RESUMO
The association of vaccines with immunostimulants such as ß-glucan, promote the production of cytokines, competent immune cells and antibodies. However, differences between ß-glucan types and trials make it difficult to understand ß-glucan's mechanism of action. In this study, three trials were carried out with control and fish fed ß-glucan, the first trial occurred at 15 days; the second trial occurred at 30 days when we associated ß-glucan and vaccine; and the third trial occurred at 15 days post-challenge with Streptococcus agalactiae in tilapia (O. niloticus) in order to investigate immune-related gene expression in the head kidney and spleen using real-time qPCR. We found increases in HSP70, IL-6, IL-1ß, TNF-α, IL-10, Lys and C3 predominantly in the head kidney, except for IgM expression, which prevailed in the spleen, under vaccinated + ß-glucan action. This demonstrates the trade-off presented by the head kidney and spleen after immunostimulation in order to produce acquired immunity, as well as an increase in HSP70 expression in vaccinated + ß-glucan fish. The results suggest that ß-glucan stimulates the immune response through damage-associated molecular patterns (DAMPs) recognition. Therefore, these dynamics of the immune response promote a more robust defense against disease.
Assuntos
Adjuvantes Imunológicos/administração & dosagem , Ciclídeos/imunologia , Rim Cefálico/efeitos dos fármacos , Baço/efeitos dos fármacos , Vacinas Estreptocócicas/administração & dosagem , beta-Glucanas/administração & dosagem , Imunidade Adaptativa , Animais , Ciclídeos/genética , Ciclídeos/microbiologia , Citocinas/genética , Doenças dos Peixes/genética , Doenças dos Peixes/imunologia , Doenças dos Peixes/prevenção & controle , Proteínas de Peixes/genética , Expressão Gênica/efeitos dos fármacos , Proteínas de Choque Térmico HSP70/genética , Rim Cefálico/imunologia , Muramidase/imunologia , Transdução de Sinais , Baço/imunologia , Infecções Estreptocócicas/genética , Infecções Estreptocócicas/imunologia , Infecções Estreptocócicas/prevenção & controle , Infecções Estreptocócicas/veterinária , Streptococcus agalactiaeRESUMO
This study aimed to evaluate the transcriptional changes occurring in isolated perfused mammary alveolar tissue in response to inoculation with S. agalactiae and to identify the most affected biological functions and pathways after 3 h. Four udders taken at slaughter from cows with healthy mammary gland were perfused ex situ with warmed and gassed Tyrode's solution. Mammary alveolar tissue samples were taken from the left fore and rear quarters (IQ-inoculated quarters) before inoculation (hour 0) and at 3 h post inoculation (hpi) and at the same times from control right fore and rear quarters (not inoculated: NIQ). A total of 1756 differentially expressed genes (DEGs) were identified between IQ and NIQ at 3 hpi using edgeR package. Within this set of DEGs, 952 were up regulated and mainly involved with innate immune response and inflammatory response, e.g., CD14, CCL5, TLR2, IL-8, SAA3, as well as in transcriptional regulation such as FOS, STAT3 and NFKBIA. Genes down-regulated (804) included those involved with lipid synthesis e.g., APOC2, SCD, FABP3 and FABP4. The most affected pathways were chemokine signaling, Wnt signaling and complement and coagulation cascades, which likely reflects the early stage response of mammary tissue to S. agalactiae infection. No significant gene expression changes were detected by RNA-Seq in the others contrasts. Real time-PCR confirmed the increase in mRNA abundance of immune-related genes: TLR2, TLR4, IL-1ß, and IL-10 at 3 hpi between IQ and NIQ. The expression profiles of Casp1 and Bax for any contrasts were unaffected whereas Bcl2 was increased in IQ, which suggests no induction of apoptosis during the first hours after infection. Results provided novel information regarding the early functional pathways and gene network that orchestrate innate immune responses to S. agalactiae infection. This knowledge could contribute to new strategies to enhance resistance to this disease, such as genomic selection.
Assuntos
Perfilação da Expressão Gênica/veterinária , Glândulas Mamárias Animais/metabolismo , Infecções Estreptocócicas/veterinária , Streptococcus agalactiae , Animais , Bovinos , Feminino , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica , Imunidade/genética , Inflamação/genética , Mastite Bovina/genética , Mastite Bovina/imunologia , Mastite Bovina/microbiologia , Reação em Cadeia da Polimerase em Tempo Real/veterinária , Infecções Estreptocócicas/genética , Infecções Estreptocócicas/imunologiaRESUMO
BACKGROUND: Streptococcus agalactiae (GBS) is a major pathogen of Nile tilapia, a global commodity of the aquaculture sector. The aims of this study were to evaluate protein expression in the main genotypes of GBS isolated from diseased fishes in Brazil using a label-free shotgun nano-liquid chromatography-ultra definition mass spectrometry (nanoLC-UDMSE) approach and to compare the differential abundance of proteins identified in strains isolated from GBS-infected fishes and humans. RESULTS: A total of 1070 protein clusters were identified by nanoLC-UDMSE in 5 fish-adapted GBS strains belonging to sequence types ST-260 and ST-927 and the non-typeable (NT) lineage and 1 human GBS strain (ST-23). A total of 1065 protein clusters corresponded to the pan-proteome of fish-adapted GBS strains; 989 of these were identified in all fish-adapted GBS strains (core proteome), and 62 were shared by at least two strains (accessory proteome). Proteins involved in the stress response and in the regulation of gene expression, metabolism and virulence were detected, reflecting the adaptive ability of fish-adapted GBS strains in response to stressor factors that affect bacterial survival in the aquatic environment and bacterial survival and multiplication inside the host cell. Measurement of protein abundance among different hosts showed that 5 and 26 proteins were exclusively found in the human- and fish-adapted GBS strains, respectively; the proteins exclusively identified in fish isolates were mainly related to virulence factors. Furthermore, 215 and 269 proteins were up- and down-regulated, respectively, in the fish-adapted GBS strains in comparison to the human isolate. CONCLUSIONS: Our study showed that the core proteome of fish-adapted GBS strains is conserved and demonstrated high similarity of the proteins expressed by fish-adapted strains to the proteome of the human GBS strain. This high degree of proteome conservation of different STs suggests that, a monovalent vaccine may be effective against these variants.
Assuntos
Doenças dos Peixes/genética , Proteoma/genética , Infecções Estreptocócicas/genética , Streptococcus agalactiae/genética , Animais , Brasil , Ciclídeos/genética , Ciclídeos/microbiologia , Doenças dos Peixes/microbiologia , Genótipo , Humanos , Filogenia , Infecções Estreptocócicas/microbiologia , Streptococcus agalactiae/patogenicidade , Virulência/genéticaRESUMO
The aim of this study was to elucidate the differential gene expression in the RNA sequencing transcriptome of isolated perfused udders collected from 4 slaughtered Holstein × Zebu crossbred dairy cows experimentally inoculated with Streptococcus agalactiae. We studied 3 different statistical tools (edgeR, baySeq, and Cuffdiff 2). In summary, 2 quarters of each udder were experimentally inoculated with Strep. agalactiae and the other 2 were used as a control. Mammary tissue biopsies were collected at times 0 and 3 h after infection. The total RNA was extracted and sequenced on an Illumina HiSeq 2000 (Illumina Inc., San Diego, CA). Transcripts were assembled from the reads aligned to the bovine UMD 3.1 reference genome, and the statistical analyses were performed using the previously mentioned tools (edgeR, baySeq, and Cuffdiff 2). Finally, the identified genes were submitted to pathway enrichment analysis. A total of 1,756, 1,161, and 3,389 genes with differential gene expression were identified when using edgeR, baySeq, and Cuffdiff 2, respectively. A total of 122 genes were identified by the overlapping of the 3 methods; however, only the platelet activation presented a significantly enriched pathway. From the results, we suggest the FCER1G, GNAI2, ORAI1, and VASP genes shared among the 3 methods in this pathway for posterior biological validation.
Assuntos
Glândulas Mamárias Animais/microbiologia , Mastite Bovina/genética , RNA/genética , Infecções Estreptocócicas/veterinária , Streptococcus agalactiae/fisiologia , Animais , Bovinos , Feminino , Genoma , Glândulas Mamárias Animais/metabolismo , Mastite Bovina/metabolismo , Mastite Bovina/microbiologia , RNA/metabolismo , Análise de Sequência de RNA , Infecções Estreptocócicas/genética , Infecções Estreptocócicas/metabolismo , Infecções Estreptocócicas/microbiologia , TranscriptomaRESUMO
Group B Streptococcus (GBS) carriage by pregnant women is the primary risk factor for early-onset GBS neonatal sepsis. Intrapartum antibiotic prophylaxis (IAP) can prevent this transmission route, and two main approaches are recommended to base the selection of pregnant women to be submitted to IAP: the risk-based and the culture-based strategies. In Brazil, compliance to such recommendations is poor, and not much is known about GBS carriage. In the present study, 3,647 pregnant women living in Rio de Janeiro State, Brazil, were screened for GBS anogenital colonization, over a period of 8 years (2008-2015). GBS was detected in 956 (26.2%) of them, and presence of vaginal discharge was the only trait associated with a higher risk for GBS colonization. Serotypes Ia (257; 37.3%) and II (137; 19.9%) were the most frequent among 689 (72.1% of the total) GBS isolates evaluated, followed by NT isolates (84; 12.1%), serotype Ib (77; 11.1%), V (63; 9.1%), III (47; 6.8%) and IV (24; 3.5%). Estimated coverage of major serotype-based GBS vaccines currently under clinical trials would vary from 65.2% to 84.3%. All 689 isolates tested were susceptible to ampicillin and vancomycin. Resistance to chloramphenicol, clindamycin, erythromycin, levofloxacin, and tetracycline was observed in 5% (35), 2% (14), 14% (97), 5% (35) and 86% (592) of the isolates, respectively. No significant fluctuations in colonization rates, serotype distribution and antimicrobial susceptibility profiles were observed throughout the period of time investigated. The culture-based approach for IAP recommendation showed to be the best choice for the population investigated when compared to the risk-based, since the first did not increase the number of pregnant women submitted to antibiotic therapy and covered a larger number of women who were actually colonized by GBS. The fact the not all isolates were available for additional characterization, and serotype IX antiserum was not available for testing represent limitations of this study. Nevertheless, to the best of our knowledge, this is the largest investigation on GBS carriage among pregnant women in Brazil up to date, and results are useful for improving GBS prevention and treatment strategies.
Assuntos
Complicações Infecciosas na Gravidez , Infecções Estreptocócicas , Streptococcus agalactiae , Adolescente , Adulto , Brasil/epidemiologia , Feminino , Humanos , Gravidez , Complicações Infecciosas na Gravidez/tratamento farmacológico , Complicações Infecciosas na Gravidez/epidemiologia , Complicações Infecciosas na Gravidez/microbiologia , Complicações Infecciosas na Gravidez/patologia , Fatores de Risco , Infecções Estreptocócicas/tratamento farmacológico , Infecções Estreptocócicas/epidemiologia , Infecções Estreptocócicas/genética , Infecções Estreptocócicas/patologia , Streptococcus agalactiae/genética , Streptococcus agalactiae/isolamento & purificaçãoRESUMO
BACKGROUND: Mastitis resistance is a complex and multifactorial trait, and its expression depends on both genetic and environmental factors, including infection pressure. The objective of this research was to determine the genetic basis of mastitis resistance to specific pathogens using a repeatability threshold probit animal model. RESULTS: The most prevalent isolated pathogens were coagulase-negative staphylococci (CNS); 39 % of records and 77 % of the animals infected at least one time in the whole period of study. There was significant genetic variation only for Streptococci (STR). In addition, there was a positive genetic correlation between STR and all pathogens together (ALL) (0.36 ± 0.22), and CNS and ALL (0.92 ± 0.04). CONCLUSION: The results of our study support the presence of significant genetic variation for mastitis caused by Streptococci and suggest the importance of discriminating between different pathogens causing mastitis due to the fact that they most likely influence different genetic traits. Low heritabilities for pathogen specific-mastitis resistance may be considered when including bacteriological status as a measure of mastitis presence to implement breeding strategies for improving udder health in dairy ewes.
Assuntos
Resistência à Doença/genética , Variação Genética , Mastite/veterinária , Doenças dos Ovinos/genética , Doenças dos Ovinos/microbiologia , Infecções Estreptocócicas/veterinária , Animais , Cruzamento , Indústria de Laticínios , Feminino , Mastite/genética , Mastite/microbiologia , Mastite/prevenção & controle , Ovinos , Doenças dos Ovinos/prevenção & controle , Infecções Estreptocócicas/genética , Infecções Estreptocócicas/microbiologia , Infecções Estreptocócicas/prevenção & controle , Streptococcus/genética , Streptococcus/isolamento & purificaçãoRESUMO
Single-nucleotide polymorphisms (SNPs) are the most common source of genetic variation within a species; however, few investigations demonstrate how naturally occurring SNPs may increase strain virulence. We recently used group A Streptococcus as a model pathogen to study bacteria strain genotype-patient disease phenotype relationships. Whole-genome sequencing of approximately 800 serotype M59 group A Streptococcus strains, recovered during an outbreak of severe invasive infections across North America, identified a disproportionate number of SNPs in the gene encoding multiple gene regulator of group A Streptococcus (mga). Herein, we report results of studies designed to test the hypothesis that the most commonly occurring SNP, encoding a replacement of arginine for histidine at codon 201 of Mga (H201R), significantly increases virulence. Whole transcriptome analysis revealed that the H201R replacement significantly increased expression of mga and 54 other genes, including many proven virulence factors. Compared to the wild-type strain, a H201R isogenic mutant strain caused significantly larger skin lesions in mice. Serial quantitative bacterial culture and noninvasive magnetic resonance imaging also demonstrated that the isogenic H201R strain was significantly more virulent in a nonhuman primate model of joint infection. These findings show that the H201R replacement in Mga increases the virulence of M59 group A Streptococcus and provide new insight to how a naturally occurring SNP in bacteria contributes to human disease phenotypes.
Assuntos
Proteínas de Bactérias , Artropatias , Mutação de Sentido Incorreto , Polimorfismo de Nucleotídeo Único , Infecções Estreptocócicas , Streptococcus pyogenes , Substituição de Aminoácidos , Animais , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Linhagem Celular , Feminino , Genoma Bacteriano , Humanos , Artropatias/genética , Artropatias/metabolismo , Artropatias/microbiologia , Artropatias/patologia , Camundongos , Camundongos Pelados , Infecções Estreptocócicas/genética , Infecções Estreptocócicas/metabolismo , Infecções Estreptocócicas/patologia , Streptococcus pyogenes/genética , Streptococcus pyogenes/metabolismo , Streptococcus pyogenes/patogenicidadeRESUMO
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
Infection with Streptococcus pyogenes (S. pyogenes) can result in several diseases, particularly in children. S. pyogenes M protein is the major virulence factor, and certain regions of its N-terminus can trigger autoimmune sequelae such as rheumatic fever in susceptible individuals with untreated group A streptococcal pharyngitis. In a previous study, we utilized a large panel of human peripheral blood cells to define the C-terminal protective epitope StreptInCor (medical identity), which does not induce autoimmune reactions. We recently confirmed the results in HLA-transgenic mice. In the present study, we extended the experimental assays to outbred animals (Swiss mice). Herein, we demonstrate high titers of StreptInCor-specific antibodies, as well as appropriate T-cell immune responses. No cross-reaction to cardiac myosin was detected. Additionally, immunized Swiss mice exhibited 87% survival one month after challenge with S. pyogenes. In conclusion, the data presented herein reinforce previous results in humans and animals and further emphasize that StreptInCor could be an effective and safe vaccine for the prevention of S. pyogenes infections.
Assuntos
Vacinas Bacterianas/imunologia , Cruzamento , Epitopos/imunologia , Infecções Estreptocócicas/prevenção & controle , Streptococcus pyogenes/imunologia , Streptococcus pyogenes/fisiologia , Vacinas de Subunidades Antigênicas/imunologia , Sequência de Aminoácidos , Animais , Especificidade de Anticorpos , Aderência Bacteriana/imunologia , Vacinas Bacterianas/química , Linhagem Celular , Proliferação de Células , Feminino , Humanos , Imunização , Imunoglobulina G/imunologia , Camundongos , Dados de Sequência Molecular , Polimorfismo Genético , Especificidade da Espécie , Baço/citologia , Infecções Estreptocócicas/sangue , Infecções Estreptocócicas/genética , Vacinas de Subunidades Antigênicas/químicaRESUMO
Publicly available genomic data are a great source of biological knowledge that can be extracted when appropriate data analysis is used. Predicting the biological function of genes is of interest to understand molecular mechanisms of virulence and resistance in pathogens and hosts and is important for drug discovery and disease control. This is commonly done by searching for similar gene expression behavior. Here, we used publicly available Streptococcus pyogenes microarray data obtained during primate infection to identify genes that have a potential influence on virulence and Phytophtora infestance inoculated tomato microarray data to identify genes potentially implicated in resistance processes. This approach goes beyond co-expression analysis. We employed a quasi-likelihood model separated by primate gender/inoculation condition to model median gene expression of known virulence/resistance factors. Based on this model, an influence analysis considering time course measurement was performed to detect genes with atypical expression. This procedure allowed for the detection of genes potentially implicated in the infection process. Finally, we discuss the biological meaning of these results, showing that influence analysis is an efficient and useful alternative for functional gene prediction.