The issues of laboratory diagnostics have been relevant since the first days of the SARS-CoV-2 pandemic and play a key role in the fight against the spread of a new coronavirus infection. A direct method for the etiological diagnosis of the causative agent of COVID-19 is the detection of SARS-CoV-2 RNA using the nucleic acid amplification method. In the context of a pandemic and the mass appeal of patients for medical care, the issues of ensuring the quality of ongoing molecular biological studies at all stages (preanalytical, analytical, postanalytical) become the most relevant. the results and timing of the study not only affect the diagnosis and treatment tactics of a particular patient, but are also the basis for the introduction of anti-epidemic measures, the adoption of organizational measures. The study is to summarize the experience in creating an effective and reliable system for managing the quality of molecular biological research in a pandemic using the example of a federal budgetary healthcare institution. The experience of the laboratory of a federal healthcare institution in the context of the COVID-19 pandemic was analyzed, errors were analyzed at the preanalytical, analytical and postanalytical stages of PCR studies with the identification of quality criteria, the impact on which significantly leads to quality improvement. The quality control system for PCR studies is based on the development of regulatory documents and instructions for the patient and laboratory staff, registration of all documents in a single information system with access to information from all structural divisions, with the possibility of uploading data to the patient's personal account on the institution's website. Quality indicators of all stages of PCR studies and measures that significantly affect the quality of laboratory studies were identified; Measures have been identified to reduce the turnaround time of a PCR test: distribution of biomaterial flows, optimization of operators' work, purchase of additional equipment, a patient feedback system, and an infection control information system. The obtained results make it possible to create a reliable quality control system, minimize the risk of obtaining erroneous research results, optimizing the work of the clinical diagnostic laboratory and increasing its productivity.

COVID-19 is a disease caused by the new coronavirus SARS-CoV-2. Outbreaks were first reported in China on December 31, 2019. Exactly one month later, the WHO declared the outbreak a public health emergency of international concern, and on March 11, it was declared a pandemic. In February, the infection began to spread rapidly to various countries, with Europe declared the center. By April 17, 2020, cases had been confirmed in all subjects of the Russian Federation. At the beginning of September 2020, the number of cases exceeded one million; at November 19, two million; at December 26, three million. At February 10, 2021, four million; at May 23, five million; at July 20, six million; at September 5, seven million; at October 18, eight million; at November 13, nine million; and at December 12, 2021, ten million. The rapid spread of the virus, accompanied by a significant increase in the number of infections and deaths. A total of about 18.6 million cases were recorded at the end of the first half of 2022. The total number of deaths from coronavirus in Russia at that time was 382,313 (2.06% of all cases). The number of tests performed by various analytical methods amounted to over 274, 5 million, i.e. 1.9 million per 1 million population. The rapid spread and the increase in new infections caused by SARS-CoV-2 made it necessary to use new epidemiological and diagnostic approaches based on fast, accurate and reliable technology for detecting the infectious agent. One such virus detection method is polymerase chain reaction with reverse transcription and real-time detection of the results. The review presents the domestic market offerings of PCR diagnostic kits and provides their comparative consumer characteristics.

Using the example of the clinical strain of R. sibirica «Bayevo 105/87», the possibility of quantitative determination of rickettsias in clinical samples from patients with Siberian tick-borne typhus by real-time polymerase chain reaction (PCR-RT) was evaluated. Cultivation was carried out in the yolk sacs of developing chicken embryos, from which a piece of the yolk sac or chorion was taken. A total of 125 samples were examined. A set of reagents "RealBest DNA Rickettsia species (kit1)" was used for PCR-RT. The obtained values of the threshold amplification cycle (Ct) were compared with the results of microscopy of smear preparations stained by the Zdrodovsky method, the values of which were divided into ranks: the I rank - single rickettsias in individual fields of vision, the II rank - single rickettsias in each field of vision, the III rank - from 10 to 25 rickettsias in each field of vision, the IV rank - from 25 to 50 rickettsias in each field of view. The median Ct value for rank I was 17.6 (16.37; 18.58), for the II - 16.0 (15.0; 16.41), for the III - 15.0 (14.0; 15.1) and for the IV - 15.0 (13.7; 14.64). A significant average correlation was established between the number of rickettsias in the preparation under microscopy and the value of the threshold cycle in PCR RT (r=-0, 4849542; p=9.968e-09). When determining the correlation between the pathomorphological characteristic and the value of the threshold cycle, its absence was established. The detection of rickettsias in the blood vessels of the chorion of developing chicken embryos was of interest. In 10 samples, the yolk sac and chorion were taken for the study, and in parallel they were examined by PCR-RT. The use of modern, more sensitive molecular biological methods allows for quantitative analysis of DNA in the chorion, while preserving the volumes of the most valuable material - the yolk sac.

The coronavirus infection continues to spread around the world. In this regard, the purpose of this work was: to develop a set of reagents for the qualitative detection of SARS-CoV-2 virus RNA. The set was developed by CJSC «Ecolab», 20 positive samples were used to develop the kit. The research method consisted of several stages: isolation of SARS-CoV-2 RNA, RNA reverse transcription reaction and PCR amplification of cDNA with simultaneous detection of the result in real time. The main characteristics of the kit: analytical sensitivity - 100%, specificity - 100%, accuracy - 100%. Thus, our method for diagnosing a new coronavirus infection based on real-time RT-PCR makes it possible to qualitatively and quickly detect betacoronavirus RNA in clinical material from patients and healthy individuals with suspected coronavirus infection and other symptoms of SARS.

A PCR assay has been developed to identify the DNA of the human herpes virus type 7. The search and selection of conserved regions was carried out by comparing the whole genome nucleotide sequences of HHV-7. A fragment duplicated in the HHV-7 genomes was chosen as a target for amplification. The performance of the assay was tested on a synthetic matrix and clinical samples. The developed assay has high sensitivity and specificity and showed good efficiency in detecting HHV-7 DNA in clinical samples.

Antinuclear antibodies (ANA) are a heterogeneous group of autoantibodies that react with various components of the cell nucleus and cytoplasm. ANA is the main serological marker for autoimmune liver disease (AILD). The aim of the study was to compare the diagnostic value of two methods of screening for the determination of ANA (indirect immunofluorescence reaction on HEp-2 cells (IIF -HEp-2) and enzyme-linked immunosorbent assay (ELISA) in the sera of AILD patients. The sera of 118 patients with AILD (51 with autoimmune hepatitis - AIH, 19 with primary biliary cholangitis - PBC, 48 with overlapping syndrome - OVERLAP), 30 patients with non-alcoholic fatty liver disease (NAFLD) and 30 healthy donors (HD) were studied. Determination of ANA by the IIF-HEp-2 method was carried out by visual assessment of samples under an AXIOSKOP 40 microscope, by ELISA - on an Alegria automatic analyzer. A weak degree of agreement between the positive and negative results of the ANA screening study using IIF-HEp-2 and ELISA (Cohen's kappa coefficient æ=0.4) was noted. Screening determination of ANA in patients with AILD by the IIF-HEp-2 method was distinguished by greater diagnostic sensitivity (DS) (68.6%) and a lower frequency of false negative results (31.4%) compared with ELISA (35.6% and 64.4 % respectively, p<0.05). The overall diagnostic specificity (DS) of the ANA study in IIF-HEp-2 was lower than with ELISA (66.7% and 86.7%, respectively, p<0.05). Both screening methods for determining ANA (IIF-HEp-2 and ELISA) were useful for diagnosing AILD (positive likelihood ratio - LR+: 2.1 and 2.6, respectively). In terms of the negative likelihood ratio (LR-), screening for ANA by the IIF-HEp-2 method, in contrast to ELISA, served as a "useful" test to exclude the diagnosis of AILD (0.5 and 0.8, respectively). The determination of ANA using IIF-HEp-2 is the most sensitive and "useful" screening test for the diagnosis of AILD, and ELISA is classified as a less "useful" screening method due to low diagnostic sensitivity and a high false-negative rate.

Due to the steady increase in the number of children with autism and the high heterogeneity of clinical groups, the diagnosis of these disorders and their severity is an urgent problem in modern medicine. In the course of the work, 126 children from 3 to 13 years old with typical neurodevelopment and with severe and mild autism spectrum disorders (ASD) were examined. Disease severity was determined according to the Childhood Autism Rating Scale (CARS). The levels of pro-/anti-inflammatory cytokines and neurotrophic factors (nerve growth factor beta and brain-derived neurotrophic factor) in blood plasma were assessed by enzyme immunoassay. Associations between indicators in each group of patients were assessed using the Spearman test and visualized as a heatmap of correlations. Statistical data processing was carried out in the R software. Significantly high levels of IL-4 in blood plasma and a decrease in the number of significant correlations within/between systems were revealed in children with mild autism compared with children with typical neurodevelopment. Such data can probably reflect the theory that some children with ASD are characterized by slow brain development, as a variant of the evolutionary norm. On the contrary, in children with severe ASD, high systemic levels of IL-6 and IFNg are shown against the background of low values of IL-10, IL-1β, TNFα and NGFβ, supported by the almost complete absence of intra/ and intersystem interactions. This may act as an indicator of maladaptation of the immune and nervous systems in severe autism, which contributes to the pathogenesis of the disease. Thus, a set of indicators: high levels of key pro-inflammatory cytokines - IL-6 and IFNg, low levels of IL-10, NGFβ and disintegration of the cytokine and nervous systems in the periphery can be proposed as an approach to indicate the severity of the condition in children with ASD.

The possibility of the preoperative level of 42 indicators characterizing the cellular composition and metabolism in blood of patients with stage III lung adenocarcinoma (AC) to predict their relapse-free survival was studied. Blood samples of 451 patients with newly diagnosed AK stage III after their surgical treatment (resection volume - R0) have been investigated. The duration of the relapse-free period (period of observation - 1 year), cellular composition of the blood, concentration of C-RP, albumin, Cyfra 21-1 antigens, SCC, TPA, chemokines CXCL5, CXCL8, pyruvate kinase TuM2 PK isoenzyme, HIF-1α and hyaluronic acid in blood serum so as the proportion of blood cells with CXCR1 and CXCR2, CD44V6 receptors in blood serum were measured. To determine the dependence of the duration of the relapse-free period after the treatment on the observation time, Kaplan-Meier graphs were built. The relationship between the determined parameters and survival was judged using single- and multi-factor Cox proportional hazard models. Comparison of groups with different risk of AK recurrence was performed using the Log Rank test and χ2. The assessment of the predictive information content of laboratory tests was carried out using ROC analysis. It was shown that the concentration of monocytes, eosinophilic leukocytes, the relative quantity of lymphocytes with CXCR1 receptor, the level of Cyfra 21-1 before surgical treatment were associated with the duration of the relapse-free period. A regression equation was compiled, which included the level of Cyfra 21-1, relative content of lymphocytes with CXCR1, and the eosinophilic leukocytes / monocytes ratio. Based on the threshold value Y=0,597, a Kaplan-Meier plot of patient survival was built and the results of it correspond to the TNM stratification. The prognostic sensitivity of the results of the equation - 85,7%, the specificity - 94,7%.

The aim of the study is to develop a method for early diagnosis of intrauterine infection (IUI). A study of markers of inflammation in the venous blood of 60 pregnant women was conducted. The study was followed by a retrospective assessment of the outcomes of pregnancies and childbirth. Of these, 33 patients with a gestation period of more than 37 weeks (full-term pregnancy) and, accordingly, 27 patients from whom the blood sample was taken at a period of less than 37 weeks - patients with the threat of premature birth (PB). PB is the main factor contributing to the development of IUI. 27 patients were diagnosed with premature rupture of the membranes (PROM). Of these, 15 are with the threat of PB. 8 of them had a diagnosed IUI. In all cases of diagnosed PROM, including those with IUI, the concentration of nitrite and nontiolate nitroso compounds (NO2-+RNO) in the mother's blood plasma was 2.3±1.2 µM, while normally it does not exceed 0.1 µM (p<0.001). Regardless of the duration of pregnancy. The use of antibiotics in the case of PROM contributed to the normalization of the concentration (NO2-+RNO). Therefore, increasing of this indicator is result of bacterial infection. Indications of other markers of inflammation: the number of leukocytes in venous blood and in a smear of vaginal contents, the level of C-RB did not significantly change in both PROM and IUI (p>0.1). Since the concentration index (NO2-+RNO) increased in almost all cases of PREM, unlike all other clinical and biochemical indicators used in modern medicine, there is an obvious sense of its use for the current monitoring of the health of pregnant women. But it is still impossible to say unequivocally about the possibility of monitoring the fetal health by concentration (NO2-+RNO) in the mother's blood.

The role of the kidneys in the metabolism and homeostasis of sulfur-containing amino acids is great, so the levels of methionine (Met), total homocysteine (tHcy) and their ratios can be of diagnostic value in chronic kidney disease (CKD), in a course of the arterial hypertension (AH). The aim of the study was to evaluate the Met/tHcy ratio in hypertensive patients with CKD. We used blood plasma of 76 patients aged 40-75 years with AH and the excretory dysfunction of the kidneys; subgroups: 1 - with proteinuria (n=37); 2 - without proteinuria with glomerular filtration rate (GFR) < 90 ml/min/1.73 m2 (n=39) and comparison group 3 - patients with AH without renal excretory dysfunction (n=28). Significantly lower Met levels were in subgroup 1. THcy levels were higher in subgroups 1 and 2 than in group 3. The Met/tHcy ratio revealed differences in subgroups 1 and 2 vs group 3. No differences were found in Arg and Lys levels. Positive correlations of the Met/tHcy ratio with the number of erythrocytes, but not with the level of hemoglobin, were revealed. In the ROC analysis, the cut-off points for the Met/tHcy ratio compared to group 3 were 3.08 for subgroup 1 and 3.36 for subgroup 2. With the progression of CKD, there is an increase in the levels of tHcy in the blood, and a decrease in the content of Met. A decrease in GFR, especially in a case with proteinuria, is accompanied by a decrease in the level of Met. The Met/tHcy ratio above 3.36 can be considered as the minimum of the balance between these sulfur-containing amino acids contents in a blood necessary for hypertensive patients with CKD.

Simultaneous quantitative measurement of mRNA of the WT1, BAALC, EVI1, PRAME and HMGA2 genes in whole blood samples reflects the specific pathological proliferative activity in acute leukemia and their ratio is promising as a diagnostic marker. The transcriptome profile of acute leukemia cells is usually assessed using NGS or microarray techniques after a preliminary procedure for isolation of mononuclear cells. However, the results of using the multiplex PCR reaction for the simultaneous determination of all above mRNAs in whole blood samples have not been published so far. Determination of mRNA of WT1, BAALC, EVI1, PRAME and HMGA2 genes in venous blood level samples by multiplex RT-PCR. The study included 127 blood samples from patients who diagnosis of acute leukemia was subsequently confirmed. In the comparison group, 87 samples of patients without oncohematological diagnosis were selected, including 31 samples (K1) with a normal blood formula and 56 samples (K2) with a violation of the cellular composition - anemia, leukocytosis and thrombocytopenia. RNA isolation and reverse transcription were performed using the Ribozol-D and Reverta-L kits (TsNIIE, Russia). Determination of the mRNA expression level of the WT1, BAALC, EVI1, PRAME and HMGA2 genes by multiplex real-time PCR using a homemade multiplex PCR kit. The mRNA level was characterized by high interindividual variation and did not correlate with the rate of circulating leukocytes or blood blasts. Expression of WT1 mRNA was observed in whole blood only in one patient from the control group and in 112 (88%) patients with leukemia and was combined with a decrease in the level of HMGA2 mRNA expression and BAALC mRNA values. In contrast to the control groups, patients with leukemia had higher levels of BAALC mRNA in AML and ALL, increased PRAME mRNA in AML and APL, but lower levels of HMGA2 in APL.

Despite of great number of investigations in the area of tinea pedis, question is opened: to what extent dermatophyte fungi are spread among modern population and does their occurrence interrelated with host age? Investigated group included 99 volunteers from 14 to 73 years old. Skin scales were collected from heel area of foot, and signs of heel skin trouble were expressed in points. In contrast to usual laboratory microscope magnification x900 we worked at x1750, what allowed to estimate not only fungal, but bacterial forms too. Average abundances of microbial morphotypes were expressed in points. Heel skin trouble increased in the process of aging (Pirsons` coefficient r=0.954). Bacilli occurred in all persons independently from age, but their abundance increased with aging (0.821). On the contrary cocci were more common and abundant in young person`s feet (-0.620). Occurrence of dermatophytes increased with age (0.891), at that relatively high values took place in young persons (10.5% with mycelium and 73.7% with spores) and in group without any heel skin trouble symptoms (7.7% and 76.9%), what allow to refer these fungi to normal habitats of foot skin.

The relationship between multiple sclerosis and the state of the human microbiome was studied, namely, the change in the representation of microbiota phylotypes, the proportion of coccal flora, the proportion of anaerobic, gram-negative, proteolytically active microflora, as well as the concentration of markers of bacterial plasmalogen and endotoxin in the blood. Microbiome studies were carried out by gas chromatography - mass spectrometry of microbial markers in the blood. A statistically significant increase in blood concentrations of the total level of microbial markers of bacterial plasmalogen and endotoxin was determined in multiple sclerosis, which may be associated with an increase in the permeability of the intestinal wall. In multiple sclerosis, the proportion of coccal, gram-negative, anaerobic microflora with a proteolytic type of metabolic activity increases. The correlations of the representation of microbiota phylotypes change due to the switching of the direct relationship Proteobacteria-Bacteroides to Proteobacteria-Firmicutes. In multiple sclerosis, Actinobacteria and Proteobacteria increase and Firmicutes decrease. Conclusion. The multiple sclerosis disease may be associated with pathological changes in the structure of the microbiome and the growth of endotoxemia, which may be one of the factors in the pathogenesis of the disease. New laboratory markers for diagnosing and predicting the course of MS have been proposed.

One of the reasons for the emergence of highly resistant strains is associated with the ability of bacteria to form biofilms on various surfaces. The formation of a biofilm by pathogens leads to a decrease in the activity of the antibiotic, an increase in the time for the production of stress response genes by bacteria, and, as a result, an increase in antimicrobial tolerance. To investigate the effect of imipenem and cefepime on the activity of biofilm forms of K. pneumoniae bacteria isolated from the wounds of patients with chronic osteomyelitis. The object of the study is clinical strains of K. pneumoniae isolated from the wounds of patients with chronic osteomyelitis. In the control series, the level of biofilm formation of K. pneumoniae strains was assessed after 48 hours of cultivation on coverslips and 96-well polystyrene plates. In the second and third series, the biofilm form of K. pneumoniae bacteria was exposed to imipenem and cefepime, and after 24 hours the activity of biofilm formation was assessed according to previously developed criteria. The structure of the emerging biofilm on the surface of the coverslip in all series of the experiment was represented by single adherent cells and microcolonies of various sizes. Cultivation with antibiotics led to a decrease in the number of microcolonies ranging in size from 10 to 10,000 µm2 in the second and third series, however, significant differences from the control series were found only when exposed to cefepime. The intensity of film formation of K. pneumoniae in the control series by the tablet method was 0.350 (0.334; 0.368) units opt.pl. When cultivating biofilms together with antibacterial drugs, the biofilm-forming activity after 24 hours of the experiment was significantly lower than in the control group in all experimental series. K. pneumoniae bacteria isolated from patients with chronic osteomyelitis, when cultivated on polystyrene plates and on the surface of coverslips, actively form a biofilm, exhibiting highly adhesive properties. The studied antibiotics were shown to have a bacteriostatic effect on biofilm forms of K. pneumoniae bacteria. The bactericidal effect of imipenem and cefepime on biofilm forms was not revealed.

The widespread use of traditional removable prosthetics is explained by the relative simplicity of the technological stages of manufacture and determines its availability. The development of prosthetic stomatitis of the oral cavity is facilitated by poor fixation and stabilization of removable orthopedic structures. Microbiome biofilms formed on the surface of dental orthopedic structures can help reduce their service life and cause an inflammatory process of the oral cavity of microbial etiology during dental prosthetics in the process of orthopedic rehabilitation. The purpose of the study: to assess the level of adaptation of patients during orthopedic rehabilitation based on the study of the microbiome and the assessment of the degree of fixation of removable lamellar dentures. Qualitative and quantitative characteristics of the microbiome of prostheses at the stages of orthopedic pealitation were assessed; facultative anaerobic species belonging to the genera Staphylococcus, Micrococcus, Enterococcus, Streptococcus, Klebsiella prevailed;noted the elimination of microorganisms of the genera Bifidobacterium and Lactobacterium, yeast-like fungi of the species Candida albicans were isolated. An analysis of the index of fixation of prostheses showed an increase depending on the duration of use; a good level of fixation of prostheses was established in groups of patients.