RESUMO
Chikungunya fever is a mosquito-borne disease of key public health importance in tropical and subtropical countries. Although severe joint pain is the most distinguishing feature of chikungunya fever, diagnosis remains difficult because the symptoms of chikungunya fever are shared by many pathogens, including dengue fever. The present study aimed to develop a new immunochromatographic diagnosis test for the detection of chikungunya virus antigen in serum. Mice were immunized with isolates from patients with Thai chikungunya fever, East/Central/South African genotype, to produce mouse monoclonal antibodies against chikungunya virus. Using these monoclonal antibodies, a new diagnostic test was developed and evaluated for the detection of chikungunya virus. The newly developed diagnostic test reacted with not only the East/Central/South African genotype but also with the Asian and West African genotypes of chikungunya virus. Testing of sera from patients suspected to have chikungunya fever in Thailand (n = 50), Laos (n = 54), Indonesia (n = 2), and Senegal (n = 6) revealed sensitivity, specificity, and real-time PCR (RT-PCR) agreement values of 89.4%, 94.4%, and 91.1%, respectively. In our study using serial samples, a new diagnostic test showed high agreement with the RT-PCR within the first 5 days after onset. A rapid diagnostic test was developed using mouse monoclonal antibodies that react with chikungunya virus envelope proteins. The diagnostic accuracy of our test is clinically acceptable for chikungunya fever in the acute phase.
Assuntos
Antígenos Virais/sangue , Febre de Chikungunya/diagnóstico , Vírus Chikungunya/isolamento & purificação , Cromatografia de Afinidade/métodos , Testes Diagnósticos de Rotina/métodos , Soro/virologia , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Antivirais/imunologia , Febre de Chikungunya/virologia , Humanos , Indonésia , Camundongos Endogâmicos BALB C , Senegal , Sensibilidade e Especificidade , Tailândia , Fatores de TempoRESUMO
Warts, caused by human papillomavirus (HPV) infection, have various clinical presentations, making them difficult to differentiate from clavus, callus, and sometimes, squamous cell carcinoma. Although skin biopsies are the gold standard, a less-invasive method of examining these lesions is desired. Ninety patients with warts and related diseases, such as clavus and callus, were recruited to explore new differentiation methods using the surface of the warts. DNA was extracted from three types of specimens in each case: surface swab, shaved hyperkeratotic scale, and post-shaved surface swab. Total DNA was successfully extracted from these three specimens and was sufficient for subsequent HPV DNA detection. We analyzed samples for the HPV type and HPV viral load using polymerase chain reaction (PCR). Fifty-five cases were PCR-positive, and HPV1a, 2a, 4, 27, 57, and 65 were detected. The amount of HPV1a DNA produced was significantly greater than that of other HPV types. Regarding the correlation between the clinical diagnosis and HPV detection, the positive agreement rate was 90.9%, the negative agreement rate was 40.0%, and the overall agreement rate was 71.1%. Ten of the 21 cases clinically diagnosed as plantar warts were PCR-negative, especially in elderly patients. This suggests that it is difficult to distinguish plantar warts from clavus and callus in clinical practice. Although the amount of HPV DNA in the removed keratinization scale was highest for all HPV types, HPV detection by swabbing before and after shaving is also useful for follow-up as well as for differential diagnosis.
Assuntos
Doenças do Pé , Infecções por Papillomavirus , Neoplasias Cutâneas , Verrugas , Humanos , Idoso , Infecções por Papillomavirus/diagnóstico , Papillomavirus Humano , DNA Viral/genética , Verrugas/diagnóstico , Papillomaviridae/genéticaRESUMO
BACKGROUND: For the diagnosis of seasonal influenza, clinicians rely on point-of-care testing (POCT) using commercially available kits developed against seasonal influenza viruses. However, POCT has not yet been established for the diagnosis of pandemic influenza A virus (H1N1pdm) infection due to the low sensitivity of the existing kits for H1N1pdm. METHODOLOGY/PRINCIPAL FINDINGS: An immunochromatography (IC) test kit was developed based on a monoclonal antibody against H1N1pdm, which does not cross-react with seasonal influenza A or B viruses. The efficacy of this kit (PDM-IC kit) for the diagnosis of H1N1pdm infection was compared with that of an existing kit for the detection of seasonal influenza viruses (SEA-IC kit). Nasal swabs (nâ=â542) were obtained from patients with flu-like syndrome at 13 clinics in Osaka, Japan during the winter of 2010/2011. Among the 542 samples, randomly selected 332 were further evaluated for viral presence by reverse transcriptase polymerase chain reaction (RT-PCR). The PDM-IC kit versus the SEA-IC kit showed higher sensitivity to and specificity for H1N1pdm, despite several inconsistencies between the two kits or between the kits and RT-PCR. Consequently, greater numbers of false-negative and false-positive cases were documented when the SEA-IC kit was employed. Significant correlation coefficients for sensitivity, specificity, and negative prediction values between the two kits were observed at individual clinics, indicating that the results could be affected by clinic-related techniques for sampling and kit handling. Importantly, many patients (especially influenza-negative cases) were prescribed anti-influenza drugs that were incongruous with their condition, largely due to physician preference for patient responses to questionnaires and patient symptomology, as opposed to actual viral presence. CONCLUSIONS/SIGNIFICANCE: Concomitant use of SEA-IC and PDM-IC kits increased the likelihood of correct influenza diagnosis. Increasing the credibility of POCT is anticipated to decrease the inappropriate dispensing of anti-influenza drugs, thereby minimizing the emergence of drug-resistant H1N1pdm strains.
Assuntos
Antivirais/uso terapêutico , Cromatografia de Afinidade/métodos , Vírus da Influenza A Subtipo H1N1/isolamento & purificação , Vírus da Influenza A Subtipo H1N1/fisiologia , Influenza Humana/diagnóstico , Influenza Humana/epidemiologia , Pandemias , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Antivirais/administração & dosagem , Antivirais/farmacologia , Criança , Pré-Escolar , Cães , Prescrições de Medicamentos/estatística & dados numéricos , Feminino , Humanos , Lactente , Recém-Nascido , Vírus da Influenza A Subtipo H1N1/efeitos dos fármacos , Vírus da Influenza A Subtipo H1N1/genética , Influenza Humana/tratamento farmacológico , Células Madin Darby de Rim Canino , Masculino , Pessoa de Meia-Idade , Sistemas Automatizados de Assistência Junto ao Leito , Valor Preditivo dos Testes , Reprodutibilidade dos Testes , Fatores de Tempo , Adulto JovemRESUMO
N¹,N¹²-diacetylspermine (DiAcSpm) is a minor component of human urine that constitutes less than 0.5% of total polyamine species in human urine. Structurally related polyamines and acetylpolyamines were separated and analyzed by HPLC and gas chromatography, and refinement of these procedures led to the identification of this minor component. Subsequent analyses of urines from cancer patients as well as healthy persons revealed that DiAcSpm is a promising candidate for a novel tumor marker. It is much more sensitive than established tumor markers in detecting colorectal and other cancers, and most importantly, is able to detect 60% of early colorectal cancers confined to mucous membranes. Serum CEA is able to detect only about 10% of colorectal cancers at this stage. Collection of urine is easy and does not give any pain to patients, which adds another merit to urinary DiAcSpm as a tumor marker. DiAcSpm-specific antibodies were then developed for simpler determination of DiAcSpm in urine, and the antibodies were used to construct an ELISA system. More recently, a reagent kit for DiAcSpm determination based on colloidal gold aggregation that can be used with automatic biochemical analyzers was also developed.
Assuntos
Bioquímica/métodos , Biomarcadores Tumorais/urina , Espermina/análogos & derivados , Animais , Biomarcadores Tumorais/sangue , Antígeno Carcinoembrionário/sangue , Bovinos , Cromatografia Líquida de Alta Pressão , Coloides , Neoplasias do Colo/sangue , Ensaio de Imunoadsorção Enzimática , Ouro , Humanos , Peptídeos/metabolismo , Sensibilidade e Especificidade , Manejo de Espécimes , Espermina/metabolismo , Espermina/urinaRESUMO
Since its emergence in April 2009, pandemic influenza A virus H1N1 (H1N1 pdm), a new type of influenza A virus with a triple-reassortant genome, has spread throughout the world. Initial attempts to diagnose the infection in patients using immunochromatography (IC) relied on test kits developed for seasonal influenza A and B viruses, many of which proved significantly less sensitive to H1N1 pdm. Here, we prepared monoclonal antibodies that react with H1N1 pdm but not seasonal influenza A (H1N1 and H3N2) or B viruses. Using two of these antibodies, one recognizing viral hemagglutinin (HA) and the other recognizing nucleoprotein (NP), we developed kits for the specific detection of H1N1 pdm and tested them using clinical specimens of nasal wash fluid or nasopharyngeal fluid from patients with influenza-like illnesses. The specificities of both IC test kits were very high (93% for the HA kit, 100% for the NP kit). The test sensitivities for detection of H1N1 pdm were 85.5% with the anti-NP antibody, 49.4% with the anti-HA antibody, and 79.5% with a commercially available influenza A virus detection assay. Use of the anti-NP antibody could allow the rapid and accurate diagnosis of H1N1 pdm infections.
Assuntos
Técnicas de Laboratório Clínico/métodos , Glicoproteínas de Hemaglutininação de Vírus da Influenza/análise , Vírus da Influenza A Subtipo H1N1/isolamento & purificação , Influenza Humana/diagnóstico , Proteínas de Ligação a RNA/análise , Kit de Reagentes para Diagnóstico , Proteínas do Core Viral/análise , Animais , Anticorpos Monoclonais/isolamento & purificação , Anticorpos Antivirais/isolamento & purificação , Feminino , Glicoproteínas de Hemaglutininação de Vírus da Influenza/imunologia , Humanos , Imunoensaio/métodos , Influenza Humana/virologia , Camundongos , Camundongos Endogâmicos BALB C , Mucosa Nasal/virologia , Nasofaringe/virologia , Proteínas do Nucleocapsídeo , Proteínas de Ligação a RNA/isolamento & purificação , Sensibilidade e Especificidade , Fatores de Tempo , Proteínas do Core Viral/isolamento & purificaçãoRESUMO
BACKGROUND: Colonoscopic examination is the common pathway for positive screening tests detecting colorectal lesions. We evaluated a specific, quantitative high-throughput automatic immunochemical fecal occult blood test (Auto iFOBT) method for colorectal cancer (CRC) screening and to determine its concordance with physician assessments informed by complete colonoscopy, the gold-standard technique for evaluation of the colonic mucosa. METHODS: 1200 CRC symptomatic patients were recruited for a retrospective investigation. Colorectal neoplasia were localized by colonoscopy and cancer outcomes were enumerated according to severity. In addition, stool samples were collected and analyzed by Auto iFOBT to derive sensitivity, specificity, and positive predictive value. Qualitative colonoscopy and Auto iFOBT results were correlated, as were cancer severities and quantitative hemoglobin concentrations. RESULTS: Ninety-one patients were found positive for CRC; 50 mucosal, 20 submucosal, and 21 advanced. At standard cutoff, sensitivity was 60%, 90%, and 95%, respectively. Specificity and positive predictive value for all neoplasia and cancers were 89.6% and 86.4%, and 60.9% and 33.7%, respectively. Cancer severities could be approximated roughly according to hemoglobin concentrations. CONCLUSIONS: Specific qualitative 2-day Auto iFOBT is an accurate tool for the detection of colorectal cancer and therefore provides the basis for a large-scale screening program.