COVID-19 seroepidemiological survey among healthcare workers in the City of Ribeirão Preto, São Paulo, Brazil

ABSTRACT Background: Coronavirus disease (COVID-19) serology testing evaluates the prevalence of COVID-19 cases. Methods: A seroepidemiological survey of COVID-19 among healthcare workers was performed (June 2020 to November 2020) in Ribeirão Preto, São Paulo, Brazil. Overall, 10,172 and 2,129 workers participated in the first and second phases, respectively. Results: First phase: 12.7% tested positive for COVID-19 (73.5% females and 35.2% aged 30-39 years), and 29.6% were nursing technicians. Second phase: 12.1% tested positive for COVID-19 (65.5% females and 33.3% aged 40-49 years), and 24.8% were nursing assistants. Conclusions: In 2020, healthcare workers in Ribeirão Preto had COVID-19 in a similar way.

Local and systemic effects caused by Crotalus durissus terrificus, Crotalus durissus collilineatus, and Crotalus durissus cascavella snake venoms in swiss mice

Abstract INTRODUCTION: Crotalus envenomations cause serious complications and can be fatal without appropriate treatment. Venom isoforms present and inter/intraspecific variations in the venom composition can result in different symptoms presented by bites by snakes from the same species but from different geographical regions. We comparatively evaluated the local and systemic effects caused by Crotalus durissus terrificus (Cdt), C.d. collilineatus (Cdcolli), and C.d. cascavella (Cdcasc) envenomation. METHODS: Venom chromatography was performed. Proteolytic, phospholipase, and LAAO activities were analyzed. Edema, myotoxicity, hepatotoxicity, nephrotoxicity, and coagulation alterations were evaluated. RESULTS: The venom SDS-PAGE analyses found the presence of convulxin, gyroxin, crotoxin, and crotamine in Cdt and Cdcolli venoms. Crotamine was not present in the Cdcasc venom. Cdt, Cdcollli, and Cdcasc venoms had no proteolytic activity. Only Cdcasc and Cdt venoms had phospholipase activity. LAAO activity was observed in Cdcolli and Cdcasc venoms. Cdcolli and Cdcasc venoms caused 36.7% and 13.3% edema increases, respectively. Cdt venom caused a 10% edema induction compared to those by other venoms. All venoms increased TOTAL-CK, MB-CK, and LDH levels (indicating muscle injury) and ALT, AST, GGT, and ALP levels (markers of liver damage) and were able to induce a neuromuscular blockade. Urea and creatinine levels were also altered in both plasma and urine, indicating kidney damage. Only Cdcolli and Cdcasc venoms increased TAPP and TAP. CONCLUSIONS: Together, these results allow us to draw a distinction between local and systemic effects caused by Crotalus subspecies, highlighting the clinical and biochemical effects produced by their respective venoms.

Epidemiological study of snakebite cases in Brazilian Western Amazonia

Abstract INTRODUCTION Brazil has the largest number of snakebite cases in South America, of which the large majority is concentrated in the Midwest and North. METHODS In this descriptive observational study, we assessed the epidemiological and clinical snakebite cases referred to the Centro de Medicina Tropical de Rondônia from September 2008 to September 2010. RESULTS We followed up 92 cases from admission until discharge, namely 81 (88%) men and 11 (12%) women, with a mean age of 37 years, and mainly from rural areas (91.3%). The snakebites occurred while performing work activities (63%) during the Amazon rainy season (78.3%). The vast majority of individuals presented from the Porto Velho microregion (84.7%). Approximately 95.6% of the snakebites were caused by snakes of the genus Bothrops, followed by two lachetics and two elapidics cases. Surgery was performed in 10 cases (9 fasciotomies in the lower limb and 1 amputation). No deaths were reported in this study, but 4 cases (4.3%) developed sequelae in the lower limb. CONCLUSIONS This study can contribute to a better understanding of envenomation in the state of Rondônia and thus can be useful for identifying real conditions that can increase the incidence of snakebites in this region. Moreover, the study results can serve as a basis for improving educational campaigns designed to prevent these types of snakebites, as well as for preserving snakes.

Biochemical characterization of a phospholipase A2 homologue from the venom of the social wasp Polybia occidentalis

Background: Wasp venoms constitute a molecular reservoir of new pharmacological substances such as peptides and proteins, biological property holders, many of which are yet to be identified. Exploring these sources may lead to the discovery of molecules hitherto unknown. This study describes, for the first time in hymenopteran venoms, the identification of an enzymatically inactive phospholipase A2 (PLA2) from the venom of the social wasp Polybia occidentalis. Methods: P. occidentalis venom was fractioned by molecular exclusion and reverse phase chromatography. For the biochemical characterization of the protein, 1D and 2D SDS-PAGE were performed, along with phospholipase activity assays on synthetic substrates, MALDI-TOF mass spectrometry and sequencing by Edman degradation. Results: The protein, called PocTX, was isolated using two chromatographic steps. Based on the phospholipase activity assay, electrophoresis and mass spectrometry, the protein presented a high degree of purity, with a mass of 13,896. 47 Da and a basic pI. After sequencing by the Edman degradation method, it was found that the protein showed a high identity with snake venom PLA2 homologues. Conclusion: This is the first report of an enzymatically inactive PLA2 isolated from wasp venom, similar to snake PLA2 homologues.

Biochemical characterization of a phospholipase A2 homologue from the venom of the social wasp Polybia occidentalis

Wasp venoms constitute a molecular reservoir of new pharmacological substances such as peptides and proteins, biological property holders, many of which are yet to be identified. Exploring these sources may lead to the discovery of molecules hitherto unknown. This study describes, for the first time in hymenopteran venoms, the identification of an enzymatically inactive phospholipase A2 (PLA2) from the venom of the social wasp Polybia occidentalis. Methods: P. occidentalis venom was fractioned by molecular exclusion and reverse phase chromatography. For the biochemical characterization of the protein, 1D and 2D SDS-PAGE were performed, along with phospholipase activity assays on synthetic substrates, MALDI-TOF mass spectrometry and sequencing by Edman degradation. Results: The protein, called PocTX, was isolated using two chromatographic steps. Based on the phospholipase activity assay, electrophoresis and mass spectrometry, the protein presented a high degree of purity, with a mass of 13,896. 47 Da and a basic pI. After sequencing by the Edman degradation method, it was found that the protein showed a high identity with snake venom PLA2 homologues. Conclusion: This is the first report of an enzymatically inactive PLA2 isolated from wasp venom, similar to snake PLA2 homologues.(AU)

Immunoproteomics of Plasmodium falciparum-infected red blood cell membrane fractions

BACKGROUND The surface of infected red blood cells (iRBCs) has been widely investigated because of the molecular complexity and pathogenesis mechanisms involved. Asymptomatic individuals are important in the field because they can perpetuate transmission as natural reservoirs and present a challenge for diagnosing malaria because of their low levels of circulating parasites. Recent studies of iRBC antibody recognition have shown that responses are quantitatively similar in symptomatic and asymptomatic infections, but no studies have characterised the plasmodial proteins targeted by this response. OBJECTIVES Our main objective was to identify Plasmodium falciparum proteins associated with iRBC ghosts recognised by antibodies in the sera of symptomatic and asymptomatic individuals in the Brazilian Amazon. METHODS We collected symptomatic and asymptomatic sera from patients residing in the Brazilian Amazon and P. falciparum iRBC ghosts to identify the proteins involved in natural antibody recognition by 2D-electrophoresis, western blotting, and high- resolution mass spectrometry. FINDINGS 2D gel-based immunoproteome analysis using symptomatic and asymptomatic sera identified 11 proteins with at least one unique peptide, such as chaperones HSP70-1 and HSP70-x, which likely are components of the secretion machinery/PTEX translocon. PfEMP1 is involved in antigenic variation in symptomatic infections and we found putative membrane proteins whose functions are unknown. MAIN FINDINGS Our results suggest a potential role of old and new proteins, such as antigenic variation proteins, iRBC remodelling, and membrane proteins, with no assigned functions related to the immune response against P. falciparum, providing insights into the pathogenesis, erythrocyte remodelling, and secretion machinery important for alternative diagnosis and/or malaria therapy.

Engenharia de anticorpos na busca por fragmentos funcionais para insumos de diagnóstico e aplicações terapêuticas / Antibody engineering in the search for functional fragments for diagnostic inputs and therapeutic applications

Anticorpos, agentes empregados no desenvolvimento de pesquisas biomédicas, no diagnóstico e na terapêutica, possuem elevada capacidade de interação aos mais variados ligantes, Estruturalmente são heterotetrameros constituídos por duas cadeias leves e duas cadeias pesadas com massa molecular de aproximadamente 150 kDa, Visando melhorar as características farmacocinéticas e minimizar possíveis reações adversas desencadeadas por imunoglobulinas de origem não humana, a engenharia molecular de anticorpos vem obtendo fragmentos de anticorpos como porções Fab, F(ab?)2, scFv e Fv, Em adição aos anticorpos convencionais, camelídeos produzem imunoglobulinas funcionais desprovidas de cadeia leve, onde o domínio variável da cadeia pesada, denominado VHH ou nanocorpo, é responsável pelo reconhecimento antigênico, Apresentando características adequadas ao desenvolvimento de fármacos com alta capacidade de neutralização, fragmentos VHHs vêm sendo propostos para uso em imunoterapia passiva ou em drug-delivery, No diagnóstico esses fragmentos podem ser aplicados na construção de biosensores ou na imagiologia, atuando na detecção de células cancerígenas, no monitoramento de tumores ou em alterações celulares...

Antibodies, agents employed for the development of biomedical research, diagnostic and therapeutic, have high ability to interact with different ligands. Structurally are heterotetramers constituted by two light and two heavy chains, with molecular weight of approximately 150 kDa. Aiming to improve the pharmacokinetic properties and minimize possible adverse reactions triggered by immunoglobulins of non-human origin, the molecular engineering of antibodies has been obtaining fragments of antibodies, such as Fab, F(ab?)2, Fv and scFv. In addition to the conventional antibodies, camelids produce functional immunoglobulins devoid of light chain, in which the variable domain, named VHH or nanocorpo, is able to recognize the antigen. With appropriate characteristics for the development of drugs with high neutralizing capacity, VHH fragments have been proposed for use in passive immunotherapy or drug-delivery. To the diagnosis, these fragments can be used to construct biosensors, in the imagiology , acting in the detection of cancer cells, tumor monitoring or cell changes...

Insecticidal activity of Leptodactylus knudseni andPhyllomedusa vaillantii crude skin secretions against the mosquitoes Anopheles darlingi andAedes aegypti

Background Mosquitoes are important vectors of several diseases, including malaria and dengue, and control measures are mostly performed using chemical insecticides. Unfortunately, mosquito resistance to commonly applied insecticides is widespread. Therefore, a prospection for new molecules with insecticidal activity based on Amazon biodiversity using the anuransLeptodactylus knudseni andPhyllomedusa vaillantii was performed against the mosquito speciesAnopheles darlingi and Aedes aegypti.Methods The granular secretion from anuran skin was obtained by manual stimulation, and lethal concentrations (LCs) for larvicidal and adulticidal tests were calculated using concentrations from 1-100 ppm. The skin secretions from the anuran species tested caused significant mortality within the first 24 hours on adults and larvae, but differed within the mosquito species.Results The skin secretions from the anuran species tested caused significant mortality within the first 24 hours on adults and larvae, but differed within the mosquito species. The calculated LC50 of L. knudseni skin secretions against An. darlingiwas 0.15 and 0.2 ppm for adults and larvae, respectively, but much higher for Ae. aegypti, i.e., 19 and 38 ppm, respectively. Interestingly, the calculated LCs50 of P. vaillantii against both mosquito species in adults were similar, 1.8 and 2.1 ppm, respectively, but the LC50 forAn. darlingi larvae was much lower (0.4 ppm) than forAe aegypti (2.1 ppm).Conclusions The present experiments indicate that skin secretions from L. knudseni and P. vaillantii contain bioactive molecules with potent insecticide activity. The isolation and characterization of skin secretions components will provide new insights for potential insecticidal molecules.(AU)

Insecticidal activity of Leptodactylus knudseni and Phyllomedusa vaillantii crude skin secretions against the mosquitoes Anopheles darlingi and Aedes aegypti

Mosquitoes are important vectors of several diseases, including malaria and dengue, and control measures are mostly performed using chemical insecticides. Unfortunately, mosquito resistance to commonly applied insecticides is widespread. Therefore, a prospection for new molecules with insecticidal activity based on Amazon biodiversity using the anurans Leptodactylus knudseni and Phyllomedusa vaillantii was performed against the mosquito species Anopheles darlingi and Aedes aegypti. The granular secretion from anuran skin was obtained by manual stimulation, and lethal concentrations(LCs) for larvicidal and adulticidal tests were calculated using concentrations from 1-100 ppm. The skin secretions from the anuran species tested caused significant mortality within the first 24 hours on adults and larvae, but differed within the mosquito species. The skin secretions from the anuran species tested caused significant mortality within the first 24 hours on adults and larvae, but differed within the mosquito species. The calculated LC50 of L. knudseni skin secretions against An. darlingi was 0.15 and 0.2 ppm for adults and larvae, respectively, but much higher for Ae. aegypti, i.e., 19 and 38 ppm, respectively. Interestingly, the calculated LCs50 of P. vaillantii against both mosquito species in adults were similar, 1.8 and 2.1 ppm, respectively, but the LC50 for An. darlingi larvae was much lower (0.4 ppm) than for Ae aegypti (2.1 ppm). The present experiments indicate that skin secretions from L. knudseni and P. vaillantii contain bioactive molecules with potent insecticide activity. The isolation and characterization of skin secretions components will provide new insights for potential insecticidal molecules.

Copaifera multijuga ethanolic extracts, oil-resin, and its derivatives display larvicidal activity against Anopheles darlingi and Aedes aegypti (Diptera: Culicidae)

Copaifera spp. is a common tree species found in the tropical region of Latin America, popularly known as copaiba or pau-d'alho. Oil-resin from different Copaifera species and its components present several biological activities such as antimicrobial, anti-inflammatory, antioxidant and insecticidal, including larvicidal activity against mosquitoes. Thus, bark and leaf ethanolic extracts, oil-resin, essential oil and alepterolic acid from Copaifera multijuga Hayne, Fabaceae, were tested as larvicides against the main malaria vector in the north of Brazil, Anopheles darlingi and also Aedes aegypti, the dengue vector. A. darlingi larval mortality was significantly higher than A. aegypti for most tested compounds. Bark and leaf extracts resulted in lower Lethal Concentrations (LC50) values for A. darlingi, 3 and 13 ppm, respectively, while the essential oil provided the lowest LC50 value for A. aegypti, 18 ppm. Despite of that, the lowest LC values were from the alepterolic acid for both species, i.e. 0.9 and 0.7 ppm for A. darlingi and A. aegypti, respectively.

Evaluation of larvicidal activity of the methanolic extracts of Piper alatabaccum branches and P. tuberculatum leaves and compounds isolated against Anopheles darlingi

Piper is a notable genus among Piperaceae due to their secondary metabolites such as lignans, amides, esters and long chain fatty acids used as anti-herbivore defenses with comparable effects of pyrethroids, that holds a promise in insect control, including malaria vectors such as Anopheles darlingi, the main vector in the North of Brazil. Methanolic extracts of Piper tuberculatum Jacq., Piperaceae, and P. alatabaccum Trel. & Yunck., Piperaceae, and some isolated compounds, i.e, 3,4,5-trimetoxy-dihydrocinamic acid, dihydropiplartine; piplartine, piplartine-dihydropiplartine and 5,5',7-trimetoxy-3',4'-metilenodioxiflavone were tested as larvicides against A. darlingi. The Lethal Concentrations (LC50 and LC90) of methanolic extracts were 194 and 333 ppm for P. tuberculatum and 235 and 401 ppm for P. alatabacum, respectively. Isolated compounds had lower LC values, e.g. the LC50 and LC90 of the piplartine-dihidropiplartine isolated from both plant species was 40 and 79 ppm, respectively.

The leishmanicidal activity of a cyclopentenedione derivative isolated from the roots of a native Amazonian pepper (Piper carniconnectivum)

The Piper species chemistry has been widely investigated and the phytochemical analyses have led to the isolation of a number of active compounds like alkaloids, terpenes and flavones among others. The aim of this study was to evaluate the leishmanicidal activity of 2-[1-hydroxy-3-phenyl-(Z,2E)-2-propenylidene]-4-methyl-4-cyclopentene-1,3-dione (DCPC), a cyclopentenedione derivative isolated from the roots of Piper carniconnectivum C. DC., Piperaceae. Leishmanicidal activity against Leishmania amazonensis promastigotes was assessed, and the risk to host cell was assessed by measuring the cytotoxicity to peritoneal macrophages from BALB/c mice in vitro. L. amazonensis promastigotes and host macrophages were cultured in the presence of 100, 50, 25, 12.5 and 6 µg/mL of the cyclopentenedione derivative for up to 96 h. At the end of this period, the inhibitory concentrations (IC50) were compared with those from untreated cultures. The IC50 for promastigotes was 4.4 µg/mL after 96 h of treatment with the derivative. The 50% cytotoxic concentration (CC50) against murine peritoneal macrophages was 129 µg/mL. These results indicate that DCPC is a promising molecule for the development of leishmanicidal drugs.

Antileishmanial activity of 3-(3,4,5-trimethoxyphenyl) propanoic acid purified from Amazonian Piper tuberculatum Jacq., Piperaceae, fruits / Atividade antileishmania do 3,4,5-trimetoxi-dihidrocinâmico purificado das frutas de Piper tuberculatum Jacq., Piperaceae, amazônica

Leishmanicidal activity of the 3-(3,4,5-trimethoxyphenyl) propanoic acid (TMPP) isolated from EtOH extracts of the Amazonian Piper turbeculatum Jacq. fruits was evaluated in vitro using Leishmania amazonensis promastigotes. The TMPP was assayed at concentrations of 1600 to 6.25 µg/mL for 24, 48, 72 and 96 h. Promastigotes viability was analyzed and the IC50 of TMPP was 145 µg/mL.

A atividade leishmanicida do ácido 3,4,5-trimetoxi-dihidrocinâmico (TMPP) isolado do extrato hidroalcoólico de frutos de Piper turbeculatum Jacq. amazônica foi testado em ensaios in vitro utilizando formas promastigotas de Leishmania amazonensis. O TMPP foi utilizado em culturas de L. amazonensis nas concentrações de 1600 a 6,25 µg/mL. A viabilidade celular das formas promastigotas foi observada em 24, 48, 72 e 96 h para cálculo da CI50. O TMPP apresentou efeito leishmanicida dose dependente para as formas promastigotas de L. amazonensis apresentando CI50 de 145 µg/mL.