RESUMO
AIMS: Allergic asthma is a chronic inflammatory lung disease characterized by a Th2-type immune response pattern. The development of nonspecific immunotherapy is one of the primary goals for the control of this disease. METHODS AND RESULTS: In this study, we evaluated the therapeutic effects of Lactococcus lactis-producing mycobacterial heat shock protein 65 (LLHsp65) in an ovalbumin (OVA)-induced allergic asthma model. OVA-challenged BALB/c mice were orally administrated with LLHsp65 for 10 consecutive days. The results demonstrate that LLhsp65 attenuates critical features of allergic inflammation, like airway hyperresponsiveness and mucus production. Likewise, the treatment decreases the pulmonary eosinophilia and the serum level of OVA-specific IgE. In addition to deviating immune responses towards Th1-cytokine profile, increase regulatory T cells, and cytokine levels, such as IL-6 and IL-10. CONCLUSIONS: Our results reveal that the mucosal immunotherapy of LLHsp65 significantly reduces the overall burden of airway allergic inflammation, suggesting a promising therapeutic strategy for allergic asthma treatment. SIGNIFICANCE AND IMPACT OF THE STUDY: This research reveals new perspectives on nonspecific immunotherapy based on the delivery of recombinant proteins by lactic acid bacteria to treat of allergic disorders.
Assuntos
Asma/tratamento farmacológico , Proteínas de Bactérias/farmacologia , Chaperonina 60/farmacologia , Inflamação/tratamento farmacológico , Lactococcus lactis/imunologia , Administração Oral , Animais , Asma/imunologia , Líquido da Lavagem Broncoalveolar/citologia , Citocinas/metabolismo , Modelos Animais de Doenças , Feminino , Hipersensibilidade/tratamento farmacológico , Imunoglobulina E/sangue , Imunoglobulina G/sangue , Imunoterapia , Lactococcus lactis/metabolismo , Pulmão/efeitos dos fármacos , Pulmão/imunologia , Pulmão/patologia , Camundongos , Camundongos Endogâmicos BALB C , Ovalbumina , Linfócitos T Reguladores/imunologiaRESUMO
AIMS: To investigate the anti-inflammatory activity of an invasive and Hp65-producing strain Lactococcus lactis NCDO2118 FnBPA+ (pXYCYT:Hsp65) in acute 2,4,6-trinitrobenzene sulphonic acid (TNBS)-induced colitis in mice as an innovative therapeutic strategy against Crohn's disease (CD). METHODS AND RESULTS: The pXYCYT:Hsp65 plasmid was transformed into the L. lactis NCDO2118 FnBPA+ strain, resulting in the L. lactis NCDO2118 FnBPA+ (pXYCYT:Hsp65) strain. Then, the functionality of the strain was evaluated in vitro for Hsp65 production by Western blotting and for invasion into Caco-2 cells. The results demonstrated that the strain was able to produce Hsp65 and efficiently invade eukaryotic cells. Subsequently, in vivo, the anti-inflammatory capacity of the recombinant strain was evaluated in colitis induced with TNBS in BALB/c mice. Oral administration of the recombinant strain was able to attenuated the severity of colitis by mainly reducing IL-12 and IL-17 levels and increasing IL-10 and secretory immunoglobulin A levels. CONCLUSIONS: The L. lactis NCDO2118 FnBPA+ (pXYCYT:Hsp65) strain contributed to a reduction in inflammatory damage in experimental CD. SIGNIFICANCE AND IMPACT OF THE STUDY: This study, which used L. lactis for the production and delivery of Hsp65, has scientific relevance because it shows the efficacy of this new strategy based on therapeutic protein delivery into mammalian enterocytes.
Assuntos
Proteínas de Bactérias/metabolismo , Chaperonina 60/metabolismo , Colite/terapia , Imunoglobulina A Secretora/metabolismo , Interleucina-10/metabolismo , Lactococcus lactis/fisiologia , Administração Oral , Animais , Proteínas de Bactérias/genética , Células CACO-2 , Chaperonina 60/genética , Colite/induzido quimicamente , Colite/imunologia , Sistemas de Liberação de Medicamentos , Feminino , Humanos , Inflamação/terapia , Lactococcus lactis/genética , Lactococcus lactis/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Ácido Trinitrobenzenossulfônico/toxicidadeRESUMO
AIMS: A regimen utilizing Bacille Calmette-Guerin (BCG) and another vaccine system as a booster may represent a promising strategy for the development of an efficient tuberculosis vaccine for adults. In a previous work, we confirmed the ability of Lactococcus lactis fibronectin-binding protein A (FnBPA+) (pValac:ESAT-6), a live mucosal DNA vaccine, to produce a specific immune response in mice after oral immunization. In this study, we examined the immunogenicity of this strain as a booster for the BCG vaccine in mice. METHODS AND RESULTS: After immunization, cytokine and immunoglobulin profiles were measured. The BCG prime L. lactis FnBPA+ (pValac:ESAT-6) boost group was the most responsive group, with a significant increase in splenic pro-inflammatory cytokines IL-17, IFN-γ, IL-6 and TNF-α compared with the negative control. CONCLUSIONS: Based on the results obtained here, we demonstrated that L. lactis FnBPA+ (pValac:ESAT-6) was able to increase the BCG vaccine general immune response. SIGNIFICANCE AND IMPACT OF THE STUDY: This work is of great scientific and social importance because it represents the first step towards the development of a booster to the BCG vaccine using L. lactis as a DNA delivery system.
Assuntos
Antígenos de Bactérias/imunologia , Vacina BCG/imunologia , Proteínas de Bactérias/imunologia , Citocinas/sangue , Interleucina-17/metabolismo , Lactococcus lactis/genética , Vacinas de DNA/imunologia , Animais , Antígenos de Bactérias/genética , Proteínas de Bactérias/genética , Feminino , Interleucina-6 , Lactococcus lactis/metabolismo , Camundongos , Fator de Necrose Tumoral alfaRESUMO
AIMS: Propionibacterium freudenreichii is an actinobacterium widely used in dairy industry during the ripening process of Swiss-type cheeses and which presents probiotic properties. P. freudenreichii is reportedly a hardy bacterium, able to survive during the cheese-making process and when subjected to digestive stresses. During this study the long-term survival (LTS) of P. freudenreichii was investigated for 11 days by means of phenotypic characterization in a culture medium without the addition of any nutrients. METHODS AND RESULTS: For 11 days, in a non-nutrient supplemented culture medium, eight strains were monitored by measuring their optical density, counting colony-forming units (CFU) and using LIVE/DEAD staining and microscopy observation. Under these conditions, all strains displayed high survival rates in the culture medium, their culturability reaching more than 9 log10 CFU ml(-1) after 2 days. After 11 days, this value ranged from 7·8 to 8·2 log10 CFU ml(-1) depending on the strain, and at least 50% of the P. freudenreichii population displayed an intact envelope. As lysis of part of a bacterial population may be a microbial strategy to recover nutrients, in CIRM-BIA 138 (the strain with the highest population at day 11), cell lysis was assessed by quantifying intact bacterial cells using qPCR targeting the housekeeping gene tuf. No lysis was observed. CONCLUSION: Taken together, our results suggest that P. freudenreichii strains use a viable but nonculturable state to adapt to the LTS phase. SIGNIFICANCE AND IMPACT OF THE STUDY: Assessing the viability of P. freudenreichii and understanding their mechanisms for survival should be of great interest regarding their potential probiotic applications.
Assuntos
Meios de Cultura/metabolismo , Propionibacterium/crescimento & desenvolvimento , Meios de Cultura/análise , Viabilidade Microbiana , Propionibacterium/metabolismoRESUMO
In this study, Lactococcus lactis was engineered to express mutated internalin A on its surface and to secrete large amounts of listeriolysin O (LLO) in order to improve its potential as a vehicle for DNA vaccination. Western blotting experiments demonstrated that the bacterium expressed LLO in both the cytoplasmic and extracellular compartments, with higher quantities found in the culture supernatants. A hemolytic assay showed that the recombinant strain secreted 250 ng active LLO/mg total protein. This mInlA/LLO-producing strain of L. lactis may be used as an alternative tool in DNA vaccination against a number of infectious diseases or in cancer therapy.
Assuntos
Proteínas de Bactérias/genética , Proteínas de Bactérias/imunologia , Toxinas Bacterianas/genética , Toxinas Bacterianas/imunologia , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/imunologia , Proteínas Hemolisinas/genética , Proteínas Hemolisinas/imunologia , Lactococcus lactis/genética , Listeria monocytogenes/imunologia , Mutação , Proteínas Recombinantes , Vacinas Bacterianas , Membrana Celular/metabolismo , Expressão Gênica , Hemólise , Lactococcus lactis/metabolismo , VacinaçãoRESUMO
The human genome project is one of the significant achievements that have provided detailed insight into our genetic legacy. During the last two decades, biomedical investigations have gathered a considerable body of evidence by detecting more than 2000 disease genes. Despite the imperative advances in the genetic understanding of various diseases, the pathogenesis of many others remains obscure. With recent advances, the laborious methodologies used to identify DNA variations are replaced by direct sequencing of genomic DNA to detect genetic changes. The ability to perform such studies depends equally on the development of high-throughput and economical genotyping methods. Currently, basically for every disease whose origen is still unknown, genetic approaches are available which could be pedigree-dependent or -independent with the capacity to elucidate fundamental disease mechanisms. Computer algorithms and programs for linkage analysis have formed the foundation for many disease gene detection projects, similarly databases of clinical findings have been widely used to support diagnostic decisions in dysmorphology and general human disease. For every disease type, genome sequence variations, particularly single nucleotide polymorphisms are mapped by comparing the genetic makeup of case and control groups. Methods that predict the effects of polymorphisms on protein stability are useful for the identification of possible disease associations, whereas structural effects can be assessed using methods to predict stability changes in proteins using sequence and/or structural information.
Assuntos
Ligação Genética , Genoma Humano , Biologia Molecular/tendências , Software , Biologia Computacional , Bases de Dados Genéticas , Predisposição Genética para Doença , Humanos , Biologia Molecular/instrumentação , Biologia Molecular/métodos , Linhagem , Polimorfismo Genético , Análise de Sequência de DNARESUMO
Allergic diseases affect up to 30% of the western population, and their prevalence is increasing. Probiotics are able to modulate the mucosal immune response, and clinical trials demonstrated that specific strains, especially lactic acid bacteria (LAB) ones, reduce allergic symptoms. Moreover, the use of recombinant probiotics has been evaluated as possible strategies for the immunotherapy of allergic diseases. The production and delivery of allergens by recombinant LAB in concert with their ability to induce a Th1-type immune response have been shown to be a promising mucosal vaccination strategy in mouse model. The aim of this article is to review the applications of probiotics in allergy immunotherapy with a special focus on recombinant LAB delivering proteins or DNA.
Assuntos
Hipersensibilidade/terapia , Imunoterapia , Probióticos/uso terapêutico , Alérgenos/genética , Alérgenos/imunologia , Animais , Bifidobacterium/genética , DNA/administração & dosagem , Humanos , Hipersensibilidade/imunologia , Imunidade nas Mucosas , Lactobacillus/genética , Camundongos , Hipersensibilidade a Leite/terapia , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/imunologiaRESUMO
Due to next-generation sequence technologies, sequencing of bacterial genomes is no longer one of the main bottlenecks in bacterial research and the number of new genomes deposited in public databases continues to increase at an accelerating rate. Among these new genomes, several belong to the same species and were generated for pan-genomic studies. A pan-genomic study allows investigation of strain phenotypic differences based on genotypic differences. Along with a need for good assembly quality, it is also fundamental to guarantee good functional genome annotation of the different strains. In order to ensure quality and standards for functional genome annotation among different strains, we developed and made available PANNOTATOR (http://bnet.egr.vcu.edu/iioab/agenote.php), a web-based automated pipeline for the annotation of closely related and well-suited genomes for pan-genome studies, aiming at reducing the manual work to generate reports and corrections of various genome strains. PANNOTATOR achieved 98 and 76% of correctness for gene name and function, respectively, as result of an annotation transfer, with a similarity cut-off of 70%, compared with a gold standard annotation for the same species. These results surpassed the RAST and BASys softwares by 41 and 21% and 66 and 17% for gene name and function annotation, respectively, when there were reliable genome annotations of closely related species. PANNOTATOR provides fast and reliable pan-genome annotation; thereby allowing us to maintain the research focus on the main genotype differences between strains.
Assuntos
Genoma Bacteriano/genética , Anotação de Sequência Molecular , Software , Bases de Dados Genéticas , Sequenciamento de Nucleotídeos em Larga EscalaRESUMO
Streptococcus agalactiae (Lancefield group B; group B streptococci) is a major pathogen that causes meningoencephalitis in fish, mastitis in cows, and neonatal sepsis and meningitis in humans. The available prophylactic measures for conserving human and animal health are not totally effective and have limitations. Effective vaccines against the different serotypes or genotypes of pathogenic strains from the various hosts would be useful. We used an in silico strategy to identify conserved vaccine candidates in 15 genomes of group B streptococci strains isolated from human, bovine, and fish samples. The degree of conservation, subcellular localization, and immunogenic potential of S. agalactiae proteins were investigated. We identified 36 antigenic proteins that were conserved in all 15 genomes. Among these proteins, 5 and 23 were shared only by human or fish strains, respectively. These potential vaccine targets may help develop effective vaccines that will help prevent S. agalactiae infection.
Assuntos
Peixes/genética , Imunoterapia Ativa , Mastite Bovina/prevenção & controle , Infecções Estreptocócicas/prevenção & controle , Animais , Bovinos , Simulação por Computador , Feminino , Genoma Bacteriano , Humanos , Mastite Bovina/genética , Mastite Bovina/microbiologia , Terapia de Alvo Molecular , Infecções Estreptocócicas/genética , Infecções Estreptocócicas/imunologia , Infecções Estreptocócicas/veterinária , Streptococcus agalactiae/efeitos dos fármacos , Streptococcus agalactiae/genética , Streptococcus agalactiae/imunologia , Streptococcus agalactiae/patogenicidadeRESUMO
BACKGROUND/AIM: Small cell neuroendocrine carcinoma of the cervix (SCNEC) is a rare disease characterized by a higher incidence of lymphatic invasion, metastasis, and recurrence compared to the squamous cell carcinoma and adenocarcinoma subtypes. Furthermore, it is prone to early distant metastasis and has a poor prognosis. Chemotherapy has an important role in the management of cervical SCNEC. The effective treatment schemes for early-stage SCNEC are local treatment with radical surgery and systemic chemotherapy. However no standard treatment regimen exists because of a rare disease. We reviewed previous reports to determine whether etoposide/platinum, which is used for histopathologically similar small cell carcinoma of the lung, is an appropriate initial chemotherapy regimen for SCNEC of the cervix. MATERIALS AND METHODS: In this review the Cochrane library sources, ClinicalTrials.gov, Web of Silence, PubMed and search engine of Google scholar were searched for all interventional studies, reviews, case reports and meta-analyses published in 1997-2021. RESULTS: Etoposide/platinum (EP) is the most commonly used regimen and paclitaxel/carboplatin is the second most common, used as a part of multimodality therapy for SCNEC of the cervix in most studies. Cisplatin/vincristine/bleomycin, cisplatin/irinotecan, cisplatin/ifosfamide/etoposide were also reported in concurrence with EP; however no clinical trials are dedicated to SCNEC. CONCLUSION: Etoposide and platinum tend to have a better prognosis compared to other regimens used for other subtypes of cervical cancer. For recurrent cervical SCNEC, treatment options for patients are very limited. The application of molecular testing for targeted mutations may suggest potential future therapies that may be useful in this disease.
Assuntos
Carcinoma Neuroendócrino , Carcinoma de Células Pequenas , Neoplasias Pulmonares , Carcinoma de Pequenas Células do Pulmão , Neoplasias do Colo do Útero , Feminino , Humanos , Carcinoma de Células Pequenas/patologia , Colo do Útero/patologia , Cisplatino , Neoplasias do Colo do Útero/patologia , Carcinoma Neuroendócrino/patologia , Etoposídeo/uso terapêutico , Platina/uso terapêutico , Doenças Raras/tratamento farmacológico , Estadiamento de Neoplasias , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Carcinoma de Pequenas Células do Pulmão/tratamento farmacológico , Ifosfamida/uso terapêutico , Neoplasias Pulmonares/patologiaRESUMO
Invasive infections caused by Corynebacterium diphtheriae in vaccinated and non-vaccinated individuals have been reported increasingly. In this study we used multilocus sequence typing (MLST) to study genetic relationships between six invasive strains of this bacterium isolated solely in the urban area of Rio de Janeiro, Brazil, during a 10-year period. Of note, all the strains rendered negative results in PCR reactions for the tox gene, and four strains presented an atypical sucrose-fermenting ability. Five strains represented new sequence types. MLST results did not support the hypothesis that invasive (sucrose-positive) strains of C. diphtheriae are part of a single clonal complex. Instead, one of the main findings of the study was that such strains can be normally found in clonal complexes with strains related to non-invasive disease. Comparative analyses with C. diphtheriae isolated in different countries provided further information on the geographical circulation of some sequence types.
Assuntos
Corynebacterium diphtheriae/genética , Difteria/microbiologia , Adolescente , Adulto , Idoso , Técnicas de Tipagem Bacteriana , Brasil/epidemiologia , Criança , Difteria/epidemiologia , Feminino , Humanos , Masculino , Tipagem de Sequências MultilocusRESUMO
Spontaneous progressive nephropathy dominated by glomerular lesions in common marmosets has been reported. However, the histopathologic characteristics, including the relationship between glomerular and tubulointerstitial lesions, have not been described in detail. In the present study, the authors examined the histopathologic characteristics of the background renal lesions in common marmosets (3 males and 9 females, 3 to 8 years old). The severity of glomerular lesions was graded into 3 classes: grade I, no alteration; grade II, hilar/focal increase of mesangial matrix; grade III, global/diffuse increase of mesangial matrix. Tubulointerstitial lesions (tubular regeneration and hyperplasia and interstitial inflammation and fibrosis) were scored according to the area of each lesion. The renal lesions were characterized by enlargement of glomeruli, expanded mesangial area with increase of periodic acid-Schiff reaction-positive matrix, tubular regeneration and hyperplasia, and interstitial inflammation and fibrosis. Glomerular lesions progressed with increasing mesangial matrix and aging. Additionally, the tubulointerstitial lesions became exacerbated with progressing glomerular lesions. Tubular hyperplasia was divided into 4 types according to the structure of the cell layer (simple or stratified-like), the area of increased lining cells (partial or entire), cytoplasmic staining (eosinophilic or basophilic), brush border and thickness of basement membrane, and the activity of cell proliferation. In conclusion, the background renal lesions in common marmosets were characterized by glomerular lesions with increase of mesangial matrix, which progressed with aging, and secondary tubulointerstitial lesions, including tubular hyperplasia. Those lesions were thus diagnosed as progressive glomerulonephropathy in common marmosets.
Assuntos
Callithrix , Hiperplasia/veterinária , Nefropatias/veterinária , Rim/patologia , Doenças dos Macacos/patologia , Fatores Etários , Animais , Membrana Basal/patologia , Feminino , Fibrose/patologia , Fibrose/veterinária , Mesângio Glomerular/patologia , Hiperplasia/patologia , Imuno-Histoquímica/veterinária , Nefropatias/patologia , Glomérulos Renais/patologia , Túbulos Renais/patologia , MasculinoRESUMO
Lactic acid bacteria (LAB) are an attractive and safe alternative for the expression of heterologous proteins, as they are nonpathogenic and endotoxin-free organisms. Lactococcus lactis, the LAB model organism, has been extensively employed in the biotechnology field for large-scale production of heterologous proteins, and its use as a "cell factory" has been widely studied. We have been particularly interested in the use of L. lactis for production of heat shock proteins (HSPs), which reportedly play important roles in the initiation of innate and adaptive immune responses. However, this activity has been questioned, as LPS contamination appears to be responsible for most, if not all, immunostimulatory activity of HSPs. In order to study the effect of pure HSPs on the immune system, we constructed recombinant L. lactis strains able to produce and properly address the Mycobacterium leprae 65-kDa HSP (Hsp65) to the cytoplasm or to the extracellular medium, using a xylose-induced expression system. Approximately 7 mg/L recombinant Hsp65 was secreted. Degradation products related to lactococcal HtrA activity were not observed, and the Limulus amebocyte lysate assay demonstrated that the amount of LPS in the recombinant Hsp65 preparations was 10-100 times lower than the permitted levels established by the U.S. Food and Drug Administration. These new L. lactis strains will allow investigation of the effects of M. leprae Hsp65 without the interference of LPS; consequently, they have potential for a variety of biotechnological, medical and therapeutic applications.
Assuntos
Proteínas de Bactérias/genética , Chaperonina 60/genética , Lactococcus lactis/metabolismo , Mycobacterium leprae/metabolismo , Sequência de Bases , Clonagem Molecular , Primers do DNA , Lactococcus lactis/genética , Mycobacterium leprae/genética , Reação em Cadeia da Polimerase , Proteínas Recombinantes/genéticaRESUMO
Corynebacterium pseudotuberculosis, the infectious agent of caseous lymphadenitis (CLA), is responsible for substantial economic losses in goat and sheep production. Molecular characterization of C. pseudotuberculosis isolates by enterobacterial repetitive intergenic consensus (ERIC)-PCR has shown promising results in genotyping strains isolated from sheep with CLA. We evaluated the genetic diversity of C. pseudotuberculosis isolates collected from the Sertão region of the Pernambuco (PE) State, Brazil, and investigated the potential of ERIC-PCR as a tool for the molecular typing of strains of C. pseudotuberculosis isolated from goats. Thirty-two C. pseudotuberculosis strains isolated from goats in the municipalities of Floresta and Ibimirim, PE, C. pseudotuberculosis type strain ATCC 19410, the 1002 vaccine strain, and a field isolate of Rhodococcus equi were fingerprinted using the primers ERIC-1R and ERIC-2 and the primer pair ERIC- 1R+ERIC-2. Using 100% similarity as the cutoff, 8, 10, and 7 genotypes were obtained with ERIC-1-PCR, ERIC-2-PCR, and ERIC-1+2-PCR, respectively. The Hunter-Gaston discriminatory index calculated for the ERIC-1-PCR was 0.75. The index for the ERIC-2-PCR was 0.88, and the index for the ERIC-1+2-PCR was 0.79. Among goat isolates of C. pseudotuberculosis, three, two and four genotypes (found by ERIC-1-PCR, ERIC-2-PCR, and ERIC-1+2-PCR, respectively) had been previously described among sheep isolates from Minas Gerais State, Brazil. These results showed that ERIC-PCR has good discriminatory power and typeability, making it a useful tool for discrimination among C. pseudotuberculosis isolates from goats.
Assuntos
Sequência Consenso/genética , Corynebacterium pseudotuberculosis/genética , Corynebacterium pseudotuberculosis/isolamento & purificação , DNA Intergênico/genética , Cabras/microbiologia , Reação em Cadeia da Polimerase/métodos , Sequências Repetitivas de Ácido Nucleico/genética , Animais , Brasil , Análise por Conglomerados , Impressões Digitais de DNA , Enterobacteriaceae/genéticaRESUMO
Enterococcus faecalis is a natural inhabitant of the human gastrointestinal tract. This bacterial species is subdominant in a healthy physiological state of the gut microbiota (eubiosis) in adults, but can become dominant and cause infections when the intestinal homeostasis is disrupted (dysbiosis). The relatively high concentrations of bile acids deoxycholate (DCA) and taurocholate (TCA) hallmark eubiosis and dysbiosis, respectively. This study aimed to better understand how E. faecalis adapts to DCA and TCA. We showed that DCA impairs E. faecalis growth and possibly imposes a continuous adjustment in the expression of many essential genes, including a majority of ribosomal proteins. This may account for slow growth and low levels of E. faecalis in the gut. In contrast, TCA had no detectable growth effect. The evolving transcriptome upon TCA adaptation showed the early activation of an oligopeptide permease system (opp2) followed by the adjustment of amino acid and nucleotide metabolisms. We provide evidence that TCA favors the exploitation of oligopeptide resources to fuel amino acid needs in limiting oligopeptide conditions. Altogether, our data suggest that the combined effects of decreased DCA and increased TCA concentrations can contribute to the rise of E. faecalis population during dysbiosis.
Assuntos
Ácidos e Sais Biliares , Enterococcus faecalis , Aminoácidos/metabolismo , Ácidos e Sais Biliares/metabolismo , Ácido Desoxicólico/metabolismo , Ácido Desoxicólico/farmacologia , Disbiose , Enterococcus faecalis/genética , Humanos , Ácido Taurocólico/metabolismo , Ácido Taurocólico/farmacologiaRESUMO
During the last decades, the majority of Brazilian Corynebacterium diphtheriae isolates were shown to be capable to metabolize sucrose, sometimes leading to erroneous identification as a non-diphtheric Corynebacterium species. The sequencing of the polymorphic region of the RNA polymerase beta subunit-encoding gene (rpoB) is an important taxonomic tool for identification of corynebacteria. The present study aimed to investigate the rpoB gene polymorphic features of sucrose-fermenting and non sucrose-fermenting strains. The results showed that sucrose-fermenting strains presented rpoB gene polymorphic regions with more than 98% similarity with the sequences deposited in the gene bank corresponding to non sucrose-fermenting strains. Data indicate that sucrose-fermenting isolates may act as a variant of C. diphtheriae biotype mitis. In addition we alert that sucrose-fermenting strains should not be discarded as contaminants mainly in countries where the possibility of isolation of this variant is higher.
Assuntos
Proteínas de Bactérias/metabolismo , Corynebacterium diphtheriae/genética , Corynebacterium diphtheriae/metabolismo , Sacarose/metabolismo , Proteínas de Bactérias/classificação , Proteínas de Bactérias/genética , Fermentação , FilogeniaRESUMO
The large number of microbial genomes deposited in databanks has opened the door for in-depth studies of organisms, including post-genomics investigations. Thanks to new generation sequencing technology, these studies have made advances that have lead to extraordinary discoveries in bacterial transcriptomics. In this review, we describe bacterial RNA sequencing studies that use these new techniques. We also examined the advantages and biases of these new generation technologies; advances in bioinformatics make it possible to overcome the biases, providing interesting and surprising results.
Assuntos
Bactérias/genética , Genoma Bacteriano , RNA Bacteriano/química , Transcriptoma , Sequência de Bases , RNA Bacteriano/genética , Análise de Sequência de RNA/métodosRESUMO
Despite the existence of a vaccine against diphtheria, this disease remains endemic and is reemerging in several regions due to many factors, including variations in genes coding for virulence factors. One common feature of virulence factors is their high concentration in pathogenicity islands (PAIs), very unstable regions acquired via horizontal gene transfer, which has lead to the emergence of various bacterial pathogens. The 13 putative PAIs in Corynebacterium diphtheriae NCTC 13129 and the reemergence of this disease point to the great variability in the PAIs of this species, which may reflect on bacterial life style and physiological versatility. We investigated the relationships between the large number of PAIs in C. diphtheriae and the possible implications of their plasticity in virulence. The GenoFrag software was used to design primers to analyze the genome plasticity of two pathogenicity islands of the reference strain (PiCds 3 and 8) in 11 different strains. We found that PiCd 3 was absent in only two strains, showing genes playing putative important roles in virulence and that only one strain harbored PiCd 8, due to its location in a putative "hotspot" for horizontal gene transfer events.
Assuntos
Corynebacterium diphtheriae/patogenicidade , Reação em Cadeia da Polimerase/métodos , Virulência/genética , Sequência de Bases , Corynebacterium diphtheriae/genética , Primers do DNA , Genes BacterianosRESUMO
BACKGROUND: Around 300 million people world-wide suffer from asthma, and the prevalence of allergic diseases has increased. Much effort has been used in the study of mechanisms involved in the immune response observed in asthma to intervene for the treatment of this condition. During inflammation in asthma, Th2 cytokines and eosinophils are essential components of the host immune system. Furthermore, for therapeutic interventions against this disease, IL-10 is an important cytokine because it has a central role in the regulation of inflammatory cascades. OBJECTIVE: To evaluate the immunomodulatory effect of Lactococcus lactis strains expressing recombinant IL-10 in a mouse model of ovalbumin (OVA)-induced acute airway inflammation. METHODS: L. lactis expressing recombinant IL-10 in a cytoplasmic (LL-CYT) or secreted form (LL-SEC) and wild-type (LL-WT) were used. IL-10 production by the recombinant strains was evaluated by ELISA. After an intranasal administration of L. lactis producing recombinant IL-10 and the induction of acute allergic airway inflammation in mice, blood samples were collected to detect IgE anti-OVA, and bronchoalveolar lavage (BAL) was harvested for eosinophil count. Additionally, the lungs were collected for the detection of the eosinophil peroxidase (EPO) activity, measurement of cytokines and chemokines and evaluation of pathology. RESULTS: Mice that received LL-CYT and LL-SEC strains showed a significant decrease in eosinophils numbers, EPO activity, anti-OVA IgE and IgG1 levels, IL-4 and CCL3 production and pulmonary inflammation and mucus hypersecretion, compared with the asthmatic group. Only the LL-CYT/OVA group showed reduced levels of IL-5, CCL2, CCL5 and CCL11. CONCLUSION: Treatment with L. lactis producing recombinant IL-10 used in this study (LL-CYT and LL-SEC) modulated experimental airway inflammation in the mouse model independently of Treg cells. Additionally, the LL-CYT strain was more efficient in the suppression of lung inflammation.
Assuntos
Terapia Genética/métodos , Hipersensibilidade/imunologia , Interleucina-10/biossíntese , Lactococcus lactis/genética , Pneumonia/imunologia , Administração Intranasal , Animais , Asma/imunologia , Asma/patologia , Separação Celular , Citocinas/análise , Citocinas/imunologia , Modelos Animais de Doenças , Ensaio de Imunoadsorção Enzimática , Feminino , Citometria de Fluxo , Vetores Genéticos , Hipersensibilidade/patologia , Imunoglobulina E/sangue , Imunoglobulina E/imunologia , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Imunoterapia/métodos , Interleucina-10/genética , Interleucina-10/imunologia , Pulmão/imunologia , Pulmão/patologia , Camundongos , Camundongos Endogâmicos BALB C , Ovalbumina/imunologia , Pneumonia/patologia , Proteínas Recombinantes/imunologia , Células Th2/imunologiaRESUMO
Previously, we isolated two strains of spontaneous oxidative (SpOx2 and SpOx3) stress mutants of Lactococcus lactis subsp cremoris. Herein, we compared these mutants to a parental wild-type strain (J60011) and a commercial starter in experimental fermented milk production. Total solid contents of milk and fermentation temperature both affected the acidification profile of the spontaneous oxidative stress-resistant L. lactis mutants during fermented milk production. Fermentation times to pH 4.7 ranged from 6.40 h (J60011) to 9.36 h (SpOx2); V(max) values were inversely proportional to fermentation time. Bacterial counts increased to above 8.50 log(10) cfu/mL. The counts of viable SpOx3 mutants were higher than those of the parental wild strain in all treatments. All fermented milk products showed post-fermentation acidification after 24 h of storage at 4 degrees C; they remained stable after one week of storage.