A competitive ELISA, using a specific monoclonal antibody, was designed to detect antibodies to Mycoplasma mycoides subsp. mycoides SC, the agent of contagious bovine pleuropneumonia. One monoclonal antibody was found suitable for such a test, `117/5', it does not cross-react with any of the other mycoplasma species tested, furthermore, its binding is inhibited by positive sera. The cutoff, 50% of inhibition, was determined using a set of negative sera from CBPP-free areas. The sensitivity was controlled with sera from artificially infected animals as well as from sera from areas where CBPP is enzootic. In both cases, cELISA compared favorably with CFT. The precocity of detection was similar but cELISA detected more positives and the positive titers seemed to persist longer than in the case of CFT. Lysis of the antigen used to coat the ELISA plates reduced the variability of fixation and improved the repeatability of the test. A field evaluation is now in progress which will determine the true sensitivity and specificity of the test and also check if antibodies are detected after vaccination.
Standard tests for liver damage testing in sheep include glutamate dehydrogenase, aminotransferases and lactate dehydrogenase. The first is a mitochondrial enzyme which is fairly liver specific, while the others lack organ specificity. Glutathione S-transferase (GST, EC 2.5.1.18) is a cytosolic enzyme which has a high activity in hepatocytes and is thus a candidate enzyme marker of liver damage. An automated procedure based on the conjugation of 1-chloro-2,4-dinitrobenzene was adapted on a centrifugal analyser and validated for sheep plasma. The limit of quantification was about 0.5 U l−1 and the coefficient of variation of between-series reproducibility was less than 4%. Reference values did not differ according to breed, sex or age (n = 139; nine males, 100 females, 30 undetermined; 127 Lacaune and 12 Manech sheep; 41 animals 3–9 months of age, 98 animals 2–6 years of age). The distribution was log-normal and the median and geometric means were 5.1 U l−1 and 10.0 U l−1, respectively. The 0.025 and 0.975 quantiles were 2.3 U l−1 and 18.4 U l−1. Plasma GST showed a circadian rhythm with a minimum (about 60% of maximum) from 17:00 to 05:00 h and was not significantly altered by repeated daily fine needle liver biopsies for 5 days. In experimental liver damage by intraruminal administration of 0.20 ml kg−1 BW CCl4, plasma GST was increased 90-fold in the 24th hour, then returned to baseline by day 4. Thus, plasma GST could be an interesting candidate marker for liver damage testing in sheep.
The association of Helicobacter pylori in the stomach, trachea and lungs with the incidence of SIDS, gastric ulcers and cancer may have a counterpart in animals. In field studies of white muscle disease (WMD) and hepatic necrosis in selenium-deficient pigs dying suddenly, veterinarians identified gastric ulcers in 40% of inspected piglets. The lesion was also commonly observed by researchers in experimentally produced vitamin E-selenium deficiency and other researchers suspected that gastric ulcers in swine may be associated with vitamin E-selenium deficiency. Mice preferentially concentrated 75selenium in peritoneal exudative cells (PEC) when 75selenium as selenium selenate was administered by stomach tube to selenium-deficient mice. Selenium concentrated in PECs as glutathione peroxidase (GSHPx). GSHPx-deficient leucocytes in peritoneal exudate failed to kill yeast cells. GSHPx deficiency has also been associated with decreased microbicidal activity of leucocytes in patients with chronic granulomatosis. The selenium-deficient swine were usually growing rapidly in crowded conditions, and, apart from WMD and hepatic necrosis, edema was prominent in the spiral colon, subcutaneous tissues, lungs and submucosa of the stomach. The elevated immunological response in the spleen and lungs of SIDS victims suggests an initial defective microbicidal propensity of the peritoneal exudative cells.
The sites of early (pre-viraemic) localization and multiplication of virus in cattle exposed to foot-and-mouth disease virus by various routes were identified by sequential sampling from the pharynx and by tissue titrations after slaughter. Virus was recovered from thepha rynx of 30 of 37 cattle exposed by direct or indirect contact or to virus aerosols for one or more days before the detection of viraemia. The distribution and amounts of virus in the tissues of 23 cattle killed before the onset of viraemia indicated that the pharyngeal area was the most likely site of initial infection and virus growth.
The rabbit is a popular model in laboratory animal medicine due to its relatively large size and docile nature. Though rabbits are sometimes mistakenly thought of as rodents, they are in their own Order, Lagomorpha. Among the many breeds of rabbits, by far the most popular in research is the New Zealand White rabbit. Research in these rabbits has contributed to advances in cardiology, orthopedics, dentistry, immunology, and more. This chapter outlines the anatomy and physiology of rabbits. Rabbits have thin, delicate skin that is generously covered with both underfur and guard hairs. The rabbit's skin, similar to the rat's, has blood vessels immediately under the dermis. Unlike the rat, however, the fascia superficialis is well differentiated due to the elastic fibers and dense collagen content. Hair grows in waves starting from the ventrum and grows dorsally and caudally. Thick fur covers the feet in place of footpads. The fragile skin can tear easily and must be handled carefully during any manipulation. Rabbits are social, nocturnal animals that, despite over 2000 years of domestication, still have a highly developed prey instinct.
Caseous lymphadenitis (CLA) is a ruminant disease caused by Corynebacterium pseudotuberculosis, a Gram-positive facultative intracellular pathogen. The present work was performed to investigate the biochemical composition, morphology and antimicrobial susceptibility pattern of C. pseudotuberculosis biofilm using Raman spectroscopy, field emission scanning electron microscopy (FESEM) and microplate biofilm assay respectively. Results showed that the 24-h-old biofilm was characterized by Raman spectral peaks at 615 cm−1 (CCC symmetric bend phenyl ring), 668 cm−1 (Valine) and 825 cm−1 (Ring breath Tyr.) whilst the 48-h-old and 72-h-old biofilms were characterized by Raman spectral peaks at 1400 cm−1 (COO sym.), 1450 cm−1 (COO sym.), 1581 cm−1 (Ring breath Trp.), 1650 cm−1 (COO asym.) and 1725 cm−1 (CO str.). Raman spectra also revealed the biochemical heterogeneity in C. pseudotuberculosis biofilm. FESEM images clearly showed the biofilm cells which were surrounded by the extracellular matrix. Treatment with nalidixic acid, streptomycin, tetracyclin, ethylenediaminetetraacetic acid (EDTA) and dimethyl sulfoxide (DMSO) significantly (p < 0.05) inhibited the viability of C. pseudotuberculosis biofilm. The present study suggests that the biochemical composition of C. pseudotuberculosis biofilm may vary across different developmental stages. Meanwhile, nalidixic acid, streptomycin, tetracyclin, EDTA and DMSO may be useful in the treatment of CLA.
Iminodipropionitrile (IDPN) was given to rats on diets with normal copper levels, 5 ppm, and to rats on diets with high copper levels, 100 ppm. The former developed neurological symptoms, namely choreoathetoid head movements, circling and retropulsion, while the latter only showed vague incoordination. Since rats, raised on a copper deficient diet alone, i.e. 0.9 ppm develop the same symptoms, we assume that IDPN chelates copper. The axons of the motor nerve cells show metachromatic degeneration in the rats given IDPN on a diet with normal copper level, but not in the rats on a high copper level.
Cell-mediated immunity has been exploited historically in the diagnosis of mycobacterial diseases through elicitation of a delayed-type hypersensitivity (DTH) reaction following intradermal injection of an antigen. Here we describe the histopathological features of the cutaneous DTH reaction and its association with intestinal pathology and systemic immune responses in sheep with Mycobacterium avium subspecies paratuberculosis (MAP) infection. A mixed mononuclear cellular infiltrate dominated the DTH reaction and was present in perivascular and periadnexal patterns. Multiple multinucleate giant cells were present in the cellular infiltrate in one sheep while plasma cells were an obvious feature in six others. Sheep with paucibacillary intestinal lesions had the greatest degrees of cutaneous induration, more severe cellular infiltration in DTH lesions and high systemic interferon (IFN)-γ production. In contrast, sheep with multibacillary intestinal lesions, and particularly those with dissemination of MAP to extra-intestinal tissues, had minimal cutaneous induration, nil to mild cellular infiltration in DTH lesions and high serum anti-MAP antibody levels. Systemic IFN-γ production generally was augmented following skin sensitization. In general, the gross and histopathological features of the cutaneous DTH response matched the stage of paratuberculosis reflected by intestinal pathology and systemic measures of humoral and cellular immunity.
An inactivated iron regulating protein (IRP) vaccine containing nine Pasteurella haemolytica serotypes was used. Two groups of Awassi lambs were used during the experiment. One group of 10 lambs was vaccinated and the second unvaccinated group of 10 lambs considered as negative control.Blood samples were collected on day 0, 2 and 4 weeks (post-vaccination) and 4 weeks after a revaccination. Antibody titers against capsular antigen of P. haemolytica serotypes A1, A2, A6, A7 and T3 was evaluated for both groups of lambs by indirect haemagglutination test (IHT). Antibody titers against P. haemolytica A2 outer membrane IRPs and the 35 kDa proteins was evaluated by enzyme linked immunosorbent assay (ELISA) and by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting.Results showed that the vaccine used induced sero-conversion against the capsular antigen, major outer membrane proteins (OMPs) and IRPs of P. haemolytica in the vaccinated lambs. Results also showed that one inoculation of the vaccine was insufficient to induce high antibody titers.
Nairobi sheep disease (NSD) virus, the prototype tick-borne virus of the genus Nairovirus, family Bunyaviridae is associated with acute hemorrhagic gastroenteritis in sheep and goats in East and Central Africa. The closely related Ganjam virus found in India is associated with febrile illness in humans and disease in livestock. The complete S, M and L segment sequences of Ganjam and NSD virus and partial sequence analysis of Ganjam viral RNA genome S, M and L segments encoding regions (396 bp, 701 bp and 425 bp) of the viral nucleocapsid (N), glycoprotein precursor (GPC) and L polymerase (L) proteins, respectively, was carried out for multiple Ganjam virus isolates obtained from 1954 to 2002 and from various regions of India.M segments of NSD and Ganjam virus encode a large ORF for the glycoprotein precursor (GPC), (1627 and 1624 amino acids in length, respectively) and their L segments encode a very large L polymerase (3991 amino acids). The complete S, M and L segments of NSD and Ganjam viruses were more closely related to one another than to other characterized nairoviruses, and no evidence of reassortment was found. However, the NSD and Ganjam virus complete M segment differed by 22.90% and 14.70%, for nucleotide and amino acid respectively, and the complete L segment nucleotide and protein differing by 9.90% and 2.70%, respectively among themselves. Ganjam and NSD virus, complete S segment differed by 9.40–10.40% and 3.2–4.10 for nucleotide and proteins while among Ganjam viruses 0.0–6.20% and 0.0–1.4%, variation was found for nucleotide and amino acids.Ganjam virus isolates differed by up to 17% and 11% at the nucleotide level for the partial S and L gene fragments, respectively, with less variation observed at the deduced amino acid level (10.5 and 2%, S and L, respectively). However, the virus partial M gene fragment (which encodes the hypervariable mucin-like domain) of these viruses differed by as much as 56% at the nucleotide level. Phylogenetic analysis of partial sequence differences suggests considerable mixing and movement of Ganjam virus strains within India, with no clear relationship between genetic lineages and virus geographic origin or year of isolation. Surprisingly, NSD virus does not represent a distinct lineage, but appears as a variant with other Ganjam virus among NSD virus group.
Goats aged 3 months were inoculated with a recent isolate of Mycoplasma agalactiae (five animals) or Mycoplasma bovis (five animals) by a combined (intratracheal+intranasal) route. Two control goats were inoculated by the same route with sterile mycoplasma broth. Animals were killed 14 or 21 days after infection. At necropsy, tracheal and lung tissue was taken for pathological and immunohistochemical examination to determine changes in the lymphocyte subpopulations in the bronchus-associated lymphoid tissue (BALT). Consolidation of the lungs was not observed in any animal. M. agalactiae or M. bovis was recovered from the respiratory tract and lung of all but two infected animals. BothMycoplasma spp. induced a moderate bronchointerstitial pneumonia, characterized by lymphoid hyperplasia of the BALT and infiltration of mononuclear cells into the alveolar walls. The predominant phagocytic cell in the pulmonary parenchyma and the airways was the macrophage. The main cellular type in the BALT was the CD3+T lymphocyte, and the ratio of CD4+: CD8+cells was >1. It is likely that cellular immune mechanisms, through the activation of CD4+T lymphocytes, plays a prominent role in the acute and subacute phase of these infections.
