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1.
Int Immunopharmacol ; 128: 111467, 2024 Feb 15.
Article in English | MEDLINE | ID: mdl-38211479

ABSTRACT

The adequate management of parasite co-infections represents a challenge that has not yet been overcome, especially considering that the pathological outcomes and responses to treatment are poorly understood. Thus, this study aimed to evaluate the impact of Schistosoma mansoni infection on the efficacy of benznidazole (BZN)-based chemotherapy in Trypanosoma cruzi co-infected mice. BALB/c mice were maintained uninfected or co-infected with S. mansoni and T. cruzi, and were untreated or treated with BZN. Body weight, mortality, parasitemia, cardiac parasitism, circulating cytokines (Th1/Th2/Th17); as well as heart, liver and intestine microstructure were analyzed. The parasitemia peak was five times higher and myocarditis was more severe in co-infected than T. cruzi-infected mice. After reaching peak, parasitemia was effectively controlled in co-infected animals. BZN successfully controlled parasitemia in both co-infected and T. cruzi-infected mice and improved body mass, cardiac parasitism, myocarditis and survival in co-infected mice. Co-infection dampened the typical cytokine response to either parasite, and BZN reduced anti-inflammatory cytokines in co-infected mice. Despite BZN normalizing splenomegaly and liver cellular infiltration, it exacerbated hepatomegaly in co-infected mice. Co-infection or BZN exerted no effect on hepatic granulomas, but increased pulmonary and intestinal granulomas. Marked granulomatous inflammation was identified in the small intestine of all schistosomiasis groups. Taken together, our findings indicate that BZN retains its therapeutic efficacy against T. cruzi infection even in the presence of S. mansoni co-infection, but with organ-specific repercussions, especially in the liver.


Subject(s)
Chagas Disease , Coinfection , Myocarditis , Nitroimidazoles , Schistosomiasis mansoni , Mice , Animals , Myocarditis/parasitology , Schistosoma mansoni , Parasitemia/drug therapy , Chagas Disease/drug therapy , Cytokines/therapeutic use , Granuloma
2.
J Virol Methods ; 317: 114742, 2023 07.
Article in English | MEDLINE | ID: mdl-37116586

ABSTRACT

The pandemic caused by COVID-19 and the emergence of new variants of SARS-CoV-2 have generated clinical and epidemiological impacts on a global scale. The use of strategies for monitoring viral circulation and identifying mutations in genomic regions involved in host interaction are important measures to mitigate viral dissemination and reduce its likely complications on population health. In this context, the objective of this work was to explore the potential of high-resolution melting (HRM) analysis combined with one-step real-time reverse transcription PCR in a closed-tube system, as a fast and convenient method of screening for SARS-CoV-2 mutations with possible implications on host-pathogen interactions. The HRM analyses allowed the distinction of the Gamma, Zeta, Alpha, Delta, and Omicron variants against the predecessors (B.1.1.28, B.1.1.33) of occurrence in Brazil. It is concluded that the molecular tool standardized here has the potential to optimize the genomic surveillance of SARS-CoV-2, and could be adapted for genomic surveillance of other pathogens, due to its ability to detect, prior to sequencing, samples suggestive of new variants, selecting them more assertively and earlier for whole genome sequencing when compared to random screening.


Subject(s)
COVID-19 , SARS-CoV-2 , Humans , SARS-CoV-2/genetics , COVID-19/diagnosis , Genomics , Real-Time Polymerase Chain Reaction , Mutation
3.
Acta Trop ; 234: 106581, 2022 Oct.
Article in English | MEDLINE | ID: mdl-35779591

ABSTRACT

In recent decades, the oral infection of Trypanosoma cruzi has gathered increased attention due to frequent outbreaks that can lead to more severe clinical signs than those usually found in the areas of vector transmission. This study addresses the main routes of infection using metacyclic trypomastigotes (MT) and blood trypomastigotes (BT). Herein, BALB/c mice were infected with the Colombian (TcI) strain via intraperitoneal (IP), oral, intragastric (IG), ocular (OC) and cutaneous (CT) routes with 106 culture-derived MT or BT. Parasitemia was intermittent and low in animals inoculated with MT, in contrast, high parasitemia levels were found in BT-mice. A tropism for the muscles was observed in oral or IG infection with BT. Differently, the parasite was widely distributed in the tissues of mice infected with MT. However, the intensity of the inflammation infiltrating the tissues was higher in oral or IG infection with BT. Animals inoculated with BT via the IG route had similar serum levels of IFN-γ and smaller IL-10 compared to those infected with MT via the IG route. TNF-α levels were higher in the serum from BT-animals, which could explain the higher intensity of heart inflammation in these animals. Our results suggest that the infective form and the route of infection differentially modulated the outcome of Trypanosoma cruzi mice infection.


Subject(s)
Chagas Disease , Communicable Diseases , Trypanosoma cruzi , Animals , Mice , Mice, Inbred BALB C , Parasitemia/parasitology
4.
Mol Immunol ; 147: 199-208, 2022 07.
Article in English | MEDLINE | ID: mdl-35644072

ABSTRACT

Viral inactivation for antibody induction purposes, among other applications, should ensure biosafety, completely avoiding the risk of infectivity, and preserving viral immunogenicity. ß-propiolactone (BPL) is one of the most used reagents for viral inactivation, despite its high toxicity and recent difficulties related to importation, experienced in Brazil during the SARS-CoV-2 pandemic. In this context, the main objectives of this work were to test different inactivation procedures for SARS-CoV-2 and to evaluate the induction of neutralizing antibodies in mice immunized with antigenic preparations obtained after viral treatment with formaldehyde (FDE), glutaraldehyde (GDE), peroxide hydrogen (H2O2), as well as with viral proteins extract (VPE), in parallel with BPL. Verification of viral inactivation was performed by subsequent incubations of the inactivated virus in Vero cells, followed by cytopathic effect and lysis plaques observation, as well as by quantification of RNA load using reverse transcription-quantitative real time polymerase chain reaction. Once viral inactivation was confirmed, cell culture supernatants were concentrated and purified. In addition, an aliquot inactivated by BPL was also subjected to viral protein extraction (VPE). The different antigens were prepared using a previously developed microemulsion as adjuvant, and were administered in a four-dose immunization protocol. Antibody production was comparatively evaluated by ELISA and Plaque Reduction Neutralization Tests (PRNT). All immunogens evaluated showed some level of IgG anti-SARS-CoV-2 antibodies in the ELISA assay, with the highest levels presented by the group immunized with FDE-inactivated viral antigen. In the PRNT results, except for VPE-antigen, all other immunogens evaluated induced some level of neutralizing anti-SARS-CoV-2 antibodies, and the FDE-antigen stood out again with the most expressive values. Taken together, the present work shows that FDE can be an efficient and affordable alternative to BPL for the production of inactivated SARS-CoV-2 viral antigen.


Subject(s)
COVID-19 , SARS-CoV-2 , Animals , Antibodies, Viral , Antigens, Viral , Chlorocebus aethiops , Disease Models, Animal , Hydrogen Peroxide , Mice , Vero Cells
5.
Acta Trop ; 228: 106314, 2022 Apr.
Article in English | MEDLINE | ID: mdl-35038424

ABSTRACT

The anti-inflammatory and cardioprotective potential of coumarin metabolites in infectious myocarditis remains overlooked. Thus, the impact of the synthetic 4-nitrobenzoylcoumarin (4NB) alone and combined with benznidazole (Bz) in a murine model of Trypanosoma cruzi-induced acute myocarditis was investigated. Swiss mice infected with T. cruzi were randomized in 8 groups: uninfected, infected untreated or treated with 50 and 100 mg/kg 4NB or Bz alone and combined. Treatments were administered by gavage for 20 days. Cytokines (IL-2, IL-6, IL-10, IL-17, TNFα, and IFN-γ), immunoglobulin reactivity index (total IgG, IgG1, IgG2a and IgG2b), atrial natriuretic peptide (ANP), parasitemia, serum transaminases, heart and liver cellularity were analyzed. T. cruzi infection induced blood parasitism, heart and liver inflammation, upregulated all cytokines, IgG reactivity index, ANP and transaminase levels, determining 43% mortality in untreated mice. Transaminase levels, mean parasitemia, heart inflammation and ANP were reduced in 4NB-treated mice, reaching a 100% survival rate. Total survival (100%) was also obtained in all combinations of Bz and 4NB, which were effective in reducing blood parasitism, transaminases, cytokines and ANP levels, IgG reactivity index, liver and heart interstitial cellularity compared to 50 mg/kg Bz. Our findings indicated that 4NB alone and combined with Bz was well tolerated, showing no evidence of hepatotoxicity. Mainly in combination, these drugs exerted protective effects against T. cruzi-induced acute myocarditis by attenuating blood parasitism, systemic and heart inflammation. Thus, combinations based on 4NB and Bz are potentially relevant to develop new and more effective drug regimens for the treatment of T. cruzi-induced myocarditis.


Subject(s)
Chagas Disease , Nitroimidazoles , Trypanocidal Agents , Trypanosoma cruzi , Animals , Anti-Inflammatory Agents/therapeutic use , Antiparasitic Agents/therapeutic use , Chagas Disease/parasitology , Disease Models, Animal , Mice , Nitroimidazoles/pharmacology , Nitroimidazoles/therapeutic use , Parasitemia/drug therapy , Parasitemia/parasitology , Trypanocidal Agents/pharmacology , Trypanocidal Agents/therapeutic use
6.
Planta Med ; 87(12-13): 1045-1060, 2021 Oct.
Article in English | MEDLINE | ID: mdl-34530481

ABSTRACT

In the present study, the ethanolic extract from aerial parts of Ageratum fastigiatum was evaluated in vitro against epimastigote forms of Trypanosoma cruzi (Y strain), promastigote forms of Leishmania amazonensis (PH8 strain), and L. chagasi (BH400 strain). The extract was also evaluated against Staphylococcus aureus (ATCC 25 923), Escherichia coli (ATCC 11 775), Pseudomonas aeruginosa (ATCC 10 145), and Candida albicans (ATCC 36 802). The phytochemical screening was performed by thin-layer chromatography and high-performance liquid chromatography. The extract was fractionated using flash preparative chromatography. The ethanolic extract showed activity against T. cruzi, L. chagasi, and L. amazonensis and antimicrobial activity against S. aureus, E. coli, P. aeruginosa, and C. albicans. The phytochemical screening revealed coumarins, terpenes/sterols, and flavonoids in the ethanolic extract. In addition, the coumarin identified as ayapin was isolated from this extract. We also performed in silico prediction of potential biological activities and targets for compounds previously found in A. fastigiatum. Several predictions were confirmed both retrospectively and prospectively by experimental results described here or elsewhere. Some activities described in the in silico target fishing approach were validated by the ethnopharmacological use and known biological properties. Some new activities and/or targets were predicted and could guide future studies. These results suggest that A. fastigiatum can be an interesting source of substances with antiparasitic and antimicrobial activities.


Subject(s)
Ageratum , Computer Simulation , Escherichia coli , Microbial Sensitivity Tests , Phytochemicals/pharmacology , Plant Extracts/pharmacology , Retrospective Studies , Staphylococcus aureus
7.
Mem Inst Oswaldo Cruz ; 115: e200287, 2021.
Article in English | MEDLINE | ID: mdl-33533869

ABSTRACT

BACKGROUND: The heat-labile nature of Dengue virus (DENV) in serum samples must be considered when applying routine diagnostic tests to avoid issues that could impact the accuracy of test results with direct implications for case management and disease reporting. OBJECTIVES: To check if pre-analytical variables, such as storage time and temperature, have an impact on the accuracy of the main routine diagnostic tests for dengue. METHODS: Virus isolation, reverse transcription real-time polymerase chain reaction (RT-PCR) and NS1 enzyme-linked immunosorbent assay (ELISA) were evaluated using 84 samples submitted to different pre-analytical conditions. FINDINGS: Sensitivity and negative predictive value were directly affected by sample storage conditions. RT-PCR and virus isolation showed greater dependence on well-conserved samples for an accurate diagnosis. Interestingly, even storage at -30ºC for a relatively short time (15 days) was not adequate for accurate results using virus isolation and RT-PCR tests. On the other hand, NS1 ELISA showed no significant reduction in positivity for aliquots tested under the same conditions as in the previous tests. MAIN CONCLUSIONS: Our results support the stability of the NS1 marker in ELISA diagnosis and indicate that the accuracy of routine tests such as virus isolation and RT-PCR is significantly affected by inadequate transport and storage conditions of serum samples.


Subject(s)
Antigens, Viral/blood , Dengue Virus/isolation & purification , Dengue/diagnosis , Enzyme-Linked Immunosorbent Assay/methods , Immunologic Tests/methods , Reverse Transcriptase Polymerase Chain Reaction/methods , Viral Nonstructural Proteins/immunology , Antibodies, Viral/blood , Antigens, Viral/immunology , Dengue/blood , Dengue/virology , Dengue Virus/genetics , Dengue Virus/immunology , Humans , Predictive Value of Tests , Sensitivity and Specificity , Viral Nonstructural Proteins/genetics
8.
J Med Entomol ; 58(1): 333-342, 2021 01 12.
Article in English | MEDLINE | ID: mdl-32785582

ABSTRACT

We provide the first evidence of a recent invasion of Aedes aegypti (Linnaeus in Hasselquist, 1762) and Aedes albopictus (Skuse 1894), followed by dengue virus, in tropical montane cities in south-eastern Brazil, Mariana, and Ouro Preto, at mid and high altitudes, respectively. Long-term temperature variation, dengue public data, and sampling of immature and adult mosquitoes (ovitraps and mosquitraps) in contrasting habitats were used to explain the distribution of Aedes in what in these two cities. From 1961 to 2014, the annual temperature increased significantly due to increases in winter temperatures. In the 1990s/2000s, the winter temperature was 1.3°C warmer than in the 1960s, when it varied from 21.2 to 18.9°C. After 2007, the winter temperatures increased and ranged from 21.6 to 21.3°C. The first autochthonous dengue cases in Mariana and Ouro Preto were in 2007, followed by few occurrences until in 2012, when the mean numbers increased three-fold, and peak at 2013. The continuous 'warmer winter' may have trigged the Aedes invasion. Aedes species benefited from higher winter temperatures, which was an important driver of their invasion of the state of Minas Gerais in the 1980s and, more recently, in the remaining montane urban habitats in this region. In both 2009 and 2011, we found more Aedes in Mariana than Ouro Preto, and more Ae. albopictus in green areas and Ae. aegypti in houses, the expected pattern for well-established populations.


Subject(s)
Aedes/physiology , Animal Distribution , Ecosystem , Weather , Aedes/growth & development , Altitude , Animals , Brazil , Cities , Climate , Larva/growth & development , Larva/physiology , Pupa/growth & development , Pupa/physiology , Seasons
9.
Parasitol Res ; 119(5): 1683-1690, 2020 May.
Article in English | MEDLINE | ID: mdl-32285265

ABSTRACT

The diagnosis of canine visceral leishmaniasis (CVL) has been a problem for public health services due to the variety of clinical signs similar to other diseases and low sensitivity and specificity of available tests. In this sense, our main objective was to develop a simple, rapid, and accurate quantitative real-time PCR (qPCR) diagnosis for CVL. Thus, low-invasive samples from bone marrow (BM), popliteal lymph nodes (PLN), and conjunctival swabs (CS) were selected from negative and VL-positive dogs, using as gold standard, immunological and parasitological tests performed with different tissues. Oligonucleotides for Leishmania infantum kDNA were designed and the limit of quantification and amplification efficiency of the qPCR were determined using tissue-specific standards produced with DNA from those different tissues, mixed with DNA from a known amount of L. infantum promastigotes. Endogenous control was used to validate a comparative Ct method, and tissue parasite concentrations were estimated by comparison with tissue-specific reference standard samples. The overall analysis of the qPCR data suggests the following ranking for tissue choice: PLN > BM > CS. Finally, we have concluded that this molecular approach simplifies and accelerates the quantitative diagnostic process because it is easy to perform, requiring no DNA dosing or standard curve application, and it shows good diagnostic parameters, especially when using popliteal lymph node samples.


Subject(s)
Dog Diseases/diagnosis , Leishmania infantum/genetics , Leishmaniasis, Visceral/diagnosis , Leishmaniasis, Visceral/veterinary , Real-Time Polymerase Chain Reaction/methods , Animals , Bone Marrow/parasitology , DNA, Kinetoplast/genetics , Dog Diseases/parasitology , Dogs , Leishmaniasis, Visceral/parasitology , Lymph Nodes/parasitology , Molecular Diagnostic Techniques/methods , Real-Time Polymerase Chain Reaction/veterinary , Sensitivity and Specificity , Spleen/parasitology
10.
Waste Manag Res ; 38(5): 485-486, 2020 May.
Article in English | MEDLINE | ID: mdl-32303151

ABSTRACT

Wastes generated in healthcare facilities have been discussed and the World Health Organization has proposed a guideline for controlling the spread of the virus that causes Coronavirus Disease 2019 (COVID-19). However, waste management outside the generating facility should be discussed in more detail, taking into account factors such as virus resistance, differences in waste management systems and the climatic conditions in each affected region. Patients infected by human coronavirus being treated at home are generating infected waste possibly discarded as domestic waste, which can pose risks to workers and the environment, depending on the conditions of transport and disposal. In particular, the spread of the coronavirus may be increased by inadequate waste management, highlighting poor handling conditions associated with inappropriate use of personal protective equipment and other unfavourable conditions presented mainly in developing countries.


Subject(s)
Coronavirus Infections , Health Facilities , Medical Waste Disposal , Solid Waste , Waste Management , Betacoronavirus , COVID-19 , Epidemics , Humans , Pandemics , Pneumonia, Viral , SARS-CoV-2
11.
Future Med Chem ; 12(7): 571-581, 2020 04.
Article in English | MEDLINE | ID: mdl-32116030

ABSTRACT

Aim: Cysteine proteases are important molecular targets involved in the replication, virulence and survival of parasitic organisms, including Trypanosoma and Leishmania species. Methodology & results: Analogs of the 7-chloro-N-[3-(morpholin-4-yl)propyl]quinolin-4-amine were synthesized and their inhibitory activity against the enzymes cruzain and rhodesain as well as against promastigotes forms of Leishmania species and epimastigotes forms of Trypanosoma cruzi were evaluated. Five compounds showed activity against both enzymes with half maximal inhibitory concentration (IC50) values ranging from 23 to 123 µM. Among these, compounds 3 and 4 displayed leishmanicidal activity; compound 4 was the most promising with IC50 values <10 µM and no cytotoxicity for uninfected cells. Conclusion: The results obtained indicate that cysteine proteases are likely to be the molecular target of compounds 3 and 4.


Subject(s)
Antiprotozoal Agents/pharmacology , Cysteine Endopeptidases/metabolism , Cysteine Proteinase Inhibitors/pharmacology , Protozoan Proteins/antagonists & inhibitors , Quinolines/pharmacology , Antiprotozoal Agents/chemical synthesis , Antiprotozoal Agents/chemistry , Cysteine Proteinase Inhibitors/chemical synthesis , Cysteine Proteinase Inhibitors/chemistry , Leishmania/drug effects , Molecular Docking Simulation , Molecular Structure , Parasitic Sensitivity Tests , Protozoan Proteins/metabolism , Quinolines/chemical synthesis , Quinolines/chemistry , Trypanosoma cruzi/drug effects
12.
Acta Trop ; 189: 30-38, 2019 Jan.
Article in English | MEDLINE | ID: mdl-30290285

ABSTRACT

It is still unclear whether the progression of acute to chronic Chagas cardiomyopathy is predominantly associated with the limited efficacy of aetiological chemotherapy, or with the pharmacological resistance profiles and pathogenicity of specific Trypanosoma cruzi strains. Thus, we tested the hypothesis that parasitic load could be a limited target of aetiological chemotherapy to prevent chronic cardiomyopathy in dogs infected by different T. cruzi strains. Animals were infected with benznidazole-susceptible (Berenice-78) and -resistant (VL-10 and AAS) strains of T. cruzi. A quantitative real-time PCR strategy was developed to comparatively quantify the parasite load of the three different strains using a single standard curve. For dogs infected with the VL-10 strain, benznidazole treatment reduced cardiac parasitism during the acute phase of infection. However, similar parasite load and collagen deposition were detected in the myocardium of treated and untreated animals in the chronic phase of the infection. In animals infected with the AAS strain, benznidazole reduced parasite load, myocarditis and type III collagen deposition in the acute phase. However, increased type III collagen deposition was verified in the chronic phase. Dogs infected with the Berenice-78 strain showed a parasitological cure and no evidence of myocardial fibrosis. Parasitic load and cardiac fibrosis presented no correlation in acute or chronic phases of T. cruzi infection. Our findings in a canine model of Chagas disease suggest that parasite burden is a limited predictor for disease progression after treatment and show that benznidazole, although not inducing parasitological cure, is able to prevent total fibrosis in the early stages of infection, as well as complete prevention of cardiac damage when it eliminates parasites at the onset of infection.


Subject(s)
Chagas Cardiomyopathy/drug therapy , Chagas Cardiomyopathy/prevention & control , Myocardium/pathology , Parasite Load , Parasitemia/drug therapy , Trypanosoma cruzi , Acute Disease , Animals , Chagas Cardiomyopathy/parasitology , Chronic Disease , Collagen Type III/metabolism , DNA, Protozoan/blood , Disease Models, Animal , Disease Progression , Dogs , Drug Resistance , Fibrosis , Heart/parasitology , Myocardium/metabolism , Nitroimidazoles/therapeutic use , Parasitemia/blood , Parasitemia/parasitology , Trypanocidal Agents/therapeutic use , Trypanosoma cruzi/genetics
13.
Acta Trop ; 167: 40-49, 2017 Mar.
Article in English | MEDLINE | ID: mdl-27993495

ABSTRACT

This study was designed to verify the relationship between IgG antibodies isotypes and myocarditis in Trypanosoma cruzi infection using mice and dogs infected with different T. cruzi strains. The animals were infected with benznidazole-susceptible Berenice-78 and benznidazole-resistant AAS and VL-10 strains. The IgG subtypes were measured in serum samples from dogs (IgG, IgG1, and IgG2) and mice (IgG, IgG1, IgG2a, and IgG2b). The infection of dogs with VL-10 strain induced the highest levels of heart inflammation while intermediate and lower levels were detected with Berenice-78 and AAS strains, respectively. Similar results were found in mice infected with VL-10, but not in those infected with AAS or Berenice-78 strains. The AAS strain induced higher levels of heart inflammation in mice, while Berenice-78 strain was not able to induce it. Correlation analysis between myocarditis and antibody reactivity index revealed very interesting results, mainly for IgG and IgG1, the latter being the most exciting. High IgG1 showed a significant correlation with myocarditis in both experimental models, being more significant in dogs (r=0.94, p<0.0001) than in mice (r=0.58, p=0.047). Overall, our data suggest that IgG1 could be a good marker to demonstrate myocarditis intensity in Chagas disease.


Subject(s)
Chagas Disease/parasitology , Immunoglobulin G/blood , Myocarditis/parasitology , Trypanosoma cruzi/immunology , Animals , Chagas Disease/blood , Chagas Disease/veterinary , Disease Models, Animal , Dogs , Male , Mice , Myocarditis/blood , Myocarditis/veterinary , Nitroimidazoles
14.
Am J Trop Med Hyg ; 95(1): 193-200, 2016 07 06.
Article in English | MEDLINE | ID: mdl-27139443

ABSTRACT

Dengue is responsible for a wide range of clinical manifestations, ranging from asymptomatic infections to severe cases. The alteration of cytokine levels correlated with clinical characteristics can help determine prognostic markers of the disease and the identification of targets for immunotherapy. We measured the viral load, serotype, and cytokine levels of 212 serum samples from patients with acute dengue infection during days 1-4 after the onset of symptoms. The patients were classified as either with hemorrhagic manifestations (HM) or with no hemorrhagic manifestations (NHM). The cytokines interleukin-6 (IL-6), IL-8, and IL-10 were increased (P < 0.05) in the dengue virus+ group, compared with the control group. A higher viral load (P < 0.05) and IL-6 was detected in the HM group compared with the NHM group. Interestingly, the NHM group demonstrated a significant positive correlation between inflammatory (IL-6 and 8) and anti-inflammatory (IL-10) cytokines, whereas the HM group did not. These findings suggest that a disturbance in the balance of inflammatory cytokines IL-6 and IL-8 with the anti-inflammatory cytokine, IL-10, combined with the high levels of IL-6 and viral load, characterize possible mechanisms related to the formation of HM.


Subject(s)
Dengue/blood , Dengue/immunology , Interleukin-10/blood , Interleukin-6/blood , Interleukin-8/blood , Adolescent , Adult , Dengue/diagnosis , Dengue Virus/isolation & purification , Female , Humans , Male , Middle Aged , RNA, Viral/isolation & purification , Viral Load , Young Adult
15.
Acta Trop ; 161: 44-54, 2016 Sep.
Article in English | MEDLINE | ID: mdl-27215760

ABSTRACT

Cardiac involvement represents the main cause of mortality among patients with Chagas disease, and the relevance of trypanocidal treatment to improving diastolic dysfunction is still doubtful. In the present study, we used a canine model infected with the benznidazole-sensitive Berenice-78 Trypanosoma cruzi strain to verify the efficacy of an etiologic treatment in reducing the parasite load and ameliorating cardiac muscle tissue damage and left ventricular diastolic dysfunction in the chronic phase of the infection. The effect of the treatment on reducing the parasite load was monitored by blood PCR and blood culture assays, and the effect of the treatment on the outcome of heart tissue damage and on diastolic function was evaluated by histopathology and echo Doppler cardiogram. The benefit of the benznidazole-treatment in reducing the parasite burden was demonstrated by a marked decrease in positive blood culture and PCR assay results until 30days post-treatment. At this time, the PCR and blood culture assays yielded negative results for 82% of the treated animals, compared with only 36% of the untreated dogs. However, a progressive increase in the parasite load could be detected in the peripheral blood for one year post-treatment, as evidenced by a progressive increase in positive results for both the PCR and the blood culture assays at follow-up. The parasite load reduction induced by treatment was compatible with the lower degree of tissue damage among animals euthanized in the first month after treatment and with the increased cardiac damage after this period, reaching levels similar to those in untreated animals at the one-year follow-up. The two infected groups also presented similar, significantly smaller values for early tissue septal velocity (E' SIV) than the non-infected dogs did at this later time. Moreover, in the treated animals, an increase in the E/E' septal tissue filling pressure ratio was observed when compared with basal values as well as with values in non-infected dogs. These findings strongly suggest that the temporary reduction in the parasite load that was induced by benznidazole treatment was not able to prevent myocardial lesions and diastolic dysfunction for long after treatment.


Subject(s)
Chagas Cardiomyopathy/drug therapy , Chagas Cardiomyopathy/parasitology , Heart/parasitology , Myocardium/pathology , Nitroimidazoles/therapeutic use , Trypanocidal Agents/pharmacology , Trypanocidal Agents/therapeutic use , Animals , Dogs , Female , Humans , Male , Models, Animal , Nitroimidazoles/pharmacology , Parasite Load , Polymerase Chain Reaction , Trypanosoma cruzi/drug effects
16.
Rev. bras. farmacogn ; 26(2): 197-202, Jan.-Apr. 2016. tab, graf
Article in English | LILACS | ID: lil-779010

ABSTRACT

ABSTRACT Myracrodruon urundeuva Allemão, Anacardiaceae, is a medicinal plant widely found in Brazil, especially in the northern region. In our previous study, the ethanolic extract from leaves of M. urundeuva showed antiviral activity against simian rotavirus SA-11. Here, the crude extract was subjected to fractionations in order to subsequently work with more concentrated and pure bioactive compounds, which were analyzed by TLC and HPLC methods to support a better understanding of their virucidal effect. The antiviral activity was evaluated using a rotavirus infection model in MA-104 cells treated with the maximum non-cytotoxic concentration of the crude extract and its fractions. Data were expressed as the percentage inhibition of viral replication calculated by the inhibition of cytopathic effect in the treated cells compared to untreated controls after 48 h of incubation. First, we conducted a fractionation, generating five fractions (F1–F5) which were submitted to antiviral assay. Then, the fraction that showed the highest virucidal effect (F3, PI = 75%) was subjected to a larger partition, yielding eighteen subfractions, which were submitted to new antiviral assays. Terpenes, flavonoids and tannins were the major secondary metabolites detected by TLC analysis in F3. SF1, a flavonoid-enriched fraction, showed the strongest in vitro activity against rotavirus (PI = 92%), preventing cytopathic effect. Chromatographic profiles were obtained by HPLC for the crude extract and SF1, the most potent subfraction. Overall, our data point to the potential anti-rotavirus activity of flavonoid-enriched fraction (SF1) of M. urundeuva leaves, corroborating the traditional use of this species to treat diarrhea and broadening our perspectives on in vivo assays in mice with SF1 isolated or associated with other fractions.

17.
PLoS One ; 10(6): e0128707, 2015.
Article in English | MEDLINE | ID: mdl-26076455

ABSTRACT

The nitroheterocyclic drugs nifurtimox and benznidazole are first-line drugs available to treat Chagas disease; however, they have limitations, including long treatment courses and toxicity. Strategies to overcome these limitations include the identification of new drugs with specific target profiles, re-dosing regimens for the current drugs, drug repositioning and combination therapy. In this work, we evaluated combination therapy as an approach for optimization of the current therapeutic regimen for Chagas disease. The curative action of benznidazole/itraconazole combinations was explored in an established infection of the mice model with the T. cruzi Y strain. The activities of the benznidazole/itraconazole combinations were compared with the results from those receiving the same dosage of each individual drug. The administration of benznidazole/itraconazole in combination eliminated parasites from the blood more efficiently than each drug alone. Here, there was a significant reduction of the number of treatment days (number of doses) necessary to induce parasitemia suppression with the benznidazole/itraconazole combination, as compared to each compound administered alone. These results clearly indicate the enhanced effects of these drugs in combination, particularly at the dose of 75 mg/kg, as the effects observed with the drug combinations were four times more effective than those of each drug used alone. Moreover, benznidazole/itraconazole treatment was shown to prevent or decrease the typical lesions associated with chronic experimental Chagas disease, as illustrated by similar levels of inflammatory cells and fibrosis in the cardiac muscle tissue of healthy and treated mice. These results emphasize the importance of exploring the potential of combination treatments with currently available compounds to specifically treat Chagas disease.


Subject(s)
Chagas Disease/parasitology , Itraconazole/pharmacology , Nitroimidazoles/pharmacology , Trypanocidal Agents/pharmacology , Trypanosoma cruzi/drug effects , Animals , Antibodies, Protozoan/immunology , Chagas Disease/drug therapy , Chagas Disease/immunology , Chagas Disease/pathology , Disease Models, Animal , Drug Therapy, Combination , Female , Immunoglobulin G/immunology , Itraconazole/administration & dosage , Mice , Myocardium/pathology , Nitroimidazoles/administration & dosage , Parasite Load , Trypanocidal Agents/administration & dosage , Trypanosoma cruzi/immunology
18.
Parasitology ; : 1-10, 2014 Jul 21.
Article in English | MEDLINE | ID: mdl-25045804

ABSTRACT

SUMMARY This study describes the role of parasite clearance time induced by benznidazole, fexinidazole and posaconazole treatments upon mice infection with a benznidazole-resistant Trypanosoma cruzi strain in the pathological outcomes. Trypanosoma cruzi-infected mice were treated with different drugs and parasite clearance time was detected by blood and tissue qPCR, to determine the dynamic relationship between the efficacy of the treatments and the intensity of heart lesion/serum inflammatory mediators. Our results indicate that anti-T. cruzi treatments were able to reduce parasite replication and consequently induce immunomodulatory effects, where the degree of the immunopathology prevention was related to the time of parasite clearance induced by different treatments. Nevertheless, in benznidazole and posaconazole treatments, parasite rebounding was detected with parasitism reaching levels similar to infected and non-treated mice; the time for parasitic rebound being earlier among benznidazole-treated mice. In parallel, an increase of cardiac lesions and plasma chemokine levels was also detected and was more accentuated in benznidazole-treated animals. Interestingly, in the presence of parasitological cure (fexinidazole treatment), basal levels of these inflammatory mediators were evidenced as well as an absence of cardiac inflammation or fibrosis. Overall, our data indicate that all treatments have positive effects on the clinical evolution of T. cruzi infection, with success in preventing cardiac alterations being drug-dependent.

19.
Toxins (Basel) ; 5(10): 1780-98, 2013 Oct 15.
Article in English | MEDLINE | ID: mdl-24131891

ABSTRACT

We report the detailed molecular characterization of two PLA2s, Lys49 and Asp49 isolated from Bothrops leucurus venom, and examined their effects against Dengue virus (DENV). The Bl-PLA2s, named BlK-PLA2 and BlD-PLA2, are composed of 121 and 122 amino acids determined by automated sequencing of the native proteins and peptides produced by digestion with trypsin. They contain fourteen cysteines with pIs of 9.05 and 8.18 for BlK- and BlD-PLA2s, and show a high degree of sequence similarity to homologous snake venom PLA2s, but may display different biological effects. Molecular masses of 13,689.220 (Lys49) and 13,978.386 (Asp49) were determined by mass spectrometry. DENV causes a prevalent arboviral disease in humans, and no clinically approved antiviral therapy is currently available to treat DENV infections. The maximum non-toxic concentration of the proteins to LLC-MK2 cells determined by MTT assay was 40 µg/mL for Bl-PLA2s (pool) and 20 µg/mL for each isoform. Antiviral effects of Bl-PLA2s were assessed by quantitative Real-Time PCR. Bl-PLA2s were able to reduce DENV-1, DENV-2, and DENV-3 serotypes in LLC-MK2 cells infection. Our data provide further insight into the structural properties and their antiviral activity against DENV, opening up possibilities for biotechnological applications of these Bl-PLA2s as tools of research.


Subject(s)
Antiviral Agents/isolation & purification , Dengue Virus/drug effects , Phospholipases A2/isolation & purification , Reptilian Proteins/isolation & purification , Snake Venoms/chemistry , Aedes , Amino Acid Sequence , Animals , Antiviral Agents/chemistry , Antiviral Agents/pharmacology , Bothrops , Cell Line , Macaca mulatta , Molecular Sequence Data , Phospholipases A2/chemistry , Phospholipases A2/pharmacology , Reptilian Proteins/chemistry , Reptilian Proteins/pharmacology , Sequence Alignment
20.
Acta Trop ; 123(3): 170-7, 2012 Sep.
Article in English | MEDLINE | ID: mdl-22609548

ABSTRACT

The lack of an accurate diagnosis has been a serious obstacle to the advancement of the anti-Trypanosoma cruzi chemotherapy and long-term infection can result in different health risks to human. PCRs are alternative methods, more sensitive than conventional parasitological techniques, which due to their low sensitivities are considered unsuitable for these purposes. The aim of this study was to investigate a sensitive diagnostic strategy to quantify blood and cardiac tissues parasites based on real-time PCR tools during acute and chronic phases of murine Chagas disease, as well as to monitor the evolution of infection in those mice under specific treatment. In parallel, fresh blood examination, immunological analysis and quantification of cardiac inflammation were also performed to confront and improve real-time PCR data. Similar profiles of parasitemia curves were observed in both quantification techniques during the acute phase of the infection. In contrast, parasites could be quantified only by real-time PCR at 60 and 120 days of infection. In cardiac tissue, real-time PCR detected T. cruzi DNA in 100% of infected mice, and using this tool a significant Pearson correlation between parasite load in peripheral blood and in cardiac tissue during acute and chronic phases was observed. Levels of serum CCL2, CCL5 and nitric oxide were coincident with parasite load but focal and diffuse mononuclear infiltrates was observed, even with significant (p<0.05) reduction of parasitism after 60 days of infection. Later, this methodology was used to monitor the evolution of infection in animals treated with itraconazole (Itz). Itz-treatment induced a reduction of parasite load in both blood and cardiac muscle at the treatment period, but after the end of chemotherapy an increase of parasitism was detected. Interestingly, inflammatory mediators levels and heart inflammation intensity had similar evolution to the parasite load, in the group of animals treated. Taken together, our data show that real-time PCR strategy used was suitable for studies of murine T. cruzi infection and may prove useful in investigations involving experimental chemotherapy of the disease and the benefits of treatment in relation to parasitism and inflammatory response.


Subject(s)
Chagas Disease/parasitology , Parasite Load/methods , Real-Time Polymerase Chain Reaction/methods , Trypanosoma cruzi/isolation & purification , Animals , Antiprotozoal Agents/administration & dosage , Blood/parasitology , Chagas Disease/drug therapy , Chemokine CCL2/blood , Chemokine CCL5/blood , Disease Models, Animal , Female , Heart/parasitology , Itraconazole/administration & dosage , Mice , Nitric Oxide/blood
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