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Onchocerciasis has been declared eliminated in Ecuador and surveillance measures are of great interest. In this study, we examined the infectivity rates of Simulium exiguum by Onchocerca volvulus in previously hyperendemic areas in Esmeraldas province of Ecuador. These areas had previously undergone mass administration of ivermectin, which led to the interruption of transmission in 2009 and the certification of elimination in 2014. The study included three communities in Río Cayapas and one in Río Canandé, and a total of 2,950 adult S. exiguum were collected in 2018. We used quantitative polymerase chain reaction with O. volvulus O-150 plasmid control DNA to analyze 59 pools. Our findings revealed that the infectivity rates were zero, indicating that the transmission of O. volvulus remained suspended in the area.
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Volvo Intestinal , Onchocerca volvulus , Oncocercose , Simuliidae , Humanos , Animais , Adulto , Oncocercose/diagnóstico , Oncocercose/epidemiologia , Oncocercose/prevenção & controle , Onchocerca volvulus/genética , Reação em Cadeia da Polimerase em Tempo Real , Equador/epidemiologia , Ivermectina/uso terapêutico , Onchocerca/genéticaRESUMO
Tick-borne diseases have increased significantly in Europe and Spain in recent years. One strategy explored for tick surveillance and control is the study of the microbiota. The focus is on understanding the relationships between pathogens and endosymbionts within the microbiota and how these relationships can alter these arthropods' vectorial capacity. Thus, it is pivotal to depict the bacterial communities composing the microbiota of ticks present in specific territories. This work aimed to describe the microbiota present in 29 adult individuals of 5 tick species collected from 4 provinces of Castilla y Leon in northwestern Spain from 2015 to 2022. DNA extraction and sequencing of the V4 hypervariable region of 16S-rRNA was performed on the tick samples, with subsequent analysis of diversity, taxonomic composition, and correlations between genera of microorganisms. There were no differences in the alpha diversity of microbiota by tick species, nor were compositional changes evident at the phylum level for microorganisms. However, interindividual differences at the microbial genus level allowed spatial differentiation of the 5 tick species included in the study. Correlation analyses showed complex interactions between different genera of microbiota members. These findings provide an initial insight into the composition of the gut microbiota of various tick species in northwestern Spain, which can contribute to establishing surveillance and control measures to reduce diseases such as rickettsiosis, Lyme disease, and Crimean-Congo hemorrhagic fever.
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Microbioma Gastrointestinal , Ixodidae , Doenças Transmitidas por Carrapatos , Carrapatos , Humanos , Animais , Carrapatos/microbiologia , Ixodidae/microbiologia , Espanha , Doenças Transmitidas por Carrapatos/epidemiologia , RNA Ribossômico 16S/genéticaRESUMO
Nowadays, safe and efficacious vaccines represent powerful and cost-effective tools for global health and economic growth. In the veterinary field, these are undoubtedly key tools for improving productivity and fighting zoonoses. However, cases of persistent infections, rapidly evolving pathogens having high variability or emerging/re-emerging pathogens for which no effective vaccines have been developed point out the continuing need for new vaccine alternatives to control outbreaks. Most licensed vaccines have been successfully used for many years now; however, they have intrinsic limitations, such as variable efficacy, adverse effects, and some shortcomings. More effective adjuvants and novel delivery systems may foster real vaccine effectiveness and timely implementation. Emerging vaccine technologies involving nanoparticles such as self-assembling proteins, virus-like particles, liposomes, virosomes, and polymeric nanoparticles offer novel, safe, and high-potential approaches to address many vaccine development-related challenges. Nanotechnology is accelerating the evolution of vaccines because nanomaterials having encapsulation ability and very advantageous properties due to their size and surface area serve as effective vehicles for antigen delivery and immunostimulatory agents. This review discusses the requirements for an effective, broad-coverage-elicited immune response, the main nanoplatforms for producing it, and the latest nanovaccine applications for fighting animal pathogens.
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BACKGROUND: Schistosomiasis has been identified as a major public health problem in tropical countries. The present study aimed to investigate the schistosomicidal effects of the methanolic extract of Argemone mexicana L. and its active component, berberine against Schistosoma mansoni on in-vitro experiments. METHODS: S. mansoni adults were used. Various concentrations of the methanolic extract (10 - 200 µg/ml) and berberine (2.5 - 50 µM) were tested from 24 to 72 h. The viability of S. mansoni was confirmed with an invertoscope-microscope. Furthermore, cytotoxic (Hemolysis test), and antioxidant (DPPH radical scavenging assay) capacities were determined. RESULTS: The viability tests on S. mansoni showed that A. mexicana at 50 µg/mL is lethal at 48 h and berberine at 10 µM is lethal at 24 h. The hemolytic activity at 1,000 µg/mL was 2.9% for A. mexicana and 90.2% for berberine. The antioxidant capacities shown by A. mexicana and berberine, were EC50 156.3 and 84.1 µg/mL, respectively. CONCLUSION: The extract of A. mexicana and berberine demonstrated high antischistosomal activities in low concentration and short exposure time on the in-vitro model.
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Strongyloidiasis is a parasitosis that represents a public health problem, in tropical regions. The present study aimed to investigate the anthelmintic effects of several extracts of Argemone mexicana, as well as its main component berberine (Ber) against the third-stage larvae (L3) of Strongyloides venezuelensis in-vitro experiments. Also, the anti-hemolytic activity of the extract, fractions, and Ber were tested in human erythrocytes. A dose-response anthelminthic bioassay demonstrated Ber as the most effective component, followed by methanolic subfraction (Fr3) and finally the crude extract of A. mexicana (Am) showing LC50 response values of 1.6, 19.5, and 92.1 µg/mL, at 96 h respectively. Also, Am, Fr3, and Ber did not produce significant hemolysis against human erythrocytes (p ≤ 0.05). Am and Fr3 showed erythrocyte protection effect capacity at the membrane level (p ≤ 0.05). Furthermore, Ber was found to have an antioxidant activity of 168.18 µg/mL. According to the results, the Fr3 of A. mexicana, and particularly Ber, exhibited potent in-vitro effects against L3 of S. venezuelensis, without hemolytic activity against human erythrocytes and presented good antioxidant capacity. In conclusion, the extracts of A. mexicana and the main component have activity against S. venezuelensis, nevertheless, further studies are required to elucidate the mechanism of action.
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Anti-Helmínticos/farmacologia , Argemone/química , Berberina/farmacologia , Extratos Vegetais/farmacologia , Strongyloides/efeitos dos fármacos , Análise de Variância , Animais , Anti-Helmínticos/química , Anti-Helmínticos/uso terapêutico , Berberina/química , Berberina/uso terapêutico , Bioensaio , Relação Dose-Resposta a Droga , Eritrócitos/efeitos dos fármacos , Fezes/parasitologia , Hemólise/efeitos dos fármacos , Humanos , Larva/efeitos dos fármacos , Dose Letal Mediana , Masculino , Extratos Vegetais/química , Extratos Vegetais/uso terapêutico , Folhas de Planta/química , Caules de Planta/química , Ratos , Ratos Wistar , Estrongiloidíase/tratamento farmacológicoRESUMO
Chagas' disease is a neglected tropical disease caused by Trypanosoma cruzi which is endemic throughout Latin America and is spread by worldwide migration. Diagnosis is currently limited to serological and molecular techniques having variations regarding their sensitivity and specificity. This work was aimed at developing a new sensitive, applicable, and cost-effective molecular diagnosis technique for loop-mediated isothermal amplification-based detection of T. cruzi (Tc-LAMP). The results led to determining a highly homologous satellite repeat region (231 bp) among parasite strains as a molecular marker for diagnosing the disease. Tc-LAMP was performed correctly for detecting parasite DNA (5 fg for the CL Brener strain and 50 fg for the DM28, TcVI, and TcI strains). Assay results proved negative for DNA from 16 helminth species and 7 protozoa, including Leishmania spp. Tc-LAMP based on the highly repeated T. cruzi satellite region is thus proposed as an important alternative for diagnosing T. cruzi infection, overcoming other methods' limitations such as their analytic capability, speed, and requiring specialized equipment or highly trained personnel. Tc-LAMP could be easily adapted for point-of-care testing in areas having limited resources.
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Doença de Chagas/diagnóstico , DNA Satélite/genética , Técnicas de Diagnóstico Molecular/métodos , Técnicas de Amplificação de Ácido Nucleico/métodos , Trypanosoma cruzi/isolamento & purificação , DNA de Protozoário/genética , Marcadores Genéticos , Humanos , Técnicas de Diagnóstico Molecular/economia , Técnicas de Amplificação de Ácido Nucleico/economia , Testes Imediatos , Sensibilidade e Especificidade , Trypanosoma cruzi/genéticaRESUMO
Bovine leukocyte antigens (BoLA) have been used as disease markers and immunological traits in cattle due to their primary role in pathogen recognition by the immune system. A higher MHC allele diversity in a population will allow presenting a broader peptide repertoire. However, loss of overall diversity due to domestication process can decrease a population's peptide repertoire. Within the context of zebu and taurine cattle populations, BoLA-DRB3 genetic diversity in Spanish Morucha and Colombian Normande cattle was analyzed and an approach to estimate functional diversity was performed. Sequence-based typing was used for identifying 29, 23, 27, and 28 alleles in Spanish Morucha, Nariño-, Boyacá-, and Cundinamarca-Normande cattle, respectively. These breeds had remarkably low heterozygosity levels and the Hardy-Weinberg principle revealed significant heterozygote deficiency. FST and DA genetic distance showed that Colombian Normande populations had greater variability than other phenotypically homogeneous breeds, such as Holstein. It was also found that Spanish Morucha cattle were strongly differentiated from other cattle breeds. Spanish Morucha had greater divergence in the peptide-binding region regarding other cattle breeds. However, peptide-binding region covariation indicated that the potential peptide repertoire seemed equivalent among cattle breeds. Despite the genetic divergence observed, the extent of the potential peptide repertoire in the cattle populations studied appears to be similar and thus their pathogen recognition potential should be equivalent, suggesting that functional diversity might persist in the face of bottlenecks imposed by domestication and breeding.
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Amphimeriasis, a fish-borne zoonotic disease caused by the liver fluke Amphimerus spp., has recently been reported as an emerging disease affecting an indigenous Ameridian group, the Chachi, living in Ecuador. The only method for diagnosing amphimeriasis was the microscopic detection of eggs from the parasite in patients' stool samples with very low sensitivity. Our group developed an ELISA technique for detection of anti-Amphimerus IgG in human sera and a molecular method based on LAMP technology (named LAMPhimerus) for specific and sensitive parasite DNA detection. The LAMPhimerus method showed to be much more sensitive than classical parasitological methods for amphimeriasis diagnosis using human stool samples for analysis. The objective of this work is to demonstrate the feasibility of using dried stool samples on filter paper as source of DNA in combination with the effectiveness of our previously designed LAMPhimerus assay for successfully Amphimerus sp. detection in clinical stool samples. A total of 102 untreated and undiluted stool samples collected from Chachi population were spread as thin layer onto common filter paper for easily transportation to our laboratory and stored at room temperature for one year until DNA extraction. When LAMPhimerus method was applied for Amphimerus sp. DNA detection, a higher number of positive results was detected (61/102; 59.80%) in comparison to parasitological methods (38/102; 37.25%), including 28/61 (45.90%) microscopy-confirmed Amphimerus sp. infections. The diagnostic parameters for the sensitivity and specificity werecalculated for our LAMPhimerus assay, which were 79.17% and 65.98%, respectively. We demonstrate, for the first time, that common filter paper is useful for easy collection and long-term storage of human stool samples for later DNA extraction and molecular analysis of human-parasitic trematode eggs. This simple, economic and easily handling method combined with the specific and sensible LAMPhimerus assay has the potential to beused as an effective molecular large-scale screening test for amphimeriasis-endemic areas.
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Fezes/parasitologia , Zoonoses/diagnóstico , Adolescente , Adulto , Idoso , Animais , Criança , Pré-Escolar , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Lactente , Masculino , Pessoa de Meia-Idade , Opisthorchidae , Adulto JovemRESUMO
BACKGROUND: Amphimeriasis is a fish-borne disease caused by the liver fluke Amphimerus spp. that has recently been reported as endemic in the tropical Pacific side of Ecuador with a high prevalence in humans and domestic animals. The diagnosis is based on the stool examination to identify parasite eggs, but it lacks sensitivity. Additionally, the morphology of the eggs may be confounded with other liver and intestinal flukes. No immunological or molecular methods have been developed to date. New diagnostic techniques for specific and sensitive detection of Amphimerus spp. DNA in clinical samples are needed. METHODOLOGY/PRINCIPAL FINDINGS: A LAMP targeting a sequence of the Amphimerus sp. internal transcribed spacer 2 region was designed. Amphimerus sp. DNA was obtained from adult worms recovered from animals and used to optimize the molecular assays. Conventional PCR was performed using outer primers F3-B3 to verify the proper amplification of the Amphimerus sp. DNA target sequence. LAMP was optimized using different reaction mixtures and temperatures, and it was finally set up as LAMPhimerus. The specificity and sensitivity of both PCR and LAMP were evaluated. The detection limit was 1 pg of genomic DNA. Field testing was done using 44 human stool samples collected from localities where fluke is endemic. Twenty-five samples were microscopy positive for Amphimerus sp. eggs detection. In molecular testing, PCR F3-B3 was ineffective when DNA from fecal samples was used. When testing all human stool samples included in our study, the diagnostic parameters for the sensitivity and specificity were calculated for our LAMPhimerus assay, which were 76.67% and 80.77%, respectively. CONCLUSIONS/SIGNIFICANCE: We have developed and evaluated, for the first time, a specific and sensitive LAMP assay for detecting Amphimerus sp. in human stool samples. The procedure has been named LAMPhimerus method and has the potential to be adapted for field diagnosis and disease surveillance in amphimeriasis-endemic areas. Future large-scale studies will assess the applicability of this novel LAMP assay.
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Fezes/parasitologia , Técnicas de Diagnóstico Molecular/métodos , Técnicas de Amplificação de Ácido Nucleico/métodos , Opisthorchidae/isolamento & purificação , Infecções por Trematódeos/diagnóstico , Animais , DNA de Helmintos/genética , DNA Espaçador Ribossômico/genética , Equador , Humanos , Opisthorchidae/genética , Sensibilidade e Especificidade , Infecções por Trematódeos/parasitologiaRESUMO
Fasciolosis is listed as one of the most important neglected tropical diseases according with the World Health Organization and is also considered as a reemerging disease in the human beings. Despite there are several studies describing the immune response induced by Fasciola hepatica in the mammalian host, investigations aimed at identifying the expression profile of genes involved in inducing hepatic injury are currently scarce. Data presented here belong to a time-course investigation of the gene expression profile in the liver of BALB/c mice infected with F. hepatica metacercariae at 7 and 21 days after experimental infection. The data published here have been deposited in NCBI's Gene Expression Omnibus and are accessible through GEO Series accession number GSE69588, previously published by Rojas-Caraballo et al. (2015) in PLoS One [1].
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Fasciolosis is considered the most widespread trematode disease affecting grazing animals around the world; it is currently recognised by the World Health Organisation as an emergent human pathogen. Triclabendazole is still the most effective drug against this disease; however, resistant strains have appeared and developing an effective vaccine against this disease has increasingly become a priority. Several bioinformatics tools were here used for predicting B- and T-cell epitopes according to the available data for Fasciola hepatica protein amino acid sequences. BALB/c mice were immunised with the synthetic peptides by using the ADAD vaccination system and several immune response parameters were measured (antibody titres, cytokine levels, T-cell populations) to evaluate their ability to elicit an immune response. Based on the immunogenicity results so obtained, seven peptides were selected to assess their protection-inducing ability against experimental infection with F. hepatica metacercariae. Twenty-four B- or T-epitope-containing peptides were predicted and chemically synthesised. Immunisation of mice with peptides so-called B1, B2, B5, B6, T14, T15 and T16 induced high levels of total IgG, IgG1 and IgG2a (p<0.05) and a mixed Th1/Th2/Th17/Treg immune response, according to IFN-γ, IL-4, IL-17 and IL-10 levels, accompanied by increased CD62L+ T-cell populations. A high level of protection was obtained in mice vaccinated with peptides B2, B5, B6 and T15 formulated in the ADAD vaccination system with the AA0029 immunomodulator. The bioinformatics approach used in the present study led to the identification of seven peptides as vaccine candidates against the infection caused by Fasciola hepatica (a liver-fluke trematode). However, vaccine efficacy must be evaluated in other host species, including those having veterinary importance.