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1.
Biochimie ; 167: 207-216, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31628976

RESUMO

Oligopeptidases B (OPB) belong to the S9 prolyl oligopeptidase family and are expressed in prokaryotes, some eukaryotes and in some higher plants. OPB is not found in any of the mammalian genomes available to date. Evidences indicate that OPB participates in the infections caused by trypanosomatids Trypanosoma cruzi, Leishmania spp. and Trypanosoma brucei spp and therefore it is considered an important virulence factor. Trypanosomatids from the genera Leishmania and Trypanosoma also present other OPB, named OPB2. A more accurate investigation of trypanosomatid OPB sequences brought attention to what could be a third OPB sequence (OPB3). This review aims to discuss biochemical, structural, phylogenetic and functional properties of OPB and its potential as target for the development of drugs against Chagas disease, leishmaniasis and African trypanosomiasis.


Assuntos
Leishmania/enzimologia , Serina Endopeptidases , Trypanosoma brucei brucei/enzimologia , Trypanosoma cruzi/enzimologia , Fatores de Virulência , Animais , Doença de Chagas/parasitologia , Humanos , Leishmaniose/parasitologia , Mamíferos , Proteínas de Protozoários/química , Proteínas de Protozoários/imunologia , Serina Endopeptidases/química , Serina Endopeptidases/classificação , Serina Endopeptidases/imunologia , Tripanossomíase Africana/parasitologia , Fatores de Virulência/química , Fatores de Virulência/classificação , Fatores de Virulência/imunologia
2.
J Appl Microbiol ; 127(6): 1848-1858, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31509624

RESUMO

AIMS: The aim of this study was to develop a rapid detection and differentiation method for pathogenic Listeria species in stone fruits. METHODS AND RESULTS: We utilized activated charcoal enrichment media (ACM) to induce overexpression and hypersecretion of pathogenic Listeria virulence proteins which can subsequently be detected via immunoblot analysis. Plum and nectarine slices spiked with either L. monocytogenes or L. ivanovii were incubated in pre-enrichment broth followed by enrichment in ACM. Secreted proteins were precipitated and subjected to SDS-PAGE and immunoblot analysis using a combination of L. monocytogenes-specific antibody (α-listeriolysin O) and antibody specific for both L. monocytogenes and L. ivanovii (α-Internalin C). As low as 1 CFU per gram of L. monocytogenes in plum and nectarine was detected, whereas a detection limit of 10 CFU per gram was achieved for L. ivanovii in each food tested following a 20-h enrichment period. Nonpathogenic Listeria species and non-Listeria bacterial pathogens tested were negative. CONCLUSIONS: These results demonstrate the highly sensitive and specific nature of the detection method for pathogenic Listeria in stone fruits using activated charcoal enrichment as well as the capability to discriminate between L. monocytogenes and L. ivanovii. SIGNIFICANCE AND IMPACT OF THE STUDY: This method is the first to identify and differentiate L. monocytogenes and L. ivanovii in select stone fruit enrichments within 24 h using immunological techniques. The rapidity and sensitivity of the method could aid in the reduction of exposure to the public in the event of an outbreak and expedite the administration of appropriate antibiotics to infected individuals.


Assuntos
Microbiologia de Alimentos/métodos , Frutas/microbiologia , Listeria/isolamento & purificação , Carvão Vegetal/química , Meios de Cultura/química , Humanos , Limite de Detecção , Listeria/classificação , Listeria/imunologia , Prunus/microbiologia , Fatores de Virulência/análise , Fatores de Virulência/imunologia
3.
Infect Immun ; 87(12)2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31527122

RESUMO

We previously demonstrated that recombinant protein PAc could be administered as an anticaries vaccine. However, the relatively weak immunogenicity of PAc limits its application. In the present study, we investigated the effect of two adjuvant combinations of chitosan plus Pam3CSK4 (chitosan-Pam3CSK4) and of chitosan plus monophosphoryl lipid A (chitosan-MPL) in the immune responses to the PAc protein in vivo and in vitro PAc-chitosan-Pam3CSK4 or PAc-chitosan-MPL promoted significantly higher PAc-specific antibody titers in serum and saliva, inhibited Streptococcus mutans colonization onto the tooth surfaces, and endowed better protection effect with significantly less caries activities than PAc alone. Chitosan-Pam3CSK4 and chitosan-MPL showed no statistically significant differences. In conclusion, our study demonstrated that the chitosan-Pam3CSK4 and chitosan-MPL combinations are promising for anticaries vaccine development.


Assuntos
Vacinas Bacterianas/imunologia , Quitosana/farmacologia , Cárie Dentária/prevenção & controle , Lipídeo A/análogos & derivados , Lipopeptídeos/farmacologia , Streptococcus mutans/imunologia , Adjuvantes Imunológicos , Animais , Cárie Dentária/microbiologia , Feminino , Imunogenicidade da Vacina/imunologia , Imunoglobulina A Secretora/análise , Lipídeo A/imunologia , Proteínas de Membrana/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Proteínas NLR/agonistas , Streptococcus mutans/patogenicidade , Receptores Toll-Like/agonistas , Vacinas Sintéticas/imunologia , Fatores de Virulência/imunologia
4.
Int J Mol Sci ; 20(18)2019 Sep 07.
Artigo em Inglês | MEDLINE | ID: mdl-31500298

RESUMO

TIR domain-containing proteins are essential for bacterial pathogens to subvert host defenses. This study describes a fish pathogen, Yersinia ruckeri SC09 strain, with a novel TIR domain-containing protein (STIR-2) that affects Toll-like receptor (TLR) function. STIR-2 was identified in Y. ruckeri by bioinformatics analysis. The toxic effects of this gene on fish were determined by in vivo challenge experiments in knockout mutants and complement mutants of the stir-2 gene. In vitro, STIR-2 downregulated the expression and secretion of IL-6, IL-1ß, and TNF-α. Furthermore, the results of NF-κB-dependent luciferase reporter system, co-immunoprecipitation, GST pull-down assays, and yeast two-hybrid assay indicated that STIR-2 inhibited the TLR signaling pathway by interacting with myeloid differentiation factor 88 (MyD88). In addition, STIR-2 promoted the intracellular survival of pathogenic Yersinia ruckeri SC09 strain by binding to the TIR adaptor protein MyD88 and inhibiting the pre-inflammatory signal of immune cells. These results showed that STIR-2 increased virulence in Y. ruckeri and suppressed the innate immune response by inhibiting TLR and MyD88-mediated signaling, serving as a novel strategy for innate immune evasion.


Assuntos
Proteínas Adaptadoras de Transporte Vesicular/genética , Doenças dos Peixes/microbiologia , Fator 88 de Diferenciação Mieloide/metabolismo , Yersiniose/veterinária , Yersinia ruckeri/patogenicidade , Proteínas Adaptadoras de Transporte Vesicular/imunologia , Animais , Aderência Bacteriana , Proteínas de Bactérias/genética , Proteínas de Bactérias/imunologia , Doenças dos Peixes/imunologia , Proteínas de Peixes/metabolismo , Regulação da Expressão Gênica , Evasão da Resposta Imune , Camundongos Knockout , Oncorhynchus mykiss , Domínios Proteicos , Transdução de Sinais , Receptores Toll-Like/metabolismo , Fatores de Virulência/genética , Fatores de Virulência/imunologia , Yersiniose/imunologia , Yersinia ruckeri/genética , Yersinia ruckeri/imunologia
5.
Microb Pathog ; 135: 103661, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31400445

RESUMO

Treponema (T.) denticola is one of the key etiological agents in the development of periodontitis. The major outer sheath protein (Msp) of T. denticola has been shown to mediate pathogenesis and to facilitate adhesion of T. denticola to mucosal surfaces. This study aimed to find short polypeptides in the amino acid sequence of Msp which may be immunogenic and might elicit protective antisera against T. denticola. The complete msp sequence was divided into six fragments and the corresponding genes were cloned and expressed. Antisera against the polypeptides were raised in rabbits and fragment 3 (F3), hereinafter called PerioVax3 was the most potent fragment of the Msp in terms of yielding high titer antiserum. An adhesion assay was done to examine the inhibitory effects of antisera on the attachment of T. denticola to human gingival fibroblasts (HGFs) and human fibronectin. Antiserum against PerioVax3 significantly inhibited attachment of T. denticola to the substratum. Also, antiserum against PerioVax3 inhibited detachment of HGFs upon T. denticola exposure. To begin examining the clinical relevance of this work, blood samples from 12 sever periodontitis patients were collected and the sera were used in western blotting against the recombinant polypeptides. Periodontitis patient antisera exclusively detected PerioVax3 in western blotting. The data suggest that PerioVax3 carries epitopes that may trigger humoral immunity against T. denticola, which may protect against its adhesion functions. The complexity of periodontitis suggests that PerioVax3 may be considered for testing as a component of an experimental multivalent periodontal vaccine in further preclinical and clinical studies.


Assuntos
Antígenos de Bactérias/imunologia , Proteínas de Bactérias/imunologia , Epitopos/imunologia , Periodontite/imunologia , Treponema denticola/imunologia , Sequência de Aminoácidos , Animais , Anticorpos Antibacterianos/sangue , Anticorpos Antibacterianos/farmacologia , Antígenos de Bactérias/sangue , Antígenos de Bactérias/genética , Aderência Bacteriana/efeitos dos fármacos , Aderência Bacteriana/imunologia , Proteínas da Membrana Bacteriana Externa/sangue , Proteínas da Membrana Bacteriana Externa/imunologia , Proteínas de Bactérias/genética , Linhagem Celular , Clonagem Molecular , Modelos Animais de Doenças , Fibroblastos , Fibronectinas , Humanos , Masculino , Periodontite/sangue , Coelhos , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Treponema denticola/genética , Vacinas , Fatores de Virulência/imunologia
6.
Vet Microbiol ; 235: 188-194, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31383301

RESUMO

Outer membrane vesicles (OMVs) are produced and secreted virtually by every known Gram-negative bacterium. Despite their non-live nature, they share antigenic characteristics with the bacteria they originate from. This, together with their relative ease of purification, casts the OMVs as a very promising and flexible tool in both human and veterinary vaccinology. The aim of the current work was to get an insight into the antigenic pattern of OMVs from the pig pathogen Actinobacillus pleuropneumoniae in the context of vaccine development. Accordingly, we designed a protocol combining 2D Western Blotting and mass spectrometric identification to robustly characterize the antigenic protein pattern of the vesicles. Our analysis revealed that A. pleuropneumoniae OMVs carry several immunoreactive virulence factors. Some of these proteins, LpoA, OsmY and MIDG2331_02184, have never previously been documented as antigenic in A. pleuropneumoniae or other pathogenic bacteria. Additionally, we showed that despite their relative abundance, proteins such as FrpB and DegQ do not contribute to the antigenic profile of A. pleuropneumoniae OMVs.


Assuntos
Infecções por Actinobacillus/veterinária , Actinobacillus pleuropneumoniae/genética , Antígenos de Bactérias/imunologia , Proteínas da Membrana Bacteriana Externa/imunologia , Actinobacillus pleuropneumoniae/imunologia , Animais , Proteínas da Membrana Bacteriana Externa/genética , Western Blotting , Espectrometria de Massas , Mutação , Proteômica , Suínos , Fatores de Virulência/imunologia
7.
Infect Immun ; 87(9)2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31285251

RESUMO

Klebsiella pneumoniae-induced liver abscess (KLA) is emerging as a leading cause of pyogenic liver abscess worldwide. In recent years, the emergence of hypervirulent K. pneumoniae (hvKp) has been strongly associated with KLA. Unlike classical K. pneumoniae, which generally infects the immunocompromised population, hvKp can cause serious and invasive infections in young and healthy individuals. hvKp isolates are often associated with the K1/K2 capsular types and possess hypermucoviscous capsules. KLA is believed to be caused by K. pneumoniae colonizing the gastrointestinal tract of the host and translocating across the intestinal barrier via the hepatic portal vein into the liver to cause liver abscess. We optimized the isolation of the liver-resident macrophages called Kupffer cells in mice and examined their importance in controlling bacterial loads during hvKp infection in healthy mice. Our study reveals the high capability of Kupffer cells to kill hvKp in vitro despite the presence of the bacterial hypermucoviscous capsule, in contrast to other macrophages, which were unable to phagocytose the bacteria efficiently. Depletion of Kupffer cells and macrophages with liposome-encapsulated clodronate (liposomal clodronate) in both an intraperitoneal and an oral mouse infection model resulted in increased bacterial loads in the livers, spleens, and lungs and increased mortality of the infected mice. Thus, Kupffer cells and macrophages are critical for the control of hvKp infection.


Assuntos
Infecções por Klebsiella/microbiologia , Klebsiella pneumoniae/patogenicidade , Macrófagos do Fígado/imunologia , Abscesso Hepático/microbiologia , Macrófagos/imunologia , Animais , Cápsulas Bacterianas , Abscesso Hepático/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Virulência , Fatores de Virulência/imunologia
8.
Int J Infect Dis ; 86: 57-64, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31255709

RESUMO

BACKGROUND: Pneumolysin (Ply), as a major virulence factor of Streptococcus pneumoniae, has attracted increased attention for its potential value in the development of next-generation protein-based pneumococcal vaccines. This study aimed to analyze the genetic and antigenic diversity that can influence the immunogenicity of vaccines. METHODS: A total of 96 pneumococcal isolate samples were obtained from children of 1-35 months old with invasive pneumococcal diseases in Shanghai Children's Medical Center (Shanghai, China). After DNA amplification by PCR and Sanger sequencing, Ply DNA sequences were analyzed by bioinformatics tools, including ClustalX, BioEdit and MEGA7. RESULTS: Two alleles, allele 1 and 2, and 10 subtypes, of which were 6 novel subtypes, were identified. Nucleotide and amino acid sequence identity among these pneumococcal isolates were >99%. Subtypes with the same amino acid sequence were more closely evolutionarily related in the phylogenetic tree. Only minor differences in the B-cell epitopes were identified in the antigenicity plots of alleles 1 and 2. The most common serotype was serotype 19A. CONCLUSIONS: The sequence diversity of Ply is limited although some allelic variations are detected. Different alleles exhibit similar antigenic patterns. Development of Ply-based vaccines may be a promising method to combat pneumococcal infection in the future.


Assuntos
Infecções Pneumocócicas/microbiologia , Estreptolisinas/genética , Estreptolisinas/imunologia , Alelos , Sequência de Aminoácidos , Variação Antigênica , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Proteínas de Bactérias/imunologia , Pré-Escolar , Variação Genética , Humanos , Lactente , Filogenia , Infecções Pneumocócicas/prevenção & controle , Vacinas Pneumocócicas , Sorogrupo , Streptococcus pneumoniae/classificação , Streptococcus pneumoniae/isolamento & purificação , Estreptolisinas/química , Fatores de Virulência/genética , Fatores de Virulência/imunologia
9.
Future Microbiol ; 14: 867-884, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31340660

RESUMO

Aim: Cryptococcus neoformans is the major agent of cryptococcosis. The main virulence factor is the polysaccharide (PS) capsule. Changes in cryptococcal PS properties have been poorly elucidated. Materials & methods: We analyzed the mechanical properties of secreted PS and intact capsules, using dynamic light scattering and optical tweezers. Results: Storage and loss moduli showed that secreted PS behaves as a viscoelastic liquid, while capsular PS behaves as a viscoelastic solid. The secreted PS remains as a viscoelastic fluid at different temperatures with thermal hysteresis after 85°C. Antibody binding altered the viscoelastic behavior of both secreted and capsular PS. Conclusion: Deciphering the mechanical aspects of these structures could reveal features that may have consequences in novel therapies against cryptococcosis.


Assuntos
Anticorpos Antifúngicos/metabolismo , Cryptococcus neoformans/química , Polissacarídeos/fisiologia , Temperatura Ambiente , Fatores de Virulência/fisiologia , Anticorpos Antifúngicos/imunologia , Cápsulas Fúngicas/química , Cápsulas Fúngicas/imunologia , Cápsulas Fúngicas/fisiologia , Pinças Ópticas , Tamanho da Partícula , Polissacarídeos/química , Polissacarídeos/imunologia , Polissacarídeos/metabolismo , Reologia , Fatores de Virulência/química , Fatores de Virulência/imunologia , Fatores de Virulência/metabolismo , Substâncias Viscoelásticas
10.
PLoS Negl Trop Dis ; 13(7): e0007578, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31306423

RESUMO

BACKGROUND: Glanders caused by Burkholderia mallei is a re-emerging zoonotic disease affecting solipeds and humans. Furthermore, B. mallei is genetically related to B. pseudomallei, which is the causative agent of melioidosis. Both facultative intracellular bacteria are classified as tier 1 select biothreat agents. Our previous study with a B. mallei ΔtonB Δhcp1 (CLH001) live-attenuated vaccine demonstrated that it is attenuated, safe and protective against B. mallei wild-type strains in the susceptible BALB/c mouse model. METHODOLOGY/PRINCIPAL FINDING: In our current work, we evaluated the protective efficacy of CLH001 against glanders and melioidosis in the more disease-resistant C57BL/6 mouse strain. The humoral as well as cellular immune responses were also examined. We found that CLH001-immunized mice showed 100% survival against intranasal and aerosol challenge with B. mallei ATCC 23344. Moreover, this vaccine also afforded significant cross-protection against B. pseudomallei K96243, with low level bacterial burden detected in organs. Immunization with a prime and boost regimen of CLH001 induced significantly greater levels of total and subclasses of IgG, and generated antigen-specific splenocyte production of IFN-γ and IL-17A. Interestingly, protection induced by CLH001 is primarily dependent on humoral immunity, while CD4+ and CD8+ T cells played a less critical protective role. CONCLUSIONS/SIGNIFICANCE: Our data indicate that CLH001 serves as an effective live attenuated vaccine to prevent glanders and melioidosis. The quantity and quality of antibody responses as well as improving cell-mediated immune responses following vaccination need to be further investigated prior to advancement to preclinical studies.


Assuntos
Proteínas de Bactérias/imunologia , Vacinas Bacterianas/imunologia , Burkholderia mallei/imunologia , Mormo/imunologia , Imunização , Melioidose/imunologia , Proteínas de Membrana/imunologia , Vacinas Atenuadas/imunologia , Animais , Anticorpos Antibacterianos/imunologia , Proteínas de Bactérias/genética , Burkholderia mallei/genética , Linfócitos T CD8-Positivos/imunologia , Modelos Animais de Doenças , Feminino , Mormo/microbiologia , Mormo/prevenção & controle , Humanos , Imunidade Humoral , Melioidose/microbiologia , Melioidose/prevenção & controle , Proteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Vacinação , Fatores de Virulência/genética , Fatores de Virulência/imunologia
11.
mBio ; 10(3)2019 06 25.
Artigo em Inglês | MEDLINE | ID: mdl-31239384

RESUMO

Human cytomegalovirus (HCMV) is a betaherpesvirus that is a significant pathogen within newborn and immunocompromised populations. Morbidity associated with HCMV infection is the consequence of viral dissemination. HCMV has evolved to manipulate the host immune system to enhance viral dissemination and ensure long-term survival within the host. The immunomodulatory protein vCXCL-1, a viral chemokine functioning primarily through the CXCR2 chemokine receptor, is hypothesized to attract CXCR2+ neutrophils to infection sites, aiding viral dissemination. Neutrophils harbor HCMV in vivo; however, the interaction between vCXCL-1 and the neutrophil has not been evaluated in vivo Using the mouse model and mouse cytomegalovirus (MCMV) infection, we show that murine neutrophils harbor and transfer infectious MCMV and that virus replication initiates within this cell type. Utilizing recombinant MCMVs expressing vCXCL-1 from the HCMV strain (Toledo), we demonstrated that vCXCL-1 significantly enhances MCMV dissemination kinetics. Through cellular depletion experiments, we observe that neutrophils impact dissemination but that overall dissemination is largely neutrophil independent. This work adds neutrophils to the list of innate cells (i.e., dendritic and macrophages/monocytes) that contribute to MCMV dissemination but refutes the hypothesis that neutrophils are the primary cell responding to vCXCL-1.IMPORTANCE An adequate in vivo analysis of HCMV's viral chemokine vCXCL-1 has been lacking. Here we generate recombinant MCMVs expressing vCXCL-1 to study vCXCL-1 function in vivo using MCMV as a surrogate. We demonstrate that vCXCL-1 increases MCMV dissemination kinetics for both primary and secondary dissemination. Additionally, we provide evidence, that the murine neutrophil is largely a bystander in the mouse's response to vCXCL-1. We confirm the hypothesis that vCXCL-1 is a HCMV virulence factor. Infection of severely immunocompromised mice with MCMVs expressing vCXCL-1 was lethal in more than 50% of infected animals, while all animals infected with parental virus survived during a 12-day period. This work provides needed insights into vCXCL-1 function in vivo.


Assuntos
Quimiocina CXCL1/imunologia , Infecções por Citomegalovirus/imunologia , Citomegalovirus/imunologia , Muromegalovirus/imunologia , Neutrófilos/virologia , Animais , Quimiocina CXCL1/genética , Interações Hospedeiro-Patógeno/imunologia , Humanos , Cinética , Camundongos , Camundongos Endogâmicos BALB C , Muromegalovirus/patogenicidade , Neutrófilos/imunologia , Receptores de Interleucina-8B/genética , Receptores de Interleucina-8B/imunologia , Fatores de Virulência/imunologia , Replicação Viral
12.
Int J Mol Sci ; 20(12)2019 Jun 21.
Artigo em Inglês | MEDLINE | ID: mdl-31234322

RESUMO

Phytophthora infestans causes the severe late blight disease of potato. During its infection process, P. infestans delivers hundreds of RXLR (Arg-x-Leu-Arg, x behalf of any one amino acid) effectors to manipulate processes in its hosts, creating a suitable environment for invasion and proliferation. Several effectors interact with host proteins to suppress host immunity and inhibit plant growth. However, little is known about how P. infestans regulates the host transcriptome. Here, we identified an RXLR effector, PITG_15718.2, which is upregulated and maintains a high expression level throughout the infection. Stable transgenic potato (Solanum tuberosum) lines expressing PITG_15718.2 show enhanced leaf colonization by P. infestans and reduced vegetative growth. We further investigated the transcriptional changes between three PITG_15718.2 transgenic lines and the wild type Désirée by using RNA sequencing (RNA-Seq). Compared with Désirée, 190 differentially expressed genes (DEGs) were identified, including 158 upregulated genes and 32 downregulated genes in PITG_15718.2 transgenic lines. Eight upregulated and nine downregulated DEGs were validated by real-time RT-PCR, which showed a high correlation with the expression level identified by RNA-Seq. These DEGs will help to explore the mechanism of PITG_15718.2-mediated immunity and growth inhibition in the future.


Assuntos
Peptídeos/imunologia , Phytophthora infestans/imunologia , Doenças das Plantas/imunologia , Solanum tuberosum/imunologia , Fatores de Virulência/imunologia , Interações Hospedeiro-Parasita , Phytophthora infestans/fisiologia , Doenças das Plantas/parasitologia , Imunidade Vegetal , Plantas Geneticamente Modificadas/crescimento & desenvolvimento , Plantas Geneticamente Modificadas/imunologia , Plantas Geneticamente Modificadas/parasitologia , Solanum tuberosum/crescimento & desenvolvimento , Solanum tuberosum/parasitologia
13.
World J Microbiol Biotechnol ; 35(6): 94, 2019 Jun 11.
Artigo em Inglês | MEDLINE | ID: mdl-31187291

RESUMO

Pseudomonas aeruginosa is the major infectious agent of concern for cystic fibrosis (CF) patients. Therefore, it is necessary to develop appropriate strategies for preventing colonization by this bacterium and/or neutralizing virulence factors. In this study, we formulated the encapsulation of exotoxin A into PLGA nanoparticles. The biological activities of the nanovaccine candidate were also characterized. Based on the results, ETA-PLGA can act as a suitable immunogen to stimulate the humoral and cellular immune response. The antibodies raised against ETA-PLGA significantly decreased bacterial titer in the spleens of the immunized mice after challenge with PAO1 strain, compared to the control groups. The encapsulation of PLGA into ETA led to a significantly higher production of INF-γ, TNF-α, IL-4, and IL-17A cytokine responses compared to the ETA group. ETA-PLGA enhanced IgG responses in immunized mice compared to ETA antigen. We concluded that encapsulation of Pseudomonas aeruginosa ETA to PLGA nanoparticles can increase its functional activity by decreasing the bacterial dissemination.


Assuntos
ADP Ribose Transferases/imunologia , Toxinas Bacterianas/imunologia , Exotoxinas/imunologia , Imunização , Nanoconjugados , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/imunologia , Infecções por Pseudomonas/prevenção & controle , Pseudomonas aeruginosa/patogenicidade , Vacinas Conjugadas , Fatores de Virulência/imunologia , ADP Ribose Transferases/uso terapêutico , Animais , Toxinas Bacterianas/uso terapêutico , Citocinas/metabolismo , Modelos Animais de Doenças , Exotoxinas/uso terapêutico , Feminino , Imunidade Celular , Imunidade Humoral , Imunoglobulina G/sangue , Interferon gama/metabolismo , Interleucina-17/metabolismo , Interleucina-4/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Nanopartículas , Tamanho da Partícula , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/uso terapêutico , Infecções por Pseudomonas/imunologia , Baço/imunologia , Baço/microbiologia , Fatores de Virulência/uso terapêutico
14.
Int J Mol Sci ; 20(11)2019 Jun 04.
Artigo em Inglês | MEDLINE | ID: mdl-31167367

RESUMO

Bacteria from the species Trueperella pyogenes are a part of the biota of skin and mucous membranes of the upper respiratory, gastrointestinal, or urogenital tracts of animals, but also, opportunistic pathogens. T. pyogenes causes a variety of purulent infections, such as metritis, mastitis, pneumonia, and abscesses, which, in livestock breeding, generate significant economic losses. Although this species has been known for a long time, many questions concerning the mechanisms of infection pathogenesis, as well as reservoirs and routes of transmission of bacteria, remain poorly understood. Pyolysin is a major known virulence factor of T. pyogenes that belongs to the family of cholesterol-dependent cytolysins. Its cytolytic activity is associated with transmembrane pore formation. Other putative virulence factors, including neuraminidases, extracellular matrix-binding proteins, fimbriae, and biofilm formation ability, contribute to the adhesion and colonization of the host tissues. However, data about the pathogen-host interactions that may be involved in the development of T. pyogenes infection are still limited. The aim of this review is to present the current knowledge about the pathogenic potential and virulence of T. pyogenes.


Assuntos
Actinomycetaceae/fisiologia , Infecções por Bactérias Gram-Positivas/microbiologia , Actinomycetaceae/classificação , Actinomycetaceae/patogenicidade , Animais , Toxinas Bacterianas/química , Toxinas Bacterianas/genética , Toxinas Bacterianas/imunologia , Reservatórios de Doenças , Genoma Bacteriano , Genômica/métodos , Infecções por Bactérias Gram-Positivas/imunologia , Infecções por Bactérias Gram-Positivas/transmissão , Interações Hospedeiro-Patógeno/imunologia , Humanos , Filogenia , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , RNA Ribossômico 16S , Relação Estrutura-Atividade , Virulência , Fatores de Virulência/química , Fatores de Virulência/genética , Fatores de Virulência/imunologia
15.
Methods Mol Biol ; 1997: 77-85, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31119618

RESUMO

Gonococcal colony typing is part a science and part an art that has been central to studies which have identified crucial virulence antigens and also demonstrated the ability of the bacteria to undergo rapid phase and antigenic variation. Without this fundamental work, modern molecular biological studies of gonococcal pathogenesis would not have been possible. Indeed colony typing is still essential when performing biological experiments with clinical and laboratory isolates and for monitoring their outcome. In this chapter, the methods for performing colony typing and techniques to optimize the process are described.


Assuntos
Técnicas de Tipagem Bacteriana/métodos , Neisseria gonorrhoeae/classificação , Variação Antigênica/imunologia , Antígenos de Bactérias/imunologia , Antígenos de Bactérias/isolamento & purificação , DNA Bacteriano/isolamento & purificação , Neisseria gonorrhoeae/patogenicidade , Fatores de Virulência/imunologia , Fatores de Virulência/isolamento & purificação
16.
Methods Mol Biol ; 1997: 431-452, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31119638

RESUMO

Experimental infection of male volunteers with Neisseria gonorrhoeae is safe and reproduces the clinical features of naturally acquired gonococcal urethritis. The human model is useful for testing the importance of putative gonococcal virulence factors for urethral infection in men and the model presents opportunities to examine host immune responses that may be exploited or improved in development and testing of gonococcal vaccines. In this chapter, we describe methods for production, characterization, and storage of N. gonorrhoeae stocks for experimental human challenge, preparation and delivery of inoculum suspensions, monitoring experimental infection, and statistical considerations for data analysis.


Assuntos
Gonorreia/imunologia , Experimentação Humana , Neisseria gonorrhoeae/patogenicidade , Uretrite/imunologia , Adulto , Proteínas de Bactérias/imunologia , Gonorreia/microbiologia , Voluntários Saudáveis , Humanos , Masculino , Uretrite/microbiologia , Fatores de Virulência/imunologia
17.
Artigo em Inglês | MEDLINE | ID: mdl-31134159

RESUMO

Vi capsular polysaccharide (Vi) is a major virulence factor of human typhoid-causing pathogen Salmonella enterica serovar Typhi (S. Typhi). It distinguishes S. Typhi from closely related non-typhoidal Salmonella serovars such as S. Typhimurium which do not normally cause systemic infection in humans. Vi not only forms a capsule around S. Typhi but it is also readily released from this pathogen. We have previously reported that Vi targets prohibitin to inhibit cellular responses activated through immune receptors. Here, we show that engagement of membrane prohibitin with Vi prevents Salmonella-induced activation of small Rho-family GTPases, Rac1, and Cdc42, and suppresses actin cytoskeletal rearrangements resulting in reduced invasion and highly subdued inflammatory responses. Cells infected with S. Typhimurium in the presence of Vi show poor activation of NF-kB and MAP-kinase pathways of intracellular signaling. Treatment with Vi brings about redistribution of Rac-1, prohibitin, and ganglioside GM1 in membrane raft domains. Vi-mediated interference with activation of Rho-family GTPases represents a previously unrecognized mechanism by which S. Typhi can limit its invasion and alarming of the host.


Assuntos
Células Epiteliais/metabolismo , Polissacarídeos Bacterianos/metabolismo , Salmonella typhi/metabolismo , Febre Tifoide/imunologia , Fatores de Virulência/metabolismo , Proteínas rho de Ligação ao GTP/metabolismo , Células Epiteliais/imunologia , Células HeLa , Humanos , Interleucina-8/metabolismo , Polissacarídeos Bacterianos/imunologia , Proteínas Repressoras/metabolismo , Salmonella typhi/imunologia , Virulência , Fatores de Virulência/imunologia , Proteína cdc42 de Ligação ao GTP , Proteínas rac1 de Ligação ao GTP
18.
Cell Host Microbe ; 25(6): 884-891.e6, 2019 06 12.
Artigo em Inglês | MEDLINE | ID: mdl-31126758

RESUMO

The capacity of Streptococcus pneumoniae to successfully transmit and colonize new human hosts is a critical aspect of pneumococcal population biology and a prerequisite for invasive disease. However, the bacterial mechanisms underlying this process remain largely unknown. To identify bacterial factors required for transmission, we conducted a high-throughput genetic screen with a transposon sequencing (Tn-seq) library of a pneumococcal strain in a ferret transmission model. Key players in both metabolism and transcriptional regulation were identified as required for efficient bacterial transmission. Targeted deletion of the putative C3-degrading protease CppA, iron transporter PiaA, or competence regulatory histidine kinase ComD significantly decreased transmissibility in a mouse model, further validating the screen. Maternal vaccination with recombinant surface-exposed PiaA and CppA alone or in combination blocked transmission in offspring and were more effective than capsule-based vaccines. These data underscore the possibility of targeting pneumococcal transmission as a means of eliminating invasive disease in the population.


Assuntos
Transmissão Vertical de Doença Infecciosa/prevenção & controle , Infecções Pneumocócicas/prevenção & controle , Infecções Pneumocócicas/transmissão , Vacinas Pneumocócicas/imunologia , Streptococcus pneumoniae/genética , Streptococcus pneumoniae/isolamento & purificação , Fatores de Virulência/genética , Animais , Modelos Animais de Doenças , Furões , Testes Genéticos , Ensaios de Triagem em Larga Escala , Camundongos , Mutagênese Insercional , Infecções Pneumocócicas/microbiologia , Vacinas Pneumocócicas/administração & dosagem , Vacinas Pneumocócicas/isolamento & purificação , Análise de Sequência de DNA , Vacinas Sintéticas/administração & dosagem , Vacinas Sintéticas/imunologia , Vacinas Sintéticas/isolamento & purificação , Fatores de Virulência/imunologia
19.
Int J Mol Sci ; 20(10)2019 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-31096563

RESUMO

High immunogenicity and systemic toxicity are the main obstacles limiting the clinical use of the therapeutic agents based on Pseudomonas aeruginosa exotoxin A. In this work, we studied the immunogenicity, general toxicity and antitumor effect of the targeted toxin DARPin-LoPE composed of HER2-specific DARPin and a low immunogenic exotoxin A fragment lacking immunodominant human B lymphocyte epitopes. The targeted toxin has been shown to effectively inhibit the growth of HER2-positive human ovarian carcinoma xenografts, while exhibiting low non-specific toxicity and side effects, such as vascular leak syndrome and liver tissue degradation, as well as low immunogenicity, as was shown by specific antibody titer. This represents prospects for its use as an agent for targeted therapy of HER2-positive tumors.


Assuntos
Epitopos de Linfócito B/imunologia , Xenoenxertos , Imunotoxinas/imunologia , Imunotoxinas/farmacologia , Proteínas Musculares/imunologia , Proteínas Nucleares/imunologia , Neoplasias Ovarianas/tratamento farmacológico , Receptor ErbB-2/imunologia , ADP Ribose Transferases/imunologia , ADP Ribose Transferases/farmacologia , Sequência de Aminoácidos , Animais , Antineoplásicos/imunologia , Antineoplásicos/farmacologia , Toxinas Bacterianas/imunologia , Toxinas Bacterianas/farmacologia , Biomarcadores Tumorais , Carcinoma/tratamento farmacológico , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Modelos Animais de Doenças , Epitopos de Linfócito B/genética , Exotoxinas/imunologia , Exotoxinas/farmacologia , Feminino , Humanos , Concentração Inibidora 50 , Fígado/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Terapia de Alvo Molecular , Proteínas Musculares/genética , Proteínas Nucleares/genética , Neoplasias Ovarianas/patologia , Proteínas Recombinantes de Fusão/imunologia , Proteínas Recombinantes de Fusão/uso terapêutico , Baço/patologia , Fatores de Virulência/imunologia , Fatores de Virulência/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto
20.
J Agric Food Chem ; 67(24): 6828-6836, 2019 Jun 19.
Artigo em Inglês | MEDLINE | ID: mdl-31136163

RESUMO

Verticillium wilt, caused by Verticillium dahliae, results in a dramatic loss of cotton yields in China. There is great potential for biocontrol to manage this destructive crop disease. In this study, we obtained the endophytic bacterium Bacillus halotolerans Y6 from Verticillium wilt-resistant cotton Gossypium barbadense Xinhai15; this bacterium possesses strong antagonistic abilities that inhibit V. dahliae spore germination and mycelial growth. The results of the enzyme activity assay, heterologous expression, and gene knockdown showed that the key virulence factor of Y6 for antagonizing V. dahliae was ß -glucanase Bgy6. To facilitate field tests of biological control, we constructed the homologous Bgy6-overexpression strain OY6. Compared with the wild-type Y6 strain, the ß-glucanase activity of OY6 was increased by 91.79%, and the inhibition rate of OY6 against V. dahliae V991 exceeded 96.7%. Moreover, the spores of V. dahliae V991 treated with OY6 showed more mucus and larger holes on the surface, as observed by scanning electron microscopy. Potting test results illustrated that both OY6 and Y6 could improve the resistance of upland cotton to Verticillium wilt. With the inoculation of V. dahliae V991 for 45 days, the disease index of G. hirsutum TM-1 treated with OY6 was only 8.33, which was significantly lower than that in plants treated with the wild-type strain Y6 (17.86) or the controls without bacteria (35.94). Our research provides a new idea for the control of Verticillium wilt in upland cotton via transforming endophytic bacteria of Verticillium wilt-resistant cotton and proposes a new solution to prevent and control Verticillium wilt.


Assuntos
Bacillus/enzimologia , Proteínas de Bactérias/genética , Endo-1,3(4)-beta-Glucanase/genética , Endófitos/enzimologia , Gossypium/microbiologia , Doenças das Plantas/imunologia , Verticillium/fisiologia , Fatores de Virulência/genética , Antibiose , Bacillus/genética , Bacillus/isolamento & purificação , Bacillus/fisiologia , Proteínas de Bactérias/metabolismo , Resistência à Doença , Endo-1,3(4)-beta-Glucanase/metabolismo , Endófitos/genética , Endófitos/isolamento & purificação , Endófitos/fisiologia , Expressão Gênica , Regulação da Expressão Gênica de Plantas , Gossypium/imunologia , Doenças das Plantas/microbiologia , Fatores de Virulência/imunologia
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