Research experience
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Jan 2005–
presentResearch: Universidade do Porto
Universidade do Porto · Institute of Molecular Pathology and Immunology (IPATIMUP) · Population GeneticsPortugal · Porto
Publications (28) View all
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Article: New Insights into the Phylogeny and Worldwide Dispersion of Two Closely Related Nematode Species, Bursaphelenchus xylophilus and Bursaphelenchus mucronatus.
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ABSTRACT: The pinewood nematode, Bursaphelenchus xylophilus, is one of the greatest threats to coniferous forests worldwide, causing severe ecological damage and economic loss. The biology of B. xylophilus is similar to that of its closest relative, B. mucronatus, as both species share food resources and insect vectors, and have very similar morphological characteristics, although little pathogenicity to conifers has been associated with B. mucronatus. Using both nuclear and mitochondrial DNA markers, we show that B. xylophilus and B. mucronatus form distinct phylogenetic groups with contrasting phylogeographic patterns. B. xylophilus presents lower levels of intraspecific diversity than B. mucronatus, as expected for a species that evolved relatively recently through geographical or reproductive isolation. Genetic diversity was particularly low in recently colonised areas, such as in southwestern Europe. By contrast, B. mucronatus displays high levels of genetic diversity and two well-differentiated clades in both mitochondrial and nuclear DNA phylogenies. The lack of correlation between genetic and geographic distances in B. mucronatus suggests intense gene flow among distant regions, a phenomenon that may have remained unnoticed due to the reduced pathogenicity of the species. Overall, our findings suggest that B. xylophilus and B. mucronatus have different demographic histories despite their morphological resemblance and ecological overlap. These results suggest that Bursaphelenchus species are a valuable model for understanding the dispersion of invasive species and the risks posed to native biodiversity and ecosystems.PLoS ONE 01/2013; 8(2):e56288. · 4.09 Impact Factor -
Article: New Method for the Simultaneous Identification of Cow, Sheep, Goat, and Water Buffalo in Dairy Products by Analysis of Short Species-Specific Mitochondrial DNA Targets.
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ABSTRACT: A novel method is presented here as an analytical tool for food control and authentication of dairy products manufactured from the milk of cow, sheep, goat, and buffalo. The method is based on multiplex polymerase chain reaction (PCR) of species-specific mitochondrial DNA (mtDNA) targets followed by fragment size analysis by capillary electrophoresis. The method includes (a) simultaneous detection of four species, (b) internal control for DNA extraction and PCR, (c) mtDNA as a target for PCR, (d) amplicons of <200 bp, and (e) flexibility in the electrophoresis and fragment size detection method. Species identification proved to be straightforward, efficient, sensitive, and robust. The method is sensitive to an at least 1% (v/v) relative proportion of milk in binary mixtures. A survey of commercial products showed that 12.5% failed to conform to the description of the contents, by either the introduction or absence of listed species, thus demonstrating the relevance of this type of testing.Journal of Agricultural and Food Chemistry 10/2012; · 2.82 Impact Factor -
Article: SPInDel: a multifunctional workbench for species identification using insertion/deletion variants.
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ABSTRACT: The majority of the available methods for the molecular identification of species use pairwise sequence divergences between the query and reference sequences (DNA barcoding). The presence of multiple insertions and deletions (indels) in the target genomic regions is generally regarded as a problem, as it introduces ambiguities in sequence alignments. However, we have recently shown that a high level of species discrimination is attainable in all taxa of life simply by considering the length of hypervariable regions defined by indel variants. Each species is tagged with a numeric profile of fragment lengths-a true numeric barcode. In this study, we describe a multifunctional computational workbench (named SPInDel for SPecies Identification by Insertions/Deletions) to assist researchers using variable-length DNA sequences, and we demonstrate its applicability in molecular ecology. The SPInDel workbench provides a step-by-step environment for the alignment of target sequences, selection of informative hypervariable regions, design of PCR primers and the statistical validation of the species-identification process. In our test data sets, we were able to discriminate all species from two genera of frogs (Ansonia and Leptobrachium) inhabiting lowland rainforests and mountain regions of South-East Asia and species from the most common genus of coral reef fishes (Apogon). Our method can complement conventional DNA barcoding systems when indels are common (e.g. in rRNA genes) without the required step of DNA sequencing. The executable files, source code, documentation and test data sets are freely available at http://www.portugene.com/SPInDel/SPInDel_webworkbench.html.Molecular Ecology Resources 09/2012; 12(6):1190-5. · 3.06 Impact Factor -
Article: Mitochondrial DNA deletions are associated with non-B DNA conformations.
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ABSTRACT: Mitochondrial DNA (mtDNA) deletions are a primary cause of mitochondrial disease and are believed to contribute to the aging process and to various neurodegenerative diseases. Despite strong observational and experimental evidence, the molecular basis of the deletion process remains obscure. In this study, we test the hypothesis that the primary cause of mtDNA vulnerability to breakage resides in the formation of non-B DNA conformations, namely hairpin, cruciform and cloverleaf-like elements. Using the largest database of human mtDNA deletions built thus far (753 different cases), we show that site-specific breakage hotspots exist in the mtDNA. Furthermore, we discover that the most frequent deletion breakpoints occur within or near predicted structures, a result that is supported by data from transgenic mice with mitochondrial disease. There is also a significant association between the folding energy of an mtDNA region and the number of breakpoints that it harbours. In particular, two clusters of hairpins (near the D-loop 3'-terminus and the L-strand origin of replication) are hotspots for mtDNA breakage. Consistent with our hypothesis, the highest number of 5'- and 3'-breakpoints per base is found in the highly structured tRNA genes. Overall, the data presented in this study suggest that non-B DNA conformations are a key element of the mtDNA deletion process.Nucleic Acids Research 05/2012; 40(16):7606-21. · 8.03 Impact Factor -
Article: Identification of mtDNA lineages of Sus scrofa by multiplex single base extension for the authentication of processed food products.
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ABSTRACT: A genetic method to identify the breed of origin could serve as a useful tool for inspecting the authenticity of the increasing number of monobreed foodstuffs, such as those derived from small local European pig breeds. Mitochondrial DNA (mtDNA) is practically the only reliable genomic target for PCR in processed products, and its haploid nature and strict maternal inheritance greatly facilitate genetic analysis. As a result of strategies that sought to improve the production traits of European pigs, most industrial breeds presently show a high frequency of Asian alleles, while the absence or low frequency of such Asian alleles has been observed in small rustic breeds from which highly prized dry-cured and other traditional products are derived. Therefore, the detection of Asian ancestry would indicate nonconformity in Protected Denomination of Origin products. This study presents a single base extension assay based on 15 diagnostic mtDNA single nucleotide polymorphisms to discriminate between Asian and European Sus scrofa lineages. The test was robust, sensitive and accurate in a wide range of processed foodstuffs and allowed accurate detection of pig genetic material and identification of maternal ancestry. A market survey suggested that nonconformity of products derived from Portuguese breeds is an unusual event at present, but regular surveys both in the local populations and in commercial products would be advisible. Taking into consideration the limitations presented by other methodologies, this mtDNA-based test probably attains the highest resolution for the direct genetic test for population of origin in Sus scrofa food products.Journal of Agricultural and Food Chemistry 06/2011; 59(13):6920-6. · 2.82 Impact Factor
