Diagnosis and manifestation of encephalitozoonosis in mice after experimental infection with different species and application of dexamethasone.

Twenty-eight BALB/c mice were infected with different strains of Encephalitozoon species (Encephalitozoon cuniculi II - mouse type, E. cuniculi III - dog type, Encephalitozoon hellem, Encephalitozoon intestinalis). Five of them were infected with E. cuniculi II (mouse type) and simultaneously immunosuppressed with dexamethasone. Clinical signs of encephalitozoonosis were not remarkable. Ascites was found in two mice of dexamethasone-treated group 14 days post-infection (p.i.). The histopathological changes were found mainly in spleen and liver in the form of lymphoepithelioid granuloma. Spores were found in faeces since day 14 p.i. and visualized by Calcoflour White M2R. After cultivation on cellular cultures (VERO E6 - monkey kidney cells, RK-13 - rabbit kidney fibroblasts), the species differentiation was performed by PCR using panmicrosporidial primers (PMP1, PMP2) and specific primers (ECUN-F, ECUN-R, V1, SI-500). The differences were recorded in the immune response of immunocompetent and immunosuppressed mice. At day 60 p.i., the titres of specific antibodies measured by indirect immunofluorescence antibody test were lower (1:4096) in dexamethasone-treated mice when compared with non-immunosuppressed animals (1:8196). The significant increases of antibody titres were recorded in particular infected groups within the experiment (P < 0.01 between day 14 p.i. and day 30 p.i., P < 0.001 between day 14 p.i. and day 60 p.i.). Experimental encephalitozoonosis in non-immunosuppressed and immunosuppressed mice provides a useful model for the study of immune response and lesions associated with these protozoans.

Immune response of E. cuniculi infected mice to aflatoxin B1.

The immunomodulatory effects of aflatoxin B1 (AFB1) on mice experimentally infected with Encephalitozoon cuniculi (E. cuniculi) were studied. Mice inoculated intraperitoneally with spores of E. cuniculi drank a daily solution of AFB1 (0.2 mg kg(-1) of body weight) for 27 days. Application of AFB1 to mice demonstrated a decrease of immunocompetent cells. On the other hand, the mice infected with E. cuniculi and given AFB1 showed significant increased number of both monocytes and CD8+ T cells, and tendency to a decrease in CD4+ T cells. Aflatoxin B1 revealed to merely modulate systemic immune response of E. cuniculi infected mice.

Prevalence of antibodies to Encephalitozoon cuniculi (microsporidia) in angora goats--a potential risk of infection for breeders.

The presence of antibodies against Encephalitozoon cuniculi in Angora goats was detected by the method of indirect immunofluorescence (IFAT). The animals reacting at the titre 1: 64 and more were considered positive. Of the total number of 48 sera examined, 4 were positive at the titre 1: 32 and 2 were positive at the titre 1: 64. The occurrence of antibodies against E. cuniculi indicates that one of the causes of disorders in the reproductive cycle in Angora goats may be microsporidia Encephalitozoon cuniculi, and that these animals may be potential sources of infection for people.

Immune response to Encephalitozoon cuniculi infection in laboratory mice.

The study was performed to determine the immune response to Encephalitozoon cuniculi infection in immunocompetent mice during 120 days of experiment. Mice infected with E. cuniculi had an increased number of CD4+ T cells up to Day 20 post infection (p.i.), but counts of CD8+ T lymphocytes were significantly lower up to Day 90 p.i. in peripheral blood. Blood monocytes were significantly increased on the Day 60 and Day 120 of infection. A lack of significant decrease of CD4+ T cells may be considered as an important event in the immune response to E. cuniculi infection in immunocompetent mice.

Catecholamines and encephalitozoonosis in rabbits.

Twenty four rabbits (Oryctolagus cuniculus f. domestica) were used to detect specific anti-Encephalitozoon cuniculi antibodies. To identify microsporidian infection, a haemolytic test in agar gel was carried out. Blood samples of animals with and without spontaneous encephalitozoonosis were evaluated, and compared for the presence of epinephrine (EPI), norepinephrine (NE), and dopamine (DA). Rabbits infected spontaneously with E. cuniculi had significantly lower levels of catecholamines than healthy animals. This decrease in catecholamines is of special interest because of their role as factors modifying the immune response. These neuromediators also have different influences on the function of immune cells.

Tissue macrophages of calves after high doses of glucan.

Relatively high doses of the soluble form of fungal glucan (50 mg/kg of body weight) were administered to two clinically healthy calves. Hepatosplenomegaly was not observed. Quantitative and significant qualitative changes were seen in macrophages in the medulla of lymph nodes as compared to the control animals. Both the fibrillar and the granular components in the vacuoles of phagocytosing cells appeared to be positive in the periodic acid-Schiff reaction (PAS); an amorphous component was PAS negative. Quantitative changes occurred also in the expression of IgM molecules on the surface of B lymphocytes. The results indicate that glucan can be used in ruminants also in a single high dose.

[Evaluation of indicators of cellular immunity in experimental infectious bovine rhinotracheitis virus infection in calves treated with glucan]. / Hodnotenie niektorých ukazovatel'ov bunecnej imunity po experimentálnej infekcii vírusom IBR u teliat osetrených glukánom.

Some parameters of cell-mediated immunity (phagocytic activity PA and phagocytic index PI, metabolic activity and migration-inhibitive factor MIF of blood leucocytes) were investigated in calves experimentally infected with IBR virus (group II) and in calves premedicated with glucan seven days before infection (group I). A significant increase in the percentage of phagocytizing cells was observed in both groups of animals on days 2-4 of the experiment (P < 0.01) in comparison with the values before infection. If the two groups of animals were confronted with each other, there was a significant difference in the percentage of phagocytizing cells on days 1 and 2 after infection (P < 0.05), with the higher average values in the calves which were only infected. An evaluation of PI showed a statistically significant difference only in the glucan-treated calves on day 7 after its administration (P < 0.05), i.e. on day zero of the experiment. The inhibition of leucocyte migration ability was found on day 1 after infection, and it persisted for four days, without any significant differences between the groups. An evaluation of the metabolic activity of leucocytes (INT-test) showed a decrease in the calves of both groups in comparison with the initial value, without any significant differences when the values were confronted with each other.

[The effect of an insoluble fungal glucan on indicators of immunity in calves]. / Vplyv nerozpustného fungálneho glukánu na niektoré ukazovatele imunity teliat.

Immunomodulative effect of insoluble fungous glucan (IFG) was observed on nine accidentally chosen calves, which were in the age from 4.5-5.5 months. The following items were observed: T, B lymphocytes, white blood cell count, index and activity of phagocytosis, induction and determination of interferon from leukocytes. In the experimental group there was a decrease in B-rosseting cells observed. On the 28-th day and on the 42-th day of experiment duration a significant increase in B-rosseting cells in comparison with control group has been observed. Average values were significantly different on the levels P < 0.01 and P < 0.05. Percentages of T-rosseting cells were almost the same during the whole experiment. In the experimental group on the 7-th and on the 14-th day the decrease has been observed. By the Student's t-test a significant difference in favour of group, where IFG was applicated, has been observed. Percentage of white blood cell count was from 47 to 72 in both groups. When phagocytic activity was evaluated, there was the higher percentage of phagocytic cells in the 7th, 14-th, 21-st, 28-th and 42-th day of experiment when compared with the 0-day value. Concerning the phagocyte index a statistical difference was observed, when compared to the initial value in favour of the experimental group. The initial disbalanced values of interferon, which have been observed during the period from the 7-th to the 21-st day, increased on the 28-th and the 42-nd day of experiment which was statistically confirmed on the following levels: P < 0.05 and P < 0.01.

[Isolation and control of the functional quality of mRNA of bovine leukocyte interferon]. / Izolácia a kontrola funkcnej kvality mRNA bovinného leukocytárneho interferónu.

For the construction of the cDNA library total cellular RNA was separated from bovine leucocytes induced with NDV (inductor of interferons) for five hours. Guanidinisothiocyanate and ultracentrifugation in the gradient of CsCl were used for the separation of the total RNA (Chirgwin et al., 1979). Bands 18S and 28S were detected in the samples of RNA by electrophoresis in an MOPS buffer (Fig. 1). mRNA was separated from the mixture of RNA via affinity chromatography on oligo(dT)-cellulose (Aviv and Leder, 1972). The function quality of mRNA BoIFN-alpha was verified by microinjection into the oocytes of Xenopus laevis using a microinjector of our own construction (Fig. 2). The microinjector was calibrated by injection of a radioactive 51Cr solution (Fig. 3). It was found out on the basis of CPE inhibition measurements that the injected oocytes synthesized 320-640 U/ml BoIFN-alpha (16-32 U/50 microliters; Tab. I).