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Cytogenetics of a New Species of Paratelmatobius Cardosoi Group (Anura: Leptodactylidae), with the Description of an Apparent Ca

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Cytogenetics of a New Species of Paratelmatobius Cardosoi Group (Anura: Leptodactylidae), with the Description of an Apparent Ca Cytogenetics of anew speciesof Paratelmatobius cardosoi group (Anura: Leptodactylidae), with thedescription of an apparent caseof pericentricin version LucianaB. Lourenço 1,PauloC.A. Garcia 2,ShirleiM. Recco-Pimentel 3 1;3Departamento deBiologia Celular, Instituto de Biologia, Universidade Estadual de Campinas (UNICAMP), 13084-971,Campinas, SP ,Brasil correspondingauthor: e-mail: [email protected] e-mail: [email protected] 2 Departamento deZoologiae Cursode Pó s-Graduaç ão, Institutode Biociê ncias, Universidade Estadual Paulista (UNESP),13506-900, Rio Claro, SP ,Brasil e-mail: [email protected] Abstract. Thekaryotype of a new species of Paratelmatobius from the P. cardosoi groupis described.As with other Paratelmatobius and Scythrophrys karyotypes, Paratelmatobius sp. (aff. cardosoi)showsa diploid numberof 24chromosomes, in addition to other similarities withthe former karyotypes.The Paratelmatobius sp. (aff. cardosoi)karyotypediffers from that of P. cardosoi inthe morphology of pair 4, the NOR location andthe C-bands in pairs 3 and8 (exclusiveto Paratelmatobius sp.)and those of pairs 7 and9 (exclusiveto P. cardosoi).Bothkaryotypes also differ in the amount of heterochromatinin pair 1. Thepresence ofinterstitial heterochromatinin the long arm ofpair1 andthe interstitial C-bands in both arms ofchromosome5 are apparently synapomorphiccharacters of P. cardosoi and Paratelmatobius sp. (aff. cardosoi),since theyare absentin the other Paratelmatobius and Scythrophrys karyotypes.In Paratelmatobius sp. (aff. cardosoi),thenucleolus organizer regionis onthe short arm ofa small metacentric chromosome(pair 9), an arrangement similar tothe NOR- bearingchromosome pair in the karyotype of P.poecilogaster andin karyotype II of Scythrophrys .Aconspicuous heteromorphismunrelated to the sex determiningmechanism was alsoobserved and probably arose froma pericentricinversion. Introduction The genus Paratelmatobius Lutzand Carvalho comprises vespecies placed in two groups(Pombal Jr .andHaddad, 1999). The P. cardosoi groupincludes P. cardosoi and P.mantiqueira while the P. lutzii groupcontains P. lutzii, P.poecilogaster ,andprobably P. gaigeae. All Paratelmatobius speciesare endemicto Braziland P.mantiqueira , P. lutzii and P. gaigeae were consideredas missingspecies by thoseauthors. © KoninklijkeBrill NV ,Leiden,2003 Amphibia-Reptilia24: 47-55 Alsoavailable online - www.brill.nl 48 LucianaB. Lourenço, PauloC.A. Garcia, ShirleiM. Recco-Pimentel Comparativecytogenetic analyses of livingspecies of Paratelmatobius (Lourenço etal., 2000), P. lutzii (De Luccaet al., 1974) and genus Scythrophrys (Lourenço etal.,in press) haveshown several similarities between these genera and have allowed the evolutionary studyof somechromosomal characters in this group. Inthe present paper, we describethe karyotype of anewspecies of Paratelmatobius in the P. cardosoi groupand reassert the evolutionary hypothesis proposed in our previous studies of Paratelmatobius and Scythrophrys (Lourenço etal., 2000, in press). An inter- estingcase of an autossomal inversion is also described. This new species, here called Paratelmatobius sp. (aff. cardosoi),was foundin theSerra doMarin the State of Paraná, Brazil,and differs from P. cardosoi byitsadvertisement call. Materialsand methods Sixmales andtwo females of Paratelmatobius sp. (aff. cardosoi)collectedfrom Piraquara, Paraná , Brazil, in October1999 were studiedcytogenetically. Chromosome preparations were obtainedfrom intestinal and testes cell suspensions,as describedby Schmid (1978) and Schmid et al.(1979). Conventional staining with a 10% Giemsa solution,the C-banding technique (King, 1980) and the Ag-NOR method(Howell and Black, 1980) were done.Chromosomes were classied according to Green andSessions (1991). All the specimens were deposited inCélio F.B. Haddad collection (CFBH), Departamento deZoologia,Universidade Estadual Paulista, Rio Claro, SP, Brasil. Results Thediploid complement of Paratelmatobius sp. (aff. cardosoi)consistsof 24 chromo- somes,with six pairs of metacentricchromosomes and sixof submetacentricchromosomes (gs 1-3,5; table1). In all the specimens analyzed, a conspicuoussecondary constriction was observedin the short arm ofhomologue9 inGiemsa-stainedmetaphases ( g. 1)and was identied asan NOR sitein silver-stainedpreparations ( g. 2).Additionally, a smaller secondaryconstriction was observedinterstitially in the long arm ofchromosome 5 in moredecondensed metaphases ( g. 1a). All the centromeric regions of the Paratelmato- bius sp. (aff. cardosoi)karyotypeshowed a smallamount of constitutiveheterochromatin (g. 3).Non-centromeric heterochromatic bands were easilyvisualized in pairs1, 3,5,and adjacentto the NOR inpair9 (g. 3).In some metaphases, a smalltelomeric C-band was observedin theshort arm ofpair8 (g. 3).Considering the small size of thisband, it was probablynot detected in allthe metaphases because of technicalproblems in performing C-banding. Aninteresting dimorphism was notedfor pair1 inoneof the females. In this specimen, pair1 was deeplyheteromorphic in all metaphases analyzed ( gs 1-3). While one homologue(1a) was asubmetacentricchromosome with an arm ratioof 1.72, the other homologue(1b) could almost be considered a subtelocentricchromosome since its arm ratiowas 2.88and, according to theclassi cation system of Greenand Sessions (1991), the Cytogeneticsof anew species of Paratelmatobiuscardosoi 49 Figure 1. Paratelmatobius sp. (aff. cardosoi)karyotype.In a,aspecimen witha homomorphicpair 1. Theinset showsa secondaryconstriction in pair 5 (arrow).In b,aspecimen witha heteromorphicpair 1. The inset shows anotherpair 1 ofthesame specimen. Notethe variable size ofmorph1b relative to1a. The secondary constriction oftheNOR inpair 9 is alsoshown in a and b (arrow). arm ratioof asubtelocentricchromosome is 3.01-7.00.These morphs of pair1 alsodiffered intheir C-banding pattern. Thus, while morph 1a showed interstitial heterochromatin inits long arm andnearly all its short arm was heterochromatic,in morph 1b, the longarm was heterochromaticand no heterochromatin was detectedin the short arm (g. 3). Intercellular variation was observedin the size of chromosome1b relative to its homologouschromosome 1a. Whilein some metaphases morphs 1a and1b showedalmost thesame size, in others, chromosome 1b was clearlysmaller than 1a (g. 1b). Themeiotic analysis of four males showed that, in 92% of 119 diakineses, the homomorphicpair 1 hada clearrod-like con guration, probably the result of aterminal associationof thelong arms (g. 4). Such an association was particularlyidenti able in 50 LucianaB. Lourenço, PauloC.A. Garcia, ShirleiM. Recco-Pimentel Figure 2. Silver-stainedkaryotype. Note the active NOR inpair 9. The inset shows the heteromorphic pair 1 in onefemale specimen. Figure 3. C-bandedkaryotype. The insets show a homomorphicpair 1, anda pair8 witha clear telomeric C-band intheshort arm. Thesmall interstitialbands of pair5 are indicatedby arrows. Table 1. Morphometricdata basedon measurements from39 metaphases of Paratelmatobius sp. (aff. cardosoi). Chromosomes 11b2 34567891011 12 r.l.14.44 11.96 11.47 10.66 10.42 9.40 8.69 6.98 5.72 5.54 5.32 4.94 4.77 a.r.1.72 2.88 1.58 2.30 1.96 1.25 1.15 1.88 1.91 1.08 1.26 1.71 1.32 c.c. sm sm m sm sm m m sm sm m m sm m r.l.:relative length(%); a.r.: arm ratio;c.c.: centromeric classication; m: metacentric; sm: submetacentric; st:subtelocentric. C-bandeddiakinesis in whichthe heterochromatic markers ofchromosome1 (almostthe wholearm andan interstitialband in thelong arm) couldbe seen( g. 4b). Cytogeneticsof anew species of Paratelmatobiuscardosoi 51 Figure 4. Giemsa-stained ( a)andC-banded ( b)diakinesesfrom a specimen withhomomorphic pair 1. Note the rod-likecon guration of this pair and the position of the short arms. Theterminal associationof the long arms is indicated(arrow). Discussion Interspecic analysis Thediploid number of Paratelmatobius sp. (aff. cardosoi)isthe same as in P. cardosoi, P.poecilogaster (Lourenço etal., 2000), and P. lutzii (De Luccaet al., 1974). The morphologyof severalchromosome pairs is also very similar among these species ( g. 5). The Paratelmatobius sp. (aff. cardosoi)karyotypediffers from thoseof P.poecilogaster and P. lutzii especiallyin pair 6, which is metacentric in Paratelmatobius sp. (aff. cardosoi)andsubtelocentric in P.poecilogaster and P. lutzii.Onthe other hand, pair 4 of Paratelmatobius sp. (aff. cardosoi)differs morphologicallyfrom thatpair 4 of P. cardosoi. Thelocation of the NOR alsodistinguishes Paratelmatobius sp. (aff. cardosoi) from P. cardosoi, since in P. cardosoi pair7 isthe NOR-bearing chromosome pair. Apart from theheterochromatic bands of pair 4 of P. cardosoi,whichare characteristic of this species,the non-centromeric bands of pairs 7 and9 of P. cardosoi are alsonot found in the Paratelmatobius sp. (aff. cardosoi)karyotype.The interstitial band in pair 3 and thetelomeric band in pair 8 thatare presentin Paratelmatobius sp. (aff. cardosoi) do not occur in P. cardosoi.Additionally,while in P. cardosoi theshort arm ofchromosome1 has aninterstitial C-band, nearly all the short arm ofchromosome1a in Paratelmatobius sp. (aff. cardosoi)isheterochromatic. All of these differences provide a clearkaryological distinctionbetween the specimens studied here and P. cardosoi,andcorroborate that Paratelmatobius sp. (aff. cardosoi)isan undescribed species. Bothkaryotypes have interstitial heterochromatin in the long arm ofpair1 (morph1a in Paratelmatobius sp.),although the amount of this heterochromatin is greater in
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