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The study aimed to optimize qPCR reactions using oligonucleotides from the first Brazilian molecular diagnostic kit for leprosy on a portable platform (Q3-Plus). In addition, we sought to develop a simplified protocol for DNA extraction that met point-of-care criteria. During optimization on the Q3-Plus, optical parameters, thresholds, and cutoffs for the 16S rRNA and RLEP targets of M. leprae were established using synthetic DNA, purified DNA from M. leprae, and pre-characterized clinical samples. For the simplified extraction protocol, different lysis solutions were evaluated using chaotropic agents, and purification was carried out by transferring the lysed material to FTA cards. The complete protocol (simplified extraction + qPCR on the portable platform) was then evaluated with pre-characterized clinical skin biopsy samples and compared with standard equipment (QuantStudio-5). LOD95% for the optimized reactions was 113.31 genome-equivalents/μL for 16S rRNA and 17.70 genome-equivalents/μL for RLEP. Among the lysis solutions, the best-performing was composed of urea (2 M), which provided good dissolution of the skin fragment and a lower Ct value, indicating higher concentrations of DNA. The complete technological solution showed a sensitivity of 52% in reactions. Our results highlight the need for additional optimization to deal with paucibacillary samples, but also demonstrate the feasibility of the portable platform for the qPCR detection of M. leprae DNA in low infrastructure settings., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Bertão-Santos et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Background:  Resection of infiltrative neuroepithelial primary brain tumors, such as low-grade gliomas (LGGs) remains a neurosurgical challenge. Usual lack of clinical deficit despite LGGs growing in eloquent brain areas may be explained by reshaping and reorganization of functional networks. The development of modern diagnostic imaging techniques could disclose better understanding of the rearrangement of the brain cortex; however, mechanisms underlying such compensation and how it occurs in the motor cortex remain unclear. This systematic review aims to analyze the neuroplasticity of motor cortex in patients with LGGs, as determined by neuroimaging and functional techniques., Methods:  Following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines, medical subject headings (MeSH) and the following terms related to neuroimaging, LGGs and neuroplasticity were used with the Boolean operators AND and OR to synonymous terms in the PubMed database. Among the 118 results, 19 studies were included in the systematic review., Results:  Motor function in patients with LGG was characterized by a compensation in the contralateral and supplementary motor areas and premotor functional networks. Furthermore, ipsilateral activation in these types of gliomas was rarely described. Moreover, some studies did not reveal statistical significance in association between functional reorganization and the postoperative period, which can be explained by the low number of patients., Conclusion:  Our findings suggest a high pattern of reorganization per different eloquent motor areas and gliomas diagnosis. Understanding this process is useful to guide safe surgical resection and to develop protocols that assess the plasticity, even though functional network rearrangement needs to be better characterized by more studies., Competing Interests: None declared., (Thieme. All rights reserved.)

Scedosporium and Lomentospora are a group of filamentous fungi with some clinically relevant species causing either localized, invasive, or disseminated infections. Understanding how the host immune response is activated and how fungi interact with the host is crucial for a better management of the infection. In this context, an α-glucan has already been described in S. boydii , which plays a role in the inflammatory response. In the present study, an α-glucan has been characterized in L. prolificans and was shown to be exposed on the fungal surface. The α-glucan is recognized by peritoneal macrophages and induces oxidative burst in activated phagocytes. Its recognition by macrophages is mediated by receptors that include Dectin-1 and Mincle, but not TLR2 and TLR4. These results contribute to the understanding of how Scedosporium 's and Lomentospora 's physiopathologies are developed in patients suffering with scedosporiosis and lomentosporiosis.

Host genetic determinants that underpin variation in susceptibility to systemic fungal infection are poorly understood. Genes responsible for complex traits can be identified by correlating variation in phenotype with allele in founder strains of wild mice with known genetic variation, assembled in genetic reference panels. In this work, we describe wide natural variation in both primary and acquired resistance to experimental pulmonary blastomycosis in eight founder strains, including 129, A/J, BL/6, CAST, NOD, NZO, PWK, and WSB of the Collaborative Cross collection, and the inbred DBA strain. These differences in susceptibility across strains were accompanied by sharp differences in the accumulation and function of immune cells in the lungs. Immune perturbations were mapped by identifying reagents that phenotypically mark immune cell populations in the distinct strains of mice. In particular, we uncovered marked differences between BL/6 and DBA/2 mouse strains in the development of acquired resistance. Our findings highlight the potential value in using genetic reference panels of mice, and particularly the BXD (recombinant inbred strains of mice from a cross of C57BL/6J and DBA/2J mice) collection harboring a cross between resistant BL/6 and susceptible DBA/2 mice, for unveiling genes linked with host resistance to fungal infection. IMPORTANCE Host genetic variation significantly impacts vulnerability to infectious diseases. While host variation in susceptibility to fungal infection with dimorphic fungi has long been recognized, genes that underpin this variation are poorly understood. We used a collection of seven mouse strains that represent nearly 90% of the genetic variation in mice to identify genetic variability among the strains in resistance to pulmonary infection with the dimorphic fungus Blastomyces dermatitidis. We analyzed differences between the strains in innate resistance by infecting naive mice and in acquired resistance by infecting vaccinated mice. We identified extreme variations in both innate and acquired resistance among the strains. In particular, we found sharp differences between C57BL/6 and DBA/2 strains in the ability to acquire vaccine-induced resistance. We also identified commercial reagents that allowed the phenotyping of immune cells from this strain collection of mice. Because there are additional mice harboring a genetic cross of the C57BL/6 and DBA/2 strains (BXD collection), such mice will permit future investigations to identify the genes that underlie differences in the ability to acquire resistance to infection.

Paracoccidioidomycosis (PCM), caused by the Paracoccidioides species, is a systemic disease endemic in several Latin American countries, mainly in Brazil, Colombia, Argentina, and Venezuela. Current treatment approaches are challenging as they require prolonged durations of antifungal drugs that have potential toxicities, and despite antifungals, relapses are common. Hence, new therapeutic approaches, such as vaccines, are being investigated. The therapeutic vaccine consisting of peptide P10 associated with lipid cationic DODAB (P10+DODAB) is effective in murine models of PCM. However, the specific immune mechanisms required for the protective response has not been fully elucidated. The present work aims at evaluating the participation of neutrophils in the immune response induced by P10+DODAB. We found that the vaccine reduced both the influx of pulmonary neutrophils and the fungal load in comparison to infected animals that did not receive this treatment. The parenchymal architecture of the lungs of P10+DODAB-treated animals was largely preserved with only a few granulomas present, and tissue cytokine analysis showed a Th1 cytokine profile with augmented levels of IL-12, IFN-γ and TNF-α, and low levels of IL-4. When neutrophils were depleted 24 h prior to each treatment, the effectiveness of the P10+DODAB vaccine was completely lost as the fungal burdens remained high and histological examination showed a marked inflammation and fungal dissemination with a dysregulated cytokine response. In conclusion, these findings indicate that neutrophils are vital to ensure the triggering of an effective immune response to P10+DODAB.

Background: Chemical mosquito control using malathion has been applied in Brazil since 1985. To obtain chemical control effectiveness, vector susceptibility insecticide monitoring is required. This study aimed to describe bioassay standardizations and determine the susceptibility profile of Ae. aegypti populations to malathion and pyriproxyfen, used on a national scale in Brazil between 2017 and 2018, and discuss the observed impacts in arbovirus control., Methods: The diagnostic-doses (DD) of pyriproxyfen and malathion were determined as the double of adult emergence inhibition (EI) and lethal doses for 99% of the Rockefeller reference strain, respectively. To monitor natural populations, sampling was performed in 132 Brazilian cities, using egg traps. Colonies were raised in the laboratory for one or two generations (F1 or F2) and submitted to susceptibility tests, where larvae were exposed to the pyriproxyfen DD (0.03 µg/l) and adults, to the malathion DD determined in the present study (20 µg), in addition to the one established by the World Health Organization (WHO) DD (50 µg) in a bottle assay. Dose-response (DR) bioassays with pyriproxyfen were performed on populations that did not achieve 98% EI in the DD assays., Results: Susceptibility alterations to pyriproxyfen were recorded in six (4.5%) Ae. aegypti populations from the states of Bahia and Ceará, with Resistance Ratios (RR 95 ) ranging from 1.51 to 3.58. Concerning malathion, 73 (55.3%) populations distributed throughout the country were resistant when exposed to the local DD 20 µg/bottle. On the other hand, no population was resistant, and only 10 (7.6%) populations in eight states were considered as exhibiting decreased susceptibility (mortality ratios between 90 and 98%) when exposed to the WHO DD (50 µg/bottle)., Conclusions: The feasibility of conducting an insecticide resistance monitoring action on a nation-wide scale was confirmed herein, employing standardized and strongly coordinated sampling methods and laboratory bioassays. Brazilian Ae. aegypti populations exhibiting decreased susceptibility to pyriproxyfen were identified. The local DD for malathion was more sensitive than the WHO DD for early decreased susceptibility detection.

Candida auris and Candida haemulonii complex ( C. haemulonii, C. haemulonii var. vulnera and C. duobushaemulonii ) are phylogenetically related species that share some physiological features and habits. In the present study, we compared the virulence of these yeast species using two different experimental models: (i) Galleria mellonella larvae to evaluate the survival rate, fungal burden, histopathology and phagocytosis index and (ii) BALB/c mice to evaluate the survival. In addition, the fungal capacity to form biofilm over an inert surface was analyzed. Our results showed that in both experimental models, the animal survival rate was lower when infected with C. auris strains than the C. haemulonii species complex. The hemocytes of G. mellonella showed a significantly reduced ability to phagocytize the most virulent strains forming the C. haemulonii species complex. Interestingly, for C. auris , it was impossible to measure the phagocytosis index due to a general lysis of the hemocytes. Moreover, it was observed a greater capability of biofilm formation by C. auris compared to C. haemulonii species complex. In conclusion, we observed that C. auris and C. haemulonii complex have different levels of pathogenicity in the experimental models employed in the present study.

Priming at the site of natural infection typically elicits a protective T cell response against subsequent pathogen encounter. Here, we report the identification of a novel fungal antigen that we harnessed for mucosal vaccination and tetramer generation to test whether we can elicit protective, antigen-specific tissue-resident memory (Trm) CD4 + T cells in the lung parenchyma. In contrast to expectations, CD69 + , CXCR3 + , CD103 - Trm cells failed to protect against a lethal pulmonary fungal infection. Surprisingly, systemic vaccination induced a population of tetramer + CD4 + T cells enriched within the pulmonary vasculature, and expressing CXCR3 and CX3CR1, that migrated to the lung tissue upon challenge and efficiently protected mice against infection. Mucosal vaccine priming of Trm may not reliably protect against mucosal pathogens.

Background: The longstanding application of insecticides for vector control without periodic monitoring of the population response to these chemicals can directly drive the selection of resistant populations of vector mosquitoes. Tocantins is the newest state of the Brazilian Federation. Despite a historically high number of dengue cases, studies and monitoring data concerning insecticide resistance in the state are lacking., Objectives: To verify the resistance status of Aedes aegypti from 10 populations distributed throughout the state connected by rivers and roads., Methods: Between 50 and 150 ovitraps were installed in house gardens within each municipality. Collection points were established based on the importance of the towns and on geographic aspects. Dose response bioassays were performed in accordance with World Health Organization guidelines. Molecular assays were conducted to detect kdr mutations, which are related to pyrethroid resistance., Findings: Of the 3,200 ovitrap paddles analysed, 25.8% contained eggs, with a total of 55,687 eggs collected. With the exception of Caseara, all evaluated populations were considered to be resistant to temephos. The data showed different levels of resistance to deltamethrin among the samples. Caseara and Guaraí showed the lowest RR95 values. On average, the NaVR1 kdr allele was most frequent (40.3%), followed by NaVS (38.1%), and NaVR2 (21.6%). Palmas, the capital of the state, had the highest frequency of kdr alleles (87.5%)., Main Conclusions: With the exception of Palmas, the towns with the highest indexes (ovitrap positivity, number and density of eggs), as well with high levels of resistance and kdr alleles were located along the BR-153 road, indicating that the flow of people and cargo can contribute to the dispersion of the vector and potentially resistance. This study contributes substantially to knowledge regarding the insecticide resistance profile of Tocantins mosquito populations; the data generated via the study could facilitate the judicious use of insecticides by vector control programs.

Lomentospora prolificans is an emerging opportunistic fungus with a high resistance to antifungal agents and it can cause localized infections in immunocompetent patients and disseminated infections with a high mortality rate in immunosuppressed patients. Glucosylceramides (GlcCer) are synthetized in the majority of known fungal pathogens. They are bioactive molecules presenting different functions, such as involvement in fungal growth and morphological transitions in several fungi. The elucidation of the primary structure of the fungal surface glycoconjugates could contribute for the understanding of the mechanisms of pathogenicity. In this work, GlcCer species were isolated from mycelium and conidia forms of L. prolificans and their chemical structures were elucidated by mass spectrometry (ESI-MS). GlcCer purified from both forms presented a major species at m/z 750 that corresponds to N -2-hydroxyhexadecanoyl-1-β-D-glucopyranosyl-9-methyl-4,8-sphingadienine. Monoclonal antibodies against GlcCer could recognize L. prolificans GlcCer species from mycelium and conidia, suggesting a conserved epitope in fungal GlcCer. In addition, in vivo assays showed that purified GlcCer species from both forms was able to induce a high secretion of pro-inflammatory cytokines by splenocytes. GlcCer species also promote the recruitment of polymorphonuclear, eosinophils, small peritoneal macrophage (SPM) and mononuclear cells to the peritoneal cavity. GlcCer species were also able to induce the oxidative burst by peritoneal macrophages with NO and superoxide radicals production, and to increase the killing of L. prolificans conidia by peritoneal macrophages. These results indicate that GlcCer species from L. prolificans are a potent immune response activator.

Culicidae colonization in laboratory is paramount to conduct studies aiming at a better understanding of mosquitoes' capacity to transmit pathogens that cause deadly diseases. Colonization requires female blood feeding, a necessary step for maturation of female's oocytes. Direct blood feeding on anesthetized mammals implies in a number of disadvantages when compared to artificial blood feeding. Consequently, laboratories worldwide have been trying to -feed female mosquitoes artificially in order to replace direct feeding. In this study, we compared the effects of direct blood feeding and artificial blood feeding on important life traits of three Culicidae species. Artificial feeding was performed using citrated or defibrinated sheep blood and citrated or defibrinated rabbit blood. Direct feeding was performed using anesthetized guinea pigs as the blood source and the experiment control. Results indicated that artificial feeding using sheep blood was not good enough to justify its use in the maintenance of laboratory colonies of Culicidae. However, artificial feeding using rabbit blood maintained a recovery rate always very close to the control, especially when blood was citrated. We concluded that artificial feeding using citrated rabbit blood can substitute direct feeding on mammals reducing the use of animals, eliminating the need to maintain a bioterium in the laboratory and reducing costs in scientific researches involving Culicidae vectors.