Antifungal Defense Research Articles

Changes in antifungal defence systems during the intermoult period in the Colorado potato beetle

Susceptibility to the fungus Metarhizium robertsii and changes in host defences were evaluated in different stages of the intermoult period (4–6 h, 34–36 h and 84–86 h post moult in IV larval instars) of the Colorado potato beetle. A significant thickening of the cuticle during larval growth was accompanied by decreases in cuticle melanization, phenoloxidase activity and epicuticular hydrocarbon contents (C28-C32). At the same time, a decrease in the conidial adhesion rate and an increase in resistance to the fungus were observed. In addition, we recorded significant elevation of the encapsulation rate and total haemocyte counts in the haemolymph during the specified period. The activity of detoxification enzymes decreased in the haemolymph but increased in the fat body during larval growth. No significant differences in the fatty acid content in the epicuticle were observed. The role of developmental disorders in susceptibility to entomopathogenic fungi is also discussed.

IκB-ζ Expression Requires Both TYK2/STAT3 Activity and IL-17-Regulated mRNA Stabilization.

Cytokine IL-17A (IL-17) acts on various cell types, including epidermal keratinocytes, and induces antimicrobial peptide and chemokine production to elicit antibacterial and antifungal defense responses. Excess IL-17 leads to inflammatory skin diseases such as psoriasis. The IκB family protein IκB-ζ mediates IL-17-induced responses. However, the mechanism controlling IκB-ζ expression in IL-17-stimulated cells remains elusive. In this study, we showed that JAK kinase TYK2 positively regulates IL-17-induced IκB-ζ expression. TYK2-deficient mice showed reduced inflammation and concomitant reduction of IκB-ζ mRNA compared with wild-type mice in imiquimod-induced skin inflammation. The analysis of the IκB-ζ promoter activity using human cell lines (HaCaT and HeLa) revealed that catalytic activity of TYK2 and its substrate transcription factor STAT3, but not IL-17, is required for IκB-ζ promoter activity. In contrast, IL-17-induced signaling, which did not activate STAT3, posttranscriptionally stabilized IκB-ζ mRNA via its 3'-untranslated region. IL-17 signaling protein ACT1 was required to counteract constitutive IκB-ζ mRNA degradation by RNase Regnase-1. These results suggested that transcriptional activation by TYK2-STAT3 pathway and mRNA stabilization by IL-17-mediated signals act separately from each other but complementarily to achieve IκB-ζ induction. Therefore, JAK/TYK2 inhibition might be of significance in regulation of IL-17-induced inflammatory reactions.

Malassezia Is Associated with Crohn’s Disease and Exacerbates Colitis in Mouse Models

Inflammatory bowel disease (IBD) is characterized by alterations in the intestinal microbiota and altered immuneresponses to gut microbiota. Evidence is accumulating that IBD is influenced by not only commensal bacteria but also commensal fungi. We characterized fungi directly associated with the intestinalmucosa in healthy people and Crohn's disease patients and identified fungi specifically abundant in patients. One of these, the common skin resident fungus Malassezia restricta, is also linked to the presence of an IBD-associated polymorphism in the gene for CARD9, a signaling adaptor important for anti-fungal defense. M.restricta elicits innate inflammatory responses largely through CARD9 and is recognized by Crohn's disease patient anti-fungal antibodies. This yeast elicits strong inflammatory cytokine production from innate cells harboring the IBD-linked polymorphism in CARD9 and exacerbates colitis via CARD9 in mouse models of disease. Collectively, these results suggest that targeting specific commensal fungi may be a therapeutic strategy for IBD.

Galectin-3 expression and effect of supplementation in neonatal mice with disseminated Candida albicans infection.

Background:Invasive candidiasis is an important cause of fungal infections in immunocompromised patients, including premature infants. The S-type lectin, galectin-3 (gal3), is increasingly recognized for its role in antifungal host defense. This study tested the hypothesis that tissue gal3 expression is affected by disseminated infection with Candida albicans and that supplementation with gal3 will provide a benefit in this setting.Methods:To determine the expression of gal3 at the tissue level in response to disseminated infection with C. albicans, adult and neonatal mice were infected using previously established models. End points were chosen that reflected substantive tissue fungal burden but before mortality.Results:No differences in gal3 were detected in tissues of adult animals relative to uninfected controls. In neonatal animals, gal3 concentration was lower in the spleen of infected animals compared to uninfected. Pretreatment of neonatal mice with recombinant gal3 was associated with reduced mortality and reduced fungal burden in the kidney, spleen and lung at 24 hours following infection.Conclusion:These findings suggest that gal3 has an active role in host defense against candidiasis and that neonatal animals can benefit from supplementation with this lectin in the setting of disseminated candidiasis.

Pseudomonas fluorescens increases mycorrhization and modulates expression of antifungal defense response genes in roots of aspen seedlings

BackgroundPlants, fungi, and bacteria form complex, mutually-beneficial communities within the soil environment. In return for photosynthetically derived sugars in the form of exudates from plant roots, the microbial symbionts in these rhizosphere communities provide their host plants access to otherwise inaccessible nutrients in soils and help defend the plant against biotic and abiotic stresses. One role that bacteria may play in these communities is that of Mycorrhizal Helper Bacteria (MHB). MHB are bacteria that facilitate the interactions between plant roots and symbiotic mycorrhizal fungi and, while the effects of MHB on the formation of plant-fungal symbiosis and on plant health have been well documented, the specific molecular mechanisms by which MHB drive gene regulation in plant roots leading to these benefits remain largely uncharacterized.ResultsHere, we investigate the effects of the bacterium Pseudomonas fluorescens SBW25 (SBW25) on aspen root transcriptome using a tripartite laboratory community comprised of Populus tremuloides (aspen) seedlings and the ectomycorrhizal fungus Laccaria bicolor (Laccaria). We show that SBW25 has MHB activity and promotes mycorrhization of aspen roots by Laccaria. Using transcriptomic analysis of aspen roots under multiple community compositions, we identify clusters of co-regulated genes associated with mycorrhization, the presence of SBW25, and MHB-associated functions, and we generate a combinatorial logic network that links causal relationships in observed patterns of gene expression in aspen seedling roots in a single Boolean circuit diagram. The predicted regulatory circuit is used to infer regulatory mechanisms associated with MHB activity.ConclusionsIn our laboratory conditions, SBW25 increases the ability of Laccaria to form ectomycorrhizal interactions with aspen seedling roots through the suppression of aspen root antifungal defense responses. Analysis of transcriptomic data identifies that potential molecular mechanisms in aspen roots that respond to MHB activity are proteins with homology to pollen recognition sensors. Pollen recognition sensors integrate multiple environmental signals to down-regulate pollenization-associated gene clusters, making proteins with homology to this system an excellent fit for a predicted mechanism that integrates information from the rhizosphere to down-regulate antifungal defense response genes in the root. These results provide a deeper understanding of aspen gene regulation in response to MHB and suggest additional, hypothesis-driven biological experiments to validate putative molecular mechanisms of MHB activity in the aspen-Laccaria ectomycorrhizal symbiosis.

Aspergeillus Fumigatus: The Effects of Adiponectin on Inflammatory Cytokine Production

Background and Hypothesis: Although inflammatory cytokines are important for antifungal defenses, excessive production significantly increases host immune pathology. It is therefore important to identify host pathways that limit detrimental inflammation in invasive fungal infection. Our prior results showed that mice with invasive aspergillosis (IA) that were deficient in the metabolic cytokine production produced more of the cytokine Tumor Necrosis Factor (TNF) than alveolar in wild-type control mice. Therefore, we hypothesize that adiponectin inhibits antifungal cytokine secretion in alveolar macrophages. 
 Experimental Design or Project Methods: To test this hypothesis, the commonly used A. fumigatus strain Af293 was purchased from the Fungal Genetics Stock Center and grown on and harvested from agar. The alveolar macrophage cell line MH-S and cytokine ELISA kits was obtained from MilliporeSigma and ThermoFisher, respectively. MH-S cells were stimulated with swollen, fixed Af293 conidia for 24 hours in the presence or absence of recombinant mouse adiponectin. After 24 hours, supernatants and cells was were collected and assayed for ILs 1a, 6, and TNF protein and mRNA , by ELISA and quantitative RT-PCR, respectively. 
 Results: Although our preliminary results suggest possible inhibition of cytokine secretion by adiponectin in response to A. fumigatus, significant differences have thus far not been observed. 
 Conclusion and Potential Impact: We are therefore currently optimizing our experimental conditions to improve antifungal cytokine secretion. These studies may ultimately assist in the discovery of novel therapeutic targets and improve the prognosis of A. fumigatus infections

Expression of a wheat β-1,3-glucanase in Pichia pastoris and its inhibitory effect on fungi commonly associated with wheat kernel

β-1,3-glucanases, the plant PR-2 family of pathogenesis-related (PR) proteins, can be constitutively expressed and induced in wheat crop to enhance its anti-fungal pathogen defense. This study aimed to investigate the inhibitory effect of wheat β-1,3-glucanase on fungi most commonly associated with wheat kernel. A β-1,3-glucanase from wheat was successfully expressed in Pichia pastoris X-33 and its biochemical and antifungal properties were characterized herein. The molecular weight of recombinant β-1,3-glucanase is approximately 33 kDa. β-1,3-glucanase displays optimal activity at pH 6.5, remaining relatively high at pH 5.5–8.0. The optimal reaction temperature of β-1,3-glucanase is 50 °C, retaining approximately 84.0% residual activity after heat-treated at 50 °C for 1 h. The steady-state kinetic parameters of β-1,3-glucanase against laminarin was determined and the Km and Vmax were 1.32 ± 0.20 mg/ml and 96.4 ± 4.4 U mg−1 protein, respectively. The inhibitory effect of purified β-1,3-glucanase against the seven fungi commonly associated with wheat kernel was assessed in vitro. β-1,3-glucanase exerted differential inhibitory effects on hyphal growth of Fusarium graminearum, Alternaria sp., A. glaucus, A. flavus, A. niger, and Penicillium sp. Spore formation and mycelial morphology of Alternaria sp., A. flavus, and A. niger were significantly affected by β-1,3-glucanase (1U). The present results would help elucidate the mechanism underlying the inhibition of wheat β-1,3-glucanases on pathogens.

Stabilizing the IL-17 response

Cytokines The inflammatory cytokine interleukin-17 (IL-17) can stimulate both antifungal host defense and autoimmunity by promoting the stability of target messenger RNAs (mRNAs). Amatya et al. found that IL-17 increased the abundance of the RNA binding protein Arid5a in mouse cells (see the Focus by Puel and Casanova). Arid5a promoted the cellular response to IL-17 by increasing the mRNA stability of a selection of IL-17–stimulated transcripts. For other transcripts, Arid5a interacted with the translation initiation factor eIF4G to augment their translation. Sci. Signal. 11 , eaat4617, eaau8876 (2018).

Intrinsic Programming of Alveolar Macrophages for Protective Antifungal Innate Immunity Against Pneumocystis Infection.

Invasive fungal infections, including Pneumocystis Pneumonia (PcP), remain frequent life-threatening conditions of patients with adaptive immune defects. While innate immunity helps control pathogen growth early during infection, it is typically not sufficient for complete protection against Pneumocystis and other human fungal pathogens. Alveolar macrophages (AM) possess pattern recognition molecules capable of recognizing antigenic and structural determinants of Pneumocystis. However, this pathogen effectively evades innate immunity to infect both immunocompetent and immunosuppressed hosts, albeit with differing outcomes. During our studies of mouse models of PcP, the FVB/N strain was identified as unique because of its ability to mount a protective innate immune response against Pneumocystis infection. In contrast to other immunocompetent strains, which become transiently infected prior to the onset of adaptive immunity, FVB/N mice rapidly eradicated Pneumocystis before an adaptive immune response was triggered. Furthermore, FVB/N mice remained highly resistant to infection even in the absence of functional T cells. The effector mechanism of innate protection required the action of functional alveolar macrophages, and the adoptive transfer of resistant FVB/N AMs, but not susceptible CB.17 AMs, conferred protection to immunodeficient mice. Macrophage IFNγ receptor signaling was not required for innate resistance, and FVB/N macrophages were found to display markers of alternative activation. IFNγ reprogrammed resistant FVB/N macrophages to a permissive M1 biased phenotype through a mechanism that required direct activation of the macrophage IFNγR. These results demonstrate that appropriately programmed macrophages provide protective innate immunity against this opportunistic fungal pathogen, and suggest that modulating macrophage function may represent a feasible therapeutic strategy to enhance antifungal host defense. The identification of resistant and susceptible macrophages provides a novel platform to study not only the mechanisms of macrophage-mediated antifungal defense, but also the mechanisms by which Pneumocystis evades innate immunity.

Emerging Aspergillus Species Almost Exclusively Associated With Primary Immunodeficiencies.

Invasive aspergillosis (IA) is the most serious mold infection encountered in patients with iatrogenic immunosuppression. IA is also a major cause of mortality and morbidity in individuals with primary immunodeficiency (PID). Although Aspergillus fumigatus is the most common etiologic agent of IA reported in PID patients, followed by A. nidulans, multiple poorly recognized Aspergillus species such as A. udagawae, A. quadrilineatus, A. pseudoviridinutans, A. tanneri, A. subramanianii, and A. fumisynnematus have been reported almost exclusively from patients with inborn defects in host antifungal defense pathways. Infection in PID patients exhibits patterns of disease progression distinct from those in iatrogenic immunosuppression. Specifically, the disease can be extrapulmonary and chronic with a tendency to disseminate in a contiguous manner across anatomical planes. It is also more refractory to standard antifungal therapy. This synopsis summarizes our understanding of emerging rare Aspergillus species that primarily affect patients with PIDs but not those with acquired immunodeficiencies.

Human MAITcells are rapidly activated by Aspergillus spp. in an APC-dependent manner.

Mucosal associated invariant Tcells (MAITcells) are innate-like Tcells (TC) which are known to be activated by several bacteria and viruses. However, activation of MAITcells by moulds, such as the opportunistic human pathogen Aspergillus, is not well described. Stimulation of human PBMC with A. fumigatus, A. flavus, or A. terreus conidia revealed that in contrast to conventional CD4+ and CD8+ TC, MAITcells responded already after 4 h of coincubation with upregulation of CD69. Furthermore, concurrent increase of CD107a expression and reduced intracellular expression of cytolytic proteins like perforin and granzyme indicated degranulation of intracellular vesicles. MAITcell activation only occurred in the presence of APC and was dependent on cell-cell contact as separation of TC and APC abrogated MAITcell activation. Furthermore, we observed that MAITcell activation by moulds requires presentation of riboflavin metabolites and depends on TCR engagement as antibody blocking of MR1, the antigen presenting molecule for MAITcells, prevented upregulation of CD69 and CD107a. In summary, we could demonstrate that MAITcells are activated by Aspergillus conidia in a TCR-dependent manner by APC. These findings reveal MAITcells as an interesting new target in antifungal defense.

Mutation in the C receptor of Interleukin 17, as a cause of Chronic Mucocutaneous Candidiasis

Background: The humoral and cellular immunity plays a preponderant role in the etiopathogenesis of esophagitis, for which the majority of the patients who present this pathology will have some type of immune system. Methods & Materials: Patient of 36 years of age, family history of Hiper IgE syndrome, consulted for abdominal pain, enterorrhagia, weight loss, dysphagia and vomiting. The physical examination shows important muguet. Regarding to personal history, it is recognized that thepatients episodes of candidiasis esophagitis from the age of 6 years and cutaneous plasmocytoma located in the wing of the nose that required surgical extirpation. Results: In endoscopic exploration, the esophagus appears hyperemic, with yellowish mucous plaques that adhere firmly, with rough and friable underlying surface after removal. Pathological anatomy informs esophagitis, with presence of yeasts compatible with Candida on the mucosal surface. In the stomach, the lamina propria presents edema with lymphocyte hypercellularity. Microbiology: Candida albicans. The patients begins treatment with Fluconazole. Laboratory: HIV negative Elisa, non-reactive HBsAg, non-reactive HCV antibodies. IgG 1727 mg/dl, IgA 271 mg/dl, IgM 154 mg/dl, IgE 172 mg/dl. CD3 2123 (76.8%), CD4 1024 (46.6%), CD8 674 (24.4%), CD19 498 (15.8%). A molecular biology study was carried out in search of mutations in the AIRE gene, sequencing of exons and adjacent intronic sequences, showing no alterations compatible with mutation. The mutation of this gene is linked to several autoimmune diseases, such as hypoparathyroidism, primary adrenocortical failure and chronic mucocutaneous candidiasis. In a new molecular study, it shows mutation in the C IL 17 receptor (IL17RC). This interleukin is related in the antifungal defense. Conclusion: Interleukin 17 participates in the development of autoimmunity, inflammation and tumor immunity, playing an important role in the defense against bacterial and fungal infections. The blocking of IL-17 signaling due to the mutation of its receptor prevents the formation of germinal centers and reduces the humoral response. The discovery of this mutation is a rarity, and plays an important role in defense against infectious diseases. In this clinical case, this mutation predisposes to the recurrence of chronic mucocutaneous candidiasis.

Calcium sequestration by fungal melanin inhibits calcium-calmodulin signalling to prevent LC3-associated phagocytosis.

LC3-associated phagocytosis (LAP) is a non-canonical autophagy pathway regulated by Rubicon, with an emerging role in immune homeostasis and antifungal host defence. Aspergillus cell wall melanin protects conidia (spores) from killing by phagocytes and promotes pathogenicity through blocking nicotinamide adenine dinucleotide phosphate (NADPH) oxidase-dependent activation of LAP. However, the signalling regulating LAP upstream of Rubicon and the mechanism of melanin-induced inhibition of this pathway remain incompletely understood. Herein, we identify a Ca2+ signalling pathway that depends on intracellular Ca2+ sources from endoplasmic reticulum, endoplasmic reticulum-phagosome communication, Ca2+ release from phagosome lumen and calmodulin (CaM) recruitment, as a master regulator of Rubicon, the phagocyte NADPH oxidase NOX2 and other molecular components of LAP. Furthermore, we provide genetic evidence for the physiological importance of Ca2+-CaM signalling in aspergillosis. Finally, we demonstrate that Ca2+ sequestration by Aspergillus melanin inside the phagosome abrogates activation of Ca2+-CaM signalling to inhibit LAP. These findings reveal the important role of Ca2+-CaM signalling in antifungal immunity and identify an immunological function of Ca2+ binding by melanin pigments with broad physiological implications beyond fungal disease pathogenesis.

Metal hyperaccumulation in the Brassicaceae species Arabidopsis halleri reduces camalexin induction after fungal pathogen attack

Metal hyperaccumulation from the soil by certain plant species can serve as a defence trait. Such hyperaccumulation might impact the expression of organic defences. Here, the induction of the phytoalexin camalexin after leaf infection with a pathogenic fungus was investigated in plants of the facultative hyperaccumulator Arabidopsis halleri grown on unamended and metal-amended soil. Reduced camalexin induction was expected in plants grown in metal-amended soil. Plants were grown for twelve weeks on soil, which was either amended with high concentrations of Zn and Cd or kept unamended, and the final leaf concentration of Zn and Cd was determined. Conidia of the pathogenic fungus Alternaria brassicae were applied on mechanically damaged leaves. Leaves were harvested after 24 h and 48 h, and the amount of induced camalexin was quantified. Plants grown on metal-amended soil hyperaccumulated Zn and Cd and induced significantly less camalexin than plants grown on unamended soils after pathogen infestation. This suggests a physiological trade-off between metal hyperaccumulation and an induced antifungal organic defence, which has, to our knowledge, not been observed before in A. halleri for other organic defence compounds. This phenomenon might partly be explained by limitations in sulphur pools and/or the interruption of (phytohormonal) defence signalling due to metal hyperaccumulation. This work highlights the importance of considering defences elicited by antagonists from various guilds when studying plant organic defence responses to specific abiotic environments.

Antifungal potential and defense gene induction in maize against Rhizoctonia root rot by seed extract of Ammi visnaga (L.) Lam.

Methanol extracts from five medicinal plants ( Eucalyptus tereticornis Sm., Ammi visnaga (L.) Lam., Azadirachta indica A. Juss., Rheum Palmatum L., and Adansonia digitata L.) were assessed in vitro for antifungal activity against Rhizoctonia solani Kuhn, the causal agent of Rhizoctonia root rot of maize. All tested extracts showed antifungal activity with varied extents. Ammi visnaga (khella) extract showed the greatest activity compared with the untreated experimental controls. Observations using transmission electron microscopy showed ultrastructural changes in hyphal cells as a response to exposure to khella extract. Gas chromatography-mass spectrometric analysis of khella extract showed the presence of 69 compounds. The antifungal properties of the extracts were mainly attributed to their content of coumarins and fatty acids. In the greenhouse experiment, treatment of maize plants with khella extract at 15% gave the least incidence of Rhizoctonia root rot. Results of DD-PCR showed up- and down- regulations of some genes in maize as response to the treatment with khela extract. Identification of the randomly selected genes from DD-PCR revealed that they were defense-related, as S-domain class receptor-like kinase 3 and glutathione-S-transferase1. Real-time PCR showed induction in the gene expression of the pathogenesis-related protein chitinases (2.36 fold) and thaumatin-like proteins (8.99 fold) by treatment with khella extract at 15%, which was greater expression than detected at 10 and 20%. This indicates triggering effects from the extract on the maize immune system against the R. solani infection in a concentration-dependent manner. The efficient, low-cost and eco-friendly characteristics of khella extract indicate that it could be used for the control of Rhizoctonia root rot of maize.

Functional analysis of a chitinase gene PnCHI1 from Panax notoginseng

Chitinases, a glycosidase enzyme that hydrolyzes chitin to N-acetylglucosamine, are widely found in plant cells, and they are an important part of plant antifungal defense system. The function of a Panax notoginseng chitinase gene PnCHI1 was characterized in this paper. Expression vector of PnCHI1 was constructed and transiently expressed in onion epidermal cells, and laser scanning confocal microscopy demonstrated that PnCHI1 was localized in the cell wall. Prokaryotic expression vector of PnCHI1 was also constructed, and recombinant protein of PnCHI1 was induced and purified. In vitro antibacterial assay showed that recombinant PnCHI1 protein had strong inhibitory activity on the mycelium growth of Fusarium solani, F. oxysporum and F. verticillioide. The function of PnCHI1 was further verified by reverse genetics. PnCHI1 expression vector was transferred into tobacco by Agrobacterium tumefaciens and expression of PnCHI1 was confirmed by qRT-PCR. It was found by leaf inoculation experiment that resistance of transgenic tobacco to F. solani was significantly increased. It is conclnded that: PnCHI1 is a chitinase localized in the cell wall, which inhibits several fungi which cause the root rot disease of P. notoginseng. Overexpression of this chitinase gene in tobacco greatly increased resistance to F. solani. PnCHI1 may be an important resistance gene in P. notoginseng that participates in the defense against root rot disease.

A Comprehensive in Silico Analysis of Regulatory SNPs of Human CLEC7A Gene and Its Validation as Genotypic and Phenotypic Disease Marker in Recurrent Vulvovaginal Infections.

Recurrent Vulvovaginal infections (RVVI) are the commonly reported microbiological syndrome affecting millions of women globally. Various molecules of innate immune system are instrumental in clearance of these microbial pathogens, thus suggested as one of the most important contributing factor in determining the disease outcome. Dendritic cell-associated C-type lectin-1 (Dectin-1) is an important molecule of innate immunity that is primarily known for its role in antifungal defenses. However, role of dectin-1 in recognition of other pathogens is also documented. The intracellular expression of dectin-1 was shown to be up-regulated by Mannose Binding Lectin (MBL)-mediated opsonophagocytosis of pathogens. Dectin-1 is encoded by CLEC7A, postulated to be a candidate gene in modulating risk of developing RVVI. In this study, we identified CLEC7A causal variants using in silico analysis. To assess their impact on susceptibility to RVVI, these causal variants along with serum dectin-1 levels (sDectin-1) were investigated using polymerase chain reaction-restriction fragment length polymorphism (PCR–RFLP) and Enzyme Linked Immnosorbent Assay (ELISA) respectively, under a case-control design. Furthermore, effect of these polymorphisms was also assessed on sMBL levels. In silico analysis revealed 9 putative functional conserved SNPs of CLEC7A. Association analysis revealed a significantly lower risk of developing RVVI and its types in carriers of CLEC7A rs3901533 G allele and its homozygous genotypes (p < 0.05). The heterozygous genotype was associated with significant protection against RVVI (p = 0.004). Haplotypes GGG and GTA showed significant protection against RVVI (p < 0.0001; p = 0.0003), Bacterial Vaginosis (p = 0.03; p = 0.002), Vulvovaginal Candidiasis (p = 0.03; p = 0.01) and Mixed Infections (p = 0.007; p = 0.04). Mean sDectin-1 levels were significantly high in RVVI and its types compared to controls (p < 0.05). Further, genotype-phenotype stratification showed significant differences within/between cases groups and controls. The CLEC7A rs3901533 polymorphism was also found to be associated with sMBL levels. The present study contributed novel insights into the role of dectin-1 in RVVI. CLEC7A rs3901533 polymorphism and high sDectin-1 levels along with low sMBL levels were found to be associated with RVVI susceptibility. Thus, screening of women with RVVI for these novel associations may lead to better diagnosis and treatment. Also genotyping method used in this study constitutes a simple and reliable assay, which can be confidently, used as a cheaper alternative for genotyping these variants in clinical settings. Finally, new restorative markers for other infectious diseases might be found by exploring nine functionally identified CLEC7A SNPs.

Melanin triggers antifungal defences.

Melanins are enigmatic pigments that have many roles, and the melanin in pathogenic fungi can aid host infection. Identification of a mammalian protein that recognizes melanin now reveals an antifungal defence pathway. Melanins are enigmatic pigments that have many roles, and the melanin in pathogenic fungi can aid host infection. Identification of a mammalian protein that recognizes melanin now reveals an antifungal defence pathway.

Arabidopsis MLO2 is a negative regulator of sensitivity to extracellular reactive oxygen species.

The atmospheric pollutant ozone (O3 ) is a strong oxidant that causes extracellular reactive oxygen species (ROS) formation, has significant ecological relevance, and is used here as a non-invasive ROS inducer to study plant signalling. Previous genetic screens identified several mutants exhibiting enhanced O3 sensitivity, but few with enhanced tolerance. We found that loss-of-function mutants in Arabidopsis MLO2, a gene implicated in susceptibility to powdery mildew disease, exhibit enhanced dose-dependent tolerance to O3 and extracellular ROS, but a normal response to intracellular ROS. This phenotype is increased in a mlo2 mlo6 mlo12 triple mutant, reminiscent of the genetic redundancy of MLO genes in powdery mildew resistance. Stomatal assays revealed that enhanced O3 tolerance in mlo2 mutants is not caused by altered stomatal conductance. We explored modulation of the mlo2-associated O3 tolerance, powdery mildew resistance, and early senescence phenotypes by genetic epistasis analysis, involving mutants with known effects on ROS sensitivity or antifungal defence. Mining of publicly accessible microarray data suggests that these MLO proteins regulate accumulation of abiotic stress response transcripts, and transcript accumulation of MLO2 itself is O3 responsive. In summary, our data reveal MLO2 as a novel negative regulator in plant ROS responses, which links biotic and abiotic stress response pathways.

Fungal and herbivore elicitation of the novel maize sesquiterpenoid, zealexin A4, is attenuated by elevated CO2.

Chemical isolation and NMR-based structure elucidation revealed a novel keto-acidic sesquiterpenoid, termed zealexin A4 (ZA4). ZA4 is elicited by pathogens and herbivory, but attenuated by heightened levels of CO 2 . The identification of the labdane-related diterpenoids, termed kauralexins and acidic sesquiterpenoids, termed zealexins, demonstrated the existence of at least ten novel stress-inducible maize metabolites with diverse antimicrobial activity. Despite these advances, the identity of co-occurring and predictably related analytes remains largely unexplored. In the current effort, we identify and characterize the first sesquiterpene keto acid derivative of β-macrocarpene, named zealexin A4 (ZA4). Evaluation of diverse maize inbreds revealed that ZA4 is commonly produced in maize scutella during the first 14days of seedling development; however, ZA4 production in the scutella was markedly reduced in seedlings grown in sterile soil. Elevated ZA4 production was observed in response to inoculation with adventitious fungal pathogens, such as Aspergillus flavus and Rhizopus microsporus, and a positive relationship between ZA4 production and expression of the predicted zealexin biosynthetic genes, terpene synthases 6 and 11 (Tps6 and Tps11), was observed. ZA4 exhibited significant antimicrobial activity against the mycotoxigenic pathogen A. flavus; however, ZA4 activity against R. microsporus was minimal, suggesting the potential of some fungi to detoxify ZA4. Significant induction of ZA4 production was also observed in response to infestation with the stem tunneling herbivore Ostrinia nubilalis. Examination of the interactive effects of elevated CO2 (E-CO2) on both fungal and herbivore-elicited ZA4 production revealed significantly reduced levels of inducible ZA4 accumulation, consistent with a negative role for E-CO2 on ZA4 production. Collectively, these results describe a novel β-macrocarpene-derived antifungal defense in maize and expand the established diversity of zealexins that are differentially regulated in response to biotic/abiotic stress.