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1.
Vet Res ; 50(1): 69, 2019 Sep 23.
Artigo em Inglês | MEDLINE | ID: mdl-31547880

RESUMO

Haemophilus parasuis is part of the microbiota of the upper respiratory tract in swine. However, virulent strains can cause a systemic disease known as Glässer's disease. Several virulence factors have been described in H. parasuis including the virulence-associated trimeric autotransporters (VtaAs). VtaA2 is up-regulated during infection and is only found in virulent strains. In order to determine its biological function, the vtaA2 gene was cloned with its native promotor region in pACYC184, and the transformed Escherichia coli was used to perform functional in vitro assays. VtaA2 was found to have a role in attachment to plastic, mucin, BSA, fibronectin and collagen. As other VtaAs from H. parasuis, the passenger domain of VtaA2 contains collagen domains. In order to examine the contribution of the collagen repeats to VtaA2 function, a recombinant vtaA2 without the central collagen domains was obtained and named vtaA2OL. VtaA2OL showed similar capacity than VtaA2 to adhere to plastic, mucin, BSA, fibronectin and plasma but a reduced capacity to adhere to collagen, suggesting that the collagen domains of VtaA2 are involved in collagen attachment. No function in cell adhesion and invasion to epithelial alveolar cell line A549 or unspecific binding to primary alveolar macrophages was found. Likewise VtaA2 had no role in serum or phagocytosis resistance. We propose that VtaA2 mediates adherence to the host by binding to the mucin, found in the upper respiratory tract mucus, and to the extracellular matrix proteins, present in the connective tissue of systemic sites, such as the serosa.


Assuntos
Aderência Bacteriana/genética , Infecções por Haemophilus/veterinária , Haemophilus parasuis/fisiologia , Doenças dos Suínos/microbiologia , Fatores de Virulência/genética , Animais , Escherichia coli/genética , Infecções por Haemophilus/microbiologia , Suínos , Virulência/genética
2.
BMC Vet Res ; 15(1): 272, 2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-31370843

RESUMO

BACKGROUND: Ceftiofur Sodium is widely used in China. Our aim was to determine Ceftiofur Sodium activity and optimize dosing regimens against the pathogen Haemophilus parasuis using an in vitro and ex vivo pharmacokinetics/pharmacodynamics modeling approach. By adopting these strategies, we wanted to extend the effective life of Ceftiofur Sodium in reduce drug-resistance in pigs. RESULTS: We established an H. parasuis infection model in pigs, and assessed the pharmacokinetics of Ceftiofur Sodium in both healthy and infected animals. After Ceftiofur Sodium (10 mg/kg, i.m.) administration to healthy and H. parasuis-infected pigs, plasma based desfuroylceftiofur (a metabolite of Ceftiofur Sodium) was measured by High Performance Liquid Chromatography. The pharmacokinetics of Ceftiofur Sodium (desfuroylceftiofur) was consistent with a two-compartment open model, with first-order absorption. We observed no significant differences (P > 0.05) in pharmacokinetic parameters between healthy and infected pigs. Pharmacodynamics data showed that Ceftiofur Sodium was highly inhibitory against H. parasuis, with MIC, MBC, and MPC values of 0.003125, 0.0125 and 0.032 µg/mL, respectively. Desfuroylceftiofur in plasma also had strong bactericidal activity. Almost all H. parasuis cultured in plasma medium of Ceftiofur Sodium-inoculated healthy pigs, at each time point, were killed within 24 h. A weaker antibacterial activity was measured in infected-pig plasma medium at 18, 24, 36, and 48 h, after Ceftiofur Sodium inoculation. Pharmacokinetic parameters were combined with ex vivo pharmacodynamic parameters, and the bacteriostatic effect (36.006 h), bactericidal effect (71.637 h) and clearance (90.619 h) within 24 h, were determined using the Hill equation. Dose-calculation equations revealed the optimal dose of Ceftiofur Sodium to be 0.599-1.507 mg/kg. CONCLUSIONS: There were no significant differences in Ceftiofur Sodium pharmacokinetic parameters between healthy and infected pigs, although pharmacokinetics/pharmacodynamics fitting curves showed obviously differences. The optimal dose of Ceftiofur Sodium was lower than recommended (3 mg/kg), which may provide improved treatments for Glässers disease, with lower drug-resistance possibility.


Assuntos
Cefalosporinas , Infecções por Haemophilus/veterinária , Modelos Biológicos , Animais , Antibacterianos/farmacocinética , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Cefalosporinas/farmacocinética , Cefalosporinas/farmacologia , Cefalosporinas/uso terapêutico , Infecções por Haemophilus/tratamento farmacológico , Infecções por Haemophilus/microbiologia , Haemophilus parasuis/efeitos dos fármacos , Suínos , Doenças dos Suínos/tratamento farmacológico , Doenças dos Suínos/microbiologia
3.
BMC Vet Res ; 15(1): 294, 2019 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-31412870

RESUMO

BACKGROUND: Glaesserella parasuis (G. parasuis) is an influential pathogen of the pig, which induces high morbidity and mortality in naive pig populations in the pig industry. Accurate and rapid detection of the agent is important for disease control. In this study, a simple recombinase polymerase amplification (RPA) with a Lateral flow (LF) strip (RPA-LF-GPS) was developed to detect G. parasuis. RESULTS: The RPA-LF-GPS can specifically detect G. parasuis a limit of 100 CFU from other common related pathogens causing arthritis in the pig. The RPA-LF-GPS assay can use boiled synovial fluid samples as a template with the same sensitivity as other DNA extraction methods. In the detection of clinic positive synovial fluid sample, RPA-LF-GPS is equally sensitive (98.1%) compared with that of PCR (90.4%) (P > 0.05). The whole procedure of the RPA-LF-GPS assay could be finished in 1 hour without professional equipment. CONCLUSIONS: RPA-LF-GPS assay is a rapid and simple method for point-of-care diagnostic testing for G. parasuis infection.


Assuntos
Infecções por Haemophilus/veterinária , Haemophilus parasuis , Recombinases/metabolismo , Doenças dos Suínos/microbiologia , Líquido Sinovial/microbiologia , Animais , Infecções por Haemophilus/microbiologia , Técnicas de Amplificação de Ácido Nucleico/veterinária , Reação em Cadeia da Polimerase/veterinária , Recombinases/química , Recombinases/genética , Sensibilidade e Especificidade , Suínos , Doenças dos Suínos/diagnóstico
4.
J Vet Diagn Invest ; 31(5): 714-718, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31347465

RESUMO

Avibacterium paragallinarum is the causative agent of infectious coryza, a highly contagious respiratory disease in chickens. Given its fastidious nature, this bacterium is difficult to recover and identify, particularly from locations colonized by normal bacterial flora. Standard PCR methods have been utilized for detection but are labor-intensive and not feasible for high-throughput testing. We evaluated a real-time PCR (rtPCR) method targeting the HPG-2 region of A. paragallinarum, and validated a high-throughput extraction for this assay. Using single-tube extraction, the rtPCR detected 4 A. paragallinarum (ATCC 29545T and 3 clinical) isolates with a limit of detection (LOD) of 10 cfu/mL and a PCR efficiency of 89-111%. Cross-reaction was not detected with 33 non-A. paragallinarum, all close relatives from the family Pasteurellaceae. Real-time PCR testing on extracts of 66 clinical samples (choana, sinus, or trachea) yielded 98.2% (35 of 36 on positives, 30 of 30 on negatives) agreement with conventional PCR. Duplicate samples tested in a 96-well format extraction in parallel with the single-tube method produced equivalent LOD on all A. paragallinarum isolates, and 96.8% agreement on 93 additional clinical samples extracted with both procedures. This A. paragallinarum rtPCR can be utilized for outbreak investigations and routine monitoring of susceptible flocks.


Assuntos
Galinhas/microbiologia , Infecções por Haemophilus/veterinária , Haemophilus paragallinarum/isolamento & purificação , Doenças das Aves Domésticas/microbiologia , Reação em Cadeia da Polimerase em Tempo Real/veterinária , Animais , Resfriado Comum/microbiologia , Resfriado Comum/veterinária , Infecções por Haemophilus/microbiologia , Haemophilus paragallinarum/genética , Sequenciamento de Nucleotídeos em Larga Escala/veterinária , Nasofaringe/microbiologia , Seios Paranasais/microbiologia , Pasteurellaceae , Doenças das Aves Domésticas/diagnóstico , Traqueia/microbiologia
5.
Artigo em Inglês | MEDLINE | ID: mdl-31174697

RESUMO

TLRs, Siglecs and CD163 are cell surface receptors that play an important role in immune response and sepsis. The objective of this study was to assess changes in the expression levels of several of these receptors (TLR2, TLR4, CD163, Siglec-1, Siglec-3, Siglec-5 and Siglec-10) on the surface of peripheral blood mononuclear cells from pigs with sepsis caused by Haemophilus parasuis. Flow cytometry was employed to analyze samples from an experimental infection and from cell cultures. A significant increase in CD163, TLR2 and Siglec-3 expression during infection was seen. However, in vitro exposure of peripheral blood monocytes to bacteria or sera from infected pigs did not increase the expression of these receptors. These changes may be due to recruitment of monocytes into the blood compartment in response to H. parasuis-induced sepsis.


Assuntos
Antígenos CD/genética , Antígenos de Diferenciação Mielomonocítica/genética , Infecções por Haemophilus/veterinária , Monócitos/imunologia , Receptores de Superfície Celular/genética , Sepse/veterinária , Lectina 3 Semelhante a Ig de Ligação ao Ácido Siálico/genética , Doenças dos Suínos/imunologia , Receptor 2 Toll-Like/genética , Animais , Antígenos CD/imunologia , Antígenos de Diferenciação Mielomonocítica/imunologia , Células Cultivadas , Infecções por Haemophilus/imunologia , Haemophilus parasuis , Monócitos/microbiologia , Receptores de Superfície Celular/imunologia , Sepse/imunologia , Sepse/microbiologia , Lectina 3 Semelhante a Ig de Ligação ao Ácido Siálico/imunologia , Suínos , Doenças dos Suínos/microbiologia , Receptor 2 Toll-Like/imunologia
6.
Artigo em Inglês | MEDLINE | ID: mdl-31106159

RESUMO

Bacterial infections activate autophagy and autophagy restricts pathogens such as Haemophilus parasuis through specific mechanisms. Autophagy is associated with the pathogenesis of H. parasuis. However, the mechanisms have not been clarified. Here, we monitored autophagy processes using confocal microscopy, western blot, and transmission electron microscopy (TEM) and found that H. parasuis SH0165 (high-virulent strain) but not HN0001 (non-virulent strain) infection enhanced autophagy flux. The AMPK/mTOR autophagy pathway was required for autophagy initiation and ATG5, Beclin-1, ATG7, and ATG16L1 emerged as important components in the generation of the autophagosome during H. parasuis infection. Moreover, autophagy induced by H. parasuis SH0165 turned to fight against invaded bacteria and inhibit inflammation. Then we further demonstrated that autophagy blocked the production of the cytokines IL-8, CCL4, and CCL5 induced by SH0165 infection through the inhibition of NF-κB, p38, and JNK MAPK signaling pathway. Therefore, our findings suggest that autophagy may act as a cellular defense mechanism in response to H. parasuis and provide a new way that autophagy protects the host against H. parasuis infection.


Assuntos
Autofagia , Mecanismos de Defesa , Células Epiteliais/imunologia , Células Epiteliais/microbiologia , Infecções por Haemophilus/veterinária , Haemophilus parasuis/imunologia , Inflamação/imunologia , Animais , Linhagem Celular , Citocinas/metabolismo , Infecções por Haemophilus/imunologia , Modelos Teóricos , Transdução de Sinais , Suínos
7.
Vet Microbiol ; 231: 40-44, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30955821

RESUMO

Haemophilus parasuis is a common colonizer of the upper respiratory tract of swine and frequently causes disease, especially in weaner pigs. To date, limited epidemiological data was available for H. parasuis from healthy pigs, which might be carriers of potential pathogenic strains. In this study, from September 2016 to October 2017, we investigated the prevalence and characteristics of H. parasuis from healthy pigs in China. Totally, we obtained 244 isolates from 1675 nasal samples from 6 provinces. H. parasuis isolation was more successful in weaner pigs (22.6%, 192/849), followed by finisher pigs (9.3%, 43/463), and sows (2.5%, 9/363). The most prevalent serovars were 7 (20.1%, 49/244), followed by 3 (14.8%, 36/244), 2 (14.3%, 35/244), 11 (12.7%, 31/244), 5/12 (5.7%, 14/244) and 4 (2.5%, 6/244). Bimodal or multimodal distributions of MICs were observed for most of the tested drugs, which suggested the presence of non-wild type populations. It was noted that the MIC90 values of tilmicosin (64 µg/ml) was relatively higher than that reported in previous studies. Our results suggest that: 1) potentially pathogenic serovars of H. parasuis are identified in healthy pigs, and 2) elevated MICs and presence of mechanisms of resistance not yet described for clinically important antimicrobial agents would increase the burden of disease caused by H. parasuis.


Assuntos
Antibacterianos/farmacologia , Infecções por Haemophilus/veterinária , Haemophilus parasuis/efeitos dos fármacos , Doenças dos Suínos/epidemiologia , Suínos/microbiologia , Matadouros , Animais , Portador Sadio/epidemiologia , Portador Sadio/microbiologia , Portador Sadio/veterinária , China/epidemiologia , Fazendas , Infecções por Haemophilus/epidemiologia , Haemophilus parasuis/isolamento & purificação , Testes de Sensibilidade Microbiana , Nariz/microbiologia , Reação em Cadeia da Polimerase , Prevalência , Sorogrupo , Doenças dos Suínos/microbiologia , Desmame
8.
J Vet Diagn Invest ; 31(3): 364-367, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30973083

RESUMO

Infectious coryza is a severe respiratory disease of chickens associated with large economic losses in affected commercial flocks. The fastidious causative pathogen, Avibacterium paragallinarum, is difficult to recover and identify, resulting in delayed diagnosis and enhanced spread of the agent. Small poultry flocks are increasingly common in rural and suburban environments. We assessed the frequency of A. paragallinarum using real-time PCR and clinical conditions present in samples from such flocks submitted to the California Animal Health and Food Safety Laboratory System (Davis, CA) in 2018. From the 294 samples collected for our study, 86 (30%) were PCR-positive for A. paragallinarum. Juvenile birds (≤1 y) were significantly more likely to be PCR-positive ( p = 0.017), and birds diagnosed with respiratory disease had lower Ct values ( p = 0.001) than those without. Concurrent infections were also identified, including with Mycoplasma gallisepticum (18.6%), M. synoviae (18.6%), infectious bronchitis virus (12.8%), and infectious laryngotracheitis virus (7.0%). Only 46.5% of PCR-positive chickens had antemortem respiratory signs, making endemic infections in these flocks highly likely. Our study demonstrates that A. paragallinarum is present in small-flock operations including those without respiratory disease and may present a risk for airborne pathogen transmission to commercial poultry operations.


Assuntos
Galinhas , Monitoramento Epidemiológico/veterinária , Infecções por Haemophilus/veterinária , Haemophilus paragallinarum/isolamento & purificação , Doenças das Aves Domésticas/epidemiologia , Criação de Animais Domésticos/métodos , Animais , California/epidemiologia , Comorbidade , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/veterinária , Infecções por Coronavirus/virologia , Infecções por Haemophilus/epidemiologia , Infecções por Haemophilus/microbiologia , Infecções por Herpesviridae/epidemiologia , Infecções por Herpesviridae/veterinária , Infecções por Herpesviridae/virologia , Herpesvirus Galináceo 1/isolamento & purificação , Sequenciamento de Nucleotídeos em Larga Escala/veterinária , Vírus da Bronquite Infecciosa/isolamento & purificação , Mycoplasma/isolamento & purificação , Infecções por Mycoplasma/epidemiologia , Infecções por Mycoplasma/microbiologia , Infecções por Mycoplasma/veterinária , Pasteurellaceae/isolamento & purificação , Infecções por Pasteurellaceae/epidemiologia , Infecções por Pasteurellaceae/microbiologia , Infecções por Pasteurellaceae/veterinária , Doenças das Aves Domésticas/microbiologia , Reação em Cadeia da Polimerase em Tempo Real/veterinária
9.
Vet Microbiol ; 230: 117-122, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30827376

RESUMO

Haemophilus parasuis is a commensal Gram-negative bacterial pathogen in the upper respiratory tract of pigs, which causes Glässer's disease. More than 15 serotypes of H. parasuis have been identified with apparent differences in virulence. In this research, we surveyed the prevalence and distribution of serotypes and known virulence genes of the H. parasuis isolates collected from sick and healthy pigs in Quang Binh and Thua Thien Hue provinces in Central Vietnam. By using bacterial isolation and polymerase chain reaction (PCR), 56 out of 814 (6.9%) samples were positive for H. parasuis. The most prevalent serotypes were serotype 5 (15/56, 26.8%), followed by serotype 2 (13/56, 23.2%) and serotype 4 (10/56, 17.9%). The vta1 was the most frequently detected virulence gene which was present in 62.5% of the strains, followed by vta3 (42.9%), vta2 (39.3%), HPM-1371 (35.7%), capD (30.4%), HPM-1372 (12.5%), lsgB and HPM-1373 (both shared 8.9%). Strong correlations between some serotypes and known virulence genes were observed, in which virulence genes HPM-1371, HPM-1372, vta3, vta2 and capD were mainly clustered in serotypes 5/12, and vta2 clustered in serotype 2. This study presents the first baseline information on the epidemiological characteristics of H. parasuis isolates from Central Vietnam.


Assuntos
Infecções por Haemophilus/veterinária , Haemophilus parasuis/genética , Haemophilus parasuis/patogenicidade , Fatores de Virulência/genética , Matadouros , Animais , Fazendas , Haemophilus parasuis/isolamento & purificação , Gado/virologia , Reação em Cadeia da Polimerase , Sorogrupo , Suínos/virologia , Doenças dos Suínos/microbiologia , Vietnã , Virulência/genética
10.
BMC Vet Res ; 15(1): 98, 2019 Mar 25.
Artigo em Inglês | MEDLINE | ID: mdl-30909903

RESUMO

BACKGROUND: Haemophilus parasuis (HPS) is the causative agent of Glässer's disease, characterized by arthritis, fibrinous polyserositis and meningitis, and resulting in worldwide economic losses in the swine industry. Baicalin (BA), a commonly used traditional Chinese medication, has been shown to possess a series of activities, such as anti-bacterial, anti-viral, anti-tumor, anti-oxidant and anti-inflammatory activities. However, whether BA has anti-apoptotic effects following HPS infection is unclear. Here, we investigated the anti-apoptotic effects and mechanisms of BA in HPS-induced apoptosis via the protein kinase C (PKC)-mitogen-activated protein kinase (MAPK) pathway in piglet's mononuclear phagocytes (PMNP). RESULTS: Our data demonstrated that HPS could induce reactive oxygen species (ROS) production, arrest the cell cycle and promote apoptosis via the PKC-MAPK signaling pathway in PMNP. Moreover, when BA was administered, we observed a reduction in ROS production, suppression of cleavage of caspase-3 in inducing apoptosis, and inhibition of activation of the PKC-MAPK signaling pathway for down-regulating p-JNK, p-p38, p-ERK, p-PKC-α and PKC-δ in PMNP triggered by HPS. CONCLUSIONS: Our data strongly suggest that BA can reverse the apoptosis initiated by HPS through regulating the PKC-MAPK signaling pathway, which represents a promising therapeutic agent in the treatment of HPS infection.


Assuntos
Antibacterianos/uso terapêutico , Flavonoides/uso terapêutico , Haemophilus parasuis/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Monócitos/metabolismo , Doenças dos Suínos/tratamento farmacológico , Animais , Animais Recém-Nascidos/metabolismo , Animais Recém-Nascidos/microbiologia , Antibacterianos/farmacologia , Apoptose/efeitos dos fármacos , Relação Dose-Resposta a Droga , Flavonoides/farmacologia , Infecções por Haemophilus/tratamento farmacológico , Infecções por Haemophilus/metabolismo , Infecções por Haemophilus/veterinária , Monócitos/efeitos dos fármacos , Monócitos/microbiologia , Espécies Reativas de Oxigênio/metabolismo , Suínos , Doenças dos Suínos/metabolismo , Doenças dos Suínos/microbiologia
11.
Artigo em Inglês | MEDLINE | ID: mdl-30476621

RESUMO

INTRODUCTION: Haemophilus parasuis, one of the major swine pathogens, has at least fifteen different types, all of which have significant economic effects on the global swine industry. The aim of this study was to establish an experimental intraperitoneal infection model for H. parasuis in neutropenic guinea pigs. METHODS: Intraperitoneal administration of cyclophosphamide and Haemophilus parasuis was conducted in guinea pigs. Clinical signs, gross pathology, and histopathology were observed in neutropenic guinea pigs infected with H. parasuis. RESULTS: Intraperitoneal administration of 100 mg/kg cyclophosphamide led to immunosuppression with white blood cells, lymphocytes, and neutrophils all <1000 mm3, while no histological tissue damage was observed. Intraperitoneal administration of 109 colony-forming units (CFU) of H. parasuis led to typical respiratory symptoms, 90% morbidity, and 20% mortality in a 72 h-period. Bacteriological screening revealed that multiple organs, including the heart, liver, spleen, lungs, kidneys, and blood, were infected with H. parasuis. The threshold loads of bacteria in blood and the lungs were (7.04 ±â€¯0.53)log10 CFU/mL and (6.24 ±â€¯0.62)log10 CFU/g, respectively, at 3 d after infection. Gross pathology examination showed celiac effusion, intestinal mucosal hemorrhage, and liver, spleen, or lung swelling, necrosis, and hemorrhage. Congestion, mild interstitial pneumonia, inflammatory exudation, and endothelial cell proliferation were observed in the histological examination. DISCUSSION: All the results suggest that we have established an experimental intraperitoneal infection model for H. parasuis in neutropenic guinea pigs. It is especially useful as a tool for pharmacokinetics, pharmacodynamics, or a pharmacokinetics/pharmacodynamics (PK/PD) model of antimicrobial agents against respiratory disease.


Assuntos
Modelos Animais de Doenças , Infecções por Haemophilus/imunologia , Haemophilus parasuis/imunologia , Imunossupressores/administração & dosagem , Modelos Animais , Neutropenia/induzido quimicamente , Animais , Ciclofosfamida/administração & dosagem , Feminino , Cobaias , Infecções por Haemophilus/microbiologia , Infecções por Haemophilus/veterinária , Haemophilus parasuis/patogenicidade , Injeções Intraperitoneais , Pulmão , Masculino , Camundongos , Neutropenia/imunologia , Suínos
12.
BMC Vet Res ; 14(1): 411, 2018 Dec 19.
Artigo em Inglês | MEDLINE | ID: mdl-30567563

RESUMO

BACKGROUND: Infectious coryza (IC) is an acute respiratory disease of growing chickens and layers caused by Avibacterium paragallinarum. The development of tools that allow rapid pathogen detection is necessary in order to avoid disease dissemination and economic losses in poultry. An Av. paragallinarum-specific Ma-4 epitope of the TonB-dependent transporter (TBDT) was selected using bioinformatic tools in order to immunize a BalbC mouse and to produce monoclonal antibodies to be used in a lateral flow test (LFT) developed for Av. paragallinarum detection in chicken nasal mucus samples. RESULTS: The 1G7G8 monoclonal antibody was able to detect TBDT in Av. paragallinarum cultures (serogroups: A, B and C) by Western blot and indirect ELISA assay. Consequently, we developed a self-pairing prototype LFT. The limit of detection of the prototype LFT using Av. paragallinarum cultures was 1 × 104 colony-forming units (CFU)/mL. Thirty-five nasal mucus samples from chickens suspected of having infectious coryza were evaluated for the LFT detection capacity and compared with bacterial isolation (B.I) and polymerase chain reaction (PCR). Comparative indicators such as sensitivity (Se), specificity (Sp), positive predictive value (PPV), negative predictive values (NPV) and the kappa index (K) were obtained. The values were 100.0% Se, 50% Sp, 65.4% PPV, 100% NPV, and 0.49 K and 83.9% Se, 100% Sp, 100% PPV, 44.4% NPV, and 0.54 K for the comparison of the LFT with B.I and PCR, respectively. Additionally, the LFT allowed the detection of Av. paragallinarum from coinfection cases of Av. paragallinarum with Gallibacterium anatis. CONCLUSIONS: The results indicate that the self-pairing prototype LFT is suitable for the detection of TBDT in Av. paragallinarum cultures as well as in field samples such as nasal mucus from Av. paragallinarum-infected chickens. Therefore, this prototype LFT could be considered a rapid and promising tool to be used in farm conditions for Av. paragallinarum diagnosis.


Assuntos
Infecções por Haemophilus/veterinária , Haemophilus paragallinarum , Doenças das Aves Domésticas/diagnóstico , Animais , Anticorpos Monoclonais/metabolismo , Galinhas/microbiologia , Infecções por Haemophilus/diagnóstico , Haemophilus paragallinarum/fisiologia , Limite de Detecção , Camundongos , Camundongos Endogâmicos BALB C , Sensibilidade e Especificidade
13.
BMC Vet Res ; 14(1): 326, 2018 Nov 06.
Artigo em Inglês | MEDLINE | ID: mdl-30400794

RESUMO

BACKGROUND: Haemophilus (Glässerella) parasuis is the etiological agent of Glässer's disease in pigs. Control of this disorder has been traditionally based on bacterins. The search for alternative vaccines has focused mainly on the study of outer membrane proteins. This study investigates the transcriptome of H. (G.) parasuis serovar 5 subjected to in vitro conditions mimicking to those existing during an infection (high temperature and iron-restriction), with the aim of detecting the overexpression of genes coding proteins exposed on bacterial surface, which could represent good targets as vaccine candidates. RESULTS: The transcriptomic approach identified 13 upregulated genes coding surface proteins: TbpA, TbpB, HxuA, HxuB, HxuC, FhuA, FimD, TolC, an autotransporter, a protein with immunoglobulin folding domains, another large protein with a tetratricopeptide repeat and two small proteins that did not contain any known domains. Of these, the first six genes coded proteins being related to iron extraction. CONCLUSION: Six of the proteins have already been tested as vaccine antigens in murine and/or porcine infection models and showed protection against H. (G.) parasuis. However, the remaining seven have not yet been tested and, consequently, they could become useful as putative antigens in the prevention of Glässer's disease. Anyway, the expression of this seven novel vaccine candidates should be shown in other serovars different from serovar 5.


Assuntos
Antígenos de Bactérias/imunologia , Infecções por Haemophilus/veterinária , Vacinas Anti-Haemophilus/imunologia , Haemophilus parasuis/genética , Doenças dos Suínos/microbiologia , Animais , Perfilação da Expressão Gênica/veterinária , Infecções por Haemophilus/microbiologia , Infecções por Haemophilus/prevenção & controle , Haemophilus parasuis/imunologia , Haemophilus parasuis/metabolismo , Análise de Sequência de RNA , Suínos , Doenças dos Suínos/imunologia , Doenças dos Suínos/prevenção & controle , Transcriptoma/genética
14.
PLoS One ; 13(11): e0205700, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30383795

RESUMO

Haemophilus parasuis is a respiratory pathogen of swine and the etiological agent of Glässer's disease. H. parasuis isolates can exhibit different virulence capabilities ranging from lethal systemic disease to subclinical carriage. To identify genomic differences between phenotypically distinct strains, we obtained the closed whole-genome sequence annotation and genome-wide methylation patterns for the highly virulent Nagasaki strain and for the non-virulent D74 strain. Evaluation of the virulence-associated genes contained within the genomes of D74 and Nagasaki led to the discovery of a large number of toxin-antitoxin (TA) systems within both genomes. Five predicted hemolysins were identified as unique to Nagasaki and seven putative contact-dependent growth inhibition toxin proteins were identified only in strain D74. Assessment of all potential vtaA genes revealed thirteen present in the Nagasaki genome and three in the D74 genome. Subsequent evaluation of the predicted protein structure revealed that none of the D74 VtaA proteins contain a collagen triple helix repeat domain. Additionally, the predicted protein sequence for two D74 VtaA proteins is substantially longer than any predicted Nagasaki VtaA proteins. Fifteen methylation sequence motifs were identified in D74 and fourteen methylation sequence motifs were identified in Nagasaki using SMRT sequencing analysis. Only one of the methylation sequence motifs was observed in both strains indicative of the diversity between D74 and Nagasaki. Subsequent analysis also revealed diversity in the restriction-modification systems harbored by D74 and Nagasaki. The collective information reported in this study will aid in the development of vaccines and intervention strategies to decrease the prevalence and disease burden caused by H. parasuis.


Assuntos
Infecções por Haemophilus/genética , Haemophilus parasuis/genética , Doenças dos Suínos/genética , Suínos/microbiologia , Sequência de Aminoácidos , Animais , Genoma/genética , Genômica , Infecções por Haemophilus/microbiologia , Infecções por Haemophilus/veterinária , Haemophilus parasuis/patogenicidade , Suínos/genética , Doenças dos Suínos/microbiologia , Virulência/genética , Fatores de Virulência/genética
15.
Vet Microbiol ; 225: 64-71, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30322536

RESUMO

Porcine reproductive and respiratory syndrome virus (PRRSV) predisposes pigs to secondary bacterial infection caused by Haemophilus parasuis. The aim of the present study was to analyse the immune response of monocyte-derived macrophages (MDMs), serving as a model of macrophages accumulating at the site of inflammation. The second part of the study was focused on the role of IFNα in the production of inflammatory cytokines in co-infected MDMs. Concurrent infection with PRRSV and H. parasuis decreased gene expression of pro-inflammatory cytokines (IL-1ß, IL-8) in MDMs in comparison with MDMs infected with PRRSV or H. parasuis alone. Our data showed that MDMs express IFNα after PRRSV infection. Thereafter, we exposed cells to the experimental addition of IFNα and a subsequent infection with H. parasuis, and detected a decreased expression/production of pro-inflammatory cytokines. Thus, we assume that IFNα, produced after PRRSV infection, could affect the immune response of monocyte-derived macrophages. Down-regulation of pro-inflammatory cytokine expression in inflammatory macrophages may allow the development of secondary bacterial infections in pigs.


Assuntos
Citocinas/imunologia , Haemophilus parasuis/imunologia , Interferon-alfa/imunologia , Macrófagos/imunologia , Vírus da Síndrome Respiratória e Reprodutiva Suína/imunologia , Animais , Lavagem Broncoalveolar , Sobrevivência Celular , Citocinas/genética , Infecções por Haemophilus/microbiologia , Infecções por Haemophilus/veterinária , Haemophilus parasuis/patogenicidade , Interferon-alfa/farmacologia , Macrófagos/efeitos dos fármacos , Macrófagos/microbiologia , Macrófagos/virologia , Síndrome Respiratória e Reprodutiva Suína/virologia , Vírus da Síndrome Respiratória e Reprodutiva Suína/patogenicidade , Suínos , Doenças dos Suínos/microbiologia , Doenças dos Suínos/virologia
16.
Res Vet Sci ; 119: 292-301, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30081339

RESUMO

Three recombinant outer membrane proteins (rOmps) from the Haemophilus parasuis Nagasaki strain (serovar 5 reference strain), rOmpP2, rOmpP5 and rOmpD15, which have previously shown protection against H. parasuis infection in mice, were cloned, expressed and evaluated as vaccine antigens in colostrum-deprived pigs. When these animals were immunized with these rOmps and were later challenged intratracheally with 108 CFUs of the Nagasaki strain, no protection was seen in terms of survival, clinical signs, pathological results and recovery of H. parasuis. We hypothesized that a possible explanation for this lack of protection could be the low number of epitopes accessible to the immune system as a consequence of their poor exposure on the bacterial surface so that the immune response would not be able to protect against experimental infection by H. parasuis when a fully susceptible animal model, such as pigs, was used.


Assuntos
Infecções por Haemophilus/veterinária , Vacinas Anti-Haemophilus/imunologia , Haemophilus parasuis/imunologia , Doenças dos Suínos/imunologia , Animais , Anticorpos Antibacterianos , Colostro , Feminino , Infecções por Haemophilus/imunologia , Infecções por Haemophilus/prevenção & controle , Camundongos , Gravidez , Suínos , Doenças dos Suínos/prevenção & controle
17.
J Vet Med Sci ; 80(7): 1047-1053, 2018 Jul 12.
Artigo em Inglês | MEDLINE | ID: mdl-29798967

RESUMO

The bacterium Haemophilus parasuis (H. parasuis) is the primary cause of Glässer's disease. Currently, there are no effective vaccines that can confer protection against all H. parasuis serovars. Therefore, the present study aimed to investigate the effect of tea polyphenols on growth, expression of virulence-related factors, and biofilm formation of H. parasuis, as well as to evaluate their protective effects against H. parasuis challenge. Our findings demonstrated that tea polyphenols can inhibit H. parasuis growth in a dose-dependent manner and attenuate the biofilm formation of H. parasuis. In addition, tea polyphenols exerted inhibitory effects on the expression of H. parasuis virulence-related factors. Moreover, tea polyphenols could confer protection against a lethal dose of H. parasuis and can reduce pathological tissue damage induced by H. parasuis. In summary, our findings demonstrated the promising use of tea polyphenols as a novel treatment for H. parasuis infection in pigs.


Assuntos
Infecções por Haemophilus/veterinária , Haemophilus parasuis/crescimento & desenvolvimento , Haemophilus parasuis/patogenicidade , Polifenóis/farmacologia , Doenças dos Suínos/tratamento farmacológico , Chá/química , Animais , Infecções por Haemophilus/tratamento farmacológico , Suínos , Virulência , Fatores de Virulência
18.
Int J Mol Sci ; 19(5)2018 Apr 27.
Artigo em Inglês | MEDLINE | ID: mdl-29702580

RESUMO

Haemophilus parasuis (H. parasuis) can cause Glässer’s disease in pigs. However, the molecular mechanism of the inflammation response induced by H. parasuis remains unclear. The high-mobility group box 1 (HMGB1) protein is related to the pathogenesis of various infectious pathogens, but little is known about whether H. parasuis can induce the release of HMGB1 in piglet peripheral blood monocytes. Baicalin displays important anti-inflammatory and anti-microbial activities. In the present study, we investigated whether H. parasuis can trigger the secretion of HMGB1 in piglet peripheral blood monocytes and the anti-inflammatory effect of baicalin on the production of HMGB1 in peripheral blood monocytes induced by H. parasuis during the inflammation response. In addition, host cell responses stimulated by H. parasuis were determined with RNA-Seq. The RNA-Seq results showed that H. parasuis infection provokes the expression of cytokines and the activation of numerous pathways. In addition, baicalin significantly reduced the release of HMGB1 in peripheral blood monocytes induced by H. parasuis. Taken together, our study showed that H. parasuis can induce the release of HMGB1 and baicalin can inhibit HMGB1 secretion in an H. parasuis-induced peripheral blood monocytes model, which may provide a new strategy for preventing the inflammatory disorders induced by H. parasuis.


Assuntos
Anti-Infecciosos/farmacologia , Anti-Inflamatórios/farmacologia , Flavonoides/farmacologia , Proteína HMGB1/metabolismo , Doenças dos Suínos/tratamento farmacológico , Animais , Anti-Infecciosos/uso terapêutico , Anti-Inflamatórios/uso terapêutico , Sequência de Bases , Citocinas/genética , Citocinas/metabolismo , Relação Dose-Resposta a Droga , Flavonoides/uso terapêutico , Proteína HMGB1/genética , Infecções por Haemophilus/tratamento farmacológico , Infecções por Haemophilus/veterinária , Haemophilus parasuis/patogenicidade , Inflamação/tratamento farmacológico , Inflamação/veterinária , Monócitos/efeitos dos fármacos , Cultura Primária de Células , Suínos
19.
Vet Immunol Immunopathol ; 199: 77-80, 2018 May.
Artigo em Inglês | MEDLINE | ID: mdl-29678233

RESUMO

Infectious coryza is an important respiratory disease of chickens around the world and is caused by Avibacterium paragallinarum. Among the three Page serovars currently recognized for this bacterium, serovar B is a major circulating serovar in China nowadays. The cross-protection ability of the Page serovar B reference strain (0222) and five local isolates was evaluated by a vaccination-challenge trial in SPF chickens. The clinical signs seen in control birds challenged by strain 0222 and isolate HB 01 were significantly different, with isolate HB 01 giving more severe clinical signs. In terms of cross-protection, the protection in the groups vaccinated with isolate HB 01 and BJ 02 was significantly higher than that in the groups vaccinated with 0222 and the other three isolates. In addition, an experimental oil adjuvant trivalent vaccine, containing field isolate HB 01 antigen, was compared for immune efficacy with two commercial trivalent infectious coryza vaccines containing internationally recognized serovar B strains. The experimental oil adjuvant trivalent vaccine elicited best protection (80%) among the three trivalent vaccines. In conclusion, the oil adjuvant vaccine, containing field isolate HB 01 may be a better choice in control of current serovar B Av. paragallinarum outbreaks in China under current circumstances.


Assuntos
Infecções por Haemophilus/veterinária , Vacinas Anti-Haemophilus/imunologia , Haemophilus paragallinarum/imunologia , Doenças das Aves Domésticas/prevenção & controle , Animais , Galinhas , Proteção Cruzada/imunologia , Infecções por Haemophilus/imunologia , Infecções por Haemophilus/microbiologia , Infecções por Haemophilus/prevenção & controle , Vacinas Anti-Haemophilus/farmacologia , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/microbiologia , Sorogrupo , Vacinas de Produtos Inativados
20.
J Vet Pharmacol Ther ; 41(5): 706-718, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-29696661

RESUMO

The aims of this study were to establish optimal doses of doxycycline (dox) against Haemophilus parasuis on the basis of pharmacokinetic-pharmacodynamic (PK-PD) integration modeling. The infected model was established by intranasal inoculation of organism in pigs and confirmed by clinical signs, blood biochemistry, and microscopic examinations. The recommended dose (20 mg/kg b.w.) was administered in pigs through intramuscular routes for PK studies. The area under the concentration 0- to 24-hr curve (AUC0-24 ), elimination half-life (T½ke ), and mean residence time (MRT) of dox in healthy and H. parasuis-infected pigs were 55.51 ± 5.72 versus 57.10 ± 4.89 µg·hr/ml, 8.28 ± 0.91 versus 9.80 ± 2.38 hr, and 8.43 ± 0.27 versus 8.79 ± 0.18 hr, respectively. The minimal inhibitory concentration (MIC) of dox against 40 H. parasuis isolates was conducted through broth microdilution method, the corresponding MIC50 and MIC90 were 0.25 and 1 µg/ml, respectively. The Ex vivo growth inhibition data suggested that dox exhibited a concentration-dependent killing mechanism. Based on the observed AUC24 hr /MIC values by modeling PK-PD data in H. parasuis-infected pigs, the doses predicted to obtain bacteriostatic, bactericidal, and elimination effects for H. parasuis over 24 hr were 5.25, 8.55, and 10.37 mg/kg for the 50% target attainment rate (TAR), and 7.26, 13.82, and 18.17 mg/kg for 90% TAR, respectively. This study provided a more optimized alternative for clinical use and demonstrated that the dosage 20 mg/kg of dox by intramuscular administration could have an effective bactericidal activity against H. parasuis.


Assuntos
Antibacterianos/farmacocinética , Doxiciclina/farmacocinética , Infecções por Haemophilus/veterinária , Haemophilus parasuis/efeitos dos fármacos , Doenças dos Suínos/tratamento farmacológico , Animais , Antibacterianos/administração & dosagem , Antibacterianos/farmacologia , Relação Dose-Resposta a Droga , Doxiciclina/administração & dosagem , Doxiciclina/farmacologia , Feminino , Infecções por Haemophilus/tratamento farmacológico , Injeções Intramusculares/veterinária , Masculino , Testes de Sensibilidade Microbiana/veterinária , Suínos , Doenças dos Suínos/microbiologia
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