Fast and Accurate Microplate Method (Biolog MT2) for Detection of Fusarium Fungicides Resistance/Sensitivity

Abstract

The need for finding fungicides against Fusarium is a key step in the chemical plant protection and using appropriate chemical agents. Existing, conventional methods of evaluation of Fusarium isolates resistance to fungicides are costly, time-consuming and potentially environmentally harmful due to usage of high amounts of potentially toxic chemicals. Therefore, the development of fast, accurate and effective detection methods for Fusarium resistance to fungicides is urgently required. MT2 microplates (BiologTM) method is traditionally used for bacteria identification and the evaluation of their ability to utilize different carbon substrates. However, to the best of our knowledge, there is no reports concerning the use of this technical tool to determine fungicides resistance of the Fusarium isolates. For this reason, the objectives of this study are to develop a fast method for Fusarium resistance to fungicides detection and to validate the effectiveness approach between both traditional hole-plate and MT2 microplates assays. In presented study MT2 microplate-based assay was evaluated for potential use as an alternative resistance detection method. This was carried out using three commercially available fungicides, containing following active substances: triazoles (tebuconazole), benzimidazoles (carbendazim) and strobilurins (azoxystrobin), in six concentrations (0, 0.0005, 0.005, 0.05, 0.1, 0.2%), for nine selected Fusarium isolates. In this study, the particular concentrations of each fungicides was loaded into MT2 microplate wells. The wells were inoculated with the Fusarium mycelium suspended in PM4-IF inoculating fluid. Before inoculation the suspension was standardized for each isolates into 75% of transmittance. Traditional hole-plate method was used as a control assay. The fungicides concentrations in control method were the following: 0, 0.0005, 0.005, 0.05, 0.5, 1, 2, 5, 10, 25, and 50%. Strong relationships between MT2 microplate and traditional hole-plate methods were observed regarding to the detection of Fusarium resistance to various fungicides and their concentrations. The tebuconazole was most potent, providing increased efficiency in the growth inhibition of all tested isolates. Almost all among tested isolates were resistant to azoxystrobin-based fungicide. Overall, the MT2 microplates method was effective and timesaving, alternative method for determining Fusarium resistance/sensitivity to fungicides, compering to traditional hole-plate approach.

Modified microplate method for rapid and efficient estimation of siderophore produced by bacteria

“…1 ). Although other workers have also reported that microplate method is time saving and efficient (Lapinski et al 1978 ; Frac et al 2016 ), but this is the first report demonstrating the efficiency of 96 well microplate method for siderophore estimation in terms of time, money and being even more efficient than the traditional spectrophotometric method.…”

Functional Diversity of Fungal Communities in Soil Contaminated with Diesel Oil

“…Tests on plates from the FF MicroPlate ® system by Biolog guarantee obtaining a faithful description of both pure cultures of microorganisms ( Mishra and Nautiyal, 2009 ; Bourdel et al, 2016 ; Frąc et al, 2016 ) and their communities in the soil environment ( Frąc et al, 2014 ; Pająk et al, 2016 ). The functional diversity of fungi isolated from the soil not polluted with DO, determined on FF MicroPlates ® , was higher than that of fungi isolated from the soil polluted with DO, especially at day 270.…”

Metabolic and Genetic Properties of Petriella setifera Precultured on Waste

“…To prepare an inoculum, each isolate was cultivated on Potato Dextrose Agar medium (PDA) (Oxoid Ltd., England) with a 3% addition of oak sawdust (SDM), beet pulp (BPM) or, wheat bran (WBM), and control medium (CLM) without any additives, at 27°C in the dark, for 25 days including 7 days with white light exposure for spore formation. 100 μl of the mycelium water suspension was added to wells on the MT2 plate, where previously 50 μl of 1% SD, BP or WB water solution was placed, following the modified procedures of Kadali et al (2012) , Taha et al (2015) and Frąc et al (2016) , in four replicates. The inoculated microplates were incubated at 27°C for 10 days.…”