Direct Hemagglutination Research Articles

Comparison of manual titrations to automated microplate and gel titration assays used in the screening of blood donors for production of Low-Titer O Whole Blood.

The popularity of Low-Titer O Whole Blood (LTOWB) for treating trauma patients requires that donor centers and transfusion services make decisions on what titer testing capabilities to institute and an appropriate titer level threshold. This study compared the titer results determined by four methods to find a rate of agreement. Isohemagglutinin titers were tested on 300 plasma samples utilizing various methods, each determining IgM antibody levels by direct hemagglutination with A1 and B reference cells. The methods used were the Beckman Coulter's PK7300, Immucor's NEO Iris microplate technology, Ortho Clinical Diagnostics (OCD) Vision, and manual titrations. Only 42.7% of the samples tested showed agreement across all methods on ABO isohemaglutinin titer levels and only 32.5% demonstrated "High titer" agreement. Sample agreement was close to 90% if the Immucor method was excluded. At a <1:256 titer level threshold, the pass rate was 94.3% for Immucor, 89.7% for the PK7300, 87.3% for manual testing, and 75.7% for OCD's gel method. Sensitivity and specificity rates at a ± 1 titer level were respectively 100% and 95.4% for OCD's gel, 73.7% and 100% for Immucor, and 100% and 99.6% for the PK7300. Overall method accuracy was 91.7% for Immucor, 90.3% for the PK7300, and 86.7% for OCD's gel method as compared to manual titration. All three automated methods perform comparably to the manual method at a ± 1 titer tolerance level. Based on these comparisons, a titer level of <1:256 would maximize LTOWB production regardless of the method used.

EVALUATION OF IMMUNOTROPIC PROPERTIES OF GLYPROLINE COMPOUNDS IN EXPERIMENTAL HYPERTHYROIDISM

Introduction. Thyroid disease is currently the most common among endocrine pathology. Hyperproduction of thyroid hormones is accompanied by stimulation of humoral reactions, contributing to the development of immune-mediated pathology under certain conditions, which actualizes the need to develop means for correcting neuroimmunoendocrine processes. Gliproline compounds analogous to neuropeptide protein molecules, among which are Thr-Lys-Pro-Arg-Pro-Gly-Pro and Pro-Gly-Pro, can be considered as such agents. The aim of the study. The aim of the study was to study the immunotropic properties of gliproline compounds Thr-Lys-Pro-Arg-Pro-Gly-Pro and Pro-Gly-Pro under experimental hyperthyroidism. Material and methods. The immunotropic properties of the gliproline compounds Thr-Lys-Pro-Arg-Pro-Gly-Pro and Pro-Gly-Pro were studied in white nonlinear male rats. Hyperthyroid status in animals was modeled by intragastric administration of L-thyroxine sodium salt pentahydrate («Sigma»; USA) at 150 μg/kg for 21 days. Laboratory animals were divided into groups (n=10): control group – intact animals (control); animals treated with L-thyroxine sodium salt pentahydrate (hyperthyroidism); rats receiving Thr-Lys-Pro-Arg-Pro-Gly-Pro (selanc) and rats receiving Pro-Gly-Pro intraperitoneally daily for 21 days at doses of 87 and 33 μg/kg/day. Results. The obtained results indicate the presence in gliproline compounds Thr-Lys-Pro-Arg-Pro-Gly-Pro and Pro-Gly-Pro of immune correcting activity manifested under the conditions of experimental hyperthyroidism, with the greatest impact on the humoral unit of immunity, which is confirmed by the presence of statistically significant changes in the cellular parameters of the spleen and titer antierythrocytic antibodies in a direct hemagglutination reaction. Conclusion. Thus, the study of the immunotropic properties of gliproline peptide compounds under experimental hyperthyroidism established the presence of immune-correcting activity in Thr-Lys-Pro-Arg-Pro-Gly-Pro and Pro-Gly-Pro substances.

Properties of lectin from tissue culture of Ungernia victoris and its ultrasonic extraction

The aim was to investigate biomass extracts of U. victoris strain UV-2 tissue culture for the content of lectins, to study their general characteristics and to optimize the method of ultrasonic extraction to obtain a soluble form of the lectin. Methods. Tissue culture method, direct hemagglutination test for detection of lectin activity (LA), determination of lectin carbohydrate specificity, ultrasonic extraction method. The results. Lectin activity was detected in the soluble (supernatant) and insoluble (sediment) fractions of the extract of biomass U. victoris tissue culture. In the soluble fraction LA was significantly less compared to that in the sediment, which indicated the presence of the membrane-bound form of the lectin. LA was characterized by pronounced species specificity: the reaction with mouse erythrocytes was the most intense. The study of carbohydrate specificity revealed a weak affinity for monosugars (galactose and galactosamine) and pronounced suppression of LA in the case of polysaccharides - hyaluronic acid, heparin and mucin. The possibilities of the ultrasound extraction method for separation the detected lectin from the the cell surface in the sediment fraction were investigated and the extraction procedure was optimized. The transition of LA into the soluble phase under the influence of ultrasound depended on the following parameters: the concentration and volume of the extract, as well as the time of exposure to ultrasound. It was established that the LA of the soluble phase doubled compared to the initial one with an extract concentration of 50 mg/ml and a volume of 20 ml under the action of ultrasound 40 kHz/70 W in the interval of 15-45 minutes. Longer ultrasound treatment had a negative effect on LA. Conclusions. Lectin activity of U. victoris tissue culture biomass extracts was discovered for the first time, which can become a promising source of lectin with a wide spectrum of pharmacological properties. The general characteristics of the lectin were given, and its species and carbohydrate specificity was revealed. The lectin was generally characterized and its species and carbohydrate specificity were established. Possibilities of ultrasound extraction were used and the procedure was optimized to obtain a soluble form of the lectin, necessary for its further fundamental and practical research.

Study of immunotropic activity of the valine-containing neuropeptide compound of the glyprolin series under “social” stress

Aim of the study was to investigate the immunotropic activity of neuropeptide compound Pro-Gly-Pro-Val under “social” stress.Material and methods. The study was conducted on nonlinear male rats (6 months old) under conditions of “social” stress, modeled by formation of submissive and aggressive types of animal behavior under conditions of constant sensory contact. The rats were divided into groups: Control I, animals housed alone in a cage and receiving water for injection in an equivalent volume; Control II, groups of stressed animals with aggressive and submissive types of behavior; III groups of stressed rats to which Pro-Gly-Pro-Val at a dose of 100 μg/kg/day; IV groups of stressed rats to which imunofan (Arg-Asp-Lys-Val-Tyr-Arg) at an average therapeutic dose of 0.7 μg/kg/day were administered intraperitoneally. The immunotropic properties of the neuropeptide compounds were evaluated according to the following indices: delayed-type hypersensitivity reaction index, direct hemagglutination reaction antibody titer and leukocytic formula values.Results. Injection of the tested compound Pro-Gly-Pro-Val under study leads to correction of the immune changes occurring against the background of “social” stress, namely, to an increase in the delayed-type hypersensitivity reaction index and the level of anti-erythrocytic antibodies, to an enhancement in the total leukocyte count and a restoration of the percentage ratio between the form elements of the leukocytic formula. Obtained results correlate with the results of studying the immunotropic action of the comparison preparation immunofan under the conditions of “social” stress.Conclusions. The results of the study of the immunotropic action of the neuropeptide compound Pro-Gly-Pro-Val under “social” stress indicate the presence of its immunocorrective activity, which is manifested by the restoration of indices of cellular and humoral reactions and white blood cells.

Оценка иммунотропной активности нейропептидного соединения Lys-Lys-Arg-Arg-Pro-Gly-Pro в условиях «социального» стресса

Objective: to investigate immunotropic activity of the argenine-containing neuropeptide compound Lys-Lys-Arg-Arg-Pro-Gly-Pro in conditions of "social" stress. Material and methods. The study was conducted on mongrel male rats of 6 months of age using a model of "social" stress based on the provision of permanent residence of animals in conditions of sensory contact, as a result of which submissive and aggressive types of behavior were formed. All animals were divided into groups: control I - rats who were in a cage one at a time and received water for injection in an equivalent volume; control II - groups of stressed animals with aggressive and submissive types of behavior; three groups of stressed rats were intraperitoneally injected with Lys-Lys-Arg-Arg-Pro-Gly-Pro at a dose of 100 mcg/kg/day and the comparison drug Imunofan (Arginyl-alpha-aspartyl-lysyl-valyl-thyrosyl-argininum) at an average therapeutic dose of 0.7 mcg/kg/day. The following indicators were evaluated: delayed hypersensitivity reaction index, antibody titer of direct hemagglutination reaction, phagocytic activity of peripheral blood neutrophils, leukocyte formula indicators. Results. The results obtained in the course of this study indicate that the arginine-containing neuropeptide compound Lys-Lys-Arg-Arg-Pro-Gly-Pro contributes to the correction of immune changes that occur against the background of exposure to chronic "social" stress, namely, an increase in the delayed hypersensitivity reaction index, the level of anti-erythrocyte antibodies, the total number of leukocytes and a decrease in phagocytic activity of peripheral blood neutrophils (phagocytic number and phagocytic index), as well as restoration of the percentage ratio of elements of the leukocyte formula. Conclusion. Thus, the study of the immunotropic activity of the argenine-containing neuropeptide compound Lys-Lys-Arg-Arg-Pro-Gly-Pro showed the presence of an immunocorrecting effect on the cellular and humoral links of immunity, which actualizes the need for a detailed study of the immune mechanisms of this compound.

An optimized microplate-based method to evaluate complement-dependent hemolysis mediated by intravenous immunoglobulins (IVIG)

Hemolytic reactions can cause serious complications after administration of Intravenous Immunoglobulin (IVIG), due to passive transfer of anti-A and anti-B IgG antibodies (isoagglutinins). A maximum allowable amount of isoagglutinins is established in the US and EU for licensed IVIG, as measured by a specified direct hemagglutination test (DHAT). Despite this limit, reports of hemolysis have increased over time, raising the question of how well the DHAT predicts clinically significant hemolysis. This study was undertaken to develop a microplate-based complement-dependent hemolysis assay (CDHA) that reproducibly measures functional hemolytic activity of IVIG, for assessment of IVIG products. An IVIG working reference reagent (NIBSC 14/160) was qualified as an assay control and for quantitation purposes. Hemolytic activities of 36 IVIG product lots encompassing seven brands and including 6 clinically hemolytic lots were measured. Hemolytic activity varied among IVIG product brands, and to a lesser extent, from lot-to-lot for individual brands. Correlation between the CDHA and DHAT was not robust which may reflect imprecision of the DHAT method or additional variables that influence complement-dependent hemolysis after opsonization. In conclusion, the CDHA provides a simple, specific, and sensitive tool for IVIG product characterization and investigation of hemolytic events by manufacturers, researchers, and regulatory authorities.

High titers of anti-A1 and anti-B antibodies among Peruvian group O platelet donors

Critical antibody titers have been described as factors associated with hemolysis in ABO plasma-incompatible platelet (PLT) transfusions. This study was carried out to describe the frequency of high-titers anti-A and antiB IgM and IgG antibodies in group O apheresis platelet donors, and to explore differences according to the donor characteristics. A cross-sectional study was carried out at the Blood Bank of a National Hospital in Peru from January to March 2019. IgM and IgG antibodies against A1 and B antigens were quantified in 339 platelet donors using the direct hemagglutination technique and the solid-phase adherence technique, respectively. For analysis purposes, two cut-off points; ≥128 and ≥64, were used to define a critical titer for IgM due to a lack of consensus. An IgG titer of ≥256 was also defined as critical. Of the donors, 22.1 % had critical IgM titers when the cut-off point was defined as ≥128. However, when the IgM cut-off was ≥64, the frequency of platelet donors with critical titers increased to 54.0 %. The frequency of donors with critical IgG titers was 23.5 %. Higher IgG titers were associated with female donors while higher IgM titers were negative associated with age. One in two or three platelet donors, depending on the cutoff point used to define a critical IgM titer, had at least one critical titer of anti-A or anti-B antibodies. Early identification of platelet donors with critical antibody titers could prevent passive transfusion of ABO antibodies to non-isogroup recipients.

Assessment of lectinic activity potentials in extracts of some tropical Euphorbiaceace

Lectins bind a variety of cells having cell surface glycoprotein or glycolipid, such as erythrocytes, leukemic cells, yeast and several types of bacteria. Several specificity groups have been identified such as mannose, galactose, N-acetyl glucoseamine, N-acetyl galactoseamine, L-fucose and N-acetyl neuraminic acid. The presence of two or more binding sites for each Lectin molecule allows the agglutination of many cell type. Sixteen (16) species of some tropical Euphorbiaceae plants were assessed for the presence of Lectins. The leaves of Acalypha torta, Acalypha wiskesiana, Acalypha hispida, Codiaeum variegatum, Euphorbia milli, Euphorbia pulcherima, Jatropha curcas and Jatropha gossypifola; the seeds of Croton tiglium, Ricinus comminus and Tetracarpidium conophorum; the stem of Euphorbia tirucalli and the tubers of Manihot esculenta (Cassava, vitamin A variety), Manihot esculenta (cassava, NR 8082 variety), Manihot esculenta (Cassava, TMX 419 variety), Manihot esculenta (Cassava, TMX 4(2) 1425 variety) were used for sourcing the Lectins. A. torta, A. wiskesiana, C. variegatum, C. tiglium, E. milli, E. pulcherima, E. tirucalli, J. curcas, J. gossypifola, R. comminus and T. conophorum agglutinated pooled washed human ABO cells in saline (direct haemagglutination) while A. hispida and the four varieties of M. esculenta showed no agglutination reaction. E. pulcherima showed specificity for B cells only while E. tirucalli showed specificity for O cells only, hence could be rightly indicated by referring to them as anti-B Ep and anti-H Et Lectins respectively (where Ep=Euphorbia pulcherima and Et= Euphorbia tirucalli). However A. torta and T. conophorum cross-reacted with pooled washed ABO cells in differing strengths and when standardized, showed that A. torta at a titre of 16 reacted specifically with O cells and T. conophorum at a titre of 128 reacted specifically with B cells. Based on this, these Lectins could be indicated as anti-H At and anti-B Tc respectively (Where At= Acalypha torta and Tc= Tetracarpidium conophorum). The protein content of the crude extracts of the sixteen (16) species were also assayed using Biuret protein assay method and the results revealed that there are no relation or association between the quantity of protein content and agglutination patterns of the extracts. This research has therefore succeeded in revealing presence of Lectinic properties in extracts of some tropical Euphorbiaceae.

Development of a Complement-Mediated Hemolysis Assay for Immune Globulin Intravenous Products and Characterization of Lots Associated with Clinical Hemolysis

Introduction: Immune Globulin Intravenous (Human) (IGIV), as known as Intravenous Gamma-Globulin (IVIg), are therapeutic polyclonal immunoglobulin preparations manufactured from pooled human plasma. Initially licensed for replacement therapy of primary immune deficiencies, IGIV indications have expanded to include various autoimmune and inflammatory conditions. Hemolytic reactions associated with IGIV infusion were historically considered as uncommon, but seem to be more frequent over the past 5-6 years, coinciding with widespread manufacturing changes to increase IgG yield, and use of higher dosages. Anti-A and anti-B IgG isoagglutinins in the IGIV preparations are believed to mediate these events. In 2014, FDA co-organized a workshop to discuss strategies to address hemolytic complications of IGIV infusions (Scott 2015). Although US products all comply with an anti-A and anti-B specification of ≤1:64, using the European Pharmacopoeia direct hemagglutination test (DHAT) (Thorp 2010), hemolytic events have persisted. The DHAT is semi-quantitative, imprecise, and does not measure hemolytic function. However, hemolysin tests have been difficult to standardize, especially for highly concentrated IGIV products (Scott 2015).This study reports development of a complement-dependent hemolysis assay (CDHA) suitable for testing IGIV products. We also tested a positive control preparation that is available from NIBSC (code 14/160), consisting of an IGIV lot that caused clinical hemolysis. Finally, we demonstrated differences among IGIV brands with respect to hemolytic activity.Methods: To provide consistent reagents, serum from whole blood donations were frozen as a large volume source of complement and a droplet-freezing method was used to store Red Blood Cells (Schmid 2011; Figure 1). Critical assay conditions were studied, including enzymatic treatment of RBC, removal of unbound IgG, complement source and concentration, divalent cation concentrations, and pH. Hemolytic activities of IGIV products and a positive control IGIV preparation were compared to the hemagglutination results tested using DHAT method. The CDHA was also used to evaluate hemolytic activity of IgG subclasses.Results: The most critical parameters for reproducible CDHA were complement source and preparation method, and assay pH. A pH of 6.5 (0.1M Tri-BIS buffer) was the most suitable condition for all of the tested IGIV analytes. Intra-laboratory variation was minimized by use of frozen RBCs and a consistent highly active complement preparation. The DHAT and CDHA results correlated with each other for most IGIV product lots (Figure 2). Hemolytic activity in IGIV lots varied among, in some cases within, brands. Enrichment of IgG subclass fractions from NIBSC 14/160 showed the highest hemolytic activity was present in IgG2 (Figure 3). In contrast, studies of recombinant anti-A monoclonal antibodies expressing the same anti-A F(ab'), but subclass-specific Fc receptors showed greatest activity of IgG3, consistent with evidence that IgG3 is the most potent complement-activating subclass. The role of IgG2 in hemolysis is not entirely surprising, as saccharide antigens often elicit IgG2 responses, and IgG2 can mediate complement activation (Diebolder 2014). Further subclass studies are planned.Conclusion: Hemolytic activity of IGIV products mediated by anti-A isoagglutinins can be reproducibly measured in IGIV products (Figure 4). Compared to the DHAT method, CDHA provides a continuous readout, is less variable, amenable to use of a standard curve, and more precise. Historical challenges to reproducibility are addressed through biological reagent standardization and attention to critical conditions. Studies comparing the CDHA and DHAT methods across a wider range of products and hemolytic lots will discern whether the CDHA provides a better measure of product safety than the current DHAT lot release assay. The CDHA can be helpful for characterization of new manufacturing methods and products, and investigation of hemolytic events. Studies of hemolysis-implicated lots may provide a basis for improvements in product safety. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Comparative study between Direct Microscopy and Indirect Haemagglutination Methods Used in Diagnosis of Urinary Schistosomiasis

Comparative study between Direct Microscopy and Indirect Haemagglutination Methods Used in Diagnosis of Urinary Schistosomiasis

Reliability of Postmortem ABO Blood Grouping: A Study of 100 Cases

Blood grouping has been one of the cornerstones for identification of biological materials in forensic investigations. Antigens of the ABO system can be detected even prior to birth. ABO blood groups can also be detected after death for a long period in many body tissues (teeth, bones, etc.). Blood has its own forensic value in many medico-legal issues. The present study looked at a total number of 100 autopsy cases brought for medico-legal investigations in the mortuary of the Department of Forensic Medicine in collaboration with the Department of Pathology at the Postgraduate Institute of Medical Sciences (PGIMS), Rohtak (Haryana, India). The blood samples were collected from right ventricle without any anticoagulant. ABO blood grouping was performed by direct haemagglutination technique using monoclonal antisera. Reverse blood grouping was also attempted. Our results showed that ABO blood group antigens can be detected from blood fluid after death, even in decomposed bodies with an estimated post-mortem interval (EPMI) of &lt;85 hours. Our results also show that as post-mortem interval increases beyond a certain limit there is a decrease in antigens on RBCs in the post-mortem blood.

ABH and Lewis blood group systems and their relation to diagnosis and risk of Helicobacter pylori infection

BackgroundHelicobacter pylori infection occurs in 50% of the world's population and represents a major risk factor for chronic gastritis, gastroduodenal ulcer and gastric cancer in developed and developing countries. The distribution of H. pylori virulence factors is diverse and varies geographically, such as the CagA and VacA genes, which have revealed association with disease status. Some findings show increased frequencies of these diseases in O Le (a-b +) and A Le (a-b +) blood type individuals, but other studies not found any relationship between these blood groups and H. pylori infection. AimThis study aimed to elucidate probable controversies described in the relationship between the ABH/Lewis blood groups and H. pylori, contributing to the severity of gastric diseases in northern the population of Belém -Pará.-Brazil. MethodsThis cross-sectional study included 288 samples of patients separate into two groups with gastric cancer and chronic gastritis. Blood, saliva, and gastric biopsy were analyzed using modified Gram and hematoxylin-eosin staining techniques, the enzyme immunoassay Elisa and Multiplex PCR. The antigens expression of ABH and Lewis systems was determined through Dot-ELISA and direct hemagglutination. Proportions were compared in univariate analysis, while the relation between putative risk factors including H. pylori status and ABO/Lewis phenotype was performed using multivariable logistic regression analyses, P-value < 0.05 was considered significant. ResultsThe findings of this study demonstrate that the likelihood of developing gastric cancer increases threefold if the individual is from A1 Le (a-b +) blood group, has premalignant changes, and infection with H. pylori virulent strains (cagA+/vacA + s1m1). ConclusionTherefore, this study found a significant association between ABO and Lewis phenotypes and H. pylori cagA status into the relevance of the development of gastric carcinogenesis.

Isohemagglutinin titering performed on an automated solid-phase and hemagglutinin-based analyzer is comparable to results obtained by manual gel testing.

Isohemagglutinins (anti-A and anti-B) mediate hemolytic transfusion reactions, antibody-mediated rejection of solid-organ transplants, and delayed engraftment after stem cell transplant. However, quantification of isohemagglutinins is often labor intensive and operator dependent, limiting availability and interfacility comparisons. We evaluated an automated, solid-phase and agglutination-based antibody titer platform versus manual gel testing. Plasma samples were obtained from 54 randomly selected patients. Titers were determined by our laboratory's standard assay (manual dilution followed by manual gel testing) and were compared to results obtained on a fully automated blood bank analyzer (Galileo NEO, Immucor). The analyzer determined immunoglobulin G (IgG) antibodies using solid-phase and immunoglobulin M (IgM) antibodies by direct hemagglutination. Isohemagglutinin titers obtained by manual gel versus the automated assay generally (>80%) agreed within one doubling dilution, and always (100%) agreed within two dilutions. Among O samples, the gel titer and the highest titer obtained with the automated assay (either IgG or IgM) were similar in paired, nonparametric analysis (p = 0.06 for anti-A; p = 0.13 for anti-B). Gel titers from group A and group B patients were slightly higher than the highest titer obtained using the automated assay (p = 0.04 for group A; p = 0.009 for group B), although these differences were within the accepted error of measurement. Manual and automated methodologies yielded similar isohemagglutinin titers. Separate quantification of IgM and IgG isohemagglutinins via automated titration may yield additional insight into hemolysis, graft survival after ABO-incompatible transplantation, and red blood cell engraftment after ABO-incompatible stem cell transplant.

PHENOTYPIC FREQUENCIES OF ABO AND RH(D) BLOOD GROUPS AMONGST HEALTHY BLOOD DONORS IN BIKANER, RAJASTHAN

Background &amp; Objective: Practically, most important blood groups are A, B, O, AB and RhD(+/-), which are essential for right blood supply to the recipients and in various studies. The incidence of ABO and Rhesus (Rh) groups varies markedly in different parts of the world and in different races. The present study was aimed at determining the prevalence of ABO &amp; RhD blood groups among healthy blood donors in Western Rajasthan.&#x0D; Material and Methods: At our centre, all the healthy blood donors were assessed and analyzed for ABO-Rh blood grouping during a period of previous 4 years (from January, 2015 to December, 2018). 119200 samples were checked for ABO and RhD blood grouping by manual saline antigen-antibody agglutination method using commercially prepared antisera antiA, antiB, antiAB, antiD, in house prepared reagent cells Acell, Bcell, Ocell and negative control. The results were reconfirmed on fully automated immunohematology analyzer (Galileo Neo, USA) with direct hemagglutination micro-well plates using antiA, antiB, antiD series-5, antiD Novaclone, Monoclonal control, Acell, Bcell and Ocell provided by the manufacturer. Discrepant samples were cross checked separately by column agglutination technique using DG Gel ABO/Rh-2D cards (Diagnostic Grifols S.A.) and tube technique.&#x0D; Results: We found that the most common blood group was B+ 39758 (33.35%) followed by O+, A+, AB+, B-, O-, A- and AB-. RhD negative donors were 10504 (8.81%) among the total donors. Phenotypic frequencies were expressed under the standard assumption of Hardy-Weinberg equilibrium.&#x0D; Conclusion: Present study concludes that the most common ABO phenotype found among healthy blood donors in Western Rajasthan (India) is group B with RhD negativity being 8.81%. Larger studies should be conducted to calculate antigenic, phenotypic and allelic frequencies in general population.&#x0D; Keywords: ABO-RhD blood group, Phenotype, Allele, Blood donors, hemagglutination.

Results of Epizootiological Monitoring of Small Mammals Habitant in Crimea Over the Period of 2015–2017

Objective of this work is to analyze the spatial distribution of various species of small mammals, caught between 2015 and 2017, and to identify the dominant species in different landscape areas, as well as their role in functioning of natural foci of zoonotic infections (tularemia, leptospirosis, tick-borne encephalitis, Lyme disease, Crimean hemorrhagic fever, hantavirus infection) in the territory of Crimea. Materials and methods. Small mammals were caught during the period of 2015–2017 and investigated using polymerase chain reaction (PCR), enzyme immunoassay (ELISA) and direct hemagglutination reaction. Results and conclusions. The dominant species of small mammals, containing the causative agents of some or other natural-focal infection are ubiquitous species, i.e. disseminated across the territory of the whole peninsula. In the presence of concomitant favorable conditions, they may contribute to the expansion of the natural foci of these infections. Positive tests for Lyme disease detected not only in small mammals caught in mountain-forest areas, but in steppe zone too may testify to the fact that the border of the natural focus of this infection is expanded. To clarify the boundaries of the natural foci of infections circulating in the territory of Crimea, it is necessary to conduct comprehensive analysis of distribution of small mammals, blood-sucking ectoparasites, and also epidemic manifestations in different natural areas of Crimea.

Comparative study between direct microscopy and indirect haemagglutination methods used in diagnosis of urinary schistosomiasis

Comparative study between direct microscopy and indirect haemagglutination methods used in diagnosis of urinary schistosomiasis

A novel rapid direct haemagglutination-inhibition assay for measurements of humoral immune response against non-haemagglutinating Fowlpox virus strains in vaccinated chickens.

Fowlpox (FP) is a serious disease in chickens caused by Fowlpox virus (FPV). One method currently used to control FPV is vaccination followed by confirmation that antibody titres are protective using the indirect haemagglutination assay (IHA). The direct haemagglutination inhibition (HI) assay is not done because most FPV strains do not agglutinate chicken red blood cells (RBCs). A novel FPV strain TPV-1 which agglutinates chicken RBCs was discovered recently and enabled a direct HI assay to be conducted using homologous sera. This study is therefore aimed at assessing the direct HI assay using a recently discovered novel haemagglutinating FPV strain TPV-1 in chickens vaccinated with a commercial vaccine containing a non-haemagglutinating FPV.Chicks vaccinated with FPV at 1 day-old had antibody geometric mean titres (GMT) of log2 3.7 at 7 days after vaccination and log2 8.0 at 28 days after vaccination when tested in the direct HI. Chickens vaccinated at 6 weeks-old had antibody geometric mean titres (GMT) of log2 5.0 at 7 days after vaccination and log2 8.4 at 28 days after vaccination when tested in the direct HI. The GMT recorded 28 days after vaccination was slightly higher in chickens vaccinated at 6-week-old than in chicks vaccinated at one-day-old. However, this difference was not significant (P > 0.05). All vaccinated chickens showed “takes”. No antibody response to FPV and “takes” were detected in unvaccinated chickens (GMT < 1). There was a slightly higher GMT in chickens of all ages throughout the observation period when the standard assay, the passive (indirect) haemagglutination was used (Overall GMT reached log2 9.3 ±.0.3 on day 28). However, the difference between the two assays was not significant (P > 0.05).ConclusionThese findings indicate that a simple and rapid direct HI assay using the FPV TPV-1 strain as antigen may be used to measure antibody levels in chickens vaccinated with non-haemagglutinating strains of FPV, and that the titres are comparable to those obtained by indirect IHA.

Immunostimulatory effect of the aqueous leaf extract of Phyllanthus niruri on the specific and nonspecific immune responses of Oreochromis mossambicus Peters.

Plant derived immunostimulants are a promising alternative to chemotherapeutics and also perhaps vaccines. In the present study, we examined the immunostimulating properties of aqueous leaf extract of Phyllanthus niruri, an Indian traditional medicinal herb, on neutrophil activation and antibody response of Oreochromis mossambicus (Peters). Serial ten-fold diluted doses of P. niruri ranging from 0.002 mg to 20 mg were administered to two groups of O. mossambicus (n=8). One group of fishes was administered with sheep red blood cells and the primary and secondary antibody responses were estimated using direct haemagglutination assay. The other group of fishes was administered heat-aggregated BSA to assess the ability of plant extract to elicit neutrophil activation. Our results indicate a significant enhancement of both neutrophil activation and antibody response. Among the various doses tested, fishes administered 20 mg/kg body weight caused the maximal enhancement of both primary and secondary antibody response and 0.002 mg/kg showed higher neutrophil activation compared to that of the control group. This short study indicates that aqueous leaf extract of P. niruri has the potential to be used as an immunostimulant and after confirming its immunostimulatory properties by a battery of tests on other nonspecific and specific parameters and disease-protective property by challenging the fish with virulent fish pathogens, it can be used either as a routine feed supplement to activate the immune system of farmed fishes or as an adjuvant to enhance the efficacy of vaccines.

Cerebral toxoplasmosis in patients with acquired immune deficiency syndrome in the neurological emergency department of a tertiary hospital

IntroductionCerebral toxoplasmosis is the most common cause of space occupying brain lesion in patients with HIV/AIDS in Brazil. In the post-HAART era, it is responsible for high rates of morbidity and mortality worldwide. Materials and methodsThis study consists of a case series of 56 patients diagnosed with cerebral toxoplasmosis whose clinical features, brain imaging and cerebrospinal fluid aspects were analyzed. ResultsCerebral toxoplasmosis led to the diagnosis of infection by the human immunodeficiency virus (HIV) in 27 (48.2%) of the patients, while 29 (51.2%) others already knew to be HIV seropositive. However, at the time of diagnosis of cerebral toxoplasmosis, only 9 (16.6%) reported being under antiretroviral therapy and 5 (8.9%) were receiving primary prophylaxis for toxoplasmosis. Headache, strength deficit and fever were the most frequent signs and symptoms throughout the study. Fifty-three patients showed changes consistent with toxoplasmosis in CT or MRI. Thirty-four (60.7%) CSF samples were positive in the indirect haemagglutination test and for the reaction of Toxoplasma gondii IgG ELISA, while 31 (55.4%) were positive in the direct haemagglutination test. Fifty (89.3%) patients underwent first-line treatment for toxoplasmosis. ConclusionCerebral toxoplasmosis is still a very relevant neurological disease in individuals with AIDS admitted to neurology emergency departments. Early diagnosis and initiation of empiric treatment and antiretroviral therapy are important for good prognosis.

RETROSPECTIVE ANALYSIS OF THE EFFECT OF VACCINATION OF MILITARY PERSONNEL WITH A DIPHTHERIA AND TETANUS ANATOXINS PREPARATION ADS-M ON THE DEVELOPMENT OF ACUTE BACTERIAL INFECTIONS OF THE UPPER AND LOWER RESPIRATORY TRACT IN THE POST-VACCINATION PERIOD (BASED ON THE EXPERIENCE OF THE MEDICAL SERVICE OF THE NORTHERN FLEET)

At the Northern Fleet, the formation of new regiments involves vaccination against diphtheria and tetanus using ADS-M anatoxin preparation. This is associated with increases in the incidences of acute bacterial infections of the upper and lower respiratory tract, including acute tonsillitis, acute bronchitis, and non-hospital pneumonia. The objective of the present study was to find out if there are causative relationships between vaccination and the infections. The study involved Northern Fleet draftees. Before and after vaccination, blood was sampled from the ulnar vein. The blood samples were tested for diphtheria and tetanus antibodies using direct hemagglutination techniques. The results suggest that the draftees are well protected immunologically against diphtheria and tetanus. Within the first month after vaccination, the incidence of infections among the draftees was significantly higher than in the subsequent months. The infected subjects showed higher titers of diphtheria and tetanus antibodies. This suggests the reasonability of exemption of draftees having high titers of diphtheria and tetanus antibodies from ADS-M vaccination. Conclusions: Vaccination of draftees with the ADS-M anatoxin preparation is not justified. ADS-M administration to draftees is associated with subsequent complications manifested as bacterial infections of the upper and lower respiratory tract, such as acute tonsillitis, acute bronchitis, non-hospital pneumonia, which are more frequent in the first month after vaccination than in the subsequent months. Draftees should be vaccinates against relevant infections before having been recruited to service.