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Biomedical subjects

S Hoshino-Shimizu

Publications and source records attributed to S Hoshino-Shimizu.

At least 19 recordsLinked to original sources
PubMed2003

Helminth antigens (Taenia solium, Taenia crassiceps, Toxocara canis, Schistosoma mansoni and Echinococcus granulosus) and cross-reactivities in human infections and immunized animals.

Helminth antigens were investigated in the search for accessible heterologous antigens capable to discriminate different helminthiases, by the enzyme linked immunosorbent assay (ELISA) and the immunoblot assay (IB). Antigens used were: Taenia solium cysticercus total saline (Tso); Taenia crassiceps cysticercus vesicular fluid (Tcra-VF); T. crassiceps cysticercus glycoproteins (Tcra-GP and Tcra-(18-14)-GP); Toxocara canis larva excretory-secretory (TES); Schistosoma mansoni adult total saline (Sm) and Echinococcus granulosus hydatid fluid (Eg). The assayed sera were from patients with: cysticercosis (n = 18); toxocariasis (n = 40); schistosomiasis (n = 19) and hydatidosis (n = 50) with proven clinical and laboratory diagnosis, and sera from rabbits immunized with Tso, Tcra-VF, TES and Eg. Cross-reactivity occurred mostly between infections caused by Taenia and Echinococcus or in immunized rabbits, by ELISA. Moreover, the cross-reactivity among helminthiases was found with the use of antigens belonging to phylogenetically related parasite species, Eg, Tso and Tcra-VF, by sharing same antigenic components. Lower cross-reactivities were obtained by IB technique, when only peptides were considered as antigens, and the use of T. crassiceps purified glycoproteins demonstrated high sensitivity and specificity in the diagnosis of human cysticercosis, similarly to that using homologous antigen (Tso) by the same technique.

Animals↗
PubMed2000

Detection of specific antibody isotypes and subtypes before and after treatment of American visceral leishmaniasis.

Sera from patients with American visceral leishmaniasis (AVL) were studied before and after treatment based on their antibody isotypes and subtypes. The study was comprised of 33 Brazilian patients with well-defined diagnosis of AVL and 39 clinically healthy individuals. Antileishmanial antibody isotypes and subtypes were observed in almost all patients, except IgA that was detected in about 63% of them. The sensitivity and specificity of the immunofluorescence assay in the detection of antibody isotypes (IgG and IgM) and subtypes (IgG1, IgG2, IgG3, and IgG4) were high with no statistical difference, ranging from 0.937 to 1.000 and from 0.954 to 1.000, respectively. All IgG antibodies and its subtypes had their levels reduced after treatment. However, the IgG4 had an early decay and its conversion to negative was significantly high in children. Moreover, the profile of IgG4 before treatment corresponded to a unimodal curve that shifted to a patent bimodal curve after treatment, indicative of therapeutic success. Thus, the IgG4 shows to be a suitable immunological marker for the assessment of chemotherapy in AVL patients or communities. Our findings suggest that IgG4 correlates with IL-4 that also decreases after therapy.

Adult↗
PubMed2000

One-step heminested PCR for amplification of Neisseria meningitidis DNA in cerebrospinal fluid.

A one-step polymerase chain reaction (Heminested-PCR) was designed to target the 16S rRNA fragment simultaneously using a set of primers for the universal bacterial group and a Neisseria meningitidis species-specific sequence for diagnostic purposes. The diagnostic features of the Heminested-PCR were evaluated in the study of 168 cerebrospinal fluid (CSF) specimens from 84 patients with a N. meningitidis infection, meningitis caused by unrelated bacteria and other etiologies (57 patients), or suspicious cases (27 patients) with clinical symptoms of bacterial meningitis but with negative results from bacteriological procedures. About 90% of patients with bacterial meningitis, including those suspicious cases, had prior antibiotic therapy. The sensitivity, specificity, positive, and negative predictive values found in relation to culture and/or microscopy were 91.7, 100, 100, 100, and 90.5%, respectively. In patients suspected of having bacterial meningitis, the Heminested-PCR revealed 51.9% (14 patients) positive for N. meningitidis infection and 40.7% (11 patients) positive for unrelated bacterial infections. The agreement of the Heminested-PCR with culture and/or microscopy was high and ranked as almost perfect (kappa indices > 0.856), in contrast to its agreement with other techniques. These findings speak in favor of the molecular diagnosis of meningococcal meningitis in patients who are culture- and/or microscopy-negative, due to their prior antibiotic treatment.

DNA Primers↗
PubMed2000

Evaluation of Henes-PCR assay for Mycobacterium detection in different clinical specimens from patients with or without tuberculosis-associated HIV infection.

The need for early diagnosis of tuberculosis, particularly in HIV-infected patients, requires the development of diagnosis methods that have a high sensitivity and specificity, as does the nucleic acid-based technology. With the purpose of improving the detection of mycobacterium in different clinical samples, we proposed and evaluated an assay based on nucleic acid-amplification: heminested-PCR (Henes-PCR). The procedure was designed to identify Mycobacterium spp., M. tuberculosis complex (MTC), and M. avium complex (MAC), although it has the potential to include more primers for the identification of other species. Analytical and clinical evaluation of Henes-PCR was performed by analysis of reference strains and 356 clinical specimens from 246 patients with pulmonary and meningitis tuberculosis and unrelated infections, including 142 HIV-infected individuals. Ninety-three percent (199) positive and 100% (143) negative results were obtained in specimens from patients with tuberculosis and non-tuberculosis infection, respectively. The overall sensitivity of Henes-PCR was 93.4%, specificity was 100%, positive and negative predictive values were 100 and 91.1%, respectively. Sensitivity and negative predictive value of Henes-PCR were significantly higher than culture procedure for microscopy-negative specimens. Even though frequency of HIV infection was higher in patients with tuberculosis, diagnostic parameters of Henes-PCR were similar between HIV-positive and HIV-negative patients. MTB was identified in 194 (98%) specimens while MAC was detected in 5 (2%) specimens. These findings suggest that Henes-PCR is a useful test for rapid detection of mycobacterium in clinically suspected cases of tuberculosis with smear-negative results.

HIV Infections↗
PubMed1999

Urinary tract infection: detection of Escherichia coli antigens in human urine with an ELIEDA immunoenzymatic assay.

Escherichia coli is the most common causative agent of urinary tract infection (UTI), and diagnosing this infection usually relies on bacteriologic methods. Nevertheless, screening methods can be useful for a rapid presumptive diagnosis even though some of these screening methods have low sensitivity or are expensive. To investigate a possible new alternative approach, an antigen-based immunoassay--enzyme-linked immunoelectrodiffusion assay (ELIEDA)--was standardized for screening for this bacterial infection. Combining counter-immunoelectrophoresis with an immunoenzymatic assay, the ELIEDA requires concentrated urine specimens, a cellulose acetate membrane, polyclonal antibodies to E. coli raised in rabbits, and peroxidase-labeled sheep antibodies to rabbit immunoglobulin G (IgG). This ELIEDA technique was evaluated using 244 urine specimens, 76 of them with E. coli, 47 with heterologous bacteria, and 121 without bacteria. In comparison to bacteriologic methods, the sensitivity, specificity, and positive and negative predictive values for the ELIEDA were 93.4%, 98.2%, 95.9%, and 97.1%, respectively. The data obtained suggest that this assay is useful for routine diagnostic screening for UTI caused by E. coli. In addition, since the ELIEDA stained membranes can be stored, this assay makes retrospective studies possible.

Animals↗
PubMed1998

Improvement of the indirect hemagglutination test for the detection of antibodies to Streptococcus pyogenes.

An indirect hemagglutination test for a seroepidemiological survey of Streptococcus pyogenes infection was standardized. This is an improved modification of the indirect hemagglutination test which utilizes an unstable reagent prepared with fresh blood cells. Two types of bacterial antigens represented by extracellular products and purified streptolysin O were assayed, but only the former antigen gave good results. Pretreatment of the bacterial antigen with 0.15 M NaOH and neutralization to pH 5.5, as well as postfixation of sensitized red cells with 0.1% glutaraldehyde at 56 degrees C for 30 min were found to be essential to give long stability to the reagent in liquid suspension, at least 9 months at 4 degrees C. A total of 564 serum samples with high, moderate and low anti-streptolysin O antibodies as determined by the neutralization assay were studied by the indirect hemagglutination test using the new reagent. The sensitivity, specificity, efficiency, positive predictive value and negative predictive value of the test in relation to the neutralization assay were 0.950, 0.975, 0.963, 0.973, and 0.955, respectively. The kappa agreement index between the two techniques was high (0.926) and ranked as "almost perfect". Antibody levels detected by both techniques also presented a high positive correlation (rs = 0.726). Five reagent batches successively produced proved to be reproducible. Thus, the improved indirect hemagglutination test seems to be useful for public health laboratories.

Antigens, Bacterial↗
PubMed1997

Serological diagnosis of Chagas' disease: a potential confirmatory assay using preserved protein antigens of Trypanosoma cruzi.

The diagnosis of Chagas' disease relies mostly on data provided by immunologic tests, but inconclusive results often require elucidation, especially in blood banks. When six different types of Trypanosoma cruzi epimastigote antigens were studied by an immunoblotting assay (IBA), a preserved protein antigen (Ag PP) was found to present the most interesting immunochemical features because of its high reactivity with anti-T. cruzi antibodies. Thus, the IBA with Ag PP (PP IBA) was assessed with panels of coded and noncoded serum samples prepared in different laboratories, including the Brazilian Reference Laboratory for Chagas' Disease. It was found that serum samples from patients proved (clinically, eletrocardiographically, serologically, and epidemiologically) to have Chagas' disease consistently recognized 12 bands (140, 100, 85, 78, 59, 57, 46, 35, 27, 23, 20, and 18 kDa) of Ag PP. In contrast, sera from nonchagasic patients, including patients with mucocutaneous leishmaniasis, were negative or reacted weakly, and one serum sample did not have more than five different bands. These bands were 78, 57, 46, 35, 27, 23, 20, or 18 kDa. A criterion was adopted to interpret the results obtained in the PP IBA. The criterion considered positive a serum sample recognizing all 12 bands and considered negative a serum sample that did not recognize any of the bands except the eight nonspecific bands mentioned above. The PP IBA indicated maximum sensitivity and specificity as well as high positive and negative predictive values. The data demonstrate that the PP IBA discriminates chagasic from nonchagasic infections and seems to be applicable as a confirmatory assay for elucidating inconclusive results obtained by standard serology.

Animals↗
PubMed1996

Humoral immunity patterns based on antibody reactivity to rotavirus antigens in Brazilian children under 5 years of age.

The age distribution of antibody to simian rotavirus (SA-11) was studied in serum specimens obtained from 399 children aged to 5 years and living in the city of Recife (PE), located in the north eastern region of Brazil. Sera were examined for group-specific rotavirus antibody using a blocking enzyme immunoassay (bELISA) and a hemagglutination inhibition antibody (HIA) test, and for anti-VP2, anti-VP4, anti-VP6, and anti-VP7 antibodies using an immunoblotting assay (IBA). Antibody prevalence was similar in all bELISA and HIA assays, showing a steep rise in the 6-to 17-month-old age groups. The results indicate early acquisition of antibody to rotavirus. The majority of children aged 2 to 4 years had bELISA (50% to 60%) and HIA (70% to 81%) antibodies. There was an association in prevalence data obtained by HIA and bELISA with immunoblotting (IBA), revealing four serologic profiles. Children with profiles I and II (60%) respectively had HAI and ELISA antibody or HAI antibody alone and all had immunoprotective antibodies to VP4 and/or VP7. These children were regarded as "immune," resembling convalescent patients with a rotavirus infection. Children with profile III (4%) had no HIA antibody and only non-protective anti-VP6 and/or VP7 antibody, and were considered to be "partially immune." Children with profile IV (36%) had no detectable antibody and were classified as "nonimmune." These children should be considered to be susceptible to rotavirus infection, with the risk of developing clinically severe diarrhea.

Antibodies, Viral↗
PubMed1996

Improvement of the slide hemagglutination test for rapid Chagas' disease screening in epidemiological surveys.

A slide hemagglutination test, here called SHAT, which is practical and economical for seroepidemiological surveys was standardized. This is an improved modification of the rapid hemagglutination test (RHA) which utilizes a short-lived reagent prepared with fresh blood cells. The reagent and conditions of the test were considerably modified and, most importantly, an alkaline-solubilized Trypanosoma cruzi epimastigote antigen reagent is proposed. The stability of the SHAT reagent was at least one year at 4 degrees C, in an appropriate liquid suspension. The SHAT was applied to 71 serum samples from patients with Chagas' disease and from 235 clinically healthy blood donors. Sensitivity, specificity and positive and negative predictive values for the selected cutoff titer corresponding to 1:4 dilution were 0.972 (0.903-0.992), 0.983 (0.957-0.993), 0.945 (0.867-0.979) and 0.991 (0.969-0.998), respectively. These values were comparable to those found for the RHA, immunofluorescence (IFT), indirect hemagglutination (IHAT) and complement fixation (CFT) tests. These data suggest that the SHAT should be useful for seroepidemiological surveys conducted at public health laboratories in developing countries.

Chagas Disease↗
PubMed1996

Detection of antibody isotypes to streptolysin O by dot ELISA.

The presence of antibody isotypes (IgG, IgA and IgM) to streptolysin O was determined by dot ELISA in 222 serum samples from patients with different levels of anti-streptolysin O (SLO) antibodies as measured by the neutralizing assay (NA), from patients with diseases not related to nonsuppurative complications of Streptococcus pyogenes infection, and from clinically healthy individuals. Immunoglobulin G antibodies were found in 72% of sera from patients with SLO antibodies higher than 333 Todd units (TU), and IgA antibodies were also detected in 53%, but no IgM antibodies were demonstrable. High copositivity (0.94), conegativity (0.97), and positive (0.96) and negative (0.96) predictive values were observed when IgG and IgA findings were combined. The dot ELISA gave highly reproducible results. The present data suggest that the assay may be of practical value for routine detection of SLO antibodies when employed with an anti-human immunoglobulin light chain peroxidase conjugate.

Enzyme-Linked Immunosorbent Assay↗
PubMed1994

Studies on the efficiency of measles virus antigen production using VERO cell culture in a microcarrier system.

1. A large amount of antigen is required to conduct seroepidemiologic surveys of measles. Thus, a process to obtain measles virus antigen using a bioreactor was standardized. 2. The virus was grown in a 3.7-1 culture of VERO cells using a Celligen cell culture system containing 2 mg/ml of microcarriers (cytodex I) at 37 degrees C and 60 rpm. The cultures infected with 0.5 m.o.i. of measles virus were harvested after the appearance of the cytopathic effect. The virus suspension was clarified and concentrated by ultracentrifugation. Intracellular and extracellular virus titers were determined by hemagglutination (HA) and by induction of a cytopathic effect in cell culture (TCID50). 3. Intracellular virus presented 5-7 x 10(6) TCID50/0.1 ml, HA activity per 50 microliters equal to 32, with a total HA activity of 4,480 HA units (HAU) and specific activity of 116 HAU/mg. In the concentrated supernatants, the HA titer of extracellular virus was 64, with a total HA activity of 1,024 HAU and a specific activity of 1,600 HAU/mg. 4. The antigen obtained was suitable for the detection of antibodies against measles virus in assays such as ELISA and DOT-ELISA (using 1 micrograms/well to ELISA and 2 micrograms/DOT). 5. The microcarrier system produced antigen sufficient for 26 ELISAs/ml compared to 5.7 ELISAs/ml obtained for the static culture system.

Animals↗
PubMed1993

High resolution of Trypanosoma cruzi amastigote antigen in serodiagnosis of different clinical forms of Chagas' disease.

The serodiagnosis of Chagas' disease, a highly prevalent disorder in South American countries, is usually made by the detection of antibodies to Trypanosoma cruzi epimastigote antigen. In this study, we assess the diagnostic performance of the immunofluorescence test with T. cruzi (Y strain) amastigote antigen from an LLC-MK2-infected cell supernatant in comparison with a test with the conventional epimastigote antigen. A total of 238 serum samples from patients in the acute and chronic phases of the disease, with the chronic indeterminate, cardiac, and digestive forms, and from nonchagasic individuals were tested for the presence of immunoglobulin G (IgG), IgM, and IgA antibodies. The reactivity of the amastigote antigen in terms of geometric mean titers was 2 to 4 times higher than that of the epimastigote antigen. Clear-cut results were obtained with the amastigote antigen, with no overlapping of true and false positives. IgG antibodies to amastigotes were found in all patients with Chagas' disease, whereas all sera from nonchagasic patients were negative, except for those from patients with visceral leishmaniasis, in which 63% cross-reactivity was observed. IgM antibodies to amastigotes were detected in 100% of sera from patients with acute Chagas' disease and in 7.5% of sera from patients with chronic Chagas' disease, whereas IgA antibodies were found in 60% of sera from patients in the acute phase and in 33% of sera from patients in the chronic phase. Despite the cross-reactivity observed with sera from visceral leishmaniasis patients, the IgG immunofluorescence test with the amastigote antigen had the highest sensitivity, specificity, and efficiency. No relationship was observed between the class-specific antibodies or their titers and the clinical forms of patients in the chronic phase. Amastigotes from the cell culture supernatant proved to be useful as an alternative antigen to epimastigotes because of their high resolution in the serodiagnosis of Chagas' disease.

Animals↗
PubMed1992

Seroepidemiology of schistosomiasis mansoni.

In population surveys in which the Schistosoma mansoni intensity of infection is low, or in localities where the schistosomiasis control program had success, the parasitologic methods lack in sensitivity. Despite of some limitations, the immunological methods are useful to provide valuable information in such field conditions. Thus, the prevalence of schistosomiasis in untreated population can be determined by the detection of IgG or IgM antibodies, as well as the incidence by the IgA antibodies, employing mainly immunofluorescence (IF) and immunoenzymatic (ELISA), and in some extent hemagglutination (HA) or even skin test. The true prevalence and incidence of schistosomiasis can be estimated using a probabilistic model equation, since knowing before-hand the sensitivity and specificity of employed test. The sensitivity and the specificity of serologic test become higher in low aged group, under 14. The geometric mean IF titers also gives a positive correlation with the intensity of infection. Presently, there are need of serologic tests which are economic and practical in seroepidemiologic inquiries, requiring no specialized personnel to collect population blood or serum samples, and also easily interpret the test results. The reagents for such tests are desired to be stable and reproducible. Moreover, it is expected that the tests can distinguish an active infection.

Adolescent↗
PubMed1992

Schistosoma mansoni cercaria and schistosomulum antigens in serodiagnosis of schistosomiasis.

Schistosoma mansoni cercaria and schistosomulum obtained in vitro were used in immunofluorescence (IF) tests and indirect hemagglutination (IHA) tests of 137 study sera, 44 from subjects infected with S. mansoni and 93 from healthy subjects residing outside areas endemic for the disease. The results of these tests were compared with those obtained by testing the same sera using conventional adult worm antigen, and also with the initial clinical and parasitologic diagnoses of the 137 subjects providing the study sera. Regarding sera from acute versus chronic cases, IF testing of the acute sera consistently detected IgA antibodies along with IgM and IgG, the last two being found consistently in chronic sera. Also, the geometric mean of the IgM antibody titers found in the IF tests was higher for acute than for chronic sera. Excluding IF IgA, which was negative for chronic cases, the sensitivity of the other types of tests (IF IgG, IF IgM, and IHA) using cercaria and schistosomulum antigens, both under evaluation, ranged from 0.773 to 0.955, the specificity ranged from 0.957 to 1.000, the efficiency ranged from 0.927 to 0.985, the predictive value of positives ranged from 0.909 to 1.000, and the predictive value of negatives ranged from 0.903 to 0.979. No statistical differences were observed between these results and those obtained with conventional adult worm antigen. This suggests that cercaria and schistosomulum antigens, both of which can be produced more quickly and cheaply than adult worm antigen, could serve as reliable alternatives to adult worm antigen in the serodiagnosis of schistosomiasis mansoni.

Acute Disease↗
PubMed1991

A novel IgM-indirect hemagglutination test for the serodiagnosis of acute toxoplasmosis.

A new indirect hemagglutination (M-Toxo-IHA) test was standardized for the serodiagnosis of acute toxoplasmosis. The reagent for this test consisted of stabilized human red cells coated with a Toxoplasma gondii heat-stable alkaline-solubilized extract which reacted predominantly with IgM antibodies found in serum samples from patients with a recent infection. Almost no reactivity was verified with serum samples from chronic or ancient toxoplasmosis. The reagent preparation was simple and obviated prior fractionation of parasite components reactive with IgM antibodies. Rheumatoid factor or high levels of IgG antibodies did not interfere in the M-Toxo-IHA test. A total of 358 serum samples was assessed for the evaluation of this test, comprising sera from patients with recent and ancient toxoplasmosis, from non-related diseases, as well as from clinically healthy individuals with no previous T. gondii infections. Serodiagnostic performance of the M-Toxo-IHA test in terms of sensitivity, specificity, efficiency, and positive and negative predictive values was verified, and the values obtained were 1.000, 0.985, 0.989, 0.952, and 1.000, respectively, in relation to IgM-immunofluorescence or IgM-capture ELISA. Six different batches of reagent, successively produced for the current study, provided reproducible results and were stable for at least 22 months at 4 degrees C.

Acute Disease↗
PubMed1991

Diffusion-in-gel enzyme-linked immunosorbent assay (DIG-ELISA) for Chagas' disease serodiagnosis.

1. Diffusion-in-gel enzyme-linked immunosorbent assay (DIG-ELISA) was standardized and evaluated for the diagnosis of Chagas' disease, in comparison with the conventional serological tests indirect immunofluorescence (IFI), passive hemagglutination (PHA) and complement fixation (CF). 2. A total of 236 serum samples positive and negative for the serodiagnosis of Chagas' disease were studied. The group included 50 serum samples serologically positive for leishmaniasis and 36 positive for malaria. 3. The best diagnostic performance of DIG-ELISA was observed when serum samples were diluted to 1:8 and a diameter of zero mm (no color) was taken as the cut-off. Under these conditions, the relative indices of sensitivity, specificity and agreement were 100%. High positive correlation coefficients were obtained between DIG-ELISA and IFI (r1 = 0.9010), PHA (r2 = 0.8943) and CF (r3 = 0.8269). 4. We conclude that DIG-ELISA provides an alternative technique for screening chagasic infections, as well as for seroepidemiological surveys mainly because it is simple, easy to carry out and does not require expensive equipment.

Animals↗
PubMed1988

Immunoglobulin A antibodies to Trypanosoma cruzi antigens in digestive forms of Chagas' disease.

In an attempt to find a serological marker for the diagnosis of chronic digestive forms of Chagas' disease, we compared amastigote and trypomastigote antigens obtained from immunosuppressed mice infected with Trypanosoma cruzi (Y strain) with conventional epimastigote antigens to search for immunoglobulin A (IgA) antibodies. A total of 255 serum samples from patients with acute and chronic (indeterminate, digestive, and cardiac) forms of Chagas' disease and with nonchagasic diseases and from healthy individuals were studied. Amastigote antigens proved to be the most adequate for our purpose, since IgA antibodies could be detected in 23 of 25 serum samples from patients with digestive forms, with relative indices of sensitivity, specificity, and efficiency of 0.920, 0.911, and 0.912, respectively. These antigens also showed high reactivity with IgA antibodies, with a geometric mean titer of 16,635 (12.7 log2). IgA antibodies were detected in 16 of 28 serum samples from patients with the acute form as well, but this clinical form is easily distinguished from the chronic form by the demonstration of IgM antibodies. Poor results were seen with trypomastigote and epimastigote antigens. The finding of IgA antibodies in about 20% of indeterminate forms and 20% of cardiac forms, although in low titers, requires further investigation to ascertain their role as an early signal of gastrointestinal lesions. In addition, the amastigote antigens described here seem more convenient for use in endemic areas than those obtained from cell cultures because of their lower cost.

Animals↗