RESUMO
Fasciolosis caused by Fasciola hepatica severely affects the efficiency of livestock production systems worldwide. In addition to the economic impact inflicted on livestock farmers, fasciolosis is an emergent zoonosis. This review emphasizes different aspects of the disease in South America. Available data on epidemiology in bovines and ovines in different countries, as well as a growing body of information on other domestic and wildlife definitive hosts, are summarized. The issue of drug resistance that compromises the long-term sustainability of current pharmacological strategies is examined from a regional perspective. Finally, efforts to develop a single-antigen recombinant vaccine in ruminants are reviewed, focusing on the cases of leucine aminopeptidase or thioredoxin glutathione reductase.
Assuntos
Doenças dos Bovinos/epidemiologia , Fasciolíase/veterinária , Doenças dos Ovinos/prevenção & controle , Animais , Bovinos , Doenças dos Bovinos/parasitologia , Doenças dos Bovinos/prevenção & controle , Resistência a Medicamentos , Fasciola hepatica/efeitos dos fármacos , Fasciola hepatica/fisiologia , Fasciolíase/epidemiologia , Fasciolíase/parasitologia , Fasciolíase/prevenção & controle , Ovinos , Doenças dos Ovinos/epidemiologia , Doenças dos Ovinos/parasitologia , América do Sul/epidemiologiaRESUMO
Cestodes show a remarkable proliferative capability that sustains the constant growth and differentiation of proglottids essential for their lifestyle. It is believed that a separate population of undifferentiated stem cells (the so-called germinative cells) are the only cells capable of proliferation during growth and development. The study of this particular cell subpopulation is hampered by the current lack of methods to isolate it. In this work, we developed a reproducible flow cytometry and cell sorting method to quantify and isolate the proliferating cells in the tetrathyridia larvae of the model cestode Mesocestoides corti, based on the DNA content of the cells. The isolated cells display the typical germinative cell morphology, and can be used for RNA isolation with a yield in the ng to µg range. We expect that this approach may facilitate the characterization of the germinative cells in M. corti and other model tapeworms.
Assuntos
Separação Celular/métodos , Citometria de Fluxo/métodos , Mesocestoides/citologia , Animais , Benzimidazóis , Ciclo Celular , Proliferação de Células , Fluoresceínas , Corantes Fluorescentes , Indicadores e Reagentes , Larva/citologia , Mesocestoides/crescimento & desenvolvimento , Camundongos , Modelos Animais , Propídio , Reprodutibilidade dos Testes , Tripsina/metabolismoRESUMO
OBJECTIVES: To estimate the national prevalence and analyse the factors associated with preconceptional folic acid supplementation, including maternal sociodemographic characteristics, region of residence, birth control use and chronic diseases requiring medical care before conception. DESIGN: Cross-sectional population-based study. SETTING: All maternity units in France. POPULATION: A nationally representative sample of women giving birth in 2010 (n = 12,646). METHODS: Data came from mothers' interviews 2-3 days after delivery. Statistical analyses included multivariable logistic regressions. MAIN OUTCOME MEASURE: Folic acid supplementation starting at least 1 month before conception. RESULTS: 14.8% (95% confidence interval [95% CI] 14.2-15.4) of women used folic acid before pregnancy; this percentage varied from 10.4% to 18.7% across regions. Supplementation was more frequent in primiparae, French citizens, women with higher educational levels and those needing medical monitoring or treatment before conception. Women who stopped contraception to become pregnant (75% of our population) used folic acid more often (intrauterine device or implant: 19%, pill: 17%, other methods which did not need medical monitoring: 17%) than other women (7%). The adjusted odds ratios were 3.3 (95% CI 2.6-4.3) for intrauterine device and implant, 2.2 (95% CI 1.8-2.6) for pill and 1.9 (95% CI 1.5-2.4) for other methods, compared with women who did not use birth control. CONCLUSION: The absence of preconceptional folic acid supplementation for most women, even those needing consultations with healthcare professionals before pregnancy, shows that campaigns to promote folic acid supplementation should address not only women but also healthcare professionals involved in birth control and obstetric care before pregnancy.
Assuntos
Suplementos Nutricionais/estatística & dados numéricos , Ácido Fólico/uso terapêutico , Cuidado Pré-Concepcional/estatística & dados numéricos , Complexo Vitamínico B/uso terapêutico , Adulto , Comportamento Contraceptivo , Estudos Transversais , Escolaridade , Feminino , França , Inquéritos Epidemiológicos , Humanos , Análise Multivariada , Paridade , GravidezRESUMO
A double-domain activation-associated secreted protein (dd-Co-ASP) isolated from the bovine small intestinal parasite Cooperia oncophora was previously shown to be an effective vaccine candidate to protect calves against a homologous challenge infection. The aim of this study was to investigate whether the dd-Co-ASP protein, purified from a Belgian C. oncophora isolate, would offer protection against a C. oncophora isolate from the southern hemisphere as well as other Cooperia species such as C. punctata in cattle and C. curticei in sheep. Two vaccination studies were performed, i.e. one in cattle and one in sheep, in which the protective effects of dd-Co-ASP, supplemented with Quil A as an adjuvant, were compared with an adjuvant control. Whereas our results showed a 75 % reduction in Cooperia spp. cumulative faecal egg counts, the results obtained in sheep demonstrated that dd-Co-ASP was ineffective in raising a protective immune response against a C. curticei challenge infection. Even though sequence analysis of the dd-Co-ASP gene revealed restricted sequence heterogeneity in the double domain ASP within and between bovine Cooperia species, the results of the vaccine study suggest that there is sufficient conservation at the protein level to yield cross-protection, holding promise for the development of a general Cooperia vaccine for use in cattle.
Assuntos
Doenças dos Bovinos , Enteropatias Parasitárias , Infecções por Nematoides , Doenças dos Ovinos , Trichostrongyloidea , Vacinas , Animais , Bovinos , Doenças dos Bovinos/prevenção & controle , Fezes , Enteropatias Parasitárias/veterinária , Infecções por Nematoides/veterinária , Contagem de Ovos de Parasitas/veterinária , Ovinos , Doenças dos Ovinos/prevenção & controle , Vacinação/veterináriaRESUMO
In this work we analyze the variability in substitution rates in the GP63 gene from Leishmania. By using a sliding window to estimate substitution rates along the gene, we found that the rate of synonymous substitutions along the GP63 gene is highly correlated with both the rate of amino acid substitution and codon bias. Furthermore, we show that comparisons involving genes that represent independent phylogenetic lines yield very similar divergence/conservation patterns, thus suggesting that deterministic forces (i.e., nonstochastic forces such as selection) generated these patterns. We present evidence indicating that the variability in substitution rates is unambiguously related to functionally relevant features. In particular, there is a clear relationship between rates and the tertiary structure of the encoded protein since all divergent segments are located on the surface of the molecule and facing one side (almost parallel to the cell membrane) on the exposed surface of the organism. Remarkably, the protein segments encoded by these variable regions encircle the active site in a funnel-like distribution. These results strongly suggest that the pattern of nucleotide divergence and, notably, of synonymous divergence is affected by functional constraints.
Assuntos
Leishmania/genética , Metaloendopeptidases/genética , Mutação Puntual , Animais , Sítios de Ligação , Domínio Catalítico , Códon , Bases de Dados Factuais , Conversão Gênica , Metaloendopeptidases/química , Modelos Moleculares , Família Multigênica , Filogenia , Conformação ProteicaRESUMO
The recently described cysteine proteinase cathepsin W, also known as lymphopain, which is expressed specifically by CD8+ T lymphocytes, is phylogenetically related to the cruzipain-like group of the C1 family of peptidases. We have constructed sequence alignments and a theoretical three dimensional homology model of cathepsin W. These have allowed the characterization of signature features of cathepsin W in particular and the cruzipain lineage in general. The signature features are (1) an extended loop structure, Gly 170-Trp 200, in the second or beta-sheet domain; (2) an additional disulfide bond, Cys 25/Cys 60; (3) an additional "orphan" cysteine, Cys 5; (4) an additional residue. Ala 11, inserted after the first beta-sheet sheet; and (5) an S2 pocket lined with Phe 68 and Phe 230 which explains the preference for substrates containing Leu at P2. Further, the model suggested that cathepsin W could exist as a dimer with the Cys 5 of each monomer forming a disulfide bond and the Arg 40 Phe 46 loop (RISFWDF) forming part of the dimeric interface. By comparing cathepsin W with other members of the cruzipain group and with other C1 peptidases, six conserved residues were identified which appear in general to be characteristic of the cruzipain group, and which differentiate cruzipain group members from other C1 peptidases including those of the related cathepsin L lineage. The signature residues of the cruzipain lineage are (cruzipain numbering) Asn 33, Trp 38, Ala 124, Leu 127, Leu 164, and Pro 174.
Assuntos
Catepsinas/química , Catepsinas/genética , Cisteína Endopeptidases/química , Cisteína Endopeptidases/genética , Linfócitos T Citotóxicos/enzimologia , Sequência de Aminoácidos , Animais , Domínio Catalítico , Catepsina W , Catepsinas/metabolismo , Sequência Conservada , Cisteína Endopeptidases/metabolismo , Humanos , Modelos Moleculares , Dados de Sequência Molecular , Filogenia , Conformação Proteica , Proteínas de Protozoários , Homologia de Sequência de Aminoácidos , Especificidade por SubstratoRESUMO
Many parasites have deployed proteinases to accomplish some of the tasks imposed by a parasitic life style, including tissue penetration, digestion of host tissue for nutrition and evasion of host immune responses. Information on proteinases from trematodes, cestodes and nematode parasites is reviewed, concentrating on those worms of major medical and economical importance. Their biochemical characterization is discussed, along with their putative biological roles and, where available, their associated genes. For example, proteinases expressed by the various stages of the schistosome life-cycle, in particular the well-characterized cercarial elastase which is involved in the penetration of the host skin and the variety of proteinases, such as cathepsin B (Sm31), cathepsin L1, cathepsin L2, cathepsin D, cathepsin C and legumain (Sm32), which are believed to be involved in the catabolism of host haemoglobin. The various endo- and exoproteinases of Fasciola hepatica, the causative agent of liver fluke disease, are reviewed, and recent reports of how these enzymes have been successfully employed in cocktail vaccines are discussed. The various proteinases of cestodes and of the diverse superfamilies of parasitic nematodes are detailed, with special attention being given to those parasites for which most is known, including species of Taenia, Echinococcus, Spirometra, Necator, Acylostoma and Haemonchus. By far the largest number of papers in the literature and entries to the sequence data bases dealing with proteinases of parasitic helminths report on enzymes belonging to the papain superfamily of cysteine proteinases. Accordingly, the final section of the review is devoted to a phylogenetic analysis of this superfamily using over 150 published sequences. This analysis shows that the papain superfamily can be divided into two major branches. Branch A contains the cathepin Bs, the cathepsin Cs and a novel family termed cathepsin Xs, while Branch B contains the cruzipains, cathepsin Ls, papain-like and aleurain/cathepsin H-like proteinases. The relationships of the helminth proteinases, and similar proteinases from protozoan parasites and other organisms, within these groups are discussed.
Assuntos
Endopeptidases/genética , Endopeptidases/metabolismo , Genes de Helmintos , Helmintíase/parasitologia , Helmintos/genética , Sequência de Aminoácidos , Animais , Endopeptidases/química , Endopeptidases/classificação , Helmintos/enzimologia , Humanos , Dados de Sequência Molecular , FilogeniaRESUMO
Cathepsin L2 is a major cysteine proteinase secreted by adult Fasciola hepatica. The enzyme differs from other reported cathepsin Ls in that it can cleave peptide substrates that contain proline in the P2 position. A cDNA was isolated from an expression library by immunoscreening with antiserum prepared against purified native cathepsin L2. This cDNA was sequenced and shown to encode a complete preprocathepsin L proteinase. Functionally active recombinant cathepsin L proteinase was expressed and secreted by Saccharomyces cerevisiae transformed with the cDNA. The recombinant enzyme was purified from large-scale fermentation broths using ultrafiltration and gel filtration chromatography on Sephacryl S200 HR columns. NH2-terminal amino acid sequencing showed that the cleavage point for activation of the recombinant pro-enzyme is identical to that of the F. hepatica-produced cathepsin L2. The mature active recombinant proteinase behaved similarly to the native enzyme when analysed by SDS-PAGE, immunoblotting and zymography and also cleaved peptides containing proline in the P2 position. Finally, the recombinant cathepsin L2 cleaved fibrinogen to form a fibrin clot, a property we described for F. hepatica cathepsin L2.
Assuntos
Catepsinas/genética , DNA Complementar/genética , DNA de Helmintos/genética , Endopeptidases , Fasciola hepatica/enzimologia , Fasciola hepatica/genética , Animais , Sequência de Bases , Catepsina L , Catepsinas/isolamento & purificação , Catepsinas/metabolismo , Cisteína Endopeptidases , Primers do DNA/genética , DNA Complementar/isolamento & purificação , DNA de Helmintos/isolamento & purificação , Precursores Enzimáticos/genética , Fasciola hepatica/crescimento & desenvolvimento , Fibrinogênio/metabolismo , Expressão Gênica , Genes de Helmintos , Concentração de Íons de Hidrogênio , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Saccharomyces cerevisiae/genética , Especificidade por SubstratoRESUMO
The analysis of dinucleotide biases in coding and flanking regions, introns, rDNA and repetitive sequences, in the flatworm Schistosoma mansoni is reported. Except for rDNA, all regions display CpG avoidance and TpG plus CpA excess, which might be evidence of the presence of 5mC. The distribution and hierarchies of dinucleotides differ from the data published for invertebrate and vertebrate coding sequences.
Assuntos
DNA/química , Oligodesoxirribonucleotídeos/análise , Schistosoma mansoni/genética , Animais , Sequência de Bases , Dados de Sequência Molecular , Sequências Repetitivas de Ácido NucleicoRESUMO
BACKGROUND: Tuberculinic response is weaker in elderly patients. To know the real tuberculinic reactivity in elderly patients living in geriatric institutions in Catalonia, Spain, and to reduce the probability of false negatives this study was carried out to detect the booster effect. METHODS: Three hundred forty-three individuals with a mean age of 81 +/- 6.5 years, residing in four geriatric centers within the Central Health Care Region of Catalonia, were studied. After performing the first Mantoux intradermoreaction this was repeated at one week in case of negativity. A third test was carried out after one more week on persistence of negativity. Qualitative study was carried out on cell immunity in those patients whose first Mantoux test was negative. RESULTS: The first Mantoux test was positive in 163 patients (47.5%) and the performance of the second and third test in cases of negativity of the previous test increased the number of positivities by 12.5% and 4.1%, respectively, up to a total of 220 patients (64%). In the patients who did not react to the first intradermoreaction and who preserved cell immunity a booster effect was obtained in 57 (52.8%). The prevalence of tuberculous infection in this group of elderly patients calculated in the group with preserved cell immunity was 81.2% while the morbidity of tuberculous disease in the positive cases in the Mantoux test was 1.8%. CONCLUSIONS: Given the high frequency of the booster effect in elderly people, systematic testing is recommended to investigate tuberculinic reactivity in patients who are initially negative on Mantoux intradermoreaction.
Assuntos
Serviços de Saúde para Idosos , Instituição de Longa Permanência para Idosos , Casas de Saúde , Teste Tuberculínico , Tuberculose/epidemiologia , Idoso , Idoso de 80 Anos ou mais , Reações Falso-Negativas , Feminino , Humanos , Imunidade Celular , Masculino , Prevalência , Espanha/epidemiologia , Tuberculose/imunologiaRESUMO
OBJECTIVES: To synthesize the available evidence regarding the incidence, causes, risk factors of post-partum haemorrhage (PPH) and the associated maternal morbidity. METHODS: Consultation of the Medline database and of national reports on maternal mortality. RESULTS: PPH is defined as a post-partum blood loss≥500mL, and severe PPH as a post-partum blood loss≥1000mL, whatever the delivery route. In population-based studies, the incidence of PPH is around 5% of deliveries when blood loss is not precisely assessed and around 10% when it is. The incidence of severe PPH is around 2%. Uterine atony if the main cause of PPH. Maternal mortality due to obstetric haemorrhage has decreased in France (currently 1.6 death/100,000 live births) but remained the first cause of maternal death (16%) and the most avoidable (80%). In high-resource countries, PPH is the main cause of acute severe maternal morbidity, and of pregnancy-related ICU admissions. In addition to the direct consequences of acute hypovolemia, PPH exposes the women to the complications of transfusion, of intensive care and to infertility in case of hysterectomy. The main risk factors for PPH are factors of uterine atony, but they are globally poorly predictive. CONCLUSION: PPH is the principal cause of severe maternal morbidity, most often due to uterine atony. Risk factors related to components of care during labor and delivery are amenable to change, and the assessment of their risks-benefits balance should take into account the associated risk of PPH.
Assuntos
Hemorragia Pós-Parto/epidemiologia , Feminino , Humanos , Hemorragia Pós-Parto/mortalidadeAssuntos
Catepsinas/antagonistas & inibidores , Catepsinas/farmacologia , Inibidores de Cisteína Proteinase/farmacologia , Endopeptidases , Precursores Enzimáticos/farmacologia , Fasciola hepatica/enzimologia , Sequência de Aminoácidos , Animais , Catepsina B/efeitos dos fármacos , Catepsina K , Catepsina L , Catepsinas/efeitos dos fármacos , Cisteína Endopeptidases , Dados de Sequência Molecular , Papaína/efeitos dos fármacosRESUMO
The liver fluke Fasciola hepatica parasitizes humans and ruminant livestock worldwide, and it is now being considered a reemerging zoonotic disease, especially in areas in which it is endemic, such as South America. This study investigates the immune response to excretory and secretory products produced by F. hepatica in a group of patients from the Peruvian Altiplano, where the disease is highly endemic. Using a proteomic approach and immunoblotting techniques, we have identified the enzymes leucine aminopeptidase (LAP) and phosphoenolpyruvate carboxykinase as immunodominant antigens recognized by sera from fasciolosis patients. An indirect enzyme-linked immunosorbent assay using recombinant LAP as the antigen was developed to check sera from individuals of this region. Our results demonstrate that LAP produces a specific and strong reaction, suggesting its potential use in the serologic diagnosis of F. hepatica infections in humans.
Assuntos
Antígenos de Helmintos/imunologia , Fasciola hepatica/isolamento & purificação , Fasciolíase/imunologia , Fasciolíase/veterinária , Epitopos Imunodominantes/imunologia , Leucil Aminopeptidase/imunologia , Doenças dos Ovinos/parasitologia , Adolescente , Animais , Antígenos de Helmintos/genética , Antígenos de Helmintos/metabolismo , Sequência de Bases , Western Blotting/métodos , Criança , Pré-Escolar , Primers do DNA , Eletroforese em Gel Bidimensional/métodos , Ensaio de Imunoadsorção Enzimática/métodos , Fasciola hepatica/genética , Fasciola hepatica/imunologia , Fasciolíase/sangue , Fasciolíase/parasitologia , Humanos , Epitopos Imunodominantes/genética , Epitopos Imunodominantes/metabolismo , Leucil Aminopeptidase/genética , Leucil Aminopeptidase/metabolismo , Dados de Sequência Molecular , Reação em Cadeia da Polimerase/métodos , Ovinos , Doenças dos Ovinos/imunologiaRESUMO
We report the analysis of the biases of CpG, TpG, and CpA of all the DNA sequences data from the Trematode Schistosoma mansoni. Our results show CpG avoidance whereas TpG and CpA frequencies are over the expected values. These characteristics are similar to the biases displayed by methylated genomes, but in platyhelminths 5mC has not been detected by biochemical methods. The possible implications of this CpG shortage are discussed.
Assuntos
DNA/genética , Fosfatos de Dinucleosídeos/genética , Schistosoma mansoni/genética , Animais , Composição de Bases , DNA Ribossômico/genética , Íntrons , Biossíntese de Proteínas , Sequências Repetitivas de Ácido NucleicoRESUMO
BACKGROUND: Transthoracic needle aspiration (TNA) with an ultrathin needle is a safe and highly specific procedure for obtaining a diagnosis in bacterial pneumonias, but its sensitivity is at best 70%. A study was performed to assess whether Streptococcus pneumoniae and Haemophilus influenzae type b antigen detection by latex agglutination from the TNA sample enhanced the diagnostic yield. METHODS: Blood cultures, TNA with an ultrathin needle (culture, Gram stain, and latex agglutination), serological tests, and pneumococcal antigen detection in the urine by counterimmunoelectrophoresis were performed in samples from 18 of 23 consecutive patients with severe community acquired pneumonia. RESULTS: The causative organism was identified in 16 cases (88%): S pneumoniae (10 cases), S pneumoniae plus H influenzae (two cases), Legionella pneumophila (three cases), and Mycoplasma pneumoniae (one case). The investigation of antigens by latex agglutination in the pulmonary aspirate increased the diagnostic yield of TNA from 50% to 78% and provided a rapid diagnosis (in less than two hours) with therapeutic implications in seven cases. Its effectiveness was not modified by prior antibiotic therapy. CONCLUSIONS: A latex agglutination test on the pulmonary aspirate enhances the diagnostic yield of TNA in severe community acquired pneumonia.