Haemophilus Pleuropneumoniae Strains Research Articles

Efficacy of a bivalent vaccine containing serovar 2 and 5 strains of Haemophilus pleuropneumoniae in pigs or in guinea pigs.

Efficacy of a bivalent vaccine containing serovar 2 and 5 strains of Haemophilus pleuropneumoniae in pigs or in guinea pigs.

Effect of heat treatment on the surface antigens of Haemophilus pleuropneumoniae.

Coagglutination and ring precipitation tests were used to study the effect of heat on the surface antigens of Haemophilus pleuropneumoniae strains employing the reference strains belonging to serotypes 1 to 7 and field isolates belonging to serotypes 1, 2, 3, 5, 6 and 7. By immunising rabbits with formalin-fixed whole-cell suspension, antibodies were obtained which sensitised Cowan I Staphylococcus aureus to coagglutinate antigen preparations which had not been heated, or heated at 56 degrees C, or boiled or autoclaved. Similar positive reactions were obtained with the ring precipitation test. Heating the cultures at 56 degrees C for one hour was best for exposing the most potent serotype-specific antigens in all the strains studied. All the reference strains and most of the field isolates possessed the thermostable type specific antigens which could withstand autoclaving for one hour. However, many field isolates belonging to serotype 1 did not possess this antigen. The apparent antigenic heterogeneity of serotype 1 strains based on the presence or absence of these thermostable antigens could be valuable in epidemiological investigations. It was shown that most potent serotype-specific antigens are present as freely diffusible material on the surface layer of the bacterial cells, which could easily be removed by washing in saline solution. Well washed bacterial cells devoid of surface materials are poor antigens. It is recommended that test strains should not be heated above 56 degrees C for serotyping because higher temperatures are liable to destroy the capsular antigen of some strains and render the culture untypeable.

Identification of serotypes of Haemophilus pleuropneumoniae by counterimmunoelectrophoresis.

Counterimmunoelectrophoresis, direct immunofluorescence and immunodiffusion procedures were used to serotype 15 strains of Haemophilus pleuropneumoniae isolated from the respiratory tract of pigs in southern Brazil. Antigens were prepared by extracting cultures with a saline solution or by the phenol-water method. Antisera were prepared in rabbits against serotypes 1, 2, 3 and 5. Thirteen of the isolates were type 5 and two were type 3. No differences were observed between the results obtained in serotyping with counter immunoelectrophoresis and direct immunodiffusion, but both procedures were significantly better than immunodiffusion except with the saline extracted antigen. Counterimmunoelectrophoresis was quicker, more sensitive and more easily performed than the other techniques.

Immunologic relationships among Haemophilus pleuropneumoniae strains of serovar 1 to 5 in guinea pigs.

Immunologic relationships among Haemophilus pleuropneumoniae strains of serovar 1 to 5 were studied in guinea pigs by the complement-fixation (CF) and the protection tests. Most of the guinea pigs, which had been injected once or twice with each monovalent vaccine, had the homologous CF titers of 1:16 or higher, and survived the intraperitoneal challenge inoculation with the homologous strain, whereas none of the animals having titers of 1:8 or below survived, irrespective of the combinations of vaccines and chalenge strains. Most of the injected guinea pigs produced the CF antibodies whose titers were 1:16 or higher against the homologous strain, whereas below 1:8 against the strains of heterologous serovar. Thus, the production of CF antibodies and protection against the challenge strains were closely related each other and showed serovar specificity in the injected guinea pigs. On the other hand, especially in the guinea pigs injected twice, varying extents of cross-protection (maximum 37.5%) was observed among the strains of serovar 1 to 5. The cross-protection pattern was apparently associated with the cross-reaction pattern in the CF test. These results suggest that existence of common CF antigen(s) among the H. Pleuropneumoniae strains correlates with the cross-protection in guinea pigs.

Production of RNA-dependent haemolysin by Haemophilus pleuropneumoniae

Five strains of Haemophilus pleuropneumoniae, out of eight strains tested, produced extracellular haemolysin(s) when grown in liquid culture in the presence, but not in the absence, of RNA. The haemolysin produced by the neotype strain was unstable, heat labile, and sensitive to degradation by pronase, trypsin, and chymotrypsin; moreover, trypan blue treated haemolysin preparations were less effective at causing erythrocyte lysis than were untreated preparations. Following growth in the absence of RNA, washed suspensions of the neotype strain produced extracellular haemolysin when incubated in the presence of RNA, glucose, and casein acid hydrolysate; extracellular haemolysin could not be detected if the incubation mixture contained chloramphenicol. The haemolysin produced by washed bacterial suspensions was similar to that produced by growing cultures in that it was unstable, heat labile, and sensitive to inactivation by the same complement of enzymes. Erythrocyte lysis induced by either haemolysin preparation was preceded by a prelytic phase, the duration of which was dependent upon haemolysin concentration and the initial temperature of the haemolysin--erythrocyte mixture. It is concluded that the haemolysin(s) produced by the neotype strain of H. pleuropneumoniae is distinct from, but closely related to both streptolysin S and the haemolysin produced by Treponema hyodysenteriae.

Serological Characterization of 8 Haemophilus Pleuropneumoniae Strains and Proposal of a New Serotype: Serotype 8

Eight strains of Haemophilus pleuropneumoniae isolated from 8 herd outbreaks of pleuropneumonia in pigs were studied by means of the slide agglutination test, the tube agglutination test, the IHA test and by gel diffusion. The 8 strains were antigenically homogeneous and serologically distinct from serotypes 1 through 7. It is therefore proposed to refer these strains to a new serotype: serotype 8, with strain 405 as the type strain. In addition to the serotype-specific capsular antigens, capsular antigen of serotype 3 (strain 1421) and serotype 6 (strain Femo) could be demonstrated in the 8 strains by means of the IHA test and by gel diffusion analyses.

Deoxyribonucleic Acid-Deoxyribonucleic Acid Hybridization Analysis of Actinobacillus actinomycetemcomitans and Haemophilus aphrophilus

Deoxyribonucleic acids (DNAs) were isolated and purified from 20 strains of Actinobacillus actinomycetemcomitans, 6 strains of Haemophilus aphrophilus, 2 strains of Haemophilus paraphrophilus, 2 strains of Haemophilus pleuropneumoniae, 2 strains of Haemophilus paraphrohaemolyticus, 2 strains of Haemophilus influenzae, and 1 strain each of Actinobacillus lignieresii, Actinobacillus suis, Haemophilus aegyptius, Haemophilus parainfluenzae, and Haemophilus parahaemolyticus. The guanine-plus-cytosine contents of the DNAs were determined, and they agreed closely with previous estimates. DNA-DNA hybridization analyses revealed that all of the strains identified as A. actinomycetemcomitans were at least 69% homologous to DNA probes from two A. actinomycetemcomitans strains (strains NCTC 9710T [T = type strain] and Y4). The H. aphrophilus and H. paraphrophilus strains were homologous to the two A. actinomycetemcomitans probes at levels of 25 to 37%. The DNAs of all A. actinomycetemcomitans strains were homologous to a DNA probe from H. aphrophilus strain NCTC 5906 at levels of 30 to 39%. The two strains of H. paraphrophilus tested were homologous to the H. aphrophilus probe at levels of 73 and 77%, indicating a very close relationship between these groups of organisms. H. parainfluenzae strain ATCC 9796 DNA seemed to be homologous to the H. aphrophilus and A. actinomycetemcomitans probes at low but significant levels (12 to 16%). All of the other strains of haemophili and actinobacilli tested gave 10% or less homology with the three test probes.

Determination of antigenic specificity and relationship among Haemophilus pleuropneumoniae serotypes by an indirect hemagglutination test.

Serological properties of antigens extracted from strains of Haemophilus pleuropneumoniae belonging to seven different serotypes were investigated. Antisera were prepared in rabbits against Formalin-treated whole cell suspensions as well as autoclaved cell suspensions. Saline and heat extracts and their alcohol precipitate antigens of H. pleuropneumoniae were used in the indirect hemagglutination test. All the antigens used were easily adsorbed directly onto sheep erythrocytes. Saline extract antigen showed maximum type specificity. Heating of the whole cell suspension revealed the cross-reactive minor antigenic determinants. Thus, the heat extract preparations had both type-specific and species-specific antigens. It is suggested that the indirect hemagglutination test may be useful for both serotyping and serodiagnosis of H. pleuropneumoniae infections in pigs.

The haemolytic activity of Haemophilus species.

The importance of the species of blood employed for detection of haemolysis in seventy-seven Haemophilus strains of human and porcine origin was studied. Significant differences in the visibility of haemolytic zones obtained on the different blood agar media were observed. In decreasing order, the suitability of the species of blood was: calf, sheep, human, rabbit, poultry and horse blood. On plates containing washed sheep or calf red cells the haemolysin of all 36 strains of Haemophilus pleuropneumoniae acted synergistically with the beta-toxin of the Staphylococcus aureus strain used as "feeder strain", giving rise to a lytic phenomenon resembling the CAMP reaction.