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1.
Ig Sanita Pubbl ; 75(4): 317-325, 2019.
Artigo em Italiano | MEDLINE | ID: mdl-31887737

RESUMO

The possible risk of hyperimmunization after tetanus vaccination is currently discussed after the National Vaccine Prevention Plan 2017-2019 confirmed the recommendation of a booster dose every ten years. Due to the ubiquitous nature of tetanus spores and the inability to obtain herd-immunity through vaccination, efforts to reduce the incidence of tetanus aim at eliminating the disease. The only way to prevent infection is vaccination followed by recommended periodic booster doses. Between 2012 and 2016, Italy notified 45% (252/564) of all cases reported by the 26 EU Member States, most of them in the over 65 age group, generally women in the rural areas. The recommendation of the antipertussis vaccine, combined with anti-tetanus, in pregnancy and the indications for antitetanic prophylaxis by vaccination or specific immunoglobulins in emergency setting, gives rise to doubts about the risk of hyperimmunization. Studies generally agree on the safety of diphtheria-tetanus-pertussis combined vaccines during the third trimester of pregnancy, and the time elapsed since the previous tetanus vaccination seems not to be related to significant differences in the incidence of adverse events or obstetrical complications. In the emergency wards, given the relatively high incidence of tetanus in Italy, the risk/benefit ratio often leads to prefer vaccination to no-intervention. The administration of tetanus immunoglobulins in subjects not vaccinated in the last 10 years seems justified by the epidemiology of tetanus in Italy.


Assuntos
Vacinas contra Difteria, Tétano e Coqueluche Acelular/efeitos adversos , Vacinas contra Difteria, Tétano e Coqueluche Acelular/imunologia , Difteria/prevenção & controle , Imunização Secundária/efeitos adversos , Tétano/prevenção & controle , Coqueluche/prevenção & controle , Anticorpos Antibacterianos/imunologia , Difteria/imunologia , Vacinas contra Difteria, Tétano e Coqueluche Acelular/administração & dosagem , Feminino , Humanos , Itália , Tétano/imunologia , Coqueluche/imunologia
2.
BMC Infect Dis ; 19(1): 940, 2019 Nov 07.
Artigo em Inglês | MEDLINE | ID: mdl-31699037

RESUMO

BACKGROUND: Bacillus anthracis causes a highly lethal infectious disease primarily due to toxin-mediated injury. Antibiotics are no longer effective to treat the accumulation of anthrax toxin, thereby new strategies of antibody treatment are essential. Two anti- anthrax protective antigen (PA) antibodies, hmPA6 and PA21, have been reported by our lab previously. METHODS: The mechanisms of the two antibodies were elucidated by Electrophoresis, Competitive Enzyme-linked immune sorbent assay, Western blot analysis and immunoprecipitation test, and in vitro, in vivo (F344 rats) treatment test. The epitopes of the two antibodies were proved by Western blot and Enzyme-linked immune sorbent assay with different domains of PA. RESULTS: In this study, we compared affinity and neutralization of these two antibodies. PA21 was better in protecting cells and rats, whereas hmPA6 had higher affinity. Furthermore, the neutralization mechanisms of the two antibodies and their recognition domains of PA were studied. The results showed that hmPA6 recognized domain IV, thus PA could not bind to cell receptors. Conversely, PA21 recognized domain II, thereby limiting heptamer oligomerization of PA63 in cells. CONCLUSIONS: Our studies elucidated the mechanisms and epitopes of hmPA6 and PA21. The present investigation can advance future use of the two antibodies in anthrax treatment or prophylaxis, and potentially as a combination treatment as the antibodies target different epitopes.


Assuntos
Anticorpos Antibacterianos/metabolismo , Anticorpos Neutralizantes/metabolismo , Bacillus anthracis/imunologia , Animais , Antraz/imunologia , Anticorpos Antibacterianos/imunologia , Anticorpos Antibacterianos/farmacologia , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/farmacologia , Antígenos de Bactérias/toxicidade , Toxinas Bacterianas/toxicidade , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Eletroforese , Epitopos/análise , Epitopos/imunologia , Imunoensaio , Ratos , Ratos Endogâmicos F344
3.
Infect Immun ; 88(1)2019 12 17.
Artigo em Inglês | MEDLINE | ID: mdl-31712267

RESUMO

Staphylococcus aureus is a causative agent of chronic biofilm-associated infections that are recalcitrant to resolution by the immune system or antibiotics. To combat these infections, an antistaphylococcal, biofilm-specific quadrivalent vaccine against an osteomyelitis model in rabbits has previously been developed and shown to be effective at eliminating biofilm-embedded bacterial populations. However, the addition of antibiotics was required to eradicate remaining planktonic populations. In this study, a planktonic upregulated antigen was combined with the quadrivalent vaccine to remove the need for antibiotic therapy. Immunization with this pentavalent vaccine followed by intraperitoneal challenge of BALB/c mice with S. aureus resulted in 16.7% and 91.7% mortality in pentavalent vaccine and control groups, respectively (P < 0.001). Complete bacterial elimination was found in 66.7% of the pentavalent cohort, while only 8.3% of the control animals cleared the infection (P < 0.05). Further protective efficacy was observed in immunized rabbits following intramedullary challenge with S. aureus, where 62.5% of the pentavalent cohort completely cleared the infection, versus none of the control animals (P < 0.05). Passive immunization of BALB/c mice with serum IgG against the vaccine antigens prior to intraperitoneal challenge with S. aureus prevented mortality in 100% of mice and eliminated bacteria in 33.3% of the challenged mice. These results demonstrate that targeting both the planktonic and biofilm stages with the pentavalent vaccine or the IgG elicited by immunization can effectively protect against S. aureus infection.


Assuntos
Antígenos de Bactérias/imunologia , Infecções Estafilocócicas/imunologia , Infecções Estafilocócicas/prevenção & controle , Vacinas Antiestafilocócicas/imunologia , Staphylococcus aureus/imunologia , Animais , Anticorpos Antibacterianos/administração & dosagem , Anticorpos Antibacterianos/imunologia , Modelos Animais de Doenças , Imunização Passiva , Imunoglobulina G/administração & dosagem , Imunoglobulina G/imunologia , Camundongos Endogâmicos BALB C , Coelhos , Vacinas Antiestafilocócicas/administração & dosagem , Análise de Sobrevida , Resultado do Tratamento
4.
Mol Immunol ; 116: 98-105, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31634816

RESUMO

Pseudomonas aeruginosa is a common nosocomial pathogen in burn patients, and rapidly acquires antibiotic resistance; thus, developing an effective therapeutic approach is the most promising strategy for combating infection. Type III secretion system (T3SS) translocates bacterial toxins into the cytosol of the targeted eukaryotic cells, which plays important roles in the virulence of P. aeruginosa infections in both acute pneumonia and burn wound models. The PcrV protein, a T3SS translocating protein, is required for T3SS function and is a well-validated target in animal models of immunoprophylactic strategies targeting P. aeruginosa. In the present study, we evaluated the protective efficacy of chicken egg yolk antibodies (IgY) raised against recombinant PcrV (r-PcrV) in both acute pneumonia and burn wound models. R-PcrV protein was generated by expressing the pcrV gene (cloned in pET-28a vector) in E. coli BL-21. Anti-PcrV IgY was obtained by immunization of hen. Anti-PcrV IgY induced greater protection in P. aeruginosamurine acute pneumonia and burn wound models than control IgY (C-IgY) and PBS groups. Anti-PcrV IgY improved opsonophagocytic killing and inhibition of bacterial invasion of host cells. Taken together, our data provide evidence that anti-PcrV IgY can be a promising therapeutic candidate for combating P. aeruginosa infections.


Assuntos
Anticorpos Antibacterianos/imunologia , Antígenos de Bactérias/imunologia , Toxinas Bacterianas/imunologia , Queimaduras/imunologia , Imunoglobulinas/imunologia , Pneumonia/imunologia , Proteínas Citotóxicas Formadoras de Poros/imunologia , Infecções por Pseudomonas/imunologia , Pseudomonas aeruginosa/imunologia , Animais , Queimaduras/microbiologia , Galinhas/imunologia , Galinhas/microbiologia , Modelos Animais de Doenças , Feminino , Imunização/métodos , Camundongos , Camundongos Endogâmicos BALB C , Pneumonia/microbiologia , Vacinação/métodos , Virulência/imunologia
5.
An Bras Dermatol ; 94(4): 405-410, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31644611

RESUMO

BACKGROUND: A high prevalence of leprosy among children under 15 years of age indicates the need to implement actions to prevent new cases of the disease. Serological tests have been developed with the aim of helping to control the disease by indicating, through seropositivity, the presence of infection. OBJECTIVE: To analyze the prevalence and factors associated with seropositivity rate for anti-NDO-LID antibodies in children under 15 years of age, contacts of leprosy patients. METHOD: We performed a cross-sectional study with 210 children under 15 years old of age. Of them, 50 were household contacts and 160 were neighborhood contacts living in the municipality of Cuiabá, state of Mato Grosso, in 2016. The data were obtained from interviews and the NDO-LID rapid test during home visits from February to July 2016. For the analysis, we used Poisson regression and prevalence ratio. RESULTS: Seropositivity in contacts was 6.2%. Variables associated with seropositive tests included sex (PR = 1.05; 95% CI: 1.01 - 1.08), race/skin color (PR = 0.95; 95% CI: 0.90 - 0.99), residence area (PR = 1.05; 95% CI: 1.01 - 1.09), and number of people per household (PR = 1.06; 95% CI: 1.02 - 1.08). STUDY LIMITATIONS: The small sample size, besides leading to wide confidence intervals, may have been a limitation for the identification of associated factors. CONCLUSIONS: The prevalence of seropositivity was high. Variables associated with NDO-LID seropositivity included female sex, not to be brown skinned, live in urban areas, and live with five or more people.


Assuntos
Anticorpos Anti-Idiotípicos/imunologia , Anticorpos Antibacterianos/imunologia , Hanseníase/epidemiologia , Hanseníase/imunologia , Adolescente , Distribuição por Idade , Fatores Etários , Anticorpos Anti-Idiotípicos/sangue , Anticorpos Antibacterianos/sangue , Brasil/epidemiologia , Criança , Pré-Escolar , Estudos Transversais , Características da Família , Feminino , Humanos , Lactente , Masculino , Características de Residência , Testes Sorológicos/métodos , Distribuição por Sexo , Fatores Socioeconômicos
6.
Mol Immunol ; 114: 612-619, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31542606

RESUMO

Enterohemorrhagic Escherichia coli (EHEC) are known as the gastrointestinal pathogens and major causes of enterohemorrhagic colitis since decades ago. There is no efficient approved vaccine against EHEC O157 and non-O157. In the present study, a recombinant candidate vaccine against enterohemorrhagic E. coli (EHEC) O157:H7 entrapped in the sodium alginate and PLGA nanoparticles and the efficiency of the immunization of these formulations were investigated. nanoparticles due to their properties like controlled cargoes release, adjuvanticity, cargo protection, increased bioavailability, etc have been noticed for drug delivery. A chimeric protein composed of HcpA, EspA, Tir and Stx2B antigens was designed, recombinantly expressed, purified and entrapped in nanoparticles. BALB/c mice were administrated with nano-formulated and free proteins. IgG titer, EHEC fecal shedding and the ability of the immune sera to neutralize Stx toxin and inhibit the bacterial attachment to Caco-2 cells were analyzed. Fecal shedding analysis demonstrated that the colonization of the bacteria in the intestine of the mice was reduced significantly (P > 0.01). Immune mice were able to tolerate up to 200 LD50 of the active Stx toxin. About 80% of the bacterial binding capacity to Caco-2 cells was declined, especially in groups immunized with nano-formulations. Considering the importance of EHEC, especially O157 serotype, on public health and the other hand, the lack of an efficient vaccine in this regard, delivery of HETS candidate vaccine with NPs can be applied to prevent the infection by the pathogen.


Assuntos
Alginatos/química , Formação de Anticorpos/imunologia , Infecções por Escherichia coli/imunologia , Escherichia coli O157/imunologia , Nanopartículas/química , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/química , Proteínas Recombinantes de Fusão/imunologia , Animais , Anticorpos Antibacterianos/imunologia , Células CACO-2 , Linhagem Celular Tumoral , Escherichia coli Êntero-Hemorrágica/imunologia , Proteínas de Escherichia coli/imunologia , Feminino , Humanos , Imunização/métodos , Imunoglobulina G/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/imunologia , Proteínas Recombinantes de Fusão/química , Vacinas Sintéticas/imunologia
7.
Lett Appl Microbiol ; 69(5): 366-372, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31508837

RESUMO

We evaluate the efficacy of recombinant DNA vaccine ABA392 against haemorrhagic septicaemia infection through intranasal administration route by targeting the mucosal immunity. The DNA vaccine was constructed and subjected to animal study using the Sprague Dawley (SD) rat. The study was divided into two major parts: (i) active and (ii) passive immunization studies, involving 30 animals for each part. Each group was then divided into five test groups: two test samples G1 and G2 with 50 and 100 µg ml-1 purified DNA vaccine; one positive control G5 with 106  CFU per ml formalin-killed PMB2; and two negative controls, G3 and G4 with normal saline and pVAX1 vector. Both studies were conducted for the determination of immunogenicity by total white blood cell count (TWBC), indirect ELISA and histopathological changes for the presence of the bronchus-associated lymphoid tissue (BALT). Our findings demonstrate that TWBC, IgA and IgG increased after each of the three vaccination regimes: groups G1, G2 and G5. Test samples G1 and G2 showed significant differences (P < 0·05) compared to the negative controls, G3 and G4, but no significant differences from the positive control G5. Groups G1, G2 and G5 showed more formation of BALT compared to the negative controls, G3 and G4. Our results show that intranasal inoculation of recombinant DNA vaccine ABA392 can provoke mucosal immunity which makes it a potential prophylactic against HS. SIGNIFICANCE AND IMPACT OF THE STUDY: New approach of combating haemorrhagic septicaemia disease among bovines by recombinant DNA vaccine is crucial to overcome the loss of edible products from the infected bovines. DNA vaccine can potentially serve as a better immunogen which would elicit both cellular and humoral immunity, and it is also stable for its molecular reproduction. This research report demonstrates an effective yet simple way of administering the DNA vaccine via the intranasal route in rats, to provoke the mucosal immunity through the development of immunoglobulins IgA, IgG and bronchus-associated lymphoid tissue which guard as the first-line defence at the host's mucosal lining.


Assuntos
Vacinas Bacterianas/administração & dosagem , Doenças dos Bovinos/prevenção & controle , Septicemia Hemorrágica/veterinária , Pasteurella multocida/imunologia , Vacinas de DNA/administração & dosagem , Administração Intranasal , Animais , Anticorpos Antibacterianos/imunologia , Vacinas Bacterianas/genética , Vacinas Bacterianas/imunologia , Bovinos , Doenças dos Bovinos/imunologia , Doenças dos Bovinos/microbiologia , DNA Recombinante/administração & dosagem , DNA Recombinante/genética , DNA Recombinante/imunologia , Ensaio de Imunoadsorção Enzimática , Septicemia Hemorrágica/imunologia , Septicemia Hemorrágica/microbiologia , Septicemia Hemorrágica/prevenção & controle , Imunização Passiva , Masculino , Pasteurella multocida/genética , Ratos , Ratos Sprague-Dawley , Vacinas de DNA/genética , Vacinas de DNA/imunologia
8.
Ann Agric Environ Med ; 26(3): 392-395, 2019 Sep 19.
Artigo em Inglês | MEDLINE | ID: mdl-31559791

RESUMO

Existing research for using the protective antigen (PA) of Bacillus anthracis as a vaccine component shows that protection against anthrax may be obtained using fragments of this protein. The aim of the research is to check whether the selected protein fragment of the protective antigen (domain 4) encoded by an appropriate nucleotide sequence of gene pag of B. anthracis, was expressed in the bacterial system of E. coli. In order to examine the selected sequence of the pag gene, a PCR reaction and a highly effective TOPO cloning strategy were used, followed by purification of the recombinant proteins and their detection by a western-blot method. In the planning of the PA4 antigen expression a higher level of effectiveness in production of small protein - domain 4 - was anticipated. As a result, the 139 amino acids protein fragment of B. anthracis PA (domain 4) was isolated. The research may have found the basis for in vivo research aimed at finding potential anthrax vaccine components.


Assuntos
Vacinas contra Antraz/imunologia , Antraz/microbiologia , Antígenos de Bactérias/imunologia , Bacillus anthracis/imunologia , Toxinas Bacterianas/imunologia , Animais , Antraz/imunologia , Antraz/prevenção & controle , Vacinas contra Antraz/administração & dosagem , Vacinas contra Antraz/genética , Vacinas contra Antraz/isolamento & purificação , Anticorpos Antibacterianos/imunologia , Anticorpos Neutralizantes/imunologia , Antígenos de Bactérias/administração & dosagem , Antígenos de Bactérias/genética , Antígenos de Bactérias/isolamento & purificação , Bacillus anthracis/química , Bacillus anthracis/genética , Toxinas Bacterianas/administração & dosagem , Toxinas Bacterianas/genética , Toxinas Bacterianas/isolamento & purificação , Western Blotting , Escherichia coli/genética , Escherichia coli/metabolismo , Expressão Gênica , Humanos , Imunização , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Domínios Proteicos
9.
APMIS ; 127(12): 753-763, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31512768

RESUMO

Iron uptake system is expressed in early stages of Acinetobacter baumannii infections under iron-restricted conditions. This study is aimed at the evaluation of immuno-protectivity of BfnH in comparison with BauA in both mature and selected fragmental proteins. The study was designed in single and combined forms of antigens. BfnH is presented in 3472 strains of A. baumannii with more than 97% identity. The preliminary immune-informatics analysis of this protein indicated a region from the ß-barrel domain including exposed loops 2-5, with antigenic score comparable to that of BfnH. There was a significant rise in the specific IgG response in all test groups. The bacterial challenge with a lethal dose of A. baumannii demonstrated partial protection of whole proteins which coincides with a significant reduction in the bacterial population colonized in the main organs and an increase in the survival level. Passive immunization of the mice brought about 50% survival in the mice groups immunized with BfnH and with a combination of BfnH and BauA. The protectivity of siderophore receptors suggests their potential immunogenic role that could be considered as a component of multivalent subunit vaccine candidates against A. baumannii.


Assuntos
Infecções por Acinetobacter/prevenção & controle , Acinetobacter baumannii/imunologia , Proteínas da Membrana Bacteriana Externa/imunologia , Imunização , Receptores de Superfície Celular/imunologia , Infecções por Acinetobacter/imunologia , Infecções por Acinetobacter/microbiologia , Animais , Anticorpos Antibacterianos/sangue , Anticorpos Antibacterianos/imunologia , Carga Bacteriana , Proteínas da Membrana Bacteriana Externa/química , Proteínas da Membrana Bacteriana Externa/genética , Vacinas Bacterianas/administração & dosagem , Vacinas Bacterianas/imunologia , Epitopos de Linfócito B , Feminino , Imunogenicidade da Vacina , Camundongos Endogâmicos BALB C , Receptores de Superfície Celular/química , Receptores de Superfície Celular/genética
10.
Infect Immun ; 87(12)2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31548318

RESUMO

The metabolic inhibition (MI) test is a classic test for the identification of mycoplasmas, used for measuring the growth-inhibiting antibodies directed against acid-producing mycoplasmas, although their mechanism still remains obscure. To determine the major antigens involved in the immune killing of Mycoplasma bovis, we used a pulldown assay with anti-M. bovis antibodies as bait and identified nine major antigens. Among these antigens, we performed the MI test and determined that the growth of M. bovis could be inhibited effectively in the presence of complement by antibodies against specifically membrane protein P81 or UgpB in the presence of complement. Using a complement killing assay, we demonstrated that M. bovis can be killed directly by complement and that antibody-dependent complement-mediated killing is more effective than that by complement alone. Complement lysis and scanning electron microscopy results revealed M. bovis rupture in the presence of complement. Together, these results suggest that the metabolic inhibition of M. bovis is antibody-dependent complement-mediated killing. This study provides new insights into mycoplasma killing by the complement system and may guide future vaccine development studies for the treatment of mycoplasma infection. Furthermore, our findings also indicate that mycoplasmas may be an appropriate new model for studying the lytic activity of membrane attack complex (MAC).


Assuntos
Anticorpos Antibacterianos/imunologia , Proteínas do Sistema Complemento/imunologia , Proteínas de Membrana/imunologia , Infecções por Mycoplasma/veterinária , Mycoplasma bovis/imunologia , Animais , Bovinos , Complexo de Ataque à Membrana do Sistema Complemento/imunologia , Microscopia Eletrônica de Varredura , Infecções por Mycoplasma/imunologia , Coelhos
11.
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
12.
J Med Microbiol ; 68(11): 1629-1640, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31553301

RESUMO

Introduction. ML1899 is conserved in all mycobacterium sp. and is a middle member of mle-ML1898 operon involved in mycolic acid modification.Aim. In the present study attempts were made to characterize ML1899 in detail.Methodology. Bioinformatics tools were used for prediction of active-site residues, antigenic epitopes and a three-dimensional model of protein. The gene was cloned, expressed and purified as His-tagged protein in Escherichia coli for biophysical/biochemical characterization. Recombinant protein was used to treat THP-1 cells to study change in production of nitric oxide (NO), reactive oxygen species (ROS), cytokines and chemokines using flowcytometry/ELISA.Results. In silico analysis predicted ML1899 as a member of α/ß hydrolase family with GXSXG-motif and Ser126, His282, Asp254 as active-site residues that were confirmed by site-directed mutagensis. ML1899 exhibited esterase activity. It hydrolysed pNP-butyrate as optimum substrate at pH 8.0 and 50 °C with 5.56 µM-1 min-1 catalytic efficiency. The enzyme exhibited stability up to 60 °C temperature and between pH 6.0 to 9.0. K m, V max and specific activity of ML1899 were calculated to be 400 µM, 40 µmoles min-1 ml-1 and 27 U mg- 1, respectively. ML1899 also exhibited phospholipase activity. The protein affected the survival of macrophages when treated at higher concentration. ML1899 enhanced ROS/NO production and up-regulated pro-inflammatory cytokines and chemokine including TNF-α, IFN-γ, IL-6 and IL-8 in macrophages. ML1899 was also observed to elicit humoral response in 69 % of leprosy patients.Conclusion. These results suggested that ML1899, an esterase could up-regulate the immune responses in favour of macrophages at a low concentration but kills the THP-1 macrophages cells at a higher concentration.


Assuntos
Proteínas de Bactérias/imunologia , Esterases/imunologia , Hanseníase/microbiologia , Mycobacterium leprae/enzimologia , Sequência de Aminoácidos , Anticorpos Antibacterianos/imunologia , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Citocinas/genética , Citocinas/imunologia , Estabilidade Enzimática , Esterases/química , Esterases/genética , Feminino , Humanos , Concentração de Íons de Hidrogênio , Cinética , Hanseníase/imunologia , Macrófagos/imunologia , Macrófagos/microbiologia , Masculino , Mycobacterium leprae/química , Mycobacterium leprae/genética , Mycobacterium leprae/imunologia , Óxido Nítrico/imunologia , Espécies Reativas de Oxigênio/imunologia , Alinhamento de Sequência
13.
Infect Immun ; 87(12)2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31501249

RESUMO

Coxiella burnetii, the etiological agent of Q fever, is a Gram-negative bacterium transmitted to humans by inhalation of contaminated aerosols. Acute Q fever is often self-limiting, presenting as a febrile illness that can result in atypical pneumonia. In some cases, Q fever becomes chronic, leading to endocarditis that can be life threatening. The formalin-inactivated whole-cell vaccine (WCV) confers long-term protection but has significant side effects when administered to presensitized individuals. Designing new vaccines against C. burnetii remains a challenge and requires the use of clinically relevant modes of transmission in appropriate animal models. We have developed a safe and reproducible C. burnetii aerosol challenge in three different animal models to evaluate the effects of pulmonary acquired infection. Using a MicroSprayer aerosolizer, BL/6 mice and Hartley guinea pigs were infected intratracheally with C. burnetii Nine Mile phase I (NMI) and demonstrated susceptibility as determined by measuring bacterial growth in the lungs and subsequent dissemination to the spleen. Histological analysis of lung tissue showed significant pathology associated with disease, which was more severe in guinea pigs. Infection using large-particle aerosol (LPA) delivery was further confirmed in nonhuman primates, which developed fever and pneumonia. We also demonstrate that vaccinating mice and guinea pigs with WCV prior to LPA challenge is capable of eliciting protective immunity that significantly reduces splenomegaly and the bacterial burden in spleen and lung tissues. These data suggest that these models can have appreciable value in using the LPA delivery system to study pulmonary Q fever pathogenesis as well as designing vaccine countermeasures to C. burnetii aerosol transmission.


Assuntos
Vacinas Bacterianas/imunologia , Coxiella burnetii/imunologia , Pulmão/microbiologia , Febre Q/veterinária , Vacinas de Produtos Inativados/imunologia , Administração Intranasal , Animais , Anticorpos Antibacterianos/imunologia , Vacinas Bacterianas/administração & dosagem , Modelos Animais de Doenças , Feminino , Cobaias , Pulmão/imunologia , Macaca mulatta , Camundongos , Camundongos Endogâmicos C57BL , Febre Q/imunologia , Febre Q/prevenção & controle , Baço/imunologia , Baço/microbiologia , Vacinas de Produtos Inativados/administração & dosagem
14.
Clin Lab ; 65(12)2019 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-31414587

RESUMO

BACKGROUND: Screening and timely treatment of precancerous gastric cancer diseases or of gastric cancer in the early stages has important significance in reducing the incidence and mortality of gastric cancer. Gastroscopy and histopathological biopsy are still the gold standards for the diagnosis of gastric diseases. But the application of astroscopy for the screening and diagnosis of gastric diseases is limited. In recent years, serum pepsinogen (PG), gastrin, and Helicobacter pylori (H. pylori) IgG antibodies have become indicators for "serological biopsy" of the gastric mucosa. METHODS: From January 2016 to January 2018, a total of 2,394 patients with digestive tract symptoms underwent gastroscopy. According to the endoscopic examination and pathological diagnosis, there were four case groups: 1,376 cases of chronic non-atrophic gastritis, 708 cases of chronic atrophic gastritis, 265 cases of gastric ulcer, and 45 cases of gastric cancer. Serological gastric biopsies were performed and analyzed. RESULTS: The serum levels of PGI in the chronic atrophic gastritis group was significantly lower than that in the chronic non-atrophic gastritis group, gastric ulcer group, and gastric cancer group (p < 0.05). The serum levels of PGII and G-17 in the gastric cancer group were significantly higher than those in the chronic non-atrophic Gastritis group, chronic atrophic gastritis group, and gastric ulcer group (p < 0.05). The PGR in the gastric cancer group was significantly lower than that in the chronic non-atrophic gastritis group, chronic atrophic Gastritis group, and gastric ulcer group (p < 0.05). The H. pylori positive rates in the chronic atrophic gastritis group and gastric cancer group were higher than those in the chronic non-atrophic gastritis group and gastric ulcer Group (p < 0.05). CONCLUSIONS: Serological gastric biopsy is closely correlated to gastric mucosal disease and can be used as a Screening tool in gastric disease.


Assuntos
Anticorpos Antibacterianos/sangue , Biomarcadores/sangue , Mucosa Gástrica/metabolismo , Gastrinas/sangue , Pepsinogênio A/sangue , Neoplasias Gástricas/sangue , Adulto , Idoso , Idoso de 80 Anos ou mais , Anticorpos Antibacterianos/imunologia , Biópsia , Feminino , Mucosa Gástrica/microbiologia , Mucosa Gástrica/patologia , Infecções por Helicobacter/sangue , Infecções por Helicobacter/diagnóstico , Infecções por Helicobacter/microbiologia , Helicobacter pylori/imunologia , Helicobacter pylori/fisiologia , Humanos , Masculino , Pessoa de Meia-Idade , Sensibilidade e Especificidade , Neoplasias Gástricas/patologia , Adulto Jovem
15.
Clin Lab ; 65(8)2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-31414746

RESUMO

BACKGROUND: Foreign body aspiration is a rare entity in adults. We presented an adult case with recurrent pulmonary infection firstly misdiagnosed as tuberculosis, which proved as foreign body aspiration in the left main stem bronchus by bronchoscopy. METHODS: Appropriate laboratory tests are carried out. The chest CT scan and bronchoscopy were performed for diagnosis. RESULTS: Serum sedimentation was increased and tuberculosis antibody was positive. The chest CT scan showed left lung consolidation and small pleural exudate on the left side. Significant calcification can be seen near the left main bronchus. The bronchoscopy demonstrated plenty of yellow sputum in left main bronchus and a peanut shell completely obstructed the left main bronchus and peripheral granulation tissue hyperplasia. The peanut shell was removed and the left main trachea was unobstructed. CONCLUSIONS: When a patient has recurrent pulmonary infection, especially at the same site, physicians should pay attention to airway obstruction caused by foreign body, cancer and other causes of airway stenosis. Bronchoscopy is crucial for the ultimate diagnosis.


Assuntos
Sedimentação Sanguínea , Erros de Diagnóstico , Corpos Estranhos/diagnóstico , Pulmão/diagnóstico por imagem , Mycobacterium tuberculosis/imunologia , Tuberculose/diagnóstico , Idoso , Anticorpos Antibacterianos/sangue , Anticorpos Antibacterianos/imunologia , Brônquios/microbiologia , Brônquios/patologia , Broncoscopia , Humanos , Pulmão/microbiologia , Masculino , Mycobacterium tuberculosis/fisiologia , Tomografia Computadorizada por Raios X , Traqueia/microbiologia , Traqueia/patologia , Tuberculose/sangue , Tuberculose/microbiologia
16.
Vet Microbiol ; 235: 270-279, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31383312

RESUMO

Lawsonia intracellularis is an obligate intracellular microorganism and the causative agent of porcine proliferative enteropathy. Due to its obligate intracellular nature, characterization of antigens and proteins involved in host-pathogen interaction and immune recognition have been difficult to achieve using conventional microbiological techniques. In this work, we used 2-dimensional gel electrophoresis coupled with Western-immunoblotting, mass spectrometry and bioinformatics to identify bacterial proteins that interact in vitro with pig intestinal cells (IPEC-1), have immunogenic properties and the potential to be used as subunit vaccine antigens. We detected eleven immunogenic bacterial proteins from which fliC (LI0710), LI1153 (annotated by NCBI as Putative protein N), and LI0649 (annotated as autotransporter) were predicted to be expressed on the outer membrane while LI0169 (oppA; annotated as ABC dipeptide transport system) was predicted to be periplasmic with a transmembrane domain forming a central pore through the plasma membrane. Genes coding for these four proteins were cloned and expressed in Escherichia coli and the corresponding recombinant proteins were purified using affinity chromatography. Porcine hyperimmune serum against whole Lawsonia lysate established that all four recombinant proteins were immunogenic. Further, rabbit hyperimmune sera generated against the vaccine strain of L. intracellularis and rabbit serum specific for each recombinant protein showed an inhibitory effect on the attachment and penetration of live, avirulent L. intracellularis, thus indicating that each protein is a potential neutralizing antibody target and a candidate for subunit vaccine formulation.


Assuntos
Anticorpos Antibacterianos/imunologia , Anticorpos Neutralizantes/imunologia , Proteínas de Bactérias/imunologia , Vacinas Bacterianas/imunologia , Infecções por Desulfovibrionaceae/veterinária , Lawsonia (Bactéria)/imunologia , Animais , Proteínas de Bactérias/genética , Western Blotting , Linhagem Celular , Biologia Computacional , Infecções por Desulfovibrionaceae/imunologia , Infecções por Desulfovibrionaceae/prevenção & controle , Feminino , Intestinos/citologia , Intestinos/microbiologia , Espectrometria de Massas , Proteômica , Coelhos , Proteínas Recombinantes/imunologia , Suínos , Doenças dos Suínos/imunologia , Doenças dos Suínos/microbiologia , Doenças dos Suínos/prevenção & controle , Vacinas de Subunidades/imunologia
17.
Int J Infect Dis ; 87: 54-59, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31419482

RESUMO

OBJECTIVE: The epidemiology and role of the anti-PcrV titer in non-cystic fibrosis patients with Pseudomonas aeruginosa airway tract infections is not fully understood. This study was performed to compare the anti-PcrV titers of patients with and without P. aeruginosa respiratory tract infections. METHODS: This prospective cohort study was conducted at Hokkaido University Hospital in Japan. Participants had blood and sputum specimens collected on admission. They were divided into two groups based on their sputum culture results. Those with a P. aeruginosa infection were assigned to the P. aeruginosa (PA) group and those without a P. aeruginosa infection were assigned to the non-PA group. Serum anti-PcrV titers were measured using a validated ELISA. RESULTS: Of the 44 participants, 15 were assigned to the PA group and 29 were assigned to the non-PA group. In the PA group, 10/15 participants (66.7%) had an anti-PcrV titer >1000ng/ml compared to 3/29 participants (10.3%) in the non-PA group (p<0.001). In the PA group, two of the five participants with an anti-PcrV titer <1000 ng/ml died of recurrent P. aeruginosa pneumonia; the other three participants did not develop pneumonia. CONCLUSION: The anti-PcrV titers in participants with P. aeruginosa infection varied considerably. Patients with low anti-PcrV titers and refractory P. aeruginosa infections need to be monitored closely.


Assuntos
Anticorpos Antibacterianos/imunologia , Antígenos de Bactérias/imunologia , Infecções por Pseudomonas/imunologia , Pseudomonas aeruginosa/imunologia , Infecções Respiratórias/imunologia , Idoso , Idoso de 80 Anos ou mais , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Japão , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Infecções por Pseudomonas/microbiologia , Pseudomonas aeruginosa/genética , Pseudomonas aeruginosa/fisiologia , Sistema Respiratório/microbiologia , Infecções Respiratórias/microbiologia
18.
Cochrane Database Syst Rev ; 8: CD011871, 2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-31425612

RESUMO

BACKGROUND: Early diagnosis of leptospirosis may contribute to the effectiveness of antimicrobial therapy and early outbreak recognition. Nucleic acid and antigen detection tests have the potential for early diagnosis of leptospirosis. With this systematic review, we assessed the sensitivity and specificity of nucleic acid and antigen detection tests. OBJECTIVES: To determine the diagnostic test accuracy of nucleic acid and antigen detection tests for the diagnosis of human symptomatic leptospirosis. SEARCH METHODS: We searched electronic databases including MEDLINE, Embase, the Cochrane Library, and regional databases from inception to 6 July 2018. We did not apply restrictions to language or time of publication. SELECTION CRITERIA: We included diagnostic cross-sectional studies and case-control studies of tests that made use of nucleic acid and antigen detection methods in people suspected of systemic leptospirosis. As reference standards, we considered the microscopic agglutination test alone (which detects antibodies against leptospirosis) or in a composite reference standard with culturing or other serological tests. Studies were excluded when the controls were healthy individuals or when there were insufficient data to calculate sensitivity and specificity. DATA COLLECTION AND ANALYSIS: At least two review authors independently extracted data from each study. We used the revised Quality Assessment of Diagnostic Accuracy Studies tool (QUADAS-2) to assess risk of bias. We calculated study-specific values for sensitivity and specificity with 95% confidence intervals (CI) and pooled the results in a meta-analysis when appropriate. We used the bivariate model for index tests with one positivity threshold, and we used the hierarchical summary receiver operating characteristic model for index tests with multiple positivity thresholds. As possible sources of heterogeneity, we explored: timing of index test, disease prevalence, blood sample type, primers or target genes, and the real-time polymerase chain reaction (PCR) visualisation method. These were added as covariates to the meta-regression models. MAIN RESULTS: We included 41 studies evaluating nine index tests (conventional PCR (in short: PCR), real-time PCR, nested PCR, PCR performed twice, loop-mediated isothermal amplification, enzyme-linked immunosorbent assay (ELISA), dot-ELISA, immunochromatography-based lateral flow assay, and dipstick assay) with 5981 participants (1834 with and 4147 without leptospirosis). Methodological quality criteria were often not reported, and the risk of bias of the reference standard was generally considered high. The applicability of findings was limited by the frequent use of frozen samples. We conducted meta-analyses for the PCR and the real-time PCR on blood products.The pooled sensitivity of the PCR was 70% (95% CI 37% to 90%) and the pooled specificity was 95% (95% CI 75% to 99%). When studies with a high risk of bias in the reference standard domain were excluded, the pooled sensitivity was 87% (95% CI 44% to 98%) and the pooled specificity was 97% (95% CI 60% to 100%). For the real-time PCR, we estimated a summary receiver operating characteristic curve. To illustrate, a point on the curve with 85% specificity had a sensitivity of 49% (95% CI 30% to 68%). Likewise, at 90% specificity, sensitivity was 40% (95% CI 24% to 59%) and at 95% specificity, sensitivity was 29% (95% CI 15% to 49%). The median specificity of real-time PCR on blood products was 92%. We did not formally compare the diagnostic test accuracy of PCR and real-time PCR, as direct comparison studies were lacking. Three of 15 studies analysing PCR on blood products reported the timing of sample collection in the studies included in the meta-analyses (range 1 to 7 days postonset of symptoms), and nine out of 16 studies analysing real-time PCR on blood products (range 1 to 19 days postonset of symptoms). In PCR studies, specificity was lower in settings with high leptospirosis prevalence. Other investigations of heterogeneity did not identify statistically significant associations. Two studies suggested that PCR and real-time PCR may be more sensitive on blood samples collected early in the disease stage. Results of other index tests were described narratively. AUTHORS' CONCLUSIONS: The validity of review findings are limited and should be interpreted with caution. There is a substantial between-study variability in the accuracy of PCR and real-time PCR, as well as a substantial variability in the prevalence of leptospirosis. Consequently, the position of PCR and real-time PCR in the clinical pathway depends on regional considerations such as disease prevalence, factors that are likely to influence accuracy, and downstream consequences of test results. There is insufficient evidence to conclude which of the nucleic acid and antigen detection tests is the most accurate. There is preliminary evidence that PCR and real-time PCR are more sensitive on blood samples collected early in the disease stage, but this needs to be confirmed in future studies.


Assuntos
Anticorpos Antibacterianos/imunologia , Leptospira/imunologia , Leptospirose/diagnóstico , Ácidos Nucleicos/sangue , Reação em Cadeia da Polimerase/métodos , Ensaio de Imunoadsorção Enzimática/métodos , Humanos , Leptospirose/sangue , Curva ROC , Sensibilidade e Especificidade
19.
J Dermatol ; 46(10): 853-858, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31432529

RESUMO

In patients with lepromatous leprosy, Mycobacterium leprae is often observed inside the human microvascular endothelial cells (HMVEC) surrounding Schwann cells (SC) at the site of lesions in the peripheral nerves. Based on this observation, it is considered that the nasal mucous may be the invasion pathway for M. leprae and HMVEC serve as an important reservoir for the bacteria before they invade SC. In light of previous research which revealed that Mce1A protein mediates bacterial invasion into nasal epithelial cells and HMVEC, we conducted a study to determine whether the invasion of M. leprae into HMVEC can be suppressed by blocking the Mce1A protein. In this study, we analyzed bacterial invasive activity by adding recombinant Escherichia coli, which express the active region (InvX:72 a.a.) of Mce1A protein on their external membrane, into cultured HMVEC, using the adhesin involved in the diffuse adherence mechanism. The number of bacteria that invaded into the cells was then measured by a colony counting method. The active region of Mce1A was divided into four sections, and hyperimmune antisera was prepared for each section for analyzing the inhibitory effect against invasion. The invasive activity was suppressed by antibodies against InvX regions 1-24 a.a., 25-46 a.a. and 58-72 a.a. This suggests that the InvX regions 1-24 a.a., 25-46 a.a. and 58-72 a.a. of Mce1A protein play an important role in the invasion of M. leprae into HMVEC and that it may be possible to suppress entry of M. leprae in HMVEC with antibodies against these regions.


Assuntos
Anticorpos Antibacterianos/imunologia , Proteínas de Bactérias/imunologia , Células Endoteliais/microbiologia , Hanseníase/imunologia , Mycobacterium leprae/imunologia , Animais , Anticorpos Antibacterianos/isolamento & purificação , Proteínas de Bactérias/genética , Linhagem Celular , Contagem de Colônia Microbiana , Humanos , Soros Imunes/imunologia , Soros Imunes/isolamento & purificação , Hanseníase/microbiologia , Hanseníase/prevenção & controle , Mycobacterium leprae/patogenicidade , Coelhos , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia
20.
Analyst ; 144(17): 5277-5283, 2019 Aug 16.
Artigo em Inglês | MEDLINE | ID: mdl-31369000

RESUMO

The faster a disease can be diagnosed, the sooner effective treatment can be initiated, motivating a drive to replace standard laboratory techniques with point-of-care technologies that return answers in minutes rather than hours. Thus motivated, we describe the development of an E-DNA scaffold sensor for the rapid and convenient measurement of antibodies diagnostic of syphilis. To achieve this (and in contrast to previous sensors of this class, which relied on single, linear epitopes for detection), we utilized a near full-length antigen as the sensor's recognition element, allowing us to simultaneously display multiple epitopes. The resultant sensor is able to detect antibodies against Treponema pallidum pallidum, the causative agent of syphilis, at clinically relevant concentrations in samples in less than 10 min. Preliminary results obtained using sero-positive and sero-negative human samples suggest the clinical sensitivity and specificity of the approach compare well to current gold-standard tests, while being simple and rapid enough to deploy at the point of care.


Assuntos
Anticorpos Antibacterianos/sangue , DNA/química , Sífilis/diagnóstico , Anticorpos Antibacterianos/imunologia , Antígenos de Bactérias/imunologia , Proteínas de Bactérias/imunologia , Sequência de Bases , Técnicas Eletroquímicas/instrumentação , Técnicas Eletroquímicas/métodos , Eletrodos , Escherichia coli/genética , Humanos , Ácidos Nucleicos Imobilizados/química , Azul de Metileno/química , Oxirredução , Treponema pallidum/química
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