LIDIA YAMAMOTO
Projetos de Pesquisa
Unidades Organizacionais
LIM/48 - Laboratório de Imunologia, Hospital das Clínicas, Faculdade de Medicina
17 resultados
Resultados de Busca
Agora exibindo 1 - 10 de 17
- Single-round multiplex PCR with species-specific mitochondrial primers of P. falciparum, P. vivax/P. simium and P. malariae/P. brasilianum: Comparison with standard techniques(2022) DOMINGUES, Wilson; SANTOS, Emilly Henrique dos; YAMAMOTO, Lidia; SANTI, Silvia Maria Di; KANUNFRE, Kelly Aparecida; OKAY, Thelma SuelyA single-round multiplex PCR (mPCR) with species-specific primers (SSP) of three mitochondrial genes of Plasmodium, namely COX I, COX III and CYT B, was compared to microscopy and 18S rRNA semi-nested PCR, nested-PCR and Real Time PCRs (*PCRs). Each parasite has between 20 and 150 mitochondria and each mitochondria has one copy of each target gene, while 18S rRNA gene is repeated 4 to 8 times. The specificity of mPCR was assessed by testing Plasmodium from rodents and birds, parasites responsible for other endemic diseases in the country such as schistosomiasis, Chagas disease and leishmaniasis in addition to microorganisms that, like Plasmodium, can cause anemia (Bartonella henselae, Babesia vogeli, Rickettsia vini). No cross-reactions were detected. From a total of 149 specimens from suspected cases of malaria were tested, 97 were positive by microscopy (49 P. falciparum, 38 P. vivax, 6 P. malariae, 4 P. falciparum/P. vivax- mixed infections) and 52 were negative; 148 samples were positive by *PCRs (49 P. falciparum, 53 P. vivax, 7 P. malariae and 39 mixed infections) and one was negative; 146 were positive by mPCR (49 P. falciparum, 56 P. vivax, 9 P. malariae and 32 mixed infections) and three were negative. The comparison of groups found statistically significant differences between microscopy vs.*PCRs or vs. mPCR (p-values <0.0001), but no difference was found between mPCR vs. *PCRs (p=0.946). The agreement in the identification of Plasmodium species was only regular, with Kappa indices of 0.407 (microscopy vs. *PCRs), 0.433 (microscopy vs. mPCR) and 0.558 (*PCRs vs. mPCR). In conclusion, the diagnostic performance of mPCR was comparable to those of *PCRs, and superior to microscopy, although the identification of Plasmodium species showed many disagreements. In conclusion, a sensitive and specific one-round SSP multiplex PCR, capable of simultaneously detecting and identifying P. falciparum, P. vivax/P. simium and P. malariae/P. brasilianum may be useful in resource-constrained countries where quantitative amplifications are not yet fully accessible.
- Accuracy of the QuantiFERON-TB Gold in Tube for diagnosing tuberculosis in a young pediatric population previously vaccinated with Bacille Calmette-Guerin(2014) VALLADA, Marcelo Genofre; OKAY, Thelma Suely; DEL NEGRO, Gilda Maria B.; ANTONIO, Claudio Amaral; YAMAMOTO, Lidia; RAMOS, Sonia Regina T. S.Objective: To evaluate the accuracy of an interferongamma release assay (QuantiFERON-TB Gold in Tube) for diagnosing Mycobacterium tuberculosis infection in a young pediatric population. Methods: 195 children previously vaccinated with BCG were evaluated, being 184 healthy individuals with no clinical or epidemiological evidence of mycobacterial infection, and 11 with Mycobacterium tuberculosis infection, according to clinical, radiological, and laboratory parameters. A blood sample was obtained from each child and processed according to the manufacturer's instructions. The assay performance was evaluated by a Receiver Operating Characteristic (ROC) curve. Results: In the group of 184 non-infected children, 130 (70.6%) were under the age of four years (mean age of 35 months). In this group, 177 children (96.2%) had negative test results, six (3.2%) had indeterminate results, and one (0.5%) had a positive result. In the group of 11 infected children, the mean age was 58.5 months, and two of them (18%) had negative results. The ROC curve had an area under the curve of 0.88 (95%CI 0.82-0.92; p<0.001), disclosing a predictive positive value of 81.8% for the test (95%CI 46.3-97.4). The assay sensitivity was 81.8% (95%CI 48.2-97.2) and the specificity was 98.8% (95%CI 96-99.8). Conclusions: In the present study, the QuantiFERON-TB Gold in Tube performance for diagnosing M. tuberculosis infection was appropriate in a young pediatric population.
- PRELIMINARY REPORT ON THE PUTATIVE ASSOCIATION OF IL10-3575 T/A GENETIC POLYMORPHISM WITH MALARIA SYMPTOMS(2016) DOMINGUES, Wilson; KANUNFRE, Kelly Aparecida; RODRIGUES, Jonatas Cristian; TEIXEIRA, Leandro Emidio; YAMAMOTO, Lidia; OKAY, Thelma SuelyOnly a small percentage of individuals living in endemic areas develop severe malaria suggesting that host genetic factors may play a key role. This study has determined the frequency of single nucleotide polymorphisms (SNPs) in some pro and anti-inflammatory cytokine gene sequences: IL6 (-174; rs1800795), IL12p40 (+1188; rs3212227), IL4 (+33; rs2070874), IL10 (-3575; rs1800890) and TGF beta 1 (+869; rs1800470), by means of PCR-RFLP. Blood samples were collected from 104 symptomatic and 37 asymptomatic subjects. Laboratory diagnosis was assessed by the thick blood smear test and nested-PCR. No association was found between IL6 (-174), IL12p40 (+1188), IL4 (+33), IL10 (-3575), TGF beta 1 (+869) SNPs and malaria symptoms. However, regarding the IL10 -3575 T/A SNP, there were significantly more AA and AT subjects, carrying the polymorphic allele A, in the symptomatic group (chi(2) = 4.54, p = 0.01, OR = 0.40 [95% CI - 0.17- 0.94]). When the analysis was performed by allele, the frequency of the polymorphic allele A was also significantly higher in the symptomatic group (chi(2) = 4.50, p = 0.01, OR = 0.45 [95% CI - 0.21-0.95]). In conclusion, this study has suggested the possibility that the IL10 - 3575 T/A SNP might be associated with the presence and maintenance of malaria symptoms in individuals living in endemic areas. Taking into account that this polymorphism is related to decreased IL10 production, a possible role of this SNP in the pathophysiology of malaria is also suggested, but replication studies with a higher number of patients and evaluation of IL10 levels are needed for confirmation.
- The performance of four molecular methods for the laboratory diagnosis of congenital toxoplasmosis in amniotic fluid samples(2013) TEIXEIRA, Leandro Emidio; KANUNFRE, Kelly Aparecida; SHIMOKAWA, Paulo Tadashi; TARGA, Lilia Spaleta; RODRIGUES, Jonatas Cristian; DOMINGUES, Wilson; YAMAMOTO, Lidia; OKAY, Thelma SuelyIntroduction: Toxoplasmosis may be life-threatening in fetuses and in immune-deficient patients. Conventional laboratory diagnosis of toxoplasmosis is based on the presence of IgM and IgG anti-Toxoplasma gondii antibodies; however, molecular techniques have emerged as alternative tools due to their increased sensitivity. The aim of this study was to compare the performance of 4 PCR-based methods for the laboratory diagnosis of toxoplasmosis. One hundred pregnant women who seroconverted during pregnancy were included in the study. The definition of cases was based on a 12-month follow-up of the infants. Methods: Amniotic fluid samples were submitted to DNA extraction and amplification by the following 4 Toxoplasma techniques performed with parasite B1 gene primers: conventional PCR, nested-PCR, multiplex-nested-PCR, and real-time PCR. Seven parameters were analyzed, sensitivity (Se), specificity (Sp), positive predictive value (PPV), negative predictive value (NPV), positive likelihood ratio (PLR), negative likelihood ratio (NLR) and efficiency (Ef). Results: Fifty-nine of the 100 infants had toxoplasmosis; 42 (71.2%) had IgM antibodies at birth but were asymptomatic, and the remaining 17 cases had non-detectable IgM antibodies but high IgG antibody titers that were associated with retinochoroiditis in 8 (13.5%) cases, abnormal cranial ultrasound in 5 (8.5%) cases, and signs/symptoms suggestive of infection in 4 (6.8%) cases. The conventional PCR assay detected 50 cases (9 false-negatives), nested-PCR detected 58 cases (1 false-negative and 4 false-positives), multiplex-nested-PCR detected 57 cases (2 false-negatives), and real-time-PCR detected 58 cases (1 false-negative). Conclusions: The real-time PCR assay was the best-performing technique based on the parameters of Se (98.3%), Sp (100%), PPV (100%), NPV (97.6%), PLR (co), NLR (0.017), and Ef (99%).
- Myocarditis in children and detection of viruses in myocardial tissue: Implications for immunosuppressive therapy(2011) CAMARGO, Paulo Roberto; OKAY, Thelma Suely; YAMAMOTO, Lidia; NEGRO, Gilda Maria Barbaro Del; LOPES, Antonio AugustoBackground: There is scarce information on the potential benefits of immunosuppression in children with myocarditis and viral genomes in myocardium. We investigated the occurrence of myocarditis in children with a preliminary diagnosis of dilated cardiomyopathy, the frequency of cardiotropic viruses in the myocardium, and the response to immunosuppression. Methods: Thirty patients (nine months to 12 years) with left ventricular ejection fraction of 22.8 +/- 4.1% were subjected to right cardiac catheterization and endomyocardial biopsy. Specimens were analyzed for the presence of inflammatory elements (Dallas criteria) and viral genome (polymerase chain reaction). Patients with active myocarditis received immunosuppressants (azatioprine and prednisone) and were recatheterized nine months later. A historical control group of nine patients with myocarditis who did not receive immunosuppressants was included. Results: Active myocarditis was diagnosed in ten patients (five with viral genomes detected). Immunosuppression resulted in a significant increase in left ventricular ejection fraction from 25.2 +/- 2.8% to 45.7 +/- 8.6% (versus 20.0 +/- 4.0% to 22.0 +/- 9.0% in historical controls, p < 0.01) and cardiac index from 3.28 +/- 0.51 L/min/m(2) to 4.40 +/- 0.49 L/min/m(2) (versus 3.50 +/- 0.40 L/min/m(2) to 3.70 +/- 0.50 L/min/m(2) in controls, p < 0.01), regardless of the presence of viral genomes (p - 0.98 and p - 0.22, respectively for the two variables). No relevant clinical events were observed. Non-inflammatory cardiomyopathy was diagnosed in 20 patients (seven with viral genomes). While on conventional therapy, there were four deaths and three assignments to transplantation, and no improvement of left ventricular ejection fraction in the remaining ones (22.5 +/- 3.6% to 27.5 +/- 10.6%). Conclusion: Children with chronic myocarditis seem to benefit from immunosuppressive therapy, regardless of the presence of viral genome in the myocardium.
- HLA-DQA1/B1 alleles as putative susceptibility markers in congenital toxoplasmosis(2016) SHIMOKAWA, Paulo Tadashi; TARGA, Lilia Spaleta; YAMAMOTO, Lidia; RODRIGUES, Jonatas Cristian; KANUNFRE, Kelly Aparecida; OKAY, Thelma SuelyHost and parasite genotypes are among the factors associated with congenital toxoplasmosis pathogenesis. As HLA class II molecules play a key role in the immune system regulation, the aim of this study was to investigate whether HLA-DQA1/B1 alleles are associated with susceptibility or protection to congenital toxoplasmosis. One hundred and twenty-two fetuses with and 103 without toxoplasmosis were studied. The two study groups were comparable according to a number of socio-demographic and genetic variables. HLA alleles were typed by PCR-SSP. In the HLA-DQA1 region, the allele frequencies showed that *01:03 and *03:02 alleles could confer susceptibility ( OR= 3.06, p = 0.0002 and OR=9.60, p= 0.0001, respectively) as they were more frequent among infected fetuses. Regarding the HLA-DQB1 region, the *05: 04 allele could confer susceptibility ( OR = 6.95, p < 0.0001). Of the 122 infected fetuses, 10 presented susceptibility haplotypes contrasting with only one in the non-infected group. This difference was not statistically significant after correction for multiple comparison ( OR = 9.37, p=0.011). In the casuistic, there were two severely damaged fetuses with high parasite loads determined in amniotic fluid samples and HLA-DQA1 susceptibility alleles. In the present study, a discriminatory potential of HLA-DQA1/B1 alleles to identify susceptibility to congenital toxoplasmosis and the most severe cases has been shown.
- Performance of a Multiplex Nested Polymerase Chain Reaction in Detecting 7 Pathogens Containing DNA in Their Genomes Associated With Congenital Infections(2020) YAMAMOTO, Lidia; AMORIM FILHO, Antonio G.; QUEIROZ, Joelma A.; CARVALHO, Mario H. B. de; RODRIGUES, Jonatas C.; KANUNFRE, Kelly A.; V, Rossana P. Francisco; OKAY, Thelma SuelyContext.-Infections are the leading cause of perinatal and infant mortality in low-income and low-resource countries, which have a higher prevalence of infections. Definitive diagnosis of congenital and perinatal infections is largely dependent upon the results of laboratory tests. Objective.-To develop a multiplex nested polymerase chain reaction (PCR) technique for the simultaneous detection of 7 pathogens containing DNA in their genomes in suspected cases of congenital infection. Design.-Eligible participants were pregnant women with positive immunoglobulin M antibodies raised to one of the pathogens in the prenatal serologic screening, associated or not with fetal ultrasound abnormalities or positive fetal serology. Neonates whose mothers did not attend prenatal care were included when they presented with symptomatology and laboratory parameters suggestive of infection. The detection rate of the multiplex nested PCR was compared with maternal, fetal, and neonatal serology, as well as placental immunohistochemistry and noncommercial amplifications. Results.-Of 161 suspected cases, the multiplex nested PCR detected 60 (37.3%), whereas the tests available in hospital laboratories detected 13 of 60 (21.7%) of the cases detected by the multiplex nested PCR, demonstrating a 4.6 times higher detection rate for the multiplex nested PCR (Fisher exact test, P < .001). Positive amplifications were to Toxoplasma gondii (32 cases), cytomegalovirus (14 cases), parvovirus B19 (5 cases), and adenovirus (5 cases). In 4 cases, 2 pathogens were simultaneously detected. All types of biological matrices were suitable for amplification. Sequencing of multiplex nested PCR products confirmed the molecular findings. Conclusions.-The multiplex nested PCR significantly increased the number of diagnosed congenital infections. Given the scarcity of DNA recovered from amniotic fluid and some neonatal samples, this multiplex nested PCR allows the simultaneous detection of 7 pathogens associated with congenital infections in a reliable, faster, cost-effective, and more sensitive way.
- Factors associated with hyperglycemia and low insulin levels in children undergoing cardiac surgery with cardiopulmonary bypass who received a single high dose of methylprednisolone(2013) ARKADER, Ronaldo; MALBOUISSON, Luiz Marcelo; NEGRO, Gilda Maria Barbaro Del; YAMAMOTO, Lidia; OKAY, Thelma SuelyOBJECTIVES: Administering steroids before cardiopulmonary bypass in pediatric heart surgery modulates systemic inflammatory response syndrome and improves postoperative recovery. However, the use of steroids aggravates hyperglycemia, which is associated with a poor prognosis. Adult patients with systemic inflammatory response syndrome usually evolve with hyperglycemia and high insulin levels, whereas >90% of pediatric patients exhibit hyperglycemia and low insulin levels. This study aims to determine: A) the metabolic and inflammatory factors that are associated with hyperglycemia and low insulin levels in children who underwent cardiac surgery with cardiopulmonary bypass and who received a single high dose of methylprednisolone and B) the best predictors of insulin variation using a mathematical model. METHODS: This preliminary study recruited 20 children who underwent heart surgery with cardiopulmonary bypass and received methylprednisolone (30 mg/kg) immediately after anesthesia. Among the 20 patients initially recruited, one was excluded because of the absence of hyperglycemia and lower insulin levels after surgery. However, these abnormalities were confirmed in the remaining 19 children. The C-peptide, CRP, IL-6, and adrenomedullin levels were measured before surgery, immediately after cardiopulmonary bypass, and on the first, second, and third days after cardiac surgery. RESULTS: IL-6, CRP, and adrenomedullin increments were observed, whereas the C-peptide levels remained within reference intervals. CONCLUSION: The multiple regression model demonstrated that in addition to age and glycemia (two well-known factors that are directly involved in glucose metabolism), adrenomedullin and IL-6 levels were independent factors associated with lower insulin concentrations. These four parameters were responsible for 64.7% of the observed insulin variances. In addition, the fact that C-peptide levels did not fall together with insulin could have grounded the medical decision not to administer insulin to patients.
- Coxsackievirus A6 strains causing an outbreak of hand-foot-and-mouth disease in Northeastern Brazil in 2018(2022) LUCHS, Adriana; AZEVEDO, Lais Sampaio de; SOUZA, Ellen Viana de; MEDEIROS, Roberta Salzone; SOUZA, Yasmin Franca Viana Pires de; TEIXEIRA, Dalane Loudal Florentino; CARNEIRO, Thiago Franco de Oliveira; ALENCAR, Gabriela Maria Fernandes de; MORAIS, Fernanda Lucia de Sousa Leite; PINTO, Diana de Fatima Alves; OKAY, Thelma Suely; YAMAMOTO, Lidia; MORAIS, Vanessa dos Santos; ARAUJO, Emerson Luiz Lima; LEAL, Elcio; COSTA, Antonio Charlys daHand-foot-and-mouth disease (HFMD) is a highly contagious viral disease commonly associated to Enteroviruses (EV). During 2018, Brazil faced massive HFMD outbreaks spread across the country. This study aimed to characterize the EV responsible for the HFMD outbreak that occurred in Paraiba State, Brazilian Northeastern region, in 2018, followed by a phylogenetic analysis to detail information on its genetic diversity. A total of 49 serum samples (one from each patient) collected from children <= 15 years old, clinically diagnosed with HFMD were tested for EV using conventional RT-PCR and RT-qPCR. EV infection was confirmed in 71.4% (35/49) of samples. The mean and median ages were 1.83 years and one year old, respectively. Twenty-two EV-positive samples were successfully sequenced and classified as EV-A species; 13 samples were also identified with the CV-A6 genotype. The phylogenetic analysis (VP1 region) of three samples revealed that the detected CV-A6 strains belonged to sub-lineage D3. The CV-A6 strains detected here clustered with strains from South America, Europe and West Asia strains that were also involved in HFMD cases during the 2017-2018 seasons, in addition to the previously detected Brazilian CV-A6 strains from 2012 to 2017, suggesting a global co-circulation of a set of different CV-A6 strains introduced in the country at different times. The growing circulation of the emerging CVA6 associated with HFMD, together with the detection of more severe cases worldwide, suggests the need for a more intense surveillance system of HFMD in Brazil. In addition, this investigation was performed exclusively on serum samples, and the analysis of whole blood samples should be considered and could have shown advantages when employed in the diagnosis of enteroviral HFMD outbreaks.
- SARS-CoV-2 infections with emphasis on pediatric patients: a narrative review(2020) YAMAMOTO, Lidia; SANTOS, Emilly Henrique dos; PINTO, Lacyane Silva; ROCHA, Mussya Cisotto; KANUNFRE, Kelly Aparecida; VALLADA, Marcelo Genofre; OKAY, Thelma SuelyThis narrative review summarizes the main aspects underlying the new coronavirus SARS-CoV-2, its epidemiology, pathophysiology, pointing to differences of SARS-CoV-2 main receptors ACE2, in terms of expression and the amount of soluble ACE2 in the circulation of children, men and women, and also in those with risk factors such as the smokers and pregnant women or presenting with comorbidities (diabetes, obesity, hypertension and other cardiovascular diseases, renal and CNS pre-existing diseases). Clinical manifestations in adults and children were also described, emphasizing the particularities already seen in children, regarding signs, symptoms, viral excretion time and the involvement of all organs and systems. The COVID-19 in the pediatric population was divided into two sections: one dedicated to previously healthy children and adolescents with COVID-19, and the other to those who live with comorbidities and acquired COVID-19. A few paragraphs were reserved to the recently described severe multisystemic inflammatory syndrome associated with COVID-19 (MIS-C) that shares certain characteristics with Kawasaki disease. Some studies on the infection in pregnant and postpartum women, as well as neonates were shown. This review has also covered the laboratory diagnosis of COVID-19, passing through the imaging diagnosis made by the chest tomography revealing ground glass patching opacities, and results of non-specific exams such as the total blood with lymphopenia, the coagulation tests with increased prothrombin times, as well as marked increments of the D-dimer, troponin and proinflammatory cytokines. In the section devoted to the specific laboratory diagnosis of COVID-19, the most used RT-PCR protocols were described and some studies on the serological diagnosis with IgA, IgM and IgG detection were detailed, including the use of rapid immunochromatographic assays and discussing the ideal period after the onset of symptoms to perform each type of test. In the end, the management of pediatric patients with COVID-19 based mainly on supportive measures has been briefly commented.