Clinical-epidemiological features and improvement of botulism laboratory diagnos-102 tics in Irkutsk region

Irkutsk Region is one of the Russian Regions where botulism cases are registered yearly. Botulism morbidity in Irkutsk Region is analysed for the ten-year period (since 2002); its clinical-epidemiological features are revealed at the present stage. Sporadic cases of botulism, are dominated. (81,9 %). From. 2006 in Irkutsk Region botulism share makes 100 % in food poisoning number. A considerable role in the infection transmission plays consumption of fish, mainly smoked or salty Baikal omul (90,9 %). In 30 % of cases injured people purchased fish at private persons in unascertained trade places, 60 % of patients ate own salting fish. In Irkutsk Region botulism is registered more often from July till November with peaks in July (58,3 %) and. September (20,8 %) that is connected, with activation of unprofitable fabricated fish selling in combination, with adverse conditions of this foodstuff storage. Botulism affects various age groups of population with, prevalence of the age category 30-49 years old. (47,8 %). Socially active able-bodied population. (72,3 %) is most often affected that increases the social importance of the disease. Mainly, moderate severe (56, 5 %) and. severe (34,8 %) disease forms were registered. Primary symptoms include two variants - dyspeptically-paralytic and. ophthalmologic-neurological forms. Analysis of botulism etiological structure showed, that the leading part in the disease occurrence in Irkutsk Region belonged to botulinum. type Е (73, 9 %). Clinical material and. foodstuff testing to botulinum toxin presence was conducted in parallel by the standard reaction of biological toxin neutralisation in white mice and dot-immunoanalysis with application of the test system that we have developed. Possibility of dot-immunoassay application in laboratory diagnostics of botulism is estimated as the express method with high specificity, sensitivity and. simplicity

Improvement of a test-system for the botulinus toxin screening in dot-immunoanalysis

We constructed a test system for dot-immunoassay (DIA) to accelerate definition and identification of botulinus toxins and also to refuse from application of laboratory animals for routine screening of clinical samples, foodstuff and environments. This system permitted to detect botulinus toxin during approximately 2 h in the tested samples. Sensitivity of this DIA in some cases exceeded the mice biotest. This improved method has minimum reaction to nonspecific exposures from the investigated biological substrata. It is simple to conduct. It is high efficient and expressive, does not require to use expensive equipment and the reactants, special training for the personnel. Lyophilization conditions for the immune reagents used for the test system preparation for botulinus toxin dot-immunoassay were selected. High sensitivity, specificity of the analysis are remained, stability of the preparations (periods of storage) is increased. This method is convenient to use in field conditions at extreme situations, in particular, in mobile autolaboratories for epidemiological survey

The production of hyperimmune pseudotuberculosis sera

Background. Pseudotuberculosis remains a serious public health problem, which dictates the need to develop simple and rapid diagnostic methods for this disease. The effectiveness of the latter largely depends on the activity and specificity of the diagnostic sera. Currently, in our country, a diagnostic serum for Yersinia pseudotuberculosis O-monovalent (serotype I, III) is produced with a regulated area of application: for an approximate agglutination reaction on glass (Pasteur NIIEM, St. Petersburg). Preparation of pseudotuberculosis sera for a wider scope of their use, in particular, as a  source of specific antibodies in the design of diagnostic immunobiological preparations and test systems for pseudotuberculosis, is relevant and in demand in healthcare practice.Aims. To obtain hyperimmune pseudotuberculosis sera, promising for use in the practice of laboratory studies for pseudotuberculosis.Materials and methods. Chinchilla rabbits aged 3–6 months, weighing 2.5–3.0 kg served as animal producers of pseudotuberculosis sera. As immunogens, we used a corpuscular antigen (CAg) - a suspension of cells of the epidemically significant Yersinia pseudotuberculosis 3704 O: 1b strain isolated by an outbreak of pseudotuberculosis in the town of Zima, Irkutsk region, inactivated by boiling, and a preparation obtained from the outer membranes, containing the main surface immunogen of the strain. For the adsorption of experimental sera, in order to minimize the nonspecific response, we used heterologous microorganisms Escherichiacoli, Salmonella typhimurium, Shigella flexneri and Y. enterocolitica O:3, which have a similarity of surface antigenic structures with the causative agent of  pseudotuberculosis. The presence of specific antibodies in experimental pseudotuberculosis sera was determined in a volumetric agglutinationreaction.Results. Optimal schemes of rabbit immunization were selected, which made it possible to obtain hyperimmune sera against Y. pseudotuberculosis 3704 O:1b antigens with an agglutination activity of 1:3200-1:6400. In low dilutions of experimental sera (1:100–1:400), obtained against corpuscular antigen and outer membrane antigens (Escherichia coli, Salmonella typhimurium, Shigella flexneri , Y. enterocolitica О:3, S . enteritidis Gartnery) was observed in the agglutination reaction (AR). After the adsorption of experimental pseudotuberculosis sera by cells of heterologous strains, there was no cross-reaction with the indicated microorganisms in the  agglutination reaction.Conclusions. The obtained hyperimmune adsorbed sera against the boiling-inactivated Y. pseudotuberculosi s 3704 O: 1b corpuscular antigen and outer membrane antigens can be used as a source of specific antibodies in the design of diagnostic immunobiological preparations for the detection of pseudotuberculosis microbe, as well as in the monitoring of the epidemic situation

Using dot-immunoassay in decoding the outbreak of pseudotuberculosis in the Tomsk region

Background. Pseudotuberculosis remains a serious healthcare problem, which determines the expediency of developing the express methods for its early diagnosis. To detect the pathogen, we designed test system for dot-immunoassay (DIA) based on antibodies labeled with silver nanoparticles (SNPs) isolated from hyperimmune rabbit serum obtained against killed cells of  Yersinia pseudotuberculosis of O:1b serovariant.The aim. To assess the possibility of using dot-immunoassay for express identification of Y. pseudotuberculosis cultures isolated from clinical material and environmental objects at the initial stage of bacteriological study during laboratory diagnosis of the disease.Methods. We used the materials from the outbreak of pseudotuberculosis in the Krylovskaya Boarding School of the Bakcharsky district of the Tomsk region in 2021. Specific antibodies from hyperimmune rabbit sera obtained against Y. pseudotuberculosis 3704 particulate antigen of O:1b serotype were labeled with SNPs and used in DIA on nitrocellulose membranes with visualization of reaction results with a solution of a physical developer. The presence of the causative agent of pseudotuberculosis in the test material was inferred by the formation of gray spots of different intensity (from 4+ to 1+).Results. All Y.  pseudotuberculosis strains isolated using bacteriological method on  the second day of the study from clinical material obtained from sick people and environmental objects were detected in DIA at concentrations ≥ 3.1 × 104 microbial cells per milliliter (m.c./ml).Conclusion. The designed test system for dot-immunoassay using SNPs as a marker of specific antibodies for the detection of Y.  pseudotuberculosis in cultures isolated from swabs from vegetables and clinical material from patients, including those  with  mixed infection, allows us to  detect a specific corpuscular antigen with a high sensitivity (≥ 3.1 × 104 m.c./ml), providing express identification of isolated cultures at the initial stage of bacteriological study

The Comparative Analysis of Phenolic Compounds Accumulation in Leaves of Various Kinds of Kalanchoe

One of actual problems of modern pharmacology is creation of new medicines on the basis of vegetable raw materials. In this plan some succulents present a great interest in particular some kinds of the genus Kalanchoe, such as Kalanchoe pinnata (K.pinnata) and Kalanchoe Daigremontiana (K.daigremontiana). Their leaves contain useful mineral salts, organic acids and the numerous phenolic compounds (PC). Education and accumulation of these PC depends on genetic features of a plant and numerous factors of environment. Besides, these representatives of a secondary metabolism cause biological, including antioxidant activity (AOA) of this or that species of a plant, that is ability its component to inhibit oxidizing free radical processes. Use of modern methods of antioxidant properties research for plant extracts or juice of this or that plant allow to study and reveal their medicinal value at higher level.In the present work the comparative analysis of measurement results of the total phenols content and their activity in leave juice of various kinds of Kalanchoe (Kalanchoe L.) is carried out by ammetric and chemiluminescence methods for the purpose of identification among them the most active producers of phenol metabolites. Objects of research were juice samples of 34 kinds of the genus Kalanchoe, grown up in a succulent collection in Timiryazev Institute of plant physiology RAS (Moscow, Russia). Among the studied samples two most active from the point of view of their antioxidant properties Kalanchoe kinds: K.scapigera and K.rhombopilosa are revealed. Both methods show considerably higher values of the phenol metabolite content in leave juice of these plants and their AOA in comparison with K.pinnata and K.daigremontiana. For possible expansion for use of specified types of Kalanchoe as sources of biologically active compounds, additional researches of biochemical structure, antibacterial, antimicrobic and other properties of these plant components is necessary. Probably, they will appear more perspective for their use in pharmacy and medicine.</p

The Comparative Analysis of Phenolic Compounds Accumulation in Leaves of Various Kinds of Kalanchoe

One of actual problems of modern pharmacology is creation of new medicines on the basis of vegetable raw materials. In this plan some succulents present a great interest in particular some kinds of the genus Kalanchoe, such as Kalanchoe pinnata (K.pinnata) and Kalanchoe Daigremontiana (K.daigremontiana). Their leaves contain useful mineral salts, organic acids and the numerous phenolic compounds (PC). Education and accumulation of these PC depends on genetic features of a plant and numerous factors of environment. Besides, these representatives of a secondary metabolism cause biological, including antioxidant activity (AOA) of this or that species of a plant, that is ability its component to inhibit oxidizing free radical processes. Use of modern methods of antioxidant properties research for plant extracts or juice of this or that plant allow to study and reveal their medicinal value at higher level.In the present work the comparative analysis of measurement results of the total phenols content and their activity in leave juice of various kinds of Kalanchoe (Kalanchoe L.) is carried out by ammetric and chemiluminescence methods for the purpose of identification among them the most active producers of phenol metabolites. Objects of research were juice samples of 34 kinds of the genus Kalanchoe, grown up in a succulent collection in Timiryazev Institute of plant physiology RAS (Moscow, Russia). Among the studied samples two most active from the point of view of their antioxidant properties Kalanchoe kinds: K.scapigera and K.rhombopilosa are revealed. Both methods show considerably higher values of the phenol metabolite content in leave juice of these plants and their AOA in comparison with K.pinnata and K.daigremontiana. For possible expansion for use of specified types of Kalanchoe as sources of biologically active compounds, additional researches of biochemical structure, antibacterial, antimicrobic and other properties of these plant components is necessary. Probably, they will appear more perspective for their use in pharmacy and medicine

The Conjugates of Phenolic Acids in Lichens of the Order Lecanorales

Lichens are symbiotic associations of a fungus (usually an ascomycete) and a photobiont, which may be an alga and/or a cyanobacterium. Lichens dominate on about 6–8% of land surface, mainly in the habitats with severe climatic conditions. Lichenized fungi are among the pioneer vegetation on bare rock or soil. Mat-forming species contribute substantially to the soil cover in tundras and high mountain elevations. Lichens are rich in water-soluble compounds which can be leached-out the lichen thalli with atmospheric depositions. We have recently described the occurrence of water-soluble phenolics in lichens (Zagoskina et al 2013). These compounds can play important role in the ecosystem functioning and primary soil formation (weathering, humification). The aim of this work was to study qualitative composition of water-soluble phenolics in the lichen species widespread in the soil cover of tundra zone. The air-dried thalli of Alectoria ochroleuca, Cetraria islandica, C.nigricans, C.nivalis, Cladonia arbuscula and C.stellaris were homogenized to powder and used for the study. Lichens were collected in Khibiny mountains, Kola Peninsula in August 2013. Phenolic compounds were extracted by distilled water (1h, 30C) and analyzed by TLC before and after the acid hydrolysis. It was found that all the lichens under the study contained the conjugates of phenol carboxylic acids. We have identified that non-phenolic part in some of these conjugates was represented by sugars and amino acids. The TLC of the hydrolizates of water extracts revealed occurrence of p-oxybenzoic acid in all of the species studied. The lichens Cetraria islandica, С.nigricans and Cladonia stellaris contained also vanillic acid. These phenolic acids are widespread in plant kingdom and are known as products of lignin decomposition in higher plants. The physiological role of water-soluble phenolics in lichens as well as their environmental role are need to be understood in future studies.</p

phenolic compounds in two genotypes of buckwheat sprouts

The focus of the experiment described in this paper has been to study the influence of cadmium on the morphophysiological and biochemical characteristics of buckwheat sprouts of various ploidity and to investigate stress responses of plants at early, physiologically sensitive stages of development. It has been established that growth of the aboveground plant organs and the root system of diploid buckwheat was more inhibited by cadmium compared to tetraploid buckwheat sprouts. Cadmium uptake by the aboveground parts of plants was insignificant and was not influenced by the ploidity level, whereas the root systems of tetraploid plants demonstrated an increased ability to accumulate cadmium compared to the roots of diploid buckwheat. According to the results of biochemical research of the buckwheat sprouts' ability to accumulate phenolic compounds of different classes and to produce phenylalanine ammonia lyase (PAL), the basic plant enzyme that catalyses phenolic biosynthesis, the tetraploid buckwheat species demonstrated an increase in all tested characteristics unlike the diploid form which showed a drop in the polyphenols content when PAL activity went up. The obtained results are relevant for further studies of phenolic biosynthesis in plant species of various ploidity levels and identifying the role of the key enzyme involved in phenolic compound metabolism that activates the phenylpropanoid pathway and can, in the case of increased cadmium exposure, be involved in the synthesis of secondary metabolites that are different from phenolic compounds.C1 [Kazantseva, Varvara; Goncharuk, Evgenia; Kartashov, Alexander; Zagoskina, Natalia] IPP RAS, KA Timiryazev Inst Plant Physiol RAS, 35 Bot Skaya St, Moscow 127276, Russia.[Mammadov, Ramazan] Pamukkale Univ, Fac Sci & Art, Dept Biol, TR-20017 Denizli, Turkey

The Conjugates of Phenolic Acids in Lichens of the Order Lecanorales

Lichens are symbiotic associations of a fungus (usually an ascomycete) and a photobiont, which may be an alga and/or a cyanobacterium. Lichens dominate on about 6–8% of land surface, mainly in the habitats with severe climatic conditions. Lichenized fungi are among the pioneer vegetation on bare rock or soil. Mat-forming species contribute substantially to the soil cover in tundras and high mountain elevations. Lichens are rich in water-soluble compounds which can be leached-out the lichen thalli with atmospheric depositions. We have recently described the occurrence of water-soluble phenolics in lichens (Zagoskina et al 2013). These compounds can play important role in the ecosystem functioning and primary soil formation (weathering, humification). The aim of this work was to study qualitative composition of water-soluble phenolics in the lichen species widespread in the soil cover of tundra zone. The air-dried thalli of Alectoria ochroleuca, Cetraria islandica, C.nigricans, C.nivalis, Cladonia arbuscula and C.stellaris were homogenized to powder and used for the study. Lichens were collected in Khibiny mountains, Kola Peninsula in August 2013. Phenolic compounds were extracted by distilled water (1h, 30C) and analyzed by TLC before and after the acid hydrolysis. It was found that all the lichens under the study contained the conjugates of phenol carboxylic acids. We have identified that non-phenolic part in some of these conjugates was represented by sugars and amino acids. The TLC of the hydrolizates of water extracts revealed occurrence of p-oxybenzoic acid in all of the species studied. The lichens Cetraria islandica, С.nigricans and Cladonia stellaris contained also vanillic acid. These phenolic acids are widespread in plant kingdom and are known as products of lignin decomposition in higher plants. The physiological role of water-soluble phenolics in lichens as well as their environmental role are need to be understood in future studies

Influence of cadmium on morphophysiological traits and accumulation of phenolic compounds in two genotypes of buckwheat sprouts

The focus of the experiment described in this paper has been to study the influence of cadmium on the morphophysiological and biochemical characteristics of buckwheat sprouts of various ploidity and to investigate stress responses of plants at early, physiologically sensitive stages of development. It has been established that growth of the aboveground plant organs and the root system of diploid buckwheat was more inhibited by cadmium compared to tetraploid buckwheat sprouts. Cadmium uptake by the aboveground parts of plants was insignificant and was not influenced by the ploidity level, whereas the root systems of tetraploid plants demonstrated an increased ability to accumulate cadmium compared to the roots of diploid buckwheat. According to the results of biochemical research of the buckwheat sprouts' ability to accumulate phenolic compounds of different classes and to produce phenylalanine ammonia lyase (PAL), the basic plant enzyme that catalyses phenolic biosynthesis, the tetraploid buckwheat species demonstrated an increase in all tested characteristics unlike the diploid form which showed a drop in the polyphenols content when PAL activity went up. The obtained results are relevant for further studies of phenolic biosynthesis in plant species of various ploidity levels and identifying the role of the key enzyme involved in phenolic compound metabolism that activates the phenylpropanoid pathway and can, in the case of increased cadmium exposure, be involved in the synthesis of secondary metabolites that are different from phenolic compounds.C1 [Kazantseva, Varvara; Goncharuk, Evgenia; Kartashov, Alexander; Zagoskina, Natalia] IPP RAS, KA Timiryazev Inst Plant Physiol RAS, 35 Bot Skaya St, Moscow 127276, Russia.[Mammadov, Ramazan] Pamukkale Univ, Fac Sci & Art, Dept Biol, TR-20017 Denizli, Turkey