Trichoderma stromaticum spores induce autophagy and downregulate inflammatory mediators in human peripheral blood-derived macrophages.

Trichoderma spp. are usually considered safe and normally used as biocontrol and biofertilization. Safety for human health is evaluated by several tests that detect various effects such as allergenicity, toxicity, infectivity, and pathogenicity. However, they do not evaluate the effects of the agent upon the immune system. The aim of this study was to investigate the interaction between T. stromaticum spores and mammalian cells to assess the immunomodulatory potential of the spores of this fungus. First, mouse macrophage cell line J774 and human macrophages were exposed to fungal spores and analyzed for structural features, through scanning and transmission electron microscopy. Then, various analysis were performed in human macrophages as to their effect in some functional and molecular aspects of the immune system through immunocytochemistry, flow cytometry and gene expression assays. We demonstrated that T. stromaticum spores induces autophagy and autophagy-related genes (ATGs) and downmodulate inflammatory mediators, including ROS, NLRP3, the cytokines IL-1ß, IL-18, IL-12 and IL-10, as well as TLR2, TLR4, miR-146b and miR-155, which may lead to an augmented susceptibility to pathogens. Our study shows the extension of damages the biofungicide Tricovab® can cause in the innate immune response. Further studies are necessary to elucidate other innate and adaptive immune responses and, consequently, the safety of this fungus when in contact with humans.

Toxoplasma gondii induces extracellular traps release in cat neutrophils.

Neutrophils respond differently to violations of the body's physiological barriers during infections. Extracellular traps comprise one of the mechanisms used by these cells to reduce the spread of pathogens to neighboring tissues, as well as ensure a high concentration of antimicrobial agents at the site of infection. To date, this innate defense mechanism has not been previously demonstrated in neutrophils of cats exposed to Toxoplasma gondii. The aim of this study was to characterize the in vitro release of neutrophil extracellular traps (NETs) when neutrophils isolated from cats were exposed to T. gondii. First, cellular viability was tested at different time points after parasite exposure. The production of reactive oxygen species (ROS) and lactate dehydrogenase and the amount of extracellular DNA were quantified. In addition, the number of parasites associated with neutrophils was determined, and the observed NETs formed were microscopically characterized. Results showed that (i) in culture, neutrophils isolated from cats presented diminished cellular viability after 4 h of incubation, and when neutrophils were incubated with T. gondii, they displayed cytotoxic effects after 3 h of interaction; (ii) neutrophils were able to release structures composed of DNA and histones, characterized as NETs under optical, immunofluorescence, and electron scanning microscopy, when stimulated with T. gondii; (iii) only 11.4% of neutrophils were able to discharge NETs during 3 h of incubation; however, it was observed through extracellular quantification of DNA that this small number of cells were able to display different behavior compared to a negative control (no parasite) group; (iv) significant differences in ROS production were observed in neutrophils exposed to T. gondii. In conclusion, our results showed that neutrophils isolated from cats exposed to T. gondii release structures composed of DNA and histones, similar to what has already been described in other neutrophil species infected with the parasite.