Differential expression of homing receptors and vascular addressins in tonsils and draining lymph nodes: Effect of Brucella infection in sheep

The differential expression of homing receptors (HR) and complementary vascular addressins was studied in T and B lymphocytes from ovine tonsils and draining lymph nodes (LN) in uninfected and Brucella melitensis-infected sheep. In uninfected sheep, CD4+CD25+ T cells expressed proportionally more L-selectin and beta1 integrin than beta7 integrin in pharyngeal and palatine tonsils and in parotid LN (PLN), retropharyngeal LN (RLN) and the peripheral prescapular LN (PSLN). In contrast, memory CD4+CD45RA- T cells expressed an equivalent proportion of the three HR in PLN and PSLN, whereas beta1 and beta7 integrins were proportionally more expressed than L-selectin in pharyngeal tonsil. beta7 integrin was proportionally more expressed than beta1 integrin or L-selectin in palatine tonsils, RLN and the mucosal mesenteric LN (MLN). beta1 integrin was proportionally more expressed in IgG+ and IgA+ cells than beta7 integrin and L-selectin in tonsils, PLN and RLN. The main endothelial addressin expressed on venules in both pharyngeal and palatine tonsils, the PLN and RLN, as well as in the PSLN, was the peripheral PNAd, while in the MLN it was MAdCAM-1. Conjunctival infection by Brucella resulted in an increase of CD4+CD25+ and CD4+CD45RA- T cell subsets, which was associated to modifications of HR expression. CD4+CD45RA- T cells expressed proportionally more beta1 and beta7 integrins than L-selectin in regional PLN and RLN, but also in PSLN. The infection induced an increase of IgG+ and IgA+ cell percentages expressing beta1 integrin in all LN, and also beta7 integrin in the RLN. PNAd continued to be expressed on venules of tonsils and draining LN after Brucella infection, and MAdCAM-1 was also weakly expressed on RLN venules. These results suggest that lymphocyte trafficking through tonsils and draining LN could involve L-selectin/PNAd interactions, as well as beta1 or beta7 integrin, possibly in interaction with VCAM-1 or MAdCAM-1. The homing of antigen-specific lymphocytes in these tissues could be modulated after conjunctival infection with Brucella, which induces the recruitment of lymphocytes that express both beta1 and/or beta7 integrin in regional and more distant LN.     

Differences between the ovine tonsils based on an immunohistochemical quantification of the lymphocyte subpopulations

In sheep, the pharyngeal first defence line against oral and inhaled antigens is organized in six tonsils. Since tonsils are regarded as secondary lymphoid tissue and part of the acquired immune system which is subjected to induction through contact with antigens, an evaluation of the different lymphocyte populations in tonsils is useful to determine a tendency of the specific tonsils to more inductive or more effective immunity. By means of immunohistochemistry, different lymphocyte populations were quantified and localized using a panel of eight antibodies, i.e. anti-CD45, anti-CD21, anti-CD2, anti-CD3, anti-CD4, anti-CD8, anti-WC1 and anti-Ki67. The CD21+ B lymphocytes were localized within the tonsillar lymphoid follicles. The CD2+/CD3+ T lymphocytes were numerous in the interfollicular regions and were aligned underneath and within the epithelium but were also observed at the CD21+ pole of the lymphoid follicles. Near the lingual and tubal tonsils, and the tonsil of the soft palate, the CD45+ cells around the seromucous glands and in the lamina propria were mainly CD3+ T cells. In all tonsils, the WC1+ gamma delta T cells formed a small lymphocyte population which harboured the lamina propria and the interfollicular region. The relative percentages of the different lymphocyte populations of the large palatine and pharyngeal tonsils, which are macroscopically the most developed, were comparable. In contrast, the lingual tonsil was significantly different from the other tonsils not only by its small size and lack of lymphoid follicles, but also by the lymphocyte populations. Based on the lymphocyte populations, the ovine tonsils can be divided in three groups with the tonsil of the soft palate, the tubal and paraepiglottic tonsil forming an intermediate between the palatine and pharyngeal tonsils as true tonsils on the one side, and the lingual tonsil as a scattered lymphocyte aggregation on the other side.     

Regulated expression of peripheral node addressin-positive high endothelial venules controls seeding of B lymphocytes into developing neonatal rabbit appendix

Young rabbit appendix is a homologue of chicken bursa of Fabricius; both are crucial sites for preimmune B-cell repertoire diversification. Here, we report that appendix regulates precursor lymphocyte recruitment for further development by modulating the sites of extravasation. The total area of peripheral node addressin-positive (PNAd(+)) high endothelial venules (HEVs) increased from 1 day to 1 week after birth, remained constant up to 2 weeks and declined to a low and persistent amount by 3 weeks. In normal 1-week and manipulated 5-week appendix where growth of follicles was retarded, PNAd(+) HEVs were present in the basolateral sides of B-cell follicles whereas, in normal 5-wk-appendix these were restricted to T-cell areas. The PNAd was expressed on the lumenal surface of HEVs. The proportions of CD62L(+) B cells in appendix declined from approximately 40% at 3 days to 2-3% at 4 weeks. In lymphocyte transfer experiments, CD62L(+) B cells were preferentially recruited compared with CD62L(-) B cells, anti-PNAd antibody blocked migration of B cells by approximately 50%, and 100 times more B cells were recruited in 1-week compared to 6-week appendix. Thus, a unique spatiotemporal expression pattern of PNAd(+) HEVs is associated with development of B-cell follicles. This regulates migration of blood-borne B-lymphocytes into developing appendix by interacting with CD62L.     

Histological studies on the ontogeny of bovine palatine and pharyngeal tonsil: germinal center formation, IgG, and IgA mRNA expression

The development and distribution of lymphocyte subsets in calf palatine and pharyngeal tonsil were examined. During prenatal development, B cells were distributed in the subepithelial area, and T cells and MHC class II⁺ cells were found in the deep layer of B-cell area, respectively, in both tonsils. At neonatal stage, lymphoid follicle containing a few CD4⁺ cells have been formed in both tonsils. IgG⁺ and IgA⁺ cells were found in the parafollicular and epithelial area. At 3 months old, many germinal centers were recognized in both tonsils. CD4⁺ cells and IgG mRNA expression were detected in light zone of germinal centers. Many IgG, and IgA mRNA expressions also could be detected in the parafollicular and subepithelial area of both tonsils. The data suggest that both tonsils have an important role of local immune defense against invading antigen after birth. The comparison of the histological characteristics of tonsil and Peyer's patch during ontogeny is also discussed.     

Characterization of age-related changes in bovine CD8+ T-cells

Evaluation of the changes induced by immunological interventions requires a baseline against which to compare those changes. The age-related changes in the CD8(+) T-cell population of cattle were studied. The results indicate that CD8(+) T-cells could be divided into γ/δ TCR1(+) and γ/δ TCR1(-) according to their expression of the γ/δ T-cell receptor. As a proportion, the CD8(+) γ/δ TCR1(+) population appears to increase with age. Within the CD8(+)γ/δ TCR1(-) a population of cells expressing a profile of surface molecules previously associated with effector memory T cells (CD45RO(+), CD62L(-), CD27(-), CD45RA(-) and CD28(-)) increases with age. Furthermore, a parallel increase with age in the proportion of CD8(+)CD45RO(+) T cells that express the cytotoxic granule protein perforin was observed. In peripheral tissues, namely lungs, it was found that the majority of CD8(+) T cells present expressed a phenotype indicative of previously primed T cells (high expression of CD45RO and perforin). In contrast, only a small population of memory CD8(+) T cells was present in lymphoid tissue where most of the CD8(+) T cells expressed a na?ve phenotype. In conclusion, in cattle, like in human, CD8(+) T cells that express a phenotype associated with antigen experience accumulate with age that may play a role in immunocompetence as the individual ages.     

Interactions between Streptococcus suis serotype 2 and cells of the myeloid lineage in the palatine tonsil of the pig

Streptococcus suis is a major pathogen in pigs and causes significant morbidity and mortality in herds world-wide. A major problem with S. suis is the presence of asymptomatic carrier animals which can spread the organism within and between herds. The palatine tonsil is one of the main sites where the organism can be recovered, both in infected and carrier animals. The use of multiple-colour immunohistology allowed identification of the cell types associated with bacteria in the tonsils of infected gnotobiotic piglets. Bacteria were never associated with T-cells or B-cells but were always associated with cells of the myeloid lineage. Expression of CD16 and CD163 on these leukocytes suggested an association with mature macrophages in tonsil, which may lead to clearance or control of the micro-organism.     

Flow cytometric analysis of canine umbilical cord blood lymphocytes

Lymphocyte subsets in canine umbilical cord blood were flow cytometrically analyzed and compared with those of the dams' peripheral blood. The proportion of CD3+ T lymphocytes, CD21+CD3- B lymphocytes, and CD3-CD21- non-T non-B lymphocytes in umbilical cord blood was 52.9%, 30.4%, and 16.7%, respectively. T lymphocyte/B lymphocyte ratio was significantly lower in the umbilical cord blood than in the dams' peripheral blood (2.1 +/- 1.4 versus 11.0 +/- 8.1, P < 0.001). In contrast, CD4+ lymphocyte/CD8+ lymphocyte ratio was significantly higher in the umbilical cord blood than in the dams' peripheral blood (7.6 +/- 2.2 versus 1.8 +/- 0.6, P<0.001). These findings clarified the phenotypic characters of canine umbilical cord blood lymphocytes.     

Decreased expression of L-selectin (CD62L) on lymphocytes in enzootic bovine leukaemia

Expression of L-selectin was determined by single- and two-colour immunofluorescence on granulocytes, peripheral blood mononuclear cells (PBMC) and blasts of bovine origin by means of a monoclonal antibody IVA94 which recognizes bovine L-selectin (CD62L). Cells were separated from peripheral blood of healthy cattle and colleagues infected with bovine leukaemia virus (BLV). BLV-infected animals comprised lymphocytotic and non-lymphocytotic cows. L-selectin was expressed on 90-98% of granulocytes in all tested animals. The percentage of PBMC expressing L-selectin was lower in cattle with persistent lymphocytosis than in non-lymphocytotic or BLV-free cattle, and inversely correlated with lymphocyte counts. The ratio of B lymphocytes stained for L-selectin was significantly decreased from 60.2 +/- 1.9% in BLV-free cattle to 43.8 +/- 3.6 and 22.5 +/- 5.7% in non-lymphocytotic and lymphocytotic cattle, respectively. B-lymphocytes stained for L-selectin exhibited about 50% reduction in L-selectin expression in BLV-infected cattle compared with BLV-free cattle, as judged by the mean fluorescence intensity (MFI). The percentage of L-selectin-positive PBMC not bearing surface immunoglobulin M (predominantly T lymphocytes) was comparable in BLV-free and BLV-infected cattle. However, L-selectin expression on T lymphocytes was reduced (about 50%) in BLV-infected cattle, as judged by the MFI. We suppose that BLV infection results in a decreased L-selectin expression on lymphocytes, and accordingly, it may contribute to deregulation of the host immune system.     

Ontogeny of systemic cellular immunity in the neonatal pig: correlation with the development of post-weaning multisystemic wasting syndrome

The aetiology of porcine post-weaning multisystemic wasting syndrome (PMWS) is poorly understood. Porcine circovirus type 2 (PCV-2) is an essential component of the experimental disease model for PMWS: however, evidence from experimental and field studies indicates that additional factors play a critical role in the aetiopathogenesis of PMWS. Current candidates include (1) immune stimulation (for example, via co-infection or vaccination), and (2) a novel infectious agent. A prospective, longitudinal case-control study was designed to investigate molecular triggers in leucocytes of neonatal piglets that may predispose to the development of PMWS. Blood samples were collected weekly from pigs (n=125) within five farms, from 1 week to 8 weeks of age: that is, before the appearance of clinical signs. Four colour flow cytometry was used to investigate changes in subsets of peripheral blood mononuclear cells, using monoclonal antibodies against the following cell associated markers; sIgG, CD3, MHCII dR, CD14, CD4a, CD8a, CD45RC, CD25, SWC3a, SWC8, CD163 and CD45. Sampling and laboratory analysis was supported by monitoring of clinical signs from 1 week to 20 weeks of age, or until disease supervened. At the conclusion of the study, 68 pigs (54%) were classified in Group 1 (no signs of clinical disease), 34 pigs (27%) in Group 2 (signs of clinical disease but not characteristic of PMWS), 17 pigs (14%) in Group 3 (suspect PMWS case) and 5 pigs (4%) in Group 4 (PMWS case). A single case of Porcine Dermatitis and Nephropathy syndrome (PDNS) was also diagnosed. Significant changes with age were demonstrated in clinically normal, neonatal pigs (Group 1), including an increase in B-cells and T-cells, and an increase in the proportion of total T-cells expressing MHCII. Within the T-cell subset, the proportion of CD8(+high) CD4(-) T-cells increased, in addition to the proportion of CD4(+) T-cells co-expressing CD8. Of the factors recorded, farm was found to have a highly significant effect on immune system development in the neonate. Comparison of Groups 1 and 4 cases identified significant differences between pigs which remained normal and those which subsequently developed PMWS. Pigs which went on to develop PMWS had a greater proportion of T-cells expressing MHCII in early life, higher mean intensity of expression of MHCII on T-cells, higher mean intensity of expression of MHCII on B cells and higher expression of CD25 on CD45RC(-) T-cells. These findings suggest that lymphocyte activation may be a key early event in the aetiology of PMWS.     

Flow cytometric analysis of an immunodeficiency disorder affecting juvenile llamas

The present study was undertaken to characterize the immune system of llamas and alpacas and establish the basis for an immunodeficiency disorder affecting juvenile llamas. Flow cytometric (FC) analysis of the immune system with a panel of monoclonal antibodies (mAbs) revealed the immune system of llamas and alpacas is similar in leukocyte subset composition to that in ruminants. Peripheral blood mononuclear cells in adults are comprised of surface immunoglobulin (sIg(+)) B-cells (31%+/-8 S.D.), alphabeta T-cells (27%+/-12 S.D.), WC1(+) gammadelta T-cells (16%+/-11 S.D.), and 5-16% monocytes. In contrast to cattle, goats, and sheep, however, the frequency of WC1(+) gammadelta T-cells is not high in juveniles but similar to the frequency in adults. Also, sIg(+) B-cells are present in high concentration in juveniles (43%+/-11 S.D. ). Expression of major histocompatibility class II molecules on resting T-cells was low or absent. Comparative analysis of peripheral blood lymphocyte composition in normal juvenile llamas and llamas presenting with the signs of the juvenile llama immunodeficiency syndrome (JLIDS) revealed the concentration of B-cells is extremely low (1-5%) in affected animals. The findings suggest JLIDS is attributable to an autosomal recessive genetic defect in the development of B-cells.     

An Immunohistochemical Study of Bovine Palatine and Pharyngeal Tonsils at 21, 60 and 300 Days of Age

An immunohistochemical study was performed on three groups of young cattle (21, 60 and 300 days of age). Tonsils (palatine and pharyngeal) and mucosae (nasal and oral) were removed. Eight monoclonal antibodies (specific for CD3, CD2, CD4, CD8, WC1, cell-surface IgM, cell-surface IgG and MHC class II molecules) and an avidin/biotin complex method on frozen sections were used. The immunological cytoarchitecture of bovine tonsils is similar to that of human tonsils. Nevertheless, these lymphoid tissues are not fully developed during the first weeks of life: T and B dependent areas not well-differentiated, few germinal centres, few intra-epithelial WC1 + T lymphocytes. In contrast, at 2 months, tonsils possess all the elements of a mucosa-associated lymphoid tissue (MALT). Tonsillar or mucosal epithelium is infiltrated by a large number of CD8+, WC1+ T lymphocytes and cells which express MHC class II molecules. Between 21 and 60 days, the number of WC1+ T lymphocytes increase markedly in the tonsillar epithelium. These results accredit the hypothesis that the presence of antigens has an effect on the localization of these lymphocytes at these sites.     

Flow cytometric analysis of colonic and small intestinal mucosal lymphocytes obtained by endoscopic biopsy in the healthy dog

Flow cytometric analysis of the lymphocyte population of the gut could provide useful information on the immune cells present in the gut that would not be easily obtained in tissue sections. However, little is known of the normal lymphocyte population in the canine gut as determined by flow cytometry, which allows for simultaneous staining of multiple cell surface antigens and identification of specific lymphocytic subsets. Therefore, intraepithelial lymphocytes were obtained from biopsies of the healthy canine proximal small intestine and colon taken with an endoscope, and flow cytometric analysis was used to characterize the lymphocyte subsets present. Endoscopic biopsy of the intestine is a minimally invasive technique commonly used for diagnostic purposes. Although CD3+ lymphocytes were the most abundant subset in both colon and small intestine, CD3+/CD8- lymphocytes predominated in the proximal small intestine, whereas CD3+/CD8+ lymphocytes did in the colon. Canine CD8+ intraepithelial lymphocytes were predominantly CD8alphabeta+ in both small intestine and colon. CD4+ intraepithelial lymphocytes were always much less numerous than CD8+ intraepithelial lymphocytes. As in man, a majority of intraepithelial lymphocytes expressed the T-cell receptor, TCRalphabeta, but TCRgammadelta was expressed by a third of intraepithelial T-cells in the proximal small intestine, and approximately 15% of those in the colon. Very few CD21+ lymphocytes were detected in samples of healthy canine colon and small intestinal intraepithelial cells. We have showed that canine intraepithelial lymphocytes are regionally specialized, and that those from the small intestine are unique in comparison to those of other species such as man and rodents due to the large numbers of CD3+/CD8- intraepithelial lymphocytes. This study provides a baseline for comparison with intraepithelial lymphocytes obtained from canine patients with intestinal disease.     

Lymphocyte subsets in porcine tonsillar crypt epithelium

The palatine tonsils are part of the mucosa-associated lymphoid tissue (MALT), strategically located in the oropharynx at the entrance of respiratory and gastrointestinal tracts, and are recognized portals of entry and sites of multiplication and persistence of several pathogens in pigs. As the tonsillar crypt epithelium is in close contact with external environment and the underlying lymphoid tissue, the characterization of the intra-epithelial lymphocyte subpopulations is essential for the understanding of initial steps of pathogenesis of several diseases. In this work we investigated specific lymphocyte subsets in the tonsillar crypt epithelium of 10 adult healthy pigs, using monoclonal antibodies against lymphocyte markers CD3, CD4, CD8, gammadelta T cell receptor and immunoglobulin light-chain in an avidin-biotin immunoperoxidase technique. The crypt epithelium was usually extensively infiltrated by a diverse population of T cells and by B cells. The degree of infiltration of each subset was variable among animals and within individual animals. In the T cell population CD4 cells and gammadelta TCR cells predominated over CD8 cells. These data suggest that the crypt lymphoepithelium is capable of participating in both cellular and humoral immune responses and that gammadelta T cells may play an important role in the defense of this mucosa.     

Cytotoxic T-lymphocyte responses in cattle infected with bovine viral diarrhea virus

A system for a reproducible in vitro restimulation of bovine viral diarrhea virus (BVDV)-specific cytotoxic T-cells (CTL) was developed. Lymphocyte cultures of BVDV-immunised cattle were stimulated with infectious BVDV isolate PT810 and recombinant bovine interleukin-2 for 12 to 25 days. A specific lysis of Concanavalin A-stimulated BVDV-infected autologous target cells was observed, whereas allogeneic BVDV-infected target cells were only marginally lysed as detected by flow cytometry. BVDV-specific lymphocyte transformation was further characterised by the expression of bovine lymphocyte activation antigens and bovine MHC class-II molecules. Secondary stimulation of CTL was influenced by in vitro production of BVDV-specific neutralising antibodies, which were secreted exclusively in BVDV-inoculated lymphocyte cultures of immunised cattle. These results demonstrate the presence of CTL in peripheral blood mononuclear cells (PBMC) of immunised cattle which can kill autologous BVDV-infected antigen-presenting cells after in vitro restimulation.     

Shortening of the bovine tongue according to regulation (EC) 999/2001 is not complying with the current legal definition of specified risk material - a macroscopical and histological preliminary study

The full elimination of all specified risk material (SRM) in food of animal origin is crucial for consumer protection and is of high priority in inner EU trade. Among other tissues, the tonsils of cattle are considered as SRM. The aim of this study was to evaluate whether the 'cut at the back of the tongue just before the tongue bones' required by EC regulation is sufficient to remove tonsils and lymphatic tissue completely. Eight skulls from cattle were collected for the simulation of a vertical cut according to the EC regulation and the detection of the target at the back of the tongue. Further, specimens of the lingual mucosa were cut out from two tongues and examined microscopically. The most caudal of these specimens was from the macroscopically visible part of the lingual tonsil. The most rostral specimen contained the most caudal Papilla vallata. Simulation of the obligatory ventro-dorsal cut yielded hits at varying locations on the dorsal surface of the tongue, sometimes including tissue of the lingual tonsil. Histological examination of the lingual mucosa gave clear evidence that lymphatic tissue resembling the tissue of a tonsil in terms of its histological organization and infiltration of the mucosal epithelium could even be found in areas with no macroscopically visible lingual tonsils.     

Shortening of the Bovine Tongue According to Regulation (EC) 999/2001 is not Complying with the Current Legal Definition of Specified Risk Material – a Macroscopical and Histological Preliminary Study

The full elimination of all specified risk material (SRM) in food of animal origin is crucial for consumer protection and is of high priority in inner EU trade. Among other tissues, the tonsils of cattle are considered as SRM. The aim of this study was to evaluate whether the ‘cut at the back of the tongue just before the tongue bones’ required by EC regulation is sufficient to remove tonsils and lymphatic tissue completely. Eight skulls from cattle were collected for the simulation of a vertical cut according to the EC regulation and the detection of the target at the back of the tongue. Further, specimens of the lingual mucosa were cut out from two tongues and examined microscopically. The most caudal of these specimens was from the macroscopically visible part of the lingual tonsil. The most rostral specimen contained the most caudal Papilla vallata. Simulation of the obligatory ventro‐dorsal cut yielded hits at varying locations on the dorsal surface of the tongue, sometimes including tissue of the lingual tonsil. Histological examination of the lingual mucosa gave clear evidence that lymphatic tissue resembling the tissue of a tonsil in terms of its histological organization and infiltration of the mucosal epithelium could even be found in areas with no macroscopically visible lingual tonsils.     

Signalment and Clinical Complaints Initiating Hospital Admission,Methods of Diagnosis,and Pathological Findings Associated with Bovine Lymphosarcoma (112 Cases)

Background: Lymphosarcoma in adult cattle has multiple manifestations. Objective: To describe the signalment, clinical complaints, and tumor location, and to evaluate utility of diagnostic tests in cattle with lymphosarcoma. Animals: Adult cattle admitted to Cornell University between January 1980 and December 2008 with a definitive diagnosis of lymphosarcoma. Methods: Retrospective case study was conducted with a search of all medical records at Cornell University for cattle diagnosed with lymphosarcoma. Categorical data were analyzed with a Wilcoxon rank‐sum tests. Sensitivities of diagnostic tests were calculated. Results: There were 106 cows and 6 bulls (median age 5 years) examined for anorexia (34%), weight loss (16%), and fever (14%). The sensitivities of antemortem diagnostic tests performed were peripheral lymph node (PLN) wedge biopsy, 100%; surgical exploration and biopsy, 100%; pleurocentesis, 80%; pericardiocentesis, 67%; PLN fine‐needle aspirate, 41%; abdominocentesis, 33%; and cerebral spinal fluid tap, 19%. Median peripheral blood lymphocyte count was 4,900 cells/μL, 10% of cattle were leukemic and 25% had lymphocytosis according to the Bendixen Key. The most frequently identified tumor locations (% of cattle) were the heart (66%), abomasum (61%), uterus (38%), kidney (32%), and epidural space (26%). Conclusions and Clinical Importance: Predilection sites were similar to previously reports but we found a higher incidence of renal tumors and lower incidence of retrobulbar tumors. Knowledge of common clinical presentations, organ involvement, and sensitivities of diagnostic tests will aid informed decisions on the most appropriate tests and interpretation of their results in clinical cases of bovine lymphosarcoma.     

Flow cytometric analysis of ovine peripheral blood lymphocytes.

Leukocyte suspensions were prepared from the peripheral blood of 12 sheep three times at two month intervals beginning at 12 months of age. Monoclonal antibodies and flow cytometric analysis were used to characterize the cells. There were no significant differences over time therefore the data from the three time intervals were pooled. The mean percentages and ranges (minimum to maximum) of the major lymphocyte subtypes were: B-cells (29.6%, 11-50), gamma delta T-cells (36.6%, 22-68), CD4+ T-cells (14.1%, 8-22) and CD8+ T-cells (12.0%, 4-22). Lymphocyte subtype percentages appeared less variable within than between individuals. Two populations of B-cells were noted; one population had more cytoplasm and light scatter characteristics similar to monocytes while a second population of B-cells was more typical of small lymphocytes. The nature of the large B-cells requires further study.     

Occurrence of staphylococcal species in clinically healthy domestic animals

The incidence of staphylococcus species in healthy animals was investigated in young and adult individuals of cattle, in pigs and in domestic fowl, using the method of selective isolation of strains. A total of 6066 samples was examined; 4567 strains were revealed and they included all known species, except Staphylococcus caseolyticus, S. saccharolyticus, S. schleiferi and S. lugdunensis. 3.8% of strains failed to be identified with any species. The test samples were taken from slaughtered animals, only in calves intravital smears of tonsils were examined. The species most frequently isolated from the tonsil tissue in adult cattle were as follows: S. aureus (19.9%), S. saprophyticus (12.4%), S. hyicus (8.8%) and S. hominis (8.6%). The following species were isolated from the lung tissue: S. saprophyticus (28.8%), S. cohnii (10.6%) and S. epidermidis (10.2%); from the mammary gland parenchyma these were the species S. saprophyticus (17.4%), S. xylosus (13.1%) and S. epidermidis (10%). In the tonsil tissue of pigs the percent proportions of S. aureus and S. hyicus were 44.5% and 31.8%, respectively; the most frequent species detected from the lungs were S. aureus (20.6%), S. saprophyticus (12.3%), S. hyicus (12.1%) and S. epidermidis (11.5%). In domestic fowl the most frequently occurring species were S. epidermidis (23.7%), S. gallinarum (16.5%) and S. aureus (15.4%). In calves the incidence of S. xylosus (37%), S. saprophyticus (19.2%) and S. cohnii (15.2%) was found to be highest. The indicence of the other staphylococcus species in all test animals can be expressed by low percentages.