Chick heterophils release DNA extracellular traps (DETs) in vitro and in vivo upon Aspergillus fumigatus conidia exposure.

Aspergillosis is a common fungal disease in avian species, causing high mortality in young chicks in agricultural farms and yards. It is caused by fungi belonging to the genus Aspergillus. Aspergillosis occurs by inhalation of fungal conidia, and in chickens, effective infection control relies on a rapid and large influx of heterophils to the lungs. Heterophils, upon different stimuli, release to the extracellular milieu their chromatin associated with several proteins that ensnare and kill different pathogens similarly to neutrophil extracellular traps. Here, we showed that Aspergillus fumigatus conidia and the peptidogalactomannan (PGM), isolated from the fungus cell wall, induce the release of DNA extracellular traps (DETs) in chicks' blood and lung heterophils. We demonstrated that reactive oxygen species, elastase and peptidyl arginine deiminase (PAD) were involved in DETs extrusion, the occurrence of DETs in the lungs of A. fumigatus-exposed chicks in vivo, and its role in chick survival. These results may contribute to developing more efficient tools for the therapeutic and diagnosis of aspergillosis.

Cooperation between apoptotic and viable metacyclics enhances the pathogenesis of Leishmaniasis.

Mimicking mammalian apoptotic cells by exposing phosphatidylserine (PS) is a strategy used by virus and parasitic protozoa to escape host protective inflammatory responses. With Leishmania amazonensis (La), apoptotic mimicry is a prerogative of the intramacrophagic amastigote form of the parasite and is modulated by the host. Now we show that differently from what happens with amastigotes, promastigotes exposing PS are non-viable, non-infective cells, undergoing apoptotic death. As part of the normal metacyclogenic process occurring in axenic cultures and in the gut of sand fly vectors, a sub-population of metacyclic promastigotes exposes PS. Apoptotic death of the purified PS-positive (PS(POS)) sub-population was confirmed by TUNEL staining and DNA laddering. Transmission electron microscopy revealed morphological alterations in PS(POS) metacyclics such as DNA condensation, cytoplasm degradation and mitochondrion and kinetoplast destruction, both in in vitro cultures and in sand fly guts. TUNEL(POS) promastigotes were detected only in the anterior midgut to foregut boundary of infected sand flies. Interestingly, caspase inhibitors modulated parasite death and PS exposure, when added to parasite cultures in a specific time window. Efficient in vitro macrophage infections and in vivo lesions only occur when PS(POS) and PS-negative (PS(NEG)) parasites were simultaneously added to the cell culture or inoculated in the mammalian host. The viable PS(NEG) promastigote was the infective form, as shown by following the fate of fluorescently labeled parasites, while the PS(POS) apoptotic sub-population inhibited host macrophage inflammatory response. PS exposure and macrophage inhibition by a subpopulation of promastigotes is a different mechanism than the one previously described with amastigotes, where the entire population exposes PS. Both mechanisms co-exist and play a role in the transmission and development of the disease in case of infection by La. Since both processes confer selective advantages to the infective microorganism they justify the occurrence of apoptotic features in a unicellular pathogen.