Novel lytic bacteriophage vB_GEC_EfS_9 against Enterococcus faecium.

Enterococcus spp. is a common commensal microorganism, however, some strains can cause opportunistic infections in humans. Treatment of Enterococcus faecium-related endocarditis, urinary and genital tract infections, meningitis, septicemia, and even neonatal sepsis is often complicated by antibiotic resistance. The spread of multi-resistant bacterial strains has renewed interest in phage therapy, which has many advantages: Its advantages include a much lower frequency of resistance development compared to antibiotics and strict specificity, which allows affecting of only their target microbes without disturbing necessary microbiome. We isolated and characterized a virulent bacteriophage which is active against Enterococcus faecium clinical strains. The phage, which was designated as vB_GEC_EfS_9 was studied in terms of its growth pattern and adsorption rate, as well as its host range. The whole genome of the phage was sequenced and analyzed. Obtained results indicate that phage vB_GEC_EfS_9 is a virulent phage which has a very good potential for therapeutic use against strains of E. faecium.

BIOLOGICAL CHARACTERIZATION OF BACTERIOPHAGES AGAINST STREPTOCOCCUS AGALACTIAE.

Streptococcus agalactiae, also known as group B streptococci, was first isolated from cow's milk with mastitis, and it was first identified in 1930 by Rebecca Lancefield. GBS or streptococcus agalacticae is a gram-positive cocci, beta-hemolytic, facultative anaerobic, which is a harmless inhabitant of the normal human microflora. About 30% of the population is an asymptomatic carrier of this microbe. However, it was considered the leading cause of neonatal invasions upon its discovery. GBS is encapsulated in a polysaccharide capsule, which is a major virulence factor, and 10 serotypes of group B streptococci are known for their immunological activity. According to the existing studies, vaginal colonization is most often caused by type Ia, III, and V in other countries of the world. Today number of invasive GBS diseases continues to increase and it remains a significant pathogen among both infants and adults. The use of broad-spectrum antibiotics has certainly played a major role in the fight against infections, although the number of antibiotic-resistant strains has increased and thus modern medicine starts new insights into alternative antibacterials. Bacteriophages are often suggested as an alternative therapeutic agent against bacterial infections. In the present study, our aim was to isolate and study bacteriophages active against group B streptococci. Between March 2019 and March 2020, 257 anovaginal swabs were collected at "Nia Oniashvili Clinic" Tbilisi, Georgia, and Clinic "Curatio"by using standard bacterial identification procedures, in particular group B streptococcus identification guideline provided by CDC we identified 87 Streptococcus agalactiae strains. We used both standard and modified methods to isolate bacteriophages and study their life cycle, which were developed by the team. In this study, two bacteriophages active against Streptococcus agalactiae were identified: vB_GBS_1 (Syphoviridae) and vB_GBS_2 (Syphoviridae). The biological characteristics, morphology, adsorption, and host range were studied.

IDENTIFICATION AND ANTIBIOTIC SUSCEPTIBILITY PATTERNS OF STREPTOCOCCUS AGALACTIAE.

GBS-Group B streptococcus or Streptococcus agalactiae is a Gram-positive coccus found in 20% of healthy women as part of normal gastrointestinal and genital tract flora. It is associated with pathogenicity in immunocompromised, elderly and pregnant adults as well as infants and neonates. GBS are encapsulated organisms and sub divided into ten antigenically distinct capsular serotypes. GBS case treatment mainly performed with Penicillin G but Erythromicyn, Vancomycin and Clindamycin are recommended for penicillin-allergic individuals. In this study between March 2019 and September 2019 an anovaginal swabs were collected from 80 women at 35-37 weeks of gestation and 39 were identified as str. agalactiae. 24 str. agalactiae strains provided by clinic "Curation" were isolated from urine, vagina and rectum. Culturing, identification of GBS were carried out according to standard microbiological methods. Antimicrobial susceptibility profiles were determined by disk diffusion method. Our results revealed that in total from 104 samples 29% were Str. agalactiae. All GBS strains were all sensitive to Penicillin, Ampicilin and Vancomycin. 25% were resistant to Erythromycin and 13% to Clindamycin. Our study has shown that the prevalence of Streptococcus agalactiae in Georgia quite high and especially with Erythromycin-resistant strains, which makes treatment of penicillin allergic patients problematic. In this case only hope is vancomycin. Fortunately, vancomycin-resistant strains have not been identified; however the problem is still relevant. Obtained data could be used as a base for further epidemiological studies and also managing targeted new approaches of treatment and prevention of GBS.

NEW TEMPERATE ENTEROCOCCUS PHAGE VB_GEC_EFS_2 WITH POTENTIAL OF LYSOGENIC CONVERSION.

Bacteriophages represent the widest group of viruses, from which only virulent phages are used as antibacterial agent. But the picture in the case of temperate phage is absolutely different; many lysogenic phages express gene products that have subtle effects on the phenotype of the host cell. This process is called lysogenic conversion. In present study we characterized new temperate Enterococcus faecium phage vB_GEC_EFS_2, which was isolated from river Mtkvari. The phage is a member of Siphoviridae family. Whole genome of phage vB_GEC-EfS_2 was sequenced and analyzed. Total length of the genome of phage vB_GEC_EFS_2 is 38 508bp, The assembly contains 65 ORFs, among them - 3 lysis genes , genes coded 13 structural proteins, 1 DNA replication-associated gene, 1 gene coded integration, 3 - lysis-lysogenic cycle regulation, 43 hypothetical proteins. One holin gene contained "Haemolysin XhIA" domain which is surface associated haemolisyn. We isolated and purified holin gene and determine its haemolitic activity alongside with vB_GEC_EfS_2 phage lysate. We clarified the XhIA domain function and role in protein's haemolytic nature and described another kind of lysogenic conversion.