Browsing by Author "Moreira, Olga"
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- Assessment of the Transmission Dynamics of Clostridioides difficile in a Farm Environment Reveals the Presence of a New Toxigenic Strain Connected to Swine ProductionPublication . Alves, Frederico; Nunes, Alexandra; Castro, Rita; Sequeira, António; Moreira, Olga; Matias, Rui; Rodrigues, João Carlos; Silveira, Leonor; Gomes, João Paulo; Oleastro, MónicaThe recent increase in community-acquired Clostridioides difficile infections discloses the shift in this bacterium epidemiology. This study aimed at establishing a transmission network involving One Health components, as well as assessing the zoonotic potential and genomic features of dominant clones. Samples were collected from different compartments of animal, human and environmental origin, from an animal production unit. C. difficile isolates were characterized for toxigenic profile by multiplex-PCR, while genetic diversity was evaluated by PCR-ribotyping and whole genome-based analysis. The overall C. difficile prevalence was 37.2% (70/188), and included samples from environmental (58.3%, 35/60) and animal (31.5%, 35/111) compartments; human samples (n = 17) taken from healthy workers were negative. A predominant clone from RT033 was found in almost 90% of the positive samples, including samples from all compartments connected to the pig production unit, with core-genome single nucleotide variant (SNV)-based Analysis supporting a clonal transmission between them (mean distance of 0.1 ± 0.1 core-SNVs). The isolates from this clone (herein designated PT RT033) were positive for all C. difficile toxin genes (tcdA, tcdB, cdtA/cdtB). The phyloGenetic positioning of this clone was clearly distinct from the classical RT033 cluster, suggesting a different evolutionary route. This new clone shares genomic features with several RTs from the clade 5 Sequence Type (ST) 11, including a complete pathogenicity locus (PaLoc) that is more similar to the one found in toxigenic strains and contrasting to the less virulent classical RT033 (tcdA-, tcdB-, cdtA + /cdtB +). The presence of a tcdA gene truncated into two ORFs, not previously described, requires further evaluation concerning toxin functionality. We hypothesize that the unique combination of genetic elements found in the PT RT033 clone may contribute to host tropism and environmental dissemination and maintenance. This study constitutes the first report of a toxigenic RT033 clone and adds to the overall knowledge on Clade 5 sequence type 11, considered the C. difficile evolutionary lineage with the highest zoonotic potential. The presence of this clone in all compartments associated with the pig production unit suggests a transmission chain involving these animals and contributes to unveil the role played by animal and environmental reservoirs in this pathogen epidemiology.
- Diversity of bacterial communities and genes encoding AMR in different environmental compartments along the food/feed chainPublication . Manageiro, Vera; Cano, Manuela; Dias, Elsa; Rosado, Tânia; Vieira, Patrícia; Reis, Lígia; Matias, Rui; Rodrigues, João; Coelho, Rodrigo; Menezes, Carina; Ferreira, Eugénia; Sequeira, António; Moreira, Olga; Caniça, ManuelaAim: The aim of this study was to determine the microbial biodiversity and naturally occurring antibiotic resistance (AR)-encoding genes background load along the food/feed chain in an open-air agricultural testing catchment (HOAL). Methods: Samples were collected in two time points during the year 2020 from various – but interconnected – environmental compartments of the food/feed chain within an HOAL catchment, such as air, pig feces, manure, soil, water, crops, and feed. All samples were homogenized, diluted and plated in selective media. Colonies were selected andidentification made by MALDI-TOF. Antibiotic susceptibility (AST) was assessed by disc diffusion and/or MIC methods. DNA was extracted for further molecular and genomic characterization. Results: A total of 753 Gram-negative strains were isolated. The highest number of bacteria were from families of Enterobacteriaceae(44.1%), Morganellaceae(15.5%), Moraxellaceae(15.5%), and Pseudomonadaceae(9.3%). Enterobacteriaceae and Pseudomonadaceae were present in all but one of the tested compartments (ground water and feed, respectively). Of notice, 43.1% of Enterobacteriaceae identified were isolated from a selective medium supplemented with colistin 0.5mg/L and 8.7% were selected with cefotaxime 2mg/L. Extended-spectrum -lactamase (ESBL)-producing strains were identified. Regarding Gram-positive bacteria, we identified mainly Enterococcaceae (46.7%), Bacillaceae (30.6%), and Streptococcaceae (13.9%). Pig faces and manure were the compartments with higher bacterial biodiversity. Phenotypic AST revealed the presence of resistances against several AR classes. Based on this data, strains were classified as multidrug-resistant. Conclusions: The investigation of clinically important AR-encoding genes by PCR-amplification and whole-genome-sequencing of selected strains will contribute to clarify the resistome and microbial biodiversity in the tested environmental compartments.
- Quantitative Proteomics Identifies Metabolic Pathways Affected by Babesia Infection and Blood Feeding in the Sialoproteome of the Vector Rhipicephalus bursaPublication . Couto, Joana; Villar, Margarita; Mateos-Hernández, Lourdes; Ferrolho, Joana; Sanches, Gustavo; Sofia Santos, Ana; Santos-Silva, Maria; Nobre, João; Moreira, Olga; Antunes, Sandra; de la Fuente, José; Domingos, AnaThe negative impact of ticks and tick-borne diseases on animals and human health is driving research to discover novel targets affecting both vectors and pathogens. The salivary glands are involved in feeding and pathogen transmission, thus are considered as a compelling target to focus research. In this study, proteomics approach was used to characterize Rhipicephalusbursa sialoproteome in response to Babesiaovis infection and blood feeding. Two potential tick protective antigens were identified and its influence in tick biological parameters and pathogen infection was evaluated. Results demonstrate that the R. bursa sialoproteome is highly affected by feeding but infection is well tolerated by tick cells. The combination of both stimuli shifts the previous scenario and a more evident pathogen manipulation can be suggested. Knockdown of ub2n led to a significative increase of infection in tick salivary glands but a brusque decrease in the progeny, revealing its importance in the cellular response to pathogen infection, which is worth pursuing in future studies. Additionally, an impact in the recovery rate of adults (62%), the egg production efficiency (45.75%), and the hatching rate (88.57 %) was detected. Building knowledge on vector and/or pathogen interplay bridges the identification of protective antigens and the development of novel control strategies.