First report of Leptosphaeria biglobosa ‘brassicae’ as causal agent of phoma leaf spot in Brassica napus (Canola) in Argentina

Canola(Brassicanapus L.) is the second largest oilseed crop in the worldproviding 13% of the world´s oil supply. This crop has been grown in Argentinasince the 1930s, and the area devoted to its cultivation varies every year,reaching a maximum of 95000 Ha in the 2012/2013 growing season. Because of theoccurrence of optimal weather conditions and soils with high fertility, theaverage yield in this region is about 2000 kg/Ha. Phoma leaf spot and Phomastem canker are considered the most important and devastating diseases in Brassica napus andother Brassicaespecies [1]. In both cases, the causal agent is a complexof two closely related fungal species, Leptosphaeria maculans and L. biglobosa. In Argentina,the presence of L. maculans incanola plants was reported for the first time in 2004 [2], but the existence ofL.biglobosa has not been recorded so far. During the 2015/2016season, we collected several samples with typical Phoma leaf spot symptoms fromcanola plants growing in fields from the north and northeastern regions of the Buenos Aires province.The necrotic lesions were circular to irregularly oval, 8 to 15 mm in diameter,pale brown in the center, grayish green at the margin and characterized withthe presence of pycnidia. Several leaf pieces with lesions were rinsed twicewith deionized sterile water and placed in a humid chamber (90 mm diameterPetri dish with a layer of filter paper soaked in deionized sterile water) during2-3 days to induce the pycnidia to exude cirri of conidia. After this period,one cirrus per sample was transferred onto PDA plates supplemented withantibiotics (15 mg/L streptomycin, 15 mg/L gentamicin and 12 mg/L tetracycline)using an inoculation needle under stereoscopic microscope. Thus, severalisolates were obtained, some of them showing rapid mycelial growth rate andpigment production on PDA medium, as showed by the isolate Tapidor of L. biglobosa thatwe used as control (kindly provided by Professor Bruce Fitt, University ofHertfordshire-UK). In order to confirm the identity of these isolates, a PCRassay using genomic DNA as template was performed to distinguish L. maculans from L. biglobosa withthe species-specific primers LmacR, LmacF,and LbigF ina three-primers strategy described by Gaetan (2005)[3]. These reactions gave a444-bp amplicon as expected for L. biglobosa ´brassicae´.In addition, these results were confirmed by sequencing the nuclear ribosomalinternal transcribed spacer (ITS) region, which showed a 99% of identity withthe sequence of L. biglobosa ´brassicae´at the GenBank database (FO905468). L. biglobosa isolateswere then tested for pathogenicity on the canola cultivars Westar and Bioaureo2286 (Nuseed). With this purpose, cotyledons of 10-day-old seedlings werepricked with a needle, and each wound inoculated with 10 μl ofa conidial suspension (107 42conidia/ml). Sterilized distilled water was used as control. Developing primaryleaves were removed every 2-3 days in order to ensure that cotyledons continueto expand. Fourteen days after inoculation, irregular and brown necroticlesions were visible at the site of inoculation. These cotyledons were detachedand placed in a humid chamber to induce pycnidia formation. After three dayscirri of conidia were transferred to a plate with PDA supplemented with antibioticsas mentioned above. The identity of these isolates of L. biglobosa wereconfirmed by pigment production on PDA medium and by PCR assay usingspecies-specific primers. To our knowledge, this is the first report of L. biglobosa ´brassicae´as a pathogen of canola in Argentina. This finding shows that in Argentina´scanola cropping areas not 50 only L. maculans but alsoL.biglobosa are the causal agents of Phoma leaf spot disease.Fil: Rossi, Franco Rubén. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); ArgentinaFil: Romero, Fernando Matias. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); ArgentinaFil: Garriz, Andrés. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); ArgentinaFil: Ruiz, Oscar Adolfo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); Argentina. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigaciones Agropecuarias. Instituto de Fisiología y Recursos Genéticos Vegetales; Argentin

Isolation of high quality RNA from soil-grown Ilex paraguariensis roots suitable for next-generation sequencing and gene expression analyses

Extraction of high quality RNA is a prerequisite for downstream application in functional genomics analyses. However, the extraction and purification of pure nucleic acids from root tissues is generally difficult due to the high concentration of carbohydrates and secondary metabolites. Furthermore, the presence of enzymatic inhibitors such as fulvic and humic acids can also negatively affect extraction quality, when extracting from clay soil-grown roots. In this work, total RNA was extracted from soil-grown roots of Ilex paraguariensis using four commercially available kits: SpectrumTM, RNeasy®, TRI Reagent®, and SV Total RNA Isolation System. Spectral measurement and electrophoresis were used to demonstrate RNA quality and quantity. The SpectrumTM and RNeasy® protocols provided the highest quantity and quality of RNA; however, the former revealed superior extraction performance. Consequently, total RNA was extracted from the roots of non-stressed and drought-stressed plants using the SpectrumTM method and six RNA-seq libraries were prepared from polyA + mRNAs by means of TruSeq mRNA library construction protocol to convert RNA to complementary DNA (cDNA). More than 80 million raw read sequences were obtained from each condition with an average read length of 150 bp. The yield and quality of the total RNA were consistently high and the RNA could be used for further analyses as demonstrated by cDNA library construction, RT-PCR, and transcriptome sequencing. Thus, SpectrumTM method can be used to isolate high quality RNA from roots of normal and drought stressed I. paraguariensis plants.Fil: Avico, Edgardo Hernán. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Nordeste. Instituto de Botánica del Nordeste. Universidad Nacional del Nordeste. Facultad de Ciencias Agrarias. Instituto de Botánica del Nordeste; ArgentinaFil: Acevedo, Raúl Maximiliano. Universidad Nacional del Nordeste. Facultad de Ciencias Agrarias. Departamento de Física y Química; ArgentinaFil: Calzadilla, Pablo Ignacio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas; ArgentinaFil: Ruiz, Oscar Adolfo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas; ArgentinaFil: Sansberro, Pedro Alfonso. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Nordeste. Instituto de Botánica del Nordeste. Universidad Nacional del Nordeste. Facultad de Ciencias Agrarias. Instituto de Botánica del Nordeste; Argentin

Interspecific hybridization improves the performance of Lotus spp. under saline stress

Salinity is one of the most frequent limiting conditions in pasture production for grazing livestock. Legumes, such as Lotus spp. with high forage quality and capable of adapting to different environments, improves pasture performance in restrictive areas. In order to determine potential cultivars with better forage traits, the current study assess the response to salt stress of L. tenuis, L. corniculatus and a novel L. tenuis x L. corniculatus accession. For this purpose, chlorophyll fluorescence, biomass production, ion accumulation and anthocyanins and proanthocyanidins levels have been evaluated in control and salt-treated plants PSII activity was affected by salt in L. tenuis, but not in L. corniculatus or hybrid plants. Analyzed accessions showed similar values of biomass, Na+ and K+ levels after salt treatment. Increasing Cl− concentrations were observed in all accessions. However, hybrid plants accumulate Cl− in stems at higher levels than their parental. At the same time, the levels of anthocyanins considerably increased in L. tenuis x L. corniculatus stems. Chloride and anthocyanin accumulation in stems could explain the best performance of hybrid plants after a long saline treatment. Finally, as proanthocyanidins levels were no affected by salt, L. tenuis x L. corniculatus plants maintained adequate levels to be used as ruminant feed. In conclusion, these results suggest that hybrid plants have a high potential to be used as forage on salt-affected lands. High Cl− and anthocyanins accumulation in Lotus spp. stems seems to be a trait associated to salinity tolerance, with the possibility of being used in legume breeding programs.Instituto de Fisiología y Recursos Genéticos VegetalesFil: Escaray, Francisco José. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); ArgentinaFil: Antonelli, Cristian Javier. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); ArgentinaFil: Carrasco, Pedro. Universitat de València. Departament de Bioquìmica y Biologia Molecular; EspañaFil: Ruiz, Oscar Adolfo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); Argentina. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Fisiología y Recursos Genéticos Vegetales; Argentin

Assessment of reference genes for real-time quantitative PCR normalization in Ilex paraguariensis leaves during drought

Reverse transcription of RNA followed by real-time quantitative PCR (qPCR) is to date the most reliable method for gene expression studies. However, to control the errors introduced along the numerous experimental procedures, it requires a normalization using internal reference genes with stable expression. To address this issue, nine candidate reference genes were investigated in Ilex paraguariensis leaves subjected to water stress. To facilitate the selection, we analysed the real-time qPCR data with three different software programs. The obtained results support the conclusion that RNA polymerase associated protein rtf1 homolog (RTF) combined with any of the following pairs is the most suitable triad of genes to compute a normalization factor: elongation factor 1-alpha + tubulin alpha chain (EF1a + α-Tub), actin + cyclophilin 38 (ACT + CYP38), or cyclophilin 38 + vacuolar protein sorting-associated protein 18 homologs (CYP38 + VPS). Our analysis constitutes the first in-depth study to identify the appropriate reference genes for the quantification of transcription in Ilex paraguariensis leaves during drought and provides essential information for further gene expression studies in this tree species.Fil: Acevedo, Raúl Maximiliano. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Nordeste. Instituto de Botánica del Nordeste. Universidad Nacional del Nordeste. Facultad de Ciencias Agrarias. Instituto de Botánica del Nordeste; ArgentinaFil: Avico, Edgardo Hernán. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Nordeste. Instituto de Botánica del Nordeste. Universidad Nacional del Nordeste. Facultad de Ciencias Agrarias. Instituto de Botánica del Nordeste; ArgentinaFil: Ruiz, Oscar Adolfo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); ArgentinaFil: Sansberro, Pedro Alfonso. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Nordeste. Instituto de Botánica del Nordeste. Universidad Nacional del Nordeste. Facultad de Ciencias Agrarias. Instituto de Botánica del Nordeste; Argentin

Lotus tenuis x L. corniculatus interspecific hybridization as a means to breed bloat-safe pastures and gain insight into the genetic control of proanthocyanidin biosynthesis in legumes

Background: Proanthocyanidins (PAs) are secondary metabolites that strongly affect plant quality traits. The concentration and the structure of these metabolites influence the palatability and nutritional value of forage legumes. Hence, modulating PAs in the leaves of forage legumes is of paramount relevance for forage breeders worldwide. The lack of genetic variation in the leaf PA trait within the most important forage species and the difficulties in engineering this pathway via the ectopic expression of regulatory genes, prompted us to pursue alternative strategies to enhance this trait in forage legumes of agronomic interest. The Lotus genus includes forage species which accumulate PAs in edible organs and can thus be used as potential donor parents in breeding programs. Results: We recovered a wild, diploid and PA-rich population of L. corniculatus and crossed with L. tenuis. The former grows in an alkaline-salty area in Spain while the latter is a diploid species, grown extensively in South American pastures, which does not accumulate PAs in the herbage. The resulting interspecific hybrids displayed several traits of outstanding agronomic relevance such as rhizome production, PA levels in edible tissues sufficient to prevent ruminal bloating (around 5 mg of PAs/g DW), biomass production similar to the cultivated parent and potential for adaptability to marginal lands. We show that PA levels correlate with expression levels of the R2R3MYB transcription factor TT2 and, in turn, with those of the key structural genes of the epicatechin and catechin biosynthetic pathways leading to PA biosynthesis. Conclusions: The L. tenuis x L. corniculatus hybrids, reported herein, represent the first example of the introgression of the PA trait in forage legumes to levels known to provide nutritional and health benefits to ruminants. Apart from PAs, the hybrids have additional traits which may prove useful to breed forage legumes with increased persistence and adaptability to marginal conditions. Finally, our study suggests the hybrids and their progeny are an invaluable tool to gain a leap forward in our understanding of the genetic control of PA biosynthesis and tolerance to stresses in legumes.Fil: Escaray, Francisco José. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); ArgentinaFil: Passeri, Valentina. Institute of Biosciences and BioResources; Italia. Consiglio Nazionale delle Ricerche; ItaliaFil: Babuin, María Florencia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); ArgentinaFil: Marco, Francisco. Universidad de Valencia; EspañaFil: Carrasco, Pedro. Universidad de Valencia; EspañaFil: Damiani, Francesco. Consiglio Nazionale delle Ricerche; Italia. Institute of Biosciences and BioResources; ItaliaFil: Pieckenstain, Fernando Luis. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); ArgentinaFil: Paolocci, Francesco. Consiglio Nazionale delle Ricerche; Italia. Institute of Biosciences and BioResources; ItaliaFil: Ruiz, Oscar Adolfo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); Argentin

Glomus intraradices improved salt tolerance in Prosopis alba seedlings by improving water use efficiency and shoot water content.

The present work was aimed at testing the hypothesis that mycorrhizal Prosopis alba, an economically impor tant tree species worldwide, presents increased salt-tolerance compared with non-mycorrhizal ones and at gaining insight into the possible mechanisms underlying that improvement. For this purpose, a randomized complete block experiment with two factors: mycorrhizal treatments with or without the arbuscular fungus Glomus intraradices and two salinity levels, 0 and 200 mM NaCl was performed. Plant growth in P. alba plants colonized by G. intraradices was less affected by salinity than that in non-arbuscular mycorrhizal (AM) plants, indicating that mycorrhizal colonization turned P. alba more tolerant to salinity. Photosynthesis was reduced by salinity in non-AM plants but not in AM ones.  Salini  ty caused a significant decrease in mean stomatal conductance and transpiration rate, in mycorrhizal plants, but not in uninoculated ones. In this work, we detected two main mechanisms intervening in the salt tolerance enhancement of P. alba by the inoculation with G. intraradices: a- maintaining the net photosynthesis level and b- control of the transpiration rate.  Taken together, the results suggest that inoculation with G. intraradices improves P. alba survival rates during the implantation period and seems to be a promising strategy to improve P. alba cultivation in saline lands.Fil: Scambato, Agustina Azul. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico la Plata. Instituto de Investigaciones Biotecnológicas - Instituto Tecnológico Chascomús. Instituto de Investigaciones Biotecnológicas (sede Chascomús); Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Biodiversidad y Biología Experimental; ArgentinaFil: Echeverria, Mariela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico La Plata. Instituto de Investigaciones Biotecnológicas - Instituto Tecnológico Chascomús. Instituto de Investigaciones Biotecnológicas (sede Chascomús); ArgentinaFil: Sansberro, Pedro Alfonso. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Nordeste. Instituto de Botánica del Nordeste (i); ArgentinaFil: Ruiz, Oscar Adolfo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico La Plata. Instituto de Investigaciones Biotecnológicas - Instituto Tecnológico Chascomús. Instituto de Investigaciones Biotecnológicas (sede Chascomús); ArgentinaFil: Menendez, Ana Bernardina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico La Plata. Instituto de Investigaciones Biotecnológicas - Instituto Tecnológico Chascomús. Instituto de Investigaciones Biotecnológicas (sede Chascomús); Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Biodiversidad y Biología Experimental; Argentin

Multiple bHLH/MYB-based protein complexes regulate proanthocyanidin biosynthesis in the herbage of Lotus spp.

[EN] The environmental impact and health of ruminants fed with forage legumes depend on the herbage's concentration and structure of proanthocyanidins (PAs). Unfortunately, the primary forage legumes (alfalfa and clover) do not contain substantial levels of PAs. No significant progress has been made to induce PAs to agronomically valuable levels in their edible organs by biotechnological approaches thus far. Building this trait requires a profound knowledge of PA regulators and their interplay in species naturally committed to accumulating these metabolites in the target organs. Against this background, we compared the shoot transcriptomes of two inter-fertile Lotus species, namely Lotus tenuis and Lotus corniculatus, polymorphic for this trait, to search for differentially expressed MYB and bHLH genes. We then tested the expression of the above-reported regulators in L. tenuis x L. corniculatus interspecific hybrids, several Lotus spp., and different L. corniculatus organs with contrasting PA levels. We identified a novel MYB activator and MYB-bHLH-based complexes that, when expressed in Nicotiana benthamiana, trans-activated the promoters of L. corniculatus anthocyanidin reductase and leucoanthocyanidin reductase 1 genes. The last are the two critical structural genes for the biosynthesis of PAs in Lotus spp. Competition between MYB activators for the transactivation of these promoters also emerged. Overall, by employing Lotus as a model genus, we refined the transcriptional network underlying PA biosynthesis in the herbage of legumes. These findings are crucial to engineering this trait in pasture legumes.This study was partially supported by CNR (Italy)-CONICET (Argentina) 2021-2022 Bilateral Agreement, by the project Pict 2021-2023 from Agencia de Promocion Cientifica y Tecnologica (Argentina), by Margin Up, funded by the European Union's Horizon research and innovation program GA number no 101082089, and by the Italian Agritech National Research Center funded by the European Union Next-Generation EU (Piano Nazionale di Ripresa e Resilienza (PNRR)- Missione 4 Componente 2, Investitmento 1.4 - D.D. 1032 17/06/2022, CN00000022). In particular, our study represents an original paper related to the Spoke 4 "Multifunctional and resilient agriculture and forestry systems for the mitigation of climate change risks" of the PNRR project. However, views and opinions expressed are those of the authors and do not necessarily reflect those of the European Union or the European Research Agency (REA). Neither the European Union nor the granting authorities can be held responsible. Open access funding provided by Consiglio Nazionale Delle Ricerche (CNR) within the CRUI-CARE Agreement.Escaray, FJ.; Valeri, MC.; Damiani, F.; Ruiz, OA.; Carrasco, P.; Paolocci, F. (2024). Multiple bHLH/MYB-based protein complexes regulate proanthocyanidin biosynthesis in the herbage of Lotus spp. Planta. 259(1). https://doi.org/10.1007/s00425-023-04281-2259

Metabolic Profiling and Metabolite Correlation Network Analysis Reveal that Fusarium solani Induces Differential Metabolic Responses in Lotus japonicus and Lotus tenuis against Severe Phosphate Starvation

Root fungal endophytes are essential mediators of plant nutrition under mild stress conditions. However, variations in the rhizosphere environment, such as nutrient depletion, could result in a stressful situation for both partners, shifting mutualistic to nonconvenient interactions. Mycorrhizal fungi and dark septate endophytes (DSEs) have demonstrated their ability to facilitate phosphate (Pi) acquisition. However, few studies have investigated other plant–fungal interactions that take place in the root environment with regard to phosphate nutrition. In the present research work, we aimed to analyze the effect of extreme Pi starvation and the fungal endophyte Fusarium solani on the model Lotus japonicus and the crop L. tenuis. We conducted metabolomics analysis based on gas chromatography-mass spectrometry (GC-MS) on plant tissues under optimal conditions, severe Pi starvation and F.solani presence. By combining statistical and correlation network analysis strategies, we demonstrated the differential outcomes of the two plant species against the combination of treatments. The combination of nutritional stress and Fusarium presence activated significant modifications in the metabolism of L. japonicus affecting the levels of sugars, polyols and some amino acids. Our results display potential markers for further inspection of the factors related to plant nutrition and plant–fungal interactions.Fil: Nieva, Amira Susana del Valle. Max Planck Institute of Molecular Plant Physiology; Alemania. Deutscher Akademischer Austauschdienst; AlemaniaFil: Romero, Fernando Matias. Universidad Nacional de San Martin. Instituto Tecnologico de Chascomus. - Consejo Nacional de Investigaciones Cientificas y Tecnicas. Centro Cientifico Tecnologico Conicet - la Plata. Instituto Tecnologico de Chascomus.; ArgentinaFil: Erban, Alexander. Max Planck Institute of Molecular Plant Physiology; AlemaniaFil: Carrasco, Pedro. Universidad de Valencia; EspañaFil: Ruiz, Oscar Adolfo. Universidad Nacional de San Martin. Instituto Tecnologico de Chascomus. - Consejo Nacional de Investigaciones Cientificas y Tecnicas. Centro Cientifico Tecnologico Conicet - la Plata. Instituto Tecnologico de Chascomus.; ArgentinaFil: Kopka, Joachim. Max Planck Institute of Molecular Plant Physiology; Alemani

Defense Responses in Two Ecotypes of Lotus japonicus against Non-Pathogenic Pseudomonas syringae

Lotus japonicus is a model legume broadly used to study many important processes as nitrogen fixing nodule formation and adaptation to salt stress. However, no studies on the defense responses occurring in this species against invading microorganisms have been carried out at the present. Understanding how this model plant protects itself against pathogens will certainly help to develop more tolerant cultivars in economically important Lotus species as well as in other legumes. In order to uncover the most important defense mechanisms activated upon bacterial attack, we explored in this work the main responses occurring in the phenotypically contrasting ecotypes MG-20 and Gifu B-129 of L. japonicus after inoculation with Pseudomonas syringae DC3000 pv. tomato. Our analysis demonstrated that this bacterial strain is unable to cause disease in these accessions, even though the defense mechanisms triggered in these ecotypes might differ. Thus, disease tolerance in MG-20 was characterized by bacterial multiplication, chlorosis and desiccation at the infiltrated tissues. In turn, Gifu B-129 plants did not show any symptom at all and were completely successful in restricting bacterial growth. We performed a microarray based analysis of these responses and determined the regulation of several genes that could play important roles in plant defense. Interestingly, we were also able to identify a set of defense genes with a relative high expression in Gifu B-129 plants under non-stress conditions, what could explain its higher tolerance. The participation of these genes in plant defense is discussed. Our results position the L. japonicus-P. syringae interaction as a interesting model to study defense mechanisms in legume species.Fil: Bordenave, César Daniel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico La Plata. Instituto de Investigaciones Biotecnológicas - Instituto Tecnológico Chascomús. Instituto de Investigaciones Biotecnológicas (sede Chascomús); Argentina;Fil: Escaray, Francisco José. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico La Plata. Instituto de Investigaciones Biotecnológicas - Instituto Tecnológico Chascomús. Instituto de Investigaciones Biotecnológicas (sede Chascomús); Argentina;Fil: Menendez, Ana Bernardina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico La Plata. Instituto de Investigaciones Biotecnológicas - Instituto Tecnológico Chascomús. Instituto de Investigaciones Biotecnológicas (sede Chascomús); Argentina;Fil: Serna, Eva. Universidad de Valencia. INCLIVA. Unidad Central en Investigación en Medicina; España;Fil: Carrazco, Pedro. Universidad de Valencia. Departamento de Bioquímica y Biología Vegetal; España;Fil: Ruiz, Oscar Adolfo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico la Plata. Instituto de Investigaciones Biotecnológicas - Instituto Tecnológico Chascomús. Instituto de Investigaciones Biotecnológicas (sede Chascomús); Argentina;Fil: Gárriz, Andrés. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico La Plata. Instituto de Investigaciones Biotecnológicas - Instituto Tecnológico Chascomús. Instituto de Investigaciones Biotecnológicas (sede Chascomús); Argentina

Phenotypic plasticity with respect to salt stress response by Lotus glaber: The role of its AM fungal and rhizobial symbionts

Our hypothesis is that Lotus glaber (a glycophytic species, highly tolerant to saline-alkaline soils) displays a plastic root phenotypic response to soil salinity that may be influenced by mycorrhizal and rhizobial microorganisms. Uninoculated plants and plants colonised by Glomus intraradices or Mesorhizobium loti were exposed to either 150 or 0 mM NaCl. General plant growth and root architectural parameters (morphology and topology) were measured and phenotypic plasticity determined at the end of the salt treatment period. Two genotypes differing in their salt tolerance capacity were used in this study. G. intraradices and M. loti reduced the total biomass of non-salinised, sensitive plants, but they did not affect that of corresponding tolerant ones. Root morphology of sensitive plants was greatly affected by salinity, whereas mycorrhiza establishment counteracted salinity effects. Under both saline conditions, the external link length and the internal link length of mycorrhizal salt-sensitive plants were higher than those of uninoculated control and rhizobial treatments. The topological trend (TT) was strongly influenced by genotype x symbiosis interaction. Under non-saline conditions, nodulated root systems of the sensitive plant genotype had a more herringbone architecture than corresponding uninoculated ones. At 150 mM NaCl, nodulated root systems of tolerant plants were more dichotomous and those of the corresponding sensitive genotype more herringbone in architecture. Notwithstanding the absence of a link between TTs and variations in plant growth, it is possible to predict a dissimilar adaptation of plants with different TTs. Root colonisation by either symbiotic microorganisms reduced the level of root phenotypic plasticity in the sensitive plant genotype. We conclude that root plasticity could be part of the general mechanism of L. glaber salt tolerance only in the case of non-symbiotic plants.Fil: Echeverria, Mariela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); ArgentinaFil: Scambato, Agustina Azul. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Biodiversidad y Biología Experimental; ArgentinaFil: Sannazzaro, Analía Inés. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); ArgentinaFil: Maiale, Santiago Javier. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); ArgentinaFil: Ruiz, Oscar Adolfo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús). Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. Instituto de Investigaciones Biotecnológicas "Dr. Raúl Alfonsín" (sede Chascomús); ArgentinaFil: Menendez, Ana Bernardina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Biodiversidad y Biología Experimental; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin