A Novel Real Time PCR Method for the Detection and Quantification of Didymella pinodella in Symptomatic and Asymptomatic Plant Hosts

Didymella pinodella is the major pathogen of the pea root rot complex in Europe. This wide host range pathogen often asymptomatically colonizes its hosts, making the control strategies challenging. We developed a real-time PCR assay for the detection and quantification of D. pinodella based on the TEF-1 alpha gene sequence alignments. The assay was tested for specificity on a 54-isolate panel representing 35 fungal species and further validated in symptomatic and asymptomatic pea and wheat roots from greenhouse tests. The assay was highly consistent across separate qPCR reactions and had a quantification/detection limit of 3.1 pg of target DNA per reaction in plant tissue. Cross-reactions were observed with DNA extracts of five Didymella species. The risk of cross contamination, however, is low as the non-targets have not been associated with pea previously and they were amplified with at least 1000-fold lower sensitivity. Greenhouse inoculation tests revealed a high correlation between the pathogen DNA quantities in pea roots and pea root rot severity and biomass reduction. The assay also detected D. pinodella in asymptomatic wheat roots, which, despite the absence of visible root rot symptoms, caused wheat biomass reduction. This study provides new insights into the complex life style of D. pinodella and can assist in better understanding the pathogen survival and spread in the environment.

Multimodal Spectroscopic Imaging of Pea Root Nodules to Assess the Nitrogen Fixation in the Presence of Biofertilizer Based on Nod-Factors

Multimodal spectroscopic imaging methods such as Matrix Assisted Laser Desorption/Ionization Mass Spectrometry Imaging (MALDI MSI), Fourier Transform Infrared spectroscopy (FT-IR) and Raman spectroscopy were used to monitor the changes in distribution and to determine semi quantitatively selected metabolites involved in nitrogen fixation in pea root nodules. These approaches were used to evaluate the effectiveness of nitrogen fixation by pea plants treated with biofertilizer preparations containing Nod factors. To assess the effectiveness of biofertilizer, the fresh and dry masses of plants were determined. The biofertilizer was shown to be effective in enhancing the growth of the pea plants. In case of metabolic changes, the biofertilizer caused a change in the apparent distribution of the leghaemoglobin from the edges of the nodule to its centre (the active zone of nodule). Moreover, the enhanced nitrogen fixation and presumably the accelerated maturation form of the nodules were observed with the use of a biofertilizer.

Untangling the Pea Root Rot Complex Reveals Microbial Markers for Plant Health

Plant health is recognised as a key element to ensure global food security. While plant breeding has substantially improved crop resistance against individual pathogens, it showed limited success for diseases caused by the interaction of multiple pathogens such as root rot in pea (Pisum sativum L.). To untangle the causal agents of the pea root rot complex and determine the role of the plant genotype in shaping its own detrimental or beneficial microbiome, fungal and oomycete root rot pathogens, as well as previously identified beneficials, i.e., arbuscular mycorrhizal fungi (AMF) and Clonostachys rosea, were qPCR quantified in diseased roots of eight differently resistant pea genotypes grown in four agricultural soils under controlled conditions. We found that soil and pea genotype significantly determined the microbial compositions in diseased pea roots. Despite significant genotype x soil interactions and distinct soil-dependent pathogen complexes, our data revealed key microbial taxa that were associated with plant fitness. Our study indicates the potential of fungal and oomycete markers for plant health and serves as a precedent for other complex plant pathosystems. Such microbial markers can be used to complement plant phenotype- and genotype-based selection strategies to improve disease resistance in one of the world’s most important pulse crops of the world.

The Mitotic Index of Cajanus cajan from Kisar Island, in the Southwest of Maluku

The mitotic index of the roots of pigeon pea can be the basis for determining the growth of pigeon pea. The purpose of this research was to determine the time of root cell division, to observe the mitotic phases, and to determine the mitotic index of pigeon pea root cells. The preparation of the pigeon pea was carried out for 4 days to grow the roots. The roots were cut off at 08.00, 08.15, and 08.30 WIT (Eastern Indonesian Time). The roots were cut 0.5-1cm. Carnoy’s solution was used as the fixative solution using the Squash technique. The prepared roots were then observed using an Olympus cx-22 microscope and an OptiLab camera with a magnification of 100x40. The data were descriptively analyzed to describe the images of mitotic phases and the mitotic index presentation in the root cells of pigeon pea. The results of this research showed that the cell division of the pigeon pea roots began at 08.00 WIT, which was marked by the presence of a lot of prophase. The next phases that appeared were prometaphase, metaphase, and anaphase which occurred from 08.15 to 08.30 with different numbers. The highest mitotic index occurred at 08.15, when most of the root cells underwent metaphase. This study succeeded in revealing that the optimum time for pigeon pea root cell division is 08.15 WIT. In the future, this research can help pigeon pea farmers in Southwest of Maluku to carry out vegetative reproduction which is closely related to this mitotic study.

Poor Competitiveness of Bradyrhizobium in Pigeon Pea Root Colonization in Indian Soils

Plant symbiosis with N 2 -fixing bacteria is key to sustainable, low-input agriculture. While there are ongoing projects aiming to increase yield of cereals using plant genetics and host-microbiota interaction engineering, the biggest potential lies in legume plants.

Biological control of Phaseolus vulgaris and Pisum sativum root rot disease using Trichoderma species

Background Root rot pathogens reported to cause considerable losses in both the quality and productivity of common bean (Phaseolus vulgaris L.) and pea (Pisum sativum L.). It is an aggressive crop disease with detriment economic influence caused by Fusarium solani and Rhizoctonia solani among other soil-borne fungal pathogens. Destructive plant diseases such as root rot have been managed in the last decades using synthetic pesticides. Main body Seeking of economical and eco-friendly alternatives to combat aggressive soil-borne fungal pathogens that cause significant yield losses is urgently needed. Trichoderma emerged as promising antagonist that inhibits pathogens including those inducing root rot disease. Detailed studies for managing common bean and pea root rot disease using different Trichoderma species (T. harzianum, T. hamatum, T. viride, T. koningii, T. asperellum, T. atroviridae, T. lignorum, T. virens, T. longibrachiatum, T. cerinum, and T. album) were reported both in vitro and in vivo with promotion of plant growth and induction of systemic defense. The wide scale application of selected metabolites produced by Trichoderma spp. to induce host resistance and/or to promote crop yield, may represent a powerful tool for the implementation of integrated pest management strategies. Conclusions Biological management of common bean and pea root rot-inducing pathogens using various species of the Trichoderma fungus might have taken place during the recent years. Trichoderma species and their secondary metabolites are useful in the development of protection against root rot to bestow high-yielding common bean and pea crops.

Impact of crop stand, Rhizobium inoculation, and foliar fertilization on pea root parameters

SummaryEcological intensification of crop production involves the use of intercrops and the rational use of inoculation and fertilization in case of intercrops including legume species. The root system plays an important role in the productivity of crops. Therefore, effects of the inoculation treatments (Nitragina) or foliar fertilization (Photrel) or a combination of both were assessed on root parameters of pea grown as pure stand or intercrops with linseed or wheat in a 3-year experiment in Poland. Crop stand composition influenced the root parameters of pea with a higher root length density (RLD) in the root fractions of 0.1–1 mm of pea in pea/linseed intercrops than in the pure stand, a higher mean root diameter (MRD) in pure pea and intercrops of pea with linseed than with wheat, and also a tendency of a higher root dry matter (RDM) in pure pea and pea/linseed than in pea/wheat in 2 out of the 3 years. RLD was higher with Photrel than with Nitragina in root fractions of 0.1–0.5 mm. Treatments did not affect the MRD, but a combination of Nitragina + Photrel increased the RDM in 1 year. Intercropping of pea with linseed and the application of a foliar fertilizer might be a strategy to improve pea root characteristics.

Regulation of the activity of adenylate cyclases by hydrogen peroxide in pea root cells Infected with pathogens and a mutualist

This study examines the effect of a range of exogenous concentrations of hydrogen peroxide on the activity of transmembrane and soluble adenylate cyclases (EC 4.6.1.1) contained in root cells of pea seedlings infected with one of the following: Rhizobium leguminosarum bv. Viciae, Pseudomonas syringae pv. Pisi, and Clavibacter michiganensis ssp. sepedonicus. The results showed that the pool of intracellular H2O2 increased when pea roots were infected with bacteria regardless of type. The study analysed the concentration of intracellular cyclic adenosine monophosphate, a product of the adenosine triphosphate cyclization reaction catalyzed by transmembrane and soluble adenylate cyclases. The concentration of intracellular cyclic adenosine monophosphate increased when infected with either Rhizobium leguminosarum bv. viciae or Clavibacter michiganensis ssp. Sepedonicus; however, the concentration decreased by 20% when infected with Pseudomonas syringae pv. Pisi. The in vitro activity of soluble and transmembrane adenylate cyclases from pea root cells inoculated with Rhizobium leguminosarum bv. viciae was H2O2 dose-dependent: 100 nM of H2O2 reduced the activity of soluble and transmembrane adenylate cyclases slightly, while 26 µM inhibited their activity by 50–60%. When infected with Pseudomonas syringae pv. pisi, the reduction in the activity of soluble and transmembrane adenylate cyclases was independent of the concentrations of H2O2 in the range investigated. When infected with Clavibacter michiganensis ssp. sepedonicus, 100 nM of H2O2 inhibited the activity of transmembrane adenylate cyclases, although enhancing the activity of soluble adenylate cyclases. On the contrary, concentrations of H2O2 of 2.6 and 26 µM increased the activity of transmembrane adenylate cyclases and inhibited the activity of soluble adenylate cyclases. It can be concluded that the specific concentration of second messengers in plant cells depends on the specificity of the biotic stressor and forms, inter alia, by their mutual influence on the components of other plant signaling systems.