Serological studies of Actinobacillus (Haemophilus) pleuropneumoniae strains of serotype 6 and their antigenic relationship with other serotypes

Serological tests such as agglutination, coagglutination, precipitation and indirect haemagglutination were used to study the antigenic relationship of reference and field strains of Actinobacillus (Haemophilus) pleuropneumoniae of serotype 6 with reference strains of other serotypes. Both cell-associated particulate and cell-free soluble antigens prepared from unheated and heat-treated bacterial suspensions of reference and field strains of serotype 6 were used in the studies. Species-specific, common antigenic determinants associated mainly with heat-treated particulate antigens of serotype 6 were cross-reactive in tube agglutination tests with almost all the serotypes. The species-specific antigens were of a minor nature because the cross-reactivities were abolished in both 2-mercaptoethanol agglutination and coagglutination tests. Cell-free saline extracts of both unheated and heat-treated suspensions of serotype 6 strains possessed epitopes specific for serotypes 3, 5 and 8 in addition to their own specific determinants. The epitopes were dominant because the reactions of strains of serotype 6 with antisera against serotypes 3, 5 and 8 persisted in almost all the serological tests used. Serotype 6 strains were antigenically closer to serotype 8 than to serotypes 3 or 5. A combination of serological tests such as coagglutination followed by 2-mercaptoethanol tube agglutination and, or, immunodiffusion tests differentiated serotype 6 strains from those of other cross-reacting serotypes.     

Serological characterization of Actinobacillus pleuropneumoniae strains isolated from pigs in Quebec.

A total of 3306 isolates of A. pleuropneumoniae originating from lung tissues of pigs that died of acute pleuropneumonia and 140 isolates recovered from tonsils or nasal cavities of apparently healthy pigs from chronically infected herds were serotyped. Various serotyping methods, such as slide agglutination, tube agglutination, ring precipitation, coagglutination, immunodiffusion, indirect hemagglutination and counterimmunoelectrophoresis either alone or in combination were used. The techniques used for serotyping continued to evolve during the last 10 years depending on the problem encountered in serotyping. Antisera prepared in rabbits against formalinized whole cell suspensions of reference strains of A. pleuropneumoniae of serotypes 1 to 12 were employed for serotyping. Serotype 1 was predominant ranging from 55 to 87% from year to year during the last 10 years with an average prevalence of 68%. Serotype 5 was second in prevalence ranging from 9 to 30% with a mean of 23%. Both subtypes of serotype 5 (5a and 5b) were present in Quebec. Serotypes 3, 6, 7, 8, 10 and 12 were isolated in small numbers together accounting for about 9%. Serotypes 4, 9 and 11 were not present. Cross-reactions were observed among isolates of serotypes 3, 6 and 8, and 1, 9 and 11 and were easily differentiated from each other by quantitation of type and group specific antigens by coagglutination and immunodiffusion tests. Serotypes 1, 5 and 7 were isolated most frequently from tonsils of pigs from chronically infected herds. Prevalence of different serotypes in different countries has also been reviewed.     

Serologic studies of Actinobacillus (Haemophilus) pleuropneumoniae strains of serotype-3 and their antigenic relationships with other A pleuropneumoniae serotypes in swine

Serotype-3 strains of Actinobacillus (Haemophilus) pleuropneumoniae possess epitopes shared with almost all other Actinobacillus serotypes. Common epitopes detected in particulate antigens were heat-labile and heat-stable and were of minor nature. Additional cross-reactive epitopes were detected in soluble and particulate antigens prepared from strains of serotypes 3, 6, and 8. Cross-reactions occurring between serotype-3 antigens and those of other serotypes were of 2 types: one associated mainly with 2-mercaptoethanol (2-ME)-sensitive IgM antibodies and the other associated mainly with 2-ME-resistant IgG antibodies. The cross-reactivity between serotypes 3, 6, and 8 was associated mainly with IgG antibodies, as shown by the results of 2-ME tube agglutination, 2-ME-indirect hemagglutination, and coagglutination tests. None of the tests was entirely satisfactory for serotyping serotype-3 isolates, mainly as a result of overlapping of type-specific antigenic determinants of serotypes 3, 6, and 8 in different combinations in the same strain. A combination of tests, using particulate and soluble antigens, may be necessary for typing of serotype-3 isolates.     

鸭疫里默氏菌一个可能新型的鉴定

用玻片凝集试验、试管凝集试验和琼脂扩散沉淀试验，对13个鸭疫里默氏菌分离株进行了血清型鉴定。玻片凝集试验中，这些菌株只与8型抗血清发生强凝集反应，试管凝集试验中，其代表菌株C882与8型参考菌株之间仅存在单向低度交叉凝集反应，且与1～19型中的其他18个血清型参考菌株无可见的交叉凝集反应；用C882吸收可消除8型抗血清与C882等菌株之间的交叉凝集反应，但不影响8型抗血清的同源凝集反应和沉淀反应能力。琼扩试验中，C882与1～19型参考菌株之间不产生可见的交叉沉淀反应。沉淀反应模式表明，以C882为代表的13株细菌的热稳定抗原具有同一性。结果表明，13株待检菌株可能属于一个新的血清型。     

Detection of strains of Haemophilus pleuropneumoniae using the coagglutination test

Three tests were used for the serological identification of the strains of Haemophilus pleuropneumoniae (Actinobacillus pleuropneumoniae) isolated from pigs and coming from 66 sites where pleuropneumonia, caused by Haemophilus, occurred in pigs. The coagglutination test was found to be the best for the identification of the causal agent; the ring precipitation test was somewhat less sensitive, and worse results were obtained when rapid slide agglutination was used. Of all the field isolates of H. pleuropneumoniae, serovar 2 occurred most frequently (56%), followed by serovar 1 (39%); one strain was identified as serovar 7. Two strains have remained unidentified. The serological identification of the strains was performed on the basis of their comparison with eight type serovars of H. pleuropneumoniae.     

Actinobacillus (Haemophilus) pleuropneumoniae serotype-8 isolates and their antigenic relationships with other A pleuropneumoniae serotypes

Antigenic relationship of Actinobacillus (Haemophilus) pleuropneumoniae serotype-8 isolates with other serotypes was studied, using tube agglutination, with and without 2-mercaptoethanol, indirect hemagglutination with and without 2-mercaptoethanol, ring precipitation, coagglutination, and immunodiffusion tests. Serotype-8 isolates possessed serotype-specific, group-specific common antigens cross-reactive with serotypes 3 and 6 and species-specific common antigens cross-reactive with other serotypes. Absorption studies were done to study the antigenic relationship of serotype 8 with serotypes 3 and 6. Rabbit antisera against whole-cell (WC) suspensions of reference strains of serotypes 3, 6, and 8 were used for absorption studies with WC and boiled WC suspensions of homologous and heterologous serotypes. Unabsorbed and absorbed sera were tested for antibodies against WC and boiled WC antigen preparations of serotype 8, using various serotests. Absorption studies revealed that serotype-8 strains possessed 2 main types of epitopes, one of which was serotype-specific and did not have cross-reactivity with other serotypes. The second type of epitopes was group specific and was cross-reactive with serotypes 3 and 6.     

Serological characterisation of Australian isolates of Actinobacillus pleuropneumoniae

SUMMARY: Using hyperimmune rabbit antiserums to 8 reference serovar strains, a total of 51 Australian isolates of Aclinobacillus pleuropneumoniae were serotyped by either a rapid slide agglutination test or a gel diffusion test. The results were: serovar 1 — 24 isolates; serovar 2 — 6 isolates; serovar 3 — 5 isolates; serovar 7 — 15 isolates; nontypable — 1 isolate. The rapid slide agglutination test was suitable for screening field isolates, as 73% of those tested reacted with only 1 of the 8 antiserums and were assigned to a particular serovar of the basis of this test alone.     

A Serological Survey of Dogs for Brucella canis in Southwestern Ontario

Sera from 2 000 dogs from southwestern Ontario were tested for antibodies to Brucella canis by a rapid slide agglutination test. The 100 positively reacting sera were tested by tube agglutination and immunoprecipitation (gel diffusion) tests. Thirty-one of these sera gave suspicious titres and one a positive titre in the tube agglutination test. Six of the rapid slide test positive sera gave positive reactions in immunoprecipitation tests. One dog was identified which was found to have a history very suggestive of B. canis infection. It was judged that 0.3% of sera tested showed serological evidence of B. canis infection. The complexities of the serological diagnosis of B. canis infection was apparent, in particular the tendency to false-positive results in the rapid slide agglutination test.     

鸭疫里默氏菌6型、12型与16型之间的交叉反应

鸭疫里默氏菌6型和12型仅在玻片凝集试验中存在较弱的交叉反应，这一交叉反应可经血清吸收试验得到消除；12型和16型在玻片凝集试验和试管凝集试验中均表现较强的双向交叉反应，但在琼扩试验中表现为较弱的单向交叉反应；6型和16型之间没有交叉反应。12型和16型之间的交叉反应和交叉沉淀反应均可经血清吸收后得以消除，但经过16型参考菌株吸收过的12型抗血清，还与6型菌株存在交叉凝集反应，只有同时用6型和16型菌株吸收，12型抗血清才具有血清型特异性。血清吸收对6型和12型抗血清的同型凝集反应能力和沉淀反应能力没有影响，但对16型抗血清的同源凝集能力和沉淀反应能力均有较大的影响，经、2型参考菌株吸收后，16型抗血清与同型抗原之间的凝集效价下降2个滴度，而且不再形成清晰的沉淀线。结果表明，6型和12型之间、12型和16型之间均存在ab-bc的抗原关系。     

Classification of Bacteroides nodosus by agglutination tests

One thousand two hundred and sixty seven isolates of Bacteroides nodosus from 292 sheep in 58 flocks were examined. Of these, 1260 could be classified by slide agglutination into 8 serogroups designated A to H. Up to 6 serogroups were detected in individual flocks, with up to 4 serogroups being detected in a single foot. Of the 292 sheep examined, 38 (13%) carried mixed serogroup infections. Determination of the range of serological types infecting a flock frequently required the examination of a number of isolates from each of a number of sheep. Cross-tube agglutination tests carried out on 44 isolates and their antiserums indicated that members of some serogroups could be divisible into subgroups or serotypes. These results suggested that 16 or more serotypes of B. nodosus might exist. The nature of the antigens responsible for both slide and tube agglutination reactions needs to be determined.     

Study of antigenic heterogeneity among Actinobacillus pleuropneumoniae serotype 7 strains

Actinobacillus pleuropneumoniae serotype 7 strains were studied for their antigenic heterogeneity using rabbit polyclonal hyperimmune sera against all the known twelve reference strains of A. pleuropneumoniae and a battery of different serological tests such as coagglutination (COA), immunodiffusion (ID), indirect hemagglutination (IHA), counterimmunoelectrophoresis (CIE), rapid dot-ELISA (RDE), serum soft-agar (SSA) and growth agglutination (GA). Reference serotype 7 strain (WF83) showed cross-reactivity with reference serotype 1B strain but not with other serotypes. Field serotype 7 strains showed cross-reactivities with serotypes 1A, 1B, 4, 9, 10, and 11 in COA, ID, and CIE tests, but not in IHA test. Two field strains of serotype 7 (90-3182 and 86-1411) which appeared to be different from the typical serotype 7 strains were selected for further antigenic characterization by SDS-PAGE, Western blot, and Tricine SDS-PAGE assays, and identified as serotypes 1 and 7, respectively. For serotyping atypical strains, it is suggested to use Western blot assay as a confirmatory test to identify serotype-specific capsular and somatic antigens.     

Serological and bacteriological investigations of chickens from flocks naturally infected with Salmonella enteritidis

Groups of 10 birds were obtained from four flocks which had shown evidence of natural salmonella infection. S enteritidis had been isolated from three flocks and S typhimurium from the fourth. Each bird was housed in a separate cage and blood samples and cloacal swabs were taken weekly to follow the course of natural infection. After four weeks the birds were killed and examined post mortem. The isolation of Salmonella species could not be related to the serological results. In individual birds the rapid slide test and tube agglutination test could not be relied upon to detect infection; the microantiglobulin test and the enzyme-linked immunosorbent assay (ELISA) were more sensitive than the other tests and detected some infected birds that were negative by the rapid slide and tube agglutination tests, and also showed high titres in some birds from which Salmonella species could not be isolated post mortem. Sera obtained from two flocks which had a history of natural S enteritidis infection were evaluated by all the tests; evidence of infection was found with the microantiglobulin and ELISA tests but not with the other tests.     

Evaluation of three slide agglutination tests for rapid identification of Staphylococcus aureus.

Three slide agglutination tests for identification of Staphylococcus aureus were compared. The agglutination tests used for evaluation were Staphaurex (Wellcome Diagnostics), Staphyslide-Test (BioMerieux), and ANI S. aureus TEST (Ani Biotech Oy). A total of 347 isolates were analyzed, including 288 strains of S. aureus, 49 of S. epidermis, 11 of S. intermedius, 12 strains of other staphylococci and 14 non-staphylococcal strains. One hundred of the S. aureus strains were isolates from cases of food poisoning, 129 from mastitis and 59 from other clinical cases. The sensitivities of the tests were also compared using diluted suspensions of S. aureus strains and with purified Protein A dilutions. The results showed that the sensitivities of the tests were 98.6%, 97.9% and 99.0% for Staphaurex, Staphyslide-test and ANI S. aureus TEST, respectively. The specificities were 100% for the Staphyslide test and 98.8% for both the ANI S. aureus TEST and the Staphaurex test. The sensitivities measured with diluted S. aureus strain suspensions and Protein A solutions were equal with the Staphaurex and ANI S. aureus TEST. All the agglutination tests studied proved to be practical, easy to use and accurate for the rapid identification of S. aureus strains from culture isolates.     

Serological diagnosis of the species Haemophilus equigenitalis using the rapid agglutination and coagglutination method

The method of rapid slide agglutination and coagglutination was tested in the detection of Haemophilus equigenitalis (Taylorella equigenitalis)--the causal agent of contagious equine metritis (CEM). It was demonstrated that both methods were suitable for the serological diagnosis of the species under study. The antisera obtained from rabbits immunized with Haemophilus equigenitalis strains treated in different ways were specific, but with different antibody titres. When cross reactions with other species of microorganisms were verified, the antisera did not react with any of the strains, even after binding them to protein A of the positive strain Staphylococcus aureus--Cowan I. Coagglutination was much more rapid and pronounced than the ordinary rapid agglutination test. It was characterized by a low consumption of specific antiserum. The specific antibodies bound to staphylococci were kept at the temperature of 4 degrees C for several months without losing agglutinin activity.     

A rapid slide agglutination test for the serodiagnosis of Brucella canis infection that employs a variant (M-) organism as antigen

An improved antigen is described for use in the serodiagnosis of canine brucellosis. The novel antigen employs a less mucoid (M-) variant of B. canis. The replacement of Brucella ovis with B. canis (M-) cells as antigen in slide agglutination tests would increase accuracy by reducing the rate of false positive results from ca. 50% to ca. 10%. Rose bengal stained B. canis (M-) cells are slightly more sensitive than the commercially available Brucella ovis as slide test antigens. Both test procedures require brief (less than 1 min) reaction with 2-mercaptoethanol in order to reduce the rate of false positive reactions.     

Serotyping of Haemophilus pleuropneumoniae by rapid slide agglutination and indirect fluorescent antibody tests in swine

One hundred and forty-one isolates of Haemophilus pleuropneumoniae from Iowa and Illinois swine were characterized morphologically and biochemically and serotyped by rapid slide agglutination (RSA) and indirect fluorescent antibody (IFA) tests. Hyperimmune antisera were produced in rabbits using inactivated whole-cell suspensions of the reference strains for H pleuropneumoniae serotypes 1 to 7 and strain 202, representing the taxon "minor group." Cross testing of the reference strains and reference antisera indicated the antisera to be essentially serotype-specific, although reactivity of some antisera with heterologous strains was observed. Cultures of the 141 isolates formed adherent or smooth colonies or mixtures of these colony forms. Adherent and smooth colony types were found in all serotypes identified. Microscopic and biochemical characteristics of all isolates were typical of those previously described for H pleuropneumoniae. The overall incidence of H pleuropneumoniae serotypes was serotype 5, 55.3%; serotype 1, 34.0%; serotype 7, 7.8%; and nontypeable, 2.8%. Comparing the 2 test procedures, 87.2% of the isolates could be typed by RSA, and 66.0% could be typed by IFA. Cross-reactions between serotype 4 antisera and serotype 5 and 7 isolates were common with the IFA test. The reactions with serotype 7, but not serotype 5, were eliminated by cross adsorption of serotype 4 antisera. There was good correlation between the 2 test procedures, but RSA was judged to be more specific and sensitive than IFA.     

血清10型鸭疫里默氏菌第5个血清亚型的分析

采用琼脂扩散沉淀试验、玻片和试管凝集试验以及血清吸收凝集试验，对6株鸭疫里默氏菌分离株进行了抗原性分析。这6株分离株被鉴定为血清10型，但它们与10型内已知的4个亚型菌株之间又存在明显的抗原差异，因此，以菌株C919为代表的6个分离株被鉴定为血清10型的第5个亚型。     

A serologic survey of a population of Georgia dogs for Brucella canis and an evaluation of the slide agglutination test.

In a serologic survey of stray and pet dog populations of Georgia, serums were screened for Brucella canis antibodies, using the slide agglutination test. If results were positive, B canis antibody titers were determined, using the standard tube agglutination test. The stray dogs had significantly (P less than 0.01) higher titers than did the pet dogs. The reactor rate was 58% higher for the slide agglutination test than for the tube agglutination test. The manufacturer's evaluation of the slide agglutination test was based on a comparison of the serologic results of that test with those of the tube agglutination test, using a comparative method that permitted the results to be interpreted as 99% agreement between the 2 tests. Reevaluation of the manufacturer's data by a different method indicated that the slide agglutination test is very accurate when the results are negative (99.7% specific) but less so when the results are positive (62.5% sensitive).     

Serology of Campylobacter fetus fetus strains from four outbreaks of ovine abortion

As a result of a Ministry of Agriculture & Fisheries survey on ovine abortions, 76 isolates of Campylobacterfetus fetus were obtained. These isolates were from four farms in the southern Hawkes Bay, with an abortion incidence of 2.8% to 9.1%. Antisera to eight different strains of C. fetus fetus were made in rabbits. Strains were then examined using whole cell tube agglutination tests and sensitised Staphylococcal Protein A slide agglutination tests. Heat labile antigens were examined by absorbing antisera with heat-treated bacteria. Two broad serogroups were found, but within-group variation was demonstrated by cross-absorbing antisera. The isolates from one farm were all of a single broad serogroup. Both serogroups were found on the remaining three farms. Evidence for the presence of two major serogroups was also obtained by immobilisation tests and antigen analysis by gel diffusion.     

Brucella suis biotype 1 infection in a dog

Brucella suis biotype 1 was isolated from the semen of a dog with hindlimb weakness and a large, firm, left epididymis. A semen sample was oligospermic, with many neutrophils, the numbers of which decreased in serial sampling. A card agglutination test for B abortus and a rapid slide agglutination test for B canis were positive. The modified 2-mercaptoethanol slide agglutination test for B canis and the agar gel immunodiffusion test, using B canis cell wall antigen, were negative. At necropsy, chronic granulomatous inflammation was found in, and B suis biotype 1 was isolated from, the left epididymis and prostate gland.