Experimental aerosol transmission of Actinobacillus pleuropneumoniae to pigs.

In order to demonstrate the possible role of aerosol in the transmission of Actinobacillus pleuropneumoniae, an experiment including 18 specific pathogen-free (SPF), 10-week-old piglets, randomly distributed into 2 adjacent units, was carried out. In these facilities, air was forced through absolute filters to prevent any contact with infectious agents. During the first 6 d post inoculation, the 2 units were connected by a rectangular opening and the air circulation was forced by the ventilation system from unit A (inoculated pigs) to unit B (non-inoculated pigs). The A. pleuropneumoniae strain (biovar 1 serovar 9) was isolated in France from an outbreak of porcine pleuropneumonia. Two different infecting doses, 10(7) cfu/animal and 10(8) cfu/animal, were inoculated by intranasal route in 6 pigs of unit A. The infection spread quickly from the inoculated pigs to the non-inoculated pigs. Clinical signs were acute during the 4 d post inoculation: hyperthermia, respiratory distress and, sometimes, death (6 pigs of the unit A and 2 pigs of the unit B). All pigs seroconverted against A. pleuropneumoniae serovar 9 within 2 weeks. Lung lesions were severe: fibrinous pleurisy and lung hemorrhages in the acute stage, pleural adherences and focal pulmonary necrosis in the chronic stage. Actinobacillus pleuropneumoniae was isolated from the tonsils and/or lungs in 16 animals. It could be also isolated from the air of the experimental unit. This study showed that A. pleuropneumoniae was readily transmitted through aerosol over a distance of at least 2.5 m.     

Isolation of Actinobacillus pleuropneumoniae serotype 2 by immunomagnetic separation

In Denmark porcine pleuropneumonia is most frequently caused by Actinobacillus pleuropneumoniae serotype 2 (60%). Isolation of A. pleuropneumoniae from nasal cavities or tonsils from carrier animals is complicated due to the mixed bacterial flora present. An immunomagnetic separation technique (IMS) using immunomagnetic beads (Dynabeads((R))) was developed for isolation of A. pleuropneumoniae serotype 2 from pure cultures and from heterogeneous suspensions. Different coating and washing procedures were evaluated in pure and mixed cultures using polyclonal (PAb) and monoclonal antibodies. The highest reisolation yield was achieved when the beads were coated with 1.5 microg PAb IgG/10(7) beads. After washing the beads for four times 9-24% of the bacteria could be reisolated depending on the amount of IgG attached to the beads and the number of beads used. The recovery was increased to 19-61% when only two washing steps were performed. The IMS was further evaluated using dilutions of A. pleuropneumoniae with added Pasteurella multocida (10(9) CFU/ml). After two washing steps 15% of the A. pleuropneumoniae cells and no P. multocida was reisolated. A detection limit of 10 CFU/ml was found in this heterogeneous suspension. No significant difference was observed when comparing the recovery of A. pleuropneumoniae from pure culture, from mixed cultures and from artificially inoculated tonsils. From 12 pigs inoculated with an aerosol of A. pleuropneumoniae serotype 2 the bacterium could not be detected from the nasal cavity or tonsils by cultivation or PCR 6 weeks later. By using IMS A. pleuropneumoniae serotype 2 could be reisolated from the tonsils of three pigs. The IMS method represents a valuable tool for isolation of A. pleuropneumoniae from tissue samples.     

Effects of different antimicrobial treatments on serum acute phase responses and leucocyte counts in pigs after a primary and a secondary challenge infection with Actinobacillus pleuropneumoniae

The susceptibility to an initial challenge and a re-challenge inoculation with Actinobacillus pleuropneumoniae was analysed in pigs that were treated with antimicrobials of different efficacies following the first exposure to A pleuropneumoniae. In brief, 30 nine-week-old specific pathogen-free pigs were allocated to five groups of six. After acclimatisation, four groups were inoculated with A pleuropneumoniae serotype 2. At the onset of clinical signs, three of the groups of pigs were treated with enrofloxacin, tetracycline or penicillin. A fourth group served as the inoculated control and the fifth group as a control group that had not been inoculated. On day 28, all five groups were re-challenged with the same strain of A pleuropneumoniae serotype 2 as had been used in the first inoculation. No treatments were carried out at this time. The acute phase responses and differential leucocyte counts were monitored in detail after both inoculations. Leucocytosis and acute phase responses in the forms of serum amyloid A, pig-major acute phase protein and haptoglobin were recorded in all of the inoculated groups after the onset of clinical signs following the first inoculation. A porcine mannan-binding lectin-A response was less evident in the pigs. Acute phase responses resembling those of the first inoculation were observed in the pigs that had not previously been inoculated and in the pigs treated with enrofloxacin. Acute phase responses were not recorded in the other three groups, where the pigs had seroconverted to A pleuropneumoniae serotype 2 following the first inoculation.     

Effect of endobronchial challenge with Actinobacillus pleuropneumoniae serotype 10 of pigs vaccinated with bacterins consisting of A. pleuropneumoniae serotype 10 grown under NAD-rich and NAD-restricted conditions

The efficacy of two bacterins containing an Actinobacillus pleuropneumoniae serotype 10 strain was evaluated. The bacterial cells constituting bacterin 1 and 2 were grown under nicotinamide adenine dinucleotide (NAD)-rich (low-adherence capacity to alveolar epithelial cell cultures) and NAD-restricted (high-adherence capacity to alveolar epithelial cell cultures) conditions, respectively. Ten pigs were vaccinated twice with the bacterin 1 and nine pigs with the bacterin 2. Ten control animals were injected twice with a saline solution. Three weeks after the second vaccination, all pigs were endobronchially inoculated with 106.5 colony-forming units (CFU) of an A. pleuropneumoniae serotype 10 strain. In the bacterin 1 and 2 group, three and two pigs died after inoculation, respectively. Only two pigs of the control group survived challenge. Surviving pigs were killed at 7 days after challenge. The percentage of pigs with severe lung lesions (> 10% of the lung affected) was 100% in the control group, 70% in the bacterin 1 group and 22% in the bacterin 2 group. Actinobacillus pleuropneumoniae was isolated from the lungs of all animals. The mean bacterial titres of the caudal lung lobes were 7.0 x 10(6) CFU/g in the control group, 6.3 x 10(5) CFU/g in the bacterin 1 group and 1.3 x 10(6) CFU/g in the bacterin 2 group. It was concluded that both bacterins induced partial protection against severe challenge. Furthermore, there are indications that the bacterin 2, containing A. pleuropneumoniae bacteria grown under conditions resulting in high in vitro adhesin, induced better protection than the bacterin 1.     

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.     

Pathological, ultrastructural, and immunohistochemical changes caused by Lelystad virus in experimentally induced infections of mystery swine disease (synonym: porcine epidemic abortion and respiratory syndrome (PEARS))

The pathogenicity and pathogenesis of Lelystad virus was studied in six 6-day-old SPF piglets. A third passage of the agent was propagated on porcine alveolar macrophages and intranasally inoculated into pigs. Pigs were killed at hours 24, 48, 60, and 72, and on days 6 and 8 after inoculation. From day 2 on pigs developed diffuse interstitial pneumonia with focal areas of catarrhal pneumonia, and from this day on splenic red pulp macrophages were enlarged and vacuolated. Lelystad virus was re-isolated from the lungs of infected pigs from day 2 after inoculation. Lelystad virus antigens were detected by immunohistochemical techniques in bronchiolar epithelium and alveolar cells, and in spleen cells of infected pigs from day 2 after inoculation. Ultrastructural examination of tissues by electron microscopy revealed degenerating alveolar macrophages and epithelial cells in lungs and nasal mucosa, with excessive vacuolation of the endoplasmic reticulum. Although the respiratory tract seems to be the target organ for this virus, macrophages in other organs, such as the spleen, can also be infected. This preference for macrophages may impair immunological defences.     

Comparative virulence of NAD-dependent and NAD-independent Actinobacillus pleuropneumoniae strains.

The virulence of a NAD-independent Actinobacillus pleuropneumoniae serotype 2 strain and NAD-dependent serotype 2, 3 and 9 strains was compared under experimental conditions. Hysterectomy-derived piglets were inoculated endobronchially with 50-500 cfu of these strains. All 23 piglets inoculated with the NAD-dependent strains developed acute disease within 12 hours post inoculation. Twenty-two of these piglets died within 24 hours after the first clinical signs. Three of nine piglets inoculated with the NAD-independent strain did not develop clinical disease. In the other six piglets, disease signs were similar as in the piglets inoculated with the NAD-dependent strains. No differences in clinical disease were observed between colostrum deprived piglets and piglets that obtained colostrum from a SPF sow.     

Effects of oral administration of tilmicosin on pulmonary inflammation in piglets experimentally infected with Actinobacillus pleuropneumoniae

OBJECTIVES: To determine the effects of oral administration of tilmicosin in piglets experimentally infected with Actinobacillus pleuropneumoniae. ANIMALS: Forty 3-week-old specific-pathogen free piglets. PROCEDURES: Piglets were assigned to 1 of 4 groups as follows: 1) uninfected sham-treated control piglets; 2) infected untreated piglets that were intratracheally inoculated with 10(7) CFUs of A pleuropneumoniae; 3) infected treated piglets that were intratracheally inoculated with A pleuropneumoniae and received tilmicosin in feed (400 ppm [microg/g]) for 7 days prior to inoculation; or 4) infected treated piglets that were intratracheally inoculated with A pleuropneumoniae and received chlortetracycline (CTC) in feed (1100 ppm [microg/gl) for 7 days prior to inoculation. Bronchoalveolar lavage (BAL) fluid and lung tissue specimens of piglets for each group were evaluated at 3 or 24 hours after inoculation. For each time point, 4 to 6 piglets/group were studied. RESULTS: Feeding of CTC and tilmicosin decreased bacterial load in lungs of infected piglets. Tilmicosin delivered in feed, but not CTC, enhanced apoptosis in porcine BAL fluid leukocytes. This was associated with a decrease in LTB4 concentrations in BAL fluid of tilmicosin-treated piglets, compared with untreated and CTC-treated piglets, and also with a significant decrease in the number of pulmonary lesions. Tilmicosin inhibited infection-induced increases in rectal temperatures, as measured in untreated and CTC-treated piglets. Pulmonary neutrophil infiltration and prostaglandin E2 concentrations in the BAL fluid were not significantly different among groups at any time. CONCLUSIONS AND CLINICAL RELEVANCE: Oral administration of tilmicosin to infected piglets induces apoptosis in BAL fluid leukocytes and decreases BAL fluid LTB4 concentrations and inflammatory lung lesions.     

Pathological,ultrastructural, and immunohistochemical changes caused by Lelystad virus in experimentally induced infections of mystery swine disease (synonym: Porcine epidemic abortion and respiratory syndrome (PEARS))

Summary

The pathogenicity and pathogenesis of Lelystad virus was studied in six 6‐day‐old SPF piglets. A third passage of the agent was propagated on porcine alveolar macrophages and intranasally inoculated into pigs. Pigs were killed at hours 24, 48, 60, and 72, and on days 6 and 8 after inoculation. From day 2 on pigs developed diffuse interstitial pneumonia with focal areas of catarrhal pneumonia, and from this day on splenic red pulp macrophages were enlarged and vacuolated. Lelystad virus was re‐isolated from the lungs of infected pigs from day 2 after inoculation. Lelystad virus antigens were detected by immunohistochemical techniques in bronchiolar epithelium and alveolar cells, and in spleen cells of infected pigs from day 2 after inoculation. Ultrastructural examination of tissues by electron microscopy revealed degenerating alveolar macrophages and epithelial cells in lungs and nasal mucosa, with excessive vacuolation of the endoplasmic reticulum.

Although the respiratory tract seems to be the target organ for this virus, macrophages in other organs, such as the spleen, can also be infected. This preference for macrophages may impair immunological defences.     

Changes in the leucocyte subpopulations of the palatine tonsillar crypt epithelium of pigs in response to Streptococcus suis type 2 infection

The tonsils are portal of entry and a site of multiplication and persistence for a variety of pathogens, including Streptococcus suis (S. suis), which is a common cause of meningitis, septicemia and arthritis in pigs. Understanding the early changes that occur in the first barrier of the tonsil, i.e. the crypt epithelium, in response to S. suis infection is critical in clarifying the pathogenesis of this disease and for the future development of efficient methods of mucosal vaccination. In this study, we investigated the early changes, from 18 to 72 h, that occur in leucocyte subpopulations of the crypt epithelium of the palatine tonsils of 3-week-old pigs in response to S. suis type 2 infection. Monoclonal antibodies against leucocyte markers CD3, CD4, CD8, gammadelta T cell receptor, lambda-immunoglobulin light-chain, myeloid cells, and major histocompatibility complex class II molecule (MHC-II) were used in an avidin-biotin immunoperoxidase technique. An increase in the number of lambda-immunoglobulin light-chain positive cells (B cell subset) was noticed in crypts of S. suis-infected animals from 18 h after infection onwards. This increase was significant at 18 and 48 h after infection. The number of CD4 and CD8 cells was greater from 18 h onwards, with a significant increase at 24 and 72 h post-infection. No significant difference in numbers of CD3, gammadelta T cell receptor and MHC-II positive cells was detected in the crypts of infected animals compared to controls. Macrophages, neutrophils and crypt epithelial cells stained positively with the myeloid marker, and the area of crypt epithelium positive for this marker was increased in the crypts of infected animals, with a significant difference detected at 24 and 72 h after infection. These results suggest that there is participation of the innate immunity in the early phase of S. suis infection, represented by neutrophils, macrophages and likely epithelial cells, and that there is a potential for the initiation of both humoral and cellular responses against S. suis within the crypt epithelium of the palatine tonsil.     

The pathogenesis of persistent turbinate atrophy induced by toxigenic Pasteurella multocida in pigs

Six one-week-old piglets were pretreated with a 1% acetic acid solution for two days in one or both nostrils. Three piglets were not treated with acetic acid. Three days after treatment all nine piglets were inoculated in both nostrils with a toxigenic type D strain of Pasteurella multocida. Three piglets were killed seven days after inoculation; one died spontaneously 13 days after inoculation and the remaining pigs were killed at approximately 90 kg body weight, i.e., five to six months of age. All acetic acid-treated animals developed severe atrophy of the turbinates in the treated nostrils. Untreated nostrils were normal. The present results showed that toxigenic P. multocida can induce turbinate atrophy that persisted until 90 kg body weight when the lesions were similar to spontaneous atrophic rhinitis in pigs. The turbinate atrophy was not accompanied by inflammatory reaction, atrophy of other bone structures, or lesions in other organs. The experiment showed furthermore that toxigenic P. multocida may be present in the tonsils of control animals without causing turbinate atrophy. A pathogenesis for atrophic rhinitis in pigs is proposed.     

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.     

Effect of oral enrofloxacin and florfenicol on pigs experimentally infected with Actinobacillus pleuropneumoniae serotype 1

This study was carried out to compare the efficacy of two oral anti-microbials as metaphylactic medication to pigs inoculated with Actinobacillus pleuropneumoniae serotype 1. Forty-two pigs with an average weight of 22.64 kg were randomly assigned to three treatment groups: group F was given doses of 40 ppm of florfenicol, group E received 150 ppm of enrofloxacin and group C received no medication. Groups F and E received medicated feed 12 h before being inoculated and for 7 days after inoculation. All the pigs were inoculated by aerosol, with 2 x 10(7) CFU/ml of A. pleuropneumoniae serotype 1 each. The average body temperature was higher in group C than in groups E and F, between 12 and 96 h after inoculation (P < 0.05). No differences were found between groups F and E in respiration pattern, nasal secretion and general condition (P > 0.05): however, differences were found in group C for respiration pattern and general condition (P < 0.05), 12 h after inoculation. There was no mortality in groups F and E, whereas a 50% mortality was recorded in group C during the first 48 h after inoculation (P < 0.05). Necropsies and bacterial cultures were performed 12 days after inoculation. Lesions were observed in five pigs of group F (35.71%) with an average damage of 1.16%; in four pigs of group E (28.57%) with 1.24%; and in 13 animals in group C (92.85%) with 34.5% of affected lung tissue (P < 0.05). The infective agent was cultured from various organs of animals in groups F and C, but not from those in group E.     

The pathogenesis of an enteric infection in pigs,experimentally induced by the coronavirus-like agent,CV 777

Sixteen 2–3-days-old caesarean-derived, colostrum-deprived piglets were each dosed oro-nasally with 2 ml of a bacteria-free filtrate containing 10⁴ pig-infectious-doses of CV 777. The piglets were killed at intervals of 12 to 120 h after infection. The coronvirus-like agent caused a local infection of the intestinal tract which resulted in villous atrophy, malabsorption and diarrhea. The pathogenesis of this infection was similar to that of transmissible gastroenteritis (TGE), a known coronaviral infection of pigs. However, where were some differences. By immunofluorescent staining, CV 777 antigens were not only detected in the epithelial cells covering the small intestinal villi, but also in the cells of the colonic surface epithelium. Occasional fluorescence was also seen in the small intestinal crypt epithelium, but the regenerative capacity of the crypts was not affected. The progress of intestinal epithelial cell infection by CV 777 was much slower than that in TGE, resulting in a longer incubation period and in less drastic epithelial cell destruction. The infection of regenerating cells occurred to a much higher degree during the late stage of a CV 777 infection than has been observed in TGE.     

猪胸膜肺炎调查及地方适用型灭活疫苗的研制

对广东珠江三角洲地区31个猪场450份血清用IHA方法进行猪胸膜肺炎放线杆菌(APP)抗体检查,阳性猪场和阳性血清分别为96.8%和58.4%;分离鉴定了7株APP,血清分型为1、7、3和4型,且以1、7型毒力较强;用平板凝集法对263份APP抗体阳性血清进行抗体反向分型试验,结果1型、7型共占76.4%。确定了APP最佳液体培养条件,培养菌液CFU为2.4×109/mL;研制的APP 1型、7型二价灭活疫苗安全、稳定。兔和仔猪免疫抗体曲线显示,油乳剂疫苗明显优于铝胶疫苗,二次免疫优于一次免疫;APP双价油乳剂灭活疫苗一次免疫仔猪,能100%抵抗APP同源1、7型强毒菌株的攻击,保护期达110 d;田间试验明显提高猪群的生产成绩。     

Transmission of Actinobacillus pleuropneumoniae in pigs under field-like conditions: emphasis on tonsillar colonisation and passively acquired colostral antibodies

The objectives of this study were to elucidate at which age tonsillar colonisation by Actinobacillus pleuropneumoniae occurs in pigs and relate this occurrence to the presence of colostral antibodies to A. pleuropneumoniae. The infection patterns were studied in an isolated cohort of pigs, which consisted of the offspring from five sows originating from a conventional pig herd. The sows were transferred to isolated research facilities before farrowing. A. pleuropneumoniae was detected on the tonsils of all sows. After a nursing period of 3 weeks, the pigs were weaned and reared isolated from other pigs until slaughter. The pigs were examined repeatedly for the presence of A. pleuropneumoniae on the tonsils and for antibodies to A. pleuropneumoniae using bacteriological and serological techniques, respectively.A. pleuropneumoniae was detected in the tonsils of one pig as early as 11 days after birth, showing that A. pleuropneumoniae can be transmitted from sow to offspring during a 3-week nursing period. The cumulative proportion of pigs carrying A. pleuropneumoniae in their tonsils increased significantly between the age of 4-12 weeks. This age period corresponded to the age at which the proportion of pigs with detectable levels of colostral antibodies to the different serotypes of A. pleuropneumoniae was declining. Since these two events take place in the same age period, we expect a possible biological association between the level of the passive immunity and the degree of tonsillar colonisation. The median duration of tonsillar colonisation was estimated to approximately 7-8 weeks.     

Pathophysiologic correlates of acute porcine pleuropneumonia

OBJECTIVE: To develop and evaluate an in vivo model to study early events in the pathogenesis of acute porcine pleuropneumonia. ANIMALS: Thirty-six 6- to 8-week-old pigs. PROCEDURE: Pigs were inoculated intranasally or endotracheally with Actinobacillus pleuropneumoniae; inoculation routes were compared by evaluation of clinical signs, gross and microscopic lung lesions, hematologic changes, serum zinc, iron, and haptoglobin concentrations, and inflammatory cytokines. RESULTS: The 2 inoculation routes resulted in similar findings, although intranasal inoculation caused unilateral gross lung lesions, whereas endotracheal inoculation caused bilateral gross lesions. Clinical signs of disease were observed < 2 hours after endotracheal inoculation and 6 to 8 hours after intranasal inoculation. Total WBC counts did not differ significantly after inoculation by either inoculation route, although band neutrophils increased significantly. The earliest findings associated with A pleuropneumoniae inoculation, irrespective of route, were decreased serum zinc and iron concentrations. Serum haptoglobin concentrations were significantly increased after inoculation. Inoculation induced rapid influx of macrophages into the lung and local induction of proinflammatory cytokines. Northern blot analysis of total RNA from lung tissue indicated that inoculated pigs had increased concentrations of interleukin (IL)-1beta, IL-1alpha, and IL-8; tumor necrosis factor messenger RNA concentration was not increased. CONCLUSIONS: Endotracheal inoculation with A pleuropneumoniae rapidly and consistently induced diffuse bilateral pneumonia; thus, this method may be useful for the study of acute pathophysiologic changes associated with bacterial pneumonia and may provide an experimental model for testing modalities for prevention and treatment of this and other respiratory tract diseases of pigs.     

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.     

The microanatomy of the palatine tonsils of the buffalo (Bos bubalus)

The palatine tonsils play a key role in initiating immune responses against antigens entering the body through the mouth. They are also replication sites of some pathogens. There is no data available about the structure of the palatine tonsils of the Egyptian water buffalo. Therefore, palatine tonsils of 14 clinically healthy buffalo bulls (2–3 years old) were examined macroscopically and microscopically using light, and transmission electron microscopes. The tonsils had an elongated kidney shape with a central invagination (tonsillar fossa) containing a single macroscopic opening leading to a small central cavity (tonsillar sinus). A number of macroscopic crypts originated from this sinus (internal crypts). Besides the tonsillar fossa, also small macroscopic crypts (external crypts) were present. The tonsils were enclosed by a thin connective tissue capsule and septa divided the tonsils into incomplete lobes. Within these encapsulated organs mucous glands were very obvious. Each crypt was highly branched and lined with stratified squamous non-keratinized epithelium. Several lymphoid cells infiltrated between the epithelial cells forming patches of lymphoepithelium. The crypt lumen contained lymphocytes, neutrophils and erythrocytes. Lymph nodules with clear germinal centers extended under the epithelial surface. Diffusely distributed lymphocytes were found in the narrow interfollicular region. High endothelial venules, interdigitating dendritic cells, macrophages and plasma cells were observed among the diffuse lymphocytes. Lymphatics filled with lymphocytes drained the tonsils.     

Campylobacter sputorum subsp mucosalis and Campylobacter hyointestinalis infections in the intestine of gnotobiotic pigs

At 4 days of age, 7 gnotobiotic pigs were orally inoculated with broth cultures of both Campylobacter sputorum subsp mucosalis (CSM) and Campylobacter hyointestinalis (CH). One pig was killed and evaluated each week for 7 weeks. Forty-eight hours after inoculation, CH and CSM were recovered from the feces of the pigs; thereafter, only CH was recovered. Organisms morphologically typical of Campylobacter sp were observed on the mucosal surface and on the crypt epithelial cells of the ileum, cecum, and colon from post-inoculation weeks (PIW) 2 through 7. Bacteria were clustered around the surface opening of goblet cells in pigs at PIW 6 and 7. Crypt epithelial cell proliferation and intracellular bacteria were not seen, except in 1 pig (killed at PIW 7) in which intracellular bacteria were seen only in the cecum. Therefore, CSM and CH did not induce porcine proliferative enteritis in gnotobiotic pigs.