Response of sheep to experimental concurrent infection with tick-borne fever (Cytoecetes phagocytophila) and louping-ill virus

The pathogenesis of concurrent Cytoecetes phagocytophila and louping-ill virus infection was studied in two experiments. In the first experiment 18 four- to seven-year-old rams were used. Ten were infected with C phagocytophila and five days later eight of these animals and the remaining eight sheep were infected with louping-ill virus. The two rams infected with C phagocytophila alone developed no clinical signs apart from a transient pyrexia, while only three of the eight rams infected with louping-ill virus alone showed mild clinical signs. In marked contrast, all eight dually infected sheep developed severe clinical signs with pronounced depression and dysentery and three died and five were killed in extremis. They developed higher titres of viraemia and the antibody response was depressed while necrotising lesions affecting a variety of organs were detected at post mortem examination. Rhizomucor pucillus was recovered from these lesions in seven of the eight sheep. A second experiment using 10 sheep, five aged seven months and five aged two to three years, confirmed the findings of the first experiment indicating that the age of the animal had not significantly influenced the initial result. It was concluded that C phagocytophila infection could enhance the pathogenicity of louping-ill virus and that, operating together, the two pathogens facilitated fungal invasion. It is postulated that sudden deaths in sheep recently transferred to tick-infested pastures may be due to this newly described syndrome.     

Serodiagnosis of Babesia motasi (Wales), Theileria recondita (Wales) and Cytoecetes phagocytophila infection in sheep

The indirect fluorescent antibody test (IFAT) was used to diagnose some tick-borne infections of sheep, Babesia motasi (Wales), Theileria recondita (Wales) and Cytoecetes phagocytophila. Antigen was prepared from blood derived from splenectomised sheep except for C phagocytophila which was derived from a normal animal. A field survey was made to assess the prevalence of B motasi and T recondita in North Wales and a comparison made between the titres using the B motasi (Wales) antigen with those previously reported. IFA titres reported in the homologous system were consistently lower than those described previously. The results of the field survey suggested that B motasi (Wales) infection is more widespread than was originally thought and more widespread than the known distribution of its vector Haemaphysalis punctata. No serological cross reactions occurred between B motasi (Wales), T recondita (Wales), C phagocytophila, B divergens, Sarcocystis ovicanis and Toxoplasma gondii.     

The effects of Ehrlichia (Cytoecetes) phagocytophila on the clinical chemistry of sheep and goats

Tick-borne fever (TBF) is a rickettsial disease of domestic and wild ruminants in temperate climates where the hard tick Ixodes ricinus is present. The disease is characterized by a high temperature and severe leukopenia. In the present study, the effects of TBF on the activity of serum alkaline phosphatase (ALP), and on the concentrations of plasma zinc, iron, total bilirubin, urea, creatinine and albumin were investigated by inoculating one group of eight sheep and one group of eight goats with the Old Sourhope (OS) strain of Ehrlichia (Cytoecetes) phagocytophila. All goats and sheep experimentally infected with E. phagocytophila reacted with fever, rickettsiaemia and leukopenia. The leukopenia was due to an acute lymphocytopenia and prolonged neutropenia. In both groups of animals. TBF was characterized by significant reductions in the activities of serum ALP and concentrations of plasma zinc, iron and albumin. However, there were significant increases in the concentrations of plasma total bilirubin, urea and creatinine in both species of animals. The reductions in ALP and iron were significantly more pronounced in sheep than in goats.     

Lymphocyte responses to mitogens and rickettsial antigens in sheep experimentally infected with Ehrlichia (Cytoecetes) phagocytophila.

Infection of sheep with Ehrlichia (Cytoecetes) phagocytophila, the causative agent of tick-borne fever (TBF), was characterised by a significant reduction in lymphocyte reactivity to the mitogens phytohaemagglutinin, concanavalin A, pokeweed mitogen and Escherichia coli lipopolysaccharide during the period of rickettsiaemia. The addition of the prostaglandin inhibitor, indomethacin, or the nitric oxide inhibitor, N(G)-monomethyl-L-arginine, had no significant effect on the suppressive effects of E. phagocytophila on lymphocyte reactivity to the mitogens. However, peripheral blood lymphocytes obtained from primed sheep proliferated in the presence of live or heat-inactivated E. phagocytophila. Antigen-specific proliferation was detected in lymphocytes samples obtained 11 to 21 days post-inoculation with E. phagocytophila.     

An electron microscopic study of Cytoecetes phagocytophila infection in Ixodes ricinus

Tick-borne fever, caused by a rickettsia-like organism, Cytoecetes phagocytophila, is transmitted by the sheep tick Ixodes ricinus. An electron microscopic technique was developed to examine I ricinus for C phagocytophila infection. Infected and uninfected ticks were obtained from a laboratory maintained tick culture. All stages of I ricinus collected from one field site were examined; 44 per cent of nymphae and 32 per cent of adults were infected with C phagocytophila, but larvae were uninfected. This supports the previously held theory of transtadial, but not transovarial transmission of tick-borne fever.     

Chemotherapy and pharmacokinetics of some antimicrobial agents in healthy dwarf goats and those infected with Ehrlichia phagocytophila (tick-borne fever)

The therapeutic efficacy and pharmacokinetics of oxytetracycline (10 mg kg-1), ampicillin (20 mg kg-1) and a combination (TSS) of trimethoprim (20 mg kg-1), sulphadimidine (50 mg kg-1) and sulphamethylphenazole (50 mg kg-1) were investigated in normal dwarf goats and in those infected with Ehrlichia phagocytophila. Goats given oxytetracycline or TSS intravenously showed improvement, whereas ampicillin was ineffective. The infected goats had significantly prolonged elimination half-life values for sulphadimidine and oxytetracycline. The disposition kinetics of ampicillin and sulphamethylphenazole showed no marked differences between the healthy and infected animals. The tick-borne fever model used in the present study can be of value in testing the therapeutic efficacy and pharmacokinetics of chemotherapeutic agents in rickettsial infections.     

Efficacy of long-acting oxytetracycline for the prevention of tick-borne fever in calves

Twenty-six calves which had not previously grazed tick-infested pasture were divided into two equal groups. On May 26, 1988 (day 0) they were turned out into a field of rough grazing where cases of redwater fever had occurred the previous spring. Seven, 14, 21 and 28 days after the start of the trial the animals in one group each received an intramuscular injection of 20 mg/kg bodyweight of long-acting oxytetracycline. During the 60 days of the trial the animals received a severe tick-borne fever challenge, in some cases combined with a redwater fever challenge. An unforeseen complicating factor was the presence of animals persistently infected with bovine viral diarrhoea virus, present in almost equal numbers in both groups. At the end of the trial the treated group weighed on average 16 kg more than the control group, a difference which was attributed to the suppression of tick-borne fever by oxytetracycline.     

Differential haematological effects of tick-borne fever in sheep and goats

Tick-borne fever (TBF) is a rickettsial disease of domestic and wild ruminants in temperate climates. It is characterized by high fever and severe leukopenia. In the present study, the possible difference in the severity of disease in sheep and goats was investigated by inoculating one group of eight goats and one group of eight sheep with the Old Sourhope (OS) strain of Ehrlichia (Cytoecetes) phagocytophila. All sheep and goats experimentally infected with E. phagocytophila reacted with fever and rickettsiaemia, but there were significant differences between goats and sheep in the severity of clinical disease, the duration and magnitude of fever, the magnitude of rickettsiaemia and the patterns of reduction in the number of total leucocytes. Sheep reacted with fever significantly earlier than goats and the febrile period lasted for a significantly longer period. In contrast, the magnitude of rickettsiaemia was significantly higher in goats than in sheep. Infection with TBF was characterized by a transient increase in the number of neutrophils, which was quickly followed by an acute reduction in the number of lymphocytes and a prolonged reduction in the number of neutrophils in both sheep and goats. In both groups of animals, infection with TBF was also characterized by significant reductions in the total number of red blood cells (RBCs), thrombocytes and packed cell volume (PCV) and the concentration of haemoglobin (Hb). However, the mean corpuscular volume (MCV) and mean corpuscular Hb (MCH) were significantly increased in sheep only.     

Tick-borne fever (Ehrlichia phagocytophila infection) on a dairy farm in Friesland

Disease outbreaks, associated with ixodid ticks, among cows on a dairy farm in Friesland, in the north of the Netherlands, were monitored during the summers of 1996, 1997, and 1998. The most important symptoms were a sudden drop in milk yield, fever, apathy, and problems with locomotion. The diagnosis 'tick-borne fever' was confirmed by finding Ehrlichia phagocytophila inclusion bodies in neutrophils of blood smears from sick animals. Significant economic losses due to the disease were recorded each year. Borrelia burgdorferi, although transmitted by the same tick, did not seem to be involved. Further research on tick-borne fever is of particular relevance in connection with recent E. phagocytophila cases diagnosed in dogs and in a human patient in the Netherlands.     

Lymphocyte subpopulations in peripheral blood of sheep experimentally infected with tick-borne fever.

The lymphocyte subpopulations in the peripheral blood of normal sheep and sheep experimentally infected with Cytoecetes phagocytophila, the causative agent of tick-borne fever, were analysed by flow cytometry, using a panel of monoclonal antibodies against specific lymphocyte epitopes. Experimental infection with tick-borne fever was characterised by a significant reduction in the total number of circulating lymphocytes six days after experimental infection (P less than 0.001). This lymphocytopenia was associated with a significant reduction in the number of B (LCAp220+) and T (CD5+) lymphocytes (P less than 0.001) but there was a significant increase in the number of cells which were neither T nor B (CD5-LCAp220-) cells (P less than 0.01). The reduction in the number of T lymphocytes was due to reduced numbers of circulating CD4+ (helper) T cells, CD8+ (cytotoxic/suppressor) T cells and those with the pan T cell marker (CD5+) but without CD4 or CD8 epitopes (CD4-CD8-). All lymphocytes returned to preinoculation levels 13 to 16 days after experimental infection.     

Prophylactic efficacy of clorsulon against Fasciola hepatica in calves and sheep

A daily oral 5 mg kg-1 dose of clorsulon for 28 days in calves given Fasciola hepatica cysts at 3, 5, and 7 days after initiation of treatment was highly effective in reducing worm burdens (98%) and preventing liver pathology. In similarly infected and treated sheep, clorsulon showed little effect as a prophylactic for delaying the onset of liver pathology. The size of flukes recovered from treated sheep was reduced. Although clorsulon prevented development of fascioliasis in treated calves, the host antibody response was qualitatively similar to that of untreated infected calves, but the magnitude of the response was reduced. Blood clorsulon levels in calves rose to 2.90 micrograms ml-1 within the first week of treatment then fluctuated between 2.65 and 2.90 micrograms ml-1 for the next two weeks. Clorsulon levels in sheep were 0.50-0.60 micrograms ml-1 lower than those in calf blood. The difference in bioavailability of clorsulon between sheep and calves may have contributed to differences in efficacy of the drug.     

Immunisation against East Coast fever by the infection and treatment method: evaluation of the use of ice baths for field delivery and appraisal of an acid formulation of long-acting tetracycline

Immunisation by the infection and treatment method using the Katete strain is currently the most efficient prophylactic technique to control East Coast fever (ECF) in the endemic areas of the Eastern Province of Zambia. The maintenance of the cold chain in liquid nitrogen up to the time of inoculation and the cost of the reference long-acting oxytetracycline (Terramycin LA, Pfizer) are the main drawbacks of the method. The work presented in this paper aims at reducing the cost of immunisation against ECF by using an ice bath for the field delivery and a cheaper long-acting oxytetracycline formulation as chemotherapeutic agent. In experimental conditions, the results from 40 calves immunised after various periods of storage on ice ranging from 4 to 32 h indicate that deferred immunisation performed with a stabilate kept on ice for up to 6h after thawing has an efficiency of 90%. Moreover, sporozoites kept on ice were still surviving 32 h after thawing. In a field trial, 91 calves were inoculated with a stabilate kept for 3.5-5.5 h after thawing and dilution whereas 86 calves were immunised using the standard method. Clinical and parasitological reactions to immunisation were monitored as well as the seroconversion. In the field trial, the deferred immunisation was more efficient than the standard method. The acid formulation of oxytetracycline that was tested was found as suitable as the reference alkaline formulation for the chemotherapeutic control of the Katete strain in ECF immunisation. One indoor trial was carried out on 10 animals and a field trial involved 93 calves.     

Functions of neutrophils in sheep experimentally infected with Ehrlichia phagocytophila

Ehrlichia phagocytophila infection in sheep is characterized by persistent neutropaenia, indicative of decreased phagocytic capacity. This predisposes infected animals to other infections. A whole blood flow cytometrical method was used to document the degree and extent of reduced phagocytic and respiratory burst activity in phagocytes during an experimental infection with E. phagocytophila, and monitored until 56 days post-infection. Six sheep at 5 months of age were inoculated with an intravenous injection of infected blood. Six age-matched sheep were used as controls. A period of reduced respiratory burst lasting up to Day 17 post-infection was recorded. The population of cells showing phagocytic activity without respiratory burst was larger in the infected animals compared to controls up to Day 45 post-infection.