Development of Resistance to Clarithromycin and Amoxicillin-Clavulanic Acid in Lactiplantibacillus plantarum In Vitro Is Followed by Genomic Rearrangements and Evolution of Virulence.

Ensuring the safety of the use of probiotics is a top priority. Obviously, in addition to studying the beneficial properties of lactic acid bacteria, considerable attention should be directed to assessing the virulence of microorganisms as well as investigating the possibility of its evolution under conditions of selective pressure. To assess the virulence of probiotics, it is now recommended to analyze the genomes of bacteria in relation to the profiles of the virulome, resistome, and mobilome as well as the analysis of phenotypic resistance and virulence in vitro. However, the corresponding procedure has not yet been standardized, and virulence analysis of strains in vivo using model organisms has not been performed. Our study is devoted to testing the assumption that the development of antibiotic resistance in probiotic bacteria under conditions of selective pressure of antimicrobial drugs may be accompanied by the evolution of virulence. In this regard, special attention is required for the widespread in nature commensals and probiotic bacteria actively used in pharmacology and the food industry. As a result of step-by-step selection from the Lactiplantibacillus plantarum 8p-a3 strain isolated from the "Lactobacterin" probiotic (Biomed, Russia), the L. plantarum 8p-a3-Clr-Amx strain was obtained, showing increased resistance simultaneously to amoxicillin-clavulanic acid and clarithromycin (antibiotics, the combined use of which is widely used for Helicobacter pylori eradication) compared to the parent strain (MIC8p-a3-Clr-Amx of 20 µg/mL and 10 µg/mL, and MIC8p-a3 of 0.5 µg/mL and 0.05 µg/mL, respectively). The results of a comparative analysis of antibiotic-resistant and parental strains indicate that the development of resistance to the corresponding antimicrobial drugs in L. plantarum in vitro is accompanied by the following: (i) significant changes in the genomic profile (point mutations as well as deletions, insertions, duplications, and displacement of DNA sequences) associated in part with the resistome and mobilome; (ii) changes in phenotypic sensitivity to a number of antimicrobial drugs; and (iii) an increase in the level of virulence against Drosophila melanogaster, a model organism for which L. plantarum is considered to be a symbiont. The data obtained by us indicate that the mechanisms of adaptation to antimicrobial drugs in L. plantarum are not limited to those described earlier and determine the need for comprehensive studies of antibiotic resistance scenarios as well as the trajectories of virulence evolution in probiotic bacteria in vivo and in vitro to develop a standardized system for detecting virulent strains of the corresponding microorganisms. IMPORTANCE Ensuring the safety of the use of probiotics is a top priority. We found that increased resistance to popular antimicrobial drugs in Lactiplantibacillus plantarum is accompanied by significant changes in the genomic profile and phenotypic sensitivity to a number of antimicrobial drugs as well as in the level of virulence of this bacterium against Drosophila. The data obtained in our work indicate that the mechanisms of antibiotic resistance in this bacterium are not limited to those described earlier and determine the need for comprehensive studies of the potential for the evolution of virulence in lactic acid bacteria in vivo and in vitro and to develop a reliable control system to detect virulent strains among probiotics.

Adaptation to Antimicrobials and Pathogenicity in Mycoplasmas: Development of Ciprofloxacin-Resistance and Evolution of Virulence in Acholeplasma laidlawii.

For the first time it was shown that the development of resistance to ciprofloxacin in vitro in Acholeplasma laidlawii, a mycoplasma which is widely spread in nature and which is the main contaminant of cell cultures and vaccines, is associated with diverse pathways of virulence evolution: virulome and virulence differ significantly between ciprofloxacin-resistant strains, including those with the same level of antimicrobial resistance.

[The age-dependent changes of reproductive function in offspring Drosophila melanogaster.]

It was analyzed a comparative study of age-dependent changes in the parameters of reproductive function and embryonic development of offspring from adults w1c-s, w1Or mutant strains and Canton-S, Oregon-R wild-type strains D. melanogaster. In investigation three age groups of adult drosophila: 3, 10 and 20-days old were used. Reciprocal crosses between young and aging parents enable to define the influence of age of the maternal and/or paternal organism on the parameters of reproductive function. It is shown that the various features of reproductive function in F1 offspring of mutants and wild-type strains are forming differently by the different influence of the age of parents. The established effect of the white mutation, either as an independent factor or in combination with the parental age, is reflected in the low values of the fertility and viability and high values of dominant lethal mutations and mortality on the metamorphosis stage and consequently the appearance of less adapt and viable offspring in the population.

[Research of Embryonic Mortality Stages of Drosophila melanogaster Depending on Age and Starvation of an Imago].

Influence of age of parents and duration of starvation on egg production and demonstration of embryonic mortality at different stages of egg development has been studied. It is shown that, with increasing age of organisms, the overall egg production reduces and the percentage of embryonic mortality increases at 0-5.5 and 5.5-17 h of development. An increase in the duration of starvation also promotes a reduction in egg production in 3- and 10-day-old adult D. melanogaster compared with short-term starvation. A statistically significant effect of factors, such as the allelic state of the white locus, the genetic background, the age of the parents, and the duration of starvation, on all studied parameters was established.