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- Probe-based metagenomic pathogen detection: advancing laboratory capacity for complex diagnosisPublication . Ferreira, Rita; Coelho, Luís; Santos, João Dourado; Sobral, Daniel; Isidro, Joana; Mixão, Verónica; Pinto, Miguel; Nunes, Alexandra; Borrego, Maria José; Lopo, Sílvia; Oleastro, Mónica; Sousa, Rita; Palminha, Paula; Veríssimo, Cristina; Gargaté, Maria João; Guiomar, Raquel; Cordeiro, Rita; Macedo, Rita; Bajanca-Lavado, Paula; Paixão, Paulo; Duarte, Sílvia; Vieira, Luís; Borges, Vítor; Gomes, João PauloProbe-based pathogen enrichment, followed by NGS, is a promising tool for complex diagnosis, overcoming traditional challenges of shotgun metagenomics, namely small microbial/human genetic material ratio and demanding computational resources. Here, we assessed the combined detection performance of two Illumina probe-based panels, the Respiratory and the Urinary Pathogen ID panels (RPIP and UPIP), using 99 clinical samples of 15 different matrices (e.g., cerebrospinal fluid, plasma, serum, urine, swabs, biopsies, etc.) available from Portuguese National Reference Laboratories. This sample set involved 114 "PCR-positive hits" (Ct values range of 9.7-41.3; median of 28.4) for 52 non-redundant human pathogens. For a more detailed bioinformatics assessment, as a complement of the Illumina turnkey solution (Explify), we applied an extended version of our INSaFLU-TELEVIR(+) metagenomics pipeline. Whereas Explify analyses resulted in an initial detection frequency of 73.7% (84/114), the subsequent application of INSaFLU-TELEVIR(+), including taxonomic classification followed by confirmatory read mapping, enabled an overall detection proportion of 79.8% (91/114) of the PCR-positive hits. This translated into a detection rate increment from 54.3% (19/35) to 65.7% (23/35) for bacteria, and from 85.3% (58/68) to 89.7% (61/68) for viruses. The implemented workflow was also very satisfactory for samples with qPCR Ct values above 30, with an overall detection frequency of 71.8% (28/39) when compared with the 92.0% (46/50) observed for those with Ct ≤ 30. In summary, this study validated and established a pioneering approach at the Portuguese National Institute of Health to support clinicians in complex diagnosis, contributing to advance diagnostic capabilities toward a more informed clinical decision and potential improvement of infectious disease outcomes.
- Influence of application methods of a whey protein-based coating with cardoon extract on cured cheese propertiesPublication . Barbosa, Cássia H.; Andrade, Mariana A.; Tomé, Sidney; Vilarinho, Fernanda; Fernando, Ana Luísa; Sanches Silva, AnaThe application of edible coatings for cheese preservation has gained attention for maintaining its quality and enhancing its shelf life. Whey protein-based coatings, with natural extracts, can protect from moisture loss, lipid oxidation, and microbial contamination. Cynara cardunculus L. (cardoon) is a Mediterranean plant whose flowers are used in cheesemaking, and the leaves, the main by-product generated, are known to have an interesting polyphenol profile. This study aims to evaluate different methods of applying a whey protein-based coating incorporated with 2% cardoon leaf extract on cured cheese. The different methods involved two groups of control cheese samples: one group remained untreated, while the other group was coated without any extract. The active samples consisted of three groups of cheese samples: one was doubledipped and air-dried, another was double-dipped and dried in a flow chamber, and a third one was single-dipped and dried in a flow chamber. The samples were kept for 30 days at 30 °C. The samples were analysed for microbial contamination, moisture content, colour, and lipid oxidation. The whey protein-based coating incorporated with cardoon leaf extract had significant effects on the packed cheese. The active coating reduced microbial counts, specifically S. aureus and moulds and yeasts, compared to the control samples. The active coating decreased luminosity and increased the yellow-blue value. But there was no significant difference in moisture content. The active coating provided moderate protection against lipid oxidation. Overall, applying whey protein-based coating incorporated with 2% cardoon leaf extract appears to be an effective method to prolong the shelf life of cheese. Incorporating natural extracts into whey protein-based coating represents a natural and effective strategy to protect cheeses against microbiological contamination and oxidation. This approach enhances the quality and safety of cheese products while meeting consumer demand for natural and sustainable food preservation methods.
- Cardoon leaves as a nutrient food source: promoting sustainability and circular economyPublication . Barbosa, Cássia H.; Andrade, Mariana A.; Tomé, Sidney; Vilarinho, Fernanda; Fernando, Ana Luísa; Sanches Silva, AnaAgro-industrial by-products are a substantial source of bioactive compounds and nutrients. The valorisation of these by-products has the potential to promote environmental sustainability and a circular economy. This practice can lead to a significant advancement in sustainable food production. Cardoon (Cynara cardunculus L.) is a multipurpose crop, and its flowers are used as a natural coagulant in the cheesemaking process in countries such as Portugal and Spain. Cardoon leaves are rich in bioactive compounds, yet they are still often discarded as waste. These leaves can be directly consumed or used as an active ingredient in food packaging, thereby promoting resource efficiency and waste reduction. This study aims to evaluate the nutritional composition of cardoon leaves and their potential as a food ingredient. Fresh cardoon leaves were analyzed according to their content of moisture, ash, total protein, total fat, total dietary fiber, and total sugars, as well as their fatty acid profile. The results, expressed based on the weight of the fresh product, show that cardoon leaves contain 2.4 g/100 g, indicating a rich mineral composition. The total fat content is low, at 0.2 g/100 g, with only 0.05 g/100 g of saturated fat. The protein content (4.2 g/100 g) is considered adequate, highlighting its potential as a plant-based protein source. Additionally, the high fiber content (7.9 g/100 g) supports digestive health, while the low sugar content (0.9 g/100 g) further enhances its nutritional profile. Although cardoon leaves are often considered waste products, they offer substantial nutritional value, boasting high levels of protein, fiber, and minerals, as well as low levels of sugar and fat. Their effective valorisation can create a more sustainable system of food production. By incorporating cardoon leaves into the human diet and food packaging, their full potential can be achieved while minimizing waste and promoting sustainable consumption.