A versatile yeast model identifies the pesticides cymoxanil and metalaxyl as risk factors for synucleinopathies.

Parkinson's disease (PD) is a neurodegenerative disorder characterized by the loss of dopaminergic neurons and the presence of Lewy bodies, which predominantly consist of aggregated forms of the protein alpha-synuclein (aSyn). While these aggregates are a pathological hallmark of PD, the etiology of most cases remains elusive. Although environmental risk factors have been identified, such as the pesticides dieldrin and MTPT, many others remain to be assessed and their molecular impacts are underexplored. This study aimed to identify pesticides that could enhance aSyn aggregation using a humanized yeast model expressing aSyn fused to GFP as a primary screening platform, which we validated using dieldrin. We found that the pesticides cymoxanil and metalaxyl induce aggregation of aSyn in yeast, which we confirmed also occurs in a model of aSyn inclusion formation using human H4 cells. In conclusion, our approach generated invaluable molecular data on the effect of pesticides, therefore providing insights into mechanisms associated with the onset and progression of PD and other synucleinopathies.

Novel yeast-based biosensor for environmental monitoring of tebuconazole.

Due to increasing demand for high and stable crop production, human populations are highly dependent on pesticide use for growing and storing food. Environmental monitoring of these agrochemicals is therefore of utmost importance, because of their collateral effects on ecosystem and human health. Even though most current-use analytical methods achieve low detection limits, they require procedures that are too complex and costly for routine monitoring. As such, there has been an increased interest in biosensors as alternative or complementary tools to streamline detection and quantification of environmental contaminants. In this work, we developed a biosensor for environmental monitoring of tebuconazole (TEB), a common agrochemical fungicide. For that purpose, we engineered S. cerevisiae cells with a reporter gene downstream of specific promoters that are expressed after exposure to TEB and characterized the sensitivity and specificity of this model system. After optimization, we found that this easy-to-use biosensor consistently detects TEB at concentrations above 5 µg L-1 and does not respond to realistic environmental concentrations of other tested azoles, suggesting it is specific. We propose the use of this system as a complementary tool in environmental monitoring programs, namely, in high throughput scenarios requiring screening of numerous samples. KEY POINTS: • A yeast-based biosensor was developed for environmental monitoring of tebuconazole. •The biosensor offers a rapid and easy method for tebuconazole detection ≥ 5 µg L-1. •The biosensor is specific to tebuconazole at environmentally relevant concentrations.

Bovine Lactoferrin-Loaded Plasmonic Magnetoliposomes for Antifungal Therapeutic Applications.

Bovine lactoferrin (bLf) is a milk-derived protein that exhibits potent broad-spectrum antifungal activity against multiple fungi. bLf is susceptible to degradation, while some of its properties depend on the tertiary structure. So, the encapsulation of bLf in stimuli-responsive therapeutic formulations provides an added value to enhance its biological activities. Plasmonic magnetoliposomes (PMLs) arise as promising nanocarriers for dual hyperthermia (magneto-photothermia) and local chemotherapy, since the combination of magnetic and gold nanoparticles (NPs) in a single nanosystem (multifunctional liposomes) enables the targeting and controlled release of loaded drugs. In this work, plasmonic magnetoliposomes (PMLs) containing manganese ferrite nanoparticles (28 nm size) and gold nanoparticles (5-7.5 nm size), functionalized with 11-mercaptoundecanoic acid or octadecanethiol, were prepared and loaded with bLf. The NPs' optical, magnetic and structural properties were measured via UV/vis/NIR absorption spectroscopy, SQUID and TEM, respectively. The Specific Absorption Rate (SAR) was calculated to assess the capabilities for magnetic and photothermal hyperthermia. Finally, the antifungal potential of bLf-loaded PMLs and their mechanism of internalization were assessed in Saccharomyces cerevisiae by counting the colony forming units and using fluorescence microscopy. The results demonstrate that PMLs are mainly internalized through an energy- and temperature-dependent endocytic process, though the contribution of a diffusion component cannot be discarded. Most notably, only bLf-loaded plasmonic magnetoliposomes display cytotoxicity with an efficiency similar to free bLf, attesting their promising potential for bLf delivery in the context of antifungal therapeutic interventions.

Lactate Induces Cisplatin Resistance in S. cerevisiae through a Rad4p-Dependent Process.

Cisplatin is a widely used antineoplastic agent that has DNA as the main target, though cellular resistance hampers its therapeutic efficacy. An emerging hallmark of cancer cells is their altered metabolism, characterized by increased glycolysis even under aerobic conditions, with increased lactate production (known as the Warburg effect). Although this altered metabolism often results in increased resistance to chemotherapy, it also provides an opportunity for targeted therapeutic intervention. It has been suggested that cisplatin cytotoxicity can be affected by tumor metabolism, though with varying effects. We therefore sought to better characterize how lactate affects cisplatin sensitivity in the simplified Saccharomyces cerevisiae model. We show that lactate renders yeast cells resistant to cisplatin, independently of growth rate or respiration ability. We further show that histone acetylation is not affected, but histone phosphorylation is decreased in lactate-containing media. Finally, we show that Rad4p, essential for nucleotide excision repair, is required for the observed phenotype and thus likely underlies the mechanism responsible for lactate-mediated resistance to cisplatin. Overall, understanding how lactate modulates cisplatin sensitivity will aid in the development of new strategies to overcome drug resistance.

Safety Evaluation of Fluoride Content in Tea Infusions Consumed in the Azores-a Volcanic Region with Water Springs naturally Enriched in Fluoride.

Tea is the second most commonly consumed beverage in the world. It is well recognized that the consumption of tea in high quantities can promote the development of fluorosis. The main objective of this study is to estimate the exposure to fluoride in the Azores through drinking tea prepared with water from different volcanic locations, by i) investigating the fluoride (F) content of various commercial brands of tea (Camellia sinensis) marketed in Azores and ii) comparing tea releasing rates of F according to brewing time, considering the fluoride concentration in the different types of water used for the infusion. Fluoride contents were determined by ion-selective electrode in 30 samples of drinking water from three different locations and in 450 samples of tea (black and green tea) from three different brands. Fluoride concentration in water ranged from 0.29 to 1.56 ppm (Porto Formoso and Sete Cidades village, respectively). Fluoride concentrations increased with brewing time, reaching the highest values in the Azorean black and green tea infusions. For all the studied brands, a negative correlation was found between tea fluoride contents and the pH of the water used to prepare the infusion. Fluoride concentration in infusions was significantly associated with the background fluoride concentration in drinking water. Since the fluoride concentration in groundwater varies accordingly to the geological conditions and tea consumption can contribute to fluoride intake, it is important to define the limits for tea consumption, particularly in fluoride-rich areas. Graphical Abstract Fluoride concentrations in black and green tea for 3 minutes of brewing time and, association between fluoride concentration and pH with brewing time.

Sensitivity of two biomarkers for biomonitoring exposure to fluoride in children and women: A study in a volcanic area.

The natural enrichment of water with fluoride is related to natural sources such as volcanic activity, with it being documented that fluorosis, an endemic and widespread disease in volcanic areas, is associated to the ingestion of high levels of fluoride through water. Thus, in this study, we aimed to define the fluoride concentration in drinking waters of volcanic origin and compare the sensitivity of urine and nail clippings as biomarkers for fluoride exposure in adults and children. Samples of drinking water from four villages in São Miguel Island (Azores) were used and the fluoride concentration was determined, as well the fluoride content in urine and toenails clippings from 66 children and 63 adults from these villages. A validated diet questionnaire, assessing sources of fluoride, was recorded for each participant. The fluoride determination in urine and nail clipping samples was made using a fluoride-specific electrode. A positive correlation was found between the fluoride daily intake and fluoride content in children urine (rs = 0.475; p < 0.001) and in their nail clippings (rs = 0.475; p < 0.001), while in adult women, the fluoride daily intake correlated positively with fluoride content nail clippings (rs = 0.495, p < 0.001). This reveals that nail clippings are more reliable as biomarkers of chronic exposure to fluoride than urine for populations of different ages (children vs. adults). Furthermore, nail clippings are more suitable than urine fluoride levels to assess long term exposure to fluoride in areas where the exposure to fluoride in drinking water is considered within, or slightly above, the recommended legal values.