cytogenetic analysis of the y chromosome of native...

Arch. Zootec. 58 (221): 93-101. 2009.Recibido: 9-10-06. Aceptado: 1-4-08.

CYTOGENETIC ANALYSIS OF THE Y CHROMOSOME OF NATIVEBRAZILIAN BOVINE BREEDS: PRELIMINARY DATA

ANÁLISIS DEL CROMOSOMA Y DE RAZAS BOVINAS NATURALIZADAS BRASILEÑAS:DATOS PRELIMINARES

Issa, É.C.1, W. Jorge2, A.A. Egito3 and J.R.B. Sereno4

1Escola de Veterinária, UFG. Campus II. C.P. 131. CEP 74001-970. Goiânia, GO. Brasil. E-mail:[email protected] de Biologia Geral. ICB/UFMG. C.P. 486. CEP 31270-901, Belo Horizonte, MG. Brasil.3Embrapa Recursos Genéticos e Biotecnologia. C.P. 02372. CEP 70770-900, Brasília, DF. Brasil.4Embrapa Cerrados. C.P. 08223. CEP 73310-970, Planaltina, DF. Brasil.

ADDITIONAL KEYWORDS

Animal conservation. Animal genetic resources.Bos taurus taurus. Bos taurus indicus.

PALABRAS CLAVE ADICIONALES

Conservación animal. Recursos genéticos anima-les. Bos taurus taurus. Bos taurus indicus.

SUMMARYThe present work is a preliminary cytogenetic

study carried out on 29 males of the Brazilianbreeds Curraleiro, Crioulo Lageano, Junqueiraand Pantaneiro, available at Embrapa GeneticResources & Biotechnology, Brasília, DF andPromissão farm, Poconé, MT. The objective wasto investigate their Y chromosome morphology,whether submetacentric or acrocentric, consi-dered as markers of the taurine and zebuinesubspecies, respectively. The results showedthat of the six Curraleiro animals analyzed, fourhad an acrocentric and two had a submetacentricY chromosome, indicating contamination by zebubreeds. All Pantaneiro bulls analyzed hadexclusively zebuine (acrocentric) Y chromosome.On the other hand, the Crioulo Lageano andJunqueira bulls had submetacentric Y chromo-somes, indicating their taurine origin.

RESUMENEl objetivo de este trabajo fue realizar un

estudio preliminar del cariotipo de bovinos derazas brasileñas: Curraleiro, Crioulo Lageano yJunqueira disponibles en la granja Parque perte-neciente a Embrapa Recursos Genéticos yBiotecnología, Brasília, DF y de los Pantaneirosubicados en el norte del Pantanal, pertenecientesa la finca Promissão, Poconé, MT. Fueron analiza-

dos los cariotipos de 29 machos a través de su tipomorfológico de cromosoma Y, submetacéntrico oacrocéntrico, tipos estos considerados comomarcadores para las subespecies taurinas y ce-buínas, respectivamente. El resultado mostró que,dentro de los seis sementales Curraleiros evalua-dos, cuatro presentaron Y de cebú (acrocéntrico)y dos de taurino (submetacéntrico), sugiriendocontaminación racial por Cebú. Sin embargo, to-dos los Pantaneiros analizados presentaron ex-clusivamente cromosoma Y de Cebú (acrocén-trico), mientras que los bovinos Crioulo Lageanoy Junqueira presentaron sólo Y submetacéntrico,confirmando su origen taurino.

INTRODUCTION

According to Epstein and Mason (1984),all domesticated cattle was originated fromBos primigenius, extinct in Poland in 1627.

In Brazil, the native bovine breedsdescend from the Iberian animals whichspread all over the country, through randomcrosses and by interaction with theenvironment. As in South America therewere no animals of the bovine species at thetime of discovery, during the long colo-nization period the cattle needed to produce

Archivos de zootecnia vol. 58, núm. 221, p. 94.

ISSA, JORGE, EGITO AND SERENO

milk and meat had to be brought from theIberian Peninsula. It was in the 16th centurythat bovines first appeared in South America,coming from Portugal and Spain (Athana-ssof, 1946a).

The breeds introduced by the colonizersbecame adapted to the new environment,forming the large herds named Crioulo(Creole), which differentiated into severalvarieties (Britto, 1995). Thus, it is difficult toprecisely state which breed(s) originatedthe different native breeds. In some cases,the information about the original popula-tions was lost over time.

According to Mariante et al. (1999), ingeneral these breeds have extremelyvaluable characteristics, such as adaptationto inhospitable conditions, includingparasites and infectious diseases, draughtand poor quantity and quality of food.Preservation of the native breeds has also ahistorical value that is the genetic memoryof the animals which helped colonizing thecountry. Genetic traces of all these breedsare still left through their crossbreeds. Theintervention of man in their reproductionand selection processes did not cause anymodification of their potential, besides thosegranted by nature.

According to Egito et al. (2002), thesearch for more productive breeds led, as ofthe late 19th and the early 20th centuries, tothe import of breeds considered exotic,which, although highly productive, hadbeen selected in regions with a temperateclimate. These breeds caused, throughabsorbing crosses, the replacement anderosion of the local breeds. Although theselocal breeds presented lower productionlevels, they stood out for their completeadaptation to the tropics, where they hadundergone long-time natural selection. It istherefore necessary to preserve such ani-mal genetic resources, in order to preventtheir loss. Genetic diversity within domesti-cated species is reflected by the variety oftypes and breeds that exist and by thevariation within each one of them, and the

loss of a single type or breed compromisesthe access to its genes and unique geneticcombinations, which may be useful toagriculture in the future.

Some native Brazilian breeds, althoughnamed differently and living in distinctregions, present similar phenotypes whichraise doubts about their identity as a distinctracial group or native type. It is not knownif these populations are genetically similaror not. Even if they belong to the samebreed, they may have accumulated differentalleles due to genetic drift, as a consequenceof geographic isolation and adaptation todifferent ecological niches (Egito et al.,2002).

CURRALEIROAccording to Athanassof (1946b), this

breed is named Curraleiro in some States,such as Goiás and Tocantins, and Pé-Duroin others, such as Piauí and Maranhão. It isa native bovine breed that has not yet beenimproved, originated from the bovinesintroduced in Brazil by the Portuguese andSpaniards during the time of colonization.The Curraleiro breed stems from Bos taurusibericus and, given its European origin, it isassumed to belong to the subspecies Bostaurus taurus (Britto, 1995).

Aiming to prevent its extinction, as wellas that of other Brazilian breeds, EMBRAPAhas developed a project for their preser-vation, to be carried out by the CentroNacional de Pesquisa de Recursos Genéti-cos e Biotecnologia (National ResearchCenter for Genetic Resources and Biotech-nology) and by the Centro de PesquisaAgropecuária do Meio-Norte (AgriculturalResearch Center of the Mid-North) (CPAN).CPAN maintains a preservation unit for theCurraleiro cattle at São João do Piauí, PI, inthe semi-arid zone of the Brazilian Northeast,with approximately 300 animals (EMATER).

Tambasco et al. (1985) found both typesof Y chromosomes (acrocentric and sub-metacentric) in the Curraleiro breed. Thefrequency of the acrocentric Y chromosome


ANALYSIS OF THE Y CHROMOSOME OF NATIVE BRAZILIAN BOVINE BREEDS

was higher than that of the submetacentricY chromosome. In 1999, Britto and Melloconfirmed this finding.

PANTANEIROPantaneiro bovine, also named Cuiabano

or Tucura, descends from the Spanish breedsintroduced in America during the coloni-zation process of the Plata Basin. Accordingto Mazza et al. (1994), the literature on thephenotypic characteristics of Pantaneirobovine, from the 16th to the 18th centuries, israther scarce. Publications from the 20th

century portray these animals as short ofstature, with short, brown, dark and red-brown hair. The authors believe that thedescriptions of the animals of the Pantanalregion, made in the 20th century, are notreliable, because in that time the crosseswith their domestic and even zebuine breedswere still occurring. Based on morphologicalmeasurements and weight development ofPantaneiro cattle taken the germoplasmabank analysis of the data confirm theirreduced body size, with average weight atbirth of 24 and 22 kg and, in the adult phase,of 375 and 298 kg, for male and females,respectively.

The remaining populations, few and farbetween, of Pantaneiro cattle can be atpresent in the Northern part of the Pantanal,in the state of Mato Grosso, in areas wheremaximum flooding takes place along theParaguay, Cuiabá, São Lourenço and BentoGomes rivers. To the South, the Pantaneirocattle are practically extinct. In this momenthave only two populations from thesebreeds in the Pantanal region. One in theNorthern in Promissão farm, that doing agood work the rescue the breeds in thisareas and in the South there are a conserva-tion in situ nucleus conducting by EmbrapaPantanal, that have a lot of research aboutthis breeds in the region.

More than three centuries of nativepasture adaptation to flooded Pantanalregions conferred the Pantaneiro bovinewith rusticity, high fertility and the ability to

survive under conditions of water and foodstress conditions.

CRIOULO LAGEANOThese bovines from Southern Brazil

(Lages, State of Santa Catarina) are thosewhich keep the greatest resemblance to thetype of bovine introduced by the conque-rors, therefore of indisputable Iberian origin.This breed has evolved by natural selectionover four centuries and almost disappearedas a consequence of indiscriminate crosses.When the Catarinense Highland wascolonized, the settlers brought the Fran-queiro cattle with them that probably matedwith the bovines which already existed there,originating the Crioulo Lageano cattle(Spritze et al., 2003).

Currently, the population of thesebovines is reduced to about 500 animals, ofwhich more than 80% belong to a singlebreeder. Research works conducted in the1980's by Embrapa Recursos Genéticos eBiotecnologia and by the Federal Universityof Santa Catarina, in collaboration with afew private breeders, showed advantagesin exploring these bovines for crosses andalso as a pure breed, under the raisingconditions of the South Brazilian Highland(Spritze et al., 2003).

Tambasco et al. (1985) studied animalsof the Crioulo Lageano breed and found ahigher frequency of the submetacentric Ythan of the acrocentric Y chromosome, asopposed to the Curraleiro breed.

JUNQUEIRAFormed in the State of Minas Gerais, the

Junqueira cattle breed is a product ofcrosses between Caracu and other Brazilianvarieties. It is currently found in theJequitinhonha River basin. The animals arerobust, with a voluminous body with longand spiraled horns (EMATER). Their tail isthick and hairy; their hair is yellow, white-and reddish-brown speckled or patchy(Athanassof, 1946b). They are resistant toparasites and abiotic stresses, like the breed



that has probably originated this one, Cara-cu, a Brazilian breed that also developed inthis region.

Junqueira is a breed developed in theinland of the State of São Paulo between the18th and the 19th centuries, with an aptitudefor meat. In the past, their long horns wereused to manufacture berrantes, a kind ofmusical instrument. Nowadays this breed iscritically endangered, with less than ahundred animals left in the whole country(Diniz and Euler, 2005). Pires et al. (2004)found only submetacentric Y chromosomein the Junqueira breed.

Until this moment don't have muchinformation in the literature regarding themorphology of the Y chromosome of theseanimals. For other hands this information isbasic and very helpful to use in animalconservation program.

The first chromosome studies in bovineswere carried out in 1892 by Bardeleben, whodescribed 2n= 16 chromosomes for thespecies. Several works performed in thedecades of 1910 and 1920 described differentchromosome numbers (Schoenfeld, 1902;Hoof, 1919; Masui, 1919; Wodsedalek,1920), until 1931, when Krallinger describedfor the first time the correct chromosomenumber of taurines as 2n= 60, withsubmetacentric X and Y chromosomes. In1964, Monnier-Cambon described the Ychromosome of zebuines as acrocentric.

Different shapes of the Y chromosome inthe same breed have been described byseveral authors (Potter et al., 1979; Moraes,1978; Pinheiro, 1979; Halnan and Watson,1982; Xin and Lin, 1993; Frisch et al., 1997).

In the Brazilian breeds, the Y chromosomehas been analyzed by a few local authors.Tambasco et al. (1985) described themorphology of the Y in the Caracu, MochoNacional, Curraleiro and Crioulo Lageanobreeds. These authors observed a dimor-phism (acrocentric and submetacentric) ofthe Y chromosome in these breeds. Brittoand Mello (1999) analyzed the Y chromo-some of the Curraleiro breed and, like the

former authors, found both morphologicaltypes. The Junqueira breed showed only asubmetacentric Y chromosome, but thisstudy was based on three animals (Pires etal., 2004). Issa et al. (2006) showed that thePantaneiro cattle breed also presents thedimorphism, most of the animals have anacrocentric Y.

Giovambattista et al. (2000) studied thegeographic distribution and the haplotypefrequency of the Y chromosomes of Bostaurus and Bos indicus in Argentine andBolivian Creole breeds, using cytogeneticand molecular techniques. The taurinehaplotype (submetacentric Y chromosome)was found in 100% of the Argentine Creolebreeds, whereas in the Bolivian breeds botha submetacentric and an acrocentric Ychromosome were found, the first morpho-logical type being more frequent than thelatter.

According to Hanotte et al. (2000), 69African bovine populations from 22countries were analyzed for a marker locus(INRA124) of the Y chromosome. From the984 males studied, 693 (70%) and 291 (30%)showed the indicus and taurus allele,respectively.

The present work had the objective ofcollaborating with Brazilian AgriculturalResearch Corporation (Embrapa) and GoiásFederal University (UFG) in the charac-terization process from the native bovinebreeds, by means of a cytogenetic study ofthe Y chromosome, and help the farmers inthe animal conservation in Brazil.

MATERIAL AND METHODS

For the analysis of the Y chromosome,we used 12 males (six Curraleiro, two CriouloLageano and four Junqueira animals) fromEmbrapa Recursos Genéticos e Biotec-nologia in Brasilia. To the Pantaneiro breedwe used 17 unrelated males from Promissãofarm, Poconé, MT.

Blood was collected from the jugularvein using 4 ml Vacutainer tubes with sodium



heparin for lymphocyte culture. About 3 mlof blood were collected in each tube. Then,the tubes were placed in a styrofoam contai-ner with reusable preserving ice (eutheticice) and recyclable ice (Serono) and shippedby mail. Incubation of the cultures started28 hours after blood collection, with a relativesuccess. Cell culture was performed usingthe standard technique with 4ml culturemedium, 1 ml fetal calf serum, 0.1 mlphytohemagglutinin and 8 drops of blood,70-72 hours of incubation, 50 minutescolchicine and 20 minutes hypotonic (KCl0.075M) treatment. After fixation in 3:1methanol/acetic acid, the material wasdripped on slides, stained with Giemsa andexamined under a regular optical microscope.At least 15 metaphases of good quality fromeach animal were examined and photo-graphed using digital microphotography.

RESULTS AND DISCUSSION

In the present work, all animals studiedpresented a karyotype with 2n= 60 chromo-somes. The autosomes were acrocentric andthe X chromosome was submetacentric, itssize being approximately the same as that ofthe pair 1 autosomes.

As for the Y chromosome, four (66.67%)of the six Curraleiro animals analyzed had anacrocentric Y chromosome (figure 1), andtwo (33.33%) had a submetacentric Ychromosome (figure 2). Tambasco et al.(1985) showed that in the Caracu, Curraleiroand Mocho Nacional breeds the frequencyof acrocentric Y chromosomes was higherthan that of submetacentric Y's. In turn, inthe Crioulo Lageano breed, the result wasthe opposite. In 1999, Britto and Mellostudied the morphology of the Y chromo-some of the Curraleiro cattle and found that68% of the bulls had an acrocentric and 32%a submetacentric Y chromosome.

We found only submetacentric Ychromosomes, both in the Crioulo Lageanoand in the Junqueira breeds (figures 3 and4). As the number of animals in our sample

was small, we believe that with a larger numberof animals we might find both acrocentric andsubmetacentric Y chromosomes.

All of the 17 animals of Pantaneiro breedanalyzed in the present work from theNorthern part of the Pantanal, had anacrocentric Y chromosome (figure 5).However, Issa et al. (2006) showed thatPantaneiro breed from Nhumirin Farm, Co-rumbá, MS, South part of the Pantanal, had

Figure 1. Karyotype of Curraleiro cattleshowing an acrocentric Y chromosome,typical of Bos taurus indicus. Chromosomeconstitution: 2n= 60,XY. (Cariotipo de ganadoCurralleiro mostrando cromosoma Y acrocéntrico,típico de Bos taurus indicus, constitucióncromosómica: 2n= 60,XY).



a dimorphism. Of the 12 animals analyzed,nine (75%) had an acrocentric Y, and three(25%) a submetacentric Y chromosome.

Up to this moment, the native Brazilianbovine breeds, which present the differenttypes of Y chromosome, present the samemorphologic and productive characteristics.It was observed in the Pantaneiro cattlebreed (Abreu et al., 2006). The aim of thiswork is to contribute to management of theanimal conservation in the most of farmsin Brazil, not to elimination animals becausethe chromosome morphology. However,

this data showing that it is possible to do aselection based on chromosome morpho-logy from Pantaneiro breeds if the farmershave interesting in doing selection to taurinetypes.

CONCLUSIONS

The Y chromosomes of the CriouloLageano and Junqueira breeds are subme-tacentric, and those of the Curraleiro breedpresent dimorphism, with a higher frequencyof acrocentric and a lower frequency of

Figure 2. Karyotype of Curraleiro cattleshowing a submetacentric Y chromosome,typical of Bos taurus taurus. Chromosomeconstitution: 2n= 60,XY. (Cariotipo de ganadoCurraleiro mostrando cromosoma Y sub. meta-céntrico, típico de Bos taurus taurus, constitucióncromosómica: 2n= 60,XY).

Figure 3. Karyotype of Crioulo Lageanocattle showing a submetacentric Y chromo-some, typical of Bos taurus taurus. Chromo-some constitution: 2n= 60,XY. (Cariotipo deGanado Crioulo Lageano mostrando cromosomaY sub. metacéntrico, típico de Bos taurus taurus,constitución cromosómica: 2n= 60,XY).



submetacentric Y chromosomes. ThePantaneiro breeds from the Northern part ofthe Pantanal showing only frequency ofacrocentric. When the zebuines wereintroduced in Brazil, in the late 19th and early20th centuries, the animals brought by thecolonizers after the discovery underwent along process of breed contamination. Thus,the acrocentric Y chromosome found in partof the animals studied was acquired throughsuch random crossings.

ACKNOWLEDGMENTSWe thank Mr. Anderson Oliveira do

Carmo and Mr. Daniel Inêz dos Santos Filhofor their collaboration with the technicalpart. We also acknowledge the financialsupport granted by Coordenação deAperfeiçoamento de Pessoal de Nível Supe-rior (CAPES) and Fundação de Apoio aoDesenvolvimento do Ensino, Ciência eTecnologia do Estado de Mato Grosso doSul (FUNDECT).

Figure 5. Karyotype of Pantaneiro cattleshowing an acrocentric Y chromosome,typical of Bos taurus indicus. Chromosomeconstitution: 2n= 60,XY. (Cariotipo de ganadoPantaneiro mostrando cromosoma Y acrocéntrico,típico de Bos taurus indicus, constitucióncromosómica: 2n= 60, XY).

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Figure 4. Karyotype of Junqueira cattleshowing a submetacentric Y chromosome,typical of Bos taurus taurus. Chromosomeconstitution: 2n=60,XY. (Cariotipo de ganadoJunqueira mostrando cromosoma Y sub. meta-céntrico, típico de Bos taurus taurus, constitucióncromosómica: 2n= 60, XY).



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