Tonsils Of Pigs Research Articles

Prevalence and characterization of pathogenic Yersinia enterocolitica in pig tonsils from different slaughterhouses

The prevalence of yadA-positive Yersinia enterocolitica was determined in 185 pig tonsils from nine slaughterhouses using both the PCR and culture method. The mean prevalence was 37%, varying from 13% to 45% when both PCR and culture-positive results were included. Of the 52 PCR-positive tonsil samples, 20 were culture-negative, while of the 48 culture-positive, 16 were PCR-negative. Using the culture method, Y. enterocolitica belonging to the bioserotype 4/O:3 was found in 61 tonsils, of which 48 were yadA-positive. Type 4/O:3 was the only pathogenic bioserotype found in this study. Most of the yadA-positive samples (85%) were recovered already after overnight enrichment. A total of 61 isolates, including 13 yadA-negative isolates from different samples, were characterized with PFGE. UsingNotI and XbaI, 20 and 17 PFGE patterns were obtained, respectively. Although the patterns were not identical, most of them played only minor deviations. A total of 26 pulsotypes, defined by combination of the various NotI andXbaI digestion profiles, were observed. Two to eight different pulsotypes were observed in each slaughterhouse, The most common pulsotypes, 1a and 4g, were found in 36% and 20% of the tonsils, respectively and these pulsotypes were widely distributed to most of the slaughterhouses. The pulsotype 1a was identified in eight out of nine slaughterhouses and the pulsotype 4g in seven slaughterhouses.

Contamination of Carcasses, Offals, and the Environment with yadA-Positive Yersinia enterocolitica in a Pig Slaughterhouse

This study was carried out in order to evaluate the contamination of the pig-slaughtering line with pathogenic Yersinia enterocolitica carrying the yadA gene. A total of 292 samples were collected from the slaughterhouse; 131 swab samples from pig carcasses, ears, livers, kidneys, and hearts; 89 swab samples from the environment; and 72 sedimentation samples from the air. All surface samples were studied with both the polymerase chain reaction (PCR) and culture methods. The contamination rate of edible pig offals was high with both methods. Using PCR, the detection rates of yadA-positive Y. enterocolitica for livers, kidneys, and hearts were 38, 86, and 63%, respectively, and using the culture method, the detection rates were 31, 69, and 50%, respectively. Pathogenic Y. enterocolitica was also detected from different environmental sites in the slaughterhouse. Using PCR, 13% of the surface samples from the environment were contaminated with yadA-positive Y. enterocolitica. PCR-positive samples were found on the brisket saw, the hook from which the pluck set (heart, lungs, esophagus, trachea, diaphragm, liver, kidneys, and tongue with tonsils) hang, the knife used for evisceration, the floors in the eviscerating area and the weighing area, the meat-cutting table, the aprons used by trimming workers, the computer used in the meat-inspection area, and the coffeemaker used by slaughterhouse workers. The respective detection rate (6%) was considerably lower when we used the culture method. Pathogenic Y. enterocolitica was isolated from the air in the bleeding area. Bioserotype 4/O:3 was the only pathogenic bioserotype isolated in this study. A total of 113 isolates of type 4/O:3 were characterized with pulsed-field gel electrophoresis using NotI and XbaI digests. By combining these profiles, nine different pulsotypes were obtained, the most common of which (1a) was found in 19 (61%) of 31 samples from different sites. This is the same type that has dominated in pig tonsils, which suggests that tonsils may be the source of Y. enterocolitica contamination in the slaughterhouse. The four pulsotypes (1a, 4g, 6g, and 19q) found on edible offals were the same as those found in tonsils, which supports our hypothesis that tonsils are the contamination source for the liver, heart, and kidneys.

Department of Clinical Studies, The Royal Veterinary and Agricultural University, Copenhagen, Denmark

In this investigation, natural infection with Mycoplasma hyosynoviae was followed in groups of individual pigs in three different herds with regard to occurrence of tonsillar carrier state, clinical arthritis and development of antibodies in serum and in synovial fluid. Antibodies were detected by a polyclonal enzyme-linked immunosorbent assay (ELISA) developed for experimental use. The infection with M. hyosynoviae progressed very differently in the three herds investigated. In one herd, the infection was apparently limited to adult pigs. In a second herd, all pigs became tonsillar carriers of M. hyosynoviae, but no mycoplasma-related arthritis nor any serological response was demonstrated within the growing-finishing period. In the third herd investigated, tonsillar infection was detected in all pigs, clinical cases of M. hyosynoviae arthritis followed and a moderate serological response was observed in some, but not all, pigs. In all three herds, M. hyosynoviae infection was carried in the tonsils of the adult pigs, but it was only occasionally transmitted from sows to piglets. Maternal antibodies were transferred to the piglets and persisted for approximately 8-12 weeks. After weaning, some pigs became infected before 20 weeks of age, while others did not. In the majority of cases, the tonsillar infection was established from 11 weeks of age or older. A latent tonsillar infection was present for a period of several weeks within the group of investigated pigs before cases of generalized infection and arthritis were seen. In some cases, generalization of M. hyosynoviae infection in the blood and in joints was observed in spite of the detection of an active serological response a few weeks earlier. The present work suggests that generalization of the infection and development of arthritis may depend on age, immunity, virulence factors and/or infection pressure; in some herds maybe combined with certain triggering mechanisms such as stress and lowered general resistance.

Early in vivo interactions of Actinobacillus pleuropneumoniae with tonsils of pigs

Twenty gnotobiotic piglets were inoculated with 5 × 10 8 colony forming units of an Actinobacillus pleuropneumoniae biotype 1-serotype 9 strain onto their tonsils. Five other piglets (controls) were inoculated with phosphate-buffered saline solution. Pigs were euthanized at 30 min, 90 min, 180 min, 6 h, 9 h, 12 h or 24 h after inoculation. At necropsy, samples were taken from the tonsils for bacteriological, histological, immuno-histochemical and electron microscopical examination. A. pleuropneumoniae was isolated from tonsils of all the infected pigs, but not from tonsils of the control pigs. Early after inoculation bacteria were mainly associated with the stratified squamous epithelium and detached epithelial cells. Vacuolization and desquamation of the epithelium was observed and many transmigrating neutrophils were present. At later times after inoculation, bacteria were found closely associated with the crypt-walls and with detached cells present in the crypts. A strong neutrophil migration was observed mainly in the deeper parts of the crypts. It is concluded that attachment of A. pleuropneumoniae to tonsillar epithelial cells probably constitutes a first step in establishing bacteria at this body site.

Serotyping and pathogenicity of Erysipelothrix strains isolated from tonsils of slaughter pigs in Thailand.

Erysipelothrix strains were isolated from the tonsils of 46 (15.0%) of 307 apparently healthy slaughter pigs in Thailand during the period of August to September, 1997. A total of 27 of the 46 Erysipelothrix isolates could be classified into 5 serovars but the remaining 19 were untypable in this study. Of the 25 isolates serologically identified as Erysipelothrix rhusiopathiae, 20, 4, and 1 isolates belonged to serovars 2, 12, and 17, respectively. Only 2 isolates from the tonsils belonged to Erysipelothrix tonsillarum and represented either serovar 7 or 10. Although the periods and the districts of the survey were limited, the information obtained in the present investigation demonstrates the presence of a variety of serovars in pigs in Thailand. Of 29 selected isolates belonging to serovars 2, 7, 10, 12, 17, and untypable, only 5 (17.2%) were virulent for both mice and pigs. Five of these virulent isolates belonging to serovars 2 and 12 killed less than 30% of mice immunized with a swine erysipelas bacterin commercially available in Thailand, suggesting that the vaccine elicited a sufficient immunity to these field isolates.

Pathogenicity of local isolates of Erysipelothrix rhusiopathiae serotypes 1, 2, 6, ! 1, 12, and type N in mice and pigs

Six local isolates of Erysipelothrix rhusiopathiae of serotypes 1, 2, 6, 11, 12 ; ?,.;d type N isolated from the tonsil of healthy pigs at Kapuk, Jakarta, slaughter house were examined for their pathgb-;ucity for mice and pigs . Forty-two groups of mice, each contains ten mice and twenty-one pigs divided into seven groups were used as experimental animals. Isolates of serotype 1, 2, 6, 11, 12, and type N were pathogen for rnil.e with LDSO of 10", 102.", 10°6 ,103.° , 10 32 and 105A colony forming unit (C .F .U .), respectively. All (100%) pigs inoculated with isolate of serotype 1 and 33.3% pig inoculated with isolate of serotype 2 were induced general urticarial lesions with depression, anorexia and followed by death. One of three (33.3%) pig inoculated with isolate of serotype 6 was induced generalized urticarial lesions without any other clinical signs. Isolates of serotype 6, 11, 12, and type N were capable of inducing localized urticarial lesions in two out ~f three (66.7%), all (100%), two out of three (66.7° n all (100%) pigs, respectively . All pigs in control uninoculated Key words : Erysipelothrix rhusiopathiae, pathogenicity, mice, pigs

Immunohistologic diagnosis of pseudorabies (Aujeszky's disease) using monoclonal antibodies.

Pseudorabies (PR) (Aujesky’s disease) has been seen in several domestic species, such as pigs, ruminants, dogs, and horses. The laboratory diagnosis of PR has been investigated using various approaches. The most frequent have been immunofluorescence (IF) and immunoperoxidase methods, histopathologic investigation, tissue culture inoculation, and in situ hybridization. Immunoperoxidase staining can also be used to diagnose virus-induced nonsuppurative encephalitis, which is hard to differentiate histopathologically. Eighty monoclonal antibodies (MAbs) against PR virus (PRV) were produced at the Istituto Zooprofilattico to develop a competitive enzyme-linked immunosorbent assay to differentiate antibodies elicited by glycoprotein (G)F-negative strain vaccines from antibodies resulting from infection with wild type viruses and to develop a suitable diagnostic tool for the identification of PRV either directly from infected tissues or from inoculated cell cultures. Eight of the 80 MAbs reacted strongly in the IF test performed on frozen tissue sections (E. Brocchi, personal communication). The reactivity of these MAbs with paraffin-embedded sections was examined to evaluate immunoperoxidase staining for PRV diagnosis in routinely samples. Moreover, a retrospective study was conducted on animals submitted for necropsy because of clinical neurologic signs. The 8 MAbs specific for PRV (Table 1) were tested immunohistochemically with formalin-fixed, paraffin-embedded pig tonsils (for which PRV had been previously detected by IF on cryostatic sections). The following procedures were used for immunohistochemical staining. Five-micrometerthick paraffin-embedded sections were dewaxed in 2 changes of xilene for 15 minute each and hydrated through graded alcohols. All sections were treated with 3% hydrogen peroxide in methanol for 30-minutes at room temperature followed by several washes in Tris-buffered saline (TBS). Nonspecific antibody binding was blocked by 30 minutes of incubation with normal horse serum 1:75 diluted in TBS. As primary antibodies, ascitic fluid TBS dilutions (1:200, 1:400, 1:800, 1:1,000) of each MAb were incubated in 2 different ways, overnight at 4 C and 1 hour at 37 C. After MAb incubation, the sections were washed with TBS and sequentially incubated with biotinylated horse anti-mouse IgG 1: 200 diluted and streptoavidin peroxidase 1500 diluted for 30 minutes each, with rinses in TBS between the incubations. Diaminobenzidine was used as substrate of the peroxidase

Development and evaluation of a selective and indicative medium for isolation of Actinobacillus pleuropneumoniae from tonsils

In order to isolate Actinobacillus pleuropneumoniae from mixed bacterial flora a selective and indicative medium was developed. The optimal concentrations of antibiotics were determined for selective chocolate agar (S-TSA) and selective blood agar (S-MBA) using a set of 25 strains of A. pleuropneumoniae. Following optimization of the media 96 isolates of A. pleuropneumoniae were tested on S-TSA and S-MBA. For isolation of A. pleuropneumoniae from 101 pig tonsils collected at slaughter the S-MBA proved significantly better than both S-TSA and the two non-selective agars tested. Furthermore, contaminating growth by bacteria other than A. pleuropneumoniae was significantly reduced on S-MBA compared to S-TSA and the two non-selective media.

Pathogenesis of swine vesicular disease after exposure of pigs to an infected environment

The pathogenesis of swine vesicular disease (SVD) has been studied following a natural route of infection. In two experiments groups of ten and eight pigs respectively were introduced into a stable contaminated with SVD virus. At various intervals after stable exposure, pigs were killed and the amount of virus was determined in serum, vesicles (if present), spleen, kidney, and in seven lymph glands representing various parts of the body. One day after the pigs were introduced into the stable, five out of eight pigs were viraemic and virus could be isolated from various tissues. At 2 d after introduction, three out of four pigs killed had vesicular lesions on the feet. The tonsils of all pigs killed between 1 to 7 d after introduction into the stable were virologically positive. Four days after introduction 50% of the pigs were serologically positive and at 7 d all pigs had developed an antibody response. This study shows that contact with a SVD virus contaminated environment can be equally as infectious as injection, or direct contact with SVD infected pigs, causing a rapid spread of the disease. Because the tonsil was shown to be highly efficient in trapping and growing circulating virus, we recommend that in addition to serological examination, virus isolation from pig tonsils should be used to study the epidemiology of SVD on farms where the infection is present.

Detection of pseudorabies viral DNA in tonsillar epithelial cells of latently infected pigs

SUMMARY The Rice strain of pseudorabies virus (prv) was intranasally instilled in pigs that were seronegative to prv. Cells were scraped or brushed from tonsillar surfaces biweekly until pigs were euthanatized at either 10 or 16 weeks after infection. The dna extracted from tonsillar cells or parenchyma were subjected to polymerase chain reaction analysis, using either a single set of oligonucleotide primers or nested primers from the prv gII glycoprotein gene. Pigs became seropositive to prv by 3 weeks after infection. The virus was isolated from the trigeminal ganglia and tonsils of pigs that were euthanatized or died 1 to 2 weeks after infection, but not from pigs that were euthanatized 10 or 16 weeks after infection. The prv gene products were consistently detected in trigeminal ganglia and tonsils of all pigs at 1, 10, and 16 weeks after infection, and sporadically in the nasal mucosa, lymph nodes, and lungs of pigs that were euthanatized or died during the first 2 weeks after infection. Cells collected biweekly from tonsillar surfaces were mostly nucleated, squamous epithelial cells with fewer lymphocytes and neutrophils. Polymerase chain reaction analysis of dna extracted from these cells revealed prv dna in a large proportion of the samples when sufficient cells were collected to provide 1 μg of extracted dna for use in the reaction mixtures. A second group of pigs had prv strain 4892 intranasally instilled. The virus was isolated from tonsillar swab specimens until 3 weeks after infection. Tonsillar brushing specimens were collected biweekly until 14 weeks after infection. Some brushing specimens contained all nucleated, squamous epithelial cells, whereas other specimens contained a mixture of epithelial cells and up to 15% neutrophils, lymphocytes, and small mononuclear cells. Results of polymerase chain reaction analysis of dna extracted from tonsillar cells collected 5,11, and 14 weeks after infection were consistently positive for prv gene products. Intact cells collected from tonsillar surfaces were placed in polymerase chain reaction mixtures with nested oligonucleotide primers from the prv gII glycoprotein gene and were subjected to multiple amplification cycles. Afterward, the specificity of the amplified prv gene products was determined by hybridization procedures, using a virus-specific oligonucleotide probe. Most nucleated, squamous epithelial cells stained positive for prv dna, suggesting that these cells were the primary source of prv gene products in tonsillar brushing specimens.

Detection of Yersinia enterocolitica O:3 in faecal samples and tonsil swabs from pigs using IMS and PCR

Dilutions of faecal samples spiked with Yersinia enterocolitica O:3 were analysed using immunomagnetic separation (IMS) followed by PCR. In 10% faecal dilutions with added Y. enterocolitica cells, the limit of detection was 200 cells g-1 faeces. Faecal samples from 38 pigs were analysed by IMS-PCR in parallel with detection and quantification of Y. enterocolitica O:3 using cold pre-enrichment culturing. Of the 15 culture-positive samples, only two were detected with IMS-PCR. These two samples contained 40-400 Y. enterocolitica O:3 cells g-1 faeces; the highest level found in the investigation. This indicated that the low sensitivity of IMS-PCR was due to low amounts of cells in the faecal samples. Swab samples from 195 pig tonsils, taken on a slaughterline were examined using IMS-PCR and culture detection. Of 164 culture-positive samples, 60 were positive with IMS-PCR. In addition, IMS-PCR was positive for three culture-negative samples. Forty-five of the samples were further examined by IMS-PCR after 7-10 d of cold pre-enrichment. All 31 culture-positive samples as well as five culture-negative samples were detected by IMS-PCR. From these data it can be concluded that IMS-PCR can be used to detect Y. enterocolitica O:3 cells after pre-enrichment, but direct detection needs further optimization of the sample preparation procedures.

Pathologic observations of pigs intranasally inoculated with serovar 1, 4 and 5 of Haemophilus parasuis using immunoperoxidase method.

Nineteen, 7- to 13-week-old pigs were inoculated intranasally with different strains of Haemophilus parasuis (serovar 1, 4 and 5), and the pathological lesions induced by each strain were compared. Eleven of thirteen pigs inoculated with either strain Nagasaki (serovar 5) or No. 4 (serovar 1) died between days 1 to 6 after inoculation, and had septicemic lesions, meningitis, or polyserositis. One of six pigs inoculated with strain SW124 (serovar 4) died with polyserositis, another one recovered after illness, and the remaining four pigs remained in good health. Five of the septicemic pigs had thrombi at many organs. Endotoxin was detected in the plasma of 10 pigs in the acute stage of infection. Using the immunoperoxidase technique, H. parasuis antigen was detected in lesions of infected pigs. In the serosal lesions the bacterial antigen was found mainly in the cytoplasm of infiltrating neutrophils and macrophages and appeared as degenerated bacteria and/or lytic bacterial material in dilated phagosomes. Many of the bacteria in the blood vessels of pigs with septicemic lesions were also degenerated. Although H. parasuis was reisolated from nasal secretions of infected pigs, the bacterial antigen could not be detected in the nasal cavities of these pigs. No lesions were observed in the parenchyma of the lung. However, H. parasuis antigen was detected in the tonsil of infected pigs.

Effect of management practices on the Streptococcus suis carrier rate in nursery swine

Summary Management practices on swine farms were analyzed to determine factor(s) associated with high prevalence of pigs that were carriers of Streptococcus suis. Samples were obtained for bacteriologic culture via direct swabbing of palatine tonsils of healthy nursery pigs on 35 farms throughout the United States. Overall, 36.7% of the pigs were determined to be carriers. Isolates of S suis were serotyped, and antimicrobial susceptibility tests were performed by use of Kirby-Bauer techniques. Streptococcus suis types 1 and 2 were most commonly isolated. All isolates were susceptible to enrofloxacin, 97% of the isolates were susceptible to ceftiofur, and 94% were susceptible to ampicillin. However, only 80% of the isolates were susceptible to penicillin, and only 18% were susceptible to tetracycline. Environmental, managerial, nutritional, and health factors were measured on each farm. Excessive temperature fluctuation, high relative humidity, crowding, and an age spread of > 2 weeks between pigs in the same room were the 4 most commonly encountered problems on farms with higher-than-average percentages of carrier pigs. Continuous flow facilities were found on 50% of these farms, and various disease problems, vitamin E/selenium deficiency, inadequate vaccination programs (attributable to the presence of atypical serotypes), and penicillin-resistant strains were found on 6 to 28% of these farms. Overall, 83% (15/18) of farms with higher-than-average percentages of carrier pigs also had a history of clinical S suis disease.

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 hemaglutination 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 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.

Mycoplasma hyosynoviae Isolation from the Upper Respiratory Tract and Tonsils of Pigs

The occurrence of Mycoplasma hyosynoviae at different locations of the upper respiratory tract and tonsils of pigs was investigated in herds with problems of arthritis apparently caused by this microorganism. The isolation of M. hyosynoviae was facilitated by the use of a medium selectively suppressing the growth of Mycoplasma hyorhinis. M. hyosynoviae was cultured from 106 of 178 tonsils of slaughterhouse pigs from 8 herds but could not be isolated from the mucosa of the nasal cavity or the oral-pharyngeal area of 100 living, 10-20 weeks old pigs in 5 of the herds. The value of the selective principles in the medium appears from the circumstance that 86 of the 106 isolates were obtained despite the presence of M. hyorhinis. It is concluded that the tonsil is a reservoir for M. hyosynoviae and is probably the location of choice for an easy demonstration of the presence of this mycoplasma in a pig herd.

Evaluation of an ELISA Test for Detection of Listeria spp.

An ELISA test and two routine culture procedures have been compared in their ability to detect Listeria spp. in food products and swabs of pig tonsils. The culture procedures used were those recommended by the Food and Drug Administration (FDA) and United States Department of Agriculture (USDA). One hundred samples of minced beef, artificially inoculated with Listeria monocytogenes, together with 149 naturally contaminated samples of minced beef, pig tonsil, pig feed and soft, whitemolded, blue-veined cheese were tested by the three methods.The USDA procedure proved to be the most sensitive detection method when samples of artificially contaminated meat containing less than 3 colony forming units (CFU) of L. monocytogenes per gram were examined. In samples where the L. monocytogenes count was above 3 CFU per gram, the ELISA test and the USDA-detection procedure proved to be equally sensitive. When naturally contaminated samples were examined, the sensitivity of the ELISA test was 92% and the specificity 80%. The detection limit of the ELISA test for demonstration of L. monocytogenes in pure cultures was found to be approximately 106 CFU per ml. The corresponding detection limit of the culture procedure was calculated to be approximately 104 CFU per ml.

Immunogenicity in Aujeszky's disease virus structural glycoprotein gVI (gp50) in swine

Aujeszky's disease virus (ADV) envelope glycoprotein gVI (gp50) was purified from virus-infected Vero cells by ion-exchange and immunoaffinity chromatography and its usefulness as a subunit vaccine was evaluated in active and passive immunization studies. Four-week-old piglets were immunized intramuscularly (IM) with purified gVI twice two weeks apart and challenged intranasally (IN) 10 days after the second immunization with 30 LD 50 (10 8PFU) of a virulent strain of ADV. Pigs, vaccinated with 100 μg of purified gVI, produced virus neutralizing antibodies and did not develop clinical signs after challenge exposure. The challenge virus was not isolated from nasal swabs and tonsils of gVI-vaccinated pigs, whereas non-vaccinated control pigs developed illness after challenge exposure with the same virulent ADV strain which was later recovered from their nasal swabs and tonsils. Pregnant sows vaccinated twice with purified gVI (IM) at a three week interval produced virus neutralizing antibodies in colostrum. Four-day-old sucking piglets born of vaccinated sows were passively protected by colostral antibodies against intranasal challenge with a lethal dose of virulent ADV. Sera from gVI-vaccinated pigs were distinguished from experimentally infected swine sera by their differential reactivity in enzyme-linked immunosorbent assay (ELISA) using four major viral glycoproteins (excluding gVI) as antigen purified by the use of lentil-lectin.

Identification of two proteins associated with virulence of Streptococcus suis type 2

The protein profiles of various cell fractions of 180 strains of Streptococcus suis type 2, which were isolated from diseased pigs, from healthy pigs when they were slaughtered, and from human patients, were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting. The isolates from diseased pigs contained two proteins that were absent in most of the isolates from healthy pigs. One of these proteins was a 136-kDa protein that was previously identified as the muramidase-released protein (MRP). This protein was predominantly detected in protoplast supernatants and culture supernatants. The second protein was a 110-kDa protein that was detected only in culture supernatants and therefore was provisionally called extracellular factor (EF). Three phenotypes of S. suis type 2 strains were recognized. Isolates from organs of diseased pigs mainly belonged to the MRP+ EF+ phenotype (77%), while isolates from tonsils of healthy pigs mainly had the MRP- EF- phenotype (86%). Most of the isolates from human patients contained MRP (89%); 74% had the MRP+ EF- phenotype. These findings confirm the results of previous investigations which demonstrated that S. suis type 2 strains differ in virulence. Monoclonal antibodies raised against the 110-kDa EF recognized proteins with higher molecular weights in culture supernatants of all of the strains with the MRP+ EF- phenotype. However, none of the strains with the MRP+ EF+ phenotype produced these high-molecular-weight proteins. Our results demonstrate that MRP and EF are associated with virulence. This suggests that one or both of these proteins are virulence factors that play a role in the pathogenesis of S. suis type 2 infections in pigs and human patients.

Colonization of the Pharyngeal Tonsil and Respiratory Tract of the Gnotobiotic Pig by a Toxigenic Strain of Pasteurella multocida Type D

Seven-day-old gnotobiotic pigs were inoculated intranasally with Pasteurella multocida and euthanatized 2, 5, 9, and 14 days after inoculation. Tissues from the oropharynx and respiratory tract of pigs were cultured quantitatively and analyzed microscopically. Pigs remained afebrile and alert, except one that died of acute fibrinopurulent pneumonia. Pasteurella multocida was isolated in greatest numbers from the pharyngeal tonsils, but only in low numbers from turbinate, trachea, lung, spleen, and liver. Significant histologic changes were limited to the tonsil. Infected pigs developed mild tonsillitis with lymphocytic hyperplasia, and accumulation of cell debris and bacteria in crypts. Capsular antigens of P. multocida, identified on tissue sections with rabbit anti-capsular polysaccharide antibody and immunocytochemical reagents, were confined to the crypt lumen. Ultrastructurally, bacteria were free within crypt material or within phagosomes of macrophages or neutrophils. In a second experiment, 5-day-old pigs were infected with Streptococcus suis type 2, followed by toxigenic Pasteurella multocida at 7 days of age; one pig died of streptococcal septicemia. Pigs developed a mild tonsillitis, and both bacteria were cultured from the tonsillar crypts for up to 14 days after infection. These studies show that a toxigenic strain of Pasteurella multocida, which is a causative agent of atrophic rhinitis, can colonize the tonsil and respiratory tract of gnotobiotic pigs for up to 14 days. In addition, colonization can occur concurrently with Streptococcus suis type 2.

Serological and pathalogenic characterization of Erysipelothrix rhusiopathiae isolates from tonsils of slaughter pigs in Indonesia

Erysipelothrix rhusiopathiae was isolated from tonsils of 245 (35.7%) of 687 apparently healthy slaughter pigs in Indonesia during the period of June 1987 to February 1988. A total of 150 of the 245 E. rhusiopathiae isolated could be serotyped within the 22 recognized serotypes. Serotype 2 was most prevalent with 23.7%, followed by Serotypes 11, 12, 1a, 5 and 6 representing 7.3, 5.3, 4.9, 4.9 and 4.1% of the isolates, respectively. The nine other serotypes (Serotypes 1b, 3, 4, 8, 9, 10, 13, 19 and 22) combined to make up 11.0% of the isolates. Antibiotic-resistant strains were not found. Of 86 selected isolates belonging to various serotypes, 76 (88.4%) were highly virulent for mice (LD 50<10 3.0 colony-forming units). In swine, 40 (51.2%) of 78 isolates induced local or generalized urticarial lesions after intradermal inoculation, and the remaining 38 isolates induced no clinical signs. Of 76 isolates used for challenge in the cross-protection study, 29 (38.2%) killed > 40% of mice immunized with an erysipelas bacterin marketed in Indonesia. A tendency to be refractory to the bacterin-induced immunity was observed in some isolates of various serotypes, but this characteristic was not consistent.