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- Molecular cloning and characterization of cDNA encoding a Translocon-Associated Protein (TRAPδ) from the root-lesion nematode Pratylenchus goodeyiPublication . Pestana, Margarida; Abrantes, Isabel M. de O.; Gouveia, ManuelaThe translocon-associated protein (TRAP) complex comprises four subunits (α, β, γ, δ) and is located in the endoplasmic reticulum membrane at translocation sites. The TRAP complex is required for the efficient translocation of substrates and to correct or eliminate misfolded proteins. In this study, we described the cloning and characterization of a cDNA encoding a TRAP from the phytoparasitic nematode Pratylenchus goodeyi (Pg). The full-length cDNA had an estimated size of 690 bp and encodes a 177 amino acid peptide. The deduced protein after sequence analysis codes for TRAPδ subunit homologous to TRAPδ from other nematodes. The Pg-TRAPδ had a signal peptide indi cating a possible involvement in the transport and binding of other proteins at the endoplasmic reticulum membrane. The increase in relative expression of Pg trapδ, assessed by semi-quantitative PCR, was induced over time in nematodes exposed to a nematostatic/ nematicide extract of Solanum nigrum, suggesting that this gene product might be influenced by response mechanisms to stress in P. goodeyi. This is the first report of the cloning and characterization of trap cDNA from plant endoparasitic nematodes.
- Nematicidal activity of Solanum sisymbriifolium and S-nigrum extracts against the root-lesion nematode Pratylenchus goodeyiPublication . Pestana, M.; Rodrigues, M.; Teixeira, L.; Abrantes, I. M. de O.; Gouveia, M.; Cordeiro, NereidaThe root-lesion nematode, Pratylenchus goodeyi, is a parasite of banana plants, frequently detected in Madeira Island (Portugal) affecting culture development and consequently the production, with economical damages. To identify the phytochemicals of Solanum sisymbriifolium and S. nigrum with nematicidal properties and determine the effect of those components on P. goodeyi, an extraction sequence of at least 10 hours each from dried plants was used. The chosen solvent sequence was: dichloromethane, acetone, ethanol and water. According to the results, both plants have in their composition chemical components mainly found in water extracts, which affects the mobility and mortality of the root-lesion nematode. S sisymbriifolium and S. nigrum have potential to be used as a natural and environmentally friendly nematicide to control P. goodeyi.
- Effect of chemical stress imposed by Solanum nigrum in calreticulin and beta-1,4- endoglucanase genes and in infectivity of Pratylenchus goodeyiPublication . Pestana, M.; Abrantes, I.; Gouveia, M.Isolation and expression of effector genes encoding proteins secreted by plant-parasitic nematodes into a host can be helpful in improving the understand ing of parasitic interactions. In this study, calreticulin, a highly conserved Ca2+-binding and multifunctional pro tein, and beta-1,4-endoglucanase, a cell wall-degrading enzyme, both known to be secreted from oesophageal gland cells and injected through the nematode stylet into host tissue, were analysed. Full-length cDNAs from calreticulin (crt) and beta-1,4-endoglucanase (eng) with an estimated size of 1549 and 1342 bp, respectively, were isolated from the root-lesion nematode Pratylenchus goodeyi (Pg) by RT-PCR and RACE tech niques. Pg-crt and Pg-eng cDNAs were characterized in silico, and their expression assessed by semi quantitative PCR in nematodes exposed to a chemical stress provided by a Solanum nigrum extract showing nematicidal activity. It was demonstrated that the plant extract down-regulated the levels of Pg-crt mRNA, whereas the transcripts of Pg-eng mRNA held steady. This extract also affected nematode behaviour towards the roots since the number of nematodes that reached and penetrated the roots diminished when the exposure time rose. These observations indicate that the nemati cidal compounds present in the plant extract were effec tive as a signal to influence the infection success of P. goodeyi in vitro and it might be tested against other phytoparasitic nematodes.