Henrique Silveira - Academia.edu (original) (raw)
Papers by Henrique Silveira
Global Heart, 2015
Background: Patients with Chagas disease are at increased risk for stroke that may result in majo... more Background: Patients with Chagas disease are at increased risk for stroke that may result in major clinical disability and death. Identification of risk factors involved in the genesis of thromboembolic events related to this disease may lead to improved therapeutic decision making and outcomes. Objectives: This study sought to assess the prevalence of ischemic cerebrovascular events (ICE) among patients with Chagas heart disease and to identify the risk factors associated with cardioembolism in this population. Methods: This study involved 330 patients, 193 were men (58%), with a mean age of 49 AE 12 years with Chagas disease classified in the chronic cardiac form of the disease. Comprehensive echocardiography was performed to search a substrate for cardioembolic events, especially apical aneurysm and intracavitary thrombus. Results: Most of the patients were classified as New York Heart Association classes I or II (75%) with mean left ventricular (LV) ejection fraction of 39 AE 14%. Sixty-seven patients had a previous ICE with the overall prevalence of 20%. Apical aneurysms were detected in 128 patients (39%), whereas LV mural thrombi were found in 48 patients (15%). In multivariate analysis including the potential predictors of ICE, apical aneurysm (adjusted odds ratio [OR]: 2.19, 95% confidence interval [CI]: 1.11 to 4.34; p ¼ 0.024) and LV thrombus (adjusted OR: 2.43, 95% CI: 1.09 to 5.42; p ¼ 0.030) emerged as important determinants of ICE, after adjusting for anticoagulation therapy. Conclusions: In a selected population referred to a tertiary center for Chagas disease that included patients with different severities of cardiac involvement, the prevalence of ICE was 20%. The presence of apical aneurysm and intracavitary thrombus were independently associated with ICE, after adjustment for other risk factors for stroke. The authors report no relationships that could be construed as a conflict of interest. This study was partly supported by grants from FAPEMIG
The diseases t ransmit ted by arthropods and other invertebrate hosts represent an enormous burde... more The diseases t ransmit ted by arthropods and other invertebrate hosts represent an enormous burden to human and animal health, if we consider that 3.3 billion people are at risk of acquiring malaria, and that , in 2010, malaria deaths were est imated at 655000 (WHO, 2012), that schistosomiasis and geohelminthiasis affect two billion humans, causing morbidity to 300 million (WHO, 2006), and that 350 million people are at risk of cont ract ing leishmaniasis (WHO, 2010), only to ment ion a few parasit ic diseases. Furthermore, the picture is wider, and the incidences of vector-borne viral diseases have been increasing sharply, even in places previously considered unlikely, as seen in recent outbreaks of West Nile and Chikungunya viruses, not forgetting dengue, a growing pandemic from which Portugal is not exempt . The prevent ion and cont rol of vector-borne diseases and of those t ransmit ted by intermediate invertebrate hosts are highly complex, as a consequence of the difficulty to ...
BackgroundCape Verde is an archipelago located off the West African coast, and is in a pre-elimin... more BackgroundCape Verde is an archipelago located off the West African coast, and is in a pre-elimination phase of malaria control. Since 2010, less than 20 Plasmodium falciparum malaria cases have been reported annually, except in 2017, when an outbreak in Praia before the rainy season led to 423 autochthonous cases. It is important to understand the genetic diversity of circulating P. falciparum to inform on drug resistance, potential transmission networks, and sources of infection, including parasite importation.MethodsEnrolled subjects involved malaria patients admitted to Dr. Agostinho Neto Hospital at Praia city, Santiago island, Cape Verde, between July and October 2017. Neighbours and family members of enrolled cases were assessed for the presence of anti-P. falciparum antibodies. Sanger sequencing and real time PCR was used to identify SNPs in genes associated with drug resistance (e.g. pfdhfr, pfdhps, pfmdr1, pfk13, pfcrt), and whole genome sequencing data was generated to in...
Malaria is a parasitic infection transmitted by Anopheles mosquitoes. Five species of the genus P... more Malaria is a parasitic infection transmitted by Anopheles mosquitoes. Five species of the genus Plasmodium cause disease in humans: P. vivax, P. malariae, P. ovale, P. knowlesi e P. falciparum being the last, the one that causes most mortality and morbidity. During 1950s and early 1960s, the global eradication of malaria was believed possible, but the successively implemented eradicationprogrammes failed and eradication was achieved only in Europe and during the 1970’s. As a consequence of the historical presence of Portugal in malaria endemic areas and due to the fact that malaria was also a health problem in Portugal, Portuguese scientists namely from IHMT, had a very relevant role in the study and control of malaria. IHMT kept a regular monitoring of the disease from 1950 onward, mostly in Angola, Mozambique and Sao Tome e Principe. With end of the colonial rule and with the creation of CMDT in 1992, the study of malaria in IHMT took a more researchoriented turn.
Biomolecules, 2020
Innovative antimalarial strategies are urgently needed given the alarming evolution of resistance... more Innovative antimalarial strategies are urgently needed given the alarming evolution of resistance to every single drug developed against Plasmodium parasites. The sulfated glycosaminoglycan heparin has been delivered in membrane feeding assays together with Plasmodium berghei-infected blood to Anopheles stephensi mosquitoes. The transition between ookinete and oocyst pathogen stages in the mosquito has been studied in vivo through oocyst counting in dissected insect midguts, whereas ookinete interactions with heparin have been followed ex vivo by flow cytometry. Heparin interferes with the parasite’s ookinete–oocyst transition by binding ookinetes, but it does not affect fertilization. Hypersulfated heparin is a more efficient blocker of ookinete development than native heparin, significantly reducing the number of oocysts per midgut when offered to mosquitoes at 5 µg/mL in membrane feeding assays. Direct delivery of heparin to mosquitoes might represent a new antimalarial strategy ...
Scientific Reports, 2018
Mosquito breeding depends on the supply of fresh vertebrate blood, a major bottleneck for largesc... more Mosquito breeding depends on the supply of fresh vertebrate blood, a major bottleneck for largescale production of Anopheles spp. Feeding alternatives to fresh blood are thus a priority for research, outdoor large-cage trials and control interventions. Several artificial meal compositions were tested and Anopheles oogenesis, egg laying and development into the next generation of adult mosquitoes were followed. We identified blood-substitute-diets that supported ovarian development, egg maturation and fertility as well as, low progeny larval mortality, and normal development of offspring into adult mosquitoes. The formulated diet is an effective artificial meal, free of fresh blood that mimics a vertebrate blood meal and represents an important advance for the sustainability of Anopheles mosquito rearing in captivity.
PLOS ONE, 2015
Allatostatin type A receptors (AST-ARs) are a group of G-protein coupled receptors activated by m... more Allatostatin type A receptors (AST-ARs) are a group of G-protein coupled receptors activated by members of the FGL-amide (AST-A) peptide family that inhibit food intake and development in arthropods. Despite their physiological importance the evolution of the AST-A system is poorly described and relatively few receptors have been isolated and functionally characterised in insects. The present study provides a comprehensive analysis of the origin and comparative evolution of the AST-A system. To determine how evolution and feeding modified the function of AST-AR the duplicate receptors in Anopheles mosquitoes, were characterised. Phylogeny and gene synteny suggested that invertebrate AST-A receptors and peptide genes shared a common evolutionary origin with KISS/GAL receptors and ligands. AST-ARs and KISSR emerged from a common gene ancestor after the divergence of GALRs in the bilaterian genome. In arthropods, the AST-A system evolved through lineage-specific events and the maintenance of two receptors in the flies and mosquitoes (Diptera) was the result of a gene duplication event. Speciation of Anophelesmosquitoes affected receptor gene organisation and characterisation of AST-AR duplicates (GPRALS1 and 2) revealed that in common with other insects, the mosquito receptors were activated by insect AST-A peptides and the iCa 2+-signalling pathway was stimulated. GPRALS1 and 2 were expressed mainly in mosquito midgut and ovaries and transcript abundance of both receptors was modified by feeding. A blood meal strongly up-regulated expression of both GPRALS in the midgut (p < 0.05) compared to glucose fed females. Based on the results we hypothesise that the AST-A system in insects shared a common origin with the vertebrate KISS system and may also share a common function as an integrator of metabolism and reproduction. Highlights: AST-A and KISS/GAL receptors and ligands shared common ancestry prior to the protostome-deuterostome divergence.
Proceedings of the 4th World Congress on Recent Advances in Nanotechnology, 2019
Scientific Reports
the saliva of hematophagous arthropods is enriched with a complex mixture of antihemostatic molec... more the saliva of hematophagous arthropods is enriched with a complex mixture of antihemostatic molecules, the biological functions of which are largely unknown. Anopheline antiplatelet protein (AApp) from malaria vector mosquito exhibits strong antiplatelet activity when bound directly to host collagen by its C-terminus and through its N-terminus with Ca 2+-binding activity. to investigate the biological functions of AApp in blood feeding behavior and malaria transmission, we generated transgenic Anopheles stephensi mosquito lines expressing anti-AApp antibody single-chain fragment (scFv) in their salivary glands. The AAPP-specific collagen-binding activity was completely abolished by AApp-scFv complex formation in the saliva. probing and prediuresis time, feeding success, blood meal size, and fecundity, which are all fitness characteristics, were significantly reduced in the transgenic mosquitoes. However, oocysts number in these mosquitoes were not significantly reduced following blood meal intake from Plasmodium berghei-infected mice. these results show that although AApp plays an important role in mosquito blood feeding, its neutralizing activity did not affect sporogonic development in our laboratory model, but its high fitness cost would pose a survival risk for parasiteinfected mosquitoes in nature.
Journal of Visualized Experiments
Malaria research requires large-scale breeding and production conditions for mosquitoes (Anophele... more Malaria research requires large-scale breeding and production conditions for mosquitoes (Anopheles spp.) in captivity. The sustainable and reliable production of mosquitoes is currently inhibited by the supply of fresh vertebrate blood. Alternatives to blood are required to promote efficient control strategies for malaria and other vector borne diseases that are transmitted by blood feeding insects. With this in mind, artificial liquid diets were formulated as substitutes for fresh vertebrate blood. Herein we report a blood-free artificial liquid diet that delivers feeding rates similar to blood and mimics the physiological effects of a fresh vertebrate blood meal. The diet induces ovarian and egg maturation of Anopheles mosquitoes and also produces good larval survival and development of functional adults. The formulated blood-free liquid diet is an important advance towards sustainable mosquito breeding in captivity and will reduce the maintenance costs of mosquito colonies and eliminate the need for fresh vertebrate blood. Video Link The video component of this article can be found at https://www.jove.com/video/60144/ 10), a protein source for egg maturation 6,12 , carbohydrates as a source of energy, and amino acids (aa) 13 that are fundamental for egg production and are a major limiting factor for mosquito fertility 14. An artificial blood free diet also needs to provide cholesterol 15 , which improves egg production. Here we describe an artificial blood-free diet for female Anopheles mosquitoes and demonstrate that it has a consistent and equivalent performance to a high-quality vertebrate blood meal.
Parasites & Vectors
Background: Elimination of malaria depends on mastering transmission and understanding the biolog... more Background: Elimination of malaria depends on mastering transmission and understanding the biological basis of Plasmodium infection in the vector. The first mosquito organ to interact with the parasite is the midgut and its transcriptomic characterization during infection can reveal effective antiplasmodial responses able to limit the survival of the parasite. The vector response to Plasmodium vivax is not fully characterized, and its specificities when compared with other malaria parasites can be of fundamental interest for specific control measures. Methods: Experimental infections were performed using a membrane-feeding device. Three groups were used: P. vivax-blood-fed, blood-fed on inactivated gametocytes, and unfed mosquitoes. Twenty-four hours after feeding, the mosquitoes were dissected and the midgut collected for transcriptomic analysis using RNAseq. Nine cDNA libraries were generated and sequenced on an Illumina HiSeq2500. Readings were checked for quality control and analysed using the Trinity platform for de novo transcriptome assembly. Transcript quantification was performed and the transcriptome was functionally annotated. Differential expression gene analysis was carried out. The role of the identified mechanisms was further explored using functional approaches. Results: Forty-nine genes were identified as being differentially expressed with P. vivax infection: 34 were upregulated and 15 were downregulated. Half of the P. vivax-related differentially expressed genes could be related to autophagy; therefore, the effect of the known inhibitor (wortmannin) and activator (spermidine) was tested on the infection outcome. Autophagic activation significantly reduced the intensity and prevalence of infection. This was associated with transcription alterations of the autophagy regulating genes Beclin, DRAM and Apg8. Conclusions: Our data indicate that P. vivax invasion of An. aquasalis midgut epithelium triggers an autophagic response and its activation reduces infection. This suggests a novel mechanism that mosquitoes can use to fight Plasmodium infection.
Emerging Infectious Diseases
In the Brazilian Amazon, the suspected source of infection in an outbreak of acute Chagas disease... more In the Brazilian Amazon, the suspected source of infection in an outbreak of acute Chagas disease involving 10 patients was Euterpe oleracea (açaí berry) juice. Patient blood and juice samples contained Trypanosoma cruzi TcIV, indicating oral transmission of the Chagas disease agent.
Parasites & vectors, Mar 13, 2018
Phlebotomine sand flies (Diptera: Psychodidae) are vectors of Leishmania species, the etiological... more Phlebotomine sand flies (Diptera: Psychodidae) are vectors of Leishmania species, the etiological agents of leishmaniasis, which is one of the most important emerging infectious diseases in the Americas. In the state of Amazonas in Brazil, anthropogenic activities encourage the presence of these insects around rural homes. The present study aimed to describe the composition and distribution of sand fly species diversity among the ecotopes (intradomicile, peridomicile and forest) in an area of American cutaneous leishmaniasis transmission and detect natural infection with Leishmania DNA to evaluate which vectors are inside houses and whether the presence of possible vectors represents a hazard of transmission. Phlebotomine sand flies were collected using light traps. A total of 2469 specimens representing 54 species, predominantly females (71.2%), were collected from four sites. Polymerase chain reaction analysis was performed on 670 samples to detect Leishmania DNA. Most of the samp...
Acta Médica Portuguesa
After more than 40 years of democracy and 30 years of European integration, Portugal has bridged ... more After more than 40 years of democracy and 30 years of European integration, Portugal has bridged the research gap it had previously. However, when compared to global and European research policies, Portugal still has a long way go regarding investment in research and development. Health Research in Portugal has been managed by the Fundação para a Ciência e Tecnologia and the National Health Institute Doctor Ricardo Jorge, and it has not been a political priority, emphasized by the absence of a national scientific research plan for health, resulting in a weak coordination of actors in the field. The strategic guidelines of the 2004 - 2010 National Health Plan are what comes closest to a health research policy, but these were not implemented by the institutions responsible for scientific research for the health sector. Trusting that adopting a strategy of incentives to stimulate health research is an added-value for the Portuguese health system, the authors present five strategic prop...
Acta Médica Portuguesa
The global debate on scientific research policy for human health has been led by World Health Org... more The global debate on scientific research policy for human health has been led by World Health Organisation with important contributions from other stakeholders such as Council on Health Research for Development, the World Bank and the Global Forum for Health Research. Recently it has been dominated by the thematic agendas of major global financiers. There is a growing interest worldwide in making better use of the evidence resulting from scientific research in health, in the decision-making process regarding health policies, which is fraught with difficulties, as it is the case in Europe. After more than 40 years of democracy and 30 years of European integration, Portugal has bridged the research gap it had previously. However, when compared to global and European research policies, Portugal still has a long way to go regarding investment in research and development.
Trends in parasitology, Sep 14, 2017
Training and innovation in the field of medical entomology are essential to mitigate the burden o... more Training and innovation in the field of medical entomology are essential to mitigate the burden of vector-borne diseases globally. However, there is a shortage of medical entomologists worldwide, and there are large discrepancies in capacity building in this field. In this article, we discuss the current situation, what is needed from the medical entomologist of today, and how we can bridge this gap.
Malaria journal, Jan 22, 2016
Strategies designed to advance towards malaria elimination rely on the detection and treatment of... more Strategies designed to advance towards malaria elimination rely on the detection and treatment of infections, rather than fever, and the interruption of malaria transmission between mosquitoes and humans. Mass drug administration with anti-malarials directed at eliminating parasites in blood, either to entire populations or targeting only those with malaria infections, are considered useful strategies to progress towards malaria elimination, but may be insufficient if applied on their own. These strategies assume a closer contact with populations, so incorporating a vector control intervention tool to those approaches could significantly enhance their efficacy. Ivermectin, an endectocide drug efficacious against a range of Anopheles species, could be added to other drug-based interventions. Interestingly, ivermectin could also be useful to target outdoor feeding and resting vectors, something not possible with current vector control tools, such as impregnated bed nets or indoor resi...
The Journal of Parasitology, May 1, 2003
Malaria infection in the mosquito vector can be modulated by the vertebrate host, mosquito factor... more Malaria infection in the mosquito vector can be modulated by the vertebrate host, mosquito factors, and interactions between different parasite populations. Modulation of parasite development can be assessed through the study of gene expression. The present report describes a specific, sensitive, and nonradioactive method that permits assessment of parasite load and quantification of circumsporozoite protein gene expression during the sporogonic stages of Plasmodium yoelii yoelii and P. y. nigeriensis. A decrease in parasite load was observed when comparing DNA of oocysts on day 7 postinfection with that of oocysts and sporozoites on day 19. On day 7, parasites (oocysts) showed a marked increase of circumsporozoite protein expression when compared with that (sporozoites and oocysts) on day 19. The method developed in this work can be a valuable tool to understand parasite interaction mechanisms that are involved in mosquito malaria infections.
Experimental Parasitology, Apr 1, 2009
Plasmodium cysteine proteases have been shown to be immunogenic and are being used as malaria pot... more Plasmodium cysteine proteases have been shown to be immunogenic and are being used as malaria potential serodiagnostic markers and vaccine targets. Genes encoding two Plasmodium chabaudi cysteine proteases chabaupain-1 (CP-1) and chabaupain-2 (CP-2) were identified and further expressed in Escherichia coli. Solubilisation of recombinant CP-1 and CP-2 was achieved by decreasing the temperature of induction. Anopheles gambiae tissues infected with Plasmodium were analyzed by Western blotting using the anti-CP-1 antibody showing that CP-1 is only present in the A. gambiae midguts being absent from other infected mosquito biological material. Anti-CP-1 anti-serum recognized a 30 kDa band in P. chabaudi, Plasmodium berghei and Plasmodium yoelii lysates but does not recognize the recombinant CP-2 extracts suggesting high antibody specificity.
Global Heart, 2015
Background: Patients with Chagas disease are at increased risk for stroke that may result in majo... more Background: Patients with Chagas disease are at increased risk for stroke that may result in major clinical disability and death. Identification of risk factors involved in the genesis of thromboembolic events related to this disease may lead to improved therapeutic decision making and outcomes. Objectives: This study sought to assess the prevalence of ischemic cerebrovascular events (ICE) among patients with Chagas heart disease and to identify the risk factors associated with cardioembolism in this population. Methods: This study involved 330 patients, 193 were men (58%), with a mean age of 49 AE 12 years with Chagas disease classified in the chronic cardiac form of the disease. Comprehensive echocardiography was performed to search a substrate for cardioembolic events, especially apical aneurysm and intracavitary thrombus. Results: Most of the patients were classified as New York Heart Association classes I or II (75%) with mean left ventricular (LV) ejection fraction of 39 AE 14%. Sixty-seven patients had a previous ICE with the overall prevalence of 20%. Apical aneurysms were detected in 128 patients (39%), whereas LV mural thrombi were found in 48 patients (15%). In multivariate analysis including the potential predictors of ICE, apical aneurysm (adjusted odds ratio [OR]: 2.19, 95% confidence interval [CI]: 1.11 to 4.34; p ¼ 0.024) and LV thrombus (adjusted OR: 2.43, 95% CI: 1.09 to 5.42; p ¼ 0.030) emerged as important determinants of ICE, after adjusting for anticoagulation therapy. Conclusions: In a selected population referred to a tertiary center for Chagas disease that included patients with different severities of cardiac involvement, the prevalence of ICE was 20%. The presence of apical aneurysm and intracavitary thrombus were independently associated with ICE, after adjustment for other risk factors for stroke. The authors report no relationships that could be construed as a conflict of interest. This study was partly supported by grants from FAPEMIG
The diseases t ransmit ted by arthropods and other invertebrate hosts represent an enormous burde... more The diseases t ransmit ted by arthropods and other invertebrate hosts represent an enormous burden to human and animal health, if we consider that 3.3 billion people are at risk of acquiring malaria, and that , in 2010, malaria deaths were est imated at 655000 (WHO, 2012), that schistosomiasis and geohelminthiasis affect two billion humans, causing morbidity to 300 million (WHO, 2006), and that 350 million people are at risk of cont ract ing leishmaniasis (WHO, 2010), only to ment ion a few parasit ic diseases. Furthermore, the picture is wider, and the incidences of vector-borne viral diseases have been increasing sharply, even in places previously considered unlikely, as seen in recent outbreaks of West Nile and Chikungunya viruses, not forgetting dengue, a growing pandemic from which Portugal is not exempt . The prevent ion and cont rol of vector-borne diseases and of those t ransmit ted by intermediate invertebrate hosts are highly complex, as a consequence of the difficulty to ...
BackgroundCape Verde is an archipelago located off the West African coast, and is in a pre-elimin... more BackgroundCape Verde is an archipelago located off the West African coast, and is in a pre-elimination phase of malaria control. Since 2010, less than 20 Plasmodium falciparum malaria cases have been reported annually, except in 2017, when an outbreak in Praia before the rainy season led to 423 autochthonous cases. It is important to understand the genetic diversity of circulating P. falciparum to inform on drug resistance, potential transmission networks, and sources of infection, including parasite importation.MethodsEnrolled subjects involved malaria patients admitted to Dr. Agostinho Neto Hospital at Praia city, Santiago island, Cape Verde, between July and October 2017. Neighbours and family members of enrolled cases were assessed for the presence of anti-P. falciparum antibodies. Sanger sequencing and real time PCR was used to identify SNPs in genes associated with drug resistance (e.g. pfdhfr, pfdhps, pfmdr1, pfk13, pfcrt), and whole genome sequencing data was generated to in...
Malaria is a parasitic infection transmitted by Anopheles mosquitoes. Five species of the genus P... more Malaria is a parasitic infection transmitted by Anopheles mosquitoes. Five species of the genus Plasmodium cause disease in humans: P. vivax, P. malariae, P. ovale, P. knowlesi e P. falciparum being the last, the one that causes most mortality and morbidity. During 1950s and early 1960s, the global eradication of malaria was believed possible, but the successively implemented eradicationprogrammes failed and eradication was achieved only in Europe and during the 1970’s. As a consequence of the historical presence of Portugal in malaria endemic areas and due to the fact that malaria was also a health problem in Portugal, Portuguese scientists namely from IHMT, had a very relevant role in the study and control of malaria. IHMT kept a regular monitoring of the disease from 1950 onward, mostly in Angola, Mozambique and Sao Tome e Principe. With end of the colonial rule and with the creation of CMDT in 1992, the study of malaria in IHMT took a more researchoriented turn.
Biomolecules, 2020
Innovative antimalarial strategies are urgently needed given the alarming evolution of resistance... more Innovative antimalarial strategies are urgently needed given the alarming evolution of resistance to every single drug developed against Plasmodium parasites. The sulfated glycosaminoglycan heparin has been delivered in membrane feeding assays together with Plasmodium berghei-infected blood to Anopheles stephensi mosquitoes. The transition between ookinete and oocyst pathogen stages in the mosquito has been studied in vivo through oocyst counting in dissected insect midguts, whereas ookinete interactions with heparin have been followed ex vivo by flow cytometry. Heparin interferes with the parasite’s ookinete–oocyst transition by binding ookinetes, but it does not affect fertilization. Hypersulfated heparin is a more efficient blocker of ookinete development than native heparin, significantly reducing the number of oocysts per midgut when offered to mosquitoes at 5 µg/mL in membrane feeding assays. Direct delivery of heparin to mosquitoes might represent a new antimalarial strategy ...
Scientific Reports, 2018
Mosquito breeding depends on the supply of fresh vertebrate blood, a major bottleneck for largesc... more Mosquito breeding depends on the supply of fresh vertebrate blood, a major bottleneck for largescale production of Anopheles spp. Feeding alternatives to fresh blood are thus a priority for research, outdoor large-cage trials and control interventions. Several artificial meal compositions were tested and Anopheles oogenesis, egg laying and development into the next generation of adult mosquitoes were followed. We identified blood-substitute-diets that supported ovarian development, egg maturation and fertility as well as, low progeny larval mortality, and normal development of offspring into adult mosquitoes. The formulated diet is an effective artificial meal, free of fresh blood that mimics a vertebrate blood meal and represents an important advance for the sustainability of Anopheles mosquito rearing in captivity.
PLOS ONE, 2015
Allatostatin type A receptors (AST-ARs) are a group of G-protein coupled receptors activated by m... more Allatostatin type A receptors (AST-ARs) are a group of G-protein coupled receptors activated by members of the FGL-amide (AST-A) peptide family that inhibit food intake and development in arthropods. Despite their physiological importance the evolution of the AST-A system is poorly described and relatively few receptors have been isolated and functionally characterised in insects. The present study provides a comprehensive analysis of the origin and comparative evolution of the AST-A system. To determine how evolution and feeding modified the function of AST-AR the duplicate receptors in Anopheles mosquitoes, were characterised. Phylogeny and gene synteny suggested that invertebrate AST-A receptors and peptide genes shared a common evolutionary origin with KISS/GAL receptors and ligands. AST-ARs and KISSR emerged from a common gene ancestor after the divergence of GALRs in the bilaterian genome. In arthropods, the AST-A system evolved through lineage-specific events and the maintenance of two receptors in the flies and mosquitoes (Diptera) was the result of a gene duplication event. Speciation of Anophelesmosquitoes affected receptor gene organisation and characterisation of AST-AR duplicates (GPRALS1 and 2) revealed that in common with other insects, the mosquito receptors were activated by insect AST-A peptides and the iCa 2+-signalling pathway was stimulated. GPRALS1 and 2 were expressed mainly in mosquito midgut and ovaries and transcript abundance of both receptors was modified by feeding. A blood meal strongly up-regulated expression of both GPRALS in the midgut (p < 0.05) compared to glucose fed females. Based on the results we hypothesise that the AST-A system in insects shared a common origin with the vertebrate KISS system and may also share a common function as an integrator of metabolism and reproduction. Highlights: AST-A and KISS/GAL receptors and ligands shared common ancestry prior to the protostome-deuterostome divergence.
Proceedings of the 4th World Congress on Recent Advances in Nanotechnology, 2019
Scientific Reports
the saliva of hematophagous arthropods is enriched with a complex mixture of antihemostatic molec... more the saliva of hematophagous arthropods is enriched with a complex mixture of antihemostatic molecules, the biological functions of which are largely unknown. Anopheline antiplatelet protein (AApp) from malaria vector mosquito exhibits strong antiplatelet activity when bound directly to host collagen by its C-terminus and through its N-terminus with Ca 2+-binding activity. to investigate the biological functions of AApp in blood feeding behavior and malaria transmission, we generated transgenic Anopheles stephensi mosquito lines expressing anti-AApp antibody single-chain fragment (scFv) in their salivary glands. The AAPP-specific collagen-binding activity was completely abolished by AApp-scFv complex formation in the saliva. probing and prediuresis time, feeding success, blood meal size, and fecundity, which are all fitness characteristics, were significantly reduced in the transgenic mosquitoes. However, oocysts number in these mosquitoes were not significantly reduced following blood meal intake from Plasmodium berghei-infected mice. these results show that although AApp plays an important role in mosquito blood feeding, its neutralizing activity did not affect sporogonic development in our laboratory model, but its high fitness cost would pose a survival risk for parasiteinfected mosquitoes in nature.
Journal of Visualized Experiments
Malaria research requires large-scale breeding and production conditions for mosquitoes (Anophele... more Malaria research requires large-scale breeding and production conditions for mosquitoes (Anopheles spp.) in captivity. The sustainable and reliable production of mosquitoes is currently inhibited by the supply of fresh vertebrate blood. Alternatives to blood are required to promote efficient control strategies for malaria and other vector borne diseases that are transmitted by blood feeding insects. With this in mind, artificial liquid diets were formulated as substitutes for fresh vertebrate blood. Herein we report a blood-free artificial liquid diet that delivers feeding rates similar to blood and mimics the physiological effects of a fresh vertebrate blood meal. The diet induces ovarian and egg maturation of Anopheles mosquitoes and also produces good larval survival and development of functional adults. The formulated blood-free liquid diet is an important advance towards sustainable mosquito breeding in captivity and will reduce the maintenance costs of mosquito colonies and eliminate the need for fresh vertebrate blood. Video Link The video component of this article can be found at https://www.jove.com/video/60144/ 10), a protein source for egg maturation 6,12 , carbohydrates as a source of energy, and amino acids (aa) 13 that are fundamental for egg production and are a major limiting factor for mosquito fertility 14. An artificial blood free diet also needs to provide cholesterol 15 , which improves egg production. Here we describe an artificial blood-free diet for female Anopheles mosquitoes and demonstrate that it has a consistent and equivalent performance to a high-quality vertebrate blood meal.
Parasites & Vectors
Background: Elimination of malaria depends on mastering transmission and understanding the biolog... more Background: Elimination of malaria depends on mastering transmission and understanding the biological basis of Plasmodium infection in the vector. The first mosquito organ to interact with the parasite is the midgut and its transcriptomic characterization during infection can reveal effective antiplasmodial responses able to limit the survival of the parasite. The vector response to Plasmodium vivax is not fully characterized, and its specificities when compared with other malaria parasites can be of fundamental interest for specific control measures. Methods: Experimental infections were performed using a membrane-feeding device. Three groups were used: P. vivax-blood-fed, blood-fed on inactivated gametocytes, and unfed mosquitoes. Twenty-four hours after feeding, the mosquitoes were dissected and the midgut collected for transcriptomic analysis using RNAseq. Nine cDNA libraries were generated and sequenced on an Illumina HiSeq2500. Readings were checked for quality control and analysed using the Trinity platform for de novo transcriptome assembly. Transcript quantification was performed and the transcriptome was functionally annotated. Differential expression gene analysis was carried out. The role of the identified mechanisms was further explored using functional approaches. Results: Forty-nine genes were identified as being differentially expressed with P. vivax infection: 34 were upregulated and 15 were downregulated. Half of the P. vivax-related differentially expressed genes could be related to autophagy; therefore, the effect of the known inhibitor (wortmannin) and activator (spermidine) was tested on the infection outcome. Autophagic activation significantly reduced the intensity and prevalence of infection. This was associated with transcription alterations of the autophagy regulating genes Beclin, DRAM and Apg8. Conclusions: Our data indicate that P. vivax invasion of An. aquasalis midgut epithelium triggers an autophagic response and its activation reduces infection. This suggests a novel mechanism that mosquitoes can use to fight Plasmodium infection.
Emerging Infectious Diseases
In the Brazilian Amazon, the suspected source of infection in an outbreak of acute Chagas disease... more In the Brazilian Amazon, the suspected source of infection in an outbreak of acute Chagas disease involving 10 patients was Euterpe oleracea (açaí berry) juice. Patient blood and juice samples contained Trypanosoma cruzi TcIV, indicating oral transmission of the Chagas disease agent.
Parasites & vectors, Mar 13, 2018
Phlebotomine sand flies (Diptera: Psychodidae) are vectors of Leishmania species, the etiological... more Phlebotomine sand flies (Diptera: Psychodidae) are vectors of Leishmania species, the etiological agents of leishmaniasis, which is one of the most important emerging infectious diseases in the Americas. In the state of Amazonas in Brazil, anthropogenic activities encourage the presence of these insects around rural homes. The present study aimed to describe the composition and distribution of sand fly species diversity among the ecotopes (intradomicile, peridomicile and forest) in an area of American cutaneous leishmaniasis transmission and detect natural infection with Leishmania DNA to evaluate which vectors are inside houses and whether the presence of possible vectors represents a hazard of transmission. Phlebotomine sand flies were collected using light traps. A total of 2469 specimens representing 54 species, predominantly females (71.2%), were collected from four sites. Polymerase chain reaction analysis was performed on 670 samples to detect Leishmania DNA. Most of the samp...
Acta Médica Portuguesa
After more than 40 years of democracy and 30 years of European integration, Portugal has bridged ... more After more than 40 years of democracy and 30 years of European integration, Portugal has bridged the research gap it had previously. However, when compared to global and European research policies, Portugal still has a long way go regarding investment in research and development. Health Research in Portugal has been managed by the Fundação para a Ciência e Tecnologia and the National Health Institute Doctor Ricardo Jorge, and it has not been a political priority, emphasized by the absence of a national scientific research plan for health, resulting in a weak coordination of actors in the field. The strategic guidelines of the 2004 - 2010 National Health Plan are what comes closest to a health research policy, but these were not implemented by the institutions responsible for scientific research for the health sector. Trusting that adopting a strategy of incentives to stimulate health research is an added-value for the Portuguese health system, the authors present five strategic prop...
Acta Médica Portuguesa
The global debate on scientific research policy for human health has been led by World Health Org... more The global debate on scientific research policy for human health has been led by World Health Organisation with important contributions from other stakeholders such as Council on Health Research for Development, the World Bank and the Global Forum for Health Research. Recently it has been dominated by the thematic agendas of major global financiers. There is a growing interest worldwide in making better use of the evidence resulting from scientific research in health, in the decision-making process regarding health policies, which is fraught with difficulties, as it is the case in Europe. After more than 40 years of democracy and 30 years of European integration, Portugal has bridged the research gap it had previously. However, when compared to global and European research policies, Portugal still has a long way to go regarding investment in research and development.
Trends in parasitology, Sep 14, 2017
Training and innovation in the field of medical entomology are essential to mitigate the burden o... more Training and innovation in the field of medical entomology are essential to mitigate the burden of vector-borne diseases globally. However, there is a shortage of medical entomologists worldwide, and there are large discrepancies in capacity building in this field. In this article, we discuss the current situation, what is needed from the medical entomologist of today, and how we can bridge this gap.
Malaria journal, Jan 22, 2016
Strategies designed to advance towards malaria elimination rely on the detection and treatment of... more Strategies designed to advance towards malaria elimination rely on the detection and treatment of infections, rather than fever, and the interruption of malaria transmission between mosquitoes and humans. Mass drug administration with anti-malarials directed at eliminating parasites in blood, either to entire populations or targeting only those with malaria infections, are considered useful strategies to progress towards malaria elimination, but may be insufficient if applied on their own. These strategies assume a closer contact with populations, so incorporating a vector control intervention tool to those approaches could significantly enhance their efficacy. Ivermectin, an endectocide drug efficacious against a range of Anopheles species, could be added to other drug-based interventions. Interestingly, ivermectin could also be useful to target outdoor feeding and resting vectors, something not possible with current vector control tools, such as impregnated bed nets or indoor resi...
The Journal of Parasitology, May 1, 2003
Malaria infection in the mosquito vector can be modulated by the vertebrate host, mosquito factor... more Malaria infection in the mosquito vector can be modulated by the vertebrate host, mosquito factors, and interactions between different parasite populations. Modulation of parasite development can be assessed through the study of gene expression. The present report describes a specific, sensitive, and nonradioactive method that permits assessment of parasite load and quantification of circumsporozoite protein gene expression during the sporogonic stages of Plasmodium yoelii yoelii and P. y. nigeriensis. A decrease in parasite load was observed when comparing DNA of oocysts on day 7 postinfection with that of oocysts and sporozoites on day 19. On day 7, parasites (oocysts) showed a marked increase of circumsporozoite protein expression when compared with that (sporozoites and oocysts) on day 19. The method developed in this work can be a valuable tool to understand parasite interaction mechanisms that are involved in mosquito malaria infections.
Experimental Parasitology, Apr 1, 2009
Plasmodium cysteine proteases have been shown to be immunogenic and are being used as malaria pot... more Plasmodium cysteine proteases have been shown to be immunogenic and are being used as malaria potential serodiagnostic markers and vaccine targets. Genes encoding two Plasmodium chabaudi cysteine proteases chabaupain-1 (CP-1) and chabaupain-2 (CP-2) were identified and further expressed in Escherichia coli. Solubilisation of recombinant CP-1 and CP-2 was achieved by decreasing the temperature of induction. Anopheles gambiae tissues infected with Plasmodium were analyzed by Western blotting using the anti-CP-1 antibody showing that CP-1 is only present in the A. gambiae midguts being absent from other infected mosquito biological material. Anti-CP-1 anti-serum recognized a 30 kDa band in P. chabaudi, Plasmodium berghei and Plasmodium yoelii lysates but does not recognize the recombinant CP-2 extracts suggesting high antibody specificity.