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Cardoso J.M.S.,University of Coimbra | Fonseca L.,University of Coimbra | Gomes P.,Next Generation Sequencing Unit | Egas C.,Next Generation Sequencing Unit | Abrantes I.,University of Coimbra
Forest Pathology | Year: 2015

In this study, a cDNA sequence corresponding to the Bursaphelenchus xylophilus unc-87 homolog gene (Bx-unc-87) was identified in B. xylophilus transcriptome and characterized. This cDNA has an 1152-bp-long open-reading frame that is putatively translated into a 383 amino acids peptide. The deduced protein (Bx-UNC-87) contains highly conserved regions of the calponin-like proteins and has high sequence identity to the calponin proteins of other nematodes. In Caenorhabditis elegans, the calponin protein encoded by gene unc-87 is required to maintain the structure of myofilaments in muscle cells of the body wall. The silencing of Bx-unc-87 gene was performed by RNAi, soaking the nematodes in a solution containing dsRNA of this gene. In addition to a no dsRNA control, a non-homologous dsRNA control was also included. The Bx-unc-87 transcription was downregulated in nematodes treated with Bx-unc-87 dsRNA in comparison with the non-dsRNA-treated nematodes. dsRNA-treated nematodes revealed some paralysis and uncoordinated movement in contrast to the regular and sinusoidal movement of the non-treated nematodes. Furthermore, reproduction was reduced in dsRNA-treated nematodes. These results suggest that this protein has a function similar to that of the C. elegans homolog. © 2015 Blackwell Verlag GmbH. Source


Egas C.,Next Generation Sequencing Unit | Barroso C.,Next Generation Sequencing Unit | Froufe H.J.C.,Next Generation Sequencing Unit | Pacheco J.,Next Generation Sequencing Unit | And 2 more authors.
Standards in Genomic Sciences | Year: 2015

Rubrobacter radiotolerans strain RSPS-4 is a slightly thermophilic member of the phylum "Actinobacteria" isolated from a hot spring in São Pedro do Sul, Portugal. This aerobic and halotolerant bacterium is also extremely resistant to gamma and UV radiation, which are the main reasons for the interest in sequencing its genome. Here, we present the complete genome sequence of strain RSPS-4 as well as its assembly and annotation. We also compare the gene sequence of this organism with that of the type strain of the species R. radiotolerans isolated from a hot spring in Japan. The genome of strain RSPS-4 comprises one circular chromosome of 2,875,491 bp with a G+C content of 66.91%, and 3 circular plasmids of 190,889 bp, 149,806 bp and 51,047 bp, harboring 3,214 predicted protein coding genes, 46 tRNA genes and a single rRNA operon. © 2014 The Author(s). Source


Simoes M.J.,Next Generation Sequencing Unit | Lobo C.,Association for Innovation and Biomedical Research on Light and Image AIBILI | Lobo C.,Centro Hospitalar iversitario Of Coimbra | Egas C.,Next Generation Sequencing Unit | And 7 more authors.
Ophthalmologica | Year: 2014

Purpose: To explore phenotype-genotype correlations that may contribute to a better understanding of diabetic retinopathy (DR). Procedures: An exploratory association study was performed to identify genetic variants associated with non-proliferative DR (NPDR) in 307 type 2 diabetic patients who were previously stratified into 3 different phenotypes of NPDR progression. The 307 patients were genotyped for 174 single nucleotide polymorphisms of 11 candidate genes (ACE, AGER, AKR1B1, ICAM1, MTHFR, NOS1, NOS3, PPARGC1A, TGFB1, TNF and VEGFA). Results: Significant associations were observed for PPARGC1A rs16874120 with phenotype A (odds ratio, OR = 0.60, 95% confidence interval, CI 0.36-0.99), ICAM1 rs1801714 with phenotype B (OR = 3.32, 95% CI 1.05-10.50) and both PPARGC1A rs10213440 (OR = 2.00, 95% CI 1.07-3.73) and MTHFR rs1801133 (OR = 1.84, 95% CI 1.08-3.11) with phenotype C. Conclusions: Results indicate that specific gene variants in ICAM1, PPARGC1A and MTHFR are associated with different NPDR phenotypes, being likely candidates to explain different disease mechanisms underlying the different phenotypes. This is the first study to show correlations between specific gene variants and NPDR phenotypes, opening new perspectives on DR. © 2014 S. Karger AG, Basel. Source


Figueiredo J.,University of Coimbra | Simoes M.J.,Next Generation Sequencing Unit | Gomes P.,Next Generation Sequencing Unit | Barroso C.,Next Generation Sequencing Unit | And 6 more authors.
PLoS ONE | Year: 2013

The pinewood nematode, Bursaphelenchus xylophilus, is native to North America but it only causes damaging pine wilt disease in those regions of the world where it has been introduced. The accurate detection of the species and its dispersal routes are thus essential to define effective control measures. The main goals of this study were to analyse the genetic diversity among B. xylophilus isolates from different geographic locations and identify single nucleotide polymorphism (SNPs) markers for geographic origin, through a comparative transcriptomic approach. The transcriptomes of seven B. xylophilus isolates, from Continental Portugal (4), China (1), Japan (1) and USA (1), were sequenced in the next generation platform Roche 454. Analysis of effector gene transcripts revealed inter-isolate nucleotide diversity that was validated by Sanger sequencing in the genomic DNA of the seven isolates and eight additional isolates from different geographic locations: Madeira Island (2), China (1), USA (1), Japan (2) and South Korea (2). The analysis identified 136 polymorphic positions in 10 effector transcripts. Pairwise comparison of the 136 SNPs through Neighbor-Joining and the Maximum Likelihood methods and 5-mer frequency analysis with the alignment-independent bilinear multivariate modelling approach correlated the SNPs with the isolates geographic origin. Furthermore, the SNP analysis indicated a closer proximity of the Portuguese isolates to the Korean and Chinese isolates than to the Japanese or American isolates. Each geographic cluster carried exclusive alleles that can be used as SNP markers for B. xylophilus isolate identification. © 2013 Figueiredo et al. Source

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