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1.
Haemophilus parasuis is the aetiological agent of Glässer’s disease and is also a commensal of the upper respiratory tract of pigs. Trimeric autotransporter (vtaA) genes have been identified in H. parasuis and divided into three groups on the basis of the translocator domain sequence. In this study, group 3 vtaA genes were demonstrated by PCR in all 157 H. parasuis isolates tested. Group 1 vtaA genes were associated with virulent strains; 52/54 (96%) group 1 vtaA negative field isolates were isolated from the nasal passages of healthy animals, whereas no group 1 vtaA negative field isolates were isolated from cases of Glässer’s disease. There was an association between absence of group 1 vtaA, sensitivity to phagocytosis and serum and classification of isolates into nasal cluster C by multilocus sequence typing. A multiplex PCR was developed for diagnosis of H. parasuis at the species level (group 3 vtaA positive) and to differentiate putative non-virulent strains (group 1 vtaA negative). When applied to field samples, the PCR confirmed a high prevalence of H. parasuis in conventionally farmed pigs and demonstrated that almost half of the animals carried potentially virulent strains.  相似文献   

2.
The present study investigated the effects of vaccinating sows and piglets or piglets alone against Haemophilus parasuis on the prevalence of H. parasuis in nasal swabs, on the humoral and cellular immune responses, and on the production parameters of piglets at 3 Korean farms with a clinical history of polyserositis caused by H. parasuis. Piglets born to vaccinated or non-vaccinated sows were subdivided into 3 groups: vaccinated sows and vaccinated pigs (VS-VP), non-vaccinated sows and vaccinated pigs (NVS-VP), and non-vaccinated sows and non-vaccinated pigs (NVS-NVP). The proportion of piglets with positive nasal swabs was significantly lower (P < 0.05) in the vaccinated animals (VS-VP and NVS-VP groups) than in the non-vaccinated animals (NVS-NVP group) at 35 and 60 d of age at the 3 farms. The overall growth performance (from 7 to 60 d of age) of the vaccinated piglets was significantly better (P < 0.05) than that of the non-vaccinated piglets at the 3 farms. Piglets in the VS-VP group had significantly higher levels (P < 0.05) of H. parasuis-specific IgG antibodies, lymphocyte proliferation, and interferon-γ-secreting cells than piglets in the NVS-VP and NVS-NVP groups on days 1, 7, 21, 35, and 60 after birth at the 3 farms.  相似文献   

3.
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4.
A whole cell formalin killed trivalent Haemophilus parasuis bacterin was tested for efficacy in four week old, weaned specific pathogen free pigs challenged under laboratory conditions. The vaccine contained three field strains of H. parasuis selected from confirmed cases of Glasser's disease. Two different formulations were evaluated in separate trials. In trial 1, ten pigs received 5 mL of bacterin subcutaneously in the neck, followed by a second 5 mL dose two weeks later. Another ten pigs served as nonvaccinated controls. One week after the second dose, all pigs were subjected to an aerosol challenge containing the strains of H. parasuis present in the vaccine. In trial 2, a broth rather than a saline based vaccine was prepared, and tested as in trial 1. In both trials, the vaccinated pigs remained healthy postchallenge, while eight of nine (Trial 1) and eight of ten (Trial 2) nonvaccinated pigs succumbed to Glasser's disease.  相似文献   

5.
Haemophilus parasuis is a colonizer of the upper respiratory tract of healthy pigs, but virulent strains can cause a systemic infection characterized by fibrinous polyserositis, commonly known as Glässer’s disease. The variability in virulence that is observed among H. parasuis strains is not completely understood, since the virulence mechanisms of H. parasuis are largely unknown. In the course of infection, H. parasuis has to survive the host pulmonary defences, which include alveolar macrophages, to produce disease. Using strains from different clinical backgrounds, we were able to detect clear differences in susceptibility to phagocytosis. Strains isolated from the nose of healthy animals were efficiently phagocytosed by porcine alveolar macrophages (PAM), while strains isolated from systemic lesions were resistant to this interaction. Phagocytosis of susceptible strains proceeded through mechanisms independent of a specific receptor, which involved actin filaments and microtubules. In all the systemic strains tested in this study, we observed a distinct capsule after interaction with PAM, indicating a role of this surface structure in phagocytosis resistance. However, additional mechanisms of resistance to phagocytosis should be explored, since we detected different effects of microtubule inhibition among systemic strains.  相似文献   

6.
Three trials were conducted to establish if young primary specific pathogen free (SPF) pigs could be protected from Glasser's disease by vaccination. Three age groups of cesarean-derived isolator-reared gnotobiotic pigs were vaccinated twice at 4 and 6, 3 and 5, and 2 and 4 wk of age respectively with a formalin killed aluminum hydroxide adsorbed bacterin prepared from three strains of Haemophilus parasuis isolated from Ontario pigs affected with Glasser's disease. When challenged two weeks later with the homologous strains of virulent bacteria, all the vaccinated pigs remained healthy, while 17/18 nonvaccinated pigs became severely sick or died between three and seven days postchallenge. The one surviving nonimmunized pig was retarded in growth. All of the nonimmunized pigs had visible lesions of polyserositis, the most common being polyarthritis (14/18). Other lesions were fibrinous meningitis, pericarditis, pleurisy and/or peritonitis. Two of the pigs died with a septicemia. Haemophilus parasuis was isolated from 15/18 nonimmunized pigs, usually from several of the affected sites. The organisms were not isolated from the immunized pigs, nor from the surviving nonimmunized pig. Attempts to detect the presence of specific antibodies against the H. parasuis strains in the sera of the immunized or exposed pigs by the passive hemagglutination test or by enzyme linked immunoassay were unsuccessful. The results of this work indicate that primary SPF pigs can be protected from Glasser's disease by vaccination as early as 2 and 4 wk of age. The nature of this protective mechanism was not established in this study.  相似文献   

7.
The purpose of this study was to determine the effect of enrofloxacin in the carrier stage of Haemophilus parasuis in naturally colonized weaned pigs. Twenty-three pigs colonized by H. parasuis received either 7.5 mg/kg body weight (BW) of enrofloxacin or a saline solution intramuscularly at weaning. Nasal and tonsillar swab samples were collected daily throughout the study and at necropsy and tested by quantitative polymerase chain reaction (qPCR). The H. parasuis isolates obtained from samples collected at necropsy were subjected to genotyping by enterobacterial repetitive intergenic consensus polymerase chain reaction (ERIC-PCR) and a multiplex PCR for the detection of the virulence-associated trimeric autotransporter (vtaA) genes. Haemophilus parasuis was detected in the nasal cavity and tonsils of pigs in the control group throughout the study. Antibiotic-treated pigs tested negative for H. parasuis at 1 d post-treatment and the proportion of nasal samples that tested positive was higher for control pigs than for treated pigs at 1, 2, 3, 4, 5, 6, and 7 d post-treatment and at 2, 4, and 5 d post-treatment for tonsil samples (P < 0.003). Genotyping by ERIC-PCR demonstrated that pigs were colonized with a common H. parasuis strain at the end of the study. Isolates were negative for the vtaA gene, which indicates the absence of vtaA virulence factor. In conclusion, enrofloxacin significantly reduced the H. parasuis load in naturally colonized pigs, but was unable to completely eliminate the organism.  相似文献   

8.
Objective Investigate the diversity of serovars of Haemophilus parasuis (Hps) present in Australian pig herds. Design Nasal swabs were used to obtain multiple isolates of Hps, which were grouped first by genotyping and then by serotyping representative isolates. Procedure Swabs were taken from the nasal cavity of just-weaned healthy pigs from multiparous sows on 12 farms and from post-weaned pigs of multiparous sows on 1 farm. On 5 of the 13 farms, nasal swabs were also obtained from pigs showing clinical signs suggestive of Glässer's disease. On a further 7 farms, nasal swabs were obtained only from pigs with clinical signs suggestive of Glässer's disease. Results A total of 556 Hps isolates were genotyped, and 150 isolates were serotyped. Hps was detected on 19 of the 20 farms, including 2 farms with a long history of freedom from Glässer's disease. Isolates of Hps belonging to serovars regarded as potentially pathogenic were obtained from healthy pigs at weaning on 8 of the 10 farms with a history of Glässer's disease outbreaks. Sampling 213 sick pigs yielded 115 isolates of Hps, 99 of which belonged to serovars that were either potentially pathogenic or of unknown pathogenicity. Only 16 isolates from these 213 sick pigs were of a serovar known to be non-pathogenic. Conclusion Healthy pigs contain a range of Hps serovars, even on farms free of Glässer's disease. Nasal swabbing of both healthy and sick pigs seems a useful method of serovar profiling of farms.  相似文献   

9.
From September 2008 to December 2010, 112 Haemophilus parasuis strains were isolated from 536 pigs with clinical signs of Glässer’s disease in South China, for a frequency of 21%. The 112 strains were subjected to serovar analysis by gel diffusion (GD) and indirect hemagglutination (IHA) tests and to genotype analysis by means of pulsed-field gel electrophoresis (PFGE). With a combination of the GD and IHA results, serovars 5 and 4 were found to be the most prevalent, at 23% and 17%, respectively, followed by serovars 2 (8%), 15 (7%), 13 (6%), and 12 (5%); 20% of the strains were nontypeable. The 112 strains were genetically diverse, with 85 genotypes identified (discriminatory index 0.992). The 89 typeable isolates belonged to 15 H. parasuis serovars displaying 63 different PFGE profiles. The 23 nontypeable strains displayed 22 different PFGE profiles. These findings confirmed that 15 serovars and diverse genotypes of H. parasuis were widely distributed in southern China.  相似文献   

10.
Haemophilus parasuis is a colonizer of the upper respiratory tract of pigs, although it is better known as the etiological agent of Gl?sser's disease. Interestingly, several strains can be isolated from a single farm, as determined by both genotyping and serotyping. However, it is not known how an outbreak and the subsequent treatment affect the population of H. parasuis strains. In this study, a farm was studied during an outbreak of Gl?sser's disease and 1 year after antimicrobial treatment and elimination of clinical signs. Bacterial isolation was attempted from nasal swabs and lesions. After isolation, antimicrobial susceptibility, serotype and genotype were determined. Two different genotyping techniques, enterobacterial repetitive intergenic consensus (ERIC)-PCR and multilocus sequence typing (MLST) were used. The H. parasuis strain that was isolated from lesions during the disease outbreak clustered with other virulent strains by both MLST and serotyping analysis. Nasal isolates were included in the corresponding nasal cluster by MLST, but they presented high variability by serotyping. These nasal isolates included serotypes previously classified as virulent and non-virulent. Finally, we found that during the antimicrobial treatment the diversity of strains isolated in the farm was affected and just one strain, which was resistant to the treatment, was detected. One year after the treatment strain diversity was back to normal (three strains).  相似文献   

11.
The aim of this study was to compare the development of Glasser's disease in sow-reared and colostrum-deprived piglets. Ninety piglets from a commercial pig farm in Spain were used. The farm was positive for Haemophilus parasuis. Fifty-two pigs were sow-reared (SR) and 38 were colostrum-deprived (CD) piglets. The animals were intratracheally inoculated with H. parasuis serovar 5 and sacrificed at 1, 2 and 3 days post-infection. To assess the development of disease, antibody titers, clinical signs, pathological lesions, microbiological isolation and PCR amplification were compared between the groups. Inoculation of SR pigs did not cause clinical signs or lesions of Glasser's disease. In SR pigs, H. parasuis isolation and specific PCR amplification from tissues showed a very low number of positive samples. In contrast, in CD pigs, inoculation resulted in the typical signs and lesions of Glasser's disease. Positive microbiological isolation and specific PCR products were obtained from the majority of the tissues tested, and no antibodies against H. parasuis were detected. The experimental infection using CD pigs describes a successful method to study this microorganism and confirms the important role that maternal antibodies play in protection against clinical signs and disease.  相似文献   

12.
The serum antibody response to an experimental infection by Haemophilus parasuis, the etiological agent of Glässer’s disease in pigs, was characterized by ELISA measuring IgM and IgGt levels against whole-cells and outer-membrane-proteins (OMPs) as antigens. Five groups of pigs were studied, four of those were previously immunized with different formulations, and the fifth was maintained as non-immunized control. All groups were challenged with 5 × 109 CFU of H. parasuis. The non-commercial bacterin induced a full protection against disease, the OMP-vaccine and the exposure to a sublethal dose of 105 CFU protected only partially, and the recombinant TbpB-vaccine conferred no protection. The humoral response in the pigs that died after infection (all controls, all those vaccinated with the recombinant TbpB, and two of both those inoculated with OMPs and those exposed to the sublethal dose) could be only measured before it, but it was irrelevant in all cases. However, a specific IgM and IgGt production was observed before challenge in all the surviving pigs, irrespective of the type of immunization received. This antibody response was even greater after H. parasuis infection, especially in those survivors receiving the sublethal dose. These results suggest a role of the antibodies developed after the different immunization protocols in preventing infection and death; therefore, the humoral immunity is protective against experimental Glässer’s disease.  相似文献   

13.
Comparison of sampling sites and detection methods for Haemophilus parasuis   总被引：3，自引：0，他引：3  
Objective To improve the isolation rate and identification procedures for Haemophilus parasuis from pig tissues. Design Thirteen sampling sites and up to three methods were used to confirm the presence of H. parasuis in pigs after experimental challenge. Procedure Colostrum‐deprived, naturally farrowed pigs were challenged intratracheally with H parasuis serovar 12 or 4. Samples taken during necropsy were either inoculated onto culture plates, processed directly for PCR or enriched prior to being processed for PCR. The recovery of H parasuis from different sampling sites and using different sampling methods was compared for each serovar. Results H parasuis was recovered from several sample sites for all serovar 12 challenged pigs, while the trachea was the only positive site for all pigs following serovar 4 challenge. The method of solid medium culture of swabs, and confirmation of the identity of cultured bacteria by PCR, resulted in 38% and 14% more positive results on a site basis for serovars 12 and 4, retrospectively, than direct PCR on the swabs. This difference was significant in the serovar 12 challenge. Conclusion Conventional culture proved to be more effective in detecting H parasuis than direct PCR or PCR on enrichment broths. For subacute (serovar 4) infections, the most successful sites for culture or direct PCR were pleural fluid, peritoneal fibrin and fluid, lung and pericardial fluid. For acute (serovar 12) infections, the best sites were lung, heart blood, affected joints and brain. The methodologies and key sampling sites identified in this study will enable improved isolation of H parasuis and aid the diagnosis of Glässer's disease.  相似文献   

14.
Biofilms are surface-associated microbial communities, which are encased in self-synthesized extracellular environment. Biofilm formation may trigger drug resistance and inflammation, resulting in persistent infections. Haemophilus parasuis is the etiological agent of a systemic disease, Glässer's disease, characterized by fibrinous polyserositis, arthritis and meningitis in pigs. The purpose of this study was to examine the correlation between biofilm and antibiotic resistance among the clinical isolates of H. parasuis. In the present study, we tested biofilm-forming ability of 110 H. parasuis isolates from various farms using polystyrene microtiter plate assays. Seventy-three isolates of H. parasuis (66.4%) showed biofilm formation and most of them performed weak biofilm-forming ability (38/73). All isolates were tested for antimicrobial susceptibility to 18 antimicrobial agents by the broth microdilution method. H. parasuis isolates showed very high resistance (>90%) to sulfanilamide, nalidixic acid, and trimethoprim. Resistance to eight antibiotics such as penicillin (41.1% vs 8.1%), ampicillin (31.5% vs 8.1%), amoxicillin (28.8% vs 5.4%), gentamicin (46.6% vs 24.3%), cefazolin (19.2% vs 2.7%), doxycycline (19.2% vs 8.1%), cefotaxime (11% vs 2.7%), and cefaclor (13.7% vs 5.4%) was comparatively higher among biofilm producers than non-biofilm producers. Pulsed-field gel electrophoresis (PFGE) analyses could distinguish various isolates. Our data indicated that H. parasuis field isolates were able to form biofilms in vitro. In addition, biofilm positive strains had positive correlation with resistance to β-lactams antibiotics. Thus, biofilm formation may play important roles during H. parasuis infections.  相似文献   

15.
The use of naturally-farrowed, artificially-reared piglets as an alternative model to study Haemophilus parasuis infections was evaluated. Two trials were performed in order to evaluate the proposed model. In trial 1, animals were vaccinated and challenged with H. parasuis. Results showed that the proposed model was effectively used to evaluate protective immunity against this organism. In trial 2, animals were challenged with different doses of H. parasuis. Results showed that the severity of clinical signs and lesions tended to increase with higher doses. The reproduction of clinical signs and lesions characteristic of H. parasuis systemic infection was successful in both trials, proving that this model is a viable alternative to specific-pathogen free and cesarean-derived, colostrum-deprived pigs.  相似文献   

16.
Cross-protection between Haemophilus parasuis serovars 2 and 5 was examined in pigs using a bacterin based vaccine, and subsequently the safety and efficacy of a bivalent vaccine were evaluated. Upon intratracheal challenge of a serovar 2 or 5 strain, pigs immunized with a monovalent vaccine were protected against challenge with a homologous serovar strain, but not with a heterologous serovar strain. Immunization with a bivalent vaccine containing both serovars 2 and 5 bacterins conferred protection in pigs against lethal challenge with each of the serovar strains. A total of 86 pigs from two SPF herds were injected with the bivalent vaccine intramuscularly twice at a four-week interval. No adverse reactions following the vaccination were observed. On day 7 after the second vaccination, vaccinated and non-vaccinated control pigs from herd A were transferred to herd B, where Glasser's disease had broken out. Pigs in the control group developed clinical signs of the disease, and 6 of 8 (75%) pigs died until slaughter, in contrast with only 4 of 46 (9%) pigs in the vaccinated group. In herd C, where there was no outbreak of Glasser's disease, complement fixation antibody titer was raised only in the vaccinated group. A challenge experiment on days 20 and 79 after the second vaccination showed that only the vaccinated pigs were protected. From these findings, the safety and efficacy of the bivalent vaccine were confirmed under laboratory and field conditions.  相似文献   

17.
An immunoproteomic analysis of the protective response of subunit and commercial vaccines in colostrum-deprived pigs against Glässer's disease was carried out. A mixture of proteins with affinity to porcine transferrin (PAPT) from Haemophilus parasuis Nagasaki strain (serovar 5) was inoculated intramuscularly (PAPTM) and intratracheally (PAPTCp), along with a commercial bacterin. PAPT were separated using 2 dimensional electrophoresis (2DE) gels and with them, 2DE Western blots were carried out. A total of 17 spots were identified as positive with sera of pigs from any of the three vaccinated groups, the highest number of immunoreactive proteins being detected in those having received PAPTCp. Among them, six proteins (FKBP-type peptidyl-prolyl cis-trans isomerase, neuraminidase exo-α-sialidase, xanthine-guanine phosphoribosyl transferase, CMP-N-acetylneuraminic acid synthetase, phenylalanyl-tRNA synthetase and glyceraldehyde 3-phosphate dehydrogenase) were found to be novel immunogens in H. parasuis. These proteins showed a high potential as candidates in future subunit vaccines against Glässer's disease. The three experimental groups developed specific systemic total IgG (IgGt), IgG1, IgG2 and IgM antibodies after immunizations. In addition, those receiving PAPTCp yielded a serum IgA response.  相似文献   

18.
The objective of this study was to determine the associations among Haemophilus parasuis, Mycoplasma hyorhinis, and porcine reproductive and respiratory syndrome (PRRS) virus (EU-field strain) infections in 95 pigs with polyserositis. A significant association between H. parasuis and M. hyorhinis was identified. H. parasuis and M. hyorhinis were significantly more often detected in PRRS virus positive pigs.  相似文献   

19.
The expression of several cytokines in spleen, pharyngeal lymph nodes, lung and brain after different immunization procedures and a challenge with 5 × 109 CFU of Haemophilus parasuis was compared. Five groups of colostrum-deprived pigs were used: vaccinated with (I) a bacterin, (II) an outer-membrane-protein-vaccine, (III) a recombinant transferring-binding protein B, (IV) exposed to a total dose of 105 CFU, and (V) not previously immunized. All pigs in groups III and V died, while all animals in group I, most of group IV and half of group II survived until the end of the experiment. IL-1α was found in significantly higher levels (p < 0.05) in spleen, lymph nodes and brain of dead pigs, which could be explained by the major severity of lesions in these animals. However, IL-4, IL-10, TNF-α and IFN-γ were expressed in significantly higher levels by survivors (for all the four cytokines in lymph nodes; for IL-4, IL-10 and TNF-α in spleen; for IL-4, TNF-α and IFN-γ in lung, and only for TNF-α in brain), thus suggesting a role of these four cytokines in the adaptive response, which might contribute to protection against H. parasuis infection.  相似文献   

20.
Haemophilus parasuis is a colonizer of the upper respiratory tract of pigs and the etiological agent of Gl?sser's disease, which is characterized by a fibrinous polyserositis, meningitis and arthritis. Gl?sser's disease has never been reported in wild boar (Sus scrofa), although antibodies against H. parasuis have been detected. The goal of this study was to confirm the presence of this bacterium in wild boar by bacterial isolation and to compare the strains to H. parasuis from domesticated pigs. Therefore, nasal swabs from 42 hunted wild boars were processed for bacterial isolation and subsequent H. parasuis identification by specific PCR, biochemical tests and 16S rRNA gene sequencing. Two different strains of H. parasuis from two wild boars were isolated. These strains belonged to serotype 2 and were included by 16S rRNA gene sequencing and MLST analysis in a cluster with other H. parasuis strains of nasal origin from domestic pigs. During this study, Actinobacillus minor and Actinobacillus indolicus, which are NAD-dependent Pasteurellaceae closely related to H. parasuis, were also isolated. Our results indicate similarities in the respiratory microbiota of wild boars and domestic pigs, and although H. parasuis was isolated from wild boars, more studies are needed to determine if this could be a source of H. parasuis infection for domestic pigs.  相似文献   

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