Genetic structure and drug resistance of <i>Mycobacterium tuberculosis</i> strains in the Kemerovo Region — Kuzbass

Background. Kemerovo Region has a high burden of tuberculosis (TB) with incidence rates twice the national average. The circulating variants of Mycobacterium tuberculosis significantly influence the TB epidemic process. Screening of epidemically significant variants of the pathogen in areas with a high burden of TB underlies epidemiological diagnosis and is necessary for the development of effective prevention measures. However, the population structure of M. tuberculosis in the Kemerovo Region — Kuzbass is poorly understood. Aims: to study genetic heterogeneity and phenotypic resistance to anti-tuberculosis drugs of M. tuberculosis strains in the Kemerovo Region. Materials and methods. The MIRU-VNTR genotyping of 163 M. tuberculosis strains isolated from TB patients in the Kemerovo Region in March–October 2022 was carried out. Cultivation of M. tuberculosis, drug susceptibility testing, and isolation of genomic DNA were carried out by standard methods. Genotypic identification was performed using MIRU-VNTR (24 loci) typing. In parallel, express genotyping was carried out: identification of isolates of the Beijing genotype (by RD105/207) and non-Beijing; subtyping Beijing using real-time PCR tests for detection of Central Asian Russian and B0/W148; identification of the non-Beijing group by real-tine PCR RT tests for LAM, S, Ural. Results. The isolates of the Beijing genotype (67.5%) were found to dominate both among newly diagnosed (64.4%) and previously treated patients (88.5%). MIRU-VNTR typing revealed 75 profiles, of which 94-32 (35.3%) and 100-32 (15.7%) were the most abundant and belonged to the Beijing genotype. Overall, 39.9% and 20.9% of isolates, respectively, were assigned to the Beijing Central Asian Russian and B0/W148 epidemic clusters, which differed significantly in MDR levels (50.8% and 85.3%, respectively; p = 0.005). The second most common were strains of the genetic family of the Euro-American lineage (L4) (31.9%): LAM (6.7%) Ural (7.4%) Haarlem (4.9%) and L4-unclassified (12.9%), MDR among of these minor genotypes was significantly lower than among Beijing genotype strains, and amounted to 11.5% (p 0.001). Strains from HIV-TB patients (56.4% of the total sample) carried an MDR profile more often (54.8%) compared to TB cases without HIV infection (35.2%) (p = 0.005), which may be due to higher proportion of Beijing genotype strains in the HIV-TB group (75.0% vs. 57.7%; p = 0.026). Complete comparability of the SNP analysis (in-house tests) to identify the main genotypes and epidemically significant Beijing subtypes was shown, which made it possible to characterize 75.5% of the sample by the express method. Conclusions. The molecular genetic screening carried out in the Kemerovo Region revealed the heterogeneity of the M. tuberculosis population, which was dominated by strains of the Beijing genotype, with the frequency of subtypes comparable with other territories of the Siberian Federal District

Core Proteome of the Minimal Cell: Comparative Proteomics of Three Mollicute Species

Mollicutes (mycoplasmas) have been recognized as highly evolved prokaryotes with an extremely small genome size and very limited coding capacity. Thus, they may serve as a model of a ‘minimal cell’: a cell with the lowest possible number of genes yet capable of autonomous self-replication. We present the results of a comparative analysis of proteomes of three mycoplasma species: A. laidlawii, M. gallisepticum, and M. mobile. The core proteome components found in the three mycoplasma species are involved in fundamental cellular processes which are necessary for the free living of cells. They include replication, transcription, translation, and minimal metabolism. The members of the proteome core seem to be tightly interconnected with a number of interactions forming core interactome whether or not additional species-specific proteins are located on the periphery. We also obtained a genome core of the respective organisms and compared it with the proteome core. It was found that the genome core encodes 73 more proteins than the proteome core. Apart of proteins which may not be identified due to technical limitations, there are 24 proteins that seem to not be expressed under the optimal conditions

The Relationship between the Structure of the Tick-Borne Encephalitis Virus Strains and Their Pathogenic Properties

<div><p>Tick-borne encephalitis virus (TBEV) is transmitted to vertebrates by taiga or forest ticks through bites, inducing disease of variable severity. The reasons underlying these differences in the severity of the disease are unknown. In order to identify genetic factors affecting the pathogenicity of virus strains, we have sequenced and compared the complete genomes of 34 Far-Eastern subtype (FE) TBEV strains isolated from patients with different disease severity (Primorye, the Russian Far East). We analyzed the complete genomes of 11 human pathogenic strains isolated from the brains of dead patients with the encephalitic form of the disease (Efd), 4 strains from the blood of patients with the febrile form of TBE (Ffd), and 19 strains from patients with the subclinical form of TBE (Sfd). On the phylogenetic tree, pathogenic Efd strains formed two clusters containing the prototype strains, Senzhang and Sofjin, respectively. Sfd strains formed a third separate cluster, including the Oshima strain. The strains that caused the febrile form of the disease did not form a separate cluster. In the viral proteins, we found 198 positions with at least one amino acid residue substitution, of which only 17 amino acid residue substitutions were correlated with the variable pathogenicity of these strains in humans and they authentically differed between the groups. We considered the role of each amino acid substitution and assumed that the deletion of 111 amino acids in the capsid protein in combination with the amino acid substitutions R16K and S45F in the NS3 protease may affect the budding process of viral particles. These changes may be the major reason for the diminished pathogenicity of TBEV strains. We recommend Sfd strains for testing as attenuation vaccine candidates.</p></div

Tertiary structures of RNA-dependent RNA polymerase (RdRp).

<p>(A) The tertiary structure of RdRp for the pathogenic strain Dalnegorsk. (B) The tertiary structure of RdRp for the Sfd strain Primorye-270. In both models, the various subdomains are colored as follows: blue – fingers, green – palm, and pink – thumb. The amino acid residue substitutions in both models are indicated by red atoms.</p

ML phylogenetic tree of TBEV strains.

<p>The tree was based on the complete genomes of strains inducing diseases of different severity. Pathogenic strains Efd are shown in black, Sfd strains in green and strains with the febrile form of TBEV are shown in red. Prototype strains are shown in blue. Numbers above or below branches indicate posterior node probabilities and bootstrap values from NJ analysis.</p

Aligned nucleotide sequences of the 5′ UTR.

<p>Pathogenic strains Efd are shown in black, Sfd strains in green and strains with the febrile form of TBEV are shown in red. Prototype strains are shown in blue. The nucleotides identical to the sequence of strain Sofjin are indicated by dots of the corresponding color.</p

Analyzed TBEV strains.

a<p>suckling mouse brain.</p>b<p>Efd, encephalitis form of disease; Ffd, febrile form; IF, Sfd, subclinical form.</p><p>Note: “?” passage history not available</p

Key amino acid substitutions in pathogenic and unapparent strains of TBEV

<p>Note: the Ffd strains that cause the febrile form of the disease are shown in italics; pathogenic Efd strains are in bold.</p><p>• - the same amino acid as in strain Sofjin-HO.</p

Tertiary structure of NS3 protease.

<p>The crystal structure of the West Nile protease (PDB code 3e90) was taken as a template for homology modeling. (A) The tertiary structure of NS3/NS2B for the pathogenic strain Dalnegorsk. (B) The tertiary structure of NS3/NS2B for the Sfd strain Primorye-270. The arrows indicate the replacement of key amino acids. The active center is indicated by the rectangle and its amino acid residues are shown as red atoms.</p

The scheme of the polyprotein and the position of key amino acid substitutions.

<p>The scheme of the polyprotein and the position of key amino acid substitutions.</p