Pyrenophora teres: Population structure, virulence and aggressiveness in Southern Russia.

The barley net blotch agent Pyrenophora teres (Died) Drechs. is one of the dominant fungal pathogens in agricultural crops worldwide. Here we aim to study the aggressiveness and virulence of P. teres populations collected at different ontogenesis stages (BBCH 30 and BBCH 47) from winter barley cultivars of various resistance types: moderately resistant, moderately susceptible and highly susceptible. We observed a direct proportional relationship between cultivar resistance and the aggressiveness of P. teres populations collected in both growth phases of the host plant. The isolates collected at an early stage of host plant development have a large difference in aggressiveness criteria: colony growth rate, sporulation intensity, latency period, plant damage degree, and the number of identified races. At the BBCH 30 growth stage, the growth rate of fungus colonies selected from a resistant cultivar is 1.2 times higher than that of a susceptible cultivar. The growth rate of colonies selected from resistant and susceptible cultivars in the earlier BBCH 30 stage is 1.04 times higher than the growth rate of colonies selected from the later phase. The sporulation intensity of fungal populations selected from a resistant cultivar is higher than that of populations selected from a susceptible cultivar (for BBCH 30-5.4 times, for BBCH 47-4.0 times); and it is 1.3 times higher in an earlier phase of plant development. Correlation between colony growth rate and spore formation rate in the BBCH 30 is r = 0.4. A high correlation level (r = 0.9) and notable difference between the variants were revealed when studying the duration of the latent period. The average value of plant damage by the P. teres from resistant cultivar is 4 times higher than from the susceptible cultivar in the BBCH 30 stage; and 12 times - in the BBCH 47 stage. There is a moderate negative correlation between the plant damage degree and the number of races identified from the fungal population, r = -0.59 for the BBCH 30, r = -0.8 for the BBCH 47. The number of races identified from P. teres populations collected in the late phase of plant growth was one third less. Our study helped to acquire new knowledge about intrapopulation processes under the influence of various factors - plant growth stage and cultivar genotype. The results obtained are the basis for the development of adaptive-integrated techniques for managing populations of the hemibiotrophic pathogen, barley net blotch.

Gene expression and metabolite analysis in barley inoculated with net blotch fungus and plant growth-promoting rhizobacteria.

Net blotch, caused by the ascomycete Drechslera teres, can compromise barley production. Beneficial bacteria strains are of substantial interest as biological agents for plant protection in agriculture. Belonging to the genus Paraburkholderia, a bacterium, referred to as strain B25, has been identified as protective for barley against net blotch. The strain Paraburkholderia phytofirmans (strain PsJN), which has no effect on the pathogen's growth, has been used as control. In this study, the expression of target genes involved in cell wall-related processes, defense responses, carbohydrate and phenylpropanoid pathways was studied under various conditions (with or without pathogen and/or with or without bacterial strains) at different time-points (0-6-12-48 h). The results show that specific genes were subjected to a circadian regulation and that the expression of most of them increased in barley infected with D. teres and/or bacterized with the strain PsJN. On the contrary, a decreased gene expression was observed in the presence of strain B25. To complement and enrich the gene expression analysis, untargeted metabolomics was carried out on the same samples. The data obtained show an increase in the production of lipid compounds in barley in the presence of the pathogen. In addition, the presence of strain B25 leads to a decrease in the production of defense compounds in this crop. The results contribute to advance the knowledge on the mechanisms occurring at the onset of D. teres infection and in the presence of a biocontrol agent limiting the severity of net blotch in barley.

Expression Analysis of Cell Wall-Related Genes in the Plant Pathogenic Fungus Drechslera teres.

Drechslera teres (D. teres) is an ascomycete, responsible for net blotch, the most serious barley disease causing an important economic impact. The cell wall is a crucial structure for the growth and development of fungi. Thus, understanding cell wall structure, composition and biosynthesis can help in designing new strategies for pest management. Despite the severity and economic impact of net blotch, this is the first study analyzing the cell wall-related genes in D. teres. We have identified key genes involved in the synthesis/remodeling of cell wall polysaccharides, namely chitin, ß-(1,3)-glucan and mixed-linkage glucan synthases, as well as endo/exoglucanases and a mitogen-activated protein kinase. We have also analyzed the differential expression of these genes in D. teres spores and in the mycelium after cultivation on different media, as well as in the presence of Paraburkholderia phytofirmans strain PsJN, a plant growth-promoting bacterium (PGPB). The targeted gene expression analysis shows higher gene expression in the spores and in the mycelium with the application of PGPB. Besides analyzing key cell-wall-related genes, this study also identifies the most suitable reference genes to normalize qPCR results in D. teres, thus serving as a basis for future molecular studies on this ascomycete.

Genetic structure of Pyrenophora teres net and spot populations as revealed by microsatellite analysis.

The population structure of the fungal pathogen Pyrenophora teres, collected mainly from different regions of the Czech and Slovak Republics, was examined using a microsatellite analyses (SSR). Among 305 P. teres f. teres (PTT) and 82 P. teres f. maculata (PTM) isolates that were collected, the overall gene diversity was similar (h = 0.12 and h = 0.13, respectively). A high level of genetic differentiation (FST = 0.46; P < 0.001) indicated the existence of population structure. Nine clusters that were found using a Bayesian approach represent the genetic structure of the studied P. teres populations. Two clusters consisted of PTM populations; PTT populations formed another seven clusters. An exact test of population differentiation confirmed the results that were generated by Structure. There was no difference between naturally infected populations over time, and genetic distance did not correlate with geographical distance. The facts that all individuals had unique multilocus genotypes and that the hypothesis of random mating could not be rejected in several populations or subpopulations serve as evidence that a mixed mating system plays a role in the P. teres life cycle. Despite the fact that the genetic differentiation value between PTT and PTM (FST = 0.30; P < 0.001) is lower than it is between the populations within each form (FST = 0.40 (PTT); FST = 0.35 (PTM); P < 0.001) and that individuals with mixed PTT and PTM genomes were found, the two forms of P. teres form genetically separate populations. Therefore, it can be assumed that these populations have most likely undergone speciation.