Frequency of Alternaria Genotypes Resistant to SDHI Fungicides in California Pistachio Orchards Determined by Real-Time PCR.

Real-time PCR methods were developed to quantify the frequency of SDHC-H134R and SDHB-H277Y mutants associated with succinate dehydrogenase inhibitors (SDHI) resistance in Alternaria populations from pistachio. The linearity of the standard curves demonstrated the applicability in the quantification of the assays. The accuracy and reliability of the qPCR protocols to determine the frequency of mutants in real samples were corroborated. Orchards visibly affected by Alternaria late blight were sampled. The frequency of mutants was determined using the qPCR assays, while the frequency of resistant phenotypes was determined using a single discriminatory dose. The statistical analysis showed that the frequencies of the mutation SDHC-H134R determined with the qPCR assay were highly correlated with those estimated with the conventional method. The survey also evidenced that resistance to boscalid is still widespread in California. Results also indicated the possible contribution of other mutations to SDHI resistance. Our results confirmed the prevalence of SDHC-H134R mutants and the occurrence of mutation SDHB-H277Y at low frequencies. The real-time PCR methods developed in this study were able to detect differences in the frequencies of resistant mutants caused by the use of chemical fungicides. Finally, the effects of two fungicide programs on the frequency of mutants resistant to SDHI and quinone outside inhibitors fungicides were studied using qPCR assays. The experiments demonstrated that the use of anilinopyrimidine and demethylation inhibitors fungicides in the same program reduced the frequency of these mutations in Alternaria populations. The qPCR methods developed and used in this study can be used to track resistance levels in the pistachio orchards on a large scale.

Neopestalotiopsis species presenting wide dye destaining activity: report of a mycelium-associated laccase.

Wastewaters from textile dyeing industries represent an ecological concern, notably due to the known toxicity of azo dyes to the local microbiome and human health. Although physicochemical approaches are the rule for the treatment of industrial effluents, biological strategies such as enzyme-mediated dye destaining is a promising alternative. Notwithstanding a broad range of microorganisms, including fungi, algae, yeast, and bacteria, display dye-destaining properties, most of the literature has focused in ligninolytic fungi, leaving other classes of organisms somehow ignored. In this study, six endophytic strains isolated from Maytenus ilicifolia were studied for their destaining activity. The phylogenetic and morphological analysis allowed the identification of strain LGMF1504 as Neopestalotiopsis sp. LGMF1504 that decolorized several commercial dyes as the result of a mycelium-associated laccase. The enzyme expression was modulated by carbon and nitrogen content in the culture medium, it was weakly affected by the presence of aromatic compounds and metal ions while some common laccase mediators improved the destaining activity onto dye substrates. The best culture condition observed for laccase activity was a basic culture medium containing 5 g L-1 starch and 15 g L-1 ammonium tartrate. The laccase activity showed low substrate specificity and almost unaltered performance in a wide range of pH values and NaCl concentrations, suggesting the potential of Neopestalotiopsis sp. LGMF1504 for biodegradation approaches.