AtbFinder Diagnostic Test System Improves Optimal Selection of Antibiotic Therapy in Persons with Cystic Fibrosis.

Cystic fibrosis (CF) is characterized by mutations of CFTR that lead to increased viscous secretions, bacterial colonization, and recurrent infections. Chronic Pseudomonas aeruginosa infection in persons with CF is associated with progressive and accelerated lung function decline despite aggressive antibiotic treatment. We report the management of respiratory infections in persons with CF with antibiotic therapy that was based on the recommendations of AtbFinder, a novel, rapid, culture-based diagnostic test system that employs a novel paradigm of antibiotic selection. AtbFinder mimics bacterial interactions with antibiotics at concentrations that can be achieved in affected tissues or organs and models conditions of interbacterial interactions within polymicrobial biofilms. This open-label, single-arm, investigator-initiated clinical study was designed to identify the efficacy of antibiotics selected using AtbFinder in persons with CF. Microbiological and clinical parameters were assessed following the change of antibiotic therapy to antibiotics selected with AtbFinder between January 2016 and December 2018 and retrospectively compared with clinical data collected between January 2013 and December 2015. We enrolled 35 persons with CF (33 with chronic P. aeruginosa colonization). Antibiotics selected using AtbFinder resulted in clearance of P. aeruginosa in 81.8% of subsequent cultures, decreased pulmonary exacerbations from 1.21 per patient per annum to 0, and an increase in predicted percent predicted forced expiratory volume in 1 s up to 28.4% from baseline. The number of systemic antibiotic courses used in patients after switching to the AtbFinder-selected therapy was reduced from 355 to 178. These findings describe the superiority of antibiotic regimens selected with AtbFinder compared with routine antimicrobial susceptibility testing.

In Vitro Activity of a Novel Antifungal Compound, MYC-053, against Clinically Significant Antifungal-Resistant Strains of Candida glabrata, Candida auris, Cryptococcus neoformans, and Pneumocystis spp.

An urgent need exists for new antifungal compounds to treat fungal infections in immunocompromised patients. The aim of the current study was to investigate the potency of a novel antifungal compound, MYC-053, against the emerging yeast and yeast-like pathogens Candida glabrata, Candida auris, Cryptococcus neoformans, and Pneumocystis species. MYC-053 was equally effective against the susceptible control strains, clinical isolates, and resistant strains, with MICs of 0.125 to 4.0 µg/ml. Notably, unlike other antifungals such as azoles, polyenes, and echinocandins, MYC-053 was effective against Pneumocystis isolates, therefore being the only synthetic antifungal that may potentially be used against Pneumocystis spp., Candida spp., and Cryptococcus spp. MYC-053 was highly effective against preformed 48-h-old C. glabrata and C. neoformans biofilms, with minimal biofilm eradication concentrations equal to 1 to 4 times the MIC. Together, these data indicated that MYC-053 may be developed into a promising antifungal agent for the treatment and prevention of invasive fungal infections caused by yeasts and yeast-like fungi.

[The choice of antibiotic according the results of standard bacteriological analysis and express-method without isolation of pure cultures.]

The efficiency of choosing antibiotics using two methods was analyzed. The first method is based on standard laboratory analysis including isolation of pure cultures of prospective agent and detection of their individual sensitivity to antibiotics. The second method is based on a new express-technique permitting to simultaneously evaluate effect of pharmaceuticals on cultivated and not cultivated yet bacteria without isolation of pure cultures. At that, the second method permits to determine efficiency of antibiotics namely in those concentrations that can be made in the given nidus of infection. The study demonstrated existence of discrepancy between results obtained by using the given methods. The cause of discrepancy can be presence of not cultivated yet bacteria in pathologic sample. In favor of this assumption testify the received data concerning samples from patients where were presented a significant number of spore-former bacteria having additional mechanisms of resistance to antibiotics. Furthermore, the implemented study established antibiotics characterized by the greatest discrepancy of results obtained by applied methods that indicates the necessity of revision of schemes of implementation of start or empiric therapy.

Effects of Multicide, Antibacterial Drug, on Staphylococcus Biomembranes.

Drug penetration into bacterial biomembranes is one of the most important factors determining the efficiency of antibacterial therapy. Multicide, antibacterial drug, is a nanomolecule 1.3-2.0 nm in size, easily penetrating into staphylococcus biomembranes and causing rapid death of bacteria. The drug efficiency depends on its concentration and duration of exposure. Bacteria die as a result of cell wall perforation, which is associated with changes in its morphology and release of DNA from bacterial cell into the environment. Our results indicate the efficiency of primary damage to bacterial wall leading to elimination of biomembranes.

[The choice of antibiotic in microbiological analysis of phlegm.]

The efficiency of application of the test-system "Vy`borAntibiotika" (AntibioticChoice) in incubation of a maximal possible number of bacteria from pathologic material in case of pneumonia was studied. The results of meta-genome analysis permitted to establish that test-system support incubation of practically all bacteria detected in phlegm, including those attributed to so far non-incubated ones. The comparison of the results was carried out concerning a standard detection of sensitivity of bacteria to antibiotics and choice of efficient medicinal according the results of application of test-system "Vy`borAntibiotika". The obtained data demonstrates that test-system permits to choose antibiotic during 6-20 hours without isolation of pure strain.

[The choice of antibiotic in microbiological study of pyoinflammatory processes.]

The efficiency of application of test-system "Antibiotic Choice" was examined concerning evaluation of sensitivity to medications of maximal possible number of bacteria from pathological samples at burn trauma without isolation of pure culture. The results of metagenome analysis demonstrated that test-system permits supporting factually all bacteria discovered in wound discharge, including ones related to not cultivated yet. The comparison was carried out concerning results of standard identification of sensitivity of bacteria to antibiotics and efficient medication according the results of application of test-system "Antibiotic Choice". The obtained results demonstrate that test-system permits choosing antibiotic during 6-20 hours without separation of pure culture.

[CHOOSING ANTIBIOTIC IN MICROBIOLOGICAL ANALYSIS OF INFECTIONS OF URINARY EXCRETION SYSTEM].

The effectiveness of application of test-system "Choice of antibiotic" was evaluated as a tool for incubation of maximal amount of bacteria from pathological material under acute cystitis. The results of meta-genome analysis established that test-system permits supporting growth of practically all bacteria detected in urine, including ones relating to "uncultivated for the present". The comparison of results of standard detection of sensitivity of bacteria to antibiotics and identification of effective pharmaceutical according the results of application of test-system "Choice of antibiotic" as well was implemented It is demonstrated that test- system permits choosing antibiotic during 6-20 hours wiihout isolation of pure strain.

Characteristics of bacterial biofilms during long-term culturing.

We studied properties of bacterial biofilms during long-term culturing. The biofilms were preserved for more than 90 days under the chosen conditions. At the same time, amounts of matrix and cells underwent periodic changes. The biomass of the biofilms decreased approximately every 7 days and then returned to the previous values.

Seed dressing with M451 promotes seedling growth in wheat and reduces root phytopathogenic fungi without affecting endophytes.

Fungal plant infections result in substantial losses to the agricultural sector. A range of fungicide seed dressings are available to control seed-borne fungal diseases; however, they lack sufficient efficacy because of intrinsic tolerance and acquired resistance. Moreover, many fungicide seed dressings can also penetrate plants, negatively affecting plant growth owing to their toxic effects on endophytes, as well as contributing to the spread of antibiotic resistance. Here, we evaluated the efficacy of M451, a member of a new class of antimicrobial agents that are not relevant to human healthcare. As a seed dressing for wheat seeds, M451 exhibited significant antifungal activity against one of the most devastating plant fungal pathogens, Fusarium spp. Furthermore, M451 was more active than the commercially used fungicide Maxim XL against both seed-borne and soil-borne F. oxysporum infection. Importantly, and unlike other antifungals, M451 seed dressing did not inhibit any of the major characteristics of wheat grains and seedlings, such as germination percentage, germination time, grain vigor, shoot- and root weight and length, but rather improved some of these parameters. The results also demonstrated that M451 had no negative impacts on endophytes and did not accumulate in grains. Thus, M451 may have potential applications as an antifungal agent in wheat cultivation.

Neutrophil Extracellular Traps (NETs): Opportunities for Targeted Therapy.

Antitumor therapy, including adoptive immunotherapy, inevitably faces powerful counteraction from advanced cancer. If hematological malignancies are currently amenable to therapy with CAR-T lymphocytes (T-cells modified by the chimeric antigen receptor), solid tumors, unfortunately, show a significantly higher degree of resistance to this type of therapy. As recent studies show, the leading role in the escape of solid tumors from the cytotoxic activity of immune cells belongs to the tumor microenvironment (TME). TME consists of several types of cells, including neutrophils, the most numerous cells of the immune system. Recent studies show that the development of the tumor and its ability to metastasize directly affect the extracellular traps of neutrophils (neutrophil extracellular traps, NETs) formed as a result of the response to tumor stimuli. In addition, the nuclear DNA of neutrophils - the main component of NETs - erects a spatial barrier to the interaction of CAR-T with tumor cells. Previous studies have demonstrated the promising potential of deoxyribonuclease I (DNase I) in the destruction of NETs. In this regard, the use of eukaryotic deoxyribonuclease I (DNase I) is promising in the effort to increase the efficiency of CAR-T by reducing the NETs influence in TME. We will examine the role of NETs in TME and the various approaches in the effort to reduce the effect of NETs on a tumor.

Optimization of the Protocol for the Isolation and Refolding of the Extracellular Domain of HER2 Expressed in Escherichia coli.

Receptor 2 of the human epidermal growth factor (HER2/neu, c-erbB2) is a 185 kDa proto-oncogene protein characterized by an overexpression in some oncological diseases, including 30% of mammary glands cancers, as well as tumors in the ovary, stomach and other organs of the human body. Since HER2- tumor status testing is the essential part of a successful cancer treatment, the expression and purification of substantial amounts of the extracellular domain (ECD) of HER2 is an important task. The production of ECD HER2 in Escherichia coli has several advantages over the use of eukaryotic expression systems, but the bulk of the recombinant product in bacteria accumulates as insoluble protein inclusion bodies. In this study, we obtained ECD HER2 in Escherichia coli as insoluble inclusion bodies and elaborated a simple, efficient, and fast protocol for the solubilization, refolding, and isolation of the protein in soluble form.