Center for Innovative Biomedicine and Biotechnology

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Publications

Pyomelanin synthesis in alternaria alternata inhibits DHN-Melanin synthesis and decreases cell wall chitin content and thickness

Publication . Fernandes, Chantal; Mota, Marta; Barros, Lillian; Dias, Maria Inês; Ferreira, Isabel C.F.R.; Piedade, Ana P.; Casadevall, Arturo; Gonçalves, Teresa

The genus Alternaria includes several of fungi that are darkly pigmented by DHNmelanin. These are pathogenic to plants but are also associated with human respiratory allergic diseases and with serious infections in immunocompromised individuals. The present work focuses on the alterations of the composition and structure of the hyphal cell wall of Alternaria alternata occuring under the catabolism of L-tyrosine and L-phenylalanine when cultured in minimal salt medium (MM). Under these growing conditions, we observed the released of a brown pigment into the culture medium. FTIR analysis demonstrates that the produced pigment is chemically identical to the pigment released when the fungus is grown in MM with homogentisate acid (HGA), the intermediate of pyomelanin, confirming that this pigment is pyomelanin. In contrast to other fungi that also synthesize pyomelanin under tyrosine metabolism, A. alternata inhibits DHN-melanin cell wall accumulation when pyomelanin is produced, and this is associated with reduced chitin cell wall content. When A. alternata is grown in MM containing L-phenylalanine, a L-tyrosine percursor, pyomelanin is synthesized but only at trace concentrations and A. alternata mycelia display an albino-like phenotype since DHN-melanin accumulation is inhibited. CmrA, the transcription regulator for the genes coding for the DHN-melanin pathway, is involved in the down-regulation of DHN-melanin synthesis when pyomelanin is being synthetized, since the CMRA gene and genes of the enzymes involved in DHN-melanin synthesis pathway showed a decreased expression. Other amino acids do not trigger pyomelanin synthesis and DHN-melanin accumulation in the cell wall is not affected. Transmission and scanning electron microscopy show that the cell wall structure and surface decorations are altered in L-tyrosine- and L-phenylalanine-grown fungi, depending on the pigment produced. In summary, growth in presence of L-tyrosine and L-phenylalanine leads to pigmentation and cell wall changes, which could be relevant to infection conditions where these amino acids are expected to be available.

2021Journal article

Open access

Exploring the potential of Hermetia illucens larvae extracts: A promising approach for dermocosmetic formulations

Publication . Filipe, Márcia Santos; Cardoso, Rossana V.C.; Ayuso, Manuel; Murta, Daniel; Díaz-Lanza, Ana M.; Rosado, Catarina; Pires, Tânia C.S.P.; Calhelha, Ricardo C.; Rijo, Patrícia

Globally, the yearly disposal of 1.3 billion tonnes of food raises environmental and public health concerns. Black soldier fly (BSF) larvae present a sustainable solution, converting organic waste into nutrient-rich biomass. The extracted oil from BSF larvae, rich in fatty acids (FA), offers an eco-friendly alternative for the cosmetic industry. In this study, larvae sourced from a Portuguese company were fed olive pomace, a by-product of olive oil production. The lipidic sample extracted revealed a composition high in oleic acid, valuable for cosmetics. Investigating the biological activity of lipid extractions from larvae fed with olive pomace is a novel approach. Notably, the n-hexane ultrasound-assisted extraction method demonstrated potent antioxidant properties, and some extracts displayed antimicrobial activity. Five non-cytotoxic extracts; three with no relevant activity (IC50 from 236 to >400 mu g/mL). These findings highlight BSF larvae as an environmentally friendly source of fatty acids, offering promising alternatives for diverse applications.

2024Journal article

Open access

Escherichia coli phylogenetic and antimicrobial pattern as an indicator of anthropogenic impact on threatened freshwater mussels

Publication . Varandas, Simone; Fernandes, Conceição; Cabecinha, Edna; Gomes, Sónia; Silva, Gabriela Jorge da; Saavedra, Maria José

Freshwater bivalves are widely used as accumulation indicators and monitoring tools for assessing contaminant effects on different levels of biological integration. This pilot study aimed to explore the phylogenetic diversity of Escherichia coli isolated from freshwater mussels (Margaritifera margaritifera and Potomida littoralis) and characterize their phenotypes and antibiotic resistance profiles. Samples were collected in the Rabaçal and Tua Rivers, in the Douro basin, Portugal—two sites representing different levels of anthropogenic contamination. Antimicrobial susceptibility testing was performed via the disk diffusion method with 21 antibiotics. Results showed that 31% of isolates were multidrug-resistant (MDR). Thus, freshwater mussels provide an effective and time-integrated approach for identifying/quantifying fecal indicators, including MDR bacteria. PCR-based assays were designed for assessing phylogenetic E. coli groups. Among the E. coli isolates, the highest prevalence (44%) was observed in group D or E, followed by group E or Clade I (25%), group A (19%), and group B1 (13%). E. coli isolated from M. margaritifera predominantly exhibited a higher prevalence of phylogroups D or E, whereas E. coli from P. littoralis showed associations with phylogroups E or clade I, B1, A, and D or E. Our results provide new insights into the phylogenetic diversity of E. coli in freshwater bivalves. Additionally, the findings highlight the possible linkage of phylogroups with the host species, the geographical location in the water stream, and human activity. Using E. coli as a bioindicator isolated from freshwater mussels helps us grasp how human activities affect the environment. This study has important implications for those interested in safeguarding water resources, especially in tackling antibiotic resistance in aquatic ecosystems.

2023Journal article

Open access

Essential oils from Côa Valley Lamiaceae species: cytotoxicity on glioblastoma cells

Publication . Marques, Mário Pedro; Delgado, Jéssica; Zuzarte, Mónica; Varela, Carla; Dias, Maria Inês; Barros, Lillian; Magalhães, Mariana; Cabral, Célia

Lavandula pedunculata (Mill.) Cav., Mentha cervina L. and Thymus mastichina (L) L. subsp. mastichina are widely used and fairly representative species of the flora and traditional uses of the Côa Valley, a Portuguese UNESCO World Heritage Site. L. pedunculata and T. mastichina are used by local populations to preserve olives, as a condiment in traditional cuisine and to aromatize bonfires on Saint John's Eve, while M. cervina is mainly used as a spice for river fish dishes. However, despite their popularity among the population and prevalence in the territory of Côa Valley, these aromatic plants are still undervalued and scientific literature on their potential as anti-cancer agents is scarce. Therefore, in this work we aim to study the therapeutic potential of the essential oils (EO) obtained from these three species, by assessing their chemical composition, secretory structures morphology and cytotoxic effect against glioblastoma cell lines

2022Conference object

Open access

Chemical Characterization and Differential Lipid-Modulating Effects of Selected Plant Extracts from Côa Valley (Portugal) in a Cell Model for Liver Steatosis

Publication . Amorim, Ricardo; Marques, Mário Pedro; Melim, Catarina; Varela, Carla; Sardão, Vilma A.; Teixeira, José; Dias, Maria Inês; Barros, Lillian; Oliveira, Paulo J.; Cabral, Célia

Côa Valley, located in the northeast of Portugal, harbors more than 500 medicinal plant species. Among them, four species stand out due to their traditional uses: Equisetum ramosissimum Desf. (hemorrhages, urethritis, hepatitis), Rumex scutatus L. subsp. induratus (Boiss. and Reut.) Malag. (inflammation, constipation), Geranium purpureum Vill., and Geranium lucidum L. (pain relief, gastric issues). Given their rich ethnomedicinal history, we evaluated their protective effects on an in vitro model of metabolic dysfunction-associated steatotic liver disease (MASLD). Methods: Decoction (D) and hydroalcoholic (EtOH80%) extracts were prepared and chemically characterized. Their safety profile and effects on lipid accumulation were assessed in palmitic acid (PA)-treated HepG2 cells using resazurin, sulforhodamine B, and Nile Red assays. Results: Chemical analysis revealed diverse phenolic compounds, particularly kaempferol derivatives in E. ramosissimum. All extracts showed minimal cytotoxicity at 25–50 µg/mL. At 100 µg/mL, only E. ramosissimum extracts maintained high cell viability. In the lipotoxicity model, E. ramosissimum decoction demonstrated the most potent effect, significantly reducing PA-induced neutral lipid accumulation in a dose-dependent manner, while other extracts showed varying degrees of activity. Conclusions: These findings highlight E. ramosissimum’s decoction, rich in kaempferol derivatives, as particularly effective in reducing lipid accumulation in this MASLD cell model while also providing a comprehensive characterization of traditionally used plants from the Côa Valley region.

2025Journal article

Open access