New Insights in Trichoderma harzianum Antagonism of Fungal Plant Pathogens by Secreted Protein Analysis

Trichoderma harzianum ALL42 were capable of overgrowing and degrading Rhizoctonia solani and Macrophomina phaseolina mycelia, coiling around the hyphae with formation of apressoria and hook-like structures. Hyphae of T. harzianum ALL42 did not show any coiling around Fusarium sp. hyphae suggesting that mycoparasitism may be different among the plant pathogens. In this study, a secretome analysis was used to identify some extracellular proteins secreted by T. harzianum ALL42 after growth on cell wall of M. phaseolina, Fusarium sp., and R. solani. The secreted proteins were analyzed by two-dimensional electrophoresis and MALDI-TOF mass spectrometry. A total of 60 T. harzianum ALL42 secreted proteins excised from the gel were analyzed from the three growth conditions. While seven cell wall-induced proteins were identified, more than 53 proteins spots remain unidentified, indicating that these proteins are either novel proteins or proteins that have not yet been sequenced. Endochitinase, β-glucosidase, α-mannosidase, acid phosphatase, α-1,3-glucanase, and proteases were identified in the gel and also detected in the supernatant of culture.     

Significant role for Fas in the pathogenesis of autoimmune diabetes

Programmed cell death represents an important pathogenic mechanism in various autoimmune diseases. Type I diabetes mellitus (IDDM) is a T cell-dependent autoimmune disease resulting in selective destruction of the beta cells of the islets of Langerhans. beta cell apoptosis has been associated with IDDM onset in both animal models and newly diagnosed diabetic patients. Several apoptotic pathways have been implicated in beta cell destruction, including Fas, perforin, and TNF-alpha. Evidence for Fas-mediated lysis of beta cells in the pathogenesis of IDDM in nonobese diabetic (NOD) mice includes: 1) Fas-deficient NOD mice bearing the lpr mutation (NOD-lpr/lpr) fail to develop IDDM; 2) transgenic expression of Fas ligand (FasL) on beta cells in NOD mice may result in accelerated IDDM; and 3) irradiated NOD-lpr/lpr mice are resistant to adoptive transfer of diabetes by cells from NOD mice. However, the interpretation of these results is complicated by the abnormal immune phenotype of NOD-lpr/lpr mice. Here we present novel evidence for the role of Fas/FasL interactions in the progression of NOD diabetes using two newly derived mouse strains. We show that NOD mice heterozygous for the FasL mutation gld, which have reduced functional FasL expression on T cells but no lymphadenopathy, fail to develop IDDM. Further, we show that NOD-lpr/lpr mice bearing the scid mutation (NOD-lpr/lpr-scid/scid), which eliminates the enhanced FasL-mediated lytic activity induced by Fas deficiency, still have delayed onset and reduced incidence of IDDM after adoptive transfer of diabetogenic NOD spleen cells. These results provide evidence that Fas/FasL-mediated programmed cell death plays a significant role in the pathogenesis of autoimmune diabetes.     

Moniliophthora perniciosa Necrosis- and Ethylene-Inducing Protein 2 (MpNep2) as a Metastable Dimer in Solution: Structural and Functional Implications

Understanding how Nep-like proteins (NLPs) behave during the cell cycle and disease progression of plant pathogenic oomycetes, fungi and bacteria is crucial in light of compelling evidence that these proteins play a role in Witches` Broom Disease (WBD) of Theobroma cacao, one of the most important phytopathological problems to afflict the Southern Hemisphere. The crystal structure of MpNep2, a member of the NLP family and the causal agent of WBD, revealed the key elements for its activity. This protein has the ability to refold after heating and was believed to act as a monomer in solution, in contrast to the related homologs MpNep1 and NPP from the oomyceteous fungus Phytophthora parasitica. Here, we identify and characterize a metastable MpNep2 dimer upon over-expression in Escherichia coli using different biochemical and structural approaches. We found using ultra-fast liquid chromatography that the MpNep2 dimer can be dissociated by heating but not by dilution, oxidation or high ionic strength. Small-angle X-ray scattering revealed a possible tail-to-tail interaction between monomers, and nuclear magnetic resonance measurements identified perturbed residues involved in the putative interface of interaction. We also explored the ability of the MpNep2 monomer to refold after heating or chemical denaturation. We observed that MpNep2 has a low stability and cooperative fold that could be an explanation for its structure and activity recovery after stress. These results can provide new insights into the mechanism for MpNep2′s action in dicot plants during the progression of WBD and may open new avenues for the involvement of NLP- oligomeric species in phytopathological disorders.     

Marine foraging ecology influences mercury bioaccumulation in deep-diving northern elephant seals

Mercury contamination of oceans is prevalent worldwide and methylmercury concentrations in the mesopelagic zone (200–1000 m) are increasing more rapidly than in surface waters. Yet mercury bioaccumulation in mesopelagic predators has been understudied. Northern elephant seals (Mirounga angustirostris) biannually travel thousands of kilometres to forage within coastal and open-ocean regions of the northeast Pacific Ocean. We coupled satellite telemetry, diving behaviour and stable isotopes (carbon and nitrogen) from 77 adult females, and showed that variability among individuals in foraging location, diving depth and δ¹³C values were correlated with mercury concentrations in blood and muscle. We identified three clusters of foraging strategies, and these resulted in substantially different mercury concentrations: (i) deeper-diving and offshore-foraging seals had the greatest mercury concentrations, (ii) shallower-diving and offshore-foraging seals had intermediate levels, and (iii) coastal and more northerly foraging seals had the lowest mercury concentrations. Additionally, mercury concentrations were lower at the end of the seven-month-long foraging trip (n = 31) than after the two-month- long post-breeding trip (n = 46). Our results indicate that foraging behaviour influences mercury exposure and mesopelagic predators foraging in the northeast Pacific Ocean may be at high risk for mercury bioaccumulation.