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.     

Progression of Mycoplasma hyosynoviae infection in three pig herds. Development of tonsillar carrier state, arthritis and antibodies in serum and synovial fluid in pigs from birth to slaughter

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.     

Experimental infection with Mycobacterium avium, serotype 2, 2. Oral infection with large doses of M. avium

Twelve pigs were inoculated orally with Mycobacterium avium. The doses used were 0.5, 2 or 10 mg daily for 5 days, or 10, 50 or 180 mg once (1 mg = 37 × 10⁶ viable units). Two pigs were used per dose, 1 of which was sacrificed 3 days, the other 28/31 days after the last inoculation (Table 1).Three days after inoculation, M. avium was found in the tonsils and in the intestinal mucosa of all 6 pigs, and in the mesenteric lymph nodes of 4. Viable unit counts for tonsils and intestinal mucosa were highest in pigs inoculated with 180 mg×1 and 10 mg×5. Histopathologically these pigs showed activation of the lymphoid tissue in the tonsils, Peyer patches and mesenteric lymph nodes. Twenty-eight/31 days after inoculation a spreading of the infection had taken place in all pigs, most often to the liver, less frequently to the spleen and the lungs. The kidneys and the musculature were not infected (Table 4). A correlation was apparent between the size of dose and the number of viable organisms in the tissues. Divided doses gave about 10 times higher viable counts than a single dose with the same total number of organisms (Table 5).No gross lesions were found 28/31 days after inoculation. Microscopic granulomatous lesions were found in the tonsils of 6 pigs, in the intestinal mucosa of 4 pigs, in the mandibular and mesenteric lymph nodes of 6 pigs, in the retropharyngeal lymph nodes of 3 pigs, and less frequently in the parotid and hepatic lymph nodes (Table 3).Five of 6 pigs were weakly sensitive to avian tuberculin PPD, 1000 t.u. per dose, when tested 22/25 days after inoculation; 1 of these pigs cross-reacted to human tuberculin (Table 2).     

THE ISOLATION OF SALMONELLA FROM JEJUNAL AND CAECAL LYMPH NODES OF SLAUGHTERED ANIMALS

One jejunal and one caecal lymph node were sampled from each of 50 cows, 40 yearling cattle, 25 sheep, 20 lambs and 45 pigs after slaughter. Salmonella, Clostridium perfringens and Staphylococcus aureus , all organisms which cause food poisoning in man, were sought by direct plating methods. The samples were also enriched and cultured for Salmonella. Organisms were cultured from 208 (58%) of the 360 lymph nodes; aerobic plate counts yielded up to 25,000 organisms per gram of tissue, although from most infected samples less than 1000 organisms per gram were cultured. Salmonella was isolated directly from 5% of samples, with counts up to 1,500 per gram. After enrichment Salmonella was isolated from nodes taken from 15 cows, 2 yearling cattle, one sheep and 8 pigs. Cl. perfringens was isolated from the caecal nodes of 2 yearling cattle and 2 pigs; S. aureus was not isolated from any sample. It was concluded that mesenteric lymph nodes may be a significant reservoir of Salmonella for transfer to meat and meat products.     

Polyclonal activation of B cells occurs in lymphoid organs from porcine reproductive and respiratory syndrome virus (PRRSV)-infected pigs

Porcine reproductive and respiratory syndrome virus (PRRSV) induces a persistent viral infection associated with an inefficient humoral immune response. A study of lymphoid B cells and specific humoral immune response was performed in blood and several lymphoid organs collected from PRRSV experimentally-infected pigs. Groups of specific pathogen-free (SPF) pigs were infected with the LHVA-93-3 isolate of PRRSV, and blood, tonsils, spleen and mediastinal lymph nodes (MLN) were collected at various times postinfection (p.i.) (3-60 days). Lymphoid cells were isolated, immunolabeled for cytofluorometric determination of B cell percentages, used for counting specific anti-PRRSV antibody secreting B cells by an ELISPOT assay, or cultured for metabolic activity. The presence of anti-PRRSV antibodies in the serum of infected pigs was determined using a commercial ELISA assay. Virus detection was performed in all tissues, including lungs, by virus isolation and RT-PCR. The results show that percentages of B cells increased in tonsils as soon as 3 days until 17 days p.i. in PRRSV-infected pigs while they increased in spleen at 3 days p.i. only, due to an increase of larger Ig(high)-producing B cells. Metabolic activity of lymphoid cells from blood and spleen increased at 3 days p.i. only while lymphoid cells from tonsils and MLN transiently decreased at that time and increased thereafter up to 60 days p.i. Anti-PRRSV antibody-secreting B cells occurred in tonsils after 10 days p.i. and strongly increased up to 60 days p.i. However, specific anti-PRRSV-secreting B cells were detected in blood and spleen after 17 days p.i and in MLN only after 45 days p.i. Specific antibodies were detectable in serum at 10 days p.i., reached the maximum level at 45 days and remained high up to 60 days p.i. Infectious virus was detected in lungs and MLN as soon as 3 days p.i., and remained detectable up to 45 days p.i. in tonsils of one pig while viral RNA was detected in most organs up to 60 days p.i. In vitro experiments revealed that inactivated virus induced a stimulation of lymphoid cells isolated from PRRSV-infected pigs while it was cytotoxic for lymphoid cells from control pigs. Taken together, these results indicate that viral infection induced simultaneously a polyclonal activation of B cells, mainly in tonsils, and an exaggerated and prolonged specific humoral immune response due to persistent viral infection in lymphoid organs.     

Mycobacterium avium abortion in a sow.

A 2-year-old sow aborted her entire near-term litter of 11. Gross and histologic examination of a fetus suggested a tuberculous infection, and a yellow-pigmented Mycobacterium avium serotype 1 was subsequently isolated from the fetal tissue. Efforts to rebreed the sow were unsuccessful. She was anergic to skin tests with purified protein derivative of M. avium on two occasions but had M. avium specific in vitro lymphocyte immunostimulation. Gross granulomatous lesions were found in the liver, kidneys, and endometrium when the sow was necropsied 5 months after the abortion. Histologic examination showed diffuse and focal non-encapsulated granulomas in lymph nodes, tonsils, kidney, liver, spleen, lung, and uterine and vaginal walls. There were a few encapsulated calcified foci in the endometrium. The centers of some granulomas in the tonsils, liver, kidneys, and some lymph nodes were caseated. The yellow-pigmented M. avium was isolated from the reproductive organs and from 11 of 12 other tissues cultured.     

Pathogenesis of encephalomyocarditis experimental infection in young piglets: a potential animal model to study viral myocarditis

The pathogenesis of encephalomyocarditis (EMC) due to the EMC virus (EMCV) was studied in 24 piglets oro-nasally infected with the field isolate B279/95. Two pigs were kept as negative controls and were euthanised at hour 0. The remaining 24 were euthanised every 6 h up to 78-h post infection (hpi). Virus isolation, histological examination and EMCV immunodetection were performed on the spleen, intestine, pancreas, liver, kidneys, heart, lungs, lymph nodes, tonsils and brain. EMCV was isolated at 6-hpi from the intestine and lymph nodes and at 12-hpi from the heart. From 6 to 12-hpi, scattered degenerate myocardiocytes were immunolabelled. Subsequently, myocarditis developed and progressively worsened. Immunopositive reaction in tonsil macrophages, observed in the early stage of infection (6-hpi), suggests that tonsils are the portal of entry, and by mean of wandering macrophages the EMC virus is then distributed through the body. Afterwards, EMCV-B279/95 replicates intensively in the cytoplasm of myocardiocytes and the acute myocarditis is strictly related to the tropism of these cells. Four pigs died spontaneously. In three animals no post mortem lesions or virus were isolated/detected, although all of them showed mild myocarditis. The experimental infection with EMCV B279/95 indicates: (i) the experimental protocol mimics the individual variability observed in natural disease, (ii) tonsils are the portal of entry of infection and the heart is the target organ, (iii) EMCV provides a valuable animal model for comparative studies on progressive viral myocarditis.     

Tonsillar lesions in cattle naturally infected with Mycobacterium bovis

The upper respiratory tract surfaces, the palatine and pharyngeal tonsils and associated lymph nodes of 32 tuberculin reactor cattle were examined pathologically and bacteriologically. Tuberculous lesions were observed histologically in the palatine tonsils of five animals and in both the palatine and pharyngeal tonsils of a sixth. Mycobacterium bovis was cultured from the tonsils of four of these animals and from the palatine or pharyngeal tonsils of a further eight cattle in which no lesions were observed. The upper respiratory tract surfaces of 10 animals were M bovis-positive.     

STUDIES ON THE PATHOGENESIS OF BOVINE EPHEMERAL FEVER

A study of the pathogenesis of bovine ephemeral fever confirmed that the major clinical signs were fever lasting no more than 2 days, with increased respiratory rate, dyspnoea and some degree of lameness. Haematological observations revealed a neutrophilia with a left shift and a lymphopaenia at the time of peak clinical reaction. The net result was a slight leucopaenia on the day after this reaction. The most prominent pathological changes involved the lungs and synovial joints. Pulmonary emphysema and alveolar collapse with bronchiolitis, degenerative changes in synovial membranes and increased synovial fluid were observed. Specific fluorescence indicating the presence of BEF viral antigen could be detected at the time of peak clinical response in individual cells in the lungs, spleen and lymph nodes as well as neutrophils. Before and after the peak fever some fluorescence was seen in cells which appeared to be reticular cells in the lymph nodes. Viral isolation in mice could be made from blood, lungs, spleen and lymph nodes over a period of no more than 3 days. It is postulated that viral growth takes place mainly in the reticuloendothelial cells in the lungs, spleen and lymph nodes and not in vascular endothelium or lymphoid cells.     

Porcine circovirus type 2 (PCV2) distribution and replication in tissues and immune cells in early infected pigs

Replication of porcine circovirus type 2 (PCV2) in pigs, as measured by spliced capsid mRNA (Cap mRNA) and viral DNA, was investigated following experimental infection. Peripheral blood mononuclear cells (PBMCs), and tissue from bronchial lymph nodes (BLN), inguinal lymph nodes (ILN), tonsils, lungs, liver, kidneys, spleen and thymus from infected pigs on different days post-infection (DPI) were assessed. PCV2 replication differed dramatically between tissues from the same infected pig. The virus actively replicated in most tested tissues at 14DPI in association with increased PCV2 associated lesions and PCV2 antigen levels, although no clinical signs correlated with PCV2 associated disease were observed in infected pigs during the course of the study. The PCV2 Cap mRNA was detected only at 13DPI in PBMCs from infected pigs, suggesting replication of the virus in circulating blood is transient and not a major site for PCV2 replication in vivo. Evaluation of the Cap mRNA and viral DNA synthesis in T and B lymphocyte and monocyte populations from PBMCs and BLN at various intervals post-inoculation revealed replication of PCV2 in all cell subpopulations; however, viral replication in B lymphocytes was greater than observed in mononuclear cells isolated from BLN at 14DPI indicating that B lymphocytes may be an important cell population for PCV2 replication. These findings further our understanding of the cell types permissive for PCV2 replication and the pathogenesis of PCV2 infection in vivo.     

Development of specific technics for the prevention of mycoplasma infections in swine

Four experimental series were run in 2 experiments with 44 unweaned piglets to test non-inactivated vaccine from ts-mutant M-60 of Mycoplasma (M.) arginini and from attenuated strains of CH-2 M. hyorhinis, EP-29 M. hyosynoviae, M. suipneumoniae, and B-1 Acholeplasma laidlawii. Similar deviations of clinical and immunological parameters were recorded from piglets inoculated with the above vaccine and infected with pathogenic mycoplasma cultures. These deviations, however, were less strongly pronounced in animals which had been inoculated. Mycoplasma species were re-isolated from bronchial lymph nodes and lungs of 62.5% of inoculated piglets. Lasting residual virulence was recorded from the attenuated mycoplasma strains. That residual virulence had no substantial impact upon growth and development of the piglets under laboratory conditions, throughout the period of observation. The above results are likely to suggest the advisability of further studies for the development of a vaccine from ts-mutants and attenuated strains of pathogens of mycoplasmosis in swine.     

Mycoplasma hyopneumoniae infection in pigs immunosuppressed by thymectomy and treatment with antithymocyte serum

The effect of immunosuppression on Mycoplasma hyopneumoniae infection was evaluated by comparing data from infected, thymectomized, and antithymocyte serum-treated pigs (group 1) with data from infected (group 2) and noninfected (group 3) healthy pigs. After groups 1 and 2 pigs were inoculated intranasally with M hyopneumoniae, mycoplasmas tended to multiply slightly more in the lungs and bronchial lymph nodes of group 1 pigs than that of group 2 pigs. Organisms were also isolated from the spleen of 1 of 3 group 1 pigs. Pneumonia developed in group 2 pigs and was characterized by massive peribronchial, peribronchiolar, and perivascular lymphoid hyperplasia and exudate consisting mainly of polymorphonuclear leukocytes in the alveoli and lumina of the bronchioles and bronchi. In group 1 pigs, perivascular and peribronchiolar cuffings by lymphocytes were less prominent, and the extent of intraluminal exudate was severe and widespread. Bronchial lymph nodes from group 2 pigs had marked hyperplasia of germinal centers and paracortical areas. In group 1 pigs, germinal centers were hyperplastic, whereas in the paracortical areas, depletion of lymphocytes was evident. Seemingly, cell-mediated immune mechanisms are important in the development of pneumonic lesions in enzootic pneumonia of pigs.     

ISOLATION OF MYCOPLASMA HYOPNEUMONIAE AND ITS ASSOCIATION WITH PNEUMONIA OF PIGS IN AUSTRALIA

Nine strains of mycoplasmas were isolated from the lungs of 5 pigs with clinical signs of naturally acquired enzootic pneumonia. Mycoplasma hyopneumoniae was isolated from the lungs of 1 pig and M. hyorhinis from the lungs of 4. An unidentified mycoplasma, which utilized arginine, grew rapidly in broth culture and produced centred colonies on solid medium, was isolated from the lungs of 4 pigs. The pathogenicity of the isolated strain of M. hyopneumoniae was determined by inoculation of pigs from an enzootic pneumonia-free herd. Enzootic pneumonia was produced in the lungs of all 5 pigs inoculated intranasally and intratracheally with broth cultures of the organism isolatied by limit dilution techniques. Enzootic pneumonia was produced in 3 of 6 pigs inoculated intranasally and intratracheally with M. hyopneumoniae purified by the passage of colonies on agar blocks. M. hyopneumoniae was isolated in pure culture from the lungs of all pigs with induced pneumonic lesions.     

Evaluation of serology,bacteriological isolation and polymerase chain reaction for the detection of pigs carrying Actinobacillus pleuropneumoniae in the upper respiratory tract after experimental infection

Pigs, asymptomatically infected with Actinobacillus pleuropneumoniae in their upper respiratory tract, can transmit the infection. Detection of such animals is indispensable to prevent the intake of the disease in a herd. This study was conducted to evaluate bacteriology, polymerase chain reaction (PCR) and serology for the detection of subclinically infected pigs. Pigs were inoculated onto the tonsils with an A. pleuropneumoniae serotype 9 strain (n=12, group 1) or phosphate buffered saline solution (PBSS) (n=5, group 2). To prevent infection of the lungs, pigs of group 1 were treated three times with sodium ceftiofur as an aerosol. A third group (n=5) was inoculated intranasally with the same strain. All animals were euthanized 30 days post-inoculation (dpi). In pigs of group 1, clinical signs were not observed. A small lung lesion was found in only one pig and A. pleuropneumoniae was isolated from this lesion. The bacterium was not isolated from the lungs of animals that did not develop lung lesions. A. pleuropneumoniae was demonstrated in tonsils of 9/12 animals using bacteriological isolation, whereas it was demonstrated in mixed bacterial cultures from tonsils of all 12 animals by PCR. In non-infected animals (group 2), clinical signs were not observed and A. pleuropneumoniae was not demonstrated in any sample. All intranasally infected animals (group 3) developed disease signs and lung lesions. High antibody titers against ApxI, ApxII and heat-stable antigens were detected in animals that developed lung lesions. Antibody titers against these antigens were low or absent in all other pigs. It was concluded that pigs carrying A. pleuropneumoniae in the upper respiratory tract generally do not show measurable antibodies in serum. Therefore, sensitive methods for the detection of the etiological agent such as PCR are required to identify carrier animals, while serological methods are not suitable.     

猪变形杆菌感染症的调查研究

1986年以来,我县某些猪场发生一种以“体温升高、呕吐、腹泻、咳嗽、气喘”为主要症侯的猪病,猪群发病率从28.7％～52.3％不等,病猪平均致死率17.5％。剖检病死猪见肺、肺门淋巴结、气管和小肠呈明显的病理性变化。从17头病猪的肺、肺门淋巴结、脾和肝分离到12株变形杆菌。这些菌能产生不耐热的肠毒素,对小白鼠都有致死作用,回归健康猪能得与自然病例同样的病症,并在发病后4天死亡。康复猪的血清对本菌的凝集价高达1∶80～640,从而确诊为猪的变形杆菌感染症。     

Mycoplasma hyosynoviae in joints with arthritis in abattoir baconers.

The occurrence of Mycoplasma hyosynoviae in synovial fluid of baconers with chronic arthritis was studied at an abattoir. Cultural examination of synovial fluid samples from diseased tarsal joints of 50 animals from 42 herds yielded M. hyosynoviae in 10 cases from 8 herds. Streptococci were found in 6 cases from 6 other herds. M. hyosynoviae antigen was found in 1 of 47 of the samples, and antibody to the mycoplasma was found in 14 of 40 of the samples by ELISA test. The presence of M. hyosynoviae in a joint was usually accompanied by the corresponding antibody. In joints with streptococcal infection antibody to M. hyosynoviae could not be found.     

Pathogenicity of Actinobacillus minor, Actinobacillus indolicus and Actinobacillus porcinus strains for gnotobiotic piglets

The purpose of the study was to evaluate the clinical significance of Actinobacillus minor, Actinobacillus porcinus and Actinobacillus indolicus strains in gnotobiotic piglets. Twenty-two 6-h-old Caesarean-delivered and colostrum-deprived piglets were intranasally and orally inoculated with 2 x 10(6) colony-forming units of an A. minor (group 2; n = 9), A. indolicus (group 3; n = 5), or A. porcinus (group 4; n = 8) strain. Six other piglets were inoculated in the same way with phosphate-buffered saline solution and used as controls (group 1). All pigs were observed for clinical signs and rectal temperatures were taken until euthanasia 7 days after inoculation. At necropsy, conchae, tonsils, lungs, brains, liver, spleen and kidneys were macroscopically examined for lesions and samples were taken for bacteriology. None of the pigs developed fever. Mild ataxia was observed in one pig from group 3 for 2 days. Clinical signs were not observed in the other animals. In none of the animals were macroscopic lesions detected at necropsy. NAD-dependent Pasteurellaceae were not isolated from control animals (group 1). The A. minor, A. indolicus and A. porcinus strains were isolated from the tonsils of one, two and one pigs, respectively. Actinobacillus porcinus was isolated from the brains of the pig with central nervous symptoms and from the conchae of another pig. The inoculation strains were not demonstrated in the other samples. It was concluded that, using these inoculation routes and dose, the A. minor, A. indolicus and A. porcinus strains had low capacity to colonize the upper respiratory tract of gnotobiotic piglets and demonstrated low or no pathogenicity in such animals.     

Occurrence of swine salmonellosis in postweaning multisystemic wasting syndrome (PMWS) affected pigs concurrently infected with porcine reproduction and respiratory syndrome virus (PRRSV)

Fourteen diseased pigs from four farms in which there had been an outbreak of salmonellosis were investigated. Granulomatous inflammation with depletion of lymphocytes was observed in the swollen lymph nodes in these pigs. Antigens to porcine circovirus type 2 (PCV2) were immunolabeled in the lesions along with detection of viral DNA as PCV2 by polymerase chain reaction (PCR). In addition, antigens to porcine reproductive respiratory syndrome virus (PRRSV) were immunodetected in the lungs and Salmonella Choleraesuis was isolated from the affected pigs. The nine salmonellosis affected pigs, five (55.6%) with salmonellosis and PMWS concurrently infected with PRRSV were much higher than those infected with salmonellosis and PMWS (22.2%) or with salmonellosis and PPPRV (22.2%).     

Differences in virulence between two strains of Streptococcus suis type II after experimentally induced infection of newborn germ-free pigs

Fifteen newborn germ-free pigs were inoculated with 2 strains, D-282 and T-15, of Streptococcus suis type II. Some pigs also were preinoculated with Bordetella bronchiseptica, which successfully predisposed them to S suis infection. The 2 streptococcal strains were differentiated by muramidase treatment, which released certain high molecular-weight proteins, termed muramidase-released proteins (MRP), from the cell wall of strain D-282, but not from the cell wall of strain T-15. Only strain D-282 (MRP-positive) induced clinical signs of disease and markedly increased neutrophil numbers in pigs. Streptococci were more frequently isolated from fecal swab specimens obtained from pigs inoculated with strain D-282 (MRP-positive) than from specimens obtained from pigs inoculated with strain T-15 (MRP-negative). Both strains were isolated from nasal swab specimens obtained from all infected pigs. Postmortem examination revealed fibrinopurulent meningitis, polyserositis, and polyarthritis in pigs inoculated with strain D-282; this strain was isolated from the CNS, serosae, visceral organs, heart, and joints. Whereas strains D-282 caused several pathologic changes, strain T-15, isolated from the lungs, caused only pneumonia. Both strains were isolated from the tonsils of all pigs. Virulence differed distinctly between the MRP-positive and the MRP-negative strains.     

Humoral and cellular immune responses of pigs inoculated with Mycoplasma hyopneumoniae

Cellular and humoral immune responses of pigs inoculated with Mycoplasma hyopneumoniae were investigated at postinoculation weeks (PIW) 2, 4, and 6. The response of blood lymphocytes (BL) and bronchial lymph node lymphocytes (LNL) to stimulation by M hyopneumoniae antigens was evaluated by a lymphocyte-stimulation test. Specific antibodies in serum and lung washing samples were assayed by ELISA. Immunoglobulin-positive cells in lungs and bronchial lymph nodes were identified by indirect fluorescent antibody test, using isotype-specific monoclonal antibodies. At PIW 0 to 6, BL from control and M hyopneumoniae-inoculated pigs were stimulated by M hyopneumoniae cells; however, BL from inoculated pigs generally had higher stimulation indices, especially at PIW 6. The response of LNL was influenced by previous exposure to M hyopneumoniae, as indicated by higher stimulation indices (P less than 0.01) of LNL from inoculated pigs killed at PIW 2 and 6. Specific ELISA antibodies to M hyopneumoniae in lung washings from inoculated pigs consisted mainly of IgG and IgA isotypes. Examination of lung sections by indirect immunofluorescence revealed that cells producing IgM and IgA were in controls as well as M hyopneumoniae-inoculated pigs, but IgG-positive cells were only in lungs of inoculated pigs. Resolution of pneumonia appeared to correlate with development of increased sensitization of BL, as well as development of marked increases in immunoglobulins, particularly IgG in lung washings at PIW 6.