Efecto de diferentes medios de cultivo sobre el desarrollo de Phytophthora nicotianae Breda de Haan

El objetivo del estudio fue determinar el efecto de diferentes medios de cultivo sobre el crecimiento, esporulación y secreción de enzimas hidrolíticas de Phytophthora nicotianae Breda de Haan. Para ello se emplearon los medios en estado sólido y líquido: Papa Dextrosa Agar (PDA), jugo de verduras (V8) y extracto de tabaco, combinados entre ellos y con suplemento de asparagina y carbonato de calcio. El mayor crecimiento del microorganismo se produjo en medio sólido con extracto de tabaco, pues alcanzó en menor tiempo el extremo de la placa; sin embargo, su crecimiento fue poco ramificado, a diferencia de los que contenían V8. Se comprobó que la biomasa producida fue mayor en los medios líquidos que contenían V8. El número de zoosporas estimadas en el momento en que el micelio alcanzó el borde de la placa fue significativamente mayor en el tratamiento que combinaba PDA con V8 y el menor valor se obtuvo al emplearse el extracto de tabaco. Al utilizarse la combinación del extracto de tabaco con V8, se obtuvieron las mayores actividades poligalacturonasa y xilanasa. Los resultados evidenciaron que los medios que favorecen la expresión de enzimas hidrolíticas, así como el crecimiento y esporulación de P. nicotianae, tienen V8 entre sus componentes, elemento de gran importancia, pues se disminuye el tiempo requerido para los ensayos y con ello el riesgo de contaminación de los experimentos. Palabras clave: Phytophthora nicotianae, medio de cultivo, crecimiento, esporulación, actividad enzimática.EFFECT OF DIFFERENT CULTURE MEDIA ON PHYTOPHTHORA NICOTIANAE BREDA DE HAAN DEVELOPABSTRACT: The effect of different culture media on Phytophthora nicotianae growth, sporulation, and hydrolytic enzymes secretion was studied. The liquid and solid culture media used were potato dextrose agar (PDA), vegetable juice (V8), and tobacco extract. They were combined one another and supplemented with asparagine and calcium carbonate. Phytophthora nicotianae growing on solid media containing only tobacco extract reached the plate edge in the shortest time; however, the mycelium was little branched, unlike the mycelium in the media containing V8. A greater biomass production was observed in liquid culture with V8. The zoospore number was significantly higher in the treatment combining PDA only with V8; the lowest number was obtained with tobacco extract. Polygalacturonase and xylanase activities were the highest when the liquid media contained the combination of tobacco extract and V8 juice. The culture medium V8 was shown to be useful to determine the enzymatic activity of the hydrolytic enzymes and the parameters of growth and sporulation of P. nicotianae; itreduced the time required for the trials and, consequently, the risk of contamination.Key words: Phytophthora nicotianae, culture media, growth, sporulation, enzymatic activity

Actividad inhibitoria de un polímero de quitosana en el crecimiento vegetativo y la reproducción asexual de un aislado de Phytophthora palmivora Butler

The aim was to evaluate the effect of chitosan on the vegetative growth and the asexual reproductive stages of the life cycle of Phytophthora palmivora Butler. Solutions of different chitosan concentrations, ranging from 0.5 to 2.5 g.l-1, were mixed with V8 culture media prior to add into Petri dishes, where a mycelial disc of the isolate was then inoculated. Morphocultural aspects and radial growth were evaluated when colonies from the control treatment reached the edge of the plates. The colony diameter of the isolate decreased with increases of chitosan levels in the culture medium, and the colony morphocultural pattern was altered showing irregular edges and sandy appearance. However, the hypha diameters observed under an optical microscope did not differ from the control treatment. Low polymer concentrations (0.5 g.l-1) stimulated sporangium production, while this was reduced by increases of chitosan levels. Sporangium length/width ratio was not affected by the compound concentration. In addition, the indirect sporangium germination was inhibited at 1g.l-1, and an aberrated zoospores movement was observed, with less speed at 0.5 g.l-1 of the polymer. Moreover, the cyst germination process was stopped after 3 hours of incubation with 0.5 g.l-1 of chitosan. Key words: Phytophthora palmivora, chitosan, sporulation.El objetivo del trabajo fue evaluar el efecto de la quitosana sobre el crecimiento vegetativo y las fases de reproducción asexual del ciclo de vida de Phytophthora palmivora Butler. Soluciones con diferentes concentraciones de quitosana, que oscilaron desde 0,5 hasta 2,5g.l-1 se mezclaron con medio de cultivo V8 antes de verterse en placas Petri, donde luego se inoculó un disco de micelio del aislado. Se evaluaron aspectos morfoculturales y el crecimiento radial cuando las colonias del tratamiento control alcanzaron el extremo de la placa. El diámetro de las colonias del aislado disminuyó con el aumento de los niveles de quitosana en el medio y se alteró el patrón morfocultural de las mismas: exhibieron bordes irregulares y de aspecto arenoso. Sin embargo, los diámetros de las hifas observadas al microscopio óptico no variaron con relación al tratamiento control. Las concentraciones bajas del polímero (0,5 g.l-1) estimularon la formación de esporangios, mientras que el incremento de los niveles de quitosana redujo estos valores. La relación largo-ancho de los esporangios no se afectó por la concentración del compuesto. La germinación indirecta de los esporangios se inhibió con 1,0g.l-1 y se observó un movimiento aberrado de las zoosporas, pues se desplazaban con menor velocidad en presencia de 0,5g.l-1 del polímero. Además, se detuvo el proceso de germinación de los quistes después de haber sido incubados durante 3 horas con 0,5g.l-1 de quitosana. Palabras clave: Phytophthora palmivora, quitosana, esporulación

Analysis of MAPK and MAPKK gene families in wheat and related Triticeae species

Background: The mitogen-activated protein kinase (MAPK) family is involved in signal transduction networks that underpin many different biological processes in plants, ranging from development to biotic and abiotic stress responses. To date this class of enzymes has received little attention in Triticeae species, which include important cereal crops (wheat, barley, rye and triticale) that represent over 20% of the total protein food-source worldwide. Results: The work presented here focuses on two subfamilies of Triticeae MAPKs, the MAP kinases (MPKs), and the MAPK kinases (MKKs) whose members phosphorylate the MPKs. In silico analysis of multiple Triticeae sequence databases led to the identification of 152 MAPKs belonging to these two sub-families. Some previously identified MAPKs were renamed to reflect the literature consensus on MAPK nomenclature. Two novel MPKs, MPK24 and MPK25, have been identified, including the first example of a plant MPK carrying the TGY activation loop sequence common to mammalian p38 MPKs. An EF-hand calcium-binding domain was found in members of the Triticeae MPK17 clade, a feature that appears to be specific to Triticeae species. New insights into the novel MEY activation loop identified in MPK11s are offered. When the exon-intron patterns for some MPKs and MKKs of wheat, barley and ancestors of wheat were assembled based on transcript data in GenBank, they showed deviations from the same sequence predicted in Ensembl. The functional relevance of MAPKs as derived from patterns of gene expression, MPK activation and MKK-MPK interaction is discussed. Conclusions: A comprehensive resource of accurately annotated and curated Triticeae MPK and MKK sequences has been created for wheat, barley, rye, triticale, and two ancestral wheat species, goat grass and red wild einkorn. The work we present here offers a central information resource that will resolve existing confusion in the literature and sustain expansion of MAPK research in the crucial Triticeae grains.Other UBCNon UBCReviewedFacult

Additional file 1: of Analysis of MAPK and MAPKK gene families in wheat and related Triticeae species

Number of MAPK publications in Brachypodium and Triticeae since 1996. A quick survey of the literature shows the number of publications focused on MAPKs in Brachypodium, wheat and barley is presented. To our knowledge, there are no publications on rye or triticale MAPKs. This survey included MKKKs; however, due to the large representation of this divergent gene family in plants, some publications on MKKKs may have been overlooked. (PDF 118 kb

Additional file 7: of Analysis of MAPK and MAPKK gene families in wheat and related Triticeae species

Alignment and domain analysis of Triticeae MPKs. The amino acid sequences from multiple genomic copies, along with the accessions reported in Additional file 3, of all Triticeae MPKs identified were aligned in Clustal Omega (www.ebi.ac.uk/Tools/msa/clustalo/) under default settings. A lower case ‘p’ next to the gene name indicates that only a partial sequence was identified for the given gene. The PK domain is in black font, and the ‘MAP kinase’ signature (specific to MPKs) is bolded where ‘MAP kinase’ flanks (F and C) are in red font; N- and C-terminal extensions in teal font. Missing sequence information, namely ‘X’ residues, are shown in pink font. Highlighted sequences are the ATP binding signature (blue), the catalytic C-loop (red), the activation T-loop (black), CD domain (teal) and EF-hand CBP (pink). Sequence deviations from the T(E/D)Y activation loop of plant MPKs are marked in yellow font. Previously reported conserved MPK domains [18] are written in blue font above the alignment, including the conserved clade-specific docking domain found at the C-terminal end of the PK signature (highlighted grey; deviations from proposed sequence in a darker shade). Deviations from these sequences are highlighted in yellow; some of these deviations were observed in the ATP binding signature and are highlighted in a bright blue. Additional conserved sequences described as putative phosphorylation sites distinct from the T-loop [18] are marked with a box. An asterix indicates a stop codon. (PDF 230 kb

Additional file 10: of Analysis of MAPK and MAPKK gene families in wheat and related Triticeae species

Summary of MAPK identifiers and complementary information obtained from eight studies. The table includes gene identifiers and complementary annotations for 8 published sets of MAPKs that were used for comparison in this work. (XLSX 53 kb

Additional file 5: of Analysis of MAPK and MAPKK gene families in wheat and related Triticeae species

Genomic sequence of Triticeae MKKs. Genomic DNA sequences were obtained from Ensembl. Font colour of predicted genes differentiate UTRs (orange), exons (blue), introns (grey), sequences upstream/downstream of the predicted gene (green). Where Ensembl predictions for exon-intron structure did not match available transcript data, alternative exons are proposed and highlighted in yellow; corresponding accessions have an ‘X’ suffix. Black nucleotides are used for genomic sequence for which Ensembl did not predict a gene sequence or for predicted ncRNA sequences, where proposed exon sequences are highlighted in yellow. The underlined nucleotides highlighted in various colours indicate nucleotide variants as defined by Ensembl. Purple font is used for GenBank sequence that were used to fill in the gaps where complete gene sequence was not identified from Ensembl. (DOCX 274 kb