Studies of Streptococcus suis type 2 infection in pigs

Investigations into the immunology, pathogenesis and epidemiology of Streptococcus suis type 2 infections were carried out in experimental pigs and in naturally-occurring field outbreaks of disease. The capsular polysaccharide from Str. suis type 2 was shown to induce opsonic antibodies in pigs when injected with Freund's incomplete adjuvant, but difficulties encountered in experimental production of the disease prevented a study of their protective effects. Problems with the bactericidal tests led to an investigation of other assays for antibodies against Str. suis type 2, namely, a phagocytic test with pig neutrophils, a mixed reverse passive antiglobulin haemagglutination test and an indirect haemagglutination test. There was evidence that with modifications both the latter tests would be useful. Transmission studies in 39 conventionally-reared and 7 hysterectomy-derived, colostrum-deprived pigs yielded interesting results with regard to the distribution of the organism in relation to the disease process. Tonsil carriage in clinically-healthy pigs was demonstrated after experimental and natural infection. Detectable carrier rates varied between 0 and 59%. The organism was shown to persist in the presence of circulating opsonic antibodies and in pigs on penicillin-medicated feed. Attempts to isolate the organism from the genital tract were unsuccessful. Medicated early weaning and classical SPF techniques applied to infected herds appeared to be effective in producing pigs free from Str. suis type 2 infection.     

Comparison of experimental models for Streptococcus suis infection of conventional pigs

Four different experimental models for Streptococcus suis-induced disease were compared to find a model that closely mimics naturally occurring disease in conventional pigs. Fourteen, 2-week old pigs free of S. suis type 2 were used in 2 experiments. In experiment 1, 3 pigs were inoculated intravenously (IV) and 3 pigs intranasally (IN) with S. suis. Two out of 3 of the IV-inoculated pigs exhibited signs of severe central nervous system disease (CNS) and were euthanized. Streptococcus suis type 2 was isolated from whole blood, joints, and serosal surfaces of both pigs. No clinical signs and no growth of S. suis were detected in the IN-inoculated pigs. In experiment 2, 4 pigs were inoculated IV and another 4 were inoculated IN with the same isolate as in experiment 1. One hour before inoculation the IN-inoculated pigs were given 5 mL of 1% acetic acid intranasally (IN-AA). All the IV-inoculated pigs showed CNS disease and lameness, and 2 of the pigs became severely affected and were euthanized. All the IN-AA inoculated pigs exhibited roughened hair coats and 2 pigs developed severe CNS disease and were euthanized. Streptococcus suis was isolated from the joints and blood of 3 pigs in the IV-inoculated group. Streptococcus suis was isolated from blood of 2 pigs, meninges of 3 pigs, and joints of 1 pig in the IN-AA inoculated group. Natural exposure to S. suis most likely occurs by the intranasal route. The IN-AA model should serve as a good model for S. suis-induced disease, because the natural route of exposure is intranasal and the IN-AA model was effective in inducing disease that mimics what is observed in the field.     

Glasser's Disease of Swine Produced by the Intratracheal Inoculation of Haemophilus suis

The intracheal inoculation of pigs with Haemophilus suis led to the production of Glasser's disease at every attempt without significant pulmonary involvement. Isolation of this organism from the experimental animals was possible only in the acute phase of the disease.

The indirect fluorescent antibody technique when applied to frozen sections of tissues obtained from the experimentally infected pigs at autopsy, revealed a few rod forms but mostly “round bodies” of H. suis in animals from which the organism was isolated, and “round bodies” only in the pigs from which the organism was not isolated.

Attention is drawn to the similarities between the lesions caused by H. suis and Mycoplasma hyorhinis, and to the confusion which may result therefrom. It is stressed that the laboratory diagnosis of these two diseases is complicated by the fact that both agents may not be isolated on the media commonly used in diagnostic laboratories. Both organisms necessitate the use of special media where the clinical and autopsy results indicate polyserositis and arthritis.

     

The infective and immunogenic properties of Salmonella cholerae suis in weaner pigs

Extract

Salmonella cholerae suis (Salmonella suipestifer) is the cause of an infectious disease of weaner pigs which manifests itself as either a chronic enteritis or as an acute septicaemia in which the organism rapidly invades the body tissues and fluids. Although pigs of all ages can contract the disease, experience in New Zealand has shown that young pigs from eight to twelve weeks old are more susceptible. In 1947. experiments were commenced at Wallaceville to examine the possibility of protecting young pigs by inoculation with vaccines prepared from strains of S. cholerae suis isolated in this country. The purpose of this paper is to record and discuss the results of these experiments.     

Antibody response to an autogenous vaccine and serologic profile for Streptococcus suis capsular type 1/2

An autogenous vaccine was developed, using sonicated bacteria, with a strain of Streptococcus suis capsular type 1/2. The objectives of this study were to evaluate the antibody response following vaccination and to assess the changes in antibody levels in pigs from a herd showing clinical signs of S. suis capsular type 1/2 infection in 6- to 8-week-old pigs. An enzyme-linked immunosorbent assay using the vaccine antigen was standardized. Results from a preliminary study involving 2 control and 4 vaccinated 4-week-old pigs indicated that all vaccinated pigs produced antibodies against 2 proteins of 34 and 43 kDa, respectively, and, in 3 out of 4 vaccinated pigs, against the 117-kDa muramidase-released protein. For the serologic profile, groups of 30 pigs from the infected herd were blood sampled at 2, 4, 6, 8, and 10 weeks of age. The lowest antibody level was observed between weeks 6 and 8, presumably corresponding to a decrease in maternal immunity. A marked increase was seen at 10 weeks of age, shortly after the onset of clinical signs in the herd. For the vaccination field trial, newly weaned, one-week-old piglets were divided into 2 groups of 200 piglets each (control and vaccinated); blood samples were collected from 36 piglets in each group at 2-week intervals for 12 weeks. A significant increase (P 0.05) in antibody response was observed 4 weeks following vaccination and the level of antibodies stayed high until the end of the experiment. In the control group, the increase was only observed at 13 weeks of age, probably in response to a natural infection. The response to the vaccine varied considerably among pigs and was attributed, in part, to the levels of maternal antibodies at the time of vaccination. No outbreak of S. suis was observed in the control or vaccinated groups, so the protection conferred by the vaccine could not be evaluated.     

Clinical and histological features of brucellin skin test responses in Brucella suis biovar 2 infected pigs

Current serological tests for swine brucellosis detect antibodies to the Brucella O-polysaccharide (O/PS). However, when infections by bacteria carrying cross-reacting O/PS occur, these tests suffer from false positive serological reactions (FPSR), and the skin test with Brucella soluble protein extracts is the best diagnostic alternative to differentiate true Brucella suis infections from FPSR in pigs. Since this test has been seldom used in B. suis infected swine, the clinical and histological features involved have not been described properly. Here, we describe the clinical and histological events in B. suis biovar 2 infected pigs skin tested with a cytosoluble O/PS free protein extract from rough Brucella abortus Tn5::per mutant. A similar extract from rough Ochrobactrum intermedium was also used for comparative purposes. No relevant differences were evidenced between the homologous and heterologous allergens, and the main clinical feature was an elevated area of the skin showing different induration degrees. Moreover, an important vascular reaction with hyperemia and haemorrhage was produced in most infected sows 24–48 h after inoculation, thus facilitating the clinical interpretation of positive reactions. Histologically, combined immediate (type III) and delayed (type IV) hypersensitivity reactions were identified as the most relevant feature of the inflammatory responses produced.     

Genetic diversity of Streptococcus suis serotypes 2 and 1/2 isolates recovered from carrier pigs in closed herds

The aim of this study was to compare, by randomly amplified polymorphic DNA (RAPD), the diversity of Streptococcus suis serotypes 1/2 and 2 isolates recovered at slaughter houses from the tonsils of clinically healthy pigs. The pigs belonged to herds with or without clinical signs of S. suis disease.

Overall, a low diversity was observed among isolates of serotype 1/2. A representative isolate recovered from a diseased animal presented a relatively high similarity (85%), with most isolates recovered from carrier pigs, from herds either with or without clinical signs of S. suis disease. For serotype 2 isolates, a relatively high degree of heterogeneity was observed in the whole population. Two subpopulations were observed for serotype 2 isolates, which arose from herds with clinical signs. Interestingly, the representative isolate coming from the diseased pig was included in a small closed cluster, with 2 isolates recovered from carrier pigs belonging to the same herd. On the other hand, most of the S. suis serotype 2 isolates originating from herds with no history of S. suis disease, were closely related (90% similarity). Furthermore, they presented different RAPD patterns from those originating from animals from the herd presenting S. suis clinical signs due to this serotype. Results suggest that, in the herds studied, clinical manifestations due to serotype 2 are probably related to the virulence of a specific isolate. Conversely, for the herd affected with serotype 1/2, clinical manifestations of the disease were more likely to be the result of inherent herd factors than the virulence of the specific isolate.

     

Immune and inflammatory responses in pigs infected with Trichuris suis and Oesophagostomum dentatum

The aim of the present study was to investigate parasite induced immune responses in pigs co-infected with Trichuris suis and Oesophagostomum dentatum as compared to mono-species infected pigs. T. suis is known to elicit a strong immune response leading to rapid expulsion, and a strong antagonistic effect on O. dentatum populations has been observed in co-infected pigs. Forty-eight helminth naïve pigs were allocated into 4 groups in a 2-factorial design. Two groups were trickle inoculated with either 10 T. suis eggs/kg/day (Group T) or 20 O. dentatum L3/kg/day (Group O). Group OT was infected with same levels of both T. suis and O. dentatum (Group OT) and Group C remained uninfected. In each group, six pigs were necropsied after 35 days and the remaining pigs after 71 days. Parasite E/S-antigen specific serum antibodies were quantified by an in-direct ELISA. qPCR was used to measure the expression of immune function related genes in the mucosa of proximal colon and the draining lymph node. Highly significant interactions were identified for O. dentatum specific IgG1 (p < 0.0001) and IgG2 (p < 0.0006) antibodies with a remarkable 2-fold higher antibody response in group OT pigs as compared to group O. These findings indicated that T. suis enhanced the antibody response against O. dentatum in Group OT. The gene expression data confirmed a strong Type 2 response to T. suis (e.g. marked increase in IL-13, ARG1 and CCL11) and clearly weaker in amplitude and/or delayed onset response to O. dentatum in the single infected group. Interactions were found between the two nematodes with regard to several cytokines, e.g. the increase in IL-13 observed in Group T was absent in Group OT (p = 0.06, proximal colon mucosa, 35 and 71 p.i.). Some of these immune response-related interactions may support, or even partially explain, the observed interactions between the two worm populations in co-infected pigs.     

Actinobacillus suis septicemia in mature swine: two outbreaks resembling erysipelas

In the winter of 1987/88 a previously unrecognized septicemic disease syndrome — actinobacillosis in mature sows and gilts — was diagnosed in two minimal-disease swine herds in southwestern Ontario. In herd 1, 34 sows, 2 boars, 13 feeder pigs, and 30 suckling pigs were affected; 11 sows, 2 feeders, and 18 suckling pigs died. In herd 2, 13 sows and 1 feeder pig were affected; 1 sow and 1 feeder pig died. The disease was manifested by moderate fever (39-40.5°C), round or rhomboid erythematous skin lesions, and inappetence. Sudden deaths without previous clinical signs were frequent. Histologically, coccobacillary thromboemboli in superficial and deep dermal vessels were associated with infarcted dermal and epidermal tissues. The causative organism, Actinobacillus suis, was isolated from the affected pigs. Treatment with commonly used antibacterial drugs was effective.

In many respects, the disease resembled acute swine erysipelas and presented diagnostic problems for this reason.

     

The isolation of Streptococcus suis types 1 and 2 from pigs in New Zealand

Extract

Madam:– I wish to advise of the recent isolation of Streptococcus suis 1 and 2 from pigs in New Zealand. Type 1 was isolated from brain and joint swabs from a three-week-old piglet. The piglet had meningitis, was laterally recumbent, paddling and had arthritis with grossly swollen joints. Type 2 was isolated from tonsils collected from apparently normal bacon pigs slaughtered at a meat works. Isolates were confirmed as either Streptococcus suis type 1 or 2 on the basis of biochemical characteristics⁽⁴⁾ and Lancefield precipitin tests using Sand R antisera as originally used by De Moor.⁽²⁾     

Streptococcus suis type 2 infections in pigs in the Netherlands (part two)

Summary

Since 1983 some pig breeding and fattening farms in the Netherlands have been faced with a considerable mortality in pigs due to Streptococcus suis type 2 infections. The most predominant clinical feature of S. suis type 2 infection is meningitis, although sudden deaths often occur. It was noted that some affected farms had imported breeding stock from the United Kingdom.

Tonsils of slaughter pigs were collected from herds with and without a history of S. suis type 2 infections. Bacteriological examination was done by using an elective‐selective medium. No significant difference was found in carrier rates of S. suis type 2 between clinically healthy and affected herds (38% vs. 45%).

A cohort study was carried out by regular bacteriological examination of tonsil biopsies on a farm with a high incidence of streptococcal meningitis. Twenty‐seven percent of the pigs were carriers of S. suis type 2 at nine weeks of age. Possible methods for disease control are discussed.     

Streptococcus suis infections in pigs in the Netherlands (Part I)

Summary

Data are presented on the incidence of various streptococcal infections in pigs in the Netherlands. 314 Strains isolated in the course of routine post‐mortem diagnosis were examined. The most frequently occurring streptococcus was S. subacidus (bio) type II which was isolated in 31.2% of the cases.

S. suis type 2 (Serogroup R) and S. equisimilis (Serogroup C) constituted 16.2% and 13.7% of the isolates respectively. Besides meningitis, endocarditis and polyserositis S. suis type 2 infections may frequently be associated with pneumonia (42%).

The biochemical profiles of the various S. suis and S. subacidus (bio) types are presented. The profile of both species is almost identical. It seems justified to use the name S. suis for strains with this characteristic profile and to abandon the name S. subacidus. Haemolysis does not appear to be a suitable characteristic to screen for S. subacidus/S.suis types. In comparing three serological methods for typing S. suis type 2, gel precipitation using Fuller's extract and slide agglutination give an almost 100% correlation. These two methods are recommended for serotyping.     

Actinobacillus suis infection in pigs in southwestern Ontario

Actinobacillus suis was isolated from tissues of 39 pigs, 2 porcine lungs, and 1 uterine swab submitted for diagnostic evaluation from 24 farms in southwestern Ontario between 1985 and 1988. These isolates represented a gradually increasing incidence of herd outbreaks caused by A. suis in southwestern Ontario. The outbreaks were typified by sudden death in suckling or recently weaned pigs; 87% of the affected pigs examined at the laboratory were between two and 28 days old. Petechial to ecchymotic hemorrhages in the thoracic and abdominal organs accompanied by serofibrinous exudates in both cavities were the most common gross lesions. The lesions were characterized histologically by bacterial thromboembolism and necrosis randomly scattered in thoracic and abdominal organs. Occasionally, bacterial thromboemboli were surrounded by centrifugally radiating, eosinophilic, club-like colonies. Diffuse necrohemorrhagic myocarditis that was more severe in the atria, and diffuse subacute meningoencephalitis, were less frequent but distinctive lesions. Multiple litters were affected in most herd outbreaks, and mortality often approached 50% in affected litters. Although the A. suis organism was susceptible to nearly every antibiotic against which it was tested, the suddenness of herd outbreaks precluded attempts at treatment.     

The Recombinant Nonstructural Polyprotein NS1 of Porcine Parvovirus (PPV) as Diagnostic Antigen in ELISA to Differentiate Infected from Vaccinated Pigs

To differentiate pigs infected with porcine parvovirus (PPV) from those vaccinated with inactivated whole-virus vaccine, an enzyme-linked immunosorbent assay (ELISA) based on detection of a nonstructural polyprotein 1 (NS1) was developed. A threshold of 0.23 optical density units was established and the assayhad high specificity (100), sensitivity (88), accuracy (90) and positive predictive value (100) using haemagglutination inhibition as the standard method. A reproducibility test revealed that the coefficients of variation of sera within-plates and between-run were less than 10%. The assayshowed no cross-reactivitywith antibodies to porcine reproductive respiratorysyndrome virus, pseudorabies virus, foot and mouth disease virus, Actinobacillus pleuropneumoniae, Toxoplasma or Chlamydia. Sera obtained from pigs infected with PPV reacted with recombinant NS1 protein in the ELISA. Sera from pigs vaccinated with inactivated whole virus did not recognize this protein in the ELISA. In contrast, antibodies against PPV whole virus were present in both PPV-infected and vaccinated animals. Serum conversion against NS1 was first detected 10 days after infection and NS1-specific antibodies were detectable up to half a year post infection. In conclusion, the PPV-NS1 ELISA can differentiate PPV-infected versus inactivated PPV-vaccinated pigs and could be applied in disease diagnosis and surveillance.     

Performance of skin tests with allergens from B. melitensis B115 and rough B. abortus mutants for diagnosing swine brucellosis

Swine brucellosis by Brucella suis biovar 2 is an emerging disease whose control is based on serological testing and culling. However, current serological tests detect antibodies to the O-polysaccharide (O/PS) moiety of Brucella smooth lipopolysaccharide (S-LPS), and thus lack specificity when infections by Yersinia enterocolitica O:9 and other gram-negative bacteria carrying cross-reacting O/PS occur. The skin test with the protein-rich brucellin extract obtained from rough B. melitensis B115 is assumed to be specific for discriminating these false positive serological reactions (FPSR). However, B115 strain, although unable to synthesize S-LPS, accumulates O/PS internally, which could cause diagnostic problems. Since the brucellin skin test has been seldom used in pigs and FPSR are common in these animals, we assessed its performance using cytosoluble protein extracts obtained from B. abortus rough mutants in manBcore or per genes (critical for O/PS biosynthesis) and B. melitensis B115. The diagnostic sensitivity and specificity were determined in B. suis biovar 2 culture positive and brucellosis free sows, and apparent prevalence in sows of unknown individual bacteriological and serological status belonging to B. suis biovar 2 naturally infected herds. Moreover, the specificity in discriminating brucellosis from FPSR was assessed in brucellosis free boars showing FPSR. The skin test with B. abortus ΔmanBcore and B. melitensis B115 allergens performed similarly, and the former one resulted in 100% specificity when testing animals showing FPSR in indirect ELISA, Rose Bengal and complement fixation serological tests. We conclude that O/PS-free genetically defined mutants represent an appropriate alternative to obtain Brucella protein extracts for diagnosing swine brucellosis.     

Streptococcus suis Type 2 Infection in Swine in Ontario: A Review of Clinical and Pathological Presentations

Over an 18 month period Streptococcus suis type 2 was isolated in pure or mixed culture in 19 disease outbreaks in pigs. Morbidity and case fatality were variable. Clinical signs were of a nervous or respiratory disease or of death with no premonitory signs. Gross and microscopic findings included one or more of fibrinous polyserositis, fibrinous or hemmorhagic bronchopneumonia, purulent meningitis, myocardial necrosis, focal myocarditis and valvular endocarditis. Brain, cerebrospinal fluid and lung were most reliable sites for isolation of the organism.     

The two-component regulatory system CiaRH contributes to the virulence of Streptococcus suis 2

Two-component regulatory systems (TCSs) are widely distributed among bacteria and enable the organisms to make coordinated changes in gene expression in response to a variety of environmental stimuli. In this work, we constructed a mutant strain of the TCS CiaRH and measured its virulence in vitro and in vivo. Compared with the wild type strain, the mutant strain exhibited a significant decrease in adherence to epithelial cells Hep-2 and PIEC. Furthermore, the deletion of CiaRH not only enhanced the bactericidal activity of RAW264.7 macrophage against Streptococcus suis 2, but also increased blood clearance of S. suis 2 in vivo. More importantly, the mutant was attenuated in vivo in CD1 mice and pigs, with reduced mortality, morbidity and impaired bacterial growth observed in specific organs. These results suggest that the CiaRH is required for S. suis 2 virulence.     

Parasite population dynamics in pigs infected with Trichuris suis and Oesophagostomum dentatum

The aim of the present study was to investigate the population dynamics and potential interactions between Trichuris suis and Oesophagostomum dentatum in experimentally co-infected pigs, by quantification of parasite parameters such as egg excretion, worm recovery and worm location. Forty-eight helminth naïve pigs were allocated into four groups. Group O was inoculated with 20 O. dentatum L₃/kg/day and Group T with 10 T. suis eggs/kg/day. Group OT was inoculated with both 20 O. dentatum L₃/kg/day and 10 T. suis eggs/kg/day, while Group C was kept as an uninfected control group. All inoculations were trickle infections administered twice weekly and were continued until slaughter. Faecal samples were collected from the rectum of all pigs at day 0, and twice weekly from 2 to 9 weeks post first infection (wpi). Six pigs from each group were necropsied 5 wpi and the remaining 6 pigs from each group were necropsied 10 wpi. The faecal egg counts (FEC) and total worm burdens of O. dentatum were dramatically influenced by the presence of T. suis, with significantly lower mean FECs and worm burdens at 5 and 10 wpi compared to single infected pigs. Furthermore, in the presence of T. suis we found that O. dentatum was located more posteriorly in the gut. The changes in the Trichuris population were less prominent, but faecal egg counts, worm counts 5 wpi (57% recovered vs. 39%) and the proportion of infected animals at 10 wpi were higher in Group OT compared to Group T. The location of T. suis was unaffected by the presence of O. dentatum. These results indicate an antagonistic interaction between T. suis and O. dentatum which is dominated by T. suis.     

Cultivation, LD(50) determination and experimental model of Streptococcus suis serotype 2 strain HA9801

The effects of nutritional components and submerged culture conditions on colony-forming unit (CFU) counts by Streptococcus suis serotype 2 strain HA9801 in flask culture was investigated, and the optimal medium and cultivation conditions was confirmed by using a 50 l bioreactor. The LD₅₀ values of HA9801 in pigs before and after fermentation were 1.8 × 10⁷ CFU, which indicated that the virulence of HA9801 was very stable in the fermentation process. In addition, an experimental model that closely mimics naturally occurring disease in conventional pigs was established.