Scielo RSS <![CDATA[Memórias do Instituto Oswaldo Cruz]]> http://www.scielo.br/rss.php?pid=0074-027620200001&lang=pt vol. 115 num. lang. pt <![CDATA[SciELO Logo]]> http://www.scielo.br/img/en/fbpelogp.gif http://www.scielo.br <![CDATA[Increasing putative vector importance of <em>Trichophoromyia</em> phlebotomines (Diptera: Psychodidae)]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100200&lng=pt&nrm=iso&tlng=pt Despite some phlebotomines being well recognised as vectors of leishmaniasis agents, vector importance of those belonging to the genus Trichophoromyia has not been extensively studied. The present study provides evidence regarding the putative vector role played by some species of Trichophoromyia on leishmanine enzootics, based on literature reports and findings obtained from field experiments conducted in the ecotopes of Pará State, Brazil. The species Th. ubiquitalis, Th. velascoi, Th. auraensis, Th. ininii and Th. brachipyga possess minimal criteria to be included in the list of suspected leishmanine vectors. However, knowledge on man-biting behavior, substantiation of vector competence and determination of epidemiological implications are limited for all of the above mentioned species. Published studies together with present data draw attention to prioritize these phlebotomine species in entomological surveillance programs and studies on experimental susceptibility to Leishmania spp. infection. <![CDATA[Characterisation of the <em>in vitro</em> activity of a Nitazoxanide-<em>N</em>-methyl-1<em>H</em>-benzimidazole hybrid molecule against albendazole and nitazoxanide susceptible and resistant strains of <em>Giardia intestinalis</em> and its <em>in vivo</em> giardicidal activity]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100300&lng=pt&nrm=iso&tlng=pt BACKGROUND It was previously demonstrated that CMC-20, a nitazoxanide and N-methyl-1H-benzimidazole hybrid molecule, had higher in vitro activity against Giardia intestinalis WB strain than metronidazole and albendazole and similar to nitazoxanide. OBJETIVES To evaluate the in vitro activity of CMC-20 against G. intestinalis strains with different susceptibility/resistance to albendazole and nitazoxanide and evaluate its effect on the distribution of parasite cytoskeletal proteins and its in vivo giardicidal activity. METHODS CMC-20 activity was tested against two isolates from patients with chronic and acute giardiasis, an experimentally induced albendazole resistant strain and a nitazoxanide resistant clinical isolate. CMC-20 effect on the distribution of parasite cytoskeletal proteins was analysed by indirect immunofluorescence and its activity was evaluated in a murine model of giardiasis. FINDINGS CMC-20 showed broad activity against susceptible and resistant strains to albendazole and nitaxozanide. It affected the parasite microtubule reservoir and triggered the parasite encystation. In this process, alpha-7.2 giardin co-localised with CWP-1 protein. CMC-20 reduced the infection time and cyst load in feces of G. muris infected mice similar to albendazole. MAIN CONCLUSIONS The in vitro and in vivo giardicidal activity of CMC-20 suggests its potential use in the treatment of giardiasis. <![CDATA[Antiparasitic and anti-inflammatory activities of ß-lapachone-derived naphthoimidazoles in experimental acute <em>Trypanosoma cruzi</em> infection]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100301&lng=pt&nrm=iso&tlng=pt BACKGROUND Chagas disease, which is caused by the protozoan Trypanosoma cruzi, is endemic to Latin America and mainly affects low-income populations. Chemotherapy is based on two nitrocompounds, but their reduced efficacy encourages the continuous search for alternative drugs. Our group has characterised the trypanocidal effect of naphthoquinones and their derivatives, with naphthoimidazoles derived from β-lapachone (N1, N2 and N3) being the most active in vitro. OBJECTIVES In the present work, the effects of N1, N2 and N3 on acutely infected mice were investigated. METHODS in vivo activity of the compounds was assessed by parasitological, biochemical, histopathological, immunophenotypical, electrocardiographic (ECG) and behavioral analyses. FINDINGS Naphthoimidazoles led to a decrease in parasitaemia (8 dpi) by reducing the number of bloodstream trypomastigotes by 25-50% but not by reducing mortality. N1 protected mice from heart injury (15 dpi) by decreasing inflammation. Bradycardia was also partially reversed after treatment with N1 and N2. Furthermore, the three compounds did not reverse hepatic and renal lesions or promote the improvement of other evaluated parameters. MAIN CONCLUSION N1 showed moderate trypanocidal and promising immunomodulatory activities, and its use in combination with benznidazole and/or anti-arrhythmic drugs as well as the efficacy of its alternative formulations must be investigated in the near future. <![CDATA[A new gene inventory of the ubiquitin and ubiquitin-like conjugation pathways in <em>Giardia intestinalis</em>]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100302&lng=pt&nrm=iso&tlng=pt BACKGROUND Ubiquitin (Ub) and Ub-like proteins (Ub-L) are critical regulators of complex cellular processes such as the cell cycle, DNA repair, transcription, chromatin remodeling, signal translation, and protein degradation. Giardia intestinalis possesses an experimentally proven Ub-conjugation system; however, a limited number of enzymes involved in this process were identified using basic local alignment search tool (BLAST). This is due to the limitations of BLAST’s ability to identify homologous functional regions when similarity between the sequences dips to &lt; 30%. In addition Ub-Ls and their conjugating enzymes have not been fully elucidated in Giardia. OBJETIVE To identify the enzymes involved in the Ub and Ub-Ls conjugation processes using intelligent systems based on the hidden Markov models (HMMs). METHODS We performed an HMM search of functional Pfam domains found in the key enzymes of these pathways in Giardia’s proteome. Each open reading frame identified was analysed by sequence homology, domain architecture, and transcription levels. FINDINGS We identified 118 genes, 106 of which corresponded to the ubiquitination process (Ub, E1, E2, E3, and DUB enzymes). The E3 ligase group was the largest group with 82 members; 71 of which harbored a characteristic RING domain. Four Ub-Ls were identified and the conjugation enzymes for NEDD8 and URM1 were described for first time. The 3D model for Ub-Ls displayed the β-grasp fold typical. Furthermore, our sequence analysis for the corresponding activating enzymes detected the essential motifs required for conjugation. MAIN CONCLUSIONS Our findings highlight the complexity of Giardia’s Ub-conjugation system, which is drastically different from that previously reported, and provides evidence for the presence of NEDDylation and URMylation enzymes in the genome and transcriptome of G. intestinalis. <![CDATA[Expression of NLRP3 inflammasome in leprosy indicates immune evasion of <em>Mycobacterium leprae</em>]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100303&lng=pt&nrm=iso&tlng=pt BACKGROUND Leprosy is an infectious-contagious disease caused by Mycobacterium leprae that remain endemic in 105 countries. This neglected disease has a wide range of clinical and histopathological manifestations that are related to the host inflammatory and immune responses. More recently, the inflammasome has assumed a relevant role in the inflammatory response against microbiological agents. However, the involvement of inflammasome in leprosy remains poorly understood. OBJECTIVES The aim is to associate biomarkers of inflammasome with the different immunopathological forms of leprosy. METHODS We performed an observational, cross-sectional, and comparative study of the immunophenotypic expression of inflammasome-associated proteins in immunopathological forms of leprosy of 99 skin lesion samples by immunohistochemistry. The intensity and percentage of NLRP3, Caspase-1, Caspases-4/5, interleukin-1β and interleukin-18 immunoreactivities in the inflammatory infiltrate of skin biopsies were evaluated. FINDINGS Strong expression of NLRP3 and inflammatory Caspases-4/5 were observed in lepromatous leprosy (lepromatous pole). In addition, were observed low expression of caspase-1, interleukin-1β, and interleukin-18 in tuberculoid and lepromatous leprosy. The interpolar or borderline form showed immunophenotype predominantly similar to the lepromatous pole. MAIN CONCLUSIONS Our results demonstrate that the NLRP3 inflammasome is inactive in leprosy, suggesting immune evasion of M. leprae. <![CDATA[Effects of El Niño-Southern Oscillation on human visceral leishmaniasis in the Brazilian State of Mato Grosso do Sul]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100304&lng=pt&nrm=iso&tlng=pt BACKGROUND Leishmaniases are considered a major public health problem in South America, specifically in Brazil. Moreover, the transmission and epidemiology of leishmaniasis are possibly associated with climatic and environmental variations. OBJECTIVE This study aimed to assess the association between the extreme climatic phenomenon El Niño-Southern Oscillation (ENSO), the maximum and minimum variations of temperature, precipitation, and soil moisture and the incidence of visceral leishmaniasis (VL) in Mato Grosso do Sul (MS), Brazil, from 2002 to 2015. METHODS The Niño 3.4 index was used for the ENSO variation. The other climatic data were obtained from the climatic tool TerraClimate. Records regarding VL were obtained from the Notification of Injury Information System. FINDINGS From 2002 to 2015, there were 3,137 cases of VL recorded in MS. The annual incidence of the disease was negatively associated with the ENSO index and soil moisture in MS. The VL incidence increased during the negative phase of ENSO and decreased during the positive phase. MAIN CONCLUSIONS The results demonstrated that the interannual cycles of incidence of human VL are significantly associated with the occurrence of the ENSO phenomenon and its phases, El Niño and La Niña. <![CDATA[Glycogen synthase kinase 3ß participates in late stages of Dengue virus-2 infection]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100305&lng=pt&nrm=iso&tlng=pt BACKGROUND Viruses can modulate intracellular signalling pathways to complete their infectious cycle. Among these, the PI3K/Akt pathway allows prolonged survival of infected cells that favours viral replication. GSK3β, a protein kinase downstream of PI3K/Akt, gets inactivated upon activation of the PI3K/Akt pathway, and its association with viral infections has been recently established. In this study, the role of GSK3β during Dengue virus-2 (DENV-2) infection was investigated. METHODS GSK3β participation in the DENV-2 replication process was evaluated with pharmacological and genetic inhibition during early [0-12 h post-infection (hpi)], late (12-24 hpi), and 24 hpi in Huh7 and Vero cells. We assessed the viral and cellular processes by calculating the viral titre in the supernatants, In-Cell Western, western blotting and fluorescence microscopy. RESULTS Phosphorylation of GSK3β-Ser9 was observed at the early stages of infection; neither did treatment with small molecule inhibitors nor pre-treatment prior to viral infection of GSK3β reduce viral titres of the supernatant at these time points. However, a decrease in viral titres was observed in cells infected and treated with the inhibitors much later during viral infection. Consistently, the infected cells at this stage displayed plasma membrane damage. Nonetheless, these effects were not elicited with the use of genetic inhibitors of GSK3β. CONCLUSIONS The results suggest that GSK3β participates at the late stages of the DENV replication cycle, where viral activation may promote apoptosis and release of viral particles. <![CDATA[Human papillomavirus type 18 E5 oncoprotein cooperates with E6 and E7 in promoting cell viability and invasion and in modulating the cellular redox state]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100306&lng=pt&nrm=iso&tlng=pt BACKGROUND High-risk human papillomaviruses (HR-HPVs) are the etiological agents of cervical cancer. Among them, types 16 and 18 are the most prevalent worldwide. The HPV genome encodes three oncoproteins (E5, E6, and E7) that possess a high transformation potential in culture cells when transduced simultaneously. In the present study, we analysed how these oncoproteins cooperate to boost key cancer cell features such as uncontrolled cell proliferation, invasion potential, and cellular redox state imbalance. Oxidative stress is known to contribute to the carcinogenic process, as reactive oxygen species (ROS) constitute a potentially harmful by-product of many cellular reactions, and an efficient clearance mechanism is therefore required. Cells infected with HR-HPVs can adapt to oxidative stress conditions by upregulating the formation of endogenous antioxidants such as catalase, glutathione (GSH), and peroxiredoxin (PRX). OBJECTIVES The primary aim of this work was to study how these oncoproteins cooperate to promote the development of certain cancer cell features such as uncontrolled cell proliferation, invasion potential, and oxidative stress that are known to aid in the carcinogenic process. METHODS To perform this study, we generated three different HaCaT cell lines using retroviral transduction that stably expressed combinations of HPV-18 oncogenes that included HaCaT E5-18, HaCaT E6/E7-18, and HaCaT E5/E6/E7-18. FINDINGS Our results revealed a statistically significant increment in cell viability as measured by MTT assay, cell proliferation, and invasion assays in the cell line containing the three viral oncogenes. Additionally, we observed that cells expressing HPV-18 E5/E6/E7 exhibited a decrease in catalase activity and a significant augmentation of GSH and PRX1 levels relative to those of E5, E6/E7, and HaCaT cells. MAIN CONCLUSIONS This study demonstrates for the first time that HPV-18 E5, E6, and E7 oncoproteins can cooperate to enhance malignant transformation. <![CDATA[Involvement of lipid microdomains in human endothelial cells infected by <em>Streptococcus agalactiae</em> type III belonging to the hypervirulent ST-17]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100307&lng=pt&nrm=iso&tlng=pt BACKGROUND Streptococcus agalactiae capsular type III strains are a leading cause of invasive neonatal infections. Many pathogens have developed mechanisms to escape from host defense response using the host membrane microdomain machinery. Lipid rafts play an important role in a variety of cellular functions and the benefit provided by interaction with lipid rafts can vary from one pathogen to another. OBJECTIVES This study aims to evaluate the involvement of membrane microdomains during infection of human endothelial cell by S. agalactiae. METHODS The effects of cholesterol depletion and PI3K/AKT signaling pathway activation during S. agalactiae-human umbilical vein endothelial cells (HUVEC) interaction were analysed by pre-treatment with methyl-β-cyclodextrin (MβCD) or LY294002 inhibitors, immunofluorescence and immunoblot analysis. The involvement of lipid rafts was analysed by colocalisation of bacteria with flotillin-1 and caveolin-1 using fluorescence confocal microscopy. FINDINGS In this work, we demonstrated the importance of the integrity of lipid rafts microdomains and activation of PI3K/Akt pathway during invasion of S. agalactiae strain to HUVEC cells. Our results suggest the involvement of flotillin-1 and caveolin-1 during the invasion of S. agalactiae strain in HUVEC cells. CONCLUSIONS The collection of our results suggests that lipid microdomain affects the interaction of S. agalactiae type III belonging to the hypervirulent ST-17 with HUVEC cells through PI3K/Akt signaling pathway. <![CDATA[Mapping the tuberculosis scientific landscape among BRICS countries: a bibliometric and network analysis]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100308&lng=pt&nrm=iso&tlng=pt BACKGROUND The five BRICS (Brazil, Russian, Indian, China, and South Africa) countries bear 49% of the world’s tuberculosis (TB) burden and they are committed to ending tuberculosis. OBJECTIVES The aim of this paper is to map the scientific landscape related to TB research in BRICS countries. METHODS Were combined bibliometrics and social network analysis techniques to map the scientific publications related to TB produced by the BRICS. Was made a descriptive statistical data covering the full period of analysis (1993-2016) and the research networks were made for 2007-2016 (8,366 records). The bubble charts were generated by VantagePoint and the networks by the Gephi 0.9.1 software (Gephi Consortium 2010) from co-occurrence matrices produced in VantagePoint. The Fruchterman-Reingold algorithm provided the networks’ layout. FINDINGS During the period 1993-2016, there were 38,315 peer-reviewed, among them, there were 11,018 (28.7%) articles related by one or more authors in a BRICS: India 38.7%; China 23.8%; South Africa 21.1%; Brazil 13.0%; and Russia 4.5% (The total was greater than 100% because our criterion was all papers with at least one author in a BRICS). Among the BRICS, there was greater interaction between India and South Africa and organisations in India and China had the highest productivity; however, South African organisations had more interaction with countries outside the BRICS. Publications by and about BRICS generally covered all research areas, especially those in India and China covered all research areas, although Brazil and South Africa prioritised infectious diseases, microbiology, and the respiratory system. MAIN CONCLUSIONS An overview of BRICS scientific publications and interactions highlighted the necessity to develop a BRICS TB research plan to increase efforts and funding to ensure that basic science research successfully translates into products and policies to help end the TB epidemic. <![CDATA[Rapid detection of <em>Mycobacterium tuberculosis</em> DNA and genetic markers for Isoniazid resistance in Ziehl-Neelsen stained slides]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100309&lng=pt&nrm=iso&tlng=pt BACKGROUND Early diagnosis of tuberculosis (TB) and identification of strains of Mycobacterium tuberculosis resistant to anti-TB drugs are considered the main factors for disease control. OBJECTIVES To standardise a real-time polymerase chain reaction (qPCR) assay technique and apply it to identify mutations involved in M. tuberculosis resistance to Isoniazid (INH) directly in Ziehl-Neelsen (ZN) stained slides. METHODS Were analysed 55 independent DNA samples extracted from clinical isolates of M. tuberculosis by sequencing. For application in TB diagnosis resistance, 59 ZN-stained slides were used. The sensitivity, specificity and Kappa index, with a 95% confidence interval (CI95%), were determined. FINDINGS The agreement between the tests was, for the katG target, the Kappa index of 0.89 (CI95%: 0.7-1.0). The sensitivity and specificity were 97.6% (CI95%: 87.7-99.9) and 91.7% (CI95%: 61.5-99.5), respectively. For inhA, the Kappa index was 0.92 (CI95%: 0.8-1.0), the sensitivity and specificity were 94.4% (CI95%: 72.7-99.8) and 97.3% (CI95%: 85.8-99.9), respectively. The use of ZN-stained slides for drug-resistant TB detection showed significant results when compared to other standard tests for drug resistance. MAIN CONCLUSIONS qPCR genotyping proved to be an efficient method to detect genes that confer M. tuberculosis resistance to INH. Thus, qPCR genotyping may be an alternative instead of sequencing. <![CDATA[<em>Leishmania infantum</em> induces high phagocytic capacity and intracellular nitric oxide production by human proinflammatory monocyte]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100310&lng=pt&nrm=iso&tlng=pt BACKGROUND The mechanism of resistance to SbIII in Leishmania is complex, multifactorial and involves not only biochemical mechanisms, but also other elements, such as the immune system of the host. OBJECTIVES In this study, putative changes in the immunological profile of human monocytes infected with wild-type (WT) and antimony (SbIII)-resistant Leishmania (Viannia) braziliensis and Leishmania (Leishmania) infantum lines were evaluated. METHODS Susceptibility assays WT and SbIII-resistant L. braziliensis and L. infantum were performed using lines THP-1 human monocytic lineage. Phagocytic capacity, cytokine profile, intracellular nitric oxide (NO) production and surface carbohydrate residues profile were performed in peripheral blood monocytes by flow cytometry. FINDINGS The phagocytic capacity and intracellular NO production by classical (CD14++CD16-) and proinflammatory (CD14++CD16+) monocytes were higher in the presence of L. infantum lines compared to L. braziliensis lines. The results also highlight proinflammatory monocytes as the cellular subpopulation of major relevance in a phagocytosis event and NO expression. It is important to note that L. infantum induced a proinflammatory cytokine profile characterised by higher levels of TNF-α in culture supernatant than L. braziliensis. Conversely, both Leishmania lines induce high levels of IL-6 in culture supernatant. Analysis of the expression profile of surface carbohydrates showed that L. braziliensis presents 4.3-fold higher expression of galactose(β1,4)N-acetylglucosamine than L. infantum line. Interestingly, the expression level of α-N-acetylgalactosamine residues was 2-fold lower in the SbIII-resistant L. braziliensis line than its counterpart WT line, indicating differences in surface glycoconjugates between these lines. MAIN CONCLUSIONS Our results showed that L. braziliensis and L. infantum induce different innate immune responses and a highly inflammatory profile, which is characteristic of infection by L. infantum, the species associated with visceral disease. <![CDATA[DNA nanovaccines prepared using LemA antigen protect Golden Syrian hamsters against <em>Leptospira</em> lethal infection]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100311&lng=pt&nrm=iso&tlng=pt BACKGROUND Nanoparticles (NPs) are viable candidates as carriers of exogenous materials into cells via transfection and can be used in the DNA vaccination strategy against leptospirosis. OBJECTIVES We evaluated the efficiency of halloysite clay nanotubes (HNTs) and amine-functionalised multi-walled carbon nanotubes (NH2-MWCNTs) in facilitating recombinant LemA antigen (rLemA) expression and protecting Golden Syrian hamsters (Mesocricetus auratus) against Leptospira interrogans lethal infection. METHODS An indirect immunofluorescent technique was used to investigate the potency of HNTs and NH2-MWCNTs in enhancing the transfection and expression efficiency of the DNA vaccine in Chinese hamster ovary (CHO) cells. Hamsters were immunised with two doses of vaccines HNT-pTARGET/lemA, NH2-MWCNTs-pTARGET/lemA, pTARGET/lemA, and empty pTARGET (control), and the efficacy was determined in terms of humoral immune response and protection against a lethal challenge. FINDINGS rLemA DNA vaccines carried by NPs were able to transfect CHO cells effectively, inducing IgG immune response in hamsters (p &lt; 0.05), and did not exhibit cytotoxic effects. Furthermore, 83.3% of the hamsters immunised with NH2-MWCNTs-pTARGET/lemA were protected against the lethal challenge (p &lt; 0.01), and 66.7% of hamsters immunised with HNT-pTARGET/lemA survived (p &lt; 0.05). MAIN CONCLUSIONS NH2-MWCNTs and HNTs can act as antigen carriers for mammalian cells and are suitable for DNA nanovaccine delivery. <![CDATA[Minor temperature shifts do not affect chromosomal ploidy but cause transcriptomic changes in <em>Leishmania braziliensis</em> promastigotes <em>in vitro</em>]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100312&lng=pt&nrm=iso&tlng=pt BACKGROUND The leishmaniases are complex neglected diseases caused by protozoan parasites of the genus Leishmania. Leishmania braziliensis is the main etiological agent of cutaneous leishmaniasis in the New World. In recent studies, genomic changes such as chromosome and gene copy number variations (CNVs), as well as transcriptomic changes have been highlighted as mechanisms used by Leishmania species to adapt to stress situations. OBJECTIVES The aim of this study was to determine the effect of short-term minor temperature shifts in the genomic and transcriptomic responses of L. braziliensis promastigotes in vitro. METHODS Growth curves, genome and transcriptome sequencing of L. braziliensis promastigotes were conducted from cultures exposed to three different temperatures (24ºC, 28ºC and 30ºC) compared with the control temperature (26ºC). FINDINGS Our results showed a decrease in L. braziliensis proliferation at 30ºC, with around 3% of the genes showing CNVs at each temperature, and transcriptomic changes in genes encoding amastin surface-like proteins, heat shock proteins and transport proteins, which may indicate a direct response to temperature stress. MAIN CONCLUSIONS This study provides evidence that L. braziliensis promastigotes exhibit a decrease in cell density, and noticeable changes in the transcriptomic profiles. However, there were not perceptible changes at chromosome CNVs and only ~3% of the genes changed their copies in each treatment. <![CDATA[A cross-sectional approach including dog owner characteristics as predictors of visceral leishmaniasis infection in dogs]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100313&lng=pt&nrm=iso&tlng=pt BACKGROUND Visceral leishmaniasis (VL) is relevant for human and animal public health. Several factors have been associated with the risk of Leishmania infantum infection in dogs. However, dog owner characteristics have been rarely explored. OBJECTIVES To estimate the prevalence and to identify the associated factors for VL infection including dog owners characteristics. METHODS A cross-sectional study was conducted including dogs from an endemic canine visceral leishmaniasis (CVL) region in the Federal District, Brazil. The infection was detected using parasitological, serological, and molecular methods. The associated factors were identified through Poisson regression modelling. FINDINGS The prevalence of infection was 26.25% [95% confidence interval (CI): 20.05 to 33.57]. The associated factors were: short coat prevalence ratio (PR) = 2.33 (95% CI: 1.02 to 5.22); presence of backyard with predominance of soil and/or vegetation PR = 4.15 (95% CI: 1.35 to 12.77); and highest gross family income score PR = 2.03 (95% CI: 1.16 to 3.54). MAIN CONCLUSION This is the first study that relates higher socioeconomic status of dog owners as an independent factor associated with higher prevalence of VL infection, along with other strongly associated factors related to receptive environment for phlebotomines. Our findings strengthen the need for exploration of the biological and behavioural bases linking dog owner characteristics to the risk of canine infection in prospective cohort studies. <![CDATA[Computational prediction and characterisation of miRNAs and their pathway genes in human schistosomiasis caused by <em>Schistosoma haematobium</em>]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100314&lng=pt&nrm=iso&tlng=pt BACKGROUND Key genes control the infectivity of the Schistosoma haematobium causing schistosomiasis. A method for understanding the regulation of these genes might help in developing new disease strategies to control schistosomiasis, such as the silencing mediated by microRNAs (miRNAs). The miRNAs have been studied in schistosome species and they play important roles in the post-transcriptional regulation of genes, and in parasite-host interactions. However, genome-wide identification and characterisation of novel miRNAs and their pathway genes and their gene expression have not been explored deeply in the genome and transcriptome of S. haematobium. OBJECTIVES Identify and characterise mature and precursor miRNAs and their pathway genes in the S. haematobium genome. METHODS Computational prediction and characterisation of miRNAs and genes involved in miRNA pathway from S. haematobium genome on SchistoDB. Conserved domain analysis was performed using PFAM and CDD databases. A robust algorithm was applied to identify mature miRNAs and their precursors. The characterisation of the precursor miRNAs was performed using RNAfold, RNAalifold and Perl scripts. FINDINGS We identified and characterised 14 putative proteins involved in miRNA pathway including ARGONAUTE and DICER in S. haematobium. Besides that, 149 mature miRNAs and 131 precursor miRNAs were identified in the genome including novel miRNAs. MAIN CONCLUSIONS miRNA pathway occurs in the S. haematobium, including endogenous miRNAs and miRNA pathway components, suggesting a role of this type of non-coding RNAs in gene regulation in the parasite. The results found in this work will open up a new avenue for studying miRNAs in the S. haematobium biology in helping to understand the mechanism of gene silencing in the human parasite Schistosome. <![CDATA[Frequency of first and second-line drug resistance-associated mutations among resistant <em>Mycobacterium tuberculosis</em> clinical isolates from São Paulo, Brazil]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100315&lng=pt&nrm=iso&tlng=pt BACKGROUND Tuberculosis (TB) is an infectious disease caused by the bacterium Mycobacterium tuberculosis, and the number of new cases of multidrug resistant TB (MDR-TB), pre extensively drug-resistant TB (pre-XDR-TB) and extensively drug-resistant TB (XDR-TB) has increased considerably worldwide. OBJECTIVES Herein, using 156 M. tuberculosis isolates from 106 patients previously classified as MDR or pre-XDR or XDR isolates, we investigated the genetic mutation profiles associated with phenotypic resistances in patients with MDR-TB, pre-XDR-TB and XDR-TB, treatment outcomes and resistance evolution. METHODS Molecular analyses were performed by partial sequencing of the rpoB, katG, gyrA, gyrB, rrs genes and analysis of the fabG-inhA promoter region. Clinical, epidemiologic and demographic data were obtained from the TB Notification database system of São Paulo (TB-WEB) and the Information System for Special Tuberculosis Treatments (SITE-TB). FINDINGS Drug resistance was attributed to previously known mutations and a novel Asp449Val mutation in gyrB was observed in four isolates from the same patient. Ten patients had more than one isolate evaluated and eight of these patients displayed resistance progression. MAIN CONCLUSIONS The present study is the first to report the frequency of mutations related to second-line drug resistance in MDR-TB, pre-XDR-TB and XDR-TB isolates. The results could lead to the improvement of available technologies for the rapid detection of drug resistant TB. <![CDATA[Characterisation of alternative expression vectors for recombinant Bacillus Calmette-Guérin as live bacterial delivery systems]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100316&lng=pt&nrm=iso&tlng=pt BACKGROUND Bacillus Calmette-Guérin (BCG) is considered a promising live bacterial delivery system. However, several proposals for rBCG vaccines have not progressed, mainly due to the limitations of the available expression systems. OBJECTIVES To obtain a set of mycobacterial vectors using a range of promoters with different strengths based on a standard backbone, previously shown to be stable. METHODS Mycobacterial expression vectors based on the pLA71 vector as backbone, were obtained inserting different promoters (PAN, PαAg, PHsp60, PBlaF* and PL5) and the green fluorescence protein (GFP) as reporter gene, to evaluate features such as their relative strengths, and the in vitro (inside macrophages) and in vivo stability. FINDINGS The relative fluorescence observed with the different vectors showed increasing strength of the promoters: PAN was the weakest in both Mycobacterium smegmatis and BCG and PBlaF* was higher than PHsp60 in BCG. The relative fluorescence observed in a macrophage cell line showed that PBlaF* and PHsp60 were comparable. It was not possible to obtain strains transformed with the extrachromosomal expression vector containing the PL5 in either species. MAIN CONCLUSION We have obtained a set of potentially stable mycobacterial vectors with a arrange of expression levels, to be used in the development of rBCG vaccines. <![CDATA[Genomic detection of a virus lineage replacement event of dengue virus serotype 2 in Brazil, 2019]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100317&lng=pt&nrm=iso&tlng=pt BACKGROUND Despite efforts to mitigate the impact of dengue virus (DENV) epidemics, the virus remains a public health problem in tropical and subtropical regions around the world. Most DENV cases in the Americas between January and July 2019 were reported in Brazil. São Paulo State in the southeast of Brazil has reported nearly half of all DENV infections in the country. OBJECTIVES To understand the origin and dynamics of the 2019 DENV outbreak. METHODS Here using portable nanopore sequencing we generated20 new DENV genome sequences from viremic patients with suspected dengue infection residing in two of the most-affected municipalities of São Paulo State, Araraquara and São José do Rio Preto. We conducted a comprehensive phylogenetic analysis with 1,630 global DENV strains to better understand the evolutionary history of the DENV lineages that currently circulate in the region. FINDINGS The new outbreak strains were classified as DENV2 genotype III (American/Asian genotype). Our analysis shows that the 2019 outbreak is the result of a novel DENV lineage that was recently introduced to Brazil from the Caribbean region. Dating phylogeographic analysis suggests that DENV2-III BR-4 was introduced to Brazil in or around early 2014, possibly from the Caribbean region. MAIN CONCLUSIONS Our study describes the early detection of a newly introduced and rapidly-expanding DENV2 virus lineage in Brazil. <![CDATA[Characterising ISWI chromatin remodeler in <em>Trypanosoma cruzi</em>]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100318&lng=pt&nrm=iso&tlng=pt BACKGROUND Imitation SWItch (ISWI) ATPase is the catalytic subunit in diverse chromatin remodeling complexes. These complexes modify histone-DNA interactions and therefore play a pivotal role in different DNA-dependent processes. In Trypanosoma cruzi, a protozoan that controls gene expression principally post-transcriptionally, the transcriptional regulation mechanisms mediated by chromatin remodeling are poorly understood. OBJECTIVE To characterise the ISWI remodeler in T. cruzi (TcISWI). METHODS A new version of pTcGW vectors was constructed to express green fluorescent protein (GFP)-tagged TcISWI. CRISPR-Cas9 system was used to obtain parasites with inactivated TcISWI gene and we determined TcISWI partners by cryomilling-affinity purification-mass spectrometry (MS) assay as an approximation to start to unravel the function of this protein. FINDINGS Our approach identified known ISWI partners [nucleoplasmin-like protein (NLP), regulator of chromosome condensation 1-like protein (RCCP) and phenylalanine/tyrosine-rich protein (FYRP)], previously characterised in T. brucei, and new components in TcISWI complex [DRBD2, DHH1 and proteins containing a domain characteristic of structural maintenance of chromosomes (SMC) proteins]. Data are available via ProteomeXchange with identifier PXD017869. MAIN CONCLUSIONS In addition to its participation in transcriptional silencing, as it was reported in T. brucei, the data generated here provide a framework that suggests a role for TcISWI chromatin remodeler in different nuclear processes in T. cruzi, including mRNA nuclear export control and chromatin compaction. Further work is necessary to clarify the TcISWI functional diversity that arises from this protein interaction study. <![CDATA[Influenza A(H1N1)pdm09 infection and viral load analysis in patients with different clinical presentations]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100319&lng=pt&nrm=iso&tlng=pt BACKGROUND Influenza viral load (VL) can be a decisive factor in determining the antiviral efficacy in viral clearance. OBJECTIVE This study aimed to evaluate the rate of infection and the role of influenza VL on the clinical spectrum of illnesses among different patient groups attended at a tertiary hospital in Brazil. METHODS Samples were collected from patients presenting acute respiratory infection from 2009 to 2013. Overall, 2262 samples were analysed and distributed into three groups: (i) asymptomatic (AS); (ii) symptomatic outpatients (OP); and (iii) hospitalised patients (HP). VL (expressed in Log10 RNA copies/mL) was calculated through a quantitative real-time one-step reverse transcription-polymerase chain reaction (RT-PCR) assay aimed at the M gene, with human RNAseP target as internal control and normalising gene of threshold cycle values. FINDINGS A total of 162 (7.16%) H1N1pdm09 positive samples were analysed. Patients aged from 0.08 to 77 years old [median ± standard deviation (SD): 12.5 ± 20.54]. Children with 5 to 11 years old presented the highest detection (p &lt; 0.0001). AS patients had the lowest VL, with a significant difference when compared with symptomatic patients (p = 0.0003). A higher VL was observed within two days of disease onset. Ten patients (HP group) received antiviral treatment and were followed up and presented a mean initial VL of 6.64 ± 1.82. A complete viral clearance for 50% of these patients was reached after 12 days of treatment. MAIN CONCLUSIONS It is important to evaluate AS patients as potential spreaders, as viral shedding was still present, even at lower VL. Our results suggest that patients with underlying diseases and severe clinical symptoms may be considered for prolonged viral treatment. <![CDATA[Infestation of an endemic arbovirus area by sympatric populations of <em>Aedes aegypti</em> and <em>Aedes albopictus</em> in Brazil]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100320&lng=pt&nrm=iso&tlng=pt BACKGROUND Aedes aegypti and Aedes albopictus are the most important arbovirus vectors in the world. OBJECTIVES This study aimed to investigate and compare the infestation pattern of these species in a neighbourhood of Recife, Brazil, endemic for arboviruses in 2005 (T1) and 2013 (T2). METHODS Infestation, distribution and relative abundance of these sympatric species were recorded by egg collection using a network of 59 sentinel ovitraps (s-ovt) at fixed sampling stations for 12 months in T1 and T2. FINDINGS A permanent occupation pattern was detected which was characterised by the presence of egg-laying females of one or both species with a high ovitrap positivity index (94.3 to 100%) throughout both years analysed. In terms of abundance, the total of eggs collected was lower (p &lt; 0.005) in T2 (146,153) than in T1 (281,103), although ovitraps still displayed a high index of positivity. The spatial distribution showed the presence of both species in 65.1% of the 148 s-ovt assessed, while a smaller number of traps exclusively contained Ae. aegypti (22%) or Ae. albopictus (13.2%) eggs. MAIN CONCLUSIONS Our comparative analysis demonstrated the robustness of the spatial occupation and permanence of Ae. aegypti and Ae. albopictus populations in this endemic urban area. <![CDATA[Chagas disease in Virgem da Lapa, Minas Gerais, Brazil: left ventricle aneurysm and the risk of death in the 24-year interval]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100321&lng=pt&nrm=iso&tlng=pt BACKGROUND Left ventricular aneurysm (LVA) is indicator of high morbidity in Chagas’ disease. A cross-sectional study performed identified LVA in 18.8% of the chronic chagasic patients (CCP). OBJECTIVE Determine the risk of death of patients with chronic chagasic cardiopathy (CCC) and LVA in 24-year interval. MATERIAL AND METHODS In 1995 a cohort of 298 CCP was evaluated by anamnesis, physical examination, EKG and ECHO and classified in groups: G0 = 86 without cardiopathy; G1 = 156 with cardiopathy without LVA and G2 = 56 with cardiopathy and LVA. 38 patients of G0 and G1 used benznidazole. Information about the deaths was obtained in the notary, death certificates, hospital records and family members. FINDINGS Were registered 113 deaths (37.9%): 107 (35.9%) attributed to cardiopathy and 6 (2.0%) to other causes (p &lt; 0.05). Amongst these 107 deaths, 10 (11.6%) occurred in G0; 49 (31.4%) occurred in G1 and 48 (85.7%) occurred in G2 (p &lt; 0.05). The risk of death was 2.7 and 7.4 times significantly higher in G2, than in G1 and G0, respectively. CONCLUSION Chronic chagasic patients with LVA and ejection fraction &lt; 45% have a higher risk of death than those without. <![CDATA[Analysing ambiguities in trypanosomatids taxonomy by barcoding]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100322&lng=pt&nrm=iso&tlng=pt BACKGROUND Biodiversity screens and phylogenetic studies are dependent on reliable DNA sequences in public databases. Biological collections possess vouchered specimens with a traceable history. Therefore, DNA sequencing of samples available at institutional collections can greatly contribute to taxonomy, and studies on evolution and biodiversity. METHODS We sequenced part of the glycosomal glyceraldehyde phosphate dehydrogenase (gGAPDH) and the SSU rRNA (V7/V8) genes from 102 trypanosomatid cultures, which are available on request at www.colprot.fiocruz.br. OBJECTIVE The main objective of this work was to use phylogenetic inferences, using the obtained DNA sequences and those from representatives of all Trypanosomatidae genera, to generate phylogenetic trees that can simplify new isolates screenings. FINDINGS A DNA sequence is provided for the first time for several isolates, the phylogenetic analysis allowed the classification or reclassification of several specimens, identification of candidates for new genera and species, as well as the taxonomic validation of several deposits. MAIN CONCLUSIONS This survey aimed at presenting a list of validated species and their associated DNA sequences combined with a short historical overview of each isolate, which can support taxonomic and biodiversity research and promote culture collections. <![CDATA[Deep sequencing of small RNAs reveals the repertoire of miRNAs and piRNAs in <em>Biomphalaria glabrata</em>]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100323&lng=pt&nrm=iso&tlng=pt BACKGROUND Biomphalaria glabrata snails are widely distributed in schistosomiasis endemic areas like America and Caribe, displaying high susceptibility to infection by Schistosoma mansoni. After the availability of B. glabrata genome and transcriptome data, studies focusing on genetic markers and small non-coding RNAs have become more relevant. The small RNAs have been considered important through their ability to finely regulate the gene expression in several organisms, thus controlling the functions like cell growth, metabolism, and susceptibility/resistance to infection. OBJECTIVE The present study aims on identification and characterisation of the repertoire of small non-coding RNAs in B. glabrata (Bgl-small RNAs). METHODS By using small RNA sequencing, bioinformatics tools and quantitative reverse transcription polymerase chain reaction (RT-qPCR), we identified, characterised, and validated the presence of small RNAs in B. glabrata. FINDINGS 89 mature miRNAs were identified and five of them were classified as Mollusk-specific. When compared to model organisms, sequences of B. glabrata miRNAs showed a high degree of conservation. In addition, several target genes were predicted for all the mature miRNAs identified. Furthermore, piRNAs were identified in the genome of B. glabrata for the first time. The B. glabrata piRNAs showed strong conservation of uridine as first nucleotide at 5’ end, besides adenine at 10th position. Our results showed that B. glabrata has diverse repertoire of circulating ncRNAs, several which might be involved in mollusk susceptibility to infection, due to their potential roles in the regulation of S. mansoni development. MAIN CONCLUSIONS Further studies are necessary in order to confirm the role of the Bgl-small RNAs in the parasite/host relationship thus opening new perspectives on interference of small RNAs in the organism development and susceptibility to infection. <![CDATA[The heterologous expression of <em>Escherichia coli</em> MutT enzyme is involved in the protection against oxidative stress in <em>Leishmania braziliensis</em>]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100324&lng=pt&nrm=iso&tlng=pt BACKGROUND Oxidative stress is responsible for generating DNA lesions and the 8-oxoguanine (8-oxoG) is the most commonly lesion found in DNA damage. When this base is incorporated during DNA replication, it could generate double-strand DNA breaks and cellular death. MutT enzyme hydrolyzes the 8-oxoG from the nucleotide pool, preventing its incorporation during DNA replication. OBJECTIVES To investigate the importance of 8-oxoG in Leishmania infantum and L. braziliensis, in this study we analysed the impact of heterologous expression of Escherichia coli MutT (EcMutT) enzyme in drug-resistance phenotype and defense against oxidative stress. METHODS Comparative analysis of L. braziliensis and L. infantum H2O2 tolerance and cell cycle profile were performed. Lines of L. braziliensis and L. infantum expressing EcMutT were generated and evaluated using susceptibility tests to H2O2 and SbIII, cell cycle analysis, γH2A western blotting, and BrdU native detection assay. FINDINGS Comparative analysis of tolerance to oxidative stress generated by H2O2 showed that L. infantum is more tolerant to exogenous H2O2 than L. braziliensis. In addition, cell cycle analysis showed that L. infantum, after treatment with H2O2, remains in G1 phase, returning to its normal growth rate after 72 h. In contrast, after treatment with H2O2, L. braziliensis parasites continue to move to the next stages of the cell cycle. Expression of the E. coli MutT gene in L. braziliensis and L. infantum does not interfere in parasite growth or in susceptibility to SbIII. Interestingly, we observed that L. braziliensis EcMutT-expressing clones were more tolerant to H2O2 treatment, presented lower activation of γH2A, a biomarker of genotoxic stress, and lower replication stress than its parental non-transfected parasites. In contrast, the EcMutT is not involved in protection against oxidative stress generated by H2O2 in L. infantum. MAIN CONCLUSIONS Our results showed that 8-oxoG clearance in L. braziliensis is important to avoid misincorporation during DNA replication after oxidative stress generated by H2O2. <![CDATA[Description of an automatic copulation induction system used to establish a free-mating laboratory colony of <em>Nyssorhynchus deaneorum</em> from Brazil]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100325&lng=pt&nrm=iso&tlng=pt BACKGROUND Nyssorhynchus deaneorum is a potential malaria vector because it has been shown to be competent to transmit Plasmodium vivax and Plasmodium falciparum, and because it exhibits antropophilic and endophilic behaviors in some regions of the Amazon. This profile makes Ny. deaneorum a useful mosquito for experiments that model Plasmodium-vector interactions in the Amazon. OBJECTIVE Herein we describe how a free-mating colony of Ny. deaneorum has been established using an automated light stimulation system. METHODS Mosquitoes were captured in São Francisco do Guaporé, Rondônia. The F1 generation was reared until adult emergence at which point copulation was induced using an automatic copulation induction system (ACIS). FINDINGS After four generations, natural mating and oviposition began to occur without light stimulation. The number of pupae and adult mosquitoes increased from the F5 to F10 generations. The new Ny. deaneorum colony exhibited susceptibility to P. vivax. MAIN CONCLUSIONS Automated light stimulation is an effective method for establishing an Ny. deaneorum colony under laboratory conditions as it produces enough adults to create a stenogamic colony. The establishment of a stable, P. vivax-susceptible colony of Ny. deaneorum makes it possible to model parasite-vector interactions and to test novel drug therapies that target parasite development in mosquitoes. <![CDATA[The influence of meteorological variables on the oviposition dynamics of <em>Aedes aegypti</em> (Diptera: Culicidae) in four environmentally distinct areas in northeast Brazil]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100326&lng=pt&nrm=iso&tlng=pt BACKGROUND Fluctuations in climate have been associated with variations in mosquito abundance. OBJECTIVES To analyse the influence of precipitation, temperature, solar radiation, wind speed and humidity on the oviposition dynamics of Aedes aegypti in three distinct environmental areas (Brasília Teimosa, Morro da Conceição/Alto José do Pinho and Dois Irmãos/Pintos) of the city of Recife and the Fernando de Noronha Archipelago northeastern Brazil. METHODS Time series study using a database of studies previously carried out in the areas. The eggs were collected using spatially distributed geo-referenced sentinel ovitraps (S-OVTs). Meteorological satellite data were obtained from the IRI climate data library. The association between meteorological variables and egg abundance was analysed using autoregressive models. FINDINGS Precipitation was positively associated with egg abundance in three of the four study areas with a lag of one month. Higher humidity (β = 45.7; 95% CI: 26.3 - 65.0) and lower wind speed (β = −125.2; 95% CI: −198.8 - −51.6) were associated with the average number of eggs in the hill area. MAIN CONCLUSIONS The effect of climate variables on oviposition varied according to local environmental conditions. Precipitation was a main predictor of egg abundance in the study settings. <![CDATA[The impact of imported malaria by gold miners in Roraima: characterizing the spatial dynamics of autochthonous and imported malaria in an urban region of Boa Vista]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100327&lng=pt&nrm=iso&tlng=pt BACKGROUND The number of malaria cases in Roraima nearly tripled from 2016 to 2018. The capital, Boa Vista, considered a low-risk area for malaria transmission, reported an increasing number of autochthonous and imported cases. OBJECTIVES This study describes a spatial analysis on malaria cases in an urban region of Boa Vista, which sought to identify the autochthonous and imported cases and associated them with Anopheles habitats and the potential risk of local transmission. METHODS In a cross-sectional study at the Polyclinic Cosme e Silva, 520 individuals were interviewed and diagnosed with malaria by microscopic examination. Using a global positional system, the locations of malaria cases by type and origin and the breeding sites of anopheline vectors were mapped and the risk of malaria transmission was evaluated by spatial point pattern analysis. FINDINGS Malaria was detected in 57.5% of the individuals and there was a disproportionate number of imported cases (90.6%) linked to Brazilian coming from gold mining sites in Venezuela and Guyana. MAIN CONCLUSIONS The increase in imported malaria cases circulating in the west region of Boa Vista, where there are positive breeding sites for the main vectors, may represent a potential condition for increased autochthonous malaria transmission in this space. <![CDATA[Evaluation of different total <em>Leishmania amazonensis</em> antigens for the development of a first-generation vaccine formulated with a Toll-like receptor-3 agonist to prevent cutaneous leishmaniasis]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100328&lng=pt&nrm=iso&tlng=pt BACKGROUND Unfortunately, no any vaccine against leishmaniasis has been developed for human use. Therefore, a vaccine based on total Leishmania antigens could be a good and economic approach; and there are different methodologies to obtain these antigens. However, it is unknown whether the method to obtain the antigens affects the integrity and immune response caused by them. OBJECTIVES to compare the protein profile and immune response generated by total L. amazonensis antigens (TLA) produced by different methods, as well as to analyse the immune response and protection by a first-generation vaccine formulated with sonicated TLA (sTLA) and polyinosinic:polycytidylic acid [Poly (I:C)]. METHODS TLA were obtained by four different methodologies and their integrity and immune response were evaluated. Finally, sTLA was formulated with Poly (I:C) and their protective immune response was measured. FINDINGS sTLA presented a conserved protein profile and induced a strong immune response. In addition, Poly (I:C) improved the immune response generated by sTLA. Finally, sTLA + Poly (I:C) formulation provided partial protection against L. amazonensis infection. MAIN CONCLUSIONS The protein profile and immune response depend on the methodology used to obtain the antigens. Also, the formulation sTLA + Poly (I:C) provides partial protection against cutaneous leishmaniasis in mice. <![CDATA[Assessment of <em>Culex pipiens</em> bioforms in the world’s southernmost distribution limit]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100400&lng=pt&nrm=iso&tlng=pt The mosquito Culex pipiens s.s. L. occurs as two bioforms that differ in physiology and behaviour affecting virus transmission cycles. To assess the occurrence of Cx. pipiens bioforms in the southernmost limit of its distribution, specimens were collected aboveground in southern Buenos Aires Province and east Patagonia, Argentina. Ten larvae and 25 adults were individually processed and identified by polymerase chain reaction (PCR) amplification of Ace-2 and CQ11 loci. Culex quinquefasciatus Say (one larva, two adults), Cx. pipiens f. molestus (one larva, one adult) and one adult of hybrid origin were identified in Buenos Aires Province; only Cx. pipiens f. molestus was recorded in Patagonia (eight larvae, 21 adults). The potential absence of bioform pipiens and its implications in arbovirus enzootic cycles is discussed. <![CDATA[Oropouche virus detection in saliva and urine]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100401&lng=pt&nrm=iso&tlng=pt Oropouche virus (OROV) is an arthropod-borne virus of the Peribunyaviridae family, transmitted to humans primarily by Culicoides paraensis. It is one of the main arboviruses infecting humans in Brazil, primarily in the Amazon Region. Here, we report the detection of OROV in the saliva and urine of a patient whose samples were collected five days after the onset of symptoms. Nucleotide sequencing and phylogenetic analysis further confirmed the results. To our knowledge, this is the first study reporting the detection of OROV in the saliva and urine of an infected patient. In addition, the results of our study expand the current knowledge pertaining to the natural history of Oropouche fever. <![CDATA[Human acute Chagas disease: changes in factor VII, activated protein C and hepatic enzymes from patients of oral outbreaks in Pará State (Brazilian Amazon)]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100402&lng=pt&nrm=iso&tlng=pt Oral transmission of Chagas disease has been increasing in Latin American countries. The present study aimed to investigate changes in hepatic function, coagulation factor levels and parasite load in human acute Chagas disease (ACD) secondary to oral Trypanosoma cruzi transmission. Clinical and epidemiological findings of 102 infected individuals attended in the State of Pará from October 2013 to February 2016 were included. The most common symptoms were fever (98%), asthenia (83.3%), face and limb edema (80.4%), headache (74.5%) and myalgia (72.5%). The hepatic enzymes alanine aminotransferase (ALT) and aspartate aminotransferase (AST) of 30 ACD patients were higher compared with controls, and this increase was independent of the treatment with benznidazole. Moreover, ACD individuals had higher plasma levels of activated protein C and lower levels of factor VII of the coagulation cascade. Patients with the highest parasite load had also the most increased transaminase levels. Also, ALT and AST were associated moderately (r = 0.429) and strongly (r = 0.595) with parasite load respectively. In conclusion, the present study raises the possibility that a disturbance in coagulation and hepatic function may be linked to human ACD. <![CDATA[<em>FLI1</em> gene influences lesion size and skin test may predict therapeutic response in cutaneous leishmaniasis]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100403&lng=pt&nrm=iso&tlng=pt Genes associated with wound healing have been shown to be risk factors for cutaneous leishmaniasis (CL) which is caused by Leishmania braziliensis. In this study, we examined whether the genes previously associated with CL influenced the clinical outcome. Patients were genotyped and retrospectively classified as responders, who were cured with a single course of pentavalent antimony (Sbv), or as refractories, who did not respond to Sbv. Patients characterised as responders showed a stronger response to the leishmanin skin test (LST) when compared to the refractory subjects (p = 0.0003). Furthermore, we observed an association between the FLI1 CC genotype and an increased size of ulcers (p = 0.0170). We suggest that the leishmanin skin test may be a predictive tool for therapeutic outcome and reinforce FLI1 as a potential influencer of susceptibility and lesion size in CL. <![CDATA[<em>Aedes aegypti</em> spreading in South America: new coldest and southernmost records]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100404&lng=pt&nrm=iso&tlng=pt The geographic distribution of Aedes (Stegomyia) aegypti (L.) in South America has been expanding during the last decades. Herein we present two new distribution records that extend its southern limits towards localities with extremer environmental conditions than reported to date. San Antonio Oeste constitutes the southernmost finding for the continent (40º44’S), whereas Tandil is the infested locality with the coldest mean annual temperature in Argentina (14.17ºC). The projection of a previous distribution model for Ae. aegypti predicts these two cities as positive and suggests several other localities with suitable conditions for vector proliferation beyond its assumed distribution limits. <![CDATA[<em>Schistosoma mansoni</em> granulomas in the skeletal striated muscles in the murine model of neuroschistosomiasis: histological findings]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100405&lng=pt&nrm=iso&tlng=pt Schistosomiasis mansoni presents many clinical manifestations during migration of schistosomes in their hosts, including diarrhea, hepatomegaly, splenomegaly, liver abscesses, skinlesions, brain tumors and myeloradiculopathy. No lesions have been reported in skeletal striated muscles due to schistosomiasis mansoni in the literature. This short communication reports the histopathological findings on skeletal musculature in a murine model of neuroeschistosomiasis mansoni. Lesions were found in the tongue, masseter muscle, buccinator muscle, digastric muscle and temporalis muscle. Worm recovery was carried out to confirm the infection. We describe here, for the first time in the literature, injuries in the skeletal musculature due to Schistosoma mansoni nfection. <![CDATA[Newly sequenced genomes of four Bacillus Calmette Guerin vaccines]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100406&lng=pt&nrm=iso&tlng=pt Bacillus Calmette Guerin (BCG) vaccines comprise a family of related strains. Whole genome sequencing has allowed the better characterisation of the differences between many of the BCG vaccines. As sequencing technologies improve, updating of publicly available sequence data becomes common practice. We hereby announce the draft genome of four commonly used BCG vaccines in Brazil, Argentina and Venezuela. <![CDATA[Coagulation modifiers targeting SARS-CoV-2 main protease Mpro for COVID-19 treatment: an <em>in silico</em> approach]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100407&lng=pt&nrm=iso&tlng=pt Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection depends on viral polyprotein processing, catalysed by the main proteinase (Mpro). The solution of the SARS-CoV-2 Mpro structure allowed the investigation of potential inhibitors. This work aims to provide first evidences of the applicability of commercially approved drugs to treat coronavirus disease-19 (COVID-19). We screened 4,334 compounds to found potential inhibitors of SARS-CoV-2 replication using an in silico approach. Our results evidenced the potential use of coagulation modifiers in COVID-19 treatment due to the structural similarity of SARS-CoV-2 Mpro and human coagulation factors thrombin and Factor Xa. Further in vitro and in vivo analysis are needed to corroborate these results. <![CDATA[Superinfection exclusion studies using West Nile virus and Culex flavivirus strains from Argentina]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100408&lng=pt&nrm=iso&tlng=pt In Argentina, many Flavivirus were recognised including West Nile virus (WNV). During 2009 several strains of Culex Flavivirus (CxFV), an insect-specific flavivirus, were isolated in the same region where circulation of WNV was detected. Hence, the objective of this study was to analyse the effect of co-infection in vitro assays using CxFV and WNV Argentinean strains in order to evaluate if CxFV could affect WNV replication. Our results showed that WNV replication was suppressed when multiplicity of infection (MOI) for CxFV was 10 or 100 times higher than WNV. Nevertheless, in vivo assays are necessary in order to evaluate the superinfection exclusion potential. <![CDATA[Infection by <em>Angiostrongylus cantonensis</em> in both humans and the snail <em>Achatina (Lissachatina) fulica</em> in the city of Macapá, in the Amazon Region of Brazil]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100409&lng=pt&nrm=iso&tlng=pt In January and February 2019, a malacological survey was conducted in the area surrounding the residence of a 12-year-old child that had contracted cerebral angiostrongyliasis in the municipality of Macapá, capital of the Amapá State, northern Brazil. The serological examination was positive for Angiostrongylus cantonensis infection, the principal etiological agent of this parasitosis. A sample of 54 molluscs was artificially and individually digested for parasitological analysis, containing 38 specimens of Achatina fulica, nine specimens of Bulimulus tenuissimus and seven specimens of Sarasinula linguaeformis. A. fulica was the most abundant mollusc, and the only species infected with A. cantonensis, as well as presenting co-infections with other nematodes. This is the first report of cerebral angiostrongyliasis in the Amazon Region, and the first record of A. fulica infected with A. cantonensis in Amapá. These findings highlight the potential risks of human angiostrongyliasis, and the need to implement public health measures to control the spread of the disease. <![CDATA[SARS-CoV-2 diagnostic diary: from rumors to the first case. Early reports of molecular tests from the military research and diagnostic institute of Rio de Janeiro]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100410&lng=pt&nrm=iso&tlng=pt Corona virus disease (COVID-19) presents a serious threat to global health. A historical timeline of early molecular diagnostics from government alert (January 22) (D) was presented. After in silico analysis, Brazilian Army Institute of Biology (IBEx-RJ) tested samples in house using real-time reverse transcriptase polymerase chain reaction (RT-PCR) (fast mode) based on Centers for Disease Control and Prevention (CDC) recommendations. First cases from Brazil, Rio de Janeiro, IBEx, and diagnosis team were reported in D36, D44, D66, and D74 respectively. Therefore, after 1300 tests, we recommend N1/N2 primer sets (CDC) for preliminary and Charité protocol confirmation in case of positive results. Moreover, every professional should be tested before starting work, in addition to weekly tests for everyone involved. <![CDATA[Phylogenetics applied to the human immunodeficiency virus type 1 (HIV-1): from the cross-species transmissions to the contact network inferences]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000100850&lng=pt&nrm=iso&tlng=pt Phylogenetic analyses were crucial to elucidate the origin and spread of the pandemic human immunodeficiency virus type 1 (HIV-1) group M virus, both during the pre-epidemic period of cryptic dissemination in human populations as well as during the epidemic phase of spread. The use of phylogenetics and phylodynamics approaches has provided important insights to track the founder events that resulted in the spread of HIV-1 strains across vast geographic areas, specific countries and within geographically restricted communities. In the recent years, the use of phylogenetic analysis combined with the huge availability of HIV sequences has become an increasingly important approach to reconstruct HIV transmission networks and understand transmission dynamics in concentrated and generalised epidemics. Significant efforts to obtain viral sequences from newly HIV-infected individuals could certainly contribute to detect rapidly expanding HIV-1 lineages, identify key populations at high-risk and understand what public health interventions should be prioritised in different scenarios. <![CDATA[ERRATUM: Gene regulatory network inference and analysis of multidrug-resistant Pseudomonas aeruginosa]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762020000101100&lng=pt&nrm=iso&tlng=pt Phylogenetic analyses were crucial to elucidate the origin and spread of the pandemic human immunodeficiency virus type 1 (HIV-1) group M virus, both during the pre-epidemic period of cryptic dissemination in human populations as well as during the epidemic phase of spread. The use of phylogenetics and phylodynamics approaches has provided important insights to track the founder events that resulted in the spread of HIV-1 strains across vast geographic areas, specific countries and within geographically restricted communities. In the recent years, the use of phylogenetic analysis combined with the huge availability of HIV sequences has become an increasingly important approach to reconstruct HIV transmission networks and understand transmission dynamics in concentrated and generalised epidemics. Significant efforts to obtain viral sequences from newly HIV-infected individuals could certainly contribute to detect rapidly expanding HIV-1 lineages, identify key populations at high-risk and understand what public health interventions should be prioritised in different scenarios.