The strain Streptomyces xinghaiensis ( fradiae ) ATCC19609 is sensitive to antibiotics of various chemical classes and is hypersensitive to the macrolide antibiotic oligomycin A. The spontaneous mutant S. fradiae -nitR + bld, resistant to nitron-oligomycin, carries a mutation in the gene of the PadR transcriptional regulator. A comparative proteomic analysis of the inverted membrane vesicles of the mutant strain and wild-type S. xinghaiensis ( fradiae ) ATCC19609 was performed. Using mass spectrometric analysis, quantitative changes in the protein fractions of two ABC transporters, leucyl aminopeptidase, alkaline phosphatase, and alanine dehydrogenase were detected in vesicles of the mutant strain compared with wild-type vesicles. As a result of transcriptional analysis, an increase in the expression levels of the genes of ABC transporters, alkaline phosphatase, and leucyl aminopeptidase and a decrease in the expression level of the alanine dehydrogenase gene in the mutant strain S. fradiae -nitR + bld compared with the wild-type strain were recorded. This suggested that the padR gene, in which the mutation was detected, may be involved in the regulation of antibiotic resistance and differentiation.

In the present work, we analyze the mutant strains of Streptomyces fradiae ATCC 19609 resistant to (33S)-azido-33-deoxyoligomycin A, obtained at a frequency of 10 –8 to 2.5 × 10 –9 . Analysis of the obtained mutant strains allows them to be divided into two groups based on the level of resistance to oligomycin A. One mutant out of 15 obtained is resistant to oligomycin A at MIC of 0.1 nm/mL, and 14 mutant strains are resistant at MIC of 0.01 nm/mL of oligomycin A (MIC for the initial strain: 0.001 nm/mL). Using PCR analysis and subsequent sequencing by the Sanger method, three candidate genes the products of which are related to resistance to oligomycin A are analyzed: helicase IV, the C subunit and the A subunit of FoF1-ATP synthase. It is found that, in the genomes of all 15 mutants, there is a single nucleotide substitution (SNP) in the helicase IV gene, while in one strain most resistant to oligomycin A there is an additional SNP in the gene of the A subunit of FoF1-ATP synthase. This mutant strain will be used to obtain the mutants of resistance to oligomycin A with the aim of further sequencing of genomes and detection of biotarget of oligomycin A.

Data of draft genome sequencing of Streptomyces fradiae-АТСС19609-Olg4R strain, resistant to (33S)-33-deoxy-33-thiocyanatooligomycin A, are presented. Comparative analysis of the genomes of S. fradiae wild-type strain ATCC19609 and S. fradiae strain ATCC19609-Olg4R showed the single nucleotide substitution that led to A(600)T change in the conservative P-loop NTPase region of class IV helicase gene, which, probably, promoted the resistance of S. fradiae strain ATCC19609-Olg4R to (33S)-33-deoxy-33-thiocyanatooligomycin A.

The paper provides the annotation and data on sequencing the antibiotic resistance genes in Streptomyces fradiae strain ATCC19609, highly sensitive to different antibiotics. Genome analysis revealed four groups of genes that determined the resistome of the tested strain. These included classical antibiotic resistance genes (nine aminoglycoside phosphotransferase genes, two beta-lactamase genes, and the genes of puromycin N-acetyltransferase, phosphinothricin N-acetyltransferase, and aminoglycoside acetyltransferase); the genes of ATP-dependent ABC transporters, involved in the efflux of antibiotics from the cell (MacB-2, BcrA, two-subunit MDR1); the genes of positive and negative regulation of transcription (whiB and padR families); and the genes of post-translational modification (serine-threonine protein kinases). A comparative characteristic of aminoglycoside phosphotransferase genes in S. fradiae ATCC19609, S. lividans TK24, and S. albus J1074, the causative agent of actinomycosis, is provided. The possibility of using the S. fradiae strain ATCC19609 as the test system for selection of the macrolide antibiotic oligomycin A derivatives with different levels of activity is demonstrated. Analysis of more than 20 semisynthetic oligomycin A derivatives made it possible to divide them into three groups according to the level of activity: inactive (>1 nmol/disk), 10 substances; with medium activity level (0.05–1 nmol/disk), 12 substances; and more active (0.01–0.05 nmol/disk), 2 substances. Important for the activity of semisynthetic derivatives is the change in the position of the 33rd carbon atom in the oligomycin A molecule.