Clinical signs and pathology shown by British sheep and cattle infected with bluetongue virus serotype 8 derived from the 2006 outbreak in northern Europe.

Four poll Dorset sheep and four Holstein-Friesian cattle were infected with the northern European strain of bluetongue virus (BTV), BTV-8, to assess its pathogenicity in UK breeds. The time course of infection was monitored in both species by using real-time reverse transcriptase-PCR (RT-PCR), conventional RT-PCR and serology. Two of the sheep developed severe clinical signs that would have been fatal in the field; the other two were moderately and mildly ill, respectively. The cattle were clinically unaffected, but had high levels of viral RNA in their bloodstream. Real-time RT-PCR detected viral RNA as early as one day after infection in the cattle and three days after infection in the sheep. Antibodies against BTV were detected by six days after infection in the sheep and eight days after infection in the cattle. Postmortem examinations revealed pathology in the cattle that was more severe than suggested by the mild clinical signs, but the pathological and clinical findings in the sheep were more consistent.

A simpler, more sensitive competitive inhibition enzyme-linked immunosorbent assay for detection of antibody to malignant catarrhal fever viruses.

An earlier competitive inhibition enzyme-linked immunosorbent assay (CI-ELISA) was developed for detection of specific antibody against malignant catarrhal fever (MCF) viruses (MCFV) in ruminants. In this study, the indirect CI-ELISA was improved by conjugating the monoclonal antibody 15-A directly with horseradish peroxidase and by developing a method of producing precoated, dried antigen plates. This new test is referred to as a direct CI-ELISA. The reformatted test yielded a significantly improved sensitivity, and the time required was reduced to about one-sixth of the previous time. Of 37 MCF cases in cattle that were confirmed by histopathology and polymerase chain reaction (PCR) assay, 37 (100%) were positive by the new test, whereas the indirect CI-ELISA detected only 23 (62%). The direct CI-ELISA detected antibody to MCFV in 100% of 48 sheep that had been defined as infected with ovine herpesvirus 2 (OvHV-2) by PCR, whereas the indirect CI-ELISA detected only 41 (85%). Comparison of antibody titers measured by the 2 assays for sera collected from OvHV-2-infected sheep and from cattle, bison, and deer with clinical sheep-associated MCF revealed that the direct CI-ELISA offered a 4-fold increase in analytical sensitivity over the indirect format. The number of seropositive animals detected by the direct CI-ELISA among apparently normal cattle and bison was 2-3 times greater than the number detected by the indirect CI-ELISA, indicating that a significant percentage of normal cattle and bison are subclincally infected with MCFV.

Prevention and treatment of Encephalitozoon cuniculi infection in rabbits with fenbendazole.

The efficacy of fenbendazole for preventing an experimental infection of Encephalitozoon cuniculi and for eliminating the spores from the central nervous system of naturally infected rabbits was investigated. Fenbendazole (20 mg/kg bodyweight daily) was administered from seven days before until two or 21 days after rabbits had been infected orally with 10(6) spores of E. cuniculi. Both regimens were effective in preventing the establishment of the parasites, as demonstrated by negative parasitic-specific serology and by the failure to isolate the parasite from brain tissue. In naturally infected, seropositive rabbits, parasites were successfully isolated from seven of nine untreated animals, but not from the brain tissue of eight animals treated with fenbendazole-medicated pellets for four weeks.

[Malignant catarrhal fever in Switzerland. 1.Epidemiology]. / Bösartiges Katarrhalfieber in der Schweiz. 1. Teil: Epidemiologie.

Malignant catarrhal fever (MCF) is a usually fatal infectious disease of cattle with global distribution. Based on the recent introduction of a diagnostic PCR assay and a competitive inhibition ELISA (ciELISA) epidemiological data were collected on field cases in Switzerland. Throughout a three-year period, an MCF incidence of 0.6@1000 was observed, with a gradient of cases from Eastern to Western Switzerland. While the cantons Wallis, Vaud and Geneva reported no and the remaining western cantons only reported a few cases, the highest incidence was observed in the cantons Appenzell Innerrhoden, Lucern, Glarus, Grison, St. Gallen, Schwyz, and Thurgau. MCF occurred seasonally and an age-related clustering was also observed. About 50% of all cases and all outbreaks with more than one animal in a single herd occurred between April and June. Animals between six months and two years were strongly over represented. Observations on four surviving cattle showed that the outcome of the disease is not invariably fatal and that these persistently infected cows can produce healthy negative calves. Investigations on the aetiology indicate that the main reservoir for OvHV-2 is in sheep and possibly goats, while cattle do not normally harbor the virus. An OvHV-2 negative sheep herd was raised from lambs, which were reared colostrum-free and in isolation from their mothers. The success rate clearly indicated that vertical intrauterine infection is not the main mode of transmission among sheep. Therefore, horizontal, seasonally occurring transmission of OvHV-2 among sheep has to be assumed.

[Malignant catarrhal fever in Switzerland: 2. Evaluation of the diagnosis]. / Bösartiges Katarrhalfieber in der Schweiz: 2. Teil: Evaluation der Diagnostik.

Malignant catarrhal fever (MCF) is a mostly fatal lymphoproliferative disease of cattle. In 1995 a PCR based method was introduced for the detection of the ovine herpesvirus 2 (OvHV-2), which is regarded as the causative agent of the sheep-associated form of the disease. This PCR can be regarded as a gold standard for the in vivo diagnosis of sheep-associated MCF in cattle (Müller-Doblies et al., 1998). This semi-nested PCR was now used as a reference test for the reassessment of diagnostic criteria in the clinical and post mortem diagnosis that could previously not be quantitated. Based on 83 suspected cases with a complete clinical record the clinical signs were weighted and grouped according to their sensitivity and specificity into lead signs indicative of MCF and frequently accompanying signs supportive for the diagnosis of MCF and general clinical signs that were less reliable for the diagnosis. Differential diagnoses are discussed, which are of particular significance due to their status as OIE list A diseases e.g. foot-and-mouth disease or rinderpest. 38 PCR confirmed cattle with MCF served for the quantitative analysis of organ lesions. For the post mortem diagnosis an essential set of organ samples is defined to permit a reliable histological diagnosis, as the gross pathology often did not give any indication for the diagnosis. These criteria should help to improve the diagnostic efficiency and to select the appropriate laboratory diagnostic procedures for MCF-suspected cattle.

Feeding by the tick, Ixodes scapularis, causes CD4(+) T cells responding to cognate antigen to develop the capacity to express IL-4.

Effects of tick feeding on an early antigen-specific T cell response were studied by monitoring a clonotypic population of adoptively transferred T cell receptor (TCR) transgenic CD4 cells responding to a tick-associated antigen. When recipient mice were infested with pathogen-free Ixodes scapularis nymphs several days prior to T cell transfer and intradermal injection of soluble cognate antigen at the feeding site, the clonotypic CD4 cells gained the ability to express the Th2 effector cytokine IL-4. Notably, this effect was not only observed in BALB/c mice predisposed towards developing Th2 responses but also in B10.D2 mice predisposed towards Th1 responsiveness. Furthermore, tick feeding was able to superimpose IL-4 expression potential onto a strong Th1 response (indicated by robust IFN-gamma expression potential) elicited by immunization with a vaccinia virus expressing the cognate antigen. The magnitude to which tick feeding was able to programme IL-4 expression potential in CD4 cells was partially reduced in mice that had been previously exposed to pathogen-free tick nymphs 6 weeks earlier, as well as when the nymphs were infected with Borrelia burgdorferi. Intradermal injection of salivary gland extract programmed IL-4 expression potential similar to that of tick infestation, suggesting that IL-4 programming activity is contained within tick saliva.

Flow cytometric assessment of allopurinol susceptibility in Leishmania infantum promastigote.

BACKGROUND: Leishmaniasis is a major tropical and subtropical parasitic disease. Sodium stibogluconate, N-methyl -D-glucamine antimoniate, amphotericin B, pentamidine, and ketoconazole are drugs used to treat this disease. Some of these drugs cause severe adverse side effects and treatment failures are common. Allopurinol, a purine analog, has been used to treat leishmaniasis, alone or combined with the previously mentioned drugs. Low cost, ease of administration (oral), and lack of toxicity make allopurinol a particularly appealing candidate. METHODS: The effect of allopurinol on Leishmania infantum (MCAN/ES/89/IPZ229/1/89, zymodeme MON1) wild-type promastigotes (wt-p229), and an altered form of these promastigotes (allo-p229) resulting from long term in vitro exposure to allopurinol, was determined by [(3)H]-thymidine incorporation assays and by diverse flow cytometric approaches. RESULTS: Allopurinol arrested the proliferative capacity of wt-p229 promastigotes, reduced the proportion of viable cells, and decreased their total protein content. In contrast, allo-p229 promastigote proliferation was only slightly decelerated and the proportion of viable cells and the protein content were not affected by the allopurinol treatment. CONCLUSIONS: The flow cytometry approach allowed us to demonstrate differences in allopurinol susceptibility of the two promastigote forms, expanding the spectrum of flow cytometry applications in studies of parasite resistance.

Field validation of laboratory tests for clinical diagnosis of sheep-associated malignant catarrhal fever.

Until recently, sheep-associated malignant catarrhal fever (SA-MCF) was diagnosed mainly on the basis of clinical presentation and histopathological changes. Using clinically diagnosed field cases, we have evaluated a seminested PCR and a competitive inhibition enzyme-linked immunosorbent assay (CI-ELISA) and compared these assays in the diagnosis of SA-MCF in cattle with histopathology as a provisional "gold standard." Samples from 44 cattle with clinical signs suggestive of SA-MCF were examined by histopathology, PCR, and CI-ELISA. In addition, samples from healthy cattle were evaluated by PCR (n = 96) and CI-ELISA (n = 75). Based on histopathology, 38 of the 44 clinical cases were classified as SA-MCF positive, 3 were classified as inconclusive, and 3 were classified as SA-MCF negative. The sensitivity of PCR was 95 to 97%, whereas the specificity ranged between 94 and 100%. The CI-ELISA showed a sensitivity of 56 to 87% and a specificity between 91 and 100%. In the field, there is good correlation between the diagnoses of SA-MCF by histopathology, PCR, and CI-ELISA. These data also confirm the close association of ovine herpesvirus 2 with SA-MCF in Switzerland.