PROTECTIVE EFFICACY OF A COMPLEX BIOPREPARATION BASED ON A SOMATIC EXTRACT OF TRICHINELLA AND IMUNOFAN IN EXPERIMENTAL TRICHINOSIS IN WHITE MICE

The purpose of this placebo-controlled randomized laboratory study was to evaluate the effectiveness of a complex biological product based on the Trichinella somatic antigen (SET) at a dose of 40 μg / mouse and an immunostimulant immunofan at a dose of 1 mg / mouse for T. spiralis larvae with experimental infection with them 40 days after the last injection of drugs. The preparations were administered twice, intramuscularly in 0.2 ml of sterile saline with an interval of 48 hours to 20 animals from 2 therapeutic groups. 10 mice of the control group received a placebo. Mice were experimentally infected with T. spiralis larvae at a dose of 30 ± 5 larvae / mouse orally. After 40-45 days, a control assessment was performed to detect T. spiralislarvae. The number of larvae of the parasite in animals treated with the complex drug was reduced by 61.98%, and in mice treated with an immunostimulant - by 31.42% (calculated from the geometric mean). The largest number of Trichinella larvae was found in the control group that received placebo.According to the results of the study, a conclusion was drawn about the effectiveness of double administration of a complex immunopreparation based on SET-40 and Imunofan, as well as on the prospects and feasibility of continuing further research in search of the most promising immunoprophylactic agents, their doses and frequency of use for trichinosis.

T-Bet Is Dependent on Stat-4 Inhibiting Acute Colitis but Not Stat-1 Using L4 Somatic Antigen of Heligmosomoides polygyrus

Helminths may alter the immunoinflammatory reactions of colitis. Proteins derived from H. polygyrus have prospective therapy for colitis. The goal of this study was to interpret the protective mechanisms of L4 somatic antigen (LSA) from Heligmosomoides polygyrus against an inflammatory response to the pathogenesis of DNBS-induced colitis. Colitis was actuated in mice by rectal instillation of DNBS. The mice were randomly divided into five groups containing control, DNBS alone, and three groups, with different doses of LSA (50, 100, and 200 μg/mL), respectively. Mice initiated colitis by rectal administration of DNBS and after that were immunized with LSA for 14 days. Mice treated with LSA inhibited wasting disease compared with DNBS only group. The percentages of cells producing IFN-γ were reduced by LSA treatment. The level of T lymphocytes CD4+IFN-γ+ cells in the LPL was significantly diminished by LSA at both 100 and 200 μg/mL groups (p<0.05). The mRNA expression of T-bet was significantly declined in LSA immunized mice, but not RORγ-T mRNA, whereas GATA-3 expression tended to increase. The activation of STAT-4 significantly reduced LSA-treated mice but not STAT-1. It can be concluded that T-bet is required for optimal production of IFN-γ in colitis.

Development of an interferon-γ release assay (IGRA) for detection of Brucella abortus and clinical diagnosis of brucellosis

Introduction: Brucellosis, caused by Brucella abortus (B. abortus), is an important zoonosis posing a great risk to both livestock and humans. Currently, most assays for clinical diagnosis of brucellosis have been developed based on serological principles; however, these assays have a number of limitations and disadvantages. Methodology: To address this concern, the aim of this study was to develop a gamma interferon (IFN-γ) release assay (IGRA) for the diagnosis of brucellosis. Towards this end, the stimulating effect induced by different somatic antigens of B. abortus on the secretion of IFN-γ was evaluated. Results: The best antigen candidate, B. abortus strain 2308, able to induce high levels of IFN-γ expression in peripheral blood (PB) cells from cattle, was used for the development of the IGRA. The optimal concentration for stimulation was determined as 1.0×107 CFU/mL. This study demonstrated that IFN-γ was detectable on day 5 post infection (p.i.) and peaked on day 14 p.i.. Finally, the IGRA developed was used for detection of B. abortus in clinical samples, and a higher level of IFN-γ was detected in Brucella-infected samples compared to vaccination samples and negative controls. Conclusions: The optimal somatic antigen for B. abortus was identified and used to establish a robust IGRA. The IGRA developed is suitable for clinical diagnosis of brucellosis, especially in the early stages of infection.

EFFECT OF SOMATIC ANISAKIS SIMPLEX EXTRACT TO DEVELOPMENT CHICK EMBRYOS

Helminthes have embryotoxic and teratogenic effects to host’s cells and tissues.Investigations were carried out mainly on embryo and fetus in mammals.Blaszkowska, J. (1998); Kadlubowski R. (2000) conducted studies of in chickembryos. Dose-dependent effect is also studied and found that small doses causeembryotoxic effect, while higher doses - teratogenic. Anisakis simplex antigenshave also embryotoxic effect has been found. The aim of our research was toexamine the effect of A. simplex somatic antigen to the development of chickembryos in its experimental introduction in various ways and at different stages ofembryogenesis. For the study chicken embryos were used at different stages ofembryogenesis: 7.5, 10.5 and 12.5 days. Methods for introducing antigen chickenembryos: in the allantois cavity, on-chorion allantois membrane and yolk sac at 0.2ml. The autopsy was performed on embryos 2 and 8 days. The eggs and embryoswere weighed and evaluated the embryonic development. As a result of the firstexperience the greatest change observed by mass, and when administered in thedevelopment of antigen in both yolk sac on day 2 (1.68 ± 0.06, unlike control 2.25± 0.20), and at 8 days ( 9.97 ± 6.21 in the experiment versus 14.89 ± 0.78 in thecontrol), respectively. In the second experiment eggs were used at 1 and 5 dayincubation embryos. The antigen was administered in both cases also in the yolksac at a dose of 0.2 ml. In the second experiment there was a delay in thedevelopment of both cases at autopsy after 2 days in non-incubated eggs missingthe development of, unlike the control, the development of which 48 hourscorresponded; including 5 daily observed decrease in weight 0.88 ± 0.22, against1.05 ± 0.05 and 8 days in a similar way. As a result of experiments on the effect ofthe Anisakis simplex somatic antigen on the development of chick embryosembryotoxic action installed. Thus antigen has the greatest effect on developmentof early embryos and putting it into the yolk sac.

Immunodiagnostic potency of homologous antigens for natural Haemonchus contortus infection in small ruminants in plate and paperenzyme linked immunosorbent assay

The objective of this study was to characterize Haemonchus contortus antigens, and to standardize and evaluate indirect plate and dot-ELISA using homologous antigens in the small ruminants. Electrophoretic separation of somatic antigen in reducing condition on 15% polyacrylamide gel resolved into 16 proteins of molecular weight ranging from 14-100 kDa. Two step ethanolic extraction of the supernatant of in-vitro culture of H. contortus yielded excretory-secretory (E-S) antigen/ cathepsin L cysteine proteinase of molecular weight 28 kDa. The animals (Goats=103; Sheep=91) were broadly kept into post-mortem (PM) and faecal examined groups and further sub-grouped based on mono or multiply helminths infection. At many occasion, the somatic antigen found to cross reacts with other helminths parasites thus minimizing the specificity of the selected tests and antigens. There was no any direct correlation between the parasites load and ELISA reactivity pattern. The prevalence rate of haemonchosis was 55.7 (34/61) in goats/ 47.6 (40/84) % in sheep as per PM examination while it was 45.63 (47/103) in goats/ 41.76% (38/91) in sheep and 36.89 (38/103) in goats/ 35.16% (32/91) in sheep using E-S antigen based plate and dot-ELISA, respectively. With E-S antigen, the overall % sensitivity, specificity, positive and negative predictive values of plate-ELISA was 89.74 (goats)/ 80.95 (sheep), 81.25 (goats)/ 91.84 (sheep), 74.47 (goats)/ 89.47 (sheep), 92.86 (goats)/ 84.91 (sheep), respectively while for dot-ELISA it was 66.67 (goats)/ 61.9 (sheep), 81.25 (goats)/ 87.76 (sheep), 68.42 (goats)/ 81.25 (sheep), 80 (goats)/ 72.88 (sheep), respectively, so the tests and E-S antigen can be recommended for the detection haemonchosis in the small ruminants.

15. KONSENTRASI PROTEIN ANTIGEN EKSKRETORI/SEKRETORI DAN SOMATIK PADA Fasciola gigantica DAN Eurytrema pancreaticum The Protein Concentration of Excretory/Secretory and Somatic Antigen from Fasciola gigantica and Eurytrema pancreaticum

The study was aimed at finding out the difference of protein content between excretory/secretory and somatic antigen of Fasciola gigantica as well as between excretory/secretory of Fasciola gigantica and Eurytrema pancreaticum. As many as 32 Fasciola gigantica and 30 Eurytrema pancreaticum were placed into 80 ml Roswell Park Memorial Institute (RPMI) 1640 and incubated for 6 hours. Excretory/secretory as well as somatic antigen was isolated and the concentration of protein was determined, using Lowry method. The result revealed the different of protein concentration. The concentrations of excretory/secretory antigen protein in Fasciola gigantica and Eurytrema pancreaticum were 3.386 mg/ml and 0.128, mg/ml respectively. The concentrations of somatic protein and Fasciola gigantica and Eurytrema pancreaticum were 8.028 mg/ml and 0.534 mg/ml, respectively. It can be concluded that the protein concentration from somatic tissue of Fasciola gigantica and Eurytrema pancreaticum is higher than the protein concentration from excretory/secretory of Fasciola gigantica and Eurytrema pancreaticum. The protein concentration from excerory/secretory antigen of Fasciola gigantica is higher than the protein concentration from excretory/secretory antigen of Eurytrema pancreaticum. ____________________________________________________________________________________________________________________ Key words: Fasciola gigantica, Eurytrema pancreaticum, protein concentation