Dense Nutrient Medium Research Articles

Structure of rhizosphere mycocomplexes and phyloplanes of cultural peanuts

Relevance . Peanut seeds are rich in oils, protein, carbohydrates, vitamins and flavonoids, therefore they are widely used in the production of food, feed, biodiesel, as well as in medicine. When cultivating peanuts, the problem of reducing the yield and sometimes the quality of seeds necessarily arises due to the impact of phytopathogenic microscopic fungi, and the main factors in the transmission of the infectious principle are soil and plant residues. The plant protection system should be based on knowledge of the structure of mycocomplexes of the rhizosphere and phylloplane. Methods . On a typical black soil on a natural infectious background in Belgorod (Russia), a sample of cultural peanuts V-2012 (Vietnam) from the collection of the Department of Biotechnology and Microbiology of the Institute of Pharmacy, Chemistry and Biology of the National Research University "BelSU" was studied. The species composition of microscopic fungi in the peanut rhizosphere was analyzed in comparison with fallow soil (control), which was found by seeding soil dilutions and leaf samples on dense nutrient media, followed by identification to species. Results . Compared with the control soil, the mycocomplex of the peanut rhizosphere (the Shannon index value at the level of 2.5 and 2.7 in the rhizosphere and control, respectively) was characterized by a lower degree of diversity and the degree of similarity of soil samples was high (the Jaccard coefficient was 50%), which means that that the cultivation of peanuts has lit- tle effect on the composition and structure of a typical black soil. In the peanut rhizosphere, a decrease in the frequency of occurrence of the phytopathogenic species F. oxysporum was observed, which may be associated with an increase in the rank in the mycocomplex of the mycophilic species T. lignorum. Aspergillus ochraceus, A. terreus, A. ustus, Candida albicans, Curvularia lunata, Fusarium oxysporum, Trichothecium roseum species were found to be sen- sitive to peanut cultivation. Peanut leaf spot was caused annually by the species Alternaria alternate , which can lead to a decrease in seed productivity up to 80%.

Diagnostics and prevention of aeromonosis Cyprinus sagrio

Relevance. Aeromonosis (carp rubella, hemorrhagic septicemia, infectious abdominal dropsy, Lublin disease) — an infectious disease of fish. Carp, common carp and their hybrids from underyearlings to broodstock are susceptible to the disease. The epizootic reaches its greatest distribution in the spring-summer period, by autumn it fades and the disease takes on a chronic course.Methods. The object of research was the fish Cyprinus carpio. In the experiments, we used the reference strain Aeromonas hydrophila ATCC 7966 of the Global Center for Bioresources, as well as isolates isolated during outbreaks of carp aeromonosis in fish farms in the Moscow region. The diagnosis for the presence of aeromonosis was established on the basis of epizootological, clinical and pathological data and the results of bacteriological studies. The study of biofilms of microorganisms was carried out during cultivation in liquid nutrient media, the CFU of microorganisms was counted and sown on the surface of solid media.Results.The total number of microorganisms (CFU, lg/g) of intestinal microbiocenoses— 54,21 ±0,05 – 66,09 ±0,12. Cultures of microorganisms A. hydrophyla, A. sobria, A. caviae were isolated from blood, heart, spleen, gills, intestine, liver and kidneys of fish. During cultivation at 37 °C for 6–72 h, the adhesion of vegetative forms of bacteria with a typical shape and size of the species and the formation of a bacterial monolayer — a diffuse layer of bacterial cells — was revealed. Viable and non-viable microorganisms were differentiated by fluorescence microscopy. On meat-peptone agar with 5,0% of defibrinated blood, as a rule, the growth of b-hemolytic colonies was observed, less often a-hemolytic colonies. When determining the enzyme DNase, a light zone was found along the inoculation line on the DNA BA medium. Growth of b-hemolytic cultures was observed on meat-peptone agar with 5,0% of defibrinated blood. When determining the DNase enzyme on the surface of the dense nutrient medium “DNA BA”, a light transparent zone was revealed around the colonies of microorganisms. Prevention of aeromonosis is based on systematic monitoring of the etiological structure of pathogens of aeromonosis; feeding in feeding and summer mother ponds begins in the spring when the water temperature rises to 14 °C.

Enhancement of the Yersinia pestis EV NIIEG Vaccine Srain Immunogenic and Protective Activity under Cultivation with Azoxymer Bromide (Polyoxidonium)

Background. The live-attenuated vaccine based on the Yersinia pestis strain EV line NIIEG is still used in Russia, providing protective efficacy against plague. Nevertheless, there is an urgent need for developing new ways to increase the immunogenicity of the Y. pestis EV NIIEG vaccine strain. In this study, the ability of direct action of immunoadjuvant azoximer bromide (polyoxidonium, PO) on the immunobiological properties of vaccine strain Y. pestis EV NIIEG during cultivation on a dense nutrient medium was evaluated.Materials & Methods. Y.pestis EV NIIEG, cultivated at 28 °С for 48 h on LB agar, Miller pH 7.2 ± 0.1 (Sigma-Aldrich, USA) with the addition of PO and without. MALDI-TOF mass-spectrometry was deployed for the obtainment of mass-spectra of ribosomal proteins from Y. pestis EV NIIEG cells on the MicroflexTM LT mass spectrometer (Bruker Daltonics, Germany). Protective efficacy was evaluated under subcutaneously challenge guinea pigs and mice BALB's with 400 LD50 doses of the Y. pestis 231, Y. pestis P-13268 Vietnam (MLD=5 CFU). Antibody titers to F1 in serum were determined using an ELISA.Results. The addition of the therapeutic concentration of PO in the cultivation medium induced a significant increase in the immunogenicity of Y. pestis EV NIIEG that resulted in enhancement of serum antibody levels against Y. pestis F1 antigen and several times the growth of protective efficacy in the bubonic plague model on two types of experimental animals. ImD50 of the vaccine strain Y. pestis EV NIIEG, cultivated with PO, was significantly (p < 0,05) lower in comparison to ImD50 for Y. pestis EV NIIEG in standard cultivation conditions. One year of storage at a temperature of 4 °С did not alter the protective properties of the vaccine strain Y. pestis EV NIIEG, cultivated with PO.Conclusions. Morphological studies confirmed the absence of influence PO introduction into the cultivation environment on the safety of the vaccine strain. MALDI-TOF MS profile of the Y. pestis EV NIIEG, cultivated with PO, had peaks characteristic features. The mass peak at m/z 3,061 was significantly down-regulated and new mass peaks at m/z 2,759, m/z 3,533 were determined. These changes are accompanied by the increase of Y. pestis EV NIIEG immunogenicity.

Thermal stability of a functional probiotic food ingredient based on encapsulated microorganisms Lactobacillus plantarum SP-A3

Recently, there has been increased interest in the production of functional probiotic food products such as cheese, yogurt and ice cream, as well as beverages and products based on meat, fruit, chocolate, etc. However, the range of such products is extremely limited, partly due to the difficulty of maintaining the viability of probiotic microorganisms. The thermal stability of the encapsulated L. plantarum SP-A3 culture was evaluated under conditions simulating the heat treatment of fortified food products. The object of the study is nearly pure culture of L. plantarum SP-A3 from the probiotic drug Lactobacterin (Mitrogen NPO). We used deep culture method on a dense nutrient medium – MRS agar at 37°C. The test culture was encapsulated in a 2 % alginate gel by B-390 apparatus (Buchi) using a 2 % calcium lactate solution as a fixative. Two types of microcapsules were obtained and studied: 200-300 µm and 400-600 µm in size. The capsules had a rounded regular shape, matte white, uniform color and were elastic. The efficiency of encapsulation with the proposed parameters was about 89%. We studied the possibility of using a model system simulating cooling of a food product after heat treatment. Both objects pass the most critical temperature ranges from 70 to 60 °C in a similar way, the use of a model system to study the survival of microorganisms when cooling a real food product after heat treatment is adequate. The study did not identify statistically significant differences between the capsule size (200-300 µm and 400-600 µm) and the survival after heat treatment (74,7±0,7% and 68,8±1,2%, respectively). Compared to unencapsulated cells (0% in three independent experiments), microencapsulated probiotics showed higher survival rates of up to 75,4%. Thus, with the use of microencapsulated probiotics, the range of probiotic foods can be expanded.

LYMPHOTROPIC ADMINISTRATION OF ANTIBACTERIAL DRUGS - A METHOD OF RATIONAL ANTIBIOTIC THERAPY.

The aim: To explore the possibilities of rationalizing antibacterial therapy by comparative analysis of the accumulation of ceftriaxone administered lymphotropically and intramuscularly. Materials and methods: The research used a method of studying the sensitivity of microorganisms to antibacterial drugs, which is based on the diffusion of antibacterial drugs from the carrier (homogenate of the test organ) in a dense nutrient medium, which, depending on the sensitivity of microorganisms, inhibits the growth of the studied culture and is accompanied by the formation of a growth inhibition zone (GIZ) in the nutrient medium. For control purposes, a commercial disk with ceftriaxone was used, which caused GIZ of microorganisms at the level of 27.05 ± 0.9 mm. Results: The homogenate of organ samples obtained from rabbits administered with the antibiotic lymphotropically, inhibited the growth of test cultures around the discs in all cases. The inhibition was slightly less than the control inoculation, but the result clearly indicated the presence of antibiotics in the test material in the quantity sufficient to inhibit the growth of the pathogen. Concurrently, after intramuscular administration of the antibiotic, biopsy specimens from various growth inhibition organs of test cultures were either not induced at all or were induced in small amounts, which was certainly insufficient for the inhibitory action of the drug administered by this method. Conclusions: The administration of the antibiotic lymphotropically promotes its accumulation in all organs in the quantity sufficient for antibacterial action, which allows us to recommend lymphotropic therapy as a rational method of antibiotic therapy.

Mycoplasma synovia is an infection among broilers at an industrial-type poultry farm

A method for diagnosing infectious synovitis among young broilers aged 11-12 weeks, obtained from an industrial-type poultry farm, was investigated in laboratory conditions. The experiments were carried out on the basis of the Microbiology Department of the All-Russian Research Veterinary Institute of Poultry Farming - a branch of the VNITIP Federal Research Center of the Russian Academy of Sciences. To perform mycoplasmatological studies, the limbs of birds with pronounced joint damage were selected and a nutrient medium was prepared by adding 20% horse blood serum, 10% yeast extract and 0.5% glucose to Edward's broth. Phenol-mouth at a concentration of 0.0001% was used as an indicator of the medium. As a result of a series of 7 "blind" passages, the appearance of opalescence and precipitation was noted, small transparent colonies were observed on a dense nutrient medium. Thus, the preliminary diagnosis of infectious synovitis (infektious synovitis) of broilers, obtained on the basis of epizootological data, clinical signs and pathoanatomic changes, was fully confirmed by a positive result of mycoplasmatological studies. According to the results, tilan was proposed for the treatment of sick poultry in doses recommended by the manufacturer, and for the prevention of the disease – strict compliance with the norms of bird planting density and indoor microclimate.

Validation of rapid method for determining the overall toxicity of feed using bioluminescent microorganisms Photobacterium phosphoreum

The article presents the results of studying the validation characteristics of the express method for determining the general toxicity of feed using bioluminescent microorganisms Photobacterium phosphoreum. The work used lyophilized culture Ph. phosphoreum (strain IMV B-7071; Sq3) and Colpoda steinii dry culture for ecological and toxicological studies of environmental objects, livestock and poultry products (RC № АВ-02438-01-11, produced by “Vidrodzhennia” LLC, Odesa). Measurement of the luminescence intensity of luminescent bacteria was performed on a luminometer EMILITE-1003A. To quantify the effect on luminescence of bacteria we used toxicity index to conclude on the degree of toxicity of the sample. When testing Colpoda steinii the mobility of ciliates was the criterion for assessing the toxicity of the studied feed samples. Validation parameters of the express method for determining the general toxicity using photoluminescent microorganisms Ph. phosphoreum were established according to ISO 16140:2003 “Microbiology of food and animal feeding stuffs — Protocol for the validation of alternative methods (IDT)”. There was conducted a comparative study of alternative method (determination of total feed toxicity using Ph. phosphoreum) and standard method (determination of toxicity using infusoria Colpoda steinii in accordance with DSTU 3570-97 “Feed grain, products of its processing, feed. Determination of toxicity”. The test object was a grain mixture (barley–wheat 50:50), toxicant — mycotoxin zearalenone. During the validation of the method, the following parameters were determined: relative specificity, relative accuracy, control of intralaboratory reproducibility, luminescence stability, linearity, convergence, detection limit and method determination limit. It has been established that the technique is specific, accurate, linear, reproducible. Optimal conditions and shelf life for Ph. phosphoreum: in tubes on a dense nutrient medium at a temperature of 4°C with monthly reseeding for 7 months, and optimal conditions and time of cultivation before the study: in tubes on a liquid nutrient medium at a temperature of 26°C 24 h after seeding. The limit of detection of the method (for zearalenone) is 0.125 μg/cm3, and the limit of determination is 0.25 mg/kg of feed

RESULTS OF THE STUDY OF THE POULTRY MANURE MICROFLORA

The microbiological composition of chicken manure has a significant impact on its further destruction. For the disposal of chicken manure by biological methods it is important to have in the manure nutrients, growth inhibitors (residues of antibiotics or disinfectants), the ratio of spore-forming and non-spore-forming microflora, as well as the presence of beneficial microflora. The aim of the work was to study the microbiological composition of chicken manure in poultry farms of Ukraine.
 Chicken manure from poultry farms of different scale and method of keeping poultry on the territory of Ukraine was studied. Selected samples were transported in a thermal container and examined microbiologically by inoculation on commonly used and selective media. To determine the amount of spore-forming microflora, a slurry suspension was prepared, sown in a mattress with a dense nutrient medium and turpentine was introduced into a cotton gauze plug. The morphology of the isolated microorganisms, acid-forming properties, catalase activity and enzymatic properties were studied according to generally accepted methods.
 It was found that in the manure of chickens, which were kept indoors, the percentage of spore-forming microorganisms ranged from 14.74 to 21.24 %, and not spore-forming – 78.76-85.26 %. In the manure of chickens that had grazed, the percentage of spore-forming microorganisms ranged from 48.93 to 78.81 %, and not spore-forming – 21.19-51.07 %. The average amount of spore-forming microflora under both conditions was 40.86 %, and not spore-forming – 59.14 %. According to the results of the work, 20 isolates were isolated, of which 15% of isolates had pronounced acid-forming properties. On MRS agar, the isolates had the appearance of small, smooth white-gray S-shaped colonies, in Gram-stained smears, small gram-positive rods were observed, arranged singly, in pairs, and in short chains. Isolates 1, 2 and 3 had a negative reaction to catalase.
 According to the results of the studies, isolate 1 was identified as Lactobacillus salivarius subsp. salivarius, isolate 2 – Lactobacillus reuteri, and isolate 3 – Lactobacillus delbrueckii.

Аcinetobacter baumannii bv Tryptophandestruens bv nov. isolated from clinical samples

The aim of the study was to determine the taxonomic status of a group consisting of atypical strains of Acinetobacter baumannii, outline relevant characteristics and methods necessary for their identification. There were examined 10 strains of A. baumannii (6 of them primary comprised) bearing similar profile of atypical features isolated from clinical samples (urine, sputum) in 2017–2019 at the Military Medical Academy. Сlinical strains of typical A. baumannii (n = 36), Acinetobacter nosocomialis (n = 14), Acinetobacter pittii (n = 9) and 1 strain of Acinetobacter calcoaceticus isolated from the external environment were used in comparative studies. Atypical strains had the characteristics of A. calcoaceticus — A. baumannii (ACB) complex bacteria and were identified as A. baumannii. The utilization of substrates as the only carbon source was studied on a dense synthetic medium added with 0.2 % substrate during incubation for 72 hours at 37°C. Carbohydrate oxidation coupled to acid formation was detected on the Hugh–Leifson medium by using a micromethod. Aromatic amino acid biotransformation was carried out in liquid and dense nutrient media assessed in chromogenic reaction. The rpoB gene was used for strain genetic characterization. Amplification of two 940 and 1210 base pair (bp)-long fragments from the rpoB gene was performed by the routine polymerase chain reaction using primers with previously described sequences. Amplification products were sequenced by Sanger using Big Dye Terminator v3.1 (Applied Biosystems, USA) and capillary electrophoresis on an automatic sequencer ABI PRISM 3130 (Applied Biosystems, USA), followed by using methods for determining the similarity levels of sequenced fragments with the rpoB gene sequences of the reference strain A. baumannii ATCC 17978 (GenBank accession no. CP053098.1). It was found that all strains belonging to atypical A. baumannii spp. had a specific set of features that distinguish them from typical strains of A. baumannii as well as other types of the ACB complex: detected biotransformation of L-tryptophan (via anthranilate pathway) and anthranilic acid under unambiguous lack of such signs in other bacteria; lack of utilized sodium hippurate and L-arabinose being unambiguously evident in other bacteria; lack of utilized L-tryptophan, putrescine, L-ornithine being utilized in the majority of strains of belonging to other bacterial species. Genetic analysis showed that the control strains of typical A. baumannii displayed 99.20–99.21% similarity within the sequenced fragments of the rpoB gene with those from the rpoB gene of the reference strain. All 10 strains of atypical A. baumannii had similar features (99.20–99.21%). At the same time, parameters of control strains from other bacterial species significantly differed: A. nosocomialis (95.10–95.97%), A. pittii (94.63–94.92%), A. calcoaceticus (93.00%). Hence, the strains of atypical and typical A. baumannii are genetically homogeneous and belong to the same species. The data presented allow us to consider this group of atypical A. baumannii strains as a new biovar. We propose the name for this new biovar — tryptophandestruens (tryptophan-destroying) stemming from the Latin word destruens — destroying. Identification of A. baumannii bv. tryptophandestruens bacteria can be carried out in laboratory of any level by using tests for L-tryptophan biotransformation as well as sodium hippurate utilization.

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Study of the influence of the composition of mold and starter microflora on the formation of organoleptic characteristics of soft-ripened cheeses

The article presents the results of a study of the characteristics of growth, incl. joint, mold cultures of Penicillium camembеrti and yeast-like fungi of Geotrichum candidum on a dense nutrient medium and the regularities of changes in the organoleptic, physicochemical and microbiological characteristics of Camembert-type cheeses in the process of production, ripening and storage, depending on the combinations of bacterial and mold starter microflora. The objects of the study were the cultures of Penicillium camembеrti and Geotrichum candidum, cheeses with white mold, made according to the type of “Camembert” cheese, produced from cow’s milk with mesophilic and thermophilic fermenting microflora, mold cultures of Penicillium camembеrti and yeast-like fungi of Geotrichum candidum. The study of cheeses was carried out in the process of ripening and storage after 15, 30, 60 days from the date of manufacture. It was found that when Penicillium camembеrti and Geotrichum candidum were co-cultivated on Petri dishes, visual assessment showed a symbiotic effect of culture development, which manifests itself in the stimulating effect of yeast-like fungi on the growth of mold fungi compared to the cultivation of each culture separately. Under the conditions of the experiment carried out, the main mold culture influencing the organoleptic characteristics of the cheese was the mold culture of Penicillium camemberti. The combination of mold cultures did not affect the improvement of the organoleptic characteristics of the cheeses. It is shown that the composition of the main acid-forming starter microflora, both mesophilic and thermophilic, has a significant effect on the lactic acid process and flavoring of Camembert-type cheese. The cheeses produced on the basis of mesophilic starter microflora were distinguished by an accelerated ripening process and less storage capacity due to deeper proteolysis. At the same time, cheeses produced on the basis of thermophilic fermenting microflora retained quality stability for up to 60 days. Thus, the use of various combinations of starter cultures will make it possible to create a flavor line of cheeses with white mold, depending on consumer preferences with different storage capacity.

Number of ecological-trophic groups of microorganisms depending on technology of winter wheat cultivation on ordinary chernozem of the Central Caucasus

Aim. To study the influence of cultivation technology on the number of ecological and trophic groups of microorganisms and the yield of winter wheat on the common chernozem of the Central Caucasus.Material and Methods. The number of microorganisms was determined by counting colonies on dense nutrient media according to generally accepted methods.Results. The number of microorganisms that transform organic and mineral forms of nitrogen and soil yeast was determined, depending on meteorological conditions, soil cultivation and fertilizers. Mineral fertilizers had the most significant effect on the activity of the soil microbiota. In years of varying moisture availability, fertilizers improved the nutrient regime of the soil, which contributed to increased plant development, microflora activity and increased winter wheat yield. In both the technologies applied, the largest number of all groups of microorganisms studied was observed when applying a complete mineral fertilizer -N52P52K52 .This dose of fertilizer also provided the highest yield of winter wheat - 6.07-6.33 t/ha. With a balanced plant nutrition regime (N52P52K52), the processes of decomposition and mineralization of plant residues proceeded at the same rate in both technologies. At the same time, without the use of fertilizers, the total activity of microorganisms in the no-till technology exceeded the minimum technology by 23.4х105 columnforming units, which indicates the beginning of the process of self-healing of the soil in the third year after the transition to direct sowing of crops.Conclusion. The number of microorganisms is one of the most sensitive indicators of the direction of complex biochemical processes, allowing us to assess the combined influence of all factors on the overall state of the soil microbiocenosis.

Experimental Substantiation of Antimicrobial Efficiency of a New Composite Polymeric Material Based on Poly(2-Hydroxyethyl Methacrylate) under the Action of Low-Intensity Current without External Power Supplies

Active use of polymeric materials has become an integral part of all areas of modern medicine. Wound dressings capable of prolonged release of drugs directly into the lesion occupy a special place among them. The possibility of using such materials in the presence of low-intensity currents without external power supplies in a comprehensive treatment program for patients with burn injuries remains promising. The aim of the work is to study experimentally the antimicrobial efficacy of a new composite polymeric material based on poly(2-hydroxyethyl methacrylate), saturated with the antiseptic decamethoxine, under conditions of low-intensity current without external power supplies. The method of free radical thermal polymerization of a mixture of liquid monomer 2-hydroxyethyl methacrylate, crosslinking agent triethylene glycol dimethacrylate, polymerization initiator azobisisobutyronitrile was used for the synthesis of composite polymeric material. In addition, fourfold volume of distilled water as a pore-forming agent and decamethoxine as an antimicrobial component were administered. Known dressings of synthetic and biological origin were selected for comparison, some of which were pre-soaked in a 0.02% solution of decamethoxine. The study of conductivity of the materials without external power supplies was performed on the surface of a dense nutrient medium in a Petri dish using VITA-01M measuring device. Determination of antibacterial properties was performed by diffusion into agar. The obtained results allowed to establish the ability of the suggested polymeric material to conduct low-intensity currents without external power supplies, exceeding the duration of other traditional dressings. Comparison of antimicrobial activity of the studied samples confirmed the synergism of the action of physical factors and a new polymer-based composite material with the addition of antimicrobial substance to inhibit the growth of the test museum and clinical strains of Staphylococcus aureus. The ability of low-intensity currents without external power supplies to potentiate the antimicrobial properties of a new composite polymeric material based on poly(2-hydroxyethyl methacrylate), modified with a pore-forming agent, with the addition of decamethoxine was experimentally established.

ИЗУЧЕНИЕ РЕЗИСТЕНТНОСТИ К ПРОТИВОТУБЕРКУЛЕЗНЫМ СРЕДСТВАМ У БОЛЬНЫХ, НАХОДЯЩИХСЯ НА СТАЦИОНАРНОМ ЛЕЧЕНИИ В ОТДЕЛЕНИИ РЕЗИСТЕТНЫХ ФОРМ ТУБЕРКУЛЕЗА ЛЕГКИХ

A comparative study of resistance to anti-tuberculosis drugs in patients with newly diagnosed pulmonary tuberculosis and in patients with chronic forms of tuberculosis who were treated in an anti-tuberculosis hospital was carried out. We examined 142 patients treated in the department of resistant forms of tuberculosis. A complete bacteriological study of sputum for Mycobacterium tuberculosis was carried out with the determination of sensitivity to anti-tuberculosis drugs during the cultivation of microorganisms on a liquid nutrient medium in a system with automated registration of bacterial growth and on a dense nutrient medium of Lowenstein-Jensen using the method of absolute concentrations. Methods of PCR analysis were also used. According to the anamnesis, two groups of patients were formed: 82 patients with newly diagnosed tuberculosis and 60 patients with a chronic course of the disease or with a relapse of the disease. According to the age structure, patients under the age of 50 prevailed in both groups. At the same time, in the first group there was a significant number of patients aged 18-29 years - 20.7%. In the second group, the number of patients aged 18-29 years was significantly less - 5%. Despite the fact that all patients were assigned to the category of multidrug-resistant pathogen and were treated in the unit of resistant forms of tuberculosis, it was important to assess the level of Mycobacterium tuberculosis resistance separately for patients in each of the selected groups. The largest differences between groups were found in the assessment of second-line drug resistance. Patients of the second group showed high resistance to fluoroquinolones - 30% and amikacin - 41.6%. In patients with newly diagnosed pulmonary tuberculosis (first group), resistance to fluoroquinolones was 9.7%, and to amikacin - 17.1%. Capreomycin was the most effective in both groups. The resistance of the causative agent of tuberculosis in patients of the first group was 4.9%, in the second - 11.6%.

ОЦЕНКА АНТИМИКОБАКТЕРИАЛЬНОЙ АКТИВНОСТИ ПРОИЗВОДНЫХ ПРОПИЛТИАДИАЗОЛОХИНАЗОЛИНА В ОТНОШЕНИИ MYCOBACTERIUM INTRACELLULARE

The compounds studied are propylthiadiazoloquinazoline derivatives, tryptanthrin analogs. Tryptantrin and its derivatives were found to have different antimycobacterial activity in vitro and in vivo. This series of experiments is devoted to the study of antimycobacterial activity of three propylthiadiazoloquinazoline derivatives obtained in the laboratory of organic synthesis and Biopharmaceutics of the Institute of Chemistry of the Academy of Sciences in Moldova using a non-tuberculous strain of Mycobacterium intracellulare, which is a part of the Mycobacterium avium complex (MAC). Species of the MAC are one of the main pathogen types causing mycobacteriosis. In this regard, propylthiadiazoloquinazoline derivatives are of interest for a comprehensive study as a potential antimycobacterial drug. To study antimycobacterial activity the method of dilutions in a dense nutrient medium of Middlebrook 7H9 broth with glycerol in Petri dishes was used. To assess antimycobacterial activity visual assessment of M. intracellulare growth in a solid nutrient medium was used. It is shown that the compounds studied inhibit M. Intracellulare growth. The most active compound was compound № 1 - 2-mercapto-5H-[1,3,4]-thiadiazolo-[2,3-b]-quinazoline-5-one.

Разработка способа биоконверсии целлюлозы Мiscanthus sinensis для получения биоэтанола

Introduction. Cellulose-containing parts of herbs are an excellent source of alternative energy and can be used to produce biological ethanol. The present research aims at improving this fundamental and promising area of biotechnology. It introduces a new consortium of microorganisms that can saccharify while fermenting the substrate.
 Study objects and methods. The research featured technical cellulose obtained from Miscanthus sinensis using hydrotropic delignification and oxidation with pertrifluoroacetic acid. The ethanol content in the culture liquid was determined using an Agilent 7890B gas chromatograph with a flame ionization detector. The biocompatibility of the strains was studied by growing a direct co-culture in a dense nutrient medium.
 Results and discussion. The research objective was to create a new microbial consortium for the single-step production of bioethanol from Miscanthus sinensis cellulose. A set of biocompatibility experiments and cultivation conditions made it possible to select the optimal producers. The two developed microbial consortia required optimal compositions of culture media, which were determined by varying the ratio of components and measuring the yield of ethanol in the resulting culture liquid.
 Conclusion. The best consortium for Miscanthus sinensis cellulose consisted of Pichia stipites Y7124, Candida shehatae NCL3501, Kluyveromyces marxianus Y-4290, and Zymomonas mobilis 113 at a ratio of 1:1:1:1. The optimal parameters of bioethanol production included: temperature = 35 ± 1°C, pH = 5.2, time = 16 ± 1 h. The most efficient culture medium had the following composition (g/l): glucose – 5.0; peptone – 5.0; yeast extract – 0.4; K2HPO4 – 1.5; (NH)2 HPO4 – 1.5; MgSO4 – 0.5.

ОЦЕНКА БИОСОВМЕСТИМОСТИ БАКТЕРИАЛЬНЫХ ИЗОЛЯТОВ – КОМПОНЕНТОВ БИОКОНСЕРВАНТОВ

The aim of this work was to study the relationship between the isolates of Bacillus subtilis EFS, Lactobacillus plantarum, Propionibacterium freudenreichii, Lactococcus lactis, and Strepto-coccus lactis, which have the potential for the development of a bacterial preparation that improves the safety of plant-based feed during harvesting and storage. The possibility of using the tested iso-lates as part of the association was identified by the drip method, which is high quality and most suitable for primary testing. The method involved direct joint cultivation of microorganisms on the surface of a dense nutrient medium and visual assessment of signs of suppression or enhancement of the growth of one culture by another. As a result of the study, the following types of interactions between microorganisms were identified: neutrality, contact progression, antagonism, and contact regression. For inclusion in the association, isolates that showed neutrality and contact progression to each other were selected. The corresponding types of relationships were characteristic of B. sub-tilis EFS, L. plantarum, P. freudenreichii, and L. lactis isolates. Based on this, six recipes were formed that are promising for further research in order to develop a biological preparation that helps to improve the safety of plant-based feed during harvesting and storage.

Effect of antibodies to agglutinogens 1 and 2, filamentous hemagglutinin and pertussis toxin on formation of Bordetella pertussis biofilms on abiotic substrate

Aim. Study of the effect of antibodies to agglutinogens 1 and 2, filamentous hemagglutinin (FHA) and pertussis toxin (PT) on the formation of biofilms by Bordetella pertussis strains on the abiotic substrate.Materials and methods. Vaccine-derived and freshly isolated strains of B. pertussis were used. Cultures of strains grown on a dense nutrient medium were used as an inoculum for obtaining biofilms. The intensity of biofilm formation in round-bottomed polystyrene 96-well plates in the presence of antisera to agglutinogens 1 and 2, antiserum to FHA, and monoclonal antibodies (MСА) to the S1, S2, and S3 subunits of PT was evaluated by staining with 0.1% gentian-violet solution.Results. Most of the studied strains were sensitive to antibodies, which was manifested in complete suppression of biofilm formation. All strains were sensitive to antiserum to agglutinogen 1, antiserum to FHA, and MCA to the S2 subunit of KT. Three out of 4 studied strains with this agglutinogen in their composition were sensitive to antiserum to agglutinogen 2: No. 475 (serotype 1.2.3), No. 317 (serotype 1.2.3) and No. 178 (serotype 1.2.0). Relative resistance to antiserum was detected only in serotype 1.2.0 strain No. 305, but with minimal dilution, the intensity of biofilm formation was 1.8 times lower than in the control group. Strains No. 703 (serotype 1.0.3) and No. 287 (serotype 1.0.3) that did not have agglutinogen 2 were resistant to antiserum. Four and 5 out of the 6 strains used were sensitive to the S1 and S3 subunits of PT, respectively. Strain No. 305 was resistant to MCA to the S1 and S3 subunits, and strain No. 287 to MCA to the S1 subunit. At the same time, the intensity of biofilm formation was 2 and 1.8 times lower than in the control at the minimum MCA dilution.Conclusion. These data indicate that the growth of biofilms of B. pertussis strains is suppressed by antibodies both to the surface structures of the microbial cell (agglutinogens 1 and 2, FHA) and to the S1, S2 and S3 subunits of PT.

Role of nitrogen deficiency on growth and development near isogenic by E genes lines of soybean co-inoculated with nitrogen-fixing bacteria

Nitrogen deficiency is a limiting factor in increasing efficiency of crop production in terrestrial ecosystems, and the transformation of inert nitrogen to forms that can be assimilated by plants is mediated by soil microorganisms. Symbiotic nitrogen-fixing bacteria and roots depend on each other and have developed various mechanisms for symbiotic coexistence. The aim of this work was to investigate the role of nitrogen deficiency on growth and development near isogenic by E genes lines of soybean (Glycine max (L.) Merr.): short-day (SD) line with genotype Е1е2е3(Е4е5Е7), and photoperiodic insensitive (PPI) line with genotype е1е2е3(Е4е5Е7) grown from seeds inoculated with active strains of Bradyrhizobium japonicum against the background of local populations of diazotrophs of the genus Azotobacter spp. and establish how the soybean – Bradyrhizobium symbiosis will develop as the genes of both microsymbionts and macrosymbionts are responsible for the formation of the symbiotic complex. Plants were grown in a vegetation chamber, in sand culture. To assess the quantitative composition of microorganisms in the rhizosphere and rhizoplanes, 6 plants were selected from each soybean line, then separation of the zones of the rhizosphere and rhizoplanes was performed using the method of washing and the resulting suspension was used for inoculation on dense nutrient media (mannitol-yeast agar medium and Ashby medium). The results of study showed that seed inoculation and co-inoculation provides faster formation of the symbiotic soybean – Bradyrhizobium complex. Differences in nodulation rates between the short-day line with genotype Е1е2е3(Е4е5Е7), and a photoperiodic insensitive line with genotype е1е2е3(Е4е5Е7) were identified. Determination of the amount of B. japonicum on the medium of mannitol-yeast agar in the rhizosphere and rhizoplane showed that inoculation by B. japonicum strain 634b caused a significant increase in the amount B. japonicum in the rhizosphere and rhizoplane in both soybean lines, comparison with non-inoculated seeds. Then, co-inoculation by B. japonicum strain 634b + Azotobacter chroococcum significantly increased the amount of B. japonicum only in the rhizoplane and decreased their number in the rhizosphere. Determination of the amount of A. chroococcum on the Ashby elective medium in the rhizosphere and rhizoplane showed that the inoculation by B. japonicum strain 634b caused a significant decrease in the amount of A. chroococcum both in the rhizosphere and in the rhizoplane of the PPI line of soybean, and in the rhizosphere the SD line, in comparison with non-inoculated seeds. That can testify to the competitive interaction of these microorganisms. However, the co-inoculation by B. japonicum strain 634b + A. chroococcum in the SD line significantly increased the number of A. chroococcum in the rhizoplane and decreased their number in the rhizosphere, in the PPI line their number decreased in the rhizoplane and increased in the rhizosphere, in comparison with non-inoculated seeds. Probably, the E genes (their dominant or recessive state) of soybean isogenic lines affect the regulation of the content and distribution of sugars. It was established that the nitrogen deficiency stimulated development of the root system of plants and the synthesized sugars were distributed predominantly to the root system growth. We suppose that the seeds’ inoculation had extended sugar consumption to the symbiont, due to which it compensates the lack of nitrogen, but leads to a slower growth of the root system.

Sensitivity of biofilms of vaccine and freshly isolated <i>Bordetella pertussis</i> strains to antibiotics

Aim. To study the sensitivity of biofilms of vaccine and freshly isolated strains of Bordetella pertussis to antibiotics.Materials and methods. Vaccine and freshly isolated strains of B. pertussis were used. Cultures of strains grown on dense nutrient medium were used as inoculate for biofilms production. The intensity of biofilm formation in round-bottomed polystyrene 96-well plates was estimated by staining with 0.1% gentian-violet solution. The following antibiotics were used in experiments: penicillins (ampicillin), cephalosporins (ceftriaxone), aminoglycosides (gentamicin), macrolides (erythromycin).Results. The highest resistance to antibiotics was demonstrated by the vaccine strain No. 305 and freshly isolated strain No. 211, sensitive only to erythromycin. Vaccine strain No. 703 was sensitive to gentamicin and ampicillin and showed resistance to erythromycin and ceftriaxone. Vaccine strain No. 475 was sensitive to all tested antibiotics. The Tohama 1 strain was resistant to ampicillin and sensitive to other antibiotics. Freshly isolated strains No. 178 and No. 162 were resistant to ceftriaxone and sensitive to gentamicin, erythromycin and penicillin. Minimal inhibitory concentrations of tested antibiotics ranged from 0.2 μg/ml to 5.0 μg/ml.Conclusion. These data indicate the heterogeneity of vaccine and freshly isolated strains of B. pertussis in sensitivity to antibiotics. The greatest activity was shown by erythromycin, which suppressed the growth of biofilms of 6 out of 7 strains. The least effective was ceftriaxone, which suppressed the growth of biofilms of only 2 strains.