K.M. Moriarty

Responses of bovine T cells to fractionated lysate and culture filtrate proteins of Mycobacterium bovis BCG.

Bacterial cell lysates and culture filtrate proteins of Mycobacterium bovis BCG were each separated in a two-dimensional system that yields soluble protein fractions immediately available for probing with T cells. The fractions were used in lymphocyte proliferation assays using blood lymphocytes from cattle immunized with either viable or gamma-irradiated BCG. Cattle immunized with either form of BCG responded similarly to fractionated lysate proteins. Cattle immunized with viable BCG responded to culture filtrate proteins that were not recognized by cattle immunized with dead BCG. Marked heterogeneity of the responses to the culture filtrate proteins was seen.

A possible deficiency of cell-mediated immunity in the opossum, Trichosurus vulpecula, in relation to tuberculosis

Abstract Extract The Australian opossum, Trichosurus vulpecula, is susceptible to experimental infection with both human and bovine strains of Mycobacterium tuberculosis (Bollinger and Bollinger, 1948). Naturally occurring infections of M. bovis have been reported in opossums in New Zealand (Ekdahl et al., 1970). Smith (1972) has described the lesions seen in such infections as being generalized, suppurative and associated with sinus formation and has mentioned the epidemiological significance of the disease in these animals.

A comparison of the polymerase chain reaction with standard laboratory methods for the detection of EHV-1 and EHV-4 in archival tissue samples.

A detection system incorporating the polymerase chain reaction was compared with the use of histopathology and virus isolation to determine the presence of equid herpesvirus type 1 or equid herpesvirus type 4 in equine tissues submitted to a diagnostic laboratory. When the polymerase chain reaction was performed, these tissues had been stored for up to 3 years. Thirty-eight tissues representing 14 cases had been stored embedded in paraffin wax. Analysis of these tissues using the PCR gave predictive values of 1.0 and 0.91 for a positive and negative result respectively, and sensitivity and specificity values of 75% and 100% respectively. Fifty-three tissues representing 28 cases had been stored immersed in 10% formalin. Analysis of these tissues gave predictive values of 0.44 and 0.42 for a positive and negative result respectively, and sensitivity and specificity values of 28% and 57% respectively. The poor results obtained with this group of tissues was attributed to contamination of the samples during wax embedding. Viral DNA could not be amplified from older tissues. These results indicate that under appropriate conditions the polymerase chain reaction is reliable when applied to tissues collected for routine diagnosis. However, it is less reliable when samples for analysis are handled collectively. Also, storage of tissues in wax blocks for 14 or more years inhibits later amplification of viral DNA from these tissues. The implications of these results to the application of the polymerase chain reaction to routine laboratory diagnosis are discussed.

A cloned DNA probe for the detection of Mycobacterium paratuberculosis.

DNA extracted from Mycobacterium paratuberculosis, which had been isolated from a cow with clinical Johnes disease, was used to make a gene library in the Escherichia Coli expression vector phage lambda gt11. Plaque-lifts were made from the library onto nitrocellulose membranes. These were screened by differential hybridization using radiolabelled chromosomal DNA from M. paratuberculosis and Mycobacterium phlei. By this method six recombinants that hybridized to M. paratuberculosis but not to M. phlei were identified. Three of these, designated lambda gt-R3, lambda gt-R4 and lambda gt-RS, containing DNA inserts of 2.5,1.5 and 3.7 kilobases (kb), respectively, were chosen for further analysis of their insert specificities. Following restriction with the endonucleases EcoRI and BamHI, the digestion fragments from the three recombinants were transferred to nitrocellulose membranes and probed with radiolabelled DNA from M. paratuberculosis and M. phlei. As expected, M. paratuberculosis DNA hybridized to all the fragments. M. phlei DNA hybridized to both the fragments that were generated from lambda gt-R3, to the single fragment from lambda gt-R4 and to two of the three fragments generated from lambda gt-RS. The fragment with which M. phlei DNA failed to hybridize was 0.45 kb in length. Multiple copies of this fragment were made in the plasmid pGEM-2; the plasmid DNA was then harvested and radiolabelled. Designated PAM-1, the radiolabelled material hybridized to a 3.7 kb fragment of EcoRI-digested M. paratuberculosis and to 2.2 kb fragments of similarly digested M. avium serovars 2 and 3. PAM-1 did not hybridize to DNA from the other four mycobacterial species examined or from Nocardia asteroides. The restriction fragment length polymorphism thus demonstrated distinguishes M. paratuberculosis from M. avium serovars 2 and 3.

An anaemic state in a horse associated with a cold-acting antibody.

Abstract Extract Auto-immune, haemolytic anacmias (AHA) of man (Dacie, 1963) and domestiicated animals (Schalm, 1965; Farrelly et al., 1966; Lapras and Oudar, 1971) are classified either as idiopathic or secondary to an underlying disease process (Pirofsky, 1969). In both categories antibodies active against the indivuals own erythrocytes are formed. These auto-antibodies are of two types being either warm or cold-acting. Warm-acting antibodies are most effective at 37°C, belong to the IgG class of immunoglobulins, and are incomplete in that, generally, they do not cause autohaemagglutination. Coldacting, or cryopathic, antibodies show maximum activitv at 4°C, are of the IgM immunoglobulin class and are capable of effecting autohaemagglutination. The two types of antibodies also differ in their prevalence. In man warm-acting antibodies occur infrequently and are always pathological (Dacie, 1963) while low titres of cold antibodies occur in most normal sera (Finland et al., 1945; Ellenhorn and Weiner, 195...

Cell-mediated immunity in opossums (Trichosurus vulpecula): the responses to purified protein derivative and an unrelated antigen.

Abstract Sir:— Since resistance to tuberculosis is a function of cell mediated immunity,(1) the susceptibility of Australian brush-tailed opossums (Trichosurus vulpecula) to mycobacterial infections(2) (3) suggests a deficiency in this form of their immunity. Alternatively, opossums might be fully competent in their cullular immunity but fail to respond specifically to mycobacterial antigens.

Alpha-naphthyl acetate esterase activity is not a specific marker for ovine T lymphocytes

Alpha-naphthyl acetate esterase (ANAE) activity was demonstrated in ovine lymphocytes harvested from blood on Ficoll-metrizoate gradients. The enzymes specificity for T lymphocytes, identified by immunofluorescent staining of T cell-specific antigens, was assessed. Correlation analysis of the results obtained using unfractionated lymphocytes from 12 sheep showed no correlation between ANAE activity and the expression of T cell antigens (r = 0.22). When lymphocytes from 4 sheep were fractionated on nylon wool columns a mean of only 43.2% of the cells in the non-adherent population were ANAE-positive whereas 94.7% of these cells were identified as T lymphocytes. Blood lymphocytes from 5 animals were separated into 3 fractions using Percoll discontinuous density gradients. No significant relationship was seen between ANAE activity and T cells in Fractions 1 and 3 (r = 0.41 and 0.21). Fraction 2 cells, however, did show a significant positive relationship (r = 0.91) between these two features but the biological significance of this relationship is unknown. It was concluded that ANAE activity is not a specific marker for ovine T lymphocytes.

Failure to Induce Contact Hypersensitivity in the Opossum, Ittrichosurus Vulpecula

Abstract Sir:- Contact hyypersensitivity (CHS; syn: contact allergy, contact dermatitis, contact eczema) is induced when chemicals (haptens) capable of binding covalently to autologous proteins are applied to the skin.(1)The hapten-protein complexes formed in this way are immunogenic and elicit the immune response underlying CHS. CHS is a form of delayed hypersensitivity. Exposure of a primed animal to the sensitising hapten results in erythema and induration at the site of skin contact, the response being maxima1 after 24 hours.

Age-related functional changes in the follicle-associated epithelium of the bursa of Fabricius in Shaver Cockerels

A study of the age-related functions of immunologically important components of the bursa of Fabricius in Shaver cockerels showed that endocytosis of carbon particles by the specialised follicle-associated epithelium was at a high level from hatching until 5 weeks of age and thereafter declined until at 18 weeks it could no longer be detected. The follicle-associated epithelium had marked non-specific esterase activity during the first 15 weeks of life as determined by a standard acid alpha-naphthyl acetate esterase method. The absolute weight of the bursa was at a maximum at 9 to 10 weeks. Involution began before 14 weeks and was complete by 22 weeks. The results indicate that the critical period for the bursa in regard to acquiring immunity from either local vaccination or environmental challenge is likely to be within the first five weeks of life.

Immune defence mechanisms.

Abstract Extract Infectious diseases and neoplasia are major threats to an animals survival. While acquired immunity is essential in resistance to infectious disease its value in providing protection against tumours is debatable. Infectious organisms can be divided arbitrarily into two types: those that tend to live extracellularly and those that live intracellularly. This distinction is appropriate, in an immunological sense, because, with few exceptions, immunity to extracellular organisms, and their products, is mediated by antibodies whereas immunity to intracellular organisms is largely a function of cell mediated immunity (CMI). CMI, in this context, can be defined as antibody-dependent, T-lymphocyte mediated immunity. This definition is important because other forms of “cell mediated” immunity occur. These are either antibody-in-dependent and mediated by cells bearing receptors for the Fc regions of reacted antibodies or else they are antibody-independent and mediated by so-called natural killer c...