Contrasting response of two Lotus corniculatus L. accessions to combined waterlogging-saline stress.

Waterlogging and salinity impair crop growth and productivity worldwide, with their combined effects being larger than the additive effects of the two stresses separately. Here, a common forage tetraploid Lotus corniculatus (cv. San Gabriel) and a diploid L. corniculatus accession, collected from a coastal area with high frequency of waterlogging-saline stress events, were evaluated for tolerance to waterlogging, salinity and these two stresses combined. We hypothesize that, due to its environmental niche, the diploid accession would show better adaptation to combined waterlogging-saline stress compared to the tetraploid L. corniculatus. Plants were evaluated under control conditions, waterlogging, salinity and a combined waterlogging-saline treatment for 33 days. Shoot and root growth were assessed, together with chlorophyll fluorescence and gas exchange measurements. Results showed that salinity and waterlogging effects were more severe for the tetraploid accession, with a larger effect being observed under the combined stress condition. Concentrations of Na+ , Cl- and K+ were measured in apical and basal leaves, and in roots. A larger accumulation of Na+ and Cl- was observed under both saline and combined stress treatments for the tetraploid L. corniculatus, for which ion toxicity effects were evident. The expression of CLC gene, coding for a Cl- transporter, was only increased in diploid L. corniculatus plants in response to the combined stress condition, suggesting that ion compartmentalization mechanisms were induced in this accession. Thus, this recently characterized L. corniculatus could be used for the introduction of new tolerance traits in other Lotus species used as forage.

Características fisiológicas de plantas hibernais com potencial fitorremediador sob influência dos herbicidas fomesafen e sulfentrazone / Physiological characteristics of winter cultivated plants with phytoremediation potential under the influence of of the herbicides fomesafem and sulfentrazone

O emprego de espécies vegetais para fitorremediação de solos contaminados com herbicidas persistentes, como os inibidores da enzima protoporfirinogênio oxidase (PROTOX), é uma alternativa interessante dos pontos de vista econômico e ambiental. O objetivo deste trabalho foi avaliar a influência de fomesafen e sulfentrazone nas características relacionadas à fisiologia de espécies hibernais compotencial para aplicação como fitorremediadoras de solo contaminado por esses herbicidas. Dois experimentos foram instalados em casa de vegetação, um com o herbicida fomesafen e outro com o sulfentrazone, no delineamento de blocos casualizados com quatro repetições. Foram testadas doses dos herbicidas fomesafen (0,0; 0,125; 0,250; e 0,5 kg ha-1) e sulfentrazone (0,0; 0,3; 0,6; e 1,2 kg ha-1),marcas comerciais Flex® e Boral 500®, respectivamente, aplicadas na pré-emergência das espécies com potencial fitorremediador (aveia-preta, ervilhaca, nabo e cornichão). Aos 45 dias após a emergência das plantas, foram avaliados o índice de clorofila, a eficiência de carboxilação (EC- mol CO2 m-2 s-1), a condutância estomática de vapores de água (Gs - mol m-1 s-1), a concentração interna de CO2 (Ci - µmolmol-1), a taxa de transpiração (E - mol H2O m-2 s-1), a quantidade de CO2 consumido (QT - µmol mol-1), ouso eficiente da água (UEA - mol CO2 mol H2O-1), a temperatura da folha ΔT (ºC), a taxa fotossintética (A- µmol m-2 s-1) e a massa seca (g vaso-1) da parte aérea. Observou-se que a ervilhaca apresenta os melhores resultados para todas as variáveis avaliadas, demonstrando assim potencial para ser testada como fitorremediadora de solos contaminados com os herbicidas fomesafen e sulfentrazone.(AU)

The use of plant species for phytoremediation of soil contaminated with persistent herbicides, such as inhibitors of the protoporphyrinogen oxidase enzyme (PROTOX), is an interesting alternative from the economic and environmental point of view. This work aimed to evaluate the influence of fomesafen and sulfentrazone on the characteristics related to the physiology of winter species with potential for phytoremediation of soil contaminated by these herbicides. Two experiments were installed in a greenhouse, one with the herbicide fomesafen and another with the sulfentrazone, in a randomized complete block design with four replications. Fomesafen (0.0, 0.125, 0.250, and 0.5 kg ha-1) and sulfentrazone (0.0, 0.3, 0.6, and 1.2 kg ha-1), commercial brands Flex® and Boral 500®, respectively, were applied pre-emergence on species with phytoremediation potential (black oats, vetch, turnip and lotus).The chlorophyll index, the carboxylation efficiency (EC - mol CO2 m-2 s-1), the stomatal conductance of water vapors (Gs - mol m-1 s-1), the internal CO2 concentration (Ci - μmol mol-1), the transpiration rate (E -mol H2O m-2 s-1), the amount of CO2 consumed (QT - μmol mol-1) and the water use efficiency (UEA - molCO2 mol H2O-1), the leaf temperature ΔT (ºC), the photosynthetic rate (A - μmol m-2 s-1) and the shoot drymass (g vase-1) were evaluated 45 days after plant emergence. It was observed that the vetch presented the best results for all evaluated variables, thus demonstrating potential to be further tested for phytoremediation of soils contaminated with the herbicides fomesafen and sulfentrazone.(AU)

Características fisiológicas de plantas hibernais com potencial fitorremediador sob influência dos herbicidas fomesafen e sulfentrazone / Physiological characteristics of winter cultivated plants with phytoremediation potential under the influence of of the herbicides fomesafem and sulfentrazone

O emprego de espécies vegetais para fitorremediação de solos contaminados com herbicidas persistentes, como os inibidores da enzima protoporfirinogênio oxidase (PROTOX), é uma alternativa interessante dos pontos de vista econômico e ambiental. O objetivo deste trabalho foi avaliar a influência de fomesafen e sulfentrazone nas características relacionadas à fisiologia de espécies hibernais compotencial para aplicação como fitorremediadoras de solo contaminado por esses herbicidas. Dois experimentos foram instalados em casa de vegetação, um com o herbicida fomesafen e outro com o sulfentrazone, no delineamento de blocos casualizados com quatro repetições. Foram testadas doses dos herbicidas fomesafen (0,0; 0,125; 0,250; e 0,5 kg ha-1) e sulfentrazone (0,0; 0,3; 0,6; e 1,2 kg ha-1),marcas comerciais Flex® e Boral 500®, respectivamente, aplicadas na pré-emergência das espécies com potencial fitorremediador (aveia-preta, ervilhaca, nabo e cornichão). Aos 45 dias após a emergência das plantas, foram avaliados o índice de clorofila, a eficiência de carboxilação (EC- mol CO2 m-2 s-1), a condutância estomática de vapores de água (Gs - mol m-1 s-1), a concentração interna de CO2 (Ci - µmolmol-1), a taxa de transpiração (E - mol H2O m-2 s-1), a quantidade de CO2 consumido (QT - µmol mol-1), ouso eficiente da água (UEA - mol CO2 mol H2O-1), a temperatura da folha ΔT (ºC), a taxa fotossintética (A- µmol m-2 s-1) e a massa seca (g vaso-1) da parte aérea. Observou-se que a ervilhaca apresenta os melhores resultados para todas as variáveis avaliadas, demonstrando assim potencial para ser testada como fitorremediadora de solos contaminados com os herbicidas fomesafen e sulfentrazone.

The use of plant species for phytoremediation of soil contaminated with persistent herbicides, such as inhibitors of the protoporphyrinogen oxidase enzyme (PROTOX), is an interesting alternative from the economic and environmental point of view. This work aimed to evaluate the influence of fomesafen and sulfentrazone on the characteristics related to the physiology of winter species with potential for phytoremediation of soil contaminated by these herbicides. Two experiments were installed in a greenhouse, one with the herbicide fomesafen and another with the sulfentrazone, in a randomized complete block design with four replications. Fomesafen (0.0, 0.125, 0.250, and 0.5 kg ha-1) and sulfentrazone (0.0, 0.3, 0.6, and 1.2 kg ha-1), commercial brands Flex® and Boral 500®, respectively, were applied pre-emergence on species with phytoremediation potential (black oats, vetch, turnip and lotus).The chlorophyll index, the carboxylation efficiency (EC - mol CO2 m-2 s-1), the stomatal conductance of water vapors (Gs - mol m-1 s-1), the internal CO2 concentration (Ci - μmol mol-1), the transpiration rate (E -mol H2O m-2 s-1), the amount of CO2 consumed (QT - μmol mol-1) and the water use efficiency (UEA - molCO2 mol H2O-1), the leaf temperature ΔT (ºC), the photosynthetic rate (A - μmol m-2 s-1) and the shoot drymass (g vase-1) were evaluated 45 days after plant emergence. It was observed that the vetch presented the best results for all evaluated variables, thus demonstrating potential to be further tested for phytoremediation of soils contaminated with the herbicides fomesafen and sulfentrazone.

Efecto in vitro del extracto de Lotus corniculatus L. sobre nemátodos gastrointestinales bovinos / In vitro effect of extract of Lotus corniculatus L. on bovine gastrointestinal nematodes

Introducción: las parasitosis son consideradas como uno de los problemas más importantes que afectan la producción bovina, debido a la disminución de los parámetros productivos y reproductivos. Objetivos: determinar el efecto del extracto Lotus corniculatus L. in vitro sobre nemátodos gastrointestinales bovinos. Métodos: se usaron las hojas en estado seco de L. corniculatus con una edad de rebrote de 45 y 70 días, el extracto fue etanólico y se obtuvo mediante lixiviación, el extracto se conservó a temperatura de 4 °C, se utilizó el extracto puro y diluciones obedeciéndose a un intervalo de 1,5 mg/mL. Para determinar el efecto del extracto se trabajó mediante la técnica de eclosión de huevos (Egg hatch assay) para los nemátodos: Bunostomum sp, Strongyloides papilosus y Trichostrongylus colubriformis, tomando como mínimo eficaz un porcentaje del 60 por ciento. Se determinó la presencia de taninos. Resultados: se encontró una inhibición de eclosión de huevos S. papilus con la concentración máxima y la dilución 1 y 2, para el extracto de 45 días de rebrote, con porcentajes de 100 por ciento ,86 por ciento y 66,5 por ciento; el extracto con 70 días de rebrote mostró un porcentaje de inhibición de eclosión únicamente[y1] con la concentración máxima y la dilución 1, con 98,5 por ciento y 78,5 por ciento. Para el T. colubriformis la única dilución que no fue eficaz fue la dilución 4 del extracto de 70 días de rebrote con un porcentaje de 50,5 por ciento y para el Bunostomun spp fue eficaz el extracto con la concentración máxima y la dilución 1 para el extracto de 45 y 70 días de rebrote, con porcentajes de 100 por ciento y 69,5 por ciento para el de 45 días y 100 por ciento y 68,5 por ciento para el de 70 días. Conclusiones: el extracto de L. corniculatus de 45 y 70 días de rebrote mostró los resultados más satisfactorios en inhibición de eclosión de huevos de T. colubriformis(AU)

Introduction: The parasites are considered as one of the most important issues affecting cattle production due to the decline in productive and reproductive parameters. Objective: Determine the effect of Lotus corniculatus L. extract on in vitro bovine gastrointestinal nematodes. Methods: Leaves were used in the dry state with L. corniculatus regrowth age 45 to 70 days, and the ethanolic extract was obtained by leaching, the extract was stored at 4 °C, the pure extract was used and dilutions obeying a range of 1.5 mg/mL. To determine the effect of the extract worked through the technique of hatching eggs (Egg hatch assay) for nematodes: Bunostomum ssp, Trichostrongylus and Strongyloides papilosus, on the effective minimum percentage of 60 percent. The presence of tannins was determined. Results: Inhibition of egg hatching S. papilus with maximum concentration and dilution 1 and 2, to extract 45 days of regrowth, with percentages of 100 percent, 86 percent and 66.5 percent respectively was found; extract regrowth 70 days showed a percent inhibition of hatching only the maximum concentration and dilution 1,98.5 percent and 78.5 percent. For T. colubriformis the only was ineffective dilution was 4 dilution extract regrowth 70 days with a percentage of 50.5 percent and for Bunostomun spp extract was effective with a high concentration and dilution 1 for extract 45 and 70 days of regrowth, with rates of 100 percent and 69.5 percent for 45 days and 100 percent and 68.5 percent for 70 days. Conclusions: L. corniculatus extract 45 and 70 days of regrowth showed the most satisfactory results in inhibition of hatching eggs of T. colubriformis(AU)

Salt effects on functional traits in model and in economically important Lotus species.

A common stress on plants is NaCl-derived soil salinity. Genus Lotus comprises model and economically important species, which have been studied regarding physiological responses to salinity. Leaf area ratio (LAR), root length ratio (RLR) and their components, specific leaf area (SLA) and leaf mass fraction (LMF) and specific root length (SRL) and root mass fraction (RMF) might be affected by high soil salinity. We characterised L. tenuis, L. corniculatus, L. filicaulis, L. creticus, L. burtii and L. japonicus grown under different salt concentrations (0, 50, 100 and 150 mm NaCl) on the basis of SLA, LMF, SRL and RMF using PCA. We also assessed effects of different salt concentrations on LAR and RLR in each species, and explored whether changes in these traits provide fitness benefit. Salinity (150 mm NaCl) increased LAR in L. burtii and L. corniculatus, but not in the remaining species. The highest salt concentration caused a decrease of RLR in L. japonicus Gifu, but not in the remaining species. Changes in LAR and RLR would not be adaptive, according to adaptiveness analysis, with the exception of SLA changes in L. corniculatus. PCA revealed that under favourable conditions plants optimise surfaces for light and nutrient acquisition (SLA and SRL), whereas at higher salt concentrations they favour carbon allocation to leaves and roots (LMF and RMF) in detriment to their surfaces. PCA also showed that L. creticus subjected to saline treatment was distinguished from the remaining Lotus species. We suggest that augmented carbon partitioning to leaves and roots could constitute a salt-alleviating mechanism through toxic ion dilution.

Response to long-term NaHCO3-derived alkalinity in model Lotus japonicus Ecotypes Gifu B-129 and Miyakojima MG-20: transcriptomic profiling and physiological characterization.

The current knowledge regarding transcriptomic changes induced by alkalinity on plants is scarce and limited to studies where plants were subjected to the alkaline salt for periods not longer than 48 h, so there is no information available regarding the regulation of genes involved in the generation of a new homeostatic cellular condition after long-term alkaline stress. Lotus japonicus is a model legume broadly used to study many important physiological processes including biotic interactions and biotic and abiotic stresses. In the present study, we characterized phenotipically the response to alkaline stress of the most widely used L. japonicus ecotypes, Gifu B-129 and MG-20, and analyzed global transcriptome of plants subjected to 10 mM NaHCO3 during 21 days, by using the Affymetrix Lotus japonicus GeneChip®. Plant growth assessment, gas exchange parameters, chlorophyll a fluorescence transient (OJIP) analysis and metal accumulation supported the notion that MG-20 plants displayed a higher tolerance level to alkaline stress than Gifu B-129. Overall, 407 and 459 probe sets were regulated in MG-20 and Gifu B-129, respectively. The number of probe sets differentially expressed in roots was higher than that of shoots, regardless the ecotype. Gifu B-129 and MG-20 also differed in their regulation of genes that could play important roles in the generation of a new Fe/Zn homeostatic cellular condition, synthesis of plant compounds involved in stress response, protein-degradation, damage repair and root senescence, as well as in glycolysis, gluconeogenesis and TCA. In addition, there were differences between both ecotypes in the expression patterns of putative transcription factors that could determine distinct arrangements of flavonoid and isoflavonoid compounds. Our results provided a set of selected, differentially expressed genes deserving further investigation and suggested that the L. japonicus ecotypes could constitute a useful model to search for common and distinct tolerance mechanisms to long-term alkaline stress response in plants.

Akaline, saline and mixed saline-alkaline stresses induce physiological and morpho-anatomical changes in Lotus tenuis shoots.

Saline, alkaline and mixed saline-alkaline conditions frequently co-occur in soil. In this work, we compared these plant stress sources on the legume Lotus tenuis, regarding their effects on shoot growth and leaf and stem anatomy. In addition, we aimed to gain insight on the plant physiological status of stressed plants. We performed pot experiments with four treatments: control without salt (pH = 5.8; EC = 1.2 dS·m(-1)) and three stress conditions, saline (100 mM NaCl, pH = 5.8; EC = 11.0 dS·m(-1)), alkaline (10 mM NaHCO3, pH = 8.0, EC = 1.9 dS·m(-1)) and mixed salt-alkaline (10 mM NaHCO3 + 100 mM NaCl, pH = 8.0, EC = 11.0 dS·m(-1)). Neutral and alkaline salts produced a similar level of growth inhibition on L. tenuis shoots, whereas their mixture exacerbated their detrimental effects. Our results showed that none of the analysed morpho-anatomical parameters categorically differentiated one stress from the other. However, NaCl- and NaHCO3 -derived stress could be discriminated to different extents and/or directions of changes in some of the anatomical traits. For example, alkalinity led to increased stomatal opening, unlike NaCl-treated plants, where a reduction in stomatal aperture was observed. Similarly, plants from the mixed saline-alkaline treatment characteristically lacked palisade mesophyll in their leaves. The stem cross-section and vessel areas, as well as the number of vascular bundles in the sectioned stem were reduced in all treatments. A rise in the number of vessel elements in the xylem was recorded in NaCl-treated plants, but not in those treated exclusively with NaHCO3.

Modulation of polyamine balance in Lotus glaber by salinity and arbuscular mycorrhiza.

In this work we investigated the involvement of Glomus intraradices in the regulation of plant growth, polyamines and proline levels of two Lotus glaber genotypes differing in salt tolerance, after longterm exposure to saline stress. The experiment consisted of a randomized block design with three factors: (1) mycorrhizal treatments (with or without AM fungus); (2) two salinity levels of 0 and 200mM NaCl; and (3) L. glaber genotype. Experiments were performed using stem cuttings derived from L. glaber individuals representing a natural population from saline lowlands. One of the most relevant results was the higher content of total free polyamines in mycorrhized plants compared to non-AM ones. Since polyamines have been proposed as candidates for the regulation of root development under saline situations, it is possible that AM plants (which contained higher polyamine levels and showed improved root growth) were better shaped to cope with salt stress. Colonization by G. intraradices also increased (Spd+Spm)/Put ratio in L. glaber roots. Interestingly, such increment in salt stressed AM plants of the sensitive genotype, was even higher than that produced by salinization or AM symbiosis separately. On the other hand, salinity but not mycorrhizal colonization influenced proline levels in both L. glaber genotypes since high proline accumulation was observed in both genotypes under salt stress conditions. Our results suggest that modulation of polyamine pools can be one of the mechanisms used by AM fungi to improve L. glaber adaptation to saline soils. Proline accumulation in response to salt stress is a good indicator of stress perception and our results suggest that it could be used as such among L. glaber genotypes differing in salt stress tolerance.