Determination of ceftiofur in bovine plasma by HPLC-DAD.

Ceftiofur sodium is a third generation broad-spectrum cephalosporin, formulated as an intramuscular injection, which is used to treat respiratory diseases in swine, ruminants and horses. The thioester bond on ceftiofur is rapidly cleaved to give desfuroylceftiofur which is further metabolized to a disulfide dimer and various desfuroylceftiofur-protein and amino acid conjugates. Methods of analysis of ceftiofur rely on cleavage by dithioerythritol to produce desfuroylceftiofur, which is further stabilized by derivatization to desfuroylceftiofur acetamide using iodoacetamide. Previous analytical methods have extracted derivatized analyte from plasma and tissue using solid-phase extraction clean-up steps followed by HPLC analysis with results reported as ceftiofur-free acid equivalents (CFAE). The simplified method presented here involves direct HPLC injection of a cleaved and derivatized sample following a protein precipitation step with calibration by external standardization and selectivity achieved based on chromatography and diode-array detection (DAD). The assay was linear over the calibration range 0.4-40 microg/ml in plasma. Intra-batch reproducibility R.S.D. was 10.3% and intra-batch sample repeatability R.S.D. was 2.1% at the 5 microg/ml level. The mean accuracy over the range of the calibration curve was -4.2% and the detection limit was 0.15 microg/ml. The assay was successfully applied to bovine plasma following intramuscular injection of ceftiofur sodium. This simplified method is suitable for pharmacokinetic applications involving ceftiofur at normal therapeutic levels.

Induction of Mx-related protein in cat peripheral blood mononuclear cells after administration of recombinant human interferon hybrid.

A 78 kDa protein was induced in cat peripheral blood mononuclear cells after in vivo administration of recombinant human interferon-alpha hybrid (rHuIFN-alpha B/D). This protein was antigenically related to the IFN-induced human (78 kDa) and mouse Mx proteins. Quantitative immunoblot analysis indicated that the induction of the cat Mx protein was dose-dependent. There was a dissociation in time between plasma levels of IFN which were transient, and levels of cat Mx protein which remained elevated at least five days after dosing. Our results provide evidence that a human IFN-alpha hybrid may be active in cats. They also indicate that the Mx protein is a sensitive, quantitative, and stable marker to follow IFN activity in vivo in cats.

Antiviral and antigenic properties of recombinant porcine interferon gamma.

Recombinant porcine interferon gamma (rPoIFN gamma) induced a dose-dependent inhibition of the cytopathic effect produced by vesicular stomatitis virus (VSV) challenge of both homologous and heterologous (bovine) cell lines. In addition, an antiviral effect of rPoIFN gamma was demonstrable against the coronavirus transmissible gastroenteritis virus (TGEV) infection of porcine epithelial cells and of pulmonary macrophages. A rabbit anti-PoIFN gamma antiserum was prepared and shown to specifically neutralize the antiviral effects of natural and recombinant porcine IFN gamma preparations. This antiserum could also neutralize recombinant bovine IFN gamma but not recombinant human IFN gamma. These results suggest antigenic homology of porcine and bovine IFN gamma but antigenic differences between these molecules and human IFN gamma.

Recombinant bovine interferon-gamma enhances expression of class I and class II bovine lymphocyte antigens.

Recombinant bovine interferon-gamma augments expression of class I and class II histocompatibility antigens on the surface membrane of bovine lymphocytes. Immunofluorescence techniques using a series of monoclonal anti-HLA antibodies demonstrate that this enhancement is detectable as early as 24 h after incubation with rBoIFN, while maximum surface expression is obtained within 3-5 days. A concentration as low as 10 units of rBoIFN is effective. Such results may be useful for characterizing the BoLA gene products.

Panleukopenia-like syndrome of FeLV caused by co-infection with FeLV and feline panleukopenia virus.

To study the effect of interferon on feline leukemia virus (FeLV) infection, 30 specific pathogen free (SPF) cats were infected with the apathogenic FeLV A Glasgow. Unexpectedly, between 5 and 8 weeks after FeLV infection, all 19 cats with persistent FeLV infection but not the FeLV-negative cats died from a panleukopenia-like syndrome. No feline panleukopenia virus (FPLV) antigen was found in feces by latex agglutination, enzyme-linked immunosorbent assay (ELISA) or immunoelectron microscopy. No enteropathogenic bacteria were found. Histopathology revealed changes resembling those of FPLV infection such as destruction of crypts and pancytopenia of bone marrow. Neither clinical signs nor seroconversion to FPLV could be induced by transmitting intestinal extracts to two SPF cats. However, FPLV antigen was demonstrated by immunofluorescence assay in intestinal cryostat sections of diseased animals. FPLV could also be demonstrated in intestinal extracts by immunoelectron microscopy, by latex agglutination and ELISA after anti-FPLV antibodies were removed from immune-complexed FPLV by ultracentrifugation over a CsCl gradient at pH 2.0. From these experiments it was concluded that the panleukopenia-like syndrome of FeLV may not be caused by FeLV alone but at least in some cases by co-infection with FeLV and FPLV. In addition, some form of 'cooperation' between FeLV and FPLV must be postulated because neither virus alone induced symptoms.

Resistance and immunity of dogs against Babesia canis in an endemic area.

Canine babesiosis is increasing in incidence and prevalence in some areas in France and is now a major problem for dogs. Sex and age do not have any influence on the animals' susceptibility to the disease. Some breeds are more resistant (Beagle, Fox terrier, Porcelain, Teckel, mongrel dogs) and others are more susceptible (Spaniel, Cocker, Griffon, Yorkshire terrier, Doberman, Pekinese); however, none of them is completely resistant. Dogs which live in endemic areas can synthesize antibodies against Babesia canis, sometimes at high levels, without any sign of disease.

Immunity of dogs against Babesia canis, its vector tick Dermacentor reticulatus, and Ixodes ricinus in endemic area.

Previous epidemiological studies allowed us to accurately define endemic areas of canine babesiosis and tick distribution in southeastern France (Martinod, 1983). Using a micro-ELISA test 100 dogs sera were tested with 3 antigens: Babesia canis, Dermacentor reticulatus and Ixodes ricinus. Antibodies against B. canis and its vector D. reticulatus were detected in an endemic area, sometimes with high levels (optical density 1.38 and 0.80 respectively). A correlation factor and regression lines were found between ELISA activity of B. canis and vector tick antigens, even for dogs which never showed any babesiosis symptoms. These results were compared with those of an area without any babesiosis. Furthermore I. ricinus antigens detected ELISA activity in sera of dogs; some cross reactions were observed between I. ricinus and D. reticulatus antigen.

The effects of recombinant bovine interferon-alpha on fertility in ewes.

Recombinant bovine interferon-alpha(I)1 (rBolFN-alpha) may be useful for enhancing fertility in sheep because it has extensive sequence homology with ovine trophoblast protein-1. To test the effectiveness of rBolFN-alpha, several experiments were performed in which bred females were given intramuscular injections of rBolFN-alpha around the time of maintenance of the corpus luteum. Treatment with rBolFN-alpha enhanced the fertility of ewes that were bred via natural service or embryo transfer of whole or demi-embryos. Interferon treatment was successful in enhancing lambing rate if injections were given twice daily from Days 11 to 18, 12 to 14, 12 to 15 or 12 to 16. Overall, the lambing rate for ewes bred via natural service was 94/126 (74.6%) for control ewes and 101/126 (80.2%) for rBolFN-alpha treated ewes. Litter size was not affected by treatment. Interferon treatment was not successful in increasing the lambing rate if given as a single injection on Day 12 or as a series of once-daily injections from Days 11 to 16. These results demonstrate that rBolFN-alpha can increase the lambing rate in ewes.

Risk assessment related to veterinary biologicals: side-effects in target animals.

Despite established control measures, the large-scale use of veterinary biologicals may involve side-effects. The most common side-effects observed include the following: injection site reactions, systemic reactions, allergic reactions, effects on the immune system, residual pathogenicity, inadequate inactivation, genetic recombination, contamination. New technologies, new harmonised regulations, and commitment to quality will ensure a continuous supply of safe and innovative products.

Prophylactic effects of recombinant bovine interferon-alpha I1 on acute Salmonella typhimurium infection in calves.

The in vivo effects of a single prophylactic dose of recombinant bovine interferon (rBoIFN)-alpha I1 in calves with salmonellosis were investigated, using a Salmonella typhimurium infection model. Treatment with rBoIFN-alpha I1 reduced the degree of septicemia compared with that in control groups, and, in one experiment, using disease of reduced severity, body temperature was lower in treated calves than in controls.

Vaccination practices in veterinary medicine: standardization versus tailored to needs?

Significant achievements have been made during the nineteenth century to improve animal health and welfare through vaccination. Vaccination standards and practices will change significantly in the next century. Vaccination programs need to be tailored to each livestock operation or individual animal in accordance with manufacturers' instructions, government regulations, scientific standards, professional organization guidelines, and veterinarian recommendations. Vaccination should always be considered in a population context. A better use of vaccine will require a different approach and a significant investment in research. Finally, all the partners involved in animal health should consider a close collaboration to meet future animal and public health challenges.

Epidemiology of canine babesiosis in relation to the activity of Dermacentor reticulatus in southern Jura (France).

Dates of onset of canine babesiosis within a hyperendemic focus in France (Rhone Valley, south of Jura and north of the Alps) were compared to the seasonal population level of the adult stage of Dermacentor reticulatus over a 12-month period (December 1982-November 1983). Cases of babesiosis occurred in spring and fall when adult D. reticulatus were active. The fluctuations of the vector tick population and the onsets of canine babesiosis were also correlated with climatic changes: no tick activity or clinical cases of disease were detected in winter (low temperature) or in summer. The slight disparities observed between both distributions might be explained by various factors such as the development of immunity against the parasites, the intervals between tick bites and appearance of symptoms of the disease, or the asynchrony between different biotopes.

Immunomodulatory effects in vivo of recombinant porcine interferon gamma on leukocyte functions of immunosuppressed pigs.

Immunological parameters of porcine peripheral blood mononuclear cells after in vivo injections of recombinant porcine interferon gamma (rPoIFN gamma) were studied in pigs immunosuppressed by dexamethasone (6 mg/kg body weight in a single injection). A 2-d period of rPoIFN gamma injected alone and intramuscularly at a dose of 1 microgram/kg body weight increased interleukin 1 (IL1) production (P less than 0.05). Recombinant porcine IFN gamma also reversed the immunosuppressive effects of dexamethasone on: i), lymphocyte responsiveness to mitogens: PHA (P less than 0.03), ConA (P less than 0.053); ii), IL1 production; and iii), IL2 production (P less than 0.05). However, rPoIFN gamma had no effect on neutrophilia induced by dexamethasone. These data show that rPoIFN gamma modulates leukocyte functions of pigs in vivo.

Induction of circulating tumor necrosis factor cannot be demonstrated during septicemic salmonellosis in calves.

The concentration of tumor necrosis factor in the circulation of calves, which were infected with Salmonella typhimurium and exhibited septicemia as indicated by clinical signs and blood culture, was measured with a radioimmunoassay. These levels were compared with those in calves before infection and in other calves that had received an intravenous dose of gram-negative endotoxin. The tumor necrosis factor levels measured in samples taken during septicemia were not different from those in samples from infected nonsepticemic calves or samples from calves before infection. In contrast, the levels of tumor necrosis factor rose rapidly in calves after treatment with endotoxin by intravenous injection.

Production of interferons by bovine and ovine cell lines infected with Theileria annulata or Theileria parva.

Three bovine cell lines and four ovine cell lines infected with Theileria parva or Theileria annulata were examined for the production of interferon (IFN). Biologically active IFN was detected in the tissue culture supernatants of four of the cell lines. Only one, a bovine cell line infected with T. parva, produced IFN-gamma as measured by specific neutralization with a monoclonal antibody to bovine IFN-gamma. This observation was confirmed by analysing RNA from the cell lines on Northern blots using an IFN-gamma cDNA probe. The other three cell lines which produced IFN were infected with T. annulata. The IFN produced by those lines was not IFN-gamma.

Effect of intrauterine and intramuscular administration of recombinant bovine interferon alpha 1 on luteal lifespan in cattle.

Intrauterine and intramuscular administration of interferon was tested for effectiveness in extending luteal lifespan in cattle. Intrauterine infusion of 1 mg of recombinant bovine interferon-alpha I1, twice daily, to lactating dairy cows from d 14 to 21 after estrus extended interestrous interval (30.4 +/- 1.91 d versus 24.8 +/- .58 d) and functional lifespan of the corpus luteum (28.4 +/- 2.01 d versus 23.6 +/- .75 d). In another experiment, twice daily intramuscular injection of 20 mg interferon to Simmental heifers from d 15 to 19 extended interestrous intervals (24.6 +/- 1.36 d versus 20.6 +/- .49 d) and functional lifespan of the corpus luteum (23.2 +/- .37 d versus 20.2 +/- .73 d). In a third experiment, pubertal dairy heifers received twice daily intramuscular injections of 0, 2.5, 5.0, or 10.0 mg/injection of interferon from d 14 to 21 after estrus. The three interferon-treated groups had longer interestrous intervals and functional luteal lifespans than the control group. Interestrous intervals were 22.0 +/- .68, 24.0 +/- 1.14, 24.6 +/- 1.17, and 25.4 +/- .97 d, respectively. The present data strengthen the theory that an interferon-alpha-like molecule can regulate luteal function in cattle. Such a regulatory compound might prove useful in schemes to reduce embryonic mortality caused by aberrant secretion of embryonic interferon.

Effect of bovine interferon on acute changes in body temperature and serum progesterone concentrations in heifers.

Bovine interferon-alpha I1 has extensive sequence and functional homology with the antiluteolytic protein, bovine trophoblast protein-1. Because of the possible use of interferon-alpha I1 as a drug that supplements embryonic secretion of bovine trophoblast protein-1, interferon-alpha I1 was tested for other biological actions that might affect its usefulness as a fertility-enhancing treatment. Experiments were performed to evaluate whether interferon-alpha I1 causes hyperthermia and an acute depression in circulating concentrations of progesterone. In four experiments, intramuscular administration of interferon-alpha I1 (range 1.25 to 20 mg) caused hyperthermia; average peak body temperatures of 40 to 40.4 degrees C occurred 2.5 to 6 h after injection. Temperatures returned to baseline 12 to 16 h later. The rise in rectal temperature could be reduced, but not totally alleviated, with concomitant administration of an inhibitor of prostaglandin synthesis. The maximal hyperthermic response was similar when interferon-alpha I1 was delivered via osmotic minipumps or through a series of intramuscular injections. The hyperthermic response decreased with repeated daily exposure to interferon-alpha I1. The increase in rectal temperatures was associated temporally with a decrease in serum progesterone. Effects of interferon-alpha I1 on body temperature and circulating progesterone could possibly limit its effectiveness in enhancing fertility.