J Mast

Development of immunocompetence of broiler chickens.

Subpopulations of T-cells, B-cells, macrophages and ellipsoid-associated reticular cells (EARC) could be demonstrated by immunohistochemical staining early in the development of chicken spleen. However, the typical structures of the spleen, such as the peri-arteriolar lymphoid sheath (PALS) and the ellipsoids with their surrounding ring of macrophages, were only formed around embryonic day (ED) 20. These structures and especially the B-cell compartment, i.e., the peri-ellipsoid lymphoid sheath (PELS) gradually matured during the first week posthatch. Therefore, we analysed at what age broiler chickens could generate a humoral response against the thymus-dependent antigen bovine serum albumin (BSA). Chickens were immunised in ovo (ED16 and ED18) and at 1, 7 and 12 days of age and subsequent BSA-specific immunoglobulin (Ig) M and IgG responses were measured up to 10 days postimmunisation (DPI). No major differences were observed in the relative growth rates, while hatchability was only slightly reduced. Only in chicks immunised on 12 days of age, IgM and IgG responses were high with a normal kinetic pattern. In chicks immunised on 7 days of age, responses were just detectable, but they were absent in chicks immunised in ovo and on the day of hatching (Day 1). In a subsequent experiment, 1-, 7- and 12-day-old chicks were BSA-immunised and Ig responses were measured for a longer period up to the age of 28 days. The IgG response of chicks immunised at 1 day of age was lower and occurred later (from 28 DPI) than the response of chicks immunised at 7 and 14 days of age (from 14 DPI). It was not increased by a booster immunisation on 29 days of age, in contrast to the response of chicks immunised at 7 and 14 days of age. These findings indicate that vaccination at 1 day of age does not activate the B-cell response resulting in antibody production and support the idea that the immune function of the late embryonic and neonatal chickens is not entirely developed due to the incomplete structural organisation of their secondary immune organs.

Characterisation of chicken monocytes, macrophages and interdigitating cells by the monoclonal antibody KUL01.

The distribution, function and ontogeny of the mononuclear phagocyte system of the chicken were characterised using the monoclonal antibody (MAb) KUL01. KUL01 specifically recognises chicken monocytes, macrophages and interdigitating cells, as well as activated microglia cells. Its tissue distribution allowed to discriminate KUL01 from all earlier described MAb, reactive with mononuclear phagocytes. The specificity of KUL01 for mononuclear phagocytes was further confirmed in functional assays: KUL01-positive macrophages in spleen and liver actively took up colloidal carbon, while monocytes and spleen and gut macrophages contained non-specific esterase and acid phosphatase activities characteristic for antigen-processing. Further, it was demonstrated that KUL01-reactive peripheral blood monocytes express MHC-II, but not CD4. In all tissues investigated, the same morphological subtypes of macrophages were detected in chicken at similar localisations as in mammals, indicating a high degree of conservation between the mononuclear phagocyte system of the chicken and of mammals.

Dynamics of immune cell infiltration in the caecal lamina propria of chickens after neonatal infection with a Salmonella enteritidis strain.

Dynamics of leucocyte infiltration and bacterial invasion in the caecal wall were studied after oral infection of 2-day-old chicks with Salmonella enterica ser. Enteritidis. Bacteria invaded the lamina propria of the caecal wall from 12h post-challenge onwards. Bacteriological examination of internal organs (liver, spleen) showed a peak in Salmonella bacteria at 3days post-infection, after which the number of bacteria decreased. Immunohistochemistry revealed macrophages and T-lymphocytes invading the caecal propria mucosae from 24h after challenge onwards, while B-lymphocytes came somewhat later, subsequently organising into follicular aggregates. An early increase in granulocytes was partly masked by the response to natural flora. While the B-lymphocyte and granulocyte populations were maintained, T-lymphocyte and macrophage populations were already reducing by 10days post-challenge. The infiltration of macrophages and T-lymphocytes in the caecal wall, followed by B-lymphocytes, is the result of an inflammatory response, caused by invading bacteria at this site. The structural maturation of the mucosa-associated lymphoid tissues is antigen driven, since B-cells organise in a follicular pattern.

The effect of vaccination with a Salmonella enteritidis aroA mutant on early cellular responses in caecal lamina propria of newly-hatched chickens.

When newly hatched chicks are inoculated with a Salmonella strain, they induce a rapid onset of resistance to intestinal colonization by other Salmonella strains. The exact mechanism of this early colonization-inhibition is not known. To study host-related contributions to this phenomenon, the kinetics of immune cell infiltration in the caecal wall was analyzed during the first 10 days after vaccination of newly hatched chickens with a Salmonella enterica serovar Enteritidis aroA mutant, and infection 1 day later with a virulent S. enterica serovar Enteritidis strain. These data were correlated with bacterial colonization and clearance of the Salmonella Enteritidis challenge strain. Bacteriological data showed that vaccinated animals had a much lower number of challenge bacteria in their organs and caecal contents the first days post-challenge, relative to unvaccinated animals. Immunohistochemical analysis of the caecal lamina propria revealed that heterophils started infiltrating the caecal lamina propria from 12 h post-vaccination. Macrophages and T-lymphocytes started infiltrating from 20 h and B-lymphocytes from 24 h post-vaccination. These data imply that immune cells already colonized the caecal wall at the time of challenge in vaccinated animals. The presence and activity of these cells in the caecal wall shortly after administration of a Salmonella Enteritidis aroA mutant might contribute to the inhibition of colonization of a virulent Salmonella strain, subsequently administered.

Chlamydia psittaci in turkeys: Pathogenesis of infections in avian serovars A, B and D

At 7 days of age, 4 groups, each of twenty specific pathogen free turkeys kept in isolation units were inoculated by aerosol with the Texas Turkey strain (avian Chlamydia psittaci serovar D), strain 92/1293 (avian Chlamydia psittaci serovar D), strain 84/55 (avian Chlamydia psittaci serovar A) or strain 89/1326 (avian Chlamydia psittaci serovar B). A fifth group of 4 specific pathogen free turkeys were sham inoculated controls. At daily intervals for 10 days and then twice weekly up to 34 days post infection, one bird in each group was killed and the target tissues and cells for replication and the sequence of events of serovar A, B and D infections was examined. In these turkeys, the primary site of replication was the respiratory tract. Chlamydial replication could be detected in the respiratory tract on day 1 post inoculation (p.i.) for group A, on day 3 p.i. for group B and on day 1 to 2 p.i. for groups D1 and D2. Subsequently, there was chlamydaemia and localisation in the digestive tract, in one or more parenchymatous organs, in the pericardium and in the conjunctivae. Specific immunoperoxidase staining revealed chamydiae in these organs in epithelial cells and in monomorphonuclear cells in all infected groups. The monomorphonuclear cells were identified as macrophages by double immunofluorescence staining. Chlamydiae were present in the same tissues for serovars A and D, but could not be demonstrated in proventriculus, duodenum, pancreas, ovaries and testes for serovar B. Furthermore, the intensity of replication was similar for all serovars. However, for serovar B in comparison with the other serovars, the bacteria appeared in most tissues 1 to 6 days later and the maximal replication in these tissues occurred 3 to 4 days later.

Enhanced specific antibody response to bovine serum albumin in pigeons due to L-carnitine supplementation.

1. Thirty adult female pigeons (Columba livia domestica) were randomly divided into 3 equal groups; the 1st and 2nd groups were immunised with bovine serum albumin (BSA) at 0 and 20 d, the 2nd group also received 1 g L-carnitine per litre of drinking water from -5 to 25 d post-immunisation (dpi) and the 3rd group, a control group, received neither treatment. 2. Body weights and serum samples were taken at 0, 5, 10, 15, 20, 25, 30 and 35 dpi. 3. Both BSA-specific IgG and IgM responses were enhanced by about 10% by L-carnitine supplementation. 4. L-carnitine supplemented pigeons showed a higher water consumption. Body weight loss during the onset of the immune response showed a slight tendency to be counteracted by L-carnitine supplementation. 5. The impact of L-carnitine on resistance and resilience to an immunological challenge is discussed.

Inclusion body disease in snakes: a review and description of three cases in boa constrictors in Belgium

Inclusion body disease, a fatal disorder in Boidae, is reviewed, and three cases in boa constrictors, the first reported cases in Belgium, are described. The snakes showed nervous signs, and numerous eosinophilic intracytoplasmic inclusions, which are considered to be characteristic of the disease, were found in the liver and pancreas. The disease is suspected to be caused by a retrovirus, but transmission electron microscopic examinations of several tissues from one of the snakes did not reveal particles with a typical retroviral morphology.

CD57, a marker for B-cell activation and splenic ellipsoid-associated reticular cells of the chicken

Abstract We have demonstrated that the ellipsoid-associated reticular cells of chicken spleen express CD57, a marker for B-cell activation. These cells are characterised by their spindle-shaped morphology, tissue distribution and the absence of certain leucocyte-specific markers. They are phagocytotic and possess high endogenous non-specific esterase activity. Previous reports failed to detect CD57 expression on ellipsoid-associated reticular cells, probably because the tissue sections were differently treated before immunohistochemistry. CD57 is also expressed by a small number of T-cells in the spleen and the caecal tonsils. This number is highly variable between individual chickens depending on the activation state of the immune system. Moreover, CD57 is expressed by bursal lymphocytes (90% or more) but not by B-cells of the peripheral blood. More interestingly, we have been able to discriminate and quantify three B-cell populations of the secondary lymphoid organs, i.e. resting B-cells, germinal centre B-cells and plasma cells, based on their expression levels of CD57 and Bu-1 (a pan B-cell marker). Thus, CD57 should be considered as a B-cell activation marker, rather than as a marker for bursal B-cells; it is also a valuable marker for the immunohistochemical study of ellipsoid-associated reticular cells of chicken spleen.

Expression of beta 2 integrins on blood leukocytes of cows with or without bovine leukocyte adhesion deficiency.

Peripheral blood leukocytes of 11 normal cows, 7 cows heterozygous and 2 heifers homozygous for bovine leukocyte adhesion deficiency (BLAD) were analysed by flow cytometry for the intensity of their beta 2 integrin expression (LFA-1(CD11a/CD18), CR3 (CD11b/CD18) and CR4 (CD11c/CD18)). BLAD-homozygotes revealed no or a very weak expression of the beta 2 integrins and had a 10-fold and 4- to 5-fold increase in absolute number of neutrophils and monocytes, respectively, whereas the absolute number of lymphocytes remained normal. The mean fluorescence intensity (MFI) of the beta 2 integrins (CD18) in heterozygous animals was 56 to 90% of this in the normal cows (MFI between 14 and 512). The difference in the expression level was most pronounced for LFA-1 on the small cluster of lymphocytes with the highest MFI for LFA-1. Repeated analysis and phorbol myristate acetate stimulation revealed that the LFA-1 expression on this high-expressing cell population of the peripheral blood allowed a ready identification of BLAD-heterozygotes by flow cytometry.