Hunting for Answers: Assessing Brucella spp. Seroprevalence and Risks in Red Deer and Wild Boar in Central Portugal.

Between 2016 and 2023, a cross-sectional study was conducted in the central region of Portugal in order to better understand the epidemiology and public health risks resulting from the handling and consumption of game animals infected with Brucella spp. The seroprevalence and risk factors for Brucella spp. seropositivity were evaluated. Antibodies against Brucella spp. were determined using a commercial enzyme-linked immunosorbent assay (ELISA) according to the manufacturer's instructions. Results showed that in the 650 serum samples collected from red deer (n = 298) and wild boars (n = 352) in Portugal, 21.7% (n = 141; 95% CI: 18.6-25.1%) tested positive. Wild boar had a significantly higher prevalence (35.5%; 95% CI: 30.5-40.8%) than red deer (5.4%, 95% CI: 3.1-8.6%; p ≤ 0.001). Risk factors for seropositivity were investigated using multivariable logistic regression models. The odds of being seropositive was 8.39 (95% CI: 4.75-14.84; p ≤ 0.001) times higher in wild boar than in red deer. Correlations between sex, age, body condition, and seropositivity could not be observed. The higher seroprevalence in wild boar suggests that this species may primarily contribute to the Brucella spp. ecology in central Portugal.

Using Analytic Hierarchy Process to Assess Beekeeping Suitability in Portuguese Controlled Areas: A First Approach.

Beekeeping management is greatly influenced by spatial factors (e.g., land use/land cover, roads, or electrical energy networks), so GIS are a powerful tool to overlap and relate a variety of spatial data levels and, consequently, a very useful tool for beekeeping activity planning. This study was developed within the intervention area of three controlled zones managed by Portuguese Beekeepers Associations. The methodology, based on multi-criteria decision analysis, integrates several criteria, such as hydrographic networks, road networks, soil occupation, solar radiation, and electromagnetic radiation sources. These criteria were proposed and evaluated through online questionnaires carried out with beekeepers. Concerning the selected criteria and the respective geographical data, the most relevant were land use/land cover and water availability, with a significance of 44% and 24%, respectively. The beekeeping suitability map enabled us to evaluate the degree of compliance for the actual location of apiaries, with 60% of the apiaries being installed in high potential areas. In the context of beekeeping planning, the potential of the techniques applied seems to be an important tool for optimizing the location of apiaries and the profitability of beekeeping.

Multitemporal Land Use and Cover Analysis Coupled with Climatic Change Scenarios to Protect the Endangered Taxon Asphodelus bento-rainhae subsp. bento-rainhae.

Climate change and land use and land cover (LULC) change are impacting the species' geographic distribution, causing range shifts and reducing suitable habitats. Asphodelus bento-rainhae subsp. bento-rainhae (AbR) is an endangered endemic plant restricted to Serra da Gardunha (Portugal), and knowledge of those changes will help to design conservation measures. MaxEnt was used to model AbR's current distribution and project it into the future, 2050, using the Shared Socioeconomic Pathway SSP3-7. The Portuguese LULC maps from 1951-1980, 1995, 2007, and 2018 were used to assess and quantify LULC changes over time. The results showed that the AbR current predicted distribution matches its actual known distribution, which will not be affected by future predicted climate change. The significant LULC changes were observed during the study periods 1951-1980 to 2018, particularly between 1951-1980 and 1995. Scrubland and Agriculture decreased by 5% and 2.5%, respectively, and Forests increased by 4% in the study area. In the occurrence area, Agriculture increased, and Forests decreased between 1980 and 2018, due to Orchard expansion (34%) and declines in Chestnut (16.9%) and Pine (11%) areas, respectively. The use of species distribution models and the LULC change analysis contributed to understanding current and future species distribution. The LULC changes will have a significant impact on future species distribution. To prevent the extinction of this endemic species in the future, it is crucial to implement conservation measures, namely species monitoring, replantation, and germplasm conservation, in addition to guidelines for habitat conservation.

The role of littoral cliffs in the niche delimitation on a microendemic plant facing climate change.

Obligate coastline taxa generally occupy very limited areas, especially when there is a close affinity with a specific coast type. Climate change can be a meaningful threat for them, reducing suitable habitat or forcing migration events. Cistus ladanifer subsp. sulcatus is an endemic plant of Portugal, known to occur only in the top of its south-western coast's prominent cliffs. In spite of being included in the annexes II and IV of the European Habitats Directive of Natura 2000 Network, this taxon is still understudied, especially regarding the effects of climate change on its distribution. To overcome such gap, Maxent was used to model the current distribution of C. ladanifer subsp. sulcatus and project its future distribution considering different General Circulation Models, periods (2050 and 2070) and Representation Concentration Pathways (4.5 and 8.5). The results suggested an extensive range contraction in the future, and extinction is a possible scenario. The proximity to littoral cliffs is crucial for this plant's occurrence, but these formations are irregularly distributed along the coast, hindering range expansions, further inhibited by a small dispersal capacity. Cistus ladanifer subsp. sulcatus will probably remain confined to south-western Portugal in the future, where it will continue to face relevant threats like human activity, reinforcing the need for its conservation.

Neuromechanisms of SARS-CoV-2: A Review.

Recent studies have suggested the neuroinvasive potential of severe acute respiratory coronavirus 2 (SARS-CoV-2). Notably, neuroinvasiveness might be involved in the pathophysiology of coronavirus disease 2019 (COVID-19). Some studies have demonstrated that synapse-connected routes may enable coronaviruses to access the central nervous system (CNS). However, evidence related to the presence of SARS-CoV-2 in the CNS, its direct impact on the CNS, and the contribution to symptoms suffered, remain sparse. Here, we review the current literature that indicates that SARS-CoV-2 can invade the nervous system. We also describe the neural circuits that are potentially affected by the virus and their possible role in the progress of COVID-19. In addition, we propose several strategies to understand, diagnose, and treat the neurological symptoms of COVID-19.

Rab7 controls lipid droplet-phagosome association during mycobacterial infection.

Lipid droplets (LDs) are organelles that have multiple roles in inflammatory and infectious diseases. LD act as essential platforms for immunometabolic regulation, including as sites for lipid storage and metabolism, inflammatory lipid mediator production, and signaling pathway compartmentalization. Accumulating evidence indicates that intracellular pathogens may exploit host LDs as source of nutrients and as part of their strategy to promote immune evasion. Notably, numerous studies have demonstrated the interaction between LDs and pathogen-containing phagosomes. However, the mechanism involved in this phenomenon remains elusive. Here, we observed LDs and PLIN2 surrounding M. bovis BCG-containing phagosomes, which included observations of a bacillus cell surrounded by lipid content inside a phagosome and LAM from mycobacteria co-localizing with LDs; these results were suggestive of exchange of contents between these compartments. By using beads coated with M.tb lipids, we demonstrated that LD-phagosome associations are regulated through the mycobacterial cell wall components LAM and PIM. In addition, we demonstrated that Rab7 and RILP, but not Rab5, localizes to LDs of infected macrophages and observed the presence of Rab7 at the site of interaction with an infected phagosome. Moreover, treatment of macrophages with the Rab7 inhibitor CID1067700 significantly inhibited the association between LDs and LAM-coated beads. Altogether, our data demonstrate that LD-phagosome interactions are controlled by mycobacterial cell wall components and Rab7, which enables the exchange of contents between LDs and phagosomes and may represent a fundamental aspect of bacterial pathogenesis and immune evasion.

Geospatial analysis applied to seroepidemiological survey of canine leishmaniosis in east-central Portugal.

Despite the high prevalence of canine Leishmania infantum infection in Portugal, significant differences associated with different risk factors can be found between geographically contiguous areas. In this study, a geographical area within the central region of Portugal (municipalities of Proença-a-Nova, Mação and Vila de Rei) was investigated. An epidemiological survey involved the analysis by an indirect enzyme-linked immunosorbent assay of serum samples collected during the anti-rabies vaccination campaign from 282 dogs. Geospatial analysis showed the distribution of geospatial prevalence of leishmaniosis and has delimited two areas (clusters) with a statistically significant higher risk of seropositivity in dogs (p =  0.003 and p = 0.027, for clusters 1 and 2, respectively). The highest seroprevalence (56.0%; CI: 41.2-70.0) was found in Vila de Rei. Five land occupation types showed a possible influence on the geographic distribution of seropositivity, with statistically significant differences between seropositive and seronegative dogs. Land occupied by temporary irrigated crops (p =  0.026), olive groves (p =  0.013), complex cultural systems and parcelling (p =  0.021), open forests, logging and new plantations (p =  0.043) and watercourses (p =  0.012) influenced the geographical distribution of canine Leishmania infection. Seropositive dogs had a greater average area of occupied land (i.e. open forests, logging and new plantations) than the seronegative ones (3.1439 km2 versus 2.5650 km2, respectively; p =  0.043).

Bioclimatic modeling in the Last Glacial Maximum, Mid-Holocene and facing future climatic changes in the strawberry tree (Arbutus unedo L.).

Increasing forest wildfires in Portugal remain a growing concern since forests in the Mediterranean region are vulnerable to recent global warming and reduction of precipitation. Therefore, a long-term negative effect is expected on the vegetation, with increasing drought and areas burnt by fires. The strawberry tree (Arbutus unedo L.) is particularly used in Portugal to produce a spirit by processing its fruits and is the main income for forestry owners. Other applications are possible due to the fruit and leaves' anti-oxidant properties and bioactive compounds production, with a potential for clinical and food uses. It is a sclerophyllous plant, dry-adapted and fire resistant, enduring the Mediterranean climate, and recently considered as a possibility for afforestation, to intensify forest discontinuity where pines and eucalypts monoculture dominate the region. To improve our knowledge about the species' spatial distribution we used 318 plots (the centroid of a 1 km2 square grid) measuring the species presence and nine environmental attributes. The seven bioclimatic variables most impacting on the species distribution and two topographic features, slope and altitude, were used. The past, current and future climate data were obtained through WorldClim. Finally, the vulnerability of the strawberry tree to the effects of global climate change was examined in the face of two emission scenarios (RCP 4.5 and 8.5), to predict distribution changes in the years 2050 and 2070, using a species distribution models (MaxEnt). The reduction of suitable habitat for this species is significant in the southern regions, considering the future scenarios of global warming. Central and northern mountainous regions are putative predicted refuges for this species. Forest policy and management should reflect the impact of climate change on the usable areas for forestry, particularly considering species adapted to the Mediterranean regions and wildfires, such as the strawberry tree. The distribution of the species in the Last Glacial Maximum (LGM) and Mid-Holocene (MH) agrees with previous genetic and paleontological studies in the region, which support putative refuges for the species. Two in the southern and coastal-central regions, since the LGM, and one in the east-central mountainous region, considered as cryptic refugia.

Microglia/Astrocytes-Glioblastoma Crosstalk: Crucial Molecular Mechanisms and Microenvironmental Factors.

In recent years, the functions of glial cells, namely, astrocytes and microglia, have gained prominence in several diseases of the central nervous system, especially in glioblastoma (GB), the most malignant primary brain tumor that leads to poor clinical outcomes. Studies showed that microglial cells or astrocytes play a critical role in promoting GB growth. Based on the recent findings, the complex network of the interaction between microglial/astrocytes cells and GB may constitute a potential therapeutic target to overcome tumor malignancy. In the present review, we summarize the most important mechanisms and functions of the molecular factors involved in the microglia or astrocytes-GB interactions, which is particularly the alterations that occur in the cell's extracellular matrix and the cytoskeleton. We overview the cytokines, chemokines, neurotrophic, morphogenic, metabolic factors, and non-coding RNAs actions crucial to these interactions. We have also discussed the most recent studies regarding the mechanisms of transportation and communication between microglial/astrocytes - GB cells, namely through the ABC transporters or by extracellular vesicles. Lastly, we highlight the therapeutic challenges and improvements regarding the crosstalk between these glial cells and GB.

Lipid Body Organelles within the Parasite Trypanosoma cruzi: A Role for Intracellular Arachidonic Acid Metabolism.

Most eukaryotic cells contain varying amounts of cytosolic lipidic inclusions termed lipid bodies (LBs) or lipid droplets (LDs). In mammalian cells, such as macrophages, these lipid-rich organelles are formed in response to host-pathogen interaction during infectious diseases and are sites for biosynthesis of arachidonic acid (AA)-derived inflammatory mediators (eicosanoids). Less clear are the functions of LBs in pathogenic lower eukaryotes. In this study, we demonstrated that LBs, visualized by light microscopy with different probes and transmission electron microscopy (TEM), are produced in trypomastigote forms of the parasite Trypanosoma cruzi, the causal agent of Chagas' disease, after both host interaction and exogenous AA stimulation. Quantitative TEM revealed that LBs from amastigotes, the intracellular forms of the parasite, growing in vivo have increased size and electron-density compared to LBs from amastigotes living in vitro. AA-stimulated trypomastigotes released high amounts of prostaglandin E2 (PGE2) and showed PGE2 synthase expression. Raman spectroscopy demonstrated increased unsaturated lipid content and AA incorporation in stimulated parasites. Moreover, both Raman and MALDI mass spectroscopy revealed increased AA content in LBs purified from AA-stimulated parasites compared to LBs from unstimulated group. By using a specific technique for eicosanoid detection, we immunolocalized PGE2 within LBs from AA-stimulated trypomastigotes. Altogether, our findings demonstrate that LBs from the parasite Trypanosoma cruzi are not just lipid storage inclusions but dynamic organelles, able to respond to host interaction and inflammatory events and involved in the AA metabolism. Acting as sources of PGE2, a potent immunomodulatory lipid mediator that inhibits many aspects of innate and adaptive immunity, newly-formed parasite LBs may be implicated with the pathogen survival in its host.

Differential TLR2 downstream signaling regulates lipid metabolism and cytokine production triggered by Mycobacterium bovis BCG infection.

The nuclear receptor PPARγ acts as a key modulator of lipid metabolism, inflammation and pathogenesis in BCG-infected macrophages. However, the molecular mechanisms involved in PPARγ expression and functions during infection are not completely understood. Here, we investigate signaling pathways triggered by TLR2, the involvement of co-receptors and lipid rafts in the mechanism of PPARγ expression, lipid body formation and cytokine synthesis in macrophages during BCG infection. BCG induces NF-κB activation and increased PPARγ expression in a TLR2-dependent manner. Furthermore, BCG-triggered increase of lipid body biogenesis was inhibited by the PPARγ antagonist GW9662, but not by the NF-κB inhibitor JSH-23. In contrast, KC/CXCL1 production was largely dependent on NF-κB but not on PPARγ. BCG infection induced increased expression of CD36 in macrophages in vitro. Moreover, CD36 co-immunoprecipitates with TLR2 in BCG-infected macrophages, suggesting its interaction with TLR2 in BCG signaling. Pretreatment with CD36 neutralizing antibodies significantly inhibited PPARγ expression, lipid body formation and PGE2 production induced by BCG. Involvement of CD36 in lipid body formation was further confirmed by decreased BCG-induced lipid body formation in CD36 deficient macrophages. Similarly, CD14 and CD11b/CD18 blockage also inhibited BCG-induced lipid body formation, whereas TNF-α synthesis was not affected. Disruption of rafts recapitulates the latter result, inhibiting lipid body formation, but not TNF-α synthesis in BCG-infected macrophages. In conclusion, our results suggest that CD36-TLR2 cooperation and signaling compartmentalization within rafts, divert host response signaling through PPARγ-dependent and NF-κB-independent pathways, leading to increased macrophage lipid accumulation and down-modulation of macrophage response.

Host cell lipid bodies triggered by Trypanosoma cruzi infection and enhanced by the uptake of apoptotic cells are associated with prostaglandin E2 generation and increased parasite growth.

Lipid bodies (lipid droplets) are lipid-rich organelles with functions in cell metabolism and signaling. Here, we investigate the mechanisms of Trypanosoma cruzi-induced lipid body formation and their contributions to host-parasite interplay. We demonstrate that T. cruzi-induced lipid body formation in macrophages occurs in a Toll-like receptor 2-dependent mechanism and is potentiated by apoptotic cell uptake. Lipid body biogenesis and prostaglandin E2 (PGE2) production triggered by apoptotic cell uptake was largely dependent of α(v)ß3 and transforming growth factor-ß signaling. T. cruzi-induced lipid bodies act as sites of increased PGE synthesis. Inhibition of lipid body biogenesis by the fatty acid synthase inhibitor C75 reversed the effects of apoptotic cells on lipid body formation, eicosanoid synthesis, and parasite replication. Our findings indicate that lipid bodies are highly regulated organelles during T. cruzi infection with roles in lipid mediator generation by macrophages and are potentially involved in T. cruzi-triggered escape mechanisms.

Neutrophils recruited to the site of Mycobacterium bovis BCG infection undergo apoptosis and modulate lipid body biogenesis and prostaglandin E production by macrophages.

Neutrophil influx to sites of mycobacterial infections is one of the first events of tuberculosis pathogenesis. However, the role of early neutrophil recruitment in mycobacterial infection is not completely understood. We investigated the rate of neutrophil apoptosis and the role of macrophage uptake of apoptotic neutrophils in a pleural tuberculosis model induced by BCG. Recruited neutrophils were shown to phagocyte BCG and a large number of neutrophils undergo apoptosis within 24 h. Notably, the great majority of apoptotic neutrophils were infected by BCG. Increased lipid body (lipid droplets) formation, accompanied by prostaglandin E(2) (PGE(2)) and TGF-beta1 synthesis, occurred in parallel to macrophage uptake of apoptotic cells. Lipid body and PGE(2) formation was observed after macrophage exposure to apoptotic, but not necrotic or live neutrophils. Blockage of BCG-induced lipid body formation significantly inhibited PGE(2) synthesis. Pre-treatment with the pan-caspase inhibitor zVAD inhibited BCG-induced neutrophil apoptosis and lipid body formation, indicating a role for apoptotic neutrophils in macrophage lipid body biogenesis in infected mice. In conclusion, BCG infection induced activation and apoptosis of infected neutrophils at the inflammatory site. The uptake of apoptotic neutrophils by macrophages leads to TGF-beta1 generation and PGE(2)-derived lipid body formation, and may have modulator roles in mycobacterial pathogenesis.

Toll-like receptor-2-mediated C-C chemokine receptor 3 and eotaxin-driven eosinophil influx induced by Mycobacterium bovis BCG pleurisy.

An acute and persistent eosinophil infiltration is observed during Mycobacterium bovis BCG pleural infection in mice. Eosinophil accumulation, lipid body formation, and eotaxin production were significantly reduced in BCG-infected Toll-like receptor-2 (TLR2)-deficient mice compared to wild-type mice. Neutralization of eotaxin or CCR3 drastically inhibited BCG-induced eosinophil accumulation and lipid body formation, indicating that BCG-induced eosinophil recruitment and activation is largely dependent of TLR2-mediated eotaxin generation.