The Potential of Allelochemicals from Microalgae for Biopesticides.

Improvements in agricultural productivity are required to meet the demand of a growing world population. Phytopathogens, weeds, and insects are challenges to agricultural production. The toxicity and widespread application of persistent synthetic pesticides poses a major threat to human and ecosystem health. Therefore, sustainable strategies to control pests are essential for agricultural systems to enhance productivity within a green paradigm. Allelochemicals are a less persistent, safer, and friendly alternative to efficient pest management, as they tend to be less toxic to non-target organisms and more easily degradable. Microalgae produce a great variety of allelopathic substances whose biocontrol potential against weeds, insects, and phytopathogenic fungi and bacteria has received much attention. This review provides up-to-date information and a critical perspective on allelochemicals from microalgae and their potential as biopesticides.

Quantification of schizophyllan directly from the fermented broth by ATR-FTIR and PLS regression.

Non-destructive methods that allow the quantification of bioproducts in a simple and quick manner during fermentation are extremely desirable from a practical point of view. Therefore, a 9 day fermentation experiment with Schizophyllum commune was carried out to investigate the possibility of using ATR-FTIR to quantify the schizophyllan biopolymer (SPG) directly from the culture medium. On each day, aliquots of the fermentation were taken, and the cell-free supernatant was analyzed by ATR-FTIR. The main objective of this step was to evaluate whether FTIR would be able to detect the appearance of specific peaks related to the production of SPG. The results of the PCA analysis showed that there was a reasonable separation of the days through the FTIR spectra. Then PCA-LDA was applied to the same dataset, which confirmed the formation of groups for each day of fermentation, after which, a calibration and test set was developed. Through a matrix generated by an experimental design with 2 factors and 5 levels, 25 samples were created with variations in the concentration of the culture medium and SPG. The ATR-FTIR spectra of this data set were modeled using PLS regression with backward selection of predictors. The results revealed that the amount of SPG produced can be quantified directly in the culture medium with excellent precision with R2CV = 0.951, R2P = 0.970, RMECV = 0.205 g, RMSEP = 0.170 g, RPDcv = 4.53 and RPDp = 5.88. The traditional method to quantify SPG is time consuming, requires several steps and uses solvents. In contrast, the method proposed in this work is a viable, faster, and a simpler alternative, which does not use reagents and does not require extensive pre-treatment of the samples.

A microplate assay for extracellular hydrolase detection.

This article presents a new qualitative method to detect enzyme activity replacing the conventional Agar-Petri dishes. This new method is a simple rapid and low-cost technique that uses 24-well microplates. The detection of hydrolases producing microorganisms in bioprospecting studies by qualitative methods is time consuming, costly and requires a large quantity of strains or enzymatic extracts. Tests with different substrate concentrations (0 to 20 g/L) in agar solution for the enzymatic hydrolysis analysis were performed to determine the best substrate concentrations in 24-well microplates. Other quantitative and analytical methods, such as enzymatic assays and thin layer chromatography, were performed to validate this new method and to compare the relationship between enzymatic activity and substrate degradation. Statistically relevant results were observed for amylase, endoglucanase and polygalacturonase enzymes, even when there was a low substrate concentration in agar, where the halo diameter was high. The results also indicated that the concentrations for efficient enzyme index measurements were 4 g/L carboxymethylcellulose for endoglucanase detection and 8 g/L for amylase and polygalacturonase assays. The results were presented according to the traditional methods for detection of enzymatic activity. This new method can be used as a general test for the detection of important industrial hydrolases. It is a faster and less costly alternative for screening microbial enzyme producing microorganisms and is useful for studying the production of microbial enzymes under different growing conditions.

Evaluation of Kluyveromyces marxianus endo-polygalacturonase activity through ATR-FTIR.

The endo-polygalacturonase enzyme (endoPG: EC 3.2.1.15) plays an important role in the fruit juice and wine industries, so the development of new tools for the quantitative and qualitative analysis of its enzymatic action is necessary. In this work, we report the development of a simple, fast and practical method that did not use any chemical reagent to identify and evaluate the action of the endoPG enzyme, produced by the yeast Kluyveromyces marxianus CCT3172, using attenuated total reflection Fourier-transform infrared (ATR-FTIR) spectroscopy combined with principal component analysis-linear discriminant analysis (PCA-LDA). This method evaluated the action of the endoPG enzyme on the polygalacturonic acid (PGA) substrate at 5 different times (0, 10, 15, 20 and 30 minutes), and at each time interval the samples were analyzed by ATR-FTIR. It was demonstrated that there was clear segregation between the samples that were and that were not subjected to the action of the endoPG enzyme, and it was also possible to distinguish the samples that were subjected to different incubation times with the enzyme. Through PCA-LDA it was possible to obtain wavelengths that are biomarkers for this enzymatic reaction and the observed changes as a function of hydrolysis duration were found to be in agreement with the breakdown of the glycosidic chain (1011 cm-1-CH-O- CH stretching) of PGA and release of oligosaccharides (1078 cm-1 C-OH elongation). The activity of the endoPG enzyme and the release of galacturonic acid were verified by the dinitrosalicylic acid (DNS) method in all samples. The efficacy of an automatic classifier using a principal component analysis-linear discriminant classifier (PCA-LDC) was evaluated to diagnose the action of the endoPG enzyme. The results showed an accuracy of 100% for the identification of the endoPG enzyme action and from 91.67% to 100% for classification according to the hydrolysis duration in which PGA was exposed to endoPG. The present study indicates that this methodology may be a new approach for the qualitative evaluation of the endoPG enzyme with the potential to be used in laboratories and industries.

Prevalence and Fluconazole Susceptibility Profile of Candida spp. Clinical Isolates in a Brazilian Tertiary Hospital in Minas Gerais, Brazil.

Candidiasis has become an important concern for clinical practice, especially with the increasing incidence of immunocompromised patients. In this scenario, the development resistance to fluconazole presents a challenge for treating these opportunistic infections. The aim of this study was to evaluate some epidemiology features of Candida infections in a Brazilian University Hospital using data, previously unavailable. We observed that 44% of the 93 clinical isolates tested, belonged to Candida albicans species and 56% belonged to non-Candida albicans species (mainly Candida tropicalis and Candida glabrata). Most strains were isolated from urine samples where C. albicans was predominantly detected. 29 strains presented a fluconazole resistance phenotype and of these, 22 were chemosensitised by FK506, a classical inhibitor of ABC transporters related to azoles resistance. These data suggest the probable role of efflux pumps in this resistance phenotype. Our study highlights the need for developing effective control measures for fungal infections, rational use of antifungal drugs and development of new molecules able to abrogate the active transport of antifungals.