[Morphologic and molecular characterization of Phomopsis longicolla(teleomorph unknown: Diaporthales) from tempered and subtropical regions of Argentina]. / Caracterización morfológica y molecular de una colección de aislamientos de Phomopsis longicolla (teleomorfo desconocido: Diaporthales) de la región templada y subtropical de Argentina.

Diaporthe (teleomorpho)-Phomopsis - (anamorph) (DP) is a fungal group of great genetic diversity with over 900 species associated to a wide host range that includes cultivated and uncultivated species, forest, fruit trees and weeds. DP isolates are hemibiotrophs and have different sources of primary inoculum as stubble and seeds to restart cycles of parasitism - saprophytism. They colonize host tissues from early plant stages and establish different nutritional relationships, acting as endophytic and necrotrophic fungi. The plasticity of the Phomopsis genus has favored its expansion to different agro-ecosystems and various hosts constituting an epidemiological risk. The objective was to validate the identity and evaluate the biological relationships among 12 isolates of P longicolla and D. phaseolorum var. sojae (anamorph P phaseoli var. sojae) obtained in different tempered and subtropical agro-environments of Argentina, in order to analyze the variability and strategies for preserving fungal biodiversity. Macromorphological attributes (such as texture and color of colonies, stroma shape and distribution, pycnidia and perythecia shape and distribution) and micro-morphological characteristics (such as size and shape of conidia, asci and ascospores) allowed identifying three new isolates as P longicolla. A complementary molecular analysis was also made to overcome the limitations derived from the morphological analysis, thus the AFP.8413 isolate was finally identified as P longicolla. The molecular characterization was useful to identify the evaluated isolates and to group them in four taxa of the Diaporthe-Phomopsis complex: ten isolates were included in P. longicolla, one isolate was included in D. phaseolorum var. sojae (anamorph P. phaseoli var. sojae), one isolate was identified as D. phaseolorum var. caulivora and two isolates were included in D. phaseolorum var. meridionalis. The use of phenotipic and molecular tools have contributed to an accurate identification of P longicolla, and comprehension about the biological relationships (homo or heterothallic hibridizations) among D. phaseolorum varieties (P phaseoli) and species of Diaporthe-Phomopsis. This allowed also a better understanding of the mechanisms of fungic plasticity, to colonize and expand their host range and genetic variability, promoting thus their biodiversity conservation. Rev. Biol. Trop. 63 (3): 871-884. Epub 2015 September 01.

Caracterización morfológica y molecular de una colección de aislamientos de Phomopsis longicolla(teleomorfo desconocido: Diaporthales)de la región templada y subtropical de Argentina

Diaporthe(telomorfo) - Phomopsis(anamorfo) (DP) constituye un grupo fúngico de amplia diversidad genética con más de 900 especies distribuidas en un amplio rango de hospedantes que incluye especies cultivadas y no cultivadas, forestales, frutales y malezas. Los aislamientos de DP son hemi-biótrofos y disponen de diferentes fuentes de inóculo primario, como el rastrojo y las semillas, para reiniciar sus ciclos de parasitismo-saprofitismo. Ellos colonizan los tejidos del hospedante desde los estadios tempranos del desarrollo y establecen relaciones nutricionales de endofitia y necrotrofia fúngica. La plasticidad del género Phomopsisha favorecido su expansión a diferentes agro-ecosistemas y hospedantes constituyendo un importante riesgo epidemiológico. El objetivo fue validar la identidad y evaluar las relaciones biológicas de 12 aislamientos de P. longicollay D. phaseolorumvar. sojaeobtenidos en distintos agro-ambientes templados y subtropicales de Argentina, para analizar la variabilidad y estrategias de conservación de la bio-diversidad fúngica. Las cualidades macro-morfo-lógicas(textura y color de colonias, forma y distribución de estromas, desarrollo, forma y distribución de cuerpos fructíferos), y los caracteres micro-morfológicos(tamaño y forma de conidios, ascos y ascosporas) permitieron identificar a tres nuevos aislamientos de P. longicollaincluidos en el complejo D/P. El análisis molecular complementario corrigió las limitaciones derivadas de la caracterización basada sólo en marcadores morfológicos y logró reubicar al aislamiento AFP.8413 de identidad dudosa, en el nodo correspondiente a P. longicolla.De esta manera, la caracterización molecular definió la identidad de los aislamientos y los ubicó en los 4 taxones del complejo DP: diez aislamientos fueron incluidos en Plo(AFP.Gpo 4.4, AFP.Gpo 3.5, AFP.Gpo 4.3, AFP.Gpo 3.4, AFP.CaA, AFP. CaB, AFP.B5L16, AFP.B4L17, AFP.227B2, AFP.8413), un aislamiento incluido en Dps(AFP.Qcol7), un aislamiento en Dpc(AFP.Dpc16) y dos aislamientos incluidos en Dpm(AFP.Dpm109 y AFP.Dpm112). La adecuada identificación de P. longicollay el avance en el conocimiento de las relaciones biológicas (hibridaciones homo o heterotálicas) entre variedades de D. phaseolorum (P. phaseoli)y especies de Diaporthe- Phomopsispermiten comprender la plasticidad para colonizar un amplio rango de hospedantes, los mecanismos de variabilidad genética y la preservación de la diversidad fúngica.

Diaporthe(teleomorpho)-Phomopsis (anamorph) (DP) is a fungal group of great genetic diversity with over 900 species associated to a wide host range that includes cultivated and uncultivated species, forest, fruit trees and weeds. DP isolates are hemi-biotrophs and have different sources of primary inoculum as stubble and seeds to restart cycles of parasitism -saprophytism. They colonize host tissues from early plant stages and establish different nutritional relationships, acting as endophytic and necrotrophic fungi. The plasticity of the Phomopsisgenus has favored its expansion to different agro-ecosystems and various hosts constituting an epidemiological risk. The objective was to validate the identity and evaluate the biological relationships among 12 isolates of P. longicollaand D. phaseolorumvar. sojae(anamorph P. phaseolivar. sojae)obtained in different tempered and subtropical agro-environments of Argentina, in order to analyze the variability and strategies for preserving fungal biodiversity. Macro-morphological attributes (such as texture and color of colonies, stroma shape and distribution, pycnidia and perythecia shape and distribution) and micro-morphological characteristics (such as size and shape of conidia, asci and ascospores) allowed identifying three new isolates as P. longicolla.A complementary molecular analysis was also made to overcome the limitations derived from the morphological analysis, thus the AFP.8413 isolate was finally identified as P. longicolla.The molecular characterization was useful to identify the evaluated isolates and to group them in four taxa of the Diaporthe-Phomopsiscomplex: ten isolates were included in P. longicolla,one isolate was included in D. phaseolorumvar. sojae(anamorph P. phaseolivar. sojae),one isolate was identified as D. phaseolorumvar. caulivoraand two isolates were included in D. phaseolorumvar. meridi-onalis.The use of phenotipic and molecular tools have contributed to an accurate identification of P. longicolla,and comprehension about the biological relationships (homo or heterothallic hibridizations) among D. phaseolo-rumvarieties (P. phaseoli)and species of Diaporthe-Phomopsis.This allowed also a better understanding of the mechanisms of fungic plasticity, to colonize and expand their host range and genetic variability, promoting thus their biodiversity conservation.

Soybean sudden death syndrome species diversity within north and South america revealed by multilocus genotyping.

Sudden death syndrome (SDS) of soybean has become a serious constraint to the production of this crop in North and South America. Phenotypic and multilocus molecular phylogenetic analyses, as well as pathogenicity experiments, have demonstrated that four morphologically and phylogenetically distinct fusaria can induce soybean SDS. Published molecular diagnostic assays for the detection and identification of these pathogens have reported these pathogens as F. solani, F. solani f. sp. glycines, or F. solani f. sp. phaseoli, primarily because the species limits of these four pathogens were only recently resolved. In light of the recent discovery that soybean SDS and Phaseolus and mung bean root rot (BRR) are caused by four and two distinct species, respectively, multilocus DNA sequence analyses were conducted to assess whether any of the published molecular diagnostic assays were species-specific. Comparative DNA sequence analyses of the soybean SDS and BRR pathogens revealed that highly conserved regions of three loci were used in the design of these assays, and therefore none were species-specific based on our current understanding of species limits within the SDS-BRR clade. Prompted by this finding, we developed a high-throughput multilocus genotyping (MLGT) assay which accurately differentiated the soybean SDS and two closely related Phaseolus and mung BRR pathogens based on nucleotide polymorphism within the nuclear ribosomal intergenic spacer region rDNA and two anonymous intergenic regions designated locus 51 and 96. The single-well diagnostic assay, employing flow cytometry and a novel fluorescent microsphere array, was validated by independent multilocus molecular phylogenetic analysis of a 65 isolate design panel. The MLGT assay was used to reproducibly type a total of 262 soybean SDS and 9 BRR pathogens. The validated MLGT array provides a unique molecular diagnostic for the accurate identification and molecular surveillance of these economically important plant pathogens.

Morphologic, Molecular, and Pathogenic Characterization of Diaporthe phaseolorum Variability in the Core Soybean-Producing Area of Argentina.

ABSTRACT Isolates of the Diaporthe/Phomopsis (D/P) complex were collected in the main soybean producing area of Argentina during the 1996-97, 1997-98, and 1998-99 growing seasons. Twenty-three morphologic characters related to type of colonies, stroma, pycnidia and conidia, presence of perithecia, and asci length were studied by principal component analysis (PCA). Genomic DNA were analyzed by the random amplified polymorphic DNA (RAPD) technique. From both studies, 18 isolates were identified as D/P complex and grouped in four major taxa: (i) Diaporthe phaseolorum var. meridionalis, (ii) D. phaseolorum var. caulivora, (iii) D. phaseolorum var. sojae, and (iv) Phomopsis longicolla. In addition to distinguishing interspecific and intraspecific variability, molecular markers allowed the detection of differences among isolates within the same variety. Pathogenicity was assayed in the greenhouse, by the toothpick method, inoculating the D/P isolates to soybean genotypes carrying different resistance genes (Rdc1, Rdc2, Rdc3, and Rdc4) against soybean stem canker (SSC). Pathogenic analysis distinguished two main groups: (i) the SSC-producing isolates, including D. phaseolorum var. meridionalis and D. phaseolorum var. caulivora, and (ii) the non-SSC-producing isolates, including D. phaseolorum var. sojae and P. longicolla. Cultivar RA-702 (susceptible control) was compatible with both D. phaseolorum var. meridionalis and D. phaseolorum var. caulivora isolates; meanwhile, Tracy-M (Rdc1 and Rdc 2 genes) was incompatible with D. phaseolorum var. meridionalis but compatible with D. phaseolorum var. caulivora isolates. The fact that Rdc1 and Rdc2 together (as in Tracy-M) confer an almost immune reaction to all assayed isolates of D. phaseolorum var. meridionalis but were ineffective against the D. phaseolorum var. caulivora isolates evaluated suggests that the virulence or avirulence genes in D. phaseolorum var. meridionalis and D. phaseolorum var. caulivora are different. Moreover, physiological races of D. phaseolorum var. meridionalis were detected by using differential soybean genotypes carrying distinct single Rdc genes. As far as we know, this is the first report on the existence of physiological races of D. phaseolorum var. meridionalis in South America. Selective pressure due to deployment of resistant host cultivars may have changed the frequency of the virulence or avirulence genes within the population of D. phaseolorum var. meridionalis. On the whole, our results show that pathogenic variability of D. phaseolorum in the core soybean-producing area of Argentina is higher than previously recognized.

Hongos celulolíticos asociados al rastrojo de trigo y su relación con algunas variables ambientales / Cellulolytic fungi associated to stubble of wheat and its relationship with some environmental variables

Se cuantificó e identificó la población de hongos celulolíticos, asociados al rastrojo de trigo, relacionándolos con algunas variables ambientales. En parcelas bajo siembra directa del grano, incluidas en un diseño en bloques aleatorizados con tres repeticiones, los restos de cosecha de trigo obtenidos de 1 m(2) se cortaron y distribuyeron en bolsas de malla. Mensualmente se procedió a determinar la pérdida de peso del rastrojo y a registrar las variables agroclimáticas. Se cuantificó e identificó la población fúngica celulolítica por el método de dilución en placas en medio selectivo. Por análisis de regresión múltiple, se examinó la relación entre las variables ambientales y el peso del rastrojo con la micota celulolítica (r(2)= 0,95). Por análisis stepwise, la temperatura edáfica, la variación de la humedad relativa y las precipitaciones fueron las variables más explicativas. Los hongos celulolíticos más aislados fueron especies de penicillium y fusarium. En este último género se destaca la presencia de f. solani y f. graminearum, potenciales patógenos de cultivos vegetales