21—59. 1984; Thode ci at, 1985; Vitturi & Catalano, 1988; DAWS5ON, M. 'C, WRIGHT. J. E. AND ATHERTON, L. M. 1973. Cyto- Vitturi, 1988). genetic analysis of pseudolinkage of LDH loci in the Chromosomal variability involving secondary con- teleost genus Salvelinus. Genetics, '13, 645—658. strictions of pair 9 was also observed among different FORESfl, F., ALMEIDA TOLEDO, L. F. AND TOLEDO, 5. A. 1981. Poly- specimens. Inter-individual variability can be attributed morphic nature of nucleolus organizer regions in fishes. to a varying amount of rDNA genes. This conclusion is Cytogenet. Ccli Genet., 31, 137—144. GARCIA, E., ALVAREZ. Sf. C. ANDTHODE,G. 1987. Chromosome supported by at least two arguments. The first is that relationships in the genus Blennius (Blennidae, Perci- secondary constrictions vary in length from one speci- formes) C-banding patterns suggest two karyoevolutional men to another but are uniform within the same indivi- pathways. Genetica, 72,27—36. dual; the second is that these areas are variable in GOLD, .R.1984. Silver-staining and heteromorphism of length with all techniques employed, thus indicating chromosomal nucleolus organizer regions in North that a change in the ribosomal genes has occurred. American cyprinid fishes. Copeia, 1,133—139. Variability in the size of active NORs regularly HARTLEY, 5. E. AND BORNE, M. T. 1984a. Chromosome poly- occurs in teleostean fish and it is mainly explained as a morphism and constitutive heterochromatin in Atlantic functionally differentiated activity of RNA genes salmon, Salino salar. Chromosoina, 89, 277—380. (Gold, 1984; Garcia ci at, 1987; Lopez ci at, 1989). HARTLEY, S. E. AND HORNE, M. ii 1984b. Chromosome relation- On the other hand in some species of the order ships in the genus Salmo. Chromosotna, 90, 229—237. HOWELL, W. M. AND BLACK, D. A. 1980. Controlled silver staining Gimnotiformes (Foresti ci a!., 1981), in Dice nira rchus of nucleolus organizer regions with a protective colloidal !abrax and D. punctaius (Prciformes) (Vitturi ci a!., developer: a 1-step method. Experientia, 36, 1014—1015. 1990) and in But/ms podas (Pleuronectiformes) H5U, t c., PATHAK, 5. AND CHEN, 'C R. 1975. The possibility of (Vitturi & Catalano, 1991), it has been suggested that latent eentromeres and a proposed nomenclature system NOR polymorphism could have a genetic rather than for total chromosome and whole arm translocations. transcriptional basis. Cytogenet. Cell Genet., 15, 4 1—49. 330 P. VITTURI ETAL.

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