María Virginia Moreno

ISSR markers detect high genetic variation among Fusarium poae isolates from Argentina and England

Fusarium poae is one of the Fusarium species isolated from cereal grains infected by Fusarium head blight (FHB), and in recent years it has been identified as a major FHB component. In this study, 97 F. poae isolates from Argentina (n = 62) and England (n = 35) were analysed by inter-simple sequence repeats (ISSR) to examine the genetic diversity and to determine whether intraspecific variation could be correlated with geographic and/or host origin. The molecular analysis showed high intraspecific variability within F. poae isolates, but did not reveal a clear relationship between variability and the host/geographic origin. Fusarium poae isolates from the same geographic region or host appeared in different subclusters. Conversely, isolates with the same haplotype were also collected from different geographic regions. However, we did observe subclusters consisting of isolates from Argentina only or from England only. Furthermore, a single seed sample was found to host different haplotypes. Analysis of molecular variance (AMOVA) indicated a high genetic variability in F. poae, with most of the genetic variability explained by differences within, rather than between Argentinean and English populations. This is the first report on genetic diversity of F. poae using ISSR markers. Moreover, ISSR fingerprinting generates highly polymorphic markers for F. poae and proved to be a useful and reliable assay for genetic variability studies.

Pathogenic and molecular variability among isolates of Pyrenophora tritici-repentis , causal agent of tan spot of wheat in Argentina

Tan spot, caused by Pyrenophora tritici-repentis, is a common disease of wheat (Triticum aestivum) responsible for economic losses in some wheat growing areas worldwide. In this study the pathogenic and genetic diversity of 51 P. tritici-repentis isolates collected from different ecological regions of Argentina were analyzed. Virulence tests were conducted on 10 selected wheat cultivars: Buck Halcón, Chris, Gabo, Glenlea, Klein Dragón, Klein Sendero, Max, ND 495, ProInta Guazú and ProInta Imperial. Data revealed significant differences between all main factors evaluated and the interactions for 19 of the isolates analyzed. Based on the reaction type of each isolate/cultivar combination, 48 different pathogenic patterns were detected. The molecular analysis using Inter-Simple Sequence Repeats (ISSR) revealed the existence of 36 different haplotypes among 37 isolates of P. tritici-repentis originally selected for this study. These results indicate that P. tritici-repentis on wheat in Argentina is a heterogeneous fungus, implying that screening wheat germoplasm for resistance for tan spot disease requires a wide range of pathogen isolates.

Cytokinin production by Pseudomonas fluorescens G20-18 determines biocontrol activity against Pseudomonas syringae in Arabidopsis

Plant beneficial microbes mediate biocontrol of diseases by interfering with pathogens or via strengthening the host. Although phytohormones, including cytokinins, are known to regulate plant development and physiology as well as plant immunity, their production by microorganisms has not been considered as a biocontrol mechanism. Here we identify the ability of Pseudomonas fluorescens G20-18 to efficiently control P. syringae infection in Arabidopsis, allowing maintenance of tissue integrity and ultimately biomass yield. Microbial cytokinin production was identified as a key determinant for this biocontrol effect on the hemibiotrophic bacterial pathogen. While cytokinin-deficient loss-of-function mutants of G20-18 exhibit impaired biocontrol, functional complementation with cytokinin biosynthetic genes restores cytokinin-mediated biocontrol, which is correlated with differential cytokinin levels in planta. Arabidopsis mutant analyses revealed the necessity of functional plant cytokinin perception and salicylic acid-dependent defence signalling for this biocontrol mechanism. These results demonstrate microbial cytokinin production as a novel microbe-based, hormone-mediated concept of biocontrol. This mechanism provides a basis to potentially develop novel, integrated plant protection strategies combining promotion of growth, a favourable physiological status and activation of fine-tuned direct defence and abiotic stress resilience.

Pyrenophora tritici-repentis, Causal Agent of Tan Spot: A Review of Intraspecific Genetic Diversity

M. V. Moreno1,2,3, S. A. Stenglein1,2,3 and A. E. Perello1,4 1Consejo Nacional de Investigaciones Cientificas y Tecnicas (CONICET) 2Laboratorio de Biologia Funcional y Biotecnologia (BIOLAB)-CEBB-CONICET, Facultad de Agronomia de Azul Universidad Nacional del Centro de la Provincia de Buenos Aires (UNCPBA) 3Microbiologia Agricola, Facultad de Agronomia, UNCPBA 4Centro de Investigaciones de Fitopatologia (CIDEFI) Facultad de Ciencias Agrarias y Forestales, Universidad Nacional de La Plata Argentina

Distribution of races and Tox genes in Pyrenophora tritici-repentis isolates from wheat in Argentina

Tan spot, caused by Pyrenophora tritici-repentis, is a common disease in wheat-growing regions of Argentina. In this study 65 isolates of P. tritici-repentis obtained from different cultivars and wheat regions of Argentina were assessed for their virulence on six wheat cultivars/lines (Glenlea, Salomouni, Katepwa, M-3, 6B365 and 6B662) and for the presence/absence of the Tox genes based on a PCR approach. Thirty-six isolates were assigned to races, of which races 4 and 8 were dominant. Results for molecular analysis of ToxA, ToxB, ToxB-like and toxb genes showed that 57 isolates possessed the ToxA gene whereas only one isolate possessed ToxA and ToxB genes. There was no correlation between races and the toxin genotypes. It is suggested that P. tritici-repentis exhibits a complex race structure in Argentina.

Natural Contamination with Mycotoxins Produced by Fusarium graminearum and Fusarium poae in Malting Barley in Argentina

Two of the most common species of toxin-producing Fusarium contaminating small cereal grains are Fusarium graminearum and F. poae; with both elaborating diverse toxins, especially deoxynivalenol (DON) and nivalenol (NIV), respectively. The objective of our work during the 2012–2014 growing seasons was to screen crops for the most commonly isolated Fusarium species and to quantify DON and NIV toxins in natural malting-barley samples from different producing areas of Argentina. We identified 1180 Fusarium isolates in the 119 samples analyzed, with 51.2% being F. graminearum, 26.2% F. poae and 22.6% other species. We found high concentrations of mycotoxins, at maximum values of 12 μg/g of DON and 7.71 μg/g of NIV. Of the samples, 23% exhibited DON at an average of 2.36 μg/g, with 44% exceeding the maximum limits (average of 5.24 μg/g); 29% contained NIV at an average of 2.36 μg/g; 7% contained both DON and NIV; and 55% were without DON or NIV. Finally, we report the mycotoxin contamination of the grain samples produced by F. graminearum and F. poae, those being the most frequent Fusarium species present. We identified the main Fusarium species affecting natural malting-barley grains in Argentina and documented the presence of many samples with elevated concentrations of DON and NIV. To our knowledge, the investigation reported here was the first to quantify the contamination by Fusarium and its toxins in natural samples of malting barley in Argentina.

Pseudopithomyces chartarum associated with wheat seeds in Argentina, pathogenicity and evaluation of toxigenic ability

Argentina is one of the top 10 world producers and exporters of wheat. In routine surveys of wheat (Triticum aestivum L.) in Buenos Aires Province, Argentina, a new disease was observed in 2012 on seeds of wheat cv. Buck Meteoro. Symptomatic grains (black points) and leaves (chlorosis and spots) were collected during the spring of that year. The objectives of the present study were to identify the causal agent, to investigate its pathogenicity in relation to nine wheat cultivars and to identify the secondary metabolites produced by fungus. Symptomatic grains were plated on potato dextrose agar (PDA). Morphological characterization of colonies and sequencing of the ITS region after DNA extraction identified it as Pseudopithomyces chartarum. For pathogenicity tests, two different isolates, P221 and P224, were inoculated on seedlings of nine wheat cultivars, which showed different disease symptoms, % of grain germination (GG), % of grain discoloration (GD) and % of weak seedlings (WS), suggesting different levels of response against Pseudopithomyces chartarum. Particularly B. Meteoro, Buck Guapo and Klein Proteo cultivars demonstrated major infection tolerance for GD and WS. In contrast, Sy 100 and Klein Pantera were most affected showing weakness, chlorosis, or reduced length of coleoptile, and 50% of necrotic symptoms. Pseudopithomyces chartarum isolates produced secondary metabolites including alternariol, alternariol mono-methyl ether, altertoxin I and altertoxine II. The fungus is a new pathogen of wheat in Argentina that can cause diseases on different cultivars as well as produce mycotoxins.