Candida albicans is a major cause of human infections, ranging from relatively simple to treat skin and mucosal diseases to systemic life-threatening invasive candidiasis. Fungal infections treatment faces three major challenges: the limited number of therapeutic options, the toxicity of the available drugs, and the rise of antifungal resistance. In this study, we demonstrate the antifungal activity and mechanism of action of peptides ToAP2 and NDBP-5.7 against planktonic cells and biofilms of C. albicans. Both peptides were active against C. albicans cells; however, ToAP2 was more active and produced more pronounced effects on fungal cells. Both peptides affected C. albicans membrane permeability and produced changes in fungal cell morphology, such as deformations in the cell wall and disruption of ultracellular organization. Both peptides showed synergism with amphotericin B, while ToAP2 also presents a synergic effect with fluconazole. Besides, ToAP2 (6.25 µM.) was able to inhibit filamentation after 24 h of treatment and was active against both the early phase and mature biofilms of C. albicans. Finally, ToAP2 was protective in a Galleria mellonella model of infection. Altogether these results point to the therapeutic potential of ToAP2 and other antimicrobial peptides in the development of new therapies for C. albicans infections. Candida albicans is a fungal species present in the normal human microbiota, colonizing several areas of the body. However, under certain circumstances, this species may become a pathogen, causing diseases that can be life-threatening 1-4. The use of broad-spectrum antibiotics, immune suppression, or changes in the local host environments are examples of situations that may favor the proliferation of C. albicans and the onset of disease 5-8. Moreover, C. albicans ability to thrive in human tissues involves metabolic and morphological changes associated with the expression of different virulence factors 9. C. albicans virulence factors include secretion of enzymes, adhesion to cell surfaces and evasion of the immune system 10,11. Two virulence factors of major clinical importance are the fungal polymorphism and its ability to form biofilms 12-14. C. albicans ability to transit between yeast and filamentous forms is crucial for pathogenesis and both fungal forms are relevant for infection 15. For instance, hyphae have a major role on tissue invasion, whereas the yeast morphology facilitates fungal dispersion 16. The different fungal morphologies are also important for the formation of C. albicans biofilms 17. Living in biofilms confers to the microorganisms several advantages, when compared to the planktonic lifestyle, including protection against immune cells, increased resistance to antimicrobials agents and other chemical, physical and environmental stressors 18,19 .

Paracoccidioides spp. are thermodimorphic fungi that cause a neglected tropical disease (paracoccidioidomycosis) that is endemic to Latin America. These fungi inhabit the soil, where they live as saprophytes with no need for a mammalian host to complete their life cycle. Despite this, they developed sophisticated virulence attributes allowing them not only to survive in host tissues but also to cause disease. A hypothesis for selective pressures driving the emergence or maintenance of virulence of soil fungi is their interaction with soil predators such as amoebae and helminths. We evaluated the presence of environmental amoeboid predators in soil from armadillo burrows where Paracoccidioides had been previously detected and tested if the interaction of Paracoccidioides with amoebae selects for fungi with increased virulence. Nematodes, ciliates, and amoebae–all potential predators of fungi–grew in cultures from soil samples. Microscopical observation and ITS sequencing identified the amoebae as Acanthamoeba spp, Allovahlkampfia spelaea, and Vermamoeba vermiformis. These three amoebae efficiently ingested, killed and digested Paracoccidioides spp. yeast cells, as did laboratory adapted axenic Acanthamoeba castellanii. Sequential co-cultivation of Paracoccidioides with A. castellanii selected for phenotypical traits related to the survival of the fungus within a natural predator as well as in murine macrophages and in vivo (Galleria mellonella and mice). These changes in virulence were linked to the accumulation of cell wall alpha-glucans, polysaccharides that mask recognition of fungal molecular patterns by host pattern recognition receptors. Altogether, our results indicate that Paracoccidioides inhabits a complex environment with multiple amoeboid predators that can exert selective pressure to guide the evolution of virulence traits.

Amphibian skin is a multifunctional organ that plays key roles in defense, breathing, and water balance. In this study, skin secretion samples of the fire salamander (Salamandra salamandra) were separated using RP-HPLC and de novo sequenced using MALDI-TOF MS/MS. Next, we used an in silico platform to screen antioxidant molecules in the framework of density functional theory. One of the identified peptides, salamandrin-I, [M + H]+ = 1406.6 Da, was selected for solid-phase synthesis; it showed free radical scavenging activity against DPPH and ABTS radicals. Salamandrin-I did not show antimicrobial activity against Gram-positive and -negative bacteria. In vitro assays using human microglia and red blood cells showed that salamandrin-I has no cytotoxicity up to the concentration of 100 µM. In addition, in vivo toxicity tests on Galleria mellonella larvae resulted in no mortality at 20 and 40 mg/kg. Antioxidant peptides derived from natural sources are increasingly attracting interest. Among several applications, these peptides, such as salamandrin-I, can be used as templates in the design of novel antioxidant molecules that may contribute to devising strategies for more effective control of neurological disease.

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RESUMO:A qualidade microbiológica do leite cru e do queijo coalho está diretamente relacionada com a produção e comercialização do produto. Esta pesquisa objetivou avaliar as condições higiênico-sanitárias por meio da detecção de microrganismos indicadores de contaminação de leite cru e queijo coalho comercializados em mercados públicos da cidade de Parnaíba, Piauí. Foram coletadas 16 amostras de leite cru e 16 amostras de queijo coalho (8 conservadas por refrigeração e 8 sem conservação) em quatro mercados públicos distintos, no período de abril a outubro de 2012. Na análise do leite cru, 11 das 16 amostras apresentaram números elevados tanto de coliformes totais como termotolerantes. Com relação à análise do queijo coalho, das 16 amostras 12 apresentavam índices elevados de coliformes totais, sendo que 11 estavam em desacordo com os padrões estabelecidos pela lei vigente da ANVISA. A conservação do queijo também foi avaliada como um parâmetro. Os resultados obtidos mostraram que o leite cru e o queijo coalho, comercializados na cidade de Parnaíba, estão sendo manipulados sob condições higiênico-sanitárias insatisfatórias, resultando em um produto final com elevados números de microrganismos indicadores de contaminação, ocasionando um risco para a saúde pública. PALAVRAS-CHAVE:Coliformes; Contaminação de Alimentos; Laticínios. EVALUATION OF HYGIENE-SANITARY CONDITIONS OF RAW MILK AND RENNET CHEESE SOLD IN PUBLIC MARKETS IN NORTHERN PIAUÍ, BRAZILABSTRACT: The microbiological quality of raw milk and rennet cheese is directly related to the production and commercialization of the product. Current research evaluated the hygiene-sanitary conditions by detecting microorganisms that indicate contamination of raw milk and rennet cheese commercialized in public markets in Parnaíba PI Brazil. Sixteen samples of raw milk and 16 samples of rennet cheese (8 samples kept under refrigeration and 8 without) were collected in four different public outlets between April and October 2012. Eleven out of 16 samples of raw milk had a high number of total and thermotolerant coliforms. Twelve out of the 16 samples of rennet cheese had high total coliforms, with eleven exceeding standards by the health department ANVISA. Cheese conservation was also assessed and results revealed that raw milk and rennet cheese are commercialized in Parnaiba without hygiene and sanitary precautions. The final product exhibited a high number of microorganisms indicating contamination, with great risks for public health.

Tryptophyllins constitute a heterogeneous group of peptides that are one of the first classes of peptides identified from amphibian's skin secretions. Here, we report the structural characterization and antioxidant properties of a novel tryptophyllin-like peptide, named PpT-2, isolated from the Iberian green frog Pelophylax perezi. The skin secretion of P. perezi was obtained by electrical stimulation and fractionated using RP-HPLC.De novo peptide sequencing was conducted using MALDI MS/MS. The primary structure of PpT-2 (FPWLLS-NH 2 ) was confirmed by Edman degradation and subsequently