Effect of aqueous extract from Scutellaria baicalensis Georgi roots on CD4+ and CD8+ T cell responses during experimental infection with Trichinella spiralis in mice.

The aim of this study was to investigate the effect of aqueous extract from Scutellaria baicalensis Georgi roots (SB) on blood parameters and immune response during an experimental trichinellosis. A total of 60 mice infected with 200 Trichinella spiralis larvae were assigned into two groups. One of them served as a control and the second received SB extract orally from day 5 before infection to day 28 after infection (dpi). Blood was sampled at 7, 14, 21 dpi. Lymphocytes obtained from the spleen and mesenteric lymph nodes (MLN) at 7, 14, 21, and 28 dpi were counted, CD4+ and CD8+ subpopulations were analyzed by flow cytometry, and lymphocyte proliferation was estimated with colorimetric (MTT) assay. The intensity of intestinal and muscle invasion was also studied. SB caused a remarkable elevation of banded neutrophils in the blood at 7 dpi. SB increased ConA-stimulated splenocyte proliferation and CD4+ and CD8+ splenocyte subsets at 14 and 21 dpi, whereas MLN lymphocyte subset stimulation involved only CD4+ at 14 dpi. After administration of SB a downward trend in the number of T. spiralis larvae in the muscle was observed. These results suggest that Scutellaria baicalensis root extract stimulates murine cellular immune response during intestinal phase of T. spiralis infection.

Suitability of selected culture media for Blastocystis spp.

Blastocystis is a common enteric protozoan of humans and various species of animals. Culture and microscopic examination of fecal samples is the conventional method for identifying four major forms of Blastocystis (vacuolar, granular, non-vacuolar or cystic). In this article, we com- pared eight liquid media for cultivation of Blastocystis spp. Study material included fecal samples from clinically healthy pigs. Significant differences in the growth of Blastocystis on individual media were observed.

Gastrointestinal nematodes in grazing dairy cattle from small and medium-sized farms in southern Poland.

This study aimed to estimate the prevalence of gastrointestinal nematodes and the intensity of infection in grazing dairy cattle from small and medium-sized farms in southern Poland. The level of antibodies against Ostertagia ostertagi in the bulk tank milk (BTM) from the animals was also assessed. Rectal fecal samples collected from 361 cows on 20 farms were examined using Willis-Schlaaf flotation and the McMaster method. BTM samples were tested for the presence of O. ostertagi antibodies using ELISA. Multiplex PCR was used to identify the third-stage larvae (L3) of gastrointestinal nematodes derived from the culture of pooled fecal samples from sampled farms. Gastrointestinal nematode eggs were found in the samples from 18 of the 20 herds with a prevalence range from 20.4 to 94.5%. The average number of eggs excreted in the feces of the herds was 200 eggs per gram (EPG). Antibodies to O. ostertagi were found in 20 of the examined herds (100%), of which 6 had optical density ratios (ODR) greater than 0.5. PCR results showed the presence of three nematode species: Ostertagia ostertagi, Cooperia oncophora and Oesophagostomum radiatum.

Effect of isosporiasis prevention with toltrazuril on long-term pig performance.

The efficacy of toltrazuril treatment was assessed in two experiments in Polish swine herds. Experiment 1 included a toltrazuril treatment group, Group A (n = 410), and untreated control, Group B (n = 386). Time to sale in Group A was 108 days versus 120 days for Group B, with average body weights at sale of 114.2 kg and 108.8 kg, respectively (P < 0.05). In experiment 2, the health status and body weight gain of 238 piglets treated with toltrazuril (Group D) were compared to 235 untreated piglets (Group K). A similar difference was observed in average body weights of slaughtered animals, being on average 104 kg in Group D and 101 kg in Group K (P < 0.01). Animals from Group D were slaughtered 5 days earlier than animals from Group K (day 166 versus day 171). Data from clinical trials suggest treatment of coccidiosis with toltrazuril offering potential for improved animal welfare and yields, however this has remained unproven in field conditions in large swine production facilities. The present study confirms the efficacy of toltrazuril treatment when used in the field and the subsequent positive impact on time to weaning, time to market, and on weight gain at all time points.

Trichinella spiralis: the influence of short chain fatty acids on the proliferation of lymphocytes, the goblet cell count and apoptosis in the mouse intestine.

This study was carried out to determine the influence of short chain fatty acids (SCFA) on spleen and mesenteric lymph node lymphocyte proliferation, goblet cells and apoptosis in the mouse small intestine during invasion by Trichinella spiralis. BALB/c mice were infected with 250 larvae of T. spiralis. An SCFA water solution containing acetic, propionic and butyric acids (30:15:20 mM) was administered orally starting 5 days before infection and ending 20 days post infection (dpi). Fragments of the jejunum were collected by dissection 7 and 10 dpi, and were examined for apoptotic cells in the lamina propria of the intestinal mucosa, and for goblet cells. The proliferation index of the cultured spleen and mesenteric lymph node lymphocytes with MTT test was also determined. The orally administered SCFA solution decreased the proliferation of mesenteric lymph node lymphocytes in the mice infected with T. spiralis at both examination times, but did not influence the proliferative activity of the spleen cells. Seven dpi, both in the spleen and mesenteric lymph nodes, the highest proliferation index of concanavalin A (Con A)-stimulated lymphocytes was found in the group of uninfected animals receiving SCFA animals. This tendency could still be seen 10 dpi in the mesenteric lymph nodes but not in the spleen, where the proliferation index in this group had significantly decreased. In vitro studies revealed, that butyric and propionic acids added to the cell cultures suppressed the proliferation of Con A-stimulated mesenteric lymph nodes and spleen lymphocytes taken from uninfected and T. spiralis-infected mice. Acetic acid stimulated proliferation of splenocytes taken from uninfected mice but did not affect lymphocyte proliferation in mesenteric lymph nodes from uninfected or infected mice. Orally administered SCFA increased the number of goblet cells found in the epithelium of the jejunum 7 dpi, but this number had decreased 10 dpi. The number of apoptotic cells in the lamina propria of the intestinal mucosa of animals infected with the T. spiralis and receiving SCFA was also lower, particularly 10 dpi. The above results show that SCFA can participate in the immune response during the course of trichinellosis in mice.

The effect of immunomodulating drugs on the percentage of apoptotic and necrotic lymphocytes in inflammatory infiltrations in the muscle tissue of mice infected with Trichinella spiralis.

In order to determine the effect of apoptosis and necrosis on the intensity of the muscular phase of infection by Trichinella spiralis, male CFW mice were orally infected with T. spiralis larvae and treated with some immunomodulating drugs: calf thymus extract (TFX), lipopolysaccharide from Escherichia coli (LPS), and dexametasone (DEX). Treatment with TFX increased the proportion of apoptotic lymphocytes and decreased the proportion of necrotic lymphocytes from 14 to 60 days after infection in mice infected with T. spiralis. Treatment with LPS increased proportions of both apoptotic and necrotic lymphocytes from 21 to 60 days after infection, especially at 28 days after infection. Treatment with DEX increased the proportion of apoptotic lymphocytes only at 28 days after infection, and significantly increased the proportion of necrotic lymphocytes at 21 days after infection. Parasite load in the affected muscle tissue was significantly lower than the control in mice treated with TFX, not significantly different from the control in mice treated with LPS, and significantly higher than the control in mice treated with DEX. The results of the study suggest that the parasite made an effort to reduce the effectivity of the host immune response in order to ensure its own survival.

Morphological and biochemical changes in the blood of horses naturally infected with Gasterophilus sp. larvae.

Gasterophilus sp. constitute a group of specific parasites, which larval forms are found in horses and other phylogenetically related species--donkeys, mules and zebras. Their invasion is associated with marked fluctuations of hematological and biochemical blood parameters, including the activity of many enzymes and the electrolyte concentration. The purpose of the study was to analyze changes in selected biochemical and morphological blood parameters of horses infected with botfly larvae. The experimental group was formed of horses infested by Gasterophilus sp. larvae whereas the control group consisted of noninfested horses. The study was performed in two periods: at the beginning and at the most advanced invasion. Significant differences between horses at early and late stage of invasion were found in terms of erythrocyte parameters. Mean red and white blood cell counts and mean hemoglobin concentration were lower in horses with late invasion compared to those with the invasion at its early stage. The values of all the aforementioned erythrocyte parameters remained within the reference limits for both groups of horses. Moreover, both in horses infected with botfly larvae and in those from the control group, biochemical parameters studied did not exceed the respective reference limits. Our results suggest that larvae of botfly which colonize the gastrointestinal tract have relatively insignificant influence on the basic hematological and biochemical parameters of horse blood.

Accidental discovery of Trypanosoma theileri in the in vitro culture of the heifer lymphocytes.

The diagnostics of the Trypanosoma sp. invasion by means of the classic methods i.e. the methods of thin smears or thick drop or even the microhematocrite method, especially when intensity of infection is low, is very difficult. In our climatic zone, trypanosomosis is usually considered as an exotic disease. An opportunistic model of the infection with the parasite and a lack of current data on the prevalence of T. theileri in the cattle in Poland cause that it is neglected as a potential reason of contamination of tissue cultures in cattle. We showed the presence of T. theileri in culture of isolated lymphocytes from one of six heifers examined. It seems that the prevalence of the invasion of the parasite is not very intense but it should be considered as a possible threat for bovine cell culture. It is also worth including this parasitosis in the differential diagnostics of other diseases that are infectious and/or proceed with symptoms of immunosuppression.

Effect of phytohaemagglutinin-P on apoptosis and necrosis in Trichinella spiralis infected mice.

Flow cytometry analyses were used to evaluate the contribution of apoptotic and necrotic lymphocytes in the selected organs of Trichinella spiralis infected mice treated with phytohaemagglutinin-P (PHA-P). The Tunnel method was used to examine apoptosis in a cryostat section from the jejunum and masseter muscle. CFW mice (Groups I and II) were infected with 200 larvae of T. spiralis. PHA-P was administered intravenously at a dose of 10mg/kg 24h prior to infection in Group II mice only. Group III mice were treated with PHA-P without T. spiralis infection, and Group IV mice were untreated controls. The lymphocytes obtained from the spleen, mesenteric lymph nodes (MLN) and muscular inflammatory infiltration on 7, 14, 21, 28, 35, 42 and 60 days post infection (DPI) were incubated with the Annexin-V-Fluos Staining Kit (Roche). The cryostat preparation made from the jejunum and masseter muscle was evaluated using a fluorescence microscope. PHA-P administration stimulated apoptosis in the jejunal mucosa and in the muscular inflammatory infiltration. In Group I mice, infected with T. spiralis only, the highest percentage of apoptotic cells was found on 7 DPI in the spleen and in MLN, and on 14 DPI among the cells of the muscular inflammatory infiltration. The peak of the necrotic lymphocytes was found on 7 DPI in the spleen, on 28 DPI in MLN, and on 21 DPI in the cells of muscular inflammatory infiltration. In Group II mice, infected with T. spiralis and treated with PHA-P, the peak in apoptotic cells occurred on 7 DPI in the spleen and in the muscular inflammatory infiltration. The highest level of necrotic lymphocytes was observed only on 7 DPI in the muscular inflammatory infiltration. Percentage of necrotic lymphocytes in the spleen was the same and in MLN it was lower than in Group I (T. spiralis only). Moreover, the number of muscle larvae in mice treated with PHA-P (Group II) was lower than in Group I (T. spiralis only).

Differential effect of IFN-tau on proliferation and distribution of lymphocyte subsets in one-way mixed lymphocyte reaction in cows and heifers.

IFN-tau is a signaling protein secreted by the bovine conceptus during the peri-implantation period and responsible for pregnancy recognition. Its main role is the prevention of pulsatile release of luteolytic PGF2alpha, but it also exerts immunomodulatory activities characteristic for other type I interferons. The aim of the study was to examine the effect of IFN-tau on the proliferation and distribution of peripheral blood lymphocyte subsets during one-way mixed lymphocyte reaction (MLR) in cows and heifers. IFN-tau inhibited the proliferative response of lymphocytes in MLR both in cows and heifers in a dose-dependent manner, but cow lymphocytes were less susceptible than those ones from heifers. It was also showed that IFN-tau differentially changed lymphocyte subsets distribution in MLR in cows and heifers. In cows, the relative percentage of CD8(+) cells after MRL in the presence of IFN-tau was significantly lower than in heifers. Differential effect of rIFN-tau on proliferation and lymphocyte subsets distribution in a one-way MRL in cows and heifers indicated that the age of the mother is an important factor in immunomodulatory effect towards developing bovine embryo.