Immunogenic characterization of vaccines based on Haemophilus parasuis Nagasaki strain, OmpP2, OmpP5 and OmpD15, in colostrum-deprived pigs experimentally challenged with the same strain.

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.

Tea polyphenols suppress growth and virulence-related factors of Haemophilus parasuis.

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.

First comparison of adjuvant for trivalent inactivated Haemophilus parasuis serovars 4, 5 and 12 vaccines against Glässer's disease.

Haemophilus parasuis has had a huge impact in the swine industry throughout the world. Inactivated bacterium for H. parasuis is a traditional vaccine that can elicit efficient protection against homologous challenges. The objective of this study was to screen for the adjuvant-enhanced immune effect of trivalent inactivated H. parasuis serovars 4, 5 and 12 (prevalent serovars in China) vaccines against Glässer's disease. The adjuvants of mineral oil, aluminum hydroxide, Montanide GEL 01 PR, Montanide IMS 1313N VG and Montanide ISA 760 VG were used to make emulsified inactivated H. parasuis serovars 4, 5 and 12, respectively. Safety, antibody titer and protective efficacy of these vaccines were examined separately in piglets, and the feasibility of microagglutination test for detecting antibody titer of H. parasuis was confirmed for the first time. Due to easy of injection, high safety, rapidly immune responses, high concentrations of antibody, and 100% of protective efficacy for piglets, Montanide GEL 01 PR adjuvant can provide more homologous serovar protection than other domestically developed inactivated vaccines and should be used as a candidate adjuvant.

Haemophilus parasuis: infection, immunity and enrofloxacin.

Haemophilus parasuis is an early colonizer of the porcine upper respiratory tract and is the etiological agent of Glasser's disease. The factors responsible for H. parasuis colonization and systemic infection are not yet well understood, while prevention and control of Glasser's disease continues to be challenging. Recent studies on innate immunity to H. parasuis have demonstrated that porcine alveolar macrophages (PAMs) are able to differentially up-regulate several genes related to inflammation and phagocytosis, and several pro-inflammatory cytokines are produced by porcine cells upon exposure to H. parasuis. The susceptibility of H. parasuis strains to phagocytosis by PAMs and the bactericidal effect of complement are influenced by the virulent phenotype of the strains. While non-virulent strains are susceptible to phagocytosis and complement, virulent strains are resistant to both. However, in the presence of specific antibodies against H. parasuis, virulent strains become susceptible to phagocytosis. More information is still needed, though, in order to better understand the host immune responses to H. parasuis. Antimicrobials are commonly used in the swine industry to help treat and control Glasser's disease. Some of the common antimicrobials have been shown to reduce colonization by H. parasuis, which may have implications for disease dynamics, development of effective immune responses and immunomodulation. Here, we provide the current state of research on innate and adaptive immune responses to H. parasuis and discuss the potential effect of enrofloxacin on the development of a protective immune response against H. parasuis infection.

Pharmacokinetic/pharmacodynamic evaluation of marbofloxacin in the treatment of Haemophilus parasuis and Actinobacillus pleuropneumoniae infections in nursery and fattener pigs using Monte Carlo simulations.

This study evaluated the theoretical clinical outcome of three marbofloxacin posology regimens in two groups of pigs (weaners and fatteners) for the treatment of Actinobacillus pleuropneumoniae (App) and Haemophilus parasuis (Hp) infection and the appearance of resistant bacteria due to the antibiotic treatment. The probability of target attainment (PTA) for pharmacokinetic/pharmacodynamics (PK/PD) ratios associated with clinical efficacy and with the appearance of antimicrobial resistance for fluoroquinolones at each minimum inhibitory concentration (MIC) or mutant prevention concentration (MPC) were calculated, respectively. The cumulative fraction of response (CFR) was calculated for the three posology regimens against App and they ranged from 91.12% to 96.37% in weaners and from 93% to 97.43% in fatteners, respectively. In the case of Hp, they ranged from 80.52% to 85.14% in weaners and from 82.01% to 88.49% in fatteners, respectively. Regarding the PTA of the PK/PD threshold associated with the appearance of antimicrobial resistance, results showed that marbofloxacin would prevent resistances in most of the animals up to the MPC value of 1 µg/mL.

Effect of enrofloxacin in the carrier stage of Haemophilus parasuis in naturally colonized pigs.

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.

Cette étude avait comme objectif de déterminer l'effet de l'enrofloxacin dans le portage d'Haemophilus parasuis chez des porcs sevrés colonisés naturellement. Vingt-trois porcs colonisés par H. parasuis ont reçu au moment du sevrage par voie intramusculaire soit de l'enrofloxacin à un dosage de 7,5 mg/kg de poids vif (BW) ou une solution saline. Des écouvillons nasaux ou des amygdales ont été prélevés quotidiennement durant l'étude et à la nécropsie et testés par réaction d'amplification en chaîne par la polymérase quantitative (qPCR). Les isolats d'H. parasuis obtenus des échantillons prélevés lors de la nécropsie ont été soumis à une analyse génotypique par PCR des séquences intergéniques consensus répétitives des entérobactéries (ERIC-PCR) ainsi qu'à une épreuve PCR multiplex pour la détection des gènes auto-transporteurs trimériques associés à la virulence (vtaA). La présence d'H. parasuis fut détectée dans la cavité nasale et les amygdales des porcs du groupe témoin tout au long de l'étude. Les porcs traités aux antibiotiques étaient négatifs pour la présence d'H. parasuis au jour 1 post-traitement et la proportion d'échantillons nasaux qui ont testé positifs était plus élevée pour les porcs témoins que pour les porcs traités aux jours 1, 2, 3, 4, 5, 6 et 7 post-traitement et aux jours 2, 4 et 5 post-traitement pour les échantillons d'amygdales (P < 0,003). Le génotypage par ERIC-PCR a permis de montrer qu'à la fin de l'étude les porcs étaient colonisés par une souche commune d'H. parasuis. Les isolats étaient négatifs pour la présence du gène vtaA, ce qui indique l'absence du facteur de virulence vtaA. En conclusion, l'enrofloxacin a diminué significativement la charge d'H. parasuis chez des porcs colonisés naturellement, mais a été incapable d'éliminer complètement le microorganisme.(Traduit par Docteur Serge Messier).

[Evaluation of the efficacy of a combination therapy of an antibiotic and a NSAID following an experimental Haemophilus parasuis infection in nursery piglets]. / Wirksamkeitsbeurteilung einer Kombinationstherapie aus Antibiose und NSAID nach experimenteller Haemophilus-parasuis-Infektion bei Absetzferkeln.

OBJECTIVE: The aim of the present study was to evaluate the combination therapy of an antibiotic (enrofloxacine-arginine, Baytril® RSi) and a non-steroidal anti-inflammatory drug (NSAID, ketoprofen, Dinalgen® 60 mg/ml) against a Haemophilus parasuis (HPS) infection in nursery piglets. MATERIAL AND METHODS: Forty-eight 3-week-old pigs were divided into four groups (group 1: non-infected controls; group 2: HPS infection; group 3: HPS infection/Baytril® RSi; group 4: HPS infection/Baytril® RSi/Dinalgen®) and housed within the isolation facility. After an acclimatization period of 10 days, the piglets in groups 2-4 were intratracheally infected with 1 x 107 colony forming units (CFU) HPS serovar 5, whereas animals of group 1 received physiological saline. Total clinical scores and joint scores were calculated daily after clinical examination. Seven days after the infection, piglets were humanely euthanized. At necropsy, pathological findings on serosal surfaces were scored according to severity and extension. RESULTS: Group 1 had the lowest clinical and pathological scores, followed by groups 4, 3 and 2. Piglets treated with the combination of an antibiotic and an NSAID showed the lowest body temperatures (significant). The average daily weight gain (ADWG) was not significantly different between the groups, but piglets of group 4 tended to reach a higher mean ADWG (340.5 g/d) than animals of the non-infected group 1 (323.8 g/d), the Baytril® RSi-treated group 3 (278.0 g/d) and the positive control group 2 (247.0 g/d). Piglets of the positive control group (group 2) achieved the highest values in the clinical, joint and serositis scores. CONCLUSION: The study demonstrated that a simultaneous treatment with enrofloxacine-arginine (Baytril® RSi) and ketoprofen had a superior therapeutic effect compared to a single antibiotic treatment with Baytril® RSi in nursery piglets experimentally infected with HPS.

New insights in cellular immune response in colostrum-deprived pigs after immunization with subunit and commercial vaccines against Glässer's disease.

Four groups of colostrum-deprived pigs were immunized with Porcilis Glässer® (PG) or with subunit vaccines developed by us (rTbpA, NPAPT(M) or NPAPT(Cp)) against Glässer's disease, and they were challenged with 3×10(8)CFU of Haemophilus parasuis. A strong reduction in CD3(+)γδTCR(+) cells was seen in non-immunized control and scarcely protected (rTbpA) groups, suggesting that these cells could represent a target of H. parasuis infection. A significant increase in CD172α(+)CD163(+) cells was detected in all groups but PG, while a reduction in SLAIIDR(+) molecules expression was observed after challenge in control animals. Significant increases in CD3ε(+)CD8α(+)CD8ß(+) and B cells were detected respectively in control and NPAPT groups, and in scarcely (rTbpA) and well-protected (NPAPT(M) and NPAPT(Cp)) groups. Finally, a greater response in CD4(+)CD8α(-) cells was observed in NPAPT(Cp) compared to NPAPT(M) and PG groups. These results state the potential of NPAPT antigen for developing effective vaccines against Glässer's disease.

Comparison of real-time PCR and culture isolation in colostrum-deprived pigs immunized and challenged with Haemophilus parasuis.

AIMS: A real-time PCR (RT-PCR) based on the detection of the infB gene of Haemophilus parasuis is compared with culture isolation (Frandoloso et al., (2011) Clin Vaccine Immunol 18, 50-58.), evaluating different subunit or commercial vaccines. METHODS AND RESULTS: Samples from different tissues of 24 experimentally infected and challenged colostrum-deprived piglets were tested. The RT-PCR gave globally a 23·3% more of positive results than culture, and all samples being positive by culture were positive by RT-PCR also. H. parasuis could not be cultured from any of the samples of the piglets included in the three vaccinated groups resulting in a strong protection, but it could be detected by RT-PCR in six samples in the group immunized with the commercial vaccine, in three in that vaccinated with native proteins with affinity to porcine transferrin (NPAPT) administered intramuscularly and in only two in that immunized with NPAPT intratracheally. CONCLUSIONS: The RT-PCR was more sensitive than culture for H. parasuis detection in the organs compared. SIGNIFICANCE AND IMPACT OF THE STUDY: The RT-PCR evidenced that NPAPT vaccines were those yielding the best protection results in terms of H. parasuis clearance.

Acute phase protein concentrations in colostrum-deprived pigs immunized with subunit and commercial vaccines against Glässer's disease.

Haemophilus parasuis is the etiological agent of Glässer's disease, which is characterized by fibrinous polyserositis, polyarthritis and meningitis in pigs. This study was focused on the characterization of the acute-phase response after immunization and infection of colostrum-deprived pigs with H. parasuis serovar 5, by measuring serum concentrations of three positive acute-phase proteins (APPs) (pig major acute-phase protein pig, MAP; haptoglobin, HPG; C-reactive protein, CRP) and one negative APP (apolipoprotein A-I, ApoA-I). Six experimental groups were established: a non-immunized but infected control group (CTL); two groups immunized with either a recombinant transferrin-binding protein (Tbp) A or TbpB fragment from H. parasuis Nagasaki strain (rTbpA and rTbpB, respectively); two groups immunized with native outer membrane proteins with affinity to porcine transferrin (NPAPT), one of them inoculated intramuscularly (NPAPTim) and the other intratracheally (NPAPTit), and the last group receiving a commercially available bacterin (PG). The greatest concentrations of the three positive APPs and the lowest concentration of the negative APP were detected in CTL group, as well as in those animals belonging to rTbpA or rTbpB groups that died in response to challenge. Significant differences (P<0.005) were found in these groups when comparing challenge with the following days after it. However, no significant differences were seen for the remaining vaccinated groups (NPAPTim, NPAPTit and PG), which were effectively protected against Glässer's disease. Therefore, APPs could be used as useful biomarkers for both evaluating disease progression and determining vaccination effectiveness.

Acute-phase protein response in pigs experimentally infected with Haemophilus parasuis.

The acute-phase protein (APP) response to an infection caused by Haemophilus parasuis, the etiological agent of Glässer's disease in pigs, was characterized measuring serum concentrations of pig major acute-phase protein (pig MAP), haptoglobin (HPT), C-reactive protein (CRP) and apolipoprotein A-I (ApoA-I) in colostrum-deprived pigs. They were divided into six experimental groups: non-immunized control group (I); immunized with a non-commercial bacterin (II); with an OMP-vaccine (III); with a sublethal dose (IV); and with two commercial bacterins (V and VI). All groups were challenged intratracheally with 5 × 10(9)CFU of H. parasuis 37 days after immunisation. The highest levels of the positive APPs (pig MAP, HPT and CRP) and the lowest levels of the negative APPs (ApoA-I) were observed in the animals that died as a consequence of the infection, both those in the non-immunized and in the immunized groups. However, the surviving animals (all of them in groups II, V and VI, two pigs in group III, and three in group IV) showed a minor variation in APP response, mainly on day 1 post-challenge (p.c.), and then tended to recover the initial values. APP response was still less pronounced in the groups of pigs previously immunized with bacterins. In conclusion, APP response can reflect Glässer-disease ongoing, showing a correlation between the severity and duration of the clinical signs and lesions and the magnitude of changes in the APP levels.

Cytokine expression in colostrum-deprived pigs immunized and challenged with Haemophilus parasuis.

The expression of several cytokines in spleen, pharyngeal lymph nodes, lung and brain after different immunization procedures and a challenge with 5 x 10(9) 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 10(5) 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-1alpha 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-alpha and IFN-gamma were expressed in significantly higher levels by survivors (for all the four cytokines in lymph nodes; for IL-4, IL-10 and TNF-alpha in spleen; for IL-4, TNF-alpha and IFN-gamma in lung, and only for TNF-alpha in brain), thus suggesting a role of these four cytokines in the adaptive response, which might contribute to protection against H. parasuis infection.

Comparison of sampling sites and detection methods for Haemophilus parasuis.

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.

Comparison between Haemophilus parasuis infection in colostrums-deprived and sow-reared piglets.

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.

Development of polymerase chain reaction and comparison with in situ hybridization for the detection of Haemophilus parasuis in formalin-fixed, paraffin-embedded tissues.

DNA extraction and nested polymerase chain reaction (PCR) were developed for the detection of Haemophilus parasuis from formalin-fixed, paraffin-embedded tissues. The results for nested PCR were compared with those determined by in situ hybridization. The optimal results obtained show that use of xylene deparaffinization, digestion with proteinase K followed by nested PCR is a reliable detection method. A distinct positive signal was detected in 20 pigs naturally infected with H. parasuis by in situ hybridization. The rate of agreement between nested PCR and in situ hybridization for the detection of H. parasuis in formalin-fixed, paraffin-embedded tissues was 100%. The nested PCR could be applied successfully to formalin-fixed, paraffin-embedded tissues for the detection of H. parasuis with bacterial isolation.

Effect of vaccinating sows and their piglets on the development of Glässer's disease induced by a virulent strain of Haemophilus parasuis serovar 5.

Ten pregnant gilts were divided into two groups of five and one group was vaccinated at 80 and 95 days of pregnancy with a commercial bacterin containing Haemophilus parasuis serovars 2, 3 and 5. Half the piglets born to each group of gilts were vaccinated at seven and 21 days of age with the same bacterin, and one week after they were weaned at five weeks, all the piglets were inoculated intratracheally with 10(6) colony-forming units of Hparasuis serovar 5. At slaughter, a significantly smaller percentage of the lungs of the pigs born to the vaccinated gilts was affected by pneumonic lesions, and significantly fewer of them had arthritic joint changes. The average daily liveweight gain of the pigs born to the vaccinated gilts was significantly greater than that of those born to the unvaccinated gilts, but the vaccination of the piglets had no effect. There was no significant difference between the feed conversion ratios of the four groups of piglets, and none between the average times they took to reach slaughter weight. The pigs born to the vaccinated gilts had higher ELISA titres to Hparasuis than those born to the unvaccinated gilts.

Interaction of Haemophilus parasuis with nasal and tracheal mucosa following intranasal inoculation of cesarean derived colostrum deprived (CDCD) swine.

Twenty-three cesarean derived, colostrum deprived pigs were obtained at 5 wk of age and inoculated intranasally with either 1.4 x 10(8) colony forming units of Haemophilus parasuis or sterile phosphate buffered saline. Pigs were euthanized at 4, 8, 12, 18, 26, or 36 h post-inoculation and tissues from the oropharynx and respiratory tract were obtained for qualitative bacterial culture, immunohistochemistry for H. parasuis antigens, and light and transmission electron microscopy. Haemophilus parasuis was consistently isolated from the nasal cavity (17/17, 100%) and trachea (13/17, 76%) and rarely isolated from the lung (3/17, 18%) and blood stream (1/17, 6%) of infected pigs. Antigens of H. parasuis were sporadically detected on the nasal mucosa (6/17, 35%) and trachea (8/17, 47%). Light microscopic lesions included submucosal and intraepithelial infiltrates of neutrophils and infrequent, patchy loss of cilia. Ultrastructural changes in nasal mucosal epithelial cells included cell protrusion, loss of cilia, and dilation of the cytocavitary network. Bacteria were infrequently identified and were either within an amorphous material at the apical surface of the cilia or were between individual cilia. These results suggest H. parasuis associates with the nasal mucosa and can induce a suppurative rhinitis with nasal mucosal epithelial cell degeneration. This process may represent an initial event in the pathogenesis of H. parasuis infection of swine.

Effects of various vaccination protocols on passive and active immunity to Pasteurella haemolytica and Haemophilus somnus in beef calves.

Two field trials were conducted in a beef cow herd in Saskatchewan to determine the effectiveness of a combined Pasteurella haemolytica and Haemophilus somnus vaccine in increasing passively and actively acquired antibodies in beef calves. Vaccination of dams at 4 and/or 7 weeks prepartum was associated with increased antibody titers to P. haemolytica and H. somnus in their serum (P < 0.05), colostrum(P < 0.05), and serum of their calves at 3 days and 1 month of age (P < 0.05). There was no significant(P > 0.05) difference in antibody titers in the colostrum and serum of calves from single or double vaccinated dams. Calves vaccinated at 1 and 2 months of age in the face of maternal antibodies toP. haemolytica and H. somnus had significantly(P < 0.05) higher antibodies to P. haemolytica and H. somnus at 4 and 6 months of age than did unvaccinated calves. Calves vaccinated at 3 and 4 months of age in the face of low levels of preexisting antibodies had significantly (P < 0.05) higher antibodies toP. haemolytica at 5 months of age and to H. somnus at 5 and 6 months of age than did unvaccinated calves. Calves vaccinated once at 4 months of age had significantly(P < 0.05) higher antibody titers toP. haemolytica and H. somnus at 4.5 months of age than did unvaccinated calves, but this difference was not apparent at 6 months of age. These results suggest that vaccination of beef cows with a combined Pasteurella haemolytica and Haemophilus somnus vaccine once at 4 weeks prepartum will significantly (P < 0.05) increase passive antibody titers toP. haemolytica and H. somnus in their calves. Double vaccination of calves with preexisting maternal antibodies at 1 and 2 months of age will increase antibody titers to P. haemolytica and H. somnus until 6 months of age. Vaccination of beef calves with low levels of preexisting antibody at 3 and 4 months of age will increase antibody titers to H. somnus until 6 months of age and to P. haemolytica until 5 months of age.However, the level of antibodies achieved by vaccination may depend on the calves being studied, the level of preexisting antibodies, and the efficiency of passive transfer.

Isolation of pathogenic strains of Haemophilus somnus from the female bovine reproductive tract.

The prevalence of Haemophilus somnus in the genital tract of slaughtered and live cows in southern Ontario was investigated. The vagina and uterus of slaughtered cows were swabbed separately. Live cows were examined and sampled in two field surveys: Centre A and Centre B. In the former, aspirated mucus secretions and in the latter, specimens obtained by guarded swabbing were examined bacteriologically. Haemophilus somnus was isolated from 28 genital tracts of 461 slaughtered (6.1%), and seven of 199 live (3.5%) cows during the centre B survey. The isolates were recovered from both normal and diseased reproductive tracts. Fourteen strains isolated from genital organs were examined for pathogenicity in vivo to test the occurrence of pathogenic isolates. In the initial stage of the in vivo study on pathogenicity, each of the fourteen isolates was examined on one calf using an intracisternal inoculation. Subsequently, one pathogenic and one nonpathogenic strain were inoculated into five calves each to statistically confirm their pathogenic potential. Of 14 genital isolates of H. somnus examined in an intracisternal calf assay, six (43%) caused a fatal peracute neurological disease, while eight were nonpathogenic. A comparative pathological study of pathogenic and nonpathogenic isolates showed that the former caused a severe fatal suppurative meningoencephalitis whereas the latter caused no lesions whatsoever or a mild leukocytic leptomeningitis. The salient data obtained in this study indicate that there are pathogenic strains of H. somnus in the genital tract of apparently normal cows as well as of those with inflammatory disease.

In vitro antimicrobial activity of sulfonamides against some porcine pathogens.

The minimal inhibitory concentrations (MIC) of sulfonamides were determined against Bordetella bronchiseptica (n = 10), Pasteurella multocida (n = 10), Haemophilus pleuropneumoniae (n = 20), and Streptococcus suis (n = 10) strains isolated from pigs with atrophic rhinitis, pneumonia, or meningitis. Sulfonamides tested in an agar dilution method were sulfachloropyridazine, sulfadiazine, sulfadimethoxine, sulfamethazine, sulfadoxine, sulfisoxazole, sulfamerazine, sulfamethoxazole, sulfamethoxypyridazine, sulfanilamide, sulfatroxazole, and sulfisomidine. Results indicated that monotherapy of S suis infections with sulfonamides should not be encouraged because the MIC50 of all sulfonamides investigated was greater than 32 micrograms/ml. The MIC50 of the sulfonamides against B bronchiseptica ranged from 0.5 to 8 micrograms/ml, against P multocida from 2 to 32 micrograms/ml, and against H pleuropneumoniae from 8 to 64 micrograms/ml. The MIC50 of sulfachloropyridazine, sulfadiazine, sulfadimethoxine, sulfamerazine, and sulfamethoxazole for the gram-negative bacteria did not exceed 16 micrograms/ml. Among these compounds, sulfamethoxazole had the highest activity. The frequently prescribed sulfamethazine had an overall low antimicrobial activity.