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- Fármacos en el agua: Sensibilización para la mitigación de sus impactos en los ecosistemas y en la sociedadPublication . Voznakova, Alena; Canle, Moisés; Geraldes, Ana MariaLa creciente presencia de residuos farmacéuticos en los ecosistemas acuáticos ha emergido como un problema ambiental significativo y poco visibilizado. Fármacos de uso común, como antibióticos, analgésicos y betabloqueantes, ingresan al ambiente a través de aguas residuales, desechos industriales y prácticas inadecuadas de disposición de medicamentos, persistiendo en cuerpos de agua superficiales y subterráneos. Los sistemas de tratamiento de aguas convencionales no están diseñados para eliminar completamente estos compuestos, lo que provoca una acumulación gradual con efectos adversos tanto en la biodiversidad y los ecosistemas asociados. Este artículo aborda los efectos de los fármacos en organismos acuáticos, tales como la toxicidad combinada y el desarrollo de resistencias antimicrobianas. Además, se explora el impacto a largo plazo en los ecosistemas, y los riesgos asociados para la salud humana. Finalmente, se subraya el rol fundamental de la educación ambiental en la creación de conciencia pública sobre los impactos de los residuos farmacéuticos y la importancia de promover cambios en los comportamientos sociales.
- Projeto Bio Ilhas: Conectando Ciência e Educação para sensibilizar para a importância da conservação dos riosPublication . Ferreira, Nathália Borges Bartoli; Sá, Isabel; Nogueira, Clotilde; Fachada, Ivone; Calheiros, Cristina; Geraldes, Ana MariaAs Soluções Baseadas na Natureza são uma estratégia crucial para a resiliência e qualidade ambiental, proporcionando benefícios ambientais, sociais, e económicos e educacionais. Estas tecnologias consistem em estratégias que utilizam processos naturais para melhorar e promover a biodiversidade e mitigar impactos antropogénicos. Um exemplo prático destas soluções são as ilhas flutuantes. Estas estruturas são projetadas para melhorar a qualidade da água em corpos hídricos degradados. Elas atuam como filtros naturais, removendo poluentes, promovendo a biodiversidade e criando habitats em áreas impactadas pela atividade humana, podendo também ser um recurso educacional para promover o conhecimento das espécies associadas aos ecossistemas aquáticos. Assim, o presente trabalho tem como objetivo apresentar o projeto Bio Ilhas. No âmbito deste projeto foram instaladas ilhas flutuantes piloto nos Rios Fervença (Bragança) e no Rio Côa (nas próximidades do Museu Côa-Parque) com a área total de 4 e 3 m2 respetivamente. Este projeto visa não só avaliar a eficácia e o potencial desta Solução Baseada na Natureza numa eventual reabilitação futura destes ecossistemas, mas também exemplificar como essas integrações da natureza em ambientes urbanos podem melhorar a qualidade de vida dos cidadãos, promover a biodiversidade e potencializar o contexto educativo, envolvendo a população em processos interativos de aprendizagem ambiental.
- Synthesis of carbon nanotubes from polyolefin waste: mass balance and gas phase compositionPublication . Roman, Fernanda F.; Silva, Adriano S.; Gomes, Helder T.The valorization of plastic solid waste (PSW) through thermochemical routes has emerged as a sustainable strategy to address the growing accumulation of these materials. Among the most attractive products, carbon nanotubes (CNTs) stand out due to their high added-value and potential applications in catalysis, sensors, and environmental remediation. CNT synthesis from plastics typically involves thermal decomposition of the polymer into a carbon-rich gas fraction, followed by its deposition on metallic nanoparticles to form CNTs. Optimizing this process requires a detailed understanding of both the mass balance and gas composition. In this work, CNTs were synthesized by chemical vapor deposition (CVD) from low-density polyethylene (LDPE), high-density polyethylene (HDPE), and polypropylene (PP) using a nickel–iron (NiFe) catalyst (synthesis details presented elsewhere [1]). The experiments were performed in a one-chamber reactor, with 1 g of plastic loaded in the upper zone (400–450 °C) and 0.2 g of NiFe in the lower zone (850 °C). The system was heated for 1 h and then held for 0.5 h under a N2 flow (50 cm3 min-1). Generated gases were analyzed in situ by gas chromatography with flame ionization detection (GC-FID). After reaction, solid and liquid fractions were collected and weighed. Characterization of similar CNTs can be consulted in previous publications [1-3]. The results are summarized in Fig. 1. As shown in Fig. 1a, the solid fraction yields were similar for all polymers (35–38%), while HDPE produced a higher liquid fraction than LDPE and PP. The gas fraction dominated in all cases, suggesting catalyst deactivation or an inadequate catalyst-to-plastic ratio. Three major gases were detected (Fig. 1b): methane >> propylene > ethylene. LDPE and PP showed comparable gas distributions, whereas HDPE generated markedly less methane and propylene, consistent with the mass balance trends.
- Human-mediated rapid evolutionary change in european honey beesPublication . Li, Fernanda; Taliadoros, Demetris; Costa, Maíra; Yadró Garcia, Carlos A.; Cunha, Larissa; Henriques, Dora; Martin-Hernandez, Raquel; de Graaf, Dirk; Webster, Matthew; Pinto, M. AliceIn its vast distributional range, spanning Africa, Europe, the Middle East, and western Asia, the honey bee Apis mellifera diversified into 30 subspecies grouped into four major evolutionary lineages. Two of these lineages, M (western/northern European) and C (southeastern European), are parapatric in Europe. However, increasingly intensified queen trading is likely eroding the natural genetic boundaries and altering the continent’s diversity patterns. To evaluate the impact of this recent human-mediated gene flow, we conducted an unprecedented survey spanning 33 countries and sampling more than 1,300 colonies, including 139 from conservation apiaries of the M-lineage subspecies A. m. mellifera. We used a dual-marker approach combining the hypervariable mitochondrial tRNALeu–cox2 intergenic region with nuclear genome-wide single-nucleotide polymorphisms (SNPs). Both markers were highly concordant at the lineage level and European scale, showing that in the native area of M-lineage, which covers western and northern Europe, C-lineage ancestry is now predominant. This pattern is congruent with widespread commercial dissemination of C-lineage subspecies (A. m. carnica, A. m. ligustica), which is leading to introgressive hybridisation and, in many regions, to almost complete replacement of native subspecies. The exceptions to this trend are the Iberian Peninsula, Ireland, and conservation apiaries, which retain almost exclusively native M-lineage ancestry. Remarkably, even within the native C-lineage range in the Mediterranean and southeastern Europe, the Italian subspecies A.m. ligustica, the most widely favoured subspecies worldwide, shows worrying levels of introgression from its C-lineage neighbour A.m. carnica. Equally striking is the widespread presence of African-lineage mitotypes, whose routes of introduction remain unclear. Altogether, these findings raise serious concerns about the genetic integrity of native subspecies and the consequences of admixture for adaptation in a rapidly changing environment shaped by climate change and emerging parasites and pathogens. Further, these changes may affect the gene pools of wild A. mellifera populations, recently classified as Endangered by the IUCN.
- Harnessing mitochondrial divergence in Apis mellifera to determine the origin of mediterranean honeyPublication . Honrado, Mónica; Henriques, Dora; Santos, Joana; Yadró Garcia, Carlos A.; Pinto, M. Alice; Amaral, Joana S.The Mediterranean region holds one of the highest levels of Apis mellifera subspecific diversity, exhibiting a rich evolutionary history shaped by long-term geographic isolation, climatic variation, and, more recently, by human-driven processes. As part of the international project MEDIBEES, we developed a DNA-metabarcoding approach to characterise the entomological origin of honey, focusing on mitochondrial lineages A (African), M (Western European), C (Eastern European), and O (Middle Eastern). To build a robust reference database, DNA was extracted from 1280 honeybees representing 16 A. mellifera subspecies (A.m. sahariensis, A.m. intermisa, A.m. siciliana, A.m. ruttneri, A.m. iberiensis, A.m. ligustica, A.m. macedonica, A.m. adami, A.m. cecropia, A.m. cypria, A.m. caucasica, A.m. meda, A.m. anatoliaca, A.m. syriaca, A.m. jemenitica, A.m. lamarcki) and whole genomes were sequenced. Mitochondrial genomes were assembled using MitoZ, resulting in 769 assemblies, which were individually aligned against the reference mitochondrial genome using MEGA. Only mitogenomes corresponding to native subspecific ancestry were retained, producing a final curated database of 355 sequences. This database was then used to calculate the fixation index (FST) pairwise values, and a 400 bp sliding window was used to identify single-nucleotide polymorphisms (SNPs) that effectively differentiate (FST>0.98) the four lineages. Three promising regions emerged for primer design: one in the COI gene, one in the ND1 gene, and one in the CYTB gene. The resulting primers were first validated using 36 honeybee samples of known lineage representing all 16 subspecies and subsequently applied to 83 honey samples from Lebanon, Jordan, Türkiye, Italy, Algeria, Malta, UAE, and Oman. Among the three markers, ND1 showed the highest discriminatory power, correctly assigning all reference honeybee samples to their respective lineage. The COI and CYTB primers also showed strong potential, and the observed error rates (5.8% in COI and 1.7% for CYTB) provide useful thresholds for interpreting the DNA-metabarcoding results in honey samples. Overall, this study demonstrates how patterns of mitochondrial divergence shaped by long-term evolutionary processes can be harnessed for applied DNA-metabarcoding on food authenticity.
- Adaptive genomic variation in honey bees from arid regionsPublication . Yadró Garcia, Carlos A.; Henriques, Dora; Haddad, N.; Obeidat, W. M.; AlShagour, B.; Muz, Mustafa Necati; Arab, A.; Eissa, A. A.; Hosri, C.; Lamghari, F.; Arruda, J.; Rufino, José; Martín Hernández, R.; Nanetti, A.; Pinto, M. AliceThe Middle East is home to a large number of Apis mellifera subspecies that have evolved under extreme aridity and desertic conditions. Therefore, they represent a unique opportunity to disentangle the genetic basis of adaptation to such challenging environmental conditions. We arranged a collection of 514 drone samples (haploid males) split into two latitudinal transects. Transect 1 comprised 342 samples: 66 A. m. lamarckii (Egypt), 197 A. m. syriaca (Jordan), 30 A. m. syriaca (Lebanon), 21 A. m. meda (southeastern Turkey), and 27 A. m. anatoliaca (Anatolia, Turkey). Transect 2 comprised 173 samples: 14 A. m. jemenitica (Oman), 9 A. m. jemenitica (UAE), 75 A. m. meda (Iran), and 75 A. m. caucasia (northeastern Turkey). For all samples, whole genomes were resequenced, and environmental and bioclimatic variables were retrieved from WorldClim. For each transect, we applied three genotype–environment association methods (Samβada, pRDA, and LFMM) and one outlier-detection approach (PCAdapt) to identify SNPs associated with environmental adaptation. SNPs were annotated, and the intersection of genes between the four methods in each transect was analysed for gene enrichment. In both transects, enriched terms included genes related to biological regulation through endocrine, neuroendocrine, and neurotransmitter pathways. The regulation of gene expression was also enriched, including several transcription factors, especially HOX genes. Transmembrane and ionic transporters also seem to play a central role in environmental adaptation in these subspecies. Finally, neurogenesis, synapse establishment, and neural system development were also enriched. When gene enrichment analysis was set to exclude electronic annotations (automatically assigned, non-curated GO terms), enriched terms included processes such as behaviour, learning or memory, cognition, and associative and olfactory learning and behaviour. Our findings point to a complex suite of regulatory and sensory 62 pathways that collectively shape the environmental adaptation of honey bee subspecies that have evolved in arid environments.
- Bioinformatics pipeline to evaluate patterns of diversity in detoxification genes in the Western honey bee (Apis mellifera)Publication . Barbosa, Daniela; Li, Fernanda; Bashir, Sana; Lopes, Ana Rita; Yadró Garcia, Carlos A.; Quaresma, Andreia; Rufino, José; Rosa-Fontana, Annelise; Verbinnen, Gilles; de Graaf, Dirk C.; De Smet, Lina; Taliadoros, Demetris; Webster, Matthew; Pinto, M. Alice; Henriques, DoraThe Western honey bee, Apis mellifera, displays significant genetic diversity in detoxification genes, which is pivotal for environmental adaptation and resilience. Herein, we developed a bioinformatics pipeline to investigate patterns of diversity in these genes, focusing on single nucleotide polymorphisms (SNPs) across A. mellifera populations, with variant annotation performed using both snpEff and the Variant Effect Predictor (VEP). Our pipeline integrates GATK, VCFtools, PLINK, bcftools, snpEff, and VEP to process genomic data systematically. Regions of interest were defined in a BED file for variant filtering. Using GATK, SNPs were extracted from a VCF file and conversion to PLINK format for population genetics analyses. Variants were filtered by minor allele frequency (MAF) and population differentiation (FST index) to identify SNPs with considerable. Variants were annotated with snpEff and VEP to predict functional impacts, enabling a comparative analysis of their annotation consistency and depth. Custom scripts were developed to map SNPs to detoxification genes, quantify SNP density, and integrated gene descriptions and lineage data. The resulting data were visualized using a combination of and generate different graphs using ggplot2 and chromoMap for chromossomal maps. Quality control steps were applied through the pipeline ensuring data reliability. Our findings reveal distinct SNP patterns in detoxification genes, highlighting candidate SNPs associated with A. mellifera subspecies-specific adaptations. The comparison of snpEff and VEP annotations provides insights into their strengths and limitations, which can help optimize software selection for genomic studies. This pipeline offers a reproducible framework for studying genetic diversity in A. mellifera that is adaptable to other species, advancing conservation and evolutionary genomics.
- Computational Insights into the Impact of NonSynonymous Mutations in the ABC Transporter Gene of Apis melliferaPublication . Bashir, Sana; Li, Fernanda; Shiraishi, Carlos S.H.; Younes, Sarra Bin; Yadró Garcia, Carlos A.; Barbosa, Daniela; Abreu, Rui M.V.; Rufino, José; Pinto, M. Alice; Henriques, DoraNon-synonymous mutations lead to amino acid substitutions that can affect a protein’s structure and function, thereby influencing biological processes such as detoxification. ATP-binding cassette (ABC) transporters (an important superfamily of detoxification genes) play a critical role in the efflux of toxic compounds in honey bees (Apis mellifera), essential for their survival in environments exposed to natural and synthetic xenobiotics. In this study, non-synonymous mutations in 21 ABC transporter genes were identified. Then, computational tools were employed to investigate the structural and functional consequences of a non-synonymous mutation in the protein encoded by the ABC transporter gene LOC411997. Protein structures were generated from a FASTA file using AlphaFold3, converted from mmCIF to PDB format, and visualised in PyMOL, where mutations were introduced. Functional site predictions were performed using Proteins Plus, and molecular dynamics simulations were conducted in YASARA to assess stability and conformational changes. Our findings suggest that a mutation that replaces threonine with isoleucine alters the protein dynamics by modifying its energy landscape and stability. The total potential energy of the wild-type protein was calculated as -3,640,842 kcal/mol. In contrast, this value increased to -3,443,695 kcal/mol for the mutant protein. This difference suggests that the mutation may affect conformation, flexibility, and biological function of the protein encoded by the ABC transporter gene LOC411997. Understanding these conformational changes at the molecular level will contribute to strategies for improving honey bee’s resilience and conservation.
- Genetic Diversity of Detoxification Genes in 18 Honey Bee SubspeciesPublication . Li, Fernanda; Barbosa, Daniela; Bashir, Sana; Moreira de Sá, Leandro; Yadró Garcia, Carlos A.; Rufino, José; Rosa-Fontana, Annelise; Verbinnen, Gilles; de Graaf, Dirk; de Smet, Lina; Taliadoros, Demetris; Webster, Matthew; Pinto, M. Alice; Henriques, DoraThe honey bees (Apis mellifera) is a key pollinator that is exposed to a wide array of xenobiotics, both natural (plant allelochemicals) and synthetic (pesticides), while foraging or through contaminated food within the hive. These compounds have both lethal and sub-lethal effects, impairing foraging activity and negatively affecting bee development and colony health. Similar to other insects, honey bees rely on detoxification pathways to metabolise xenobiotics into less toxic or more readily excretable forms. This process is a key mechanism underlying insecticide resistance and is influenced by genetic variation. Therefore, investigating polymorphisms in detoxification-related genes is a promising approach to predict species-specific responses to pesticide exposure. Five major gene families are involved in xenobiotic detoxification: cytochrome P450 monooxygenases (CYPs), carboxyl/cholinesterases (CCEs), glutathione Stransferases (GSTs), ATP-binding cassette transporters (ABCs), and uridine 5′-diphospho-glucuronosyltransferases (UGTs). In this study, we examined the genomic detoxification inventory of over 1,600 individuals representing 18 A. mellifera subspecies representing the four main evolutionary lineages. For each lineage and subspecies, single-nucleotide polymorphism (SNP) loci were identified within these genes, allele frequency and FST (fixation index) were calculated. Additionally, all variants were annotated to assess their potential impact on protein function. Findings from this study have the potential to inform breeding and conservation strategies by identifying populations more vulnerable to chemical stressors, ultimately supporting honey bee health in changing environments.
- A Computational Approach to Study Non-Synonymous MutationsPublication . Bashir, Sana; Li, Fernanda; Shiraishi, Carlos S.H.; Yadró Garcia, Carlos A.; Barbosa, Daniela; Abreu, Rui M.V.; Rufino, José; Pinto, M. Alice; Henriques, DoraNon-synonymous SNPs (Single Nucleotide Polymorphisms) result in amino acid substitutions within proteins. While some may have minimal impact on protein structure and function, others can significantly alter stability, conformation, and biological activity. Therefore, it is crucial to predict how SNP-induced changes affect protein structure and function. Here, we developed a novel pipeline that integrates bioinformatics and molecular modeling techniques to evaluate the structural and functional consequences of a non-synonymous SNP in a protein. Initially, Protein Plus was used to predict the potential active sites, which will help to determine whether a mutation is located within the functional binding regions. Identification of active sites is necessary to prioritize mutations that may alter protein structure and function. PolyPhen-2 and I-Mutant were used to evaluate the functional impact of mutations and the thermodynamic stability of the proteins. Next, to visualize the structural changes, AlphaFold3 was used to generate highconfidence 3D models for wild and mutant proteins. Finally, molecular dynamics (MD) simulations were conducted using YASARA to explore the dynamic behavior and stability of both mutant and wild-type proteins. MD simulations provide valuable insight into structural flexibility, potential conformational shifts, and the overall impact of the mutation on protein function. This computational pipeline provides a detailed framework to evaluate the structural and energetic consequences of non-synonymous mutations.