Rapid detection of microorganisms by peptide nucleic acids

Peptide nucleic acid (PNA) molecules are DNA mimics, where the negatively charged sugarphosphate backbone is replaced by an achiral, neutral polyamide backbone formed by repetitive units of N–(2-aminoethyl) glycine. Due to their superior hybridization properties, PNA probes to detect pathogens by fluorescence in situ hybridization (FISH) have been challenging DNA probes over the last few years. In our lab, we have already designed and developed several new probes for the specific detection of bacterial species such as Helicobacter pylori, Cronobacter spp., Staphylococcus epidermidis, Salmonella spp. and Proteus spp. [1, 2]. During development and validation, probes are tested against several related species, and have been shown to be highly specific for the microorganisms of interest. All techniques were optimized in slides and then adapted for different types of samples, depending on the microorganism: H. pylori probe has been developed to work on gastric biopsies and will soon be tested in a clinical trial for a potentially commercial application; Cronobacter spp. is a major contaminant of milk-based powdered infant formula, and as such a probe to detect the pathogen after pre-enrichment of contaminated milk was devised; S. epidermidis, which is frequently present on the skin of humans, had methods developed for its identification in blood samples and catheters; and analysis of interest for Salmonella and Proteus spp. included pipes of drinking water distribution systems and urinary samples. Future work with PNA probes will involve simultaneous detection of several species in a single sample and quantitative signal detection by flow cytometry

Additive effect involving a new locus of benomyl resistance in Aspergillus nidulans

Most of the fungicides based on the benzimidazole nucleus, including benomyl, thiabendazole and thiophanate, are systemic and because they control many important fungal diseases

Tracking persistence of Helicobacter pylori in potable water biofilms using novel 16S rRNA peptide nucleic acid probes

A two stage chemostat system was used to study the pathogenic bacteria H. pylori association to drinking water biofilms. After allowing one week old biofilms to be grown on stainless steel coupons the system was inoculated with the pathogen that was detected using the recently established technique of 16S rRNA peptide nucleic acid (PNA) fluorescence in situ hybridization (FISH). Results show that H. pylori can successfully incorporate within biofilms and its presence was detected for up to five days, either in the basal layer or inside typical biofilm structures such as stacks or fronds. The PNA FISH protocol appears to be a promising new technique for the in situ visualization of microorganisms in biofilms, especially because the hydrophobic nature of the PNA molecule allows a better diffusion through the constituents of the biofilm matrix allowing an improved discrimination of microorganisms inside these naturally occurring structures. A setback in the application of this methodology was the presence of autofluorescent microorganisms. This problem can be minimized by comparing the morphologic characteristics of these suspected false positives with typical H. pylori morphology. If questions subsist, the visualization of the biofilm under different filter blocks can also improve the degree of certainty in the identification, since the reporter probe has usually a very distinctive pattern of fluorescence when compared to the autofluorescent microorganisms

Caracterizacao agronomica e culinaria de sete variedades de macaxeira.

Este trabalho objetivou estudar o comportamento produtivo e culunário das principais variedades de macaxeira cultivadas no Meio-Norte do Brasil, visando eleger as melhores e difundi-las através da distribuição de pequenas quantidades de manivas.bitstream/item/83440/1/CT1100001.pd

Development of an optimized technique for the recovery of H. pylori from water and drinking water biofilms

At present, the route of transmission is perhaps one of the most controversialareas of Helicobacter pylori research. Drinking water and associated biofilms have been suggested as possible environmental reservoirs for the bacterium, however, successful and consistent detection of the bacterium in real systems has not yet been accomplished. A new protocol that offers improved recovery ratings of H. pylori from water and associated-biofilms has been developed and evaluated. This protocol includes optimized incubation atmospheres and a new recovery culture medium, as well as an assessmentof existing techniques for the detachment of H. pylori from surfaces. Even though obvious advantages in terms of cell counts are obtained using this method, certain problems, such as contamination by faster-growing species of the culture medium in heavily contaminated waters may still hinder a successful recovery of the pathogen.Future research will involve further improvement of the protocol and monitoring of real drinking water distribution systems.Unit of Helicobacter/Campylobacter of the Center of Bacteriology of INSA para a Ciência e a Tecnologia (FCT) ; European Commission Research Project SAFER

System analysis of metabolism in Helicobacter pylori

Systems biology integrates different levels of information for understanding biological systems. The availability of the genome sequence of Helicobacter pylori has allowed the construction of a genome-scale metabolic model for this organism. In order to study the behaviour of H. pylori and understand the mechanisms associated with infection using systems biology tools and controlled cultivation conditions, fermentations in a chemically defined medium were performed and several conditions were tested. The experimental data obtained were compared with simulated data generated by the existing model. The simultaneous use of both approaches allows to correct the in silico model and, on the other hand, to rationally adjust the medium components. The improvement of the genome-scale metabolic model will allow the identification of potential targets in order to design more effective drugs for the inactivation of H. pylori

Quando suspender o tratamento com inseticidas na cultura algodoeira.

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