Passive Hemagglutination Test Research Articles

Development of novel Streptococcus equi vaccines with an assessment of their immunizing potentials and protective efficacies

Strangles is a highly contagious disease of the equine upper respiratory tract caused by Streptococcus equi subspecies. Streptococcus equi subsp. equi (S. equi) and Streptococcus equi subsp. zooepidemicus (S. zooepidemicus) was isolated, as local, hot, and field strains, from horses clinically suffering from respiratory distress. The isolated Streptococci were identified using bacteriological and molecular techniques. Four formulations of inactivated S. equi vaccines were developed and evaluated. The first formulation was prepared using the S. equi isolates, adjuvanted with MONTANIDE GEL adjuvant, while the second formulation was adjuvanted with MONTANIDE ISA-70 adjuvant. The other 2 formulations were inactivated combined vaccines prepared from both S. equi and S. zooepidemicus isolates. The 3rd formulation was the combined isolates adjuvanted with MONTANIDE GEL while the 4th formulation was the combined isolates adjuvanted with MONTANIDE ISA-70. The developed vaccines’ physical properties, purity, sterility, safety, and potency were ensured. The immunizing efficacy was determined in isogenic BALB/c mice and white New Zealand rabbits using the passive hemagglutination test. Also, the antibodies’ titer of the combined S. equi and S. zooepidemicus vaccine adjuvanted with MONTANIDE ISA-70 in foals was tracked using an indirect enzyme-linked immunosorbent assay. The protective efficacy of the developed vaccines was determined using a challenge test in both laboratory and field animal models, where a 75% protection rate was achieved. The combined vaccine proved to be more efficacious than the monovalent vaccine. Also, the MONTANIDE ISA-70 adjuvant provided significant protective efficacy than the MONTANIDE GEL. The current work is introducing a very promising mitigative and strategic controlling solution for strangles.

In ovo Vaccination in Ducks against Aflatoxin-Ovalbumin Conjugate

Background: Aflatoxicosis is a major disease affecting ducklings especially due to their high susceptibility during the post-natal stage. There is no vaccination schedule for this disease as aflatoxins are haptens and incapable of generating immune response by itself. Methods: In this study to combat this malady, in ovo vaccination with an aflatoxin-chicken ovalbumin conjugate was done on 25th day of incubation to generate adequate immune response during the post-natal period of ducklings. An isolate of Aspergillus flavus was used for harvesting of the aflatoxin. The purified crude toxin was checked by TLC to observe the presence of all 4 isomers. Chicken ovalbumin was used as carrier protein due to its similarity with duck ovalbumin as well as to avoid autoimmunity. The conjugate was then encapsulated with chitosan to prevent clearance of the antigen as well as to provide a balanced Th1/Th2 response. A booster dose of the conjugate was given on 14th day post hatch along with sheep red blood cells. The antibody titre was checked by passive haemagglutination test. Result: The results seen i.e., Log2 antibody titre of 1.33±0.21 proved that an immune response was generated. This could provide protection to the ducklings against aflatoxins. Also, this is the first time in ovo vaccination in ducks with aflatoxin-chicken ovalbumin conjugate has been carried out.

Protection of Khaki Campbell Ducks against Duck Plague Using an Inactivated Duck Plague Vaccine

Duck plague (DP) or duck viral enteritis is a fatal viral disease of ducks that causes huge economic losses in the duck industry. The present study was performed to determine the immune response and protective efficacy of an inactivated DP vaccine prepared from a local virulent DP virus. A virulent DP virus was obtained from the laboratory repository of the Department of Microbiology and Hygiene, Bangladesh Agricultural University, Mymensingh (Bangladesh). The DP virus (EID50 105.3/ml) was inactivated using 0.04% formalin. The alum (40 g/L) was added to the inactivated DP virus as an adjuvant. A total of 60 Khaki Campbell male ducks aged 17 weeks were randomly divided into three groups. Ducks of groups A (n = 20) and B (n = 20) were vaccinated intramuscularly in the breast muscle with 1 ml of inactivated DP vaccine and a live attenuated DP vaccine, respectively. Ducks of group C (n = 20) were kept as unvaccinated control. Booster vaccination was administered at 2 weeks after primary vaccination. Antibody titers of vaccinated ducks were measured at 7, 14, 21, and 28 days post-vaccination (DPV) using a passive haemagglutination (PHA) test. Ducks of both vaccinated and unvaccinated groups were challenged with 1 ml virulent DP virus (EID50 104.3/ml) at 28 DPV. Clinical signs, morbidity and mortality, and gross pathological lesions of vaccinated and control ducks were observed for 10 days post-challenge to evaluate the protective efficacy of inactivated DP vaccine. The mean PHA antibody titers of vaccinated ducks of group A at 7, 14, 21, and 28 DPV were 5 ± 0.43, 26 ± 1.71, 43 ± 3.4, and 54 ± 3.28, respectively. Ducks in group B had mean serum PHA antibody titers of 21 ± 1.71, 41 ± 3.28, 52 ± 3.41, and 84 ± 7.25 at 7, 14, 21, and 28 DPV, respectively. No mortality or gross pathological lesions were observed in vaccinated ducks after they were subjected to a challenge infection. Additionally, no significant difference was observed between groups A and B in terms of the challenge infection. The mortality rate of the control group of ducks was 70%. Hemorrhage in the trachea and intestine and necrotic foci in the liver were seen in unvaccinated control ducks (group C). Experimentally developed inactivated DP vaccine induced a protective serum antibody titer and conferred 100% protection against virulent challenge infection up to 10 days observation period.

Аgglutination test and passive hemagglutination test as immunological methods of antigenisisty assessment of vaccines against poultry salmonellosis

Salmonellosis ranks first among acute intestinal infections with an established pathogen in Ukraine. Poultry products are the most frequent source of human salmonellosis is the highest in both the world and Ukraine. Therefore, Salmonella-specific prevention measures in most countries are an issue of great importance. The study's goal was to compare the levels of antibodies against Salmonella to the enteritidis, Typhimurium, and infantis monoantigens in the serum of poultry immunized with the experimental bivalent, the trivalent vaccine against salmonellosis using the Agglutination test (AT) and Passive Hemagglutination test (PHA). The research of two Salmonella inactivated emulsified vaccines – bivalent (S. enteritidis) and trivalent (S. enteritidis, S. typhimurium and S. infantis) was conducted on one of the poultry farms of the Lviv region. The birds were vaccinated two times in the 14th day's interval. The sera samples were taken on the 14, 21, and 28th days after the last shot. The antibody (agglutinin) levels were assessed in the AT and PHA. Both vaccines (bivalent and trivalent) inducted an intense immune response in birds' organisms. The average antibody titers were 1 : 512 –1 : 717 using the AT and 1 : 4096 – 1 : 5734 using the PHA. The highest antibody levels were detected for S. Enteritidis: 1 : 5734 for trivalent vaccine and 1 : 5120 for bivalent one using PHA. AT antibody levels were lower: 1 : 717 for the trivalent vaccine and 1 : 640 for the bivalent. The antibody levels to S. typhimurium were: 1 : 4915 for trivalent vaccine and 1 : 4710 for bivalent (PHA) and 1 : 640 for trivalent vaccine 1 : 589 for bivalent on (AT). The lowest antibody levels were detected in S. Infantis in both tests. An interesting fact of S. infantis agglutinins presence in serum samples from poultry vaccinated with the bivalent vaccine in both tests may be explained by cross-immunity formed by S. enteritidis – S. typhimurium vaccine.

Evaluation and Correlation of Rabies Vaccine Potency Using the National Institute of Health, Rapid Focus Fluorescent Inhibition, and Passive Hemagglutination Tests.

Rabies vaccine preparations are quantitatively assayed for potency using the in-vivo challenge National Institute of Health (NIH), the main test that consumes a high number of animals, takes a long time, and has wide variability. The Rapid focus fluorescent inhibition (RFFIT) and the passive hemagglutination (PHA) tests, the two serologically based tests, were also used for such purpose. In this study, we aimed to evaluate and correlate the potency of the NIH, RFFIT, and PHA tests according to the World Health Organization (WHO) validity criteria, aiming to validate the use of RFFIT or PHA test as a substitute to the NIH test for determining the potency of commercially available Rabies vaccine preparations. The results showed that, the three tests can be successfully used; however, a higher correlation between RFFIT and NIH than PHA and NIH was recorded (Pearson correlation = 1). The potency of rabies vaccine preparations using NIH, RFFIT, and PHA were 3.73, 3.51, and 4.50, respectively. NIH is the main test for the determination of vaccine potency carried out by conducting 25 experiments and consuming about 5,000 mice compared to 1,200 mice used with RFFIT and 1,000 mice used with PHA test. Taken together, we concluded that (i) in some tested preparations, both RFFIT and PHA tests gave comparable results, and they can be used interchangeably; (ii) RFFIT could successfully replace NIH test, but not PHA; (iii) RFFIT and PHA tests are faster, more accurate, more economic, and more sensitive than NIH; nevertheless, PHA needs further investigations; and (iv) both RFFIT and NIH tests complement and reinforce each other as they provide a comprehensive picture of the product potency.

Immune Responses Induced in Chickens by Capsular Extract of P. Multocida Isolated From Farm Rat

An experiment was conducted to investigate the immune response induced in chickens by capsular extract of Pasteurella multocida isolated from rats wandering in and around the poultry farms. The rat isolate of P. multocida was isolated and identified by cultural, morphological, and biochemical characteristics, followed by capsular extract preparation and experimental vaccine development. The isolated P. multocida was found Gram-negative, non-motile, non-spore forming rod occurring singly or pains and occasionally as chains or filaments in Gram’s-staining method. The isolates consistently produced acid from dextrose, sucrose and mannitol but not fermented maltose or lactose. The Capsular antigen was extracted and confirmed by acriflavine test. Finally, experimental fowl cholera vaccine was prepared. Primary vaccination was performed at the dose rate of 5.6×107 CFU/ml through intramuscular and subcutaneous routes in birds of group A (10 birds) and group B (10 birds) and group C (10 birds) were control birds. Secondary vaccination was similarly performed after 15 days of primary vaccination in groups A and B. The levels of pre-vaccination and post-vaccination sera were determined by passive haemagglutination test. The passive haemagglutination antibody titre was recorded on 15 and 35 days of post vaccination in groups A and B. It was demonstrated that experimental capsular extract fowl cholera vaccine conferred 100% protection (p<0.01) against challenge infection and found to be safe. It could be suggested that after thorough field trial, the experimentally prepared capsular extract FC vaccine using rat isolate of P. multocida may be used side by side with conventional FC vaccine.
 Res. Agric., Livest. Fish.8(1): 117-124, April 2021

Acid-base balance, blood gases and haematobiochemical profiles in camels (camelus dromedarius) with trypanosomosis

This study was carried out to investigate the status of acid-base balance and blood gases in camels with trypanosomosis compared to healthy camels. The haematobiochemical profiles were also reported in both groups. Forty-two camels with chronic weight loss, ventral oedema and ascites were examined. Passive haemagglutination test showed that 38 of the 42 camels (90%) were positive for Trypanosoma evansi. Compared to a value of 7.54+0.16 in healthy camels, the blood pH in diseased camels was 7.37+0.051. The partial pressure of carbon dioxide (PCO2) was higher in camels with trypanosomosis than healthy camels. On the contrary, the oxygen partial pressure (PO2) was lower in camels with trypanosomosis. The base excess (BE) was also lower in diseased camels than healthy ones. Similarly the bicarbonate (HCO3) was lower in diseased camels. In a similar pattern, the total carbon dioxide (TCO2) was lower in diseased than healthy group. The oxygen saturation (SO2) decreased significantly in camels with trypanosomosis when compared to healthy group. Concerning the haematological parameters, leukocytosis, neutrophilia and lymphopenia was found in diseased camels. The RBCs count, haemoglobin and haematocrit decreased significantly in camels with trypanosomosis. Concerning, the biochemical parameters albumin and phosphorus decreased significantly and globulin and magnesium increased significantly in diseased camels. The serum activity of alkaline phosphatase, y-glutamyl transferase and creatine kinase increased significantly in diseased camels compared to healthy camels. In conclusion, camels with trypanosomosis have metabolic acidosis, and the HCO3 was lower than healthy camels. The PCO2 was higher, while PO2, BE, HCO3, TCO2 and SO2 were lower in camels with trypanosomosis compared to healthy camels.

Histological analysis of morphostructural changes in bronchial and lung tissues in cattle fetuses during chlamydia infection in Perm Krai and Tyumen Oblast of the Russian Federation

Between 2017 and 2019, 543 cattle (cows` aborted fetuses, stillborn calves) were studied in Perm Krai and Tyumen Oblast of the Russian Federation for the existence of chlamydia infection. Blood serum samples and fingerprints were analyzed from the animals, and a positive result was registered in 48 animals, which is about 9% of the population. Chlamydia was diagnosed on the basis of clinical signs of disease, passive hemagglutination test and microbiological studies. From 23 aborted and stillborn fetuses, biological material was taken for electron microscopic examination. In order to study pulmonary tissue injuries under the influence of chlamydia, a histological analysis of bronchi and lung aborted and stillborn calves was carried out. All three components of the mother-placenta-fertility system were found to be affected by bovine chlamydia. A characteristic pathological-anatomical sign of chlamydia in fetal death is the formation of common swelling, both tissue and cavity localization. Histological examination of organs and tissues traces changes in the general pathological nature of the developing vascular level, alternative processes, immunopathological reactions, systemic inflammatory changes in histochemical barriers. Morphological changes in the pulmonary tissue of the fetuses of different gestation terms indicate damage to the aero-hematic barrier with the possibility of respiratory disorders in the uterine period.

Measles immunity in different population groups

Objective: to examine the state of the immunity to measles in different age groups.Materials and methods: In 2018, 4444 people were examined at the Diagnostic Center (virological). Among them, 3783 people were examined using the passive haemagglutination test for measles (manufactured by Pasteur Research Institute of Epidemiology and Microbiology, Russia). In the remaining 661 cases, the IgG to measles were detected using the enzyme-linked immunosorbent assay (ELISA) by VektoMeaseles IgG test (manufactured by Vector-Best, Russia). The correlation between the measles IgG level (ELISA) and the age was examined in 518 patients. Results: In this study, the immunity to measles was shown to be insufficient in all groups of observed people. Even among medical staff, nearly 10% had no protective level of measles antibodies. We have shown that the correlation between the measles IgG level and the age is statistically significant, so that the number of seronegative persons in different age groups differs significantly. Conclusion: The highest ratio of seronegative individuals was found in the age group between 18 and 25 years (52,33%), which can lead to serious measles outbreaks. Hence, this study confirms a strong need for additional immunization in all groups and especially in young population.

Immunopathological study of the fungus cryptococcus neoformans in mice

The study aimed to investigate the effect of cryptococcal sonicated antigen and chitosan on the infection with this fungus which isolated from human skin lesions. Isolate of Cryptococcus neoformans was used to prepare the sonicated antigen, also 60 white mice from both sexes, at average age 8-10 weeks were getted and divided randomly into 4 groups equally and treated as following: First group: fifteen mice were immunized with 0.5ml of whole sonicated Ags(Wsc Ag) (protein concentration14mg/ml) subcutaneous two doses, one week interval.Second group: fifteen mice were fed diet supplemented with chitosan 1g/kg diet for 6 weeks.Third group: Mice were immunized as 1st group and at the same time fed diet supplemented with chitosan as 2nd group. Fourth group: Mice, which seted as control positive group.
 At day 28-30 post-immunization, delayed hypersensitivity and indirect passive haemagglutination tests were done, then 1st, 2nd, 3rd and 4th groups were administrated intraperitoneally with 0.3 ml of fungal suspension containing 1X107 fungal cells, all animals were sacrificed at day 30 post-infection. The results showed that the immunized animals and fed diet containing chitosan had values of delayed hypersensitivity test(1.99±0.02 at 24hours and 2.11±0.03 at 48 hours)and value of antibodies(160±29.6) higher than that of immunized animals only(1.75±0.03 at 24 hours and 1.81±0.03 at 48 hours)and value of antibodies(136±28.2).The immunized animals with whole sonicated cryptococcal antigens (1st group) expressed high values of delayed type hypersensitivity and high levels of Abs titer, as well as the chitosan stimulate the immune system in the second group and the best results of immunization were getted in the third group while all animals of control positive(fourth group) were died at the first week post-infection.
 The current study showed that all the non-immunized and infected animals of the fourth group were died during the first 2 weeks post-infection with severe fungal cells isolated from internal organs and severe suppurative inflammatory reaction in the examined organs during early stage of infection with granulomatous lesions appear at day 14 post-infection while no mortality rate was seen in animal fed diet supplement with chitosan with or without immunization(2nd and 3rd groups) moderate fungal isolation and granulomatous lesions were recorded in the examined organs of animals sacrificed at day 30 post-infection while the immunized animals and fed diet containing chitosan showed that there is no fungal isolation and nor fungal cells in the histological sections with few histological degeneration and small granulomatous lesions and sometimes absence of them in the examined organs. It concluded that cryptococcal sonicated antigen and chitosan have important role in protective immunity against C.neoformans infection.

Clinical and epidemiological characteristic of chronic brucellosis

Aim. To study clinical epidemiological and laboratory features of chronic brucellosis in the Republic of Tatarstan in ten-year aspect. Methods. 59 patients infected with various forms of brucellosis in 2007-2017 were examined. Clinical laboratory and instrumental diagnosis of brucellosis was confirmed by the immunoassay (EIA) with determination of IgM and IgG antibodies, passive hemagglutination test with a brucellar diagnosticum, Coombs test, Wright and Hedelson agglutination test. Results. Clinically 91 % of patients had asthenic-vegetative syndrome, 55 % - mild intoxication symptoms, 89 % - articular syndrome, 49 % - fibrositis. EIA revealed in 91 % of patients IgG (38 %) and IgM (53 %) antibodies to causative agents of brucellosis, 25 % of patients had positive Wright agglutination test, and 30 % - positive Hedelson agglutination test. In 9 % of cases the diagnosis was confirmed by Coombs test and in 26 % by passive hemagglutination test with brucellar diagnosticum. The retrospective analysis with clinical cases of patients with chronic brucellosis indicates introduced cases in 19 % (from the republics of Central Asia and Transcaucasia), local cases in 81 % (from the Republic of Tatarstan), their occupational character (57 %), the mixed (contact and alimentary) route of infection (21 %), and 64 % with clinically primary involvement of the musculoskeletal system and peripheral nervous system, i.e. prevalence of the mixed form of chronic brucellosis. Conclusion. Chronic brucellosis in the Republic of Tatarstan is characterized by high risk of introduced cases, occupational history, prevalence of the mixed route of infection in females and working-age patients; with the features of systemic disease involving the musculoskeletal and peripheral nervous system against the background of mild syndrome of intoxication and moderate asthenic-vegetative syndrome. Divergence of the results of serological diagnostics requires careful studying of duration of infection, features of the immune response in each case on follow-up.

Comparative efficacy of BAU-fowl cholera and DLS-fowl cholera vaccines in indigenous chicken

The present study was conducted to determine the immune response induced in indigenous chicken produced against BAU-FC and DLS-FC vaccines with their efficacy study against Pasteurella multocida. A total of forty (40) chickens were selected and divided into Group A (15), Group B (15) and Group C (10). Group A and B were vaccinated with BAU-FCV and DLS-FCV, respectively at the dose rate of 0.5 ml through SC at six weeks of age followed by boostering at 10 weeks of age while Group C was kept as unvaccinated control. Sera samples were collected after primary and booster vaccination and antibody titre was determined by Passive hemagglutination (PHA) test. The mean PHA titres recorded at 4 weeks after primary vaccination was 51.20 ± 7.84 in birds of group A and 38.40 ± 6.40 in birds of Group B. After booster vaccination, mean PHA titer was found 140.80 ± 31.35 at 16 weeks of age in case of BAU-FC vaccinated group and 115.20 ± 12.80 in case of DLS-FC vaccinated group. The mean PHA titer was 204.80 ± 31.35 and 179.20 ± 31.35 at 19 weeks of age in birds of BAU-FC and DLS-FC vaccinated group, respectively. Birds of all groups were challenged with virulent P. multocida at 17 weeks of age. It was observed that vaccinated chickens showed maximal resistance (100%) following challenge with virulent whereas unvaccinated control birds failed to resist the challenge infection. It can be assumed from the findings of present research work that both BAU-FCV and DLS-FCV are able to protect indigenous chicken from the outbreak of avian pasteurellosis and BAU-FV vaccine showed relatively higher immuno-protective titre than that of DLS-FC vaccine.Res. Agric., Livest. Fish.5(2): 193-199, August 2018

STANDARDIZATION OF EFFECTIVE DOSE OF FOWL CHOLERA VACCINE IN PIGEON IN BANGLADESH

An experiment was conducted to determine the effective dose of formalin killed (FK) fowl cholera (FC) vaccines prepared with virulent avian Pasteurella multocida (PM 38) serotype 1 (X-73) collected from the laboratory of the Department of Microbiology and Hygiene, BAU, Mymensingh. To determine the effective dose of vaccine, 7 weeks old 30 pigeons were immunized and each group consists of 5 birds. The groups are represented by A, B, C, D, E and F. The birds belonging to groups (A-E) were vaccinated with different doses of vaccine, after two weeks of first, second immunization and challenge experiment, blood was collected from all vaccinated birds, and serum was analyzed to determine antibody titer against P. multocida by passive hemagglutination test (PHA). The PHA titer after two weeks of first vaccination were 16±3.92, 17.6±3.92, 25.6±3.92, 32±8.76, 35.2±7.84 of group A,B,C,D and E, respectively at the dose of 0.2ml (0.26×108 CFU)/birds, 0.4ml (0.5×108 CFU)/birds, 0.8 ml (1.04×108 CFU)/birds, 1ml (1.3×108 CFU)/birds, respectively. The PHA titer of prevaccination and control birds was <4. The PHA titer after 2 weeks of second vaccination or boostering were 32±8.76, 35.2±7.84, 44.8±7.84, 57.6±18.66, 70.4±15.68, of group A,B,C,D and E, respectively. After 2 weeks of challenge infection, the mean PHA titer were 44.8±7.84, 51.2±7.84, 70.4±15.68, 102.4±15.68 and 140.8±31.34 of group A,B,C,D and E, respectively. In this experiment, the antibody titer of the vaccinated pigeons with 0.4, 0.6, 0.8 and 1ml per bird via intramuscular route were higher than that of the pigeons vaccinated with 0.4ml/bird, 0.6ml/bird, 0.8ml/bird and 1ml/bird were satisfactory in terms of protective potential against P. multocida. For prevention and control of avian pasteurellosis 0.4ml to o.6ml (0.52×108 CFU to 0.78×108 CFU)/birds of vaccine may be used instead of 1ml (1.3×108 CFU)/birds for better immunization of pigeon against fowl cholera infection.

Description of Hemoproteins and Elemental Homeostasis in Brain Tumors

The role of element homeostasis in neoplastic disease pathogenesis is be-yond question. The imbalance of trace elements precisely underlies the ini-tiation and promotion of tumor pathology. The aim of the study was to in-vestigate blood and tissue macroelements, microelements and hemoproteins level in brain tumors and their intermolecular interactions. Samples of blood and brain tumor tissues were investigated. Detection of myoglobin level was implemented by the reaction of passive hemagglutination and immunoturbidimetric test. Catalase activity was determined by the method of Beer and Sizer. Free radical activity was determined by the method of induced biochemiluminescence. Microelements level was investigated by usage of atomic emission spectrometry. To build the networks of studied hemoprotein interactions with signaling pathways of proteins, expressed in brain tumors, molecular interaction databases (STRING, BioGrid) were used. Modern databases of signaling pathways (KEGG) suggest that in normal cells hypoxia can lead to HIF-1A protein synthesis. ROS synthesis inhibits the PHD enzyme and triggers the release of calcium ions, and increases proliferation. Calcium ions are triggering factor of apoptosis and cell proliferation. Myoglobin can possibly be the cell adaptation factor towards hypoxia, oxidative stress and element homeostasis violation, and myoglobin level decreasing can additionally stimulate proliferation, by apoptosis inhibition.

抗白血球抗体スクリーニングに関する研究 : 1 間接血球凝集反応による抗 HLA 血清の検討

抗白血球抗体スクリーニングに関する研究 : 1 間接血球凝集反応による抗 HLA 血清の検討

Using standard serology blood tests to diagnose latent syphilis

Goal. To conduct a comparative assessment of the results of regulated serological tests obtained as a result of blood tests in patients suffering from latent syphilis. Materials and methods. The authors examined 187 patient medical records with newly diagnosed latent syphilis in FGBU GNTsDK (State Research Center for Dermatology, Venereology and Cosmetology), Health Ministry of the Russian Federation, in 2006-2015. The results of patient blood tests were analyzed with the use of non-treponemal (microprecipitation test/RPR) and treponemal (passive hemagglutination test, immune-enzyme assay (IgA, IgM, IgG), IFabs, immunofluorescence test and Treponema pallidum immobilization test) serology tests. Results. According to the results of blood tests of latent syphilis patients, the largest number of positive results was obtained as a result of treponemal serology tests such as immune-enzyme assay (100%), passive hemagglutination test (100%) and IFabs (100%). The greatest number of negative results was observed in non-treponemal (microprecipitation test/RPR) serology tests: in 136 (72.7%) patients; evidently positive results (4+) test results were obtained in 8 (4.3%) patients only. According to the results of a comparative analysis of blood tests in patients suffering from latent syphilis obtained with the use of treponemal serology tests, the greatest number of evidently positive results (4+) was noted for the passive hemagglutination test (67.9%). Negative treponemal test results were obtained with the use of the immunofluorescence test and Treponema pallidum immobilization test (21.9% and 11.8% of cases, respectively). Moreover, weakly positive results prevailed for the immunofluorescence test: in 65 (34.7%) patients. Conclusion. These data confirm that the following treponemal tests belong to the most reliable ones for revealing patients suffering from latent syphilis: immune-enzyme assay, passive hemagglutination test and IFabs.

Efficacy of Indirect Enzyme Immunosorbent Assay and Passive Haemagglutination Test for the Diagnosis of Bovine Herpes Virus-1 (BHV-1) Infection

A total of 255 sera were collected from unvaccinated crossbred (174) and indigenous (81) cattle and buffalo having history of respiratory and reproductive disorders. All the sera sample were subjected to indirect ELISA and passive haemagglutination (PHA) test for the detection of BHV-1 antibodies. Indirect ELISA and PHA test detected significantly (P<0.01) varied prevalence of BHV-1 antibody i.e. 60.39% and 35.69%, respectively. The sensitivity and specificity of PHA test in comparison with indirect ELISA was 55.19% and 94.06%, respectively. Moderate agreement (kappa = 0.442) was noticed between ELISA and PHA for the diagnosis of BHV-1 antibodies. Significantly (p< 0.01) higher prevalence of BHV-1 antibodies was observed in crossbred (70.11%) as compared to indigenous cattle and buffalo (46.91%). Hence, it is be concluded from present investigation that the indirect ELISA could be reliable and sensitive test than PHA test to screen BHV-1 antibody.

Peculiarities of Immunological Structure Formation in Plague Carriers of the Central-Caucasian High-Mountain Natural Plague Focus

Objective of the study is to analyze immunological structure of the mountain souslik, habitant in various zones of natural focality in the Central-Caucasian natural plague focus. Materials and methods . The data on serological investigations conducted on the mountain souslik, within the periods of 1974–1988 and 2001–2004 have been obtained from FGHI “Kabardino-Balkaria Plague Control Station” of the Rospotrebnadzor. Applying passive hemagglutination test and indirect hemagglutination test, with the help of Excel Microsoft Office 2010, evaluated have been 63147 blood survey results. Conclusions. It is established that the highest ratio of the souslik with low antibody titer in blood, which characterizes initial contact with plague infection, is observed at the peak of epizootic activity. In the mountain steppe of the eastern focal part, the apex coincides with June, in the western part, as well as in the Alpine and Subalpine zones of the eastern one – with August. Animals with high antibody titers, i.e. secondarily infected, emerge in vast numbers at the peak of mountain souslik lethality. In the mountain steppe this period falls upon June, i.e. simultaneously with the peak of epizootic activity. In the western part lethality apex is also observed in June, but maximum epizootic activity takes place in August. In Alpine and Subalpine zones of the eastern focal part, peaks of epizootic activity and lethality among the souslik populations concur and fall upon August. Isolation of plague microbe cultures alongside with positive serological tests is reasonably more often observed in the mountain steppe of the river Baksan, than in other areas of natural focality.

MOLECULAR AND BIOLOGICAL CHARACTERISTICS, AND CULTIVATION OF AN ATTENUATED STRAIN 1974-VNIIVViM OF RIFT VALLEY FEVER VIRUS

In recent decades, a threat of spreading Rift Valley fever (RVF) to Asia and Europe raises serious concerns. Within many years, vaccines against RVF were developed in several ways including designing attenuated vaccine versions, inactivated or genetically engineered vaccines. A serious hazard of the RVF agent requires developing sufficiently immunogenic preparations providing both RVF specific prevention and safety precautions in production procedures. Therefore, this work was aimed at molecular and biological characterization of an attenuated RVF virus (RVFV) strain 1974VNIIVViM including search for high-tech cell culture systems for the virus growth and determination of its antigenic and/or immunogenic relationship with a virulent strain Entebbe. The results of the investigations showed that the virus accumulated in continuous cell cultures like saiga kidney (SK), Siberian mountain goat kidney (PSGK-60) and MDVK (calve kidney cell culture) at high titers with infectious (8.53±0.21 lg MICLD50/cm) and antigenic (1:64 to 1:128 in the passive hemagglutination test, PHT) activity levels. We determined optimal regime of the virus growth in BHK-21/13 (Syrian hamster kidney cell culture) using a roller culture method: the culture bottle rotation speed of 12 to 15 r.p.h.; the infection dosage of 0.01 to 0.001 MLD50/cell; the first 12 hours of the culture at pH = 6.4 to 6.8. Further, pH is maintained at 7.2 to 7.6 for 48 to 72 hours of culture at 37 С, which provides the raw virus production at high infectivity (8.5 to 8.9±0.2 lg MICLD50/cm) and antigenicity (1:128 to 1:256 in PHT and 1:625 to 1:3125 in solid phase ELISA) levels. Phylogenetic analysis of the nucleotide sequences of 3 sites in S and/or M segments showed that it was closely related to strains Smithburn and Entebbe allowing their grouping into a separate cluster. Comparative analysis of nucleotide sequences of genes encoding glycoproteins Gn and Gc, and the nucleoprotein N, shows that most of the substitutions in the sequences of the genes of the RVFV strain 1974-VNIIVViM are synonymous and do not alter the primary amino acid sequence, with insertion and deletion missing. In the nucleotide sequence encoding glycoprotein Gn of strain 1974-VNIIVViM we found 8 substitutions relative to a similar sequence of M segment of a virulent strain Entebbe, and 14 substitutions when compared with M segment of strain Smithburn. A lower (as compared with other attenuated strains) amount of significant amino acid substitutions in RVFV antigenic components was revealed; thus, strain 1974-VNIIVViM selection as a source of immunodominant proteins and/or raw virus for vaccine production seems to be well-grounded.

Hepatitis C virus genotypes in Kenya

Hepatitis C virus is a great public-health concern worldwide. Phylogenetic analysis of the HCV genome has identified six different genotypes that have generally been divided into several subtypes. There is very little information on HCV seroprevalence and genotypes in Kenya. To determine the genotypes of HCV circulating in Kenya, blood donor samples were serologically tested and confirmed by polymerase chain reaction (PCR). Positive samples were cloned and sequenced, and phylogenetic analysis conducted to determine the HCV genotypes. One hundred Murex-seropositive samples were re-tested using a passive hemagglutination test, and 16 of these were identified as seropositive. Further testing of all of the samples by PCR identified only 10 of the 16 samples as positive. Thus, only 10% (10/100) of the samples were viremic. Six were from females (60%), and four were from males (40%). The mean age of the positive donors was considerably low, at 25 +/- 9 years. Genotypic testing indicated the presence of genotype 1a (10%) and genotype 2b (90%). This study reports on HCV genotypes in a blood donor population in Kenya where little had been done to provide information on HCV genotypes.