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1.
Chem Commun (Camb) ; 56(10): 1605-1607, 2020 Feb 04.
Artigo em Inglês | MEDLINE | ID: mdl-31939465

RESUMO

We report the first preparation of a monoclonal antibody (mAb) that can immobilize a palladium (Pd)-complex. The allylic amination reaction using a supramolecular catalyst consisting of the Pd-complex and mAb selectively gives the (R)-enantiomer product with an enantiomeric excess (ee) of 98 ± 2%. This is in sharp contrast to the reaction catalyzed by a conventional Pd-catalyst (ee < 2%).


Assuntos
Anticorpos Monoclonais/química , Complexos de Coordenação/química , Paládio/química , Compostos Alílicos/síntese química , Compostos Alílicos/química , Aminação , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/metabolismo , Benzilaminas/síntese química , Benzilaminas/química , Catálise , Bovinos , Complexos de Coordenação/imunologia , Complexos de Coordenação/metabolismo , Reações Cruzadas/imunologia , Feminino , Gastrópodes/química , Hemocianinas/química , Camundongos Endogâmicos BALB C , Ligação Proteica , Ródio/química , Soroalbumina Bovina/química , Estereoisomerismo , Água/química
2.
Food Chem ; 303: 125379, 2020 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-31446361

RESUMO

In this study, heterologous competitive antigens (HCAs) suitable for improving the sensitivity of ELISA were successfully screened based on their cross-reactivities (CRs) with 19 quinolone analogues; each containing the norfloxacin amino derivative (NOR0) coupled with bovine serum albumin (BSA) as a coating antigen. HCAs prepared with hapten analogues (CRs of 0.77%-49.92%) remarkably enhanced the sensitivity of the subsequent ELISA. ELISA sensitivity for NOR detection improved 26-fold when moxifloxacin-BSA was used as a heterologous coating antigen relative to when NOR0-BSA was used as a homologous coating antigen. This work, therefore, represents a detailed screening method to select suitable heterologous competitive antigens that improve ELISA sensitivity. Secondly, we present new theoretical tools to estimate hapten structures for use in the method, which may also be applied to improve the sensitivity of other immunoassays.


Assuntos
Antígenos Heterófilos/análise , Ensaio de Imunoadsorção Enzimática/métodos , Quinolonas/análise , Reações Cruzadas , Haptenos , Simulação de Acoplamento Molecular , Norfloxacino/análogos & derivados , Soroalbumina Bovina/química
3.
Acta Trop ; 201: 105201, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31562846

RESUMO

Zika virus (ZIKV) is a mosquito-borne flavivirus that has caused recent large outbreaks in the Americas. Given its association with severe congenital defects including microcephaly, distinguishing infections caused by ZIKV from those caused by dengue virus (DENV) is of primordial importance. The objectives of this study were to evaluate the recombinant proteins rEIII-ZIKV (Envelope protein domain III) and rNS1ß-leader-ZIKV (non-structural protein 1) for the serological diagnosis of ZIKV in the Mexican population. We also evaluated potential cross-reactivity in commercial enzyme-linked immunosorbent assays (ELISA) based on the ZIKV NS1 and DENV NS1 proteins. rEIII-ZIKV and rNS1ß-leader-ZIKV proteins were tested with sera from 30 PCR-confirmed ZIKV cases, 50 ZIKV-naive, DENV-exposed subjects with no acute febrile disease, (asymptomatic subjects, AS), and 50 ZIKV-naive and DENV naive AS. Commercial ELISA tests were evaluated with sera from 57 ZIKV and 20 DENV PCR-confirmed cases, and 50 ZIKV-naive, DENV-exposed AS. In-house ELISA assays showed that IgM antibody levels against rEIII-ZIKV and rNS1ß-ZIKV were higher in ZIKV naive, DENV-exposed AS than in acutely infected ZIKV individuals. IgG reactivity was highest for rEIII-ZIKV, and indistinguishable between acutely infected ZIKV cases and DENV exposed AS. Positivity for the Euroimmun Zika IgM assay at 7-10 days was considerably higher in DENV-naive ZIKV patients (86%) than in DENV-exposed ZIKV patients (33%), while 39% of the latter had false-negative anti-ZIKV IgG before 7 days of onset. DENV-exposed ZIKV patients presented lower anti-ZIKV IgM and higher IgG responses similar to a secondary dengue response. Forty-four percent of DENV- exposed acute ZIKV patients were DENV IgM positive with the Panbio Dengue assay, and two (15%) of the DENV-naive ZIKV patients presented false DENV IgG conversion. Given the extensive cross-reactivity to both the NS1 and EDIII proteins in current serological methods, the development of sensitive and specific serological tests to distinguish ZIKV from DENV infections is an urgent priority.


Assuntos
Anticorpos Antivirais/sangue , Vírus da Dengue/imunologia , Zika virus/imunologia , Reações Cruzadas , Ensaio de Imunoadsorção Enzimática/métodos , Humanos , Testes Sorológicos
4.
Vet Parasitol ; 276: 108994, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31778941

RESUMO

Neospora caninum is a protozoan parasite (Phylum Apicomplexa) that has been recently suggested as a relevant cause of reproductive disorders in small ruminants. The aim of the present study is to develop and validate a new serological test based on time resolved fluorescency using N. caninum GRA7 recombinant antigen (GRA7-TRFIA) for the detection of N. caninum antibodies in sheep. A total of 346 serum samples (208 from experimentally infected sheep, 117 from a dairy farm with a previous history of Neospora-associated abortion, and 21 negative sera) were used. The validation of the new assay was performed by the evaluation of assay precision, analytical sensitivity (Se), accuracy and cross reactivity. In the experimentally infected sheep, antibody kinetics was compared between GRA7-TRFIA and an in house N. caninum tachyzoite soluble extract-based ELISA (NcSALUVET ELISA) by Wilcoxon matched-pairs signed rank test. The cut-off and diagnostic Se and specificity (Sp) of GRA7-TRFIA was estimated by ROC analysis with field samples. In addition, concordance and correlation between GRA7-TRFIA and a commercial ELISA and NcSALUVET ELISA were assessed by kappa value and Spearman correlation coefficient, respectively. Overall, GRA7-TRFIA showed an adequate precision, analytical Se and accuracy to detect anti-N. caninum antibodies in ovine serum, and no cross reactivity with the closely related protozoan Toxoplasma gondii. In naturally infected sheep, 100% Se and 95.35% Sp were obtained for a cut-off point of 62.68 Units of Fluorometry for N. caninum (UFN). Moreover, GRA7-TRFIA allowed earlier detection of N. caninum infection than NcSALUVET ELISA in experimentally infected sheep.


Assuntos
Anticorpos Antiprotozoários/sangue , Antígenos de Protozoários/imunologia , Coccidiose/veterinária , Neospora/imunologia , Doenças dos Ovinos/imunologia , Animais , Coccidiose/imunologia , Reações Cruzadas , Ensaio de Imunoadsorção Enzimática/veterinária , Feminino , Imunoensaio de Fluorescência por Polarização , Fluorometria/veterinária , Imunoglobulina G/sangue , Sensibilidade e Especificidade , Ovinos , Doenças dos Ovinos/parasitologia
5.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 35(9): 789-793, 2019 Sep.
Artigo em Chinês | MEDLINE | ID: mdl-31750819

RESUMO

Objective To develop the colloidal gold immunochromatography test strip for qualitatively detecting the hemoglobin (Hb) in human feces. Methods Mouse anti-human Hb monoclonal antibody SPR-5 marked by colloidal gold was coated in glass fiber membrane, and then the mouse anti-human Hb SP-5 monoclonal antibody and goat anti-mouse IgG were immobilized in testing (T) line and control (C) line located in nitrocellulose membranes, respectively. With this double antibody sandwich technique and immunochromatography test, the Hb antigen would react with both antibodies coated in the T line and C line and cause two colour reactions if the detected sample was positive, whereas the antigen-antibody combination and colour reaction only showed up in the C line when the sample was negative. Results The minimum detection limit of this test strip for human Hb was 21 ng/mL and no cross reactions were found in chick Hb, rabbit Hb, sheep Hb, pig Hb and cow Hb. Conclusion The test strips can improve the detection rate of fecal occult blood obviously and avoid false-positive results.


Assuntos
Cromatografia de Afinidade , Coloide de Ouro , Fitas Reagentes , Animais , Anticorpos Monoclonais , Bovinos , Reações Cruzadas , Feminino , Humanos , Sangue Oculto , Coelhos , Sensibilidade e Especificidade , Ovinos , Especificidade da Espécie , Suínos
6.
J Agric Food Chem ; 67(46): 12918-12926, 2019 Nov 20.
Artigo em Inglês | MEDLINE | ID: mdl-31668066

RESUMO

The triosephosphate isomerase (TIM), Scy p 8, is a crab allergen and shows cross-reactivity in the shellfish. Here, recombinant Scy p 8 was expressed, and its crystal structure was determined at a resolution of 1.8 Å. The three-dimensional structure of Scy p 8 is primarily composed of a (ß/α)8-barrel motif prototype. Additionally, Scy p 8 showed cross-reactivity with high sequential and secondary structural identity among TIMs from shellfish species. The site-directed mutagenesis of critical amino acids of conformational epitopes was carried out, and the mutants of Trp 168 and Lys 237 to Ala reduced immunoglobulin E (IgE)-binding activity by approximately 30%, compared with wild-type TIM in an inhibition ELISA; however, it still induced basophil activation despite the interpatient variability between patients. These results can help to provide an accurate template for the analysis of the IgE binding and establish meaningful relationships between structure and allergenicity.


Assuntos
Braquiúros/enzimologia , Epitopos/química , Triose-Fosfato Isomerase/imunologia , Sequência de Aminoácidos , Animais , Braquiúros/química , Braquiúros/genética , Braquiúros/imunologia , Reações Cruzadas , Cristalização , Epitopos/genética , Epitopos/imunologia , Humanos , Imunoglobulina E/imunologia , Conformação Molecular , Conformação Proteica , Alinhamento de Sequência , Frutos do Mar/análise , Hipersensibilidade a Frutos do Mar/imunologia , Triose-Fosfato Isomerase/química , Triose-Fosfato Isomerase/genética
7.
Vet Parasitol ; 276: 108962, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31704559

RESUMO

Sarcocystis neurona is the major cause of the equine protozoal myeloencephalitis (EPM) in the Americas and has opossums of the genus Didelphis as definitive hosts. Most isolates of Sarcocystis sp. shed by opossums in Brazil differ genetically from the known species of Sarcocystis. These Brazilian isolates behave similarly as Sarcocystis falcatula, which causes sarcocystosis in birds, and for this reason, have been classified as Sarcocystis falcatula-like. Genes coding for the immunodominant surface antigens SAG2, SAG3 and SAG4 of S. falcatula-like are similar to those from S. neurona. It is unknown the Sarcocystis species that causes EPM in Brazil, as S. neurona has never been genetically confirmed in Brazilian horses. All cases associated with EPM in Brazil were diagnosed by immunological tests, which are not specific for S. neurona infection. It is possible that S. falcatula-like may infect horses in Brazil. The aims of the current study were to test the susceptibility of gerbils (Meriones unguiculatus) to experimental infections with S. neurona and S. falcatula-like, and to investigate potential serologic cross-reactivity to these parasites by immunofluorescent antibody test (IFAT) and Western blot (WB). A total of 27 gerbils, distributed in five experimental groups (G1-G5), were employed in this work (G1: 4 negative controls; G2: 6 infected with S. neurona merozoites, G3: 6 infected with S. falcatula-like merozoites; G4 and G5 (5 and 6, respectively, infected with different doses of sporocysts). None of the 17 animals that seroconverted for the parasites in IFAT presented any visualized organism or Sarcocystis DNA in the examined tissues. No serologic cross-reactivity was observed using IFAT. However, sera from animals infected with S. falcatula-like and S. neurona presented the same pattern of antigenic recognition when S. neurona merozoites were used as antigen in WB, including reactivity to proteins of 30 and 16 kDa, regarded as specific markers for S. neurona-infected animals. Gerbils did not sustain infection by these parasites, although produced antibodies after inoculation. These results are suggestive that other animal species that are exposed to S. falcatula-like, including horses, may present serologic cross-reactivity to S. neurona in WB. IFAT was demonstrated to be more specific that WB for the detection of antibodies to S. falcatula-like and S. neurona in the experimental conditions of this study.


Assuntos
Antígenos de Protozoários/imunologia , Sarcocystis/imunologia , Sarcocistose/imunologia , Animais , Antígenos de Superfície/imunologia , Western Blotting/veterinária , Linhagem Celular , Galinhas , Reações Cruzadas , Didelphis/parasitologia , Encefalomielite/imunologia , Encefalomielite/parasitologia , Encefalomielite/veterinária , Feminino , Imunofluorescência/veterinária , Gerbillinae , Epitopos Imunodominantes/imunologia , Reação em Cadeia da Polimerase , Sarcocistose/parasitologia , Sarcocistose/patologia , Células Vero
8.
Arerugi ; 68(9): 1141-1147, 2019.
Artigo em Japonês | MEDLINE | ID: mdl-31723111

RESUMO

We present a case of early childhood-onset pork-cat syndrome possibly due to sensitization by both cats and dogs. A 6-year-old girl was referred to our hospital because of repetitive episodes of urticaria when she consumed pork meat. The patient lived with a dog and the ground floor of her house was a veterinary clinic run by her veterinarian parents. Blood tests demonstrated high specific IgE (≥50UA/ml) against cat dander, dog dander, pork, Sus s 1, Fel d 2, Can f 1, Can f 2, and Can f 3. The skin prick test was positive for raw pork and beef. Western blotting analysis detected hot spots on 67-kDa proteins in pork meat and cat dander extract. Cross-reactivity between these two proteins was confirmed by an inhibition test. Furthermore, crossreactivity between pork meat and dog dander extract was also noted. Taken together, the diagnosis of porkcat syndrome was made, and both cats and dogs were suggested to have led to the sensitization. The patient was advised to only eat well-cooked pork, and has been followed thereafter without additional reactions. The previously reported cases of this syndrome developed during adolescence and young adulthood because a considerable period from the sensitization to the development cross-reactivity with pork meat is required. To our best knowledge, this is the youngest reported case of pork-cat syndrome among English and Japanese literatures. The nomenclature of this syndrome as pet animal-meat syndrome improves the understanding of the underlying pathogenesis of cross-reactivity between animal albumins and meat albumins.


Assuntos
Hipersensibilidade Alimentar/imunologia , Carne Vermelha , Alérgenos , Animais , Gatos , Bovinos , Criança , Reações Cruzadas , Cães , Feminino , Humanos , Imunoglobulina E/sangue , Testes Cutâneos , Suínos , Adulto Jovem
9.
Mol Immunol ; 116: 199-207, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31731097

RESUMO

A 38 kDa ß-1,3-glucanase allergen from Cryptomeria japonica pollen (CJP38) was recombinantly produced in E. coli and purified to homogeneity with the use of Ni-affinity resin. CJP38 hydrolyzed ß-1,3-glucans such as CM-curdlan and laminarioligosaccharides in an endo-splitting manner. The optimum pH and temperature for ß-1,3-glucanase activity were approximately 4.5 and 50 °C, respectively. The enzyme was stable at 30-60 °C and pH 4.0-10.5. Furthermore, CJP38 catalyzed a transglycosylation reaction to yield reaction products with a molecular weight higher than those of the starting laminarioligosaccharide substrates. The three-dimensional structure of CJP38 was determined using X-ray crystallography at 1.5 Å resolution. CJP38 exhibited the typical (ß/α)8 TIM-barrel motif, similar to allergenic ß-1,3-glucanases from banana (Mus a 5) and rubber tree latex (Hev b 2). Amino acid sequence alignment of these proteins indicated that the two-consensus IgE epitopes identified on the molecular surfaces of Mus a 5 and Hev b 2 were highly conserved in CJP38. Their conformations and surface locations were quite similar for these proteins. Sequence and structural conservation of these regions suggest that CJP38 is a candidate allergen responsible for the pollen-latex-fruit syndrome relating to Japanese cedar pollinosis.


Assuntos
Alérgenos/química , Antígenos de Plantas/química , Cryptomeria/química , Pólen/química , Alérgenos/imunologia , Sequência de Aminoácidos , Antígenos de Plantas/imunologia , Reações Cruzadas/imunologia , Cryptomeria/imunologia , Cristalografia por Raios X/métodos , Epitopos/química , Epitopos/imunologia , Escherichia coli/imunologia , Humanos , Concentração de Íons de Hidrogênio , Imunoglobulina E/química , Imunoglobulina E/imunologia , Látex/química , Látex/imunologia , Musa/química , Musa/imunologia , Proteínas de Plantas/química , Proteínas de Plantas/imunologia , Pólen/imunologia , Rinite Alérgica Sazonal/imunologia , Temperatura Ambiente
10.
Ther Umsch ; 76(6): 293-299, 2019 Nov.
Artigo em Alemão | MEDLINE | ID: mdl-31762417

RESUMO

Allergy assessment: when and how? Abstract. Allergies are common in the clinical practice. The most important allergens are pollen, house dust mites, animal dander, mold and allergens attributable to a particular work environment. The medical history is a very important part for diagnosing an allergy, because sensitizations detected by skin prick tests and laboratory tests are common but not always symptomatic and therefore without clinical relevance. There are 3 options to manage allergic diseases: avoidance of allergens, symptomatic treatment and - in selected cases - a causal treatment like allergen-specific immunotherapy.


Assuntos
Alérgenos/imunologia , Hipersensibilidade , Antígenos de Dermatophagoides , Reações Cruzadas , Humanos , Hipersensibilidade/diagnóstico , Testes Cutâneos/métodos
11.
BMC Infect Dis ; 19(1): 895, 2019 Oct 28.
Artigo em Inglês | MEDLINE | ID: mdl-31660874

RESUMO

BACKGROUND: Leishmania infantum, the etiological agent of visceral leishmaniasis, is a neglected zoonosis that requires validation and standardization of satisfactory diagnostic methodologies. Thus, the aim of the present study was to evaluate the effectiveness of cathepsin L-like protease as a target for making molecular diagnoses and as a phylogenetic marker enabling to understand the intraspecies variations and evolutionary history of L. infantum in Brazil. METHODS: We used 44 isolates of L. infantum. The cathepsin L-like gene fragments were amplified, sequenced, manually aligned and analyzed using inference methods. The sequences generated were used to search and design oligonucleotide primers to be used in reactions specific to the target parasite. RESULTS: The cathepsin L-like gene did not show any intraspecies variability among the isolates analyzed. The pair of primers proposed amplified the target deoxyribonucleic acid (DNA) of L. infantum isolates and were effective for DNA amplification at concentrations of as low as 10- 11 ng/µl. The proposed marker did not present cross-reactions with other hemoparasites. When used for making the diagnosis in a panel of clinical samples from dogs, a positivity rate of 49.03% (102/208) was obtained, versus 14.42% (30/208) for a ribosomal internal transcribed spacer (ITS) marker. In samples from sandflies, the rate was 6.25% and from humans, 14.28%. CONCLUSIONS: The results described in this work allow us to infer that CatLeish-PCR is a sensitive and specific marker for use in diagnostic trials of L. infantum and in clinical and epidemiological surveys.


Assuntos
Catepsinas/genética , Leishmania infantum/enzimologia , Leishmaniose Visceral/diagnóstico , Filogenia , Animais , Sequência de Bases , Biomarcadores , Brasil , Ensaios Enzimáticos Clínicos/normas , Reações Cruzadas/imunologia , Primers do DNA/genética , DNA de Protozoário/genética , Doenças do Cão/parasitologia , Cães , Humanos , Leishmania infantum/classificação , Doenças Negligenciadas , Reação em Cadeia da Polimerase , Psychodidae/parasitologia , Padrões de Referência , Zoonoses/parasitologia
12.
Cancer Immunol Immunother ; 68(11): 1881-1889, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31595324

RESUMO

Adoptive cell transfer (ACT) using T cell receptor (TCR) gene-modified T cells is an exciting and rapidly evolving field. Numerous preclinical and clinical studies have demonstrated various levels of feasibility, safety, and efficacy using TCR-engineered T cells to treat cancer and viral infections. Although evidence suggests their use can be effective, to what extent and how to improve these therapeutics are still matters of investigation. As TCR affinity has been generally accepted as the central role in defining T cell specificity and sensitivity, selection for and generation of high affinity TCRs has remained a fundamental approach to design more potent T cells. However, traditional methods for affinity-enhancement by random mutagenesis can induce undesirable cross-reactivity causing on- and off-target adverse events, generate exhausted effectors by overstimulation, and ignore other kinetic and cellular parameters that have been shown to impact antigen specificity. In this Focussed Research Review, we comment on the preclinical and clinical potential of TCR gene-modified T cells, summarize our contributions challenging the role TCR affinity plays in antigen recognition, and explore how structure-guided design can be used to manipulate antigen specificity and TCR cross-reactivity to improve the safety and efficacy of TCR gene-modified T cells used in ACT.


Assuntos
Citotoxicidade Imunológica/imunologia , Genes Codificadores dos Receptores de Linfócitos T/imunologia , Imunoterapia , Neoplasias/terapia , Linfócitos T/imunologia , Linfócitos T/transplante , Animais , Especificidade de Anticorpos , Reações Cruzadas , Genes Codificadores dos Receptores de Linfócitos T/genética , Humanos , Neoplasias/imunologia , Neoplasias/metabolismo , Especificidade do Receptor de Antígeno de Linfócitos T , Linfócitos T/metabolismo
13.
Exp Parasitol ; 206: 107771, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31585116

RESUMO

A PCR targeting mitochondrial cytochrome oxidase subunit III (cox3) for molecular detection of Babesia gibsoni infection in dogs has been developed in this study. Fifty blood samples from suspected clinical cases from dogs, brought to the veterinary college clinics, were examined for presence of B. gibsoni using conventional diagnosis by microscopic examination of Giemsa stained thin blood smears. In addition, species specific PCRs targeting ITS-1 region (BgITS-1 PCR) and nested PCR targeting 18S ribosomal RNA gene (Bg18SnPCR) were carried out. A 634 bp PCR fragment of B. gibsoni cox3 gene was amplified in positive samples from three geographical locations of Satara, Wai and Pune in Maharashtra state of India. From analysis of the sequence of the B. gibsoni cox3 gene, we found that the Indian isolate had 96-98% similarity to the isolate from Japan and China. Post sequencing, de-novo diagnostic primer pair for species specific amplification of 164 bp fragment of B. gibsonicox3 was designed and the PCR was standardized. The diagnostic results of de-novo Bgcox3 PCR were compared with BgITS-1 PCR and Bg18S nPCR. Thin blood smears detected 22% (11/50) samples positive for small form of Babesia species. The BgITS-1 PCR detected 25% samples (15/50) as positive and Bg18S nPCR detected 80% (40/50) B. gibsoni positive samples. The de-novo Bgcox3 PCR detected 66% (33/50) samples positive for B. gibsoni (at 95% CI). The analytical sensitivity of cox3 PCR was evaluated as 0.000003% parasitaemia or 09 parasites in 100  µl of blood. The de-novo diagnostic cox3 PCR did not cross react with control positive DNA from other haemoprotozoa and rickettsia like B. vogeli, Hepatozoon canis, Trypanosoma evansi, Ehrlichia canis and Anaplasma platys. Statistically, cox3 PCR had better diagnostic efficiency than ITS-1 PCR in terms of sensitivity (p = 0.0006). No statistically significant difference between results of cox3 PCR and 18S nPCR was observed (p = 0.1760). Kappa values estimated for each test pair showed fair to moderate agreement between the observations. Specificity of Bgcox3 PCR was 100% when compared with microscopy or BgITS-1 PCR. Sensitivity of Bgcox3 PCR was 100% when compared with that of Bg18S nPCR.


Assuntos
Babesia/isolamento & purificação , Babesiose/diagnóstico , Doenças do Cão/diagnóstico , Complexo IV da Cadeia de Transporte de Elétrons/genética , Mitocôndrias/enzimologia , Animais , Babesia/classificação , Babesia/genética , Babesiose/parasitologia , Sequência de Bases , Reações Cruzadas , DNA Espaçador Ribossômico/química , Doenças do Cão/parasitologia , Cães , Eritrócitos/parasitologia , Funções Verossimilhança , Filogenia , Reação em Cadeia da Polimerase/veterinária , Valor Preditivo dos Testes , RNA Ribossômico 18S/análise , Sensibilidade e Especificidade , Alinhamento de Sequência/veterinária
14.
BMC Infect Dis ; 19(1): 884, 2019 Oct 23.
Artigo em Inglês | MEDLINE | ID: mdl-31646973

RESUMO

BACKGROUND: We recently conducted a serosurvey of newly arrived workers in Taiwan from four Southeast Asian countries which revealed that 1% of the migrant workers had laboratory-confirmed recent Zika virus (ZIKV) infection. Taiwan, where Aedes mosquitoes are prevalent, has a close relationship with Southeast Asian countries. Up to now, 21 imported cases of ZIKV infection have been reported in Taiwan, but there has been no confirmed indigenous case. The aim of this serosurvey was to assess whether there was unrecognized ZIKV infections in Taiwan. METHODS: A total of 212 serum samples collected in a cross-sectional seroepidemiologic study conducted during the end of the 2015 dengue epidemic in Tainan, Taiwan, were analyzed. Anti-ZIKV IgM and IgG were tested using commercial enzyme-linked immunosorbent assays (ELISAs). Plaque reduction neutralization tests (PRNTs) for ZIKV and four dengue virus (DENV) serotypes were performed for samples with positive anti-ZIKV antibodies. A confirmed case of ZIKV infection was defined by ZIKV PRNT90 titer ratio ≥ 4 compared to four DENV serotypes. RESULTS: The mean age of the 212 participants was 54.0 years (standard deviation 13.7 years), and female was predominant (67.0%). Anti-ZIKV IgM and IgG were detected in 0 (0%) and 9 (4.2%) of the 212 participants, respectively. For the 9 samples with anti-ZIKV IgG, only 1 sample had 4 times higher ZIKV PRNT90 titers compared to PRNT90 titers against four dengue virus serotypes; this individual denied having traveled abroad. CONCLUSIONS: The results suggest that undetected indigenous ZIKV transmission might have occurred in Taiwan. The findings also suggest that the threat of epidemic transmission of ZIKV in Taiwan does exist due to extremely low-level of herd immunity. Our study also indicates that serological tests for ZIKV-specific IgG remain a big challenge due to cross-reactivity, even in dengue non-endemic countries.


Assuntos
Infecção por Zika virus/epidemiologia , Adulto , Idoso , Anticorpos Anti-Idiotípicos/sangue , Reações Cruzadas , Estudos Transversais , Dengue/epidemiologia , Dengue/imunologia , Ensaio de Imunoadsorção Enzimática/métodos , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Testes de Neutralização , Estudos Soroepidemiológicos , Testes Sorológicos/métodos , Taiwan/epidemiologia , Infecção por Zika virus/imunologia
15.
Mol Immunol ; 116: 140-150, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31654938

RESUMO

BACKGROUND: Non-specific lipid transfer proteins (LTPs) are important allergens in fruits, pollen, vegetables, nuts and latex. Due to their compact structure, LTPs are highly resistant to heat treatment. Here, Art v 3 from mugwort pollen and Pru p 3 from peach were used as model allergens to in-depth investigate structural and immunological properties upon thermal treatment at different buffer conditions. METHODS: Recombinant Art v 3 and Pru p 3 were purified from E. coli and incubated at 95 °C up to 120 min using sodium phosphate buffer pH 3.4 or 7.3. Physicochemical properties of allergens were analyzed in circular dichroism spectroscopy, Fourier transform infrared spectroscopy, dynamic light scattering, size exclusion chromatography, and mass spectrometry. The crystal structure of Art v 3.0201 was determined to 1.9 Šresolution. IgG and IgE binding was investigated in ELISA using murine and LTP allergic patients' sera. RESULTS: Highly pure and homogenous recombinant allergens were obtained from bacterial production. The crystal structure of Art v 3.0201 revealed an antiparallel four helix bundle with a C-terminal extension mediating an asymmetric, transient dimer interface and differently sized cavities. Both allergens showed high thermal stability at acidic conditions. In contrast, extensive heat treatment in neutral buffer induced irreversible structural changes due to lanthionine-based cysteine rearrangement. This fostered loss of the typical α-helical structure, increased molecular size and abrogation of IgG and IgE binding epitopes. Pru p 3 lost its structural integrity at shorter heat stress duration than Art v 3, which did however only partially affect the molecule's IgE binding epitopes. CONCLUSION: During thermal treatment, susceptibility to structural changes of the LTP-fold is highly dependent on the surrounding environment but also on intrinsic features of individual LTPs. This is a crucial fact to consider when processing LTP-containing food or food products as this will directly influence their allergenic potential.


Assuntos
Alanina/análogos & derivados , Antígenos de Plantas/metabolismo , Proteínas de Transporte/metabolismo , Cisteína/metabolismo , Proteínas de Plantas/metabolismo , Sulfetos/metabolismo , Alanina/metabolismo , Sequência de Aminoácidos , Animais , Artemisia/metabolismo , Reações Cruzadas/fisiologia , Epitopos/metabolismo , Escherichia coli/metabolismo , Hipersensibilidade Alimentar/metabolismo , Humanos , Imunoglobulina E/metabolismo , Imunoglobulina G/metabolismo , Camundongos , Pólen/metabolismo , Prunus/metabolismo
17.
Nat Commun ; 10(1): 4316, 2019 09 20.
Artigo em Inglês | MEDLINE | ID: mdl-31541110

RESUMO

Zika virus (ZIKV) and dengue virus (DENV) are co-endemic in many parts of the world, but the impact of ZIKV infection on subsequent DENV infection is not well understood. Here we show in rhesus macaques that the time elapsed after ZIKV infection affects the immune response to DENV infection. We show that previous ZIKV exposure increases the magnitude of the antibody and T cell responses against DENV. The time interval between ZIKV and subsequent DENV infection further affects the immune response. A mid-convalescent period of 10 months after ZIKV infection results in higher and more durable antibody and T cell responses to DENV infection than a short period of 2 months. In contrast, previous ZIKV infection does not affect DENV viremia or pro-inflammatory status. Collectively, we find no evidence of a detrimental effect of ZIKV immunity in a subsequent DENV infection. This supports the implementation of ZIKV vaccines that could also boost immunity against future DENV epidemics.


Assuntos
Dengue/imunologia , Interações Hospedeiro-Patógeno/imunologia , Linfócitos T/imunologia , Infecção por Zika virus/imunologia , Animais , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Linhagem Celular , Reações Cruzadas/imunologia , Citocinas/metabolismo , Vírus da Dengue/imunologia , Humanos , Imunidade , Imunidade Celular , Macaca mulatta/imunologia , Masculino , Fatores de Tempo , Viremia , Zika virus/imunologia
18.
Isr Med Assoc J ; 21(7): 444-448, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31507118

RESUMO

BACKGROUND: Although cross-reactions between Epstein-Barr virus (EBV) and human systemic lupus erythematosus (SLE) autoantigens occur, a complete analysis of the potential EBV peptide cross-reactome has not been performed. OBJECTIVES: To analyze the whole EBV proteome searching for peptides common to SLE-related proteins and endowed with an immunological potential. METHODS: Fifty-one SLE-related proteins were analyzed for hexapeptide sharing with EBV proteome using publicly available databases. RESULTS: An extremely high number of hexapeptides are shared between 34 human SLE autoantigens and EBV proteins. The peptide sharing mostly occurs with complement components C4 and Interleukin-10 (IL-10). CONCLUSIONS: This study thoroughly describes the EBV vs. SLE autoantigens peptide overlap and powerfully supports cross-reactivity as a major mechanism in EBV-associated SLE etiopathogenesis.


Assuntos
Autoantígenos/imunologia , Infecções por Vírus Epstein-Barr/imunologia , Lúpus Eritematoso Sistêmico/imunologia , Peptídeos/imunologia , Proteoma , Complemento C4/imunologia , Reações Cruzadas/imunologia , Bases de Dados Factuais , Herpesvirus Humano 4/imunologia , Humanos , Interleucina-10/imunologia
19.
Infect Immun ; 87(12)2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31548320

RESUMO

Lipopolysaccharides (LPSs) of Gram-negative bacteria comprise lipid A, core, and O-polysaccharide (OPS) components. Studies have demonstrated that LPSs isolated from the pathogenic species Burkholderia pseudomallei and Burkholderia mallei and from less-pathogenic species, such as Burkholderia thailandensis, are potent immune stimulators. The LPS structure of B. pseudomallei, the causative agent of melioidosis, is highly conserved in isolates from Thailand; however, the LPSs isolated from other, related species have not been characterized to enable understanding of their immune recognition and antigenicities. Here, we describe the structural and immunological characteristics of the LPSs isolated from eight Burkholderia species and compare those for B. pseudomallei to those for the other seven species. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS), gas chromatography (GC), SDS-PAGE, Toll-like receptor 4 (TLR4) stimulation, and immunoblot analysis were performed on these Burkholderia species. MALDI-TOF profiles demonstrated that Burkholderia lipid A contains predominantly penta-acylated species modified with 4-amino-4-deoxy-arabinose residues at both terminal phosphate groups. The lipid A could be differentiated based on mass differences at m/z 1,511, 1,642, 1,773, and 1,926 and on fatty acid composition. LPSs of all species induced TLR4-dependent NF-κB responses; however, while SDS-PAGE analysis showed similar LPS ladder patterns for B. pseudomallei, B. thailandensis, and B. mallei, these patterns differed from those of other Burkholderia species. Interestingly, immunoblot analysis demonstrated that melioidosis patient sera cross-reacted with OPSs of other Burkholderia species. These findings can be used to better understand the characteristics of LPS in Burkholderia species, and they have implications for serological diagnostics based on the detection of antibodies to OPS.


Assuntos
Burkholderia mallei/imunologia , Burkholderia pseudomallei/imunologia , Burkholderia/imunologia , Lipídeo A/imunologia , Receptor 4 Toll-Like/metabolismo , Amino Açúcares/química , Anticorpos Antibacterianos/imunologia , Reações Cruzadas/imunologia , Humanos , Lipídeo A/química , Melioidose/imunologia , Melioidose/microbiologia , Conformação Molecular , Polissacarídeos Bacterianos/imunologia , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz
20.
Mol Immunol ; 114: 497-512, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31518854

RESUMO

Compounding with the problem of frequent antigenic shift and occasional drift of the segmented genome of Avian Influenza Virus (AIV), vaccines based on major surface glycoproteins such as haemagglutinin (HA) to counter heterosubtypic AIV infection in chickens remain unsuccessful. In contrast, neuraminidase (NA), the second most abundant surface glycoprotein present in viral capsid is less mutable and, in some instances, successful in eliciting inter-species cross-reactive antibody responses. However, without selective activation of B-cells and T-cells, the ability of NA to induce strong cell mediated immune responses is limited, thus NA based vaccines cannot singularly address the risk of virus escape from host defence. To this end, the highly conserved ectodomain of influenza matrix protein-2 (M2e) has emerged as an attractive cross-protective vaccine target. The present study describes the potential of recombinant Lactococcus lactis (rL. lactis) in expressing functional influenza NA or M2e proteins and conferring effective mucosal and systemic immune responses in the intestine as well as in the upper respiratory airways (trachea) of chickens. In addition, lavages collected from trachea and intestine of birds administered with rL. lactis expressing influenza NA or M2e protein were found to protect MDCK cells against avian influenza type A/PR/8/34 (H1N1) virus challenge. Although minor, the differences in the expression of pro-inflammatory cytokines gene transcripts targeted in this study among the birds administered with either empty or rL. lactis could be attributed to the activation of innate response by L. lactis.


Assuntos
Galinhas/imunologia , Imunidade nas Mucosas/imunologia , Influenza Aviária/imunologia , Lactococcus lactis/imunologia , Neuraminidase/imunologia , Proteínas da Matriz Viral/imunologia , Animais , Anticorpos Antivirais/imunologia , Linfócitos B/imunologia , Galinhas/virologia , Reações Cruzadas/imunologia , Vírus da Influenza A Subtipo H1N1/imunologia , Vacinas contra Influenza/imunologia , Linfócitos T/imunologia , Vacinação/métodos
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