Background and Aim: Brucellosis remains one of the most common zoonoses. The current anti-brucellosis measures are largely deemed ineffective due to a lack of specificity of conventional serological tests. This study evaluated the use of Brucella outer membrane protein (Omp)19 for serodiagnostic testing. Materials and Methods: The antigenicity of recombinant Brucella Omp19, Omp25, and Omp31 was examined in serum samples from mice and rabbits immunized with Omp19 or Brucella abortus 19 whole cell (WC) and 12 and 152 cows experimentally or naturally infected with brucellosis, respectively. Serum samples were collected from 151 cows that were vaccinated with B. abortus 19 and 12 unvaccinated heifers that were maintained on a brucellosis-free farm. Results: Immunization with Omp19 resulted in antibody production in mice after a single injection without the use of adjuvant. Serum antibodies obtained from rabbits immunized with inactivated B. abortus strain 19 WC targeted Omps by enzyme-linked immunosorbent assay (ELISA) and Western blot. Antibodies targeting Omp19 were identified in all B. abortus strain 544 experimentally infected cows at day 14 post-inoculation (p.i.); Omp25 was detected by ELISA at day 28 p.i., while an ELISA targeting Omp31 was negative for 25% of cows at this time point. Omp19 and Omp25 were readily detected by sera from cows from a new epizootic focus. Antibodies recognizing Omps were also detected in >50% of the animals maintained in a brucellosis-free herd at 10 months after vaccination. Conclusion: Brucella Omp19 in combination with Omp25 and Omp31 may be utilized as target antigens in an ELISA designed for serological testing of unvaccinated cattle.
The aim of this study was to evaluate the serological diagnostic potential of the Brucella recombinant outer membrane (rOMP25, rOMP31) and periplasmic proteins (rBP26, rSOD) in a comparative way using an indirect enzyme-linked immunosorbent assay (i-ELISA). Rabbit and/or mouse antibodies to Brucella whole cell and/or soluble protein preparations recognized all recombinant proteins used, which confirms the expression of target antigens in E. coli in active form. The recombinant proteins showed different antigenicity to antibodies of cattle kept on a brucellosis-affected (endemic) farm and/or a new focus of infection. Thus, the presence of anti-Brucella antibodies was confirmed by i-ELISA/rSOD in 79% of cows from endemic conditions with positive results by conventional serological tests (RBPT and/or CFT). However, antibodies specific to this protein were detected in only 14% of seropositive animals kept in the hotbed of a new brucellosis infection. Moreover, rSOD-specific antibodies were not detected in the sera of vaccinated cattle from a brucellosis-free farm, whereas antibodies to other recombinant proteins were found in 2%-8% of animals. Using recombinant proteins in immunoassays significantly reduced the number of cows positive for brucellosis. Furthermore, there was not a single protein among the rOMPs that would show the total positive results of all proteins used. Thus, the development of reliable ELISA tests for the diagnosis of brucellosis requires further comprehensive study of the recombinant proteins in order to design a multiprotein antigen that consists of a combination of several proteins with diagnostic potential.
Monoclonal antibody specific for an epitope of cretory-secretory antigen protein of Opisthorchis felineus (Rivolta, 1884) (Trematoda: Opisthorchiidae) with a molecular weight of 28 kDa was used in a sandwich enzyme-linked immunosorbent assay (ELI-SA) for immobilisation of liver fluke specific antigen to the solid phase. Examination of human sera by this ELISA compared with commercial assays demonstrated that the monoclonal antibody epitope is located within this significant parasite protein. Anti-idiotypic antibody specific for the paratope of this monoclonal antibody was obtained by a hybridoma technique. Mimicking an epitope of excretory-secretory antigen of O. felineus, it had the capacity to bind specific antibody and elicit an antibody response. The value of anti-idiotypic antibody as a substitute for the liver fluke antigen was tested by ELISA using serum samples of infected dogs. Anti-idiotypic antibody proved to be of value in both an indirect-ELISA and a competitive-ELISA for diagnosis of opisthorchiasis. Mature trematodes were isolated from all infected animals. The faecal egg counts were negative in dogs with a relatively small number of parasites, despite finding antibodies in serum by ELISA. Substitution of parasite antigen with anti-idiotype avoids the use of experimental animals and also reduces time-consuming steps of antigen preparation.
Aim of the present study was to provide presence of opisthorchiid metacercariae in cyprinid fish Leuciscus idus in Nura-Sarysu river, Kazakhstan. Infection rate of the ides by the metacercariae was 42%. The metacercariae, similar morphologically to those of the liver flukes, were found: elliptical in shape, 0.19–0.25×0.15–0.22 mm, oral and ventral suckers nearly equal size, and excretory bladder O-shape with black content, occupying posterior part of the body. The metacercariae were divided into 2 groups with differences in size and thickness of cyst wall. Adult flukes were recovered from the Syrian hamsters infected with the opisthorch metacercariae and identified with morphological characters to Opisthorchis felineus and Metorchis bilis. DNA sequences of ITS1, ITS2, and cox1 supported the taxonomic assignment.
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