Advances in microbial diagnosis: using mass spectrometry for proteomics and genomics applications

Since the last two decades mass spectrometry (MS) has been applied to analyse the chemical cellular components of microorganisms, providing rapid and discriminatory proteomic profiles for their species identification and, in some cases, subtyping. The application of MS for the microbial diagnosis is currently well-established. The remarkable reproducibility and objectivity of this method is based on the measurement of constantly expressed and highly abundant proteins, mainly important conservative ribosomal proteins, which are used as markers to generate a cellular fingerprint. Mass spectrometry based on matrix-assisted laser desorption ionization-time of flight (MALDI- TOF) technique has been an important tool for the microbial diagnostic. However, some technical limitation concerning both MALDI-TOF and its used protocols for sample preparation have fostered the research of new mass spectrometry systems (e.g. LC MS/MS). LC MS/MS is able to generate online mass spectra of specific ions with further online sequencing of these ions, which include both specific proteins and DNA fragments. In this work a set of data for yeasts and filamentous fungi diagnostic obtained through an international collaboration project involving partners from Argentina, Brazil, Chile and Portugal will be presented and discussed.

Use of a polyphasic approach including MALDI-TOF MS for identification of Aspergillus section Flavi strains isolated from food commodities in Brazil

Brazil is one the largest producers and exporters of food commodities in the world. The evaluation of fungi capable of spoilage and the production mycotoxins in these commodities is an important issue that can be of help in bioeconomic development. The present work aimed to identify fungi of the genus Aspergillus section Flavi isolated from different food commodities in Brazil. Thirty-five fungal isolates belonging to the section Flavi were identified and characterised. Different classic phenotypic and genotypic methodologies were used, as well as a novel approach based on proteomic profiles produced by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS). Type or reference strains for each taxonomic group were included in this study. Three isolates that presented discordant identification patterns were further analysed using the internal transcribed spacer (ITS) region and calmodulin gene sequences. The data obtained from the phenotypic and spectral analyses divide the isolates into three groups, corresponding to taxa closely related to Aspergillus flavus, Aspergillus parasiticus, and Aspergillus tamarii. Final polyphasic fungal identification was achieved by joining data from molecular analyses, classical morphology, and biochemical and proteomic profiles generated by MALDI-TOF MS.

Pharmacological evaluation of plant extracts from Chapada Diamantina (Bahia, Brazil): Focus on antioxidant and cytotoxic properties

Tese de Doutoramento em Biologia de Plantas MAP - Bioplant

Enzymatic hydrophobic modification of jute fibers via grafting to reinforce composites

Horseradish peroxidase (HRP)/H2O2 system catalyzes the free-radical polymerization of aromatic compounds such as lignins and gallate esters. In this work, dodecyl gallate (DG) was grafted onto the surfaces of lignin-rich jute fabrics by HRP-mediated oxidative polymerization with an aim to enhance the hydrophobicity of the fibers. The DG-grafted jute fibers and reaction products of their model compounds were characterized by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-TOF MS), attenuated total reflection Fourier transform infrared spectroscopy (ATR-FTIR), X-ray photoelectron spectroscopy (XPS), scanning electron microscopy (SEM), thermogravimetric analysis (TGA), and differential scanning calorimetry (DSC). The results clearly indicated the grafting of DG to the jute fiber by HRP. Furthermore, the hydrophobicity of jute fabrics was determined by measuring the wetting time and static contact angle. Compared to the control sample, the wetting time and static contact angle of the grated fabrics changed from ~1 s to 1 h and from ~0° to 123.68°, respectively. This clearly proved that the hydrophobicity of jute fabrics improved considerably. Conditions of the HRP-catalyzed DG-grafting reactions were optimized in terms of the DG content of modified jute fabrics. Moreover, the results of breaking strength and elongation of DG-grafted jute/ polypropylene (PP) composites demonstrated improved reinforcement of the composite due to enzymatic hydrophobic modification of jute fibers.