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Ferreira M.-I.,University of Lisbon | Conceicao N.,University of Lisbon | David T.S.,Instituto Nacional Of Investigacao Agraria E Veterinaria Ip | Nadezhdina N.,Mendel University in Brno
Acta Horticulturae | Year: 2013

Sap flow (SF) records for 2.5 years, in shallow and deep oriented roots as well as in lignotuber of rainfed olive trees, in a traditional Portuguese orchard, can demonstrate their relative role during dry and wet seasons. Deep water reserves were found to be mobilized within the soil profile during the dry periods, namely at the end of very dry summer. The role of the lignotuber, a storage organ for starch and nutrients supporting the demands for regrowth, was also evident as a water storage organ. More relevant is the role of lignotubers in coping with dry periods due to their likely connection to sources of water in deeper layers in soil and/or subsoil, partially buffering the lack of soil water, at a time shallow roots had no access to it. © ISHS 2013.

Veigas B.,CIGMH | Jacob J.M.,CENIMAT I3N | Costa M.N.,CENIMAT I3N | Santos D.S.,CENIMAT I3N | And 6 more authors.
Lab on a Chip - Miniaturisation for Chemistry and Biology | Year: 2012

Tuberculosis (TB) remains one of the most serious infectious diseases in the world and the rate of new cases continues to increase. The development of cheap and simple methodologies capable of identifying TB causing agents belonging to the Mycobacterium tuberculosis Complex (MTBC), at point-of-need, in particular in resource-poor countries where the main TB epidemics are observed, is of paramount relevance for the timely and effective diagnosis and management of patients. TB molecular diagnostics, aimed at reducing the time of laboratory diagnostics from weeks to days, still require specialised technical personnel and labour intensive methods. Recent nanotechnology-based systems have been proposed to circumvent these limitations. Here, we report on a paper-based platform capable of integrating a previously developed Au-nanoprobe based MTBC detection assay - we call it "Gold on Paper". The Au-nanoprobe assay is processed and developed on a wax-printed microplate paper platform, allowing unequivocal identification of MTBC members and can be performed without specialised laboratory equipment. Upon integration of this Au-nanoprobe colorimetric assay onto the 384-microplate, differential colour scrutiny may be captured and analysed with a generic "smartphone" device. This strategy uses the mobile device to digitalise the intensity of the colour associated with each colorimetric assay, perform a Red Green Blue (RGB) analysis and transfer relevant information to an off-site lab, thus allowing for efficient diagnostics. Integration of the GPS location metadata of every test image may add a new dimension of information, allowing for real-time epidemiologic data on MTBC identification. © 2012 The Royal Society of Chemistry.

Ahmed A.,Royal Tropical Institute KIT | Ferreira A.S.,Instituto Nacional Of Investigacao Agraria E Veterinaria Ip | Hartskeerl R.A.,Royal Tropical Institute KIT
Methods in Molecular Biology | Year: 2014

Leptospirosis is a major zoonosis with worldwide distribution. Conventional serological typing is arduous and time consuming. Genotyping is increasingly applied for the typing and identification of leptospires and contributes to genetic and virulence divergence and molecular epidemiological characteristics such as host versus leptospires population interactions and dynamics. Presently, multilocus sequence typing (MLST) is the most robust approach. In this chapter, we describe the practical steps of two major multilocus sequence typing methods for leptospires. The first method (denoted as the 6 L scheme) is based on genotyping by phylogeny using concatenated sequences derived from six loci, including genes that encode outer membrane proteins and rrs and can be used for typing pathogenic species and strains of intermediate species. The second method (referred to as the 7 L scheme) uses seven loci on housekeeping genes and allows the analysis of seven major Leptospira pathogenic species. The 7 L scheme is web based and includes the option to analyze sequence types (STs). © Springer Science+Business Media New York 2015.

Botelho A.,Instituto Nacional Of Investigacao Agraria E Veterinaria Ip | Canto A.,Instituto Nacional Of Investigacao Agraria E Veterinaria Ip | Leao C.,Instituto Nacional Of Investigacao Agraria E Veterinaria Ip | Cunha M.V.,Instituto Nacional Of Investigacao Agraria E Veterinaria Ip | Cunha M.V.,University of Lisbon
Methods in Molecular Biology | Year: 2014

Typical CRISPR (clustered, regularly interspaced, short palindromic repeat) regions are constituted by short direct repeats (DRs), interspersed with similarly sized non-repetitive spacers, derived from transmissible genetic elements, acquired when the cell is challenged with foreign DNA. The analysis of the structure, in number and nature, of CRISPR spacers is a valuable tool for molecular typing since these loci are polymorphic among strains, originating characteristic signatures. The existence of CRISPR structures in the genome of the members of Mycobacterium tuberculosis complex (MTBC) enabled the development of a genotyping method, based on the analysis of the presence or absence of 43 oligonucleotide spacers separated by conserved DRs. This method, called spoligotyping, consists on PCR amplification of the DR chromosomal region and recognition after hybridization of the spacers that are present. The workflow beneath this methodology implies that the PCR products are brought onto a membrane containing synthetic oligonucleotides that have complementary sequences to the spacer sequences. Lack of hybridization of the PCR products to a specific oligonucleotide sequence indicates absence of the correspondent spacer sequence in the examined strain. Spoligotyping gained great notoriety as a robust identification and typing tool for members of MTBC, enabling multiple epidemiological studies on human and animal tuberculosis. © Springer Science+Business Media New York 2015.

Cunha M.V.,Instituto Nacional Of Investigacao Agraria E Veterinaria Ip | Cunha M.V.,University of Lisbon | Inacio J.,Instituto Nacional Of Investigacao Agraria E Veterinaria Ip | Inacio J.,University of Brighton
Methods in Molecular Biology | Year: 2014

Terrestrial, aquatic, and aerial animals, either domestic or wild, humans, and plants all face similar health threats caused by infectious agents. Multifaceted anthropic pressure caused by an increasingly growing and resource-demanding human population has affected biodiversity at all scales, from the DNA molecule to the pathogen, to the ecosystem level, leading to species declines and extinctions and, also, to host-pathogen coevolution processes.Technological developments over the last century have also led to quantic jumps in laboratorial testing that have highly impacted animal health and welfare, ameliorated animal management and animal trade, safeguarded public health, and ultimately helped to “secure” biodiversity. In particular, the field of molecular diagnostics experienced tremendous technical progresses over the last two decades that significantly have contributed to our ability to study microbial pathogens in the clinical and research laboratories. This chapter highlights the strengths, weaknesses, opportunities, and threats (or challenges) of molecular technologies in the framework of a veterinary microbiology laboratory, in view of the latest advances. © Springer Science+Business Media New York 2015.

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