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.

PECULIARITIES OF ACTIVATION OF COMPENSATORY-ADAPTIVE PROCESSES IN ADULT RAT LIVER CAUSED BY UNILATERAL NEPHRECTOMY.

In order to determine the general patterns of activation of the inter organ compensatory-adaptive processes, the peculiarities of activation of compensatory processes caused by unilateral nephrectomy in adult rat liver at the initial stage of hepatopathy (first three days after the operation) have been studied. In particular, it has been established that only small but visible abnormalities in the spatial relationship of sinusoids and hepatic plates are revealed morphologically in rat liver histology. The increasing of the functional load caused by changes in histoarchitecture, at these times, does not stimulate hepatocyte proliferation. At the same time, it is revealed that at the initial stage of hepatopathy caused by unilateral nephrectomy, preference is given to polyploidy from compensatory-adaptive processes charachterised to liver. In particular, it has been shown that liver responses mainly by quantitative increases in binucleated cells (2cx2) to the functional load induced by unilateral nephrectomy at an early stage.

DUCTULAR REACTION AT THE EARLY AND LATE STAGES OF BILIARY OBSTRUCTION: IS THE MECHANISM THE SAME?

Ductular reaction (DR) is one of the classical features of biliary obstruction in humans as well as in experimental models. It is the histological phenomenon expressed by the increased number of ductular profiles (DPs), which is especially intensively studied in rodents' model of common bile duct ligation (CBDL). However, some issues related with DR in long-term CBDL are not clear enough. The liver tissue of male Wistar rats (150-200 g) subjected to CBDL were studied histologically (H&E). The architectonics of the biliary tract was studied by investigation of the corrosion casts obtained by retrograde injection (trough CBD) the solidified mass prepared on the basis of "Protacryl M" widely implemented in orthopedic dentistry. The casts were investigated by electro-optic device ProScope-HR with magnification X10, X50 and X200. DR accompanying CBDO is featured by the development of new DPs, the number of which is permanently increased in association with the prolongation of the terms of Cholestasis. DPs continue to develop in all areas of liver lobules - from portal/periportal region up to the regions containing central veins (hepatic veins tributaries of small calibers). The part of DPs has wide lumens, while the lumens of another part of DPs are hardly notable. In spite of evident ductular metaplasia of hepatic tissue, the small groups of normal by form and structure hepatocytes forming the separate small islets could be revealed. Histological features of these hepatocytes support the proposition that these cells maintain the functional activity on the 4th-5th weeks after CBDO. The results of investigations of biliary corrosion casts on the 4th and 5th weeks of CBDO show that solidifying mass cannot reach the small ducts/ductules and especially bile canaliculi. This fact confirms the interruption of ductular-canalicular junctions (IDCJ). We propose that due to IDCJ the increased pressure in preserved bile canaliculi should stipulate the mitotic activity and biliary trans-differentiation of hepatocytes on the late terms of CBDO and support the generation of ductal-like structures and appearance of new DPs. These ductules generally are not drained into the entire biliary tree.

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.