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OBJECTIVES: This study aimed to investigate the utility of the RAND/UCLA appropriateness method (RAM) in validating expert consensus-based multiple-choice questions (MCQs) on electrocardiogram (ECG). METHODS: According to the RAM user's manual, nine panelists comprising various experts who routinely handle ECGs were asked to reach a consensus in three phases: a preparatory phase (round 0), an online test phase (round 1), and a face-to-face expert panel meeting (round 2). In round 0, the objectives and future timeline of the study were elucidated to the nine expert panelists with a summary of relevant literature. In round 1, 100 ECG questions prepared by two skilled cardiologists were answered, and the success rate was calculated by dividing the number of correct answers by 9. Furthermore, the questions were stratified into "Appropriate," "Discussion," or "Inappropriate" according to the median score and interquartile range (IQR) of appropriateness rating by nine panelists. In round 2, the validity of the 100 ECG questions was discussed in an expert panel meeting according to the results of round 1 and finally reassessed as "Appropriate," "Candidate," "Revision," and "Defer." RESULTS: In round 1 results, the average success rate of the nine experts was 0.89. Using the median score and IQR, 54 questions were classified as " Discussion." In the expert panel meeting in round 2, 23% of the original 100 questions was ultimately deemed inappropriate, although they had been prepared by two skilled cardiologists. Most of the 46 questions categorized as "Appropriate" using the median score and IQR in round 1 were considered "Appropriate" even after round 2 (44/46, 95.7%). CONCLUSIONS: The use of the median score and IQR allowed for a more objective determination of question validity. The RAM may help select appropriate questions, contributing to the preparation of higher-quality tests.
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Eletrocardiografia , Humanos , Consenso , Reprodutibilidade dos Testes , Competência Clínica/normas , Avaliação Educacional/métodos , Cardiologia/normasRESUMO
BACKGROUND: The prevalence of Short Birth Interval (SBI) is higher in Low- and Middle-Income countries (LMICs), including Bangladesh. Previous studies in LMICs have estimated the effects of SBI on child mortality by comparing two unequal groups of mothers based on their socio-economic status. This approach may lead to overestimation or underestimation of the true effect of birth interval on child mortality, particularly when sample sizes are relatively small. OBJECTIVE: We determined the effects of SBI on several forms of child mortality in Bangladesh by comparing two equal groups created by applying the propensity score matching technique. METHODS: This study analyzed data from 5,941 mothers and 1,594 health facilities extracted from the 2017/18 Bangladesh Demographic and Health Survey and the 2017 Bangladesh Health Facility Survey. The exposure variable was SBI (defined as the interval between two subsequent births <33 months: yes, no), while the outcome variables were neonatal mortality (defined as mortality within 28 days of birth: yes, no), infant mortality (defined as mortality within 1 year of birth: yes, no), and under-five mortality (defined as mortality within 5 years of birth: yes, no). Multilevel Poisson regression based on inverse probability treatment weights was used to determine the association between exposure and outcome variables. RESULTS: The prevalence rates of neonatal, infant, and under-five mortality were 48.8, 30.8, and 23.1 per 1000 live births, respectively. Newborns of SBI mothers were found to have a 63% higher likelihood of neonatal mortality (aPR, 1.63; 95% CI, 1.08-2.46) compared to newborns of non-SBI mothers. Furthermore, the prevalence of infant mortality and under-five mortality was 1.45 times higher (aPR, 1.45; 95% CI, 1.01-2.08) and 2.82 times higher (aPR, 2.82; 95% CI, 2.16-3.70), respectively, among babies born in a short interval of their immediately preceding sibling as compared to babies born in a normal interval of their immediately preceding sibling. CONCLUSIONS: Findings of this study indicate that SBI is an important predictor of child mortality. Consequently, around 1 million children born in a short interval every year in Bangladesh are at risk of dying before reaching their fifth birthday. This indicates a challenge for Bangladesh to achieve the SDG 3 target to reduce neonatal and under-five mortality to 12 and 25 deaths per 1000 live births, respectively. Hence, awareness-building programs about the adverse effects of SBI and strengthening existing healthcare facilities are important.
Assuntos
Intervalo entre Nascimentos , Mortalidade da Criança , Lactente , Criança , Feminino , Humanos , Recém-Nascido , Pontuação de Propensão , Bangladesh/epidemiologia , Mortalidade Infantil , Fatores SocioeconômicosRESUMO
Bacteriophage T4 is decorated with 155 180 Å-long fibers of the highly antigenic outer capsid protein (Hoc). In this study, we describe a near-atomic structural model of Hoc by combining cryo-electron microscopy and AlphaFold structure predictions. It consists of a conserved C-terminal capsid-binding domain attached to a string of three variable immunoglobulin (Ig)-like domains, an architecture well-preserved in hundreds of Hoc molecules found in phage genomes. Each T4-Hoc fiber attaches randomly to the center of gp23* hexameric capsomers in one of the six possible orientations, though at the vertex-proximal hexamers that deviate from 6-fold symmetry, Hoc binds in two preferred orientations related by 180° rotation. Remarkably, each Hoc fiber binds to all six subunits of the capsomer, though the interactions are greatest with three of the subunits, resulting in the off-centered attachment of the C-domain. Biochemical analyses suggest that the acidic Hoc fiber (pI, ~4-5) allows for the clustering of virions in acidic pH and dispersion in neutral/alkaline pH. Hoc appears to have evolved as a sensing device that allows the phage to navigate its movements through reversible clustering-dispersion transitions so that it reaches its destination, the host bacterium, and persists in various ecological niches such as the human/mammalian gut.
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Bacteriófagos , Animais , Humanos , Bacteriófagos/genética , Bacteriófagos/metabolismo , Microscopia Crioeletrônica/métodos , Proteínas do Capsídeo/química , Capsídeo/metabolismo , Bacteriófago T4/genética , Bacteriófago T4/química , Ligação Proteica , MamíferosRESUMO
The Sustainable Development Goals 3 targets significant reductions in maternal and under-five deaths by 2030. The prevalence of these deaths is significantly associated with short birth intervals (SBI). Identification of factors associated with SBI is pivotal for intervening with appropriate programmes to reduce occurrence of SBI and associated adverse consequences. This study aimed to determine the factors associated with SBI in Bangladesh. A total of 5,941 women included in the 2017/18 Bangladesh Demographic and Health Survey 2017/18 and 1,524 healthcare facilities included in the 2017 Bangladesh Health Facility were linked and analysed. The sample was selected based on the availability of the birth interval data between the two most recent subsequent live birth. SBI was defined as an interval between consecutive births of 33 months or less, as recommended by the World Health Organization and was the outcome variable. Several individual-, households-, and community-level factors were considered as exposure variables. We used descriptive statistics to summarise respondents' characteristics and multilevel Poisson regression to assess the association between the outcome variable with exposure variables. Around 26% of live births occurred in short intervals, with a further higher prevalence among younger, uneducated, or rural women. The likelihoods of SBI were lower among women aged 20-34 years (PR, 0.14; 95% CI, 0.11-0.17) and ≥35 years (PR, 0.03; 95% CI, 0.02-0.05) as compared to the women aged 19 years or less. Women from households with the richest wealth quintile experienced lower odds of SBI (PR, 0.61; 95% CI, 0.45-0.85) compared to those from the poorest wealth quintile. The prevalences of SBI were higher among women whose second most recent child died (PR, 5.23; 95% CI, 4.18-6.55), those who were living in Chattogram (PR, 1.52; 95% CI, 1.12-2.07) or Sylhet (PR, 2.83, 95% CI, 2.08-3.86) divisions. Availability of modern contraceptives at the nearest healthcare facilities was 66% protective to the occurrence of SBI (PR, 0.34; 95% CI, 0.22-0.78). Also, the prevalence of SBI increased around 85% (PR, 1.85; 95% CI, 1.33-2.18) for every kilometer increase in the distance of nearby health facilities from women's homes. Targeted and tailored regional policies and programmes are needed to increase the awareness of SBI and associated adverse health outcomes and availability of modern contraception in the healthcare facilities.
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Short Birth Interval (SBI, defined as < 33 months interval between the two most recent births or < 24 months between one live birth to the next pregnancy) is a public health problem in most low- and lower-middle-income countries. Understanding geographic variations in SBI, particularly SBI hot spots and associated factors, may help intervene with tailored programs. This study identified the geographical hot spots of SBI in Bangladesh and the factors associated with them. We analyzed women's data extracted from the 2017/18 Bangladesh Demographic and Health Survey and the healthcare facility data extracted from the 2017 Service Provision Assessment. SBI was the outcome variable, and it was defined as an interval between consecutive births of 33 months or less, as recommended by the World Health Organization. The characteristics of mothers and their partners were the explanatory variables. Moran's I was used to examine the spatial variation of SBI in Bangladesh whereas the Getis-Ord [Formula: see text](d) was used to determine the hot spots of SBI. The Geographical Weighted Regression (GWR) was used to assess the predictors of SBI at the enumeration areas' level. The variables included in the GWR were selected using the exploratory regression and ordinary least square regression model. Data of 5941 women were included in the analyses. Around 26% of the total births in Bangladesh had occurred in short intervals. A majority of the SBI hot spots were found in the Sylhet division, and almost all SBI cold spots were in the Rajshahi and Khulna divisions. No engagement with formal income-generating activities, high maternal parity, and history of experiencing the death of a child were significantly associated with SBI in the Sylhet division. Women's age of 34 years or less at the first birth was a protective factor of SBI in the Rajshahi and Khulna divisions. The prevalence of SBI in Bangladesh is highly clustered in the Sylhet division. We recommend introducing tailored reproductive health care services in the hot spots instead of the existing uniform approach across the country.
Assuntos
Intervalo entre Nascimentos , Mães , Adulto , Bangladesh/epidemiologia , Criança , Feminino , Instalações de Saúde , Humanos , Gravidez , Fatores Socioeconômicos , Inquéritos e QuestionáriosRESUMO
Methods: Eight databases, PubMed, CINAHL, Web of Science, Embase, PsycINFO, Cochrane Library, Popline, and Maternity and Infant Care, were searched, covering the period of January 2000 to January 2022. Studies that had examined the association between SBI and any form of child mortality were included. The findings of the included studies were summarized through fixed-effects or random-effects meta-analysis and the model was selected based on the heterogeneity index. Results: A total of 51 studies were included. Of them, 19 were conducted in Ethiopia, 10 in Nigeria and 7 in Bangladesh. Significant higher likelihoods of stillbirth (odds ratio (OR) = 2.11; 95% confidence interval (CI) = 1.32-3.38), early neonatal mortality (OR = 1.58; 95% CI = 1.04-2.41), perinatal mortality (OR = 1.71; 95% CI = 1.32-2.21), neonatal mortality (OR = 1.85; 95% CI = 1.68-2.04), post-neonatal mortality (OR = 3.01; 95% CI = 1.43-6.33), infant mortality (OR = 1.92; 95% CI = 1.77-2.07), child mortality (OR = 1.67; 95% CI = 1.27-2.19) and under-five mortality (OR = 1.95; 95% CI = 1.56-2.44) were found among babies born in short birth intervals than those who born in normal intervals. Conclusions: SBI significantly increases the risk of child mortality in LMICs. Programmes to reduce pregnancies in short intervals need to be expanded and strengthened. Reproductive health interventions aimed at reducing child mortality should include proper counselling on family planning, distribution of appropriate contraceptives and increased awareness of the adverse effects of SBI on maternal and child health.
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Intervalo entre Nascimentos , Mortalidade da Criança , Criança , Países em Desenvolvimento , Feminino , Humanos , Lactente , Mortalidade Infantil , Recém-Nascido , Mortalidade Perinatal , GravidezRESUMO
Prognostic molecular subgrouping of glioblastoma is an ongoing effort and the current classification includes IDH-wild-type and IDH-mutant entities, the latter showing significantly better prognosis. We performed a comparative integrated analysis of the FGFR glioblastoma subgroup consisting of 5 cases from a prospective 101-patient-cohort. FGFR alterations included FGFR2-TACC2 and FGFR2 amplifications arising in a multifocal IDH-mutant glioblastoma with unexpected 2.5-month patient survival, novel FGFR3 carboxy-terminal duplication and FGFR3-TLN1 fusion, and two previously described FGFR3-TACC3 fusions. The FGFR2 tumors showed additional mutations in SERPINE1/PAI-1 and MMP16, as part of extensive extracellular matrix remodeling programs. Whole transcriptomic analysis revealed common proliferation but distinct morphogenetic gene expression programs that correlated with tumor histology. The kinase program revealed EPHA3, LTK and ALK receptor tyrosine kinase overexpression in individual FGFR tumors. Paradoxically, all FGFR-fused glioblastomas shared strong PI3K and MAPK pathway suppression effected by SPRY, DUSP and AKAP12 inhibitors, whereas the FGFR2-TACC2 tumor elicited also EGFR suppression by ERRFI1 upregulation. This integrated analysis outlined the proliferation and morphogenetic expression programs in FGFR glioblastoma, and identified four novel, clinically targetable FGFR2 and FGFR3 alterations that confer aggressive phenotype and trigger canonical pathway feedback inhibition, with important therapeutic implications.
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Neoplasias Encefálicas/genética , Glioblastoma/genética , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/genética , Receptor Tipo 3 de Fator de Crescimento de Fibroblastos/genética , Idoso , Neoplasias Encefálicas/patologia , Feminino , Amplificação de Genes , Glioblastoma/patologia , Humanos , Masculino , Pessoa de Meia-Idade , Proteínas de Fusão Oncogênica , Fenótipo , Prognóstico , Estudos Prospectivos , TranscriptomaRESUMO
Epithelioid glioblastoma is a recognized glioblastoma variant, recently added to the World Health Organization brain tumor classification, with similar prognosis as the classic variant and B-Raf V600E mutations in 50% of the cases. We identified a new subset of epithelioid glioblastoma with periventricular location and subependymal giant cell astrocytoma (SEGA)-like morphology. Genomic profiling of these tumors revealed driver mutations in NF1, subclonal mutations in TSC1, and a novel driver mutation in MTOR, suggesting upregulation of the MAPK/TSC1/mTOR pathway. Strong mTOR activation was confirmed by immunohistochemistry for the mTOR kinase target 4E-BP1. TSC1 and MTOR mutations have been previously described in low-grade glioma, such as SEGA, and focal cortical dysplasia, respectively, that display large cells with abundant cytoplasm, most likely resulting from the biogenetic signaling of mTOR. Unlike these, the mutations in SEGA-like glioblastoma occurred in the context of other genetic aberrations present in high-grade neoplasms, including in the CDKN2A/B, PIK3R1, PIK3CA and EGFR genes. For one patient with two temporally distinct specimens, the subclonal TSC1 pathogenic mutation was detected only in the specimen showing SEGA-like morphology, indicating requirement for mTOR activation as trigger for specific epithelioid/SEGA-like morphology. As FDA-approved kinase inhibitors are available and target many steps of the MAPK/mTOR pathway, recognition of this new subset of periventricular high-grade gliomas with clear phenotypic-genotypic correlates is essential for prompt biomarker testing and appropriate targeted therapeutic management of these patients.
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A sensitive and specific IgG4 enzyme-linked immunosorbent assay (ELISA) with urine samples has been reported. To confirm elimination of bancroftian filariasis, the ELISA was used in a study conducted in Yongjia County and Gaoan City, People's Republic of China, where filariasis elimination was declared, with 10,409 students 5-16 years of age. The antibody positive rates were 0.08% in Yongjia and 0.34% in Gaoan. All positive samples were re-examined and found to be negative. Our results show that this ELISA is practical and useful for confirmation of the elimination of filariasis. If similar results are obtained in different filariasis-endemic countries, this method may be useful in global filariasis elimination programs.
Assuntos
Brugia pahangi/isolamento & purificação , Filariose Linfática/urina , Ensaio de Imunoadsorção Enzimática/métodos , Imunoglobulina G/urina , Adolescente , Animais , Criança , Pré-Escolar , China/epidemiologia , Filariose Linfática/epidemiologia , Filariose Linfática/parasitologia , Feminino , Humanos , Masculino , Estudos SoroepidemiológicosRESUMO
To detect IgG antibody in the serodiagnosis of visceral leishmaniasis (VL), a recombinant antigen rK39, which is part of a Leishmania chagasi kinesin-related protein, has been used successfully and showed high sensitivity and specificity. We report production of a recombinant protein rKRP42, which is part of an L. donovani kinesin-related protein and a homolog of rK39, and its application in an enzyme-linked immunosorbent assay (ELISA) for the diagnosis of VL. When rKRP42 and rK39 were compared, amino acid sequence analysis showed 89.3% identity and 98.7% homology, with rKRP42 having 39 more amino acids than rK39. The ELISA using rKRP42 showed a sensitivity of 94.6% (70 positive samples among 74 from VL patients) and a specificity of 99.3% (148 negative samples among 149 samples from Japanese controls), whereas the sensitivity of the commercial rK39 dipstick test was 93.2% (69 positive samples among 74 from patients with VL). The rKRP42 is a promising new antigen in developing immunodiagnostic methods for VL.
Assuntos
Anticorpos Antiprotozoários/sangue , Cinesinas/genética , Leishmania donovani/genética , Leishmaniose Visceral/diagnóstico , Proteínas de Protozoários/genética , Proteínas Recombinantes/genética , Sequência de Aminoácidos , Animais , Antígenos de Protozoários/química , Antígenos de Protozoários/genética , Antígenos de Protozoários/imunologia , Ensaio de Imunoadsorção Enzimática/métodos , Genes de Protozoários/genética , Humanos , Cinesinas/química , Cinesinas/imunologia , Leishmania donovani/imunologia , Proteínas de Protozoários/química , Proteínas de Protozoários/imunologia , Proteínas Recombinantes/química , Proteínas Recombinantes/imunologia , Sensibilidade e Especificidade , Alinhamento de Sequência , Homologia de Sequência de AminoácidosRESUMO
The large intestine of a rat has been neglected almost completely as a site of Strongyloides sp. infection. We reported that adult Strongyloides ratti remained in the large intestine for more than 80 days, producing more number of infective larvae than small intestine adults, and therefore hypothesized that parasitism in this site could be a survival strategy. In wild rats, however, no study has focused on large intestine infections of Strongyloides. The present study revealed that 32.4% of 68 wild rats, Rattus norvegicus, had the infection of S. ratti in the large intestine, with an average of 4.7 worms. These worms harbored normal eggs in the uterus. In a laboratory experiment with S. ratti and Wister rats, daily output of infective larvae by 4.7 females in the large intestine was estimated to be 4,638.4, suggesting that a few parasites could play a role in the parasite transmission. Five species of nematode found in the wild rats showed seasonality in infection intensity, with highest intensities in March-May. The number of S. ratti in the large intestine was also highest in these months.
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Enteropatias Parasitárias/veterinária , Intestino Grosso/parasitologia , Ratos/parasitologia , Doenças dos Roedores/parasitologia , Strongyloides ratti/isolamento & purificação , Estrongiloidíase/veterinária , Animais , Animais Selvagens , Ascaridídios/isolamento & purificação , Infecções por Ascaridida/epidemiologia , Infecções por Ascaridida/parasitologia , Infecções por Ascaridida/veterinária , Feminino , Enteropatias Parasitárias/epidemiologia , Enteropatias Parasitárias/parasitologia , Japão/epidemiologia , Masculino , Nippostrongylus/isolamento & purificação , Prevalência , Doenças dos Roedores/epidemiologia , Estações do Ano , Infecções por Strongylida/epidemiologia , Infecções por Strongylida/parasitologia , Infecções por Strongylida/veterinária , Estrongiloidíase/epidemiologia , Estrongiloidíase/parasitologiaRESUMO
A new direct agglutination test (DAT) for use with urine samples for the diagnosis of visceral leishmaniasis (VL) has been developed and compared with the conventional DAT with serum samples and our previously reported enzyme-linked immunosorbent assay (ELISA) with urine samples (urine ELISA). The new DAT, in which anti-human IgG was used as enhancing antibody, was tested with urine samples from 75 VL patients and 225 non-VL patients and healthy people. The sensitivity of the new DAT (90.7%), was almost the same as that of the conventional DAT (91.0%) and the urine ELISA (93.3%). The specificity of the new DAT (96.4%) was nearly identical with that of the urine ELISA (97.3%). A urine-based DAT has several advantages over the conventional DAT: sample collection is non-invasive and it can process larger numbers of samples with smaller amounts of antigen.
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Testes de Aglutinação/métodos , Anticorpos Antiprotozoários/urina , Leishmania donovani/isolamento & purificação , Leishmaniose Visceral/urina , Adulto , Animais , Antígenos de Protozoários , Ensaio de Imunoadsorção Enzimática , Humanos , Imunoglobulina G , Leishmaniose Visceral/sangue , Leishmaniose Visceral/parasitologia , Sensibilidade e EspecificidadeRESUMO
Sucrose synthase (Sus) (EC 2.4.1.13) is a key enzyme for the sugar accumulation that is critical to form fruit quality. In this study, extensive data-mining and PCR amplification confirmed that there are at least six Sus genes (CitSus1-6) in the citrus genome. Gene structure and phylogeny analysis showed an evolutionary consistency with other plant species. The six Sus genes contain 12-15 exons and 11-14 introns and were evenly distributed into the three plant Sus groups (CitSus1 and CitSus2 in the Sus I group, CitSus3 and CitSus6 in the Sus II group, and CitSus4 and CitSus5 in the Sus III group). Transcripts of these six CitSus genes were subsequently examined. For tissues and organs, CitSus1 and 2 were predominantly expressed in fruit juice sacs (JS) whereas CitSus3 and 4 were predominantly expressed in early leaves (immature leaves), and CitSus5 and 6 were predominantly expressed in fruit JS and in mature leaves. During fruit development, CitSus5 transcript increased significantly and CitSus6 transcript decreased significantly in fruit JS. In the fruit segment membrane (SM), the transcript levels of CitSus2 and 5 were markedly higher and the abundant levels of CitSus3 and 6 gradually decreased. Moreover, transcript levels of CitSus1-4 examined were higher and the CitSus5 transcript level was lower in the fruit SM than in fruit JS, while CitSus6 had a similar transcript level in fruit JS and SM. In addition, transcripts of CitSus1-6 responded differently to dehydration in mature leaves or to mild drought stress in fruit JS and SM. Finally, the possible roles of Sus genes in the regulation of sugar accumulation are discussed; however, further study is required.
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Citrus/genética , Genoma de Planta/genética , Glucosiltransferases/genética , Proteínas de Plantas/genética , Transcriptoma , Metabolismo dos Carboidratos/genética , Citrus/enzimologia , Secas , Éxons/genética , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Glucosiltransferases/classificação , Glucosiltransferases/metabolismo , Íntrons/genética , Isoenzimas/classificação , Isoenzimas/genética , Isoenzimas/metabolismo , Filogenia , Proteínas de Plantas/classificação , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sacarose/metabolismo , Água/metabolismo , Água/farmacologiaRESUMO
We reported a highly sensitive and specific enzyme-linked immunosorbent assay (ELISA) that detects immunoglobulin G (IgG) in urine using rKRP42 antigen for the diagnosis of visceral leishmaniasis (VL). The ELISA was applied to study chronological change in antibody titers in five study areas in Rajshahi district, Bangladesh. A total of 585 subjects without a past VL history were examined at least three times in the 30-month follow-up period; of these subjects, 137 (23.4%) subjects became ELISA-positive at least one time during the study. Among the positive cases, 40 (29.2%) subjects developed clinical VL, and 31 (77.5%) of these subjects showed IgG titers of ≥ 1,000 U more than one time in the study period. Considering only the first ELISA results, 22 subjects with IgG titers of ≥ 1,000 U could be found, and 21 (95.5%) of these subjects turned out to be clinical cases. The high urinary IgG titers (≥ 1,000 U) will help predict possible clinical VL cases and thus, identify an outbreak in its earlier stage.
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Antígenos de Protozoários , Surtos de Doenças , Ensaio de Imunoadsorção Enzimática/métodos , Imunoglobulina G/urina , Leishmania donovani/imunologia , Leishmaniose Visceral/diagnóstico , Proteínas Recombinantes , Anticorpos Antiprotozoários/urina , Antígenos de Protozoários/genética , Antígenos de Protozoários/imunologia , Bangladesh/epidemiologia , Feminino , Humanos , Imunoglobulina G/imunologia , Leishmania donovani/genética , Leishmaniose Visceral/epidemiologia , Leishmaniose Visceral/parasitologia , Leishmaniose Visceral/urina , Masculino , Valor Preditivo dos Testes , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Sensibilidade e EspecificidadeRESUMO
This study was performed to isolate actinomycete colonies having antibacterial activity from soil samples collected from different places around Rajshahi, Bangladesh. Thirty actinomycete colonies were isolated in pure culture from five soil samples using Starch-casein-nitrate-agar medium. The isolates were grouped in five color series based on their aerial mycelia color and screened for their antibacterial activity against a range of test bacteria. Sixteen isolates (53.3%) were found to have moderate to high activity against four gram-positive and four gram-negative bacteria. Since many isolates showed inhibitory activity against indicator bacteria, it is suggestive that Bangladeshi soil could be an interesting source to explore for antibacterial secondary metabolites.
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We have applied a loop-mediated isothermal amplification (LAMP) technique to detect Leishmania donovani DNA. The LAMP technique detected 1 fg of L. donovani DNA, which was 10-fold more sensitive than a conventional polymerase chain reaction (PCR). All nested PCR-positive blood samples from visceral leishmaniasis patients were positive with the LAMP technique, and DNA samples from L. infantum, L. major, L. mexicana, L. tropica, L. braziliensis, Plasmodium falciparum, and healthy humans were negative with the LAMP technique. The advantages of the LAMP method are its shorter reaction time, a lack of requirement of sophisticated equipment, and visual judgment of positivity based on the turbidity of reaction mixture. Our LAMP technique can be a better alternative to a conventional PCR, especially under field conditions.
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DNA de Protozoário/genética , Leishmania donovani/genética , Técnicas de Amplificação de Ácido Nucleico/métodos , Animais , Humanos , Sensibilidade e EspecificidadeRESUMO
We recently reported the production of the recombinant kinesin-related protein of Leishmania donovani with a molecular weight of 42 kd (rKRP42) and the value of the antigen in serum-based ELISA for the diagnosis of visceral leishmaniasis (VL). In this study, the rKRP42 antigen was validated with ELISA using urine samples (rKRP42 urine ELISA). The urine-based ELISA showed 94% sensitivity (108 positives among 115 VL samples) and 99.6% specificity (239 negatives among 240 non-VL samples). The sensitivity and specificity are almost similar to our previous results by ELISA with acetone-treated L. donovani promastigote antigen and direct agglutination test, both methods being done by use of urine samples. A comparison of the rKRP42 urine ELISA with the commercially available urinary antigen detection kit (KAtex) using 108 VL samples showed much higher sensitivity of the ELISA (96.3%) than KAtex (55.6%). The use of the rKRP42 antigen with urine samples will facilitate epidemiologic studies.
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Anticorpos Antiprotozoários/urina , Antígenos de Protozoários/imunologia , Ensaio de Imunoadsorção Enzimática/métodos , Leishmania donovani/imunologia , Leishmaniose Visceral/diagnóstico , Animais , Humanos , Imunoglobulina G/urina , Kit de Reagentes para Diagnóstico , Proteínas Recombinantes/imunologia , Sensibilidade e Especificidade , Manejo de EspécimesRESUMO
A diagnostic method has been developed to detect anti-Leishmania donovani immunoglobulin G (IgG) in urine by enzyme-linked immunosorbent assay (ELISA). In measuring anti-L. donovani IgG, IgA, and IgM in urine, the method performed best in the detection of IgG. The sensitivity and specificity of the assay were determined with panels of urine samples from 62 visceral leishmaniasis (VL) patients, 59 healthy controls from areas of endemicity, 53 healthy controls from areas of nonendemicity, 59 malaria patients, 13 tuberculosis patients, 23 cutaneous leishmaniasis patients, and 7 patients with other diseases. Using L. donovani promastigote crude antigen, the test had 93.5% sensitivity (58 positives of 62 VL patient samples) and 89.3% specificity (191 negatives of 214 non-VL patient samples). The ELISA with acetone-treated L. donovani promastigote antigen raised the sensitivity and specificity to 95.0 and 95.3%, respectively. Western blot analysis revealed that most of the samples that cross-reacted with crude antigen in ELISA did not recognize any antigenic component of L. donovani crude antigen. We also checked 40 serum samples from the same group of VL patients for anti-L. donovani IgG and got 90.0% sensitivity with both crude and acetone-treated antigens. As collection of urine is much easier than collection of serum, the detection of anti-L. donovani IgG in urine with acetone-treated antigen will be useful in epidemiological studies. It could be an adjunct of laboratory diagnosis.