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Previous issue date: 2008-03-20 / Sequ?ncias de DNA usadas na identifica??o de material biol?gico t?m alcan?ado consider?vel
popularidade nos ?ltimos anos, especialmente no contexto dos c?digos de barras de DNA. Aferir a
esp?cie de origem em amostras de pelos, penas, peles e particularmente fezes ? um passo
fundamental para quem estuda a ecologia e evolu??o de diversos animais com este tipo de amostra.
Este ? o caso em carn?voros, cujos h?bitos furtivos e baixas densidades populacionais de algumas
esp?cies evidenciam a import?ncia de estudos baseados em amostras n?o-invasivas. Entretanto a
atual escassez de ensaios padronizados de identifica??o de carn?voros freq?entemente dificulta a
aplica??o dessas amostras em larga escala e compara??es de resultados entre diferentes localidades.
No presente estudo n?s avaliamos dois segmentos curtos (<250 pb) de DNA mitochondrial (mtDNA)
localizados nos genes ATP sintase 6 e citocromo oxidase I com potencial de servirem como
marcadores-padr?o para identifica??o de carn?voros. Entre um e 11 indiv?duos de 66 esp?cies de
carn?voros foram seq?enciados para um ou ambos os segmentos do mtDNA e analisados usando tr?s
diferentes m?todos (?rvore de dist?ncia, dist?ncia gen?tica e an?lise de caracteres). Em geral,
indiv?duos conspec?ficos apresentaram menor dist?ncia gen?tica entre si do que em rela??o a outras
esp?cies, formando agrupamentos monofil?ticos. Exce??es foram algumas esp?cies que divergiram
recentemente, algumas das quais ainda puderam ser identificadas pelo m?todo de caracteres,
hapl?tipos esp?cie-espec?ficos, ou reduzindo a abrang?ncia geogr?fica das compara??es (restringindo
a an?lise a uma regi?o zoogeogr?fica). An?lises in silico, usando um segmento curto do citocromo b
freq?entemente empregado em carn?voros, tamb?m foram realizadas para comparar o desempenho
deste segmento em rela??o aos outros dois propostos. N?s ent?o testamos o desempenho destes
segmentos na identifica??o de fezes de carn?voros por meio de tr?s estudos de caso: (i) fezes de
felinos de zool?gico, objetivando-se verificar o potencial de contamina??o das seq?encias com DNA da
presa (coelho); (ii) fezes coletadas no Cerrado brasileiro contendo restos de presas (p?los, ossos,
penas), supostamente proveniente de lobo-guar?, objetivando-se investigar a efici?ncia de identifica??o
do predador e ocorr?ncia de interfer?ncia do DNA da presa na identifica??o; e (iii) fezes coletadas em
uma reserva na Mata Atl?ntica, tamb?m com o objetivo de avaliar a efici?ncia de identifica??o. Apesar
de diferen?as em alguns aspectos de sua performance, nossos resultados indicam que os dois
segmentos propostos t?m um bom potencial de servir como marcadores moleculares eficientes para
identifica??o acurada de amostras de carn?voros ao n?vel de esp?cie. / DNA sequences for species-level identification of biological materials have achieved considerable
popularity in the last few years, especially in the context of the DNA barcoding initiative. Species
assignment of biological samples such as hairs, feathers, pelts and particularly faeces is a crucial step
for those interested in studying ecology and evolution of many species with these samples. This is
especially the case for carnivores, whose elusive habits and low densities highlight the importance of
studies based on noninvasive samples. However, the current lack of standardized assays for
carnivore identification often poses challenges to the large-scale application of this approach, as well
as the cross-comparison of results among sites. Here we evaluate the potential of two short (<250 pb)
mitochondrial DNA (mtDNA) segments located within the genes ATP synthase 6 and cytochrome
oxidase I as standardized markers for carnivore identification. Between one and eleven individuals of
66 carnivore species were sequenced for one or both of these mtDNA segments and analyzed using
three different approaches (tree-based, distance-based and character-based), in conjunction with
sequences retrieved from public databases. In most cases, conspecific individuals had lower genetic
distances from each other relative to other species, resulting in diagnosable monophyletic clusters.
Notable exceptions were the more recently diverged species, some of which could still be identified
using diagnostic character attributes, species-specific haplotypes, or by reducing the geographic
scope of the comparison (restricting the analysis to a single zoogeographic region). Additional in silico
analyses using a short cytochrome b segment frequently employed in carnivore identification were
also performed aiming to compare performance to that of our two focal markers. We then tested the
performance of these segments in the identification of carnivore faeces via three case studies: (i) felid
faeces collected in a controlled zoo experiment, aimed at assessing whether DNA from rabbit prey
would contaminate the resulting sequences; (ii) field-collected faeces from the Brazilian Cerrado
presumed to be from maned wolves and containing prey remains (hairs, bones, feathers), aimed at
investigating the efficiency of predator identification and occurrence of prey DNA interference; and (iii)
field-collected scats from an Atlantic Forest study site, also addressing the issue of PCR success rate
and identification efficiency. In spite of some relevant differences in some aspects of their
performance, our results indicate that both of our focal segments have a good potential to serve as
efficient molecular markers for accurate species-level identification of carnivore samples.
| Identifer | oai:union.ndltd.org:IBICT/oai:tede2.pucrs.br:tede/8077 |
| Date | 20 March 2008 |
| Creators | Chaves, Paulo Bomfim |
| Contributors | Eizirik, Eduardo |
| Publisher | Pontif?cia Universidade Cat?lica do Rio Grande do Sul, Programa de P?s Gradua??o em Zoologia, PUCRS, Brasil, Escola de Ci?ncias |
| Source Sets | IBICT Brazilian ETDs |
| Language | English |
| Detected Language | English |
| Type | info:eu-repo/semantics/publishedVersion, info:eu-repo/semantics/masterThesis |
| Format | application/pdf |
| Source | reponame:Biblioteca Digital de Teses e Dissertações da PUC_RS, instname:Pontifícia Universidade Católica do Rio Grande do Sul, instacron:PUC_RS |
| Rights | info:eu-repo/semantics/openAccess |
| Relation | -6482652380601267558, 500 |
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