Unraveling the adaptive strategies of Mycoplasma hominis through proteogenomic profiling of clinical isolates.

Introduction: Mycoplasma hominis (M. hominis) belongs to the class Mollicutes, characterized by a very small genome size, reduction of metabolic pathways, including transcription factors, and the absence of a cell wall. Despite this, they adapt well not only to specific niches within the host organism but can also spread throughout the body, colonizing various organs and tissues. The adaptation mechanisms of M. hominis, as well as their regulatory pathways, are poorly understood. It is known that, when adapting to adverse conditions, Mycoplasmas can undergo phenotypic switches that may persist for several generations. Methods: To investigate the adaptive properties of M. hominis related to survival in the host, we conducted a comparative phenotypic and proteogenomic analysis of eight clinical isolates of M. hominis obtained from patients with urogenital infections and the laboratory strain H-34. Results: We have shown that clinical isolates differ in phenotypic features from the laboratory strain, form biofilms more effectively and show resistance to ofloxacin. The comparative proteogenomic analysis revealed that, unlike the laboratory strain, the clinical isolates possess several features related to stress survival: they switch carbon metabolism, activating the energetically least advantageous pathway of nucleoside utilization, which allows slowing down cellular processes and transitioning to a starvation state; they reconfigure the repertoire of membrane proteins; they have integrative conjugative elements in their genomes, which are key mediators of horizontal gene transfer. The upregulation of the methylating subunit of the restriction-modification (RM) system type I and the additional components of RM systems found in clinical isolates suggest that DNA methylation may play a role in regulating the adaptation mechanisms of M. hominis in the host organism. It has been shown that based on the proteogenomic profile, namely the genome sequence, protein content, composition of the RM systems and additional subunits HsdM, HsdS and HsdR, composition and number of transposable elements, as well as the sequence of the main variable antigen Vaa, we can divide clinical isolates into two phenotypes: typical colonies (TC), which have a high growth rate, and atypical (aTC) mini-colonies, which have a slow growth rate and exhibit properties similar to persisters. Discussion: We believe that the key mechanism of adaptation of M. hominis in the host is phenotypic restructuring, leading to a slowing down cellular processes and the formation of small atypical colonies. This is due to a switch in carbon metabolism and activation the pathway of nucleoside utilization. We hypothesize that DNA methylation may play a role in regulating this switch.

Shifts in the Human Gut Microbiota Structure Caused by Quadruple Helicobacter pylori Eradication Therapy.

The human gut microbiome plays an important role both in health and disease. Use of antibiotics can alter gut microbiota composition, which can lead to various deleterious events. Here we report a whole genome sequencing metagenomic/genomic study of the intestinal microbiota changes caused by Helicobacter pylori (HP) eradication therapy. Using approaches for metagenomic data analysis we revealed a statistically significant decrease in alpha-diversity and relative abundance of Bifidobacterium adolescentis due to HP eradication therapy, while the relative abundance of Enterococcus faecium increased. We have detected changes in general metagenome resistome profiles as well: after HP eradication therapy, the ermB, CFX group, and tetQ genes were overrepresented, while tetO and tetW genes were underrepresented. We have confirmed these results with genome-resolved metagenomic approaches. MAG (metagenome-assembled genomes) abundance profiles have changed dramatically after HP eradication therapy. Focusing on ermB gene conferring resistance to macrolides, which were included in the HP eradication therapy scheme, we have shown a connection between antibiotic resistance genes (ARGs) and some overrepresented MAGs. Moreover, some E. faecium strains isolated from stool samples obtained after HP eradication have manifested greater antibiotic resistance in vitro in comparison to other isolates, as well as the higher number of ARGs conferring resistance to macrolides and tetracyclines.

Draft genomes of Enterococcus faecium strains isolated from human feces before and after eradication therapy against Helicobacter pylori.

The abundance of Enterococci in the human intestinal microbiota environment is usually < 0.1% of the total bacterial fraction. The multiple resistance to antibiotics of the opportunistic Enterococcus spp. is alarming for the world medical community because of their high prevalence among clinically significant strains of microorganisms. Enterococci are able to collect different mobile genetic elements and transmit resistance to antibiotics to wide range of Gram-positive and Gram-negative species of microorganisms, including the transmission of vancomycin resistance to methicillin-resistant strains of Staphylococcus aureus. The number of infections caused by antibiotics resistant strains of Enterococcus spp. is increasing. Here we present a draft genomes of Enterococcus faecium strains. These strains were isolated from human feces before and after (1 month) Helicobacter pylori eradication therapy. The samples were subject to whole-genome sequencing using Illumina HiSeq. 2500 platform. The data is available at NCBI https://www.ncbi.nlm.nih.gov/bioproject/PRJNA412824.

Data on gut metagenomes of the patients with Helicobacter pylori infection before and after the antibiotic therapy.

Antibiotic therapy can lead to the disruption of gut microbiota community with possible negative outcomes for human health. One of the diseases for which the treatment scheme commonly included antibiotic intake is Helicobacter pylori infection. The changes in taxonomic and functional composition of microbiota in patients can be assessed using "shotgun" metagenomic sequencing. Ten stool samples were collected from 4 patients with Helicobacter pylori infection before and directly after the H. pylori eradication course. Additionally, for two of the subjects, the samples were collected 1 month after the end of the treatment. The samples were subject to "shotgun" (whole-genome) metagenomic sequencing using Illumina HiSeq platform. The reads are deposited in the ENA (project ID: PRJEB18265).