Relevance of the bacteriophage adherence to mucus model for Pseudomonas aeruginosa phages.

Pseudomonas aeruginosa infections are getting increasingly serious as antimicrobial resistance spreads. Phage therapy may be a solution to the problem, especially if improved by current advances on phage-host studies. As a mucosal pathogen, we hypothesize that P. aeruginosa and its phages are linked to the bacteriophage adherence to mucus (BAM) model. This means that phage-host interactions could be influenced by mucin presence, impacting the success of phage infections on the P. aeruginosa host and consequently leading to the protection of the metazoan host. By using a group of four different phages, we tested three important phenotypes associated with the BAM model: phage binding to mucin, phage growth in mucin-exposed hosts, and the influence of mucin on CRISPR immunity of the bacterium. Three of the tested phages significantly bound to mucin, while two had improved growth rates in mucin-exposed hosts. Improved phage growth was likely the result of phage exploitation of mucin-induced physiological changes in the host. We could not detect CRISPR activity in our system but identified two putative anti-CRISPR proteins coded by the phage. Overall, the differential responses seen for the phages tested show that the same bacterial species can be targeted by mucosal-associated phages or by phages not affected by mucus presence. In conclusion, the BAM model is relevant for phage-bacterium interactions in P. aeruginosa, opening new possibilities to improve phage therapy against this important pathogen by considering mucosal interaction dynamics.IMPORTANCESome bacteriophages are involved in a symbiotic relationship with animals, in which phages held in mucosal surfaces protect them from invading bacteria. Pseudomonas aeruginosa is one of the many bacterial pathogens threatening humankind during the current antimicrobial resistance crisis. Here, we have tested whether P. aeruginosa and its phages are affected by mucosal conditions. We discovered by using a collection of four phages that, indeed, mucosal interaction dynamics can be seen in this model. Three of the tested phages significantly bound to mucin, while two had improved growth rates in mucin-exposed hosts. These results link P. aeruginosa and its phages to the bacteriophage adherence to the mucus model and open opportunities to explore this to improve phage therapy, be it by exploiting the phenotypes detected or by actively selecting mucosal-adapted phages for treatment.

Characterization of a Bacteriophage GEC_vB_Bfr_UZM3 Active against Bacteroides fragilis.

Bacteroides fragilis is a commensal gut bacterium that is associated with a number of blood and tissue infections. It has not yet been recognized as one of the drug-resistant human pathogens, but cases of the refractory infections, caused by strains that are not susceptible to the common antibiotic regimes established for B. fragilis, have been more frequently reported. Bacteriophages (phages) were found to be a successful antibacterial alternative to antibiotic therapy in many cases of multidrug-resistant (MDR) bacterial infections. We have characterized the bacteriophage GEC_vB_Bfr_UZM3 (UZM3), which was used for the treatment of a patient with a chronic osteomyelitis caused by a B. fragilis mixed infection. Studied biological and morphological properties of UZM3 showed that it seems to represent a strictly lytic phage belonging to a siphovirus morphotype. It is characterized by high stability at body temperature and in pH environments for about 6 h. Whole genome sequencing analysis of the phage UZM3 showed that it does not harbor any known virulence genes and can be considered as a potential therapeutic phage to be used against B. fragilis infections.

Development of an in vitro homeostasis model between airway epithelial cells, bacteria and bacteriophages: a time-lapsed observation of cell viability and inflammatory response.

Bacteriophages represent the most extensive group of viruses within the human virome and have a significant impact on general health and well-being by regulating bacterial population dynamics. Staphylococcus aureus, found in the anterior nostrils, throat and skin, is an opportunistic pathobiont that can cause a wide range of diseases, from chronic inflammation to severe and acute infections. In this study, we developed a human cell-based homeostasis model between a clinically isolated strain of S. aureus 141 and active phages for this strain (PYOSa141) isolated from the commercial Pyophage cocktail (PYO). The cocktail is produced by Eliava BioPreparations Ltd. (Tbilisi, Georgia) and is used as an add-on therapy for bacterial infections, mainly in Georgia. The triptych interaction model was evaluated by time-dependent analysis of cell death and inflammatory response of the nasal and bronchial epithelial cells. Inflammatory mediators (IL-8, CCL5/RANTES, IL-6 and IL-1ß) in the culture supernatants were measured by enzyme-linked immunosorbent assay and cell viability was determined by crystal violet staining. By measuring trans-epithelial electrical resistance, we assessed the epithelial integrity of nasal cells that had differentiated under air-liquid interface conditions. PYOSa141 was found to have a prophylactic effect on airway epithelial cells exposed to S. aureus 141 by effectively down-regulating bacterial-induced inflammation, cell death and epithelial barrier disruption in a time-dependent manner. Overall, the proposed model represents an advance in the way multi-component biological systems can be simulated in vitro.

In Vitro Evaluation of the Therapeutic Potential of Phage VA7 against Enterotoxigenic Bacteroides fragilis Infection.

Since the beginning of the 20th century, bacteriophages (phages), i.e., viruses that infect bacteria, have been used as antimicrobial agents for treating various infections. Phage preparations targeting a number of bacterial pathogens are still in use in the post-Soviet states and are experiencing a revival in the Western world. However, phages have never been used to treat diseases caused by Bacteroides fragilis, the leading agent cultured in anaerobic abscesses and postoperative peritonitis. Enterotoxin-producing strains of B. fragilis have been associated with the development of inflammatory diarrhea and colorectal carcinoma. In this study, we evaluated the molecular biosafety and antimicrobial properties of novel phage species vB_BfrS_VA7 (VA7) lysate, as well as its impact on cytokine IL-8 production in an enterotoxigenic B. fragilis (ETBF)-infected colonic epithelial cell (CEC) culture model. Compared to untreated infected cells, the addition of phage VA7 to ETBF-infected CECs led to significantly reduced bacterial counts and IL-8 levels. This in vitro study confirms the potential of phage VA7 as an antibacterial agent for use in prophylaxis or in the treatment of B. fragilis infections and associated colorectal carcinoma.

Investigation of Salmonella Phage-Bacteria Infection Profiles: Network Structure Reveals a Gradient of Target-Range from Generalist to Specialist Phage Clones in Nested Subsets.

Bacteriophages that lyse Salmonella enterica are potential tools to target and control Salmonella infections. Investigating the host range of Salmonella phages is a key to understand their impact on bacterial ecology, coevolution and inform their use in intervention strategies. Virus-host infection networks have been used to characterize the "predator-prey" interactions between phages and bacteria and provide insights into host range and specificity. Here, we characterize the target-range and infection profiles of 13 Salmonella phage clones against a diverse set of 141 Salmonella strains. The environmental source and taxonomy contributed to the observed infection profiles, and genetically proximal phages shared similar infection profiles. Using in vitro infection data, we analyzed the structure of the Salmonella phage-bacteria infection network. The network has a non-random nested organization and weak modularity suggesting a gradient of target-range from generalist to specialist species with nested subsets, which are also observed within and across the different phage infection profile groups. Our results have implications for our understanding of the coevolutionary mechanisms shaping the ecological interactions between Salmonella phages and their bacterial hosts and can inform strategies for targeting Salmonella enterica with specific phage preparations.

Characterization of Salmonella Isolates from Various Geographical Regions of the Caucasus and Their Susceptibility to Bacteriophages.

Non-typhoidal Salmonella present a major threat to animal and human health as food-borne infectious agents. We characterized 91 bacterial isolates from Armenia and Georgia in detail, using a suite of assays including conventional microbiological methods, determining antimicrobial susceptibility profiles, matrix assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry, serotyping (using the White-Kauffmann-Le Minor scheme) and genotyping (repetitive element sequence-based PCR (rep-PCR)). No less than 61.5% of the isolates were shown to be multidrug-resistant. A new antimicrobial treatment strategy is urgently needed. Phage therapy, the therapeutic use of (bacterio-) phages, the bacterial viruses, to treat bacterial infections, is increasingly put forward as an additional tool for combatting antibiotic resistant infections. Therefore, we used this representative set of well-characterized Salmonella isolates to analyze the therapeutic potential of eleven single phages and selected phage cocktails from the bacteriophage collection of the Eliava Institute (Georgia). All isolates were shown to be susceptible to at least one of the tested phage clones or their combinations. In addition, genome sequencing of these phages revealed them as members of existing phage genera (Felixounavirus, Seunavirus, Viunavirus and Tequintavirus) and did not show genome-based counter indications towards their applicability against non-typhoidal Salmonella in a phage therapy or in an agro-food setting.

Georgia's healthcare system and integration of complementary medicine.

BACKGROUND: An increasing number of people in the Republic of Georgia use complementary and alternative medicine (CAM). CAM has long been practiced in the country, but is not currently part of the formal medical system. The aim of this study was to explore patients' CAM use and their perspectives of CAM complementation of existing services in Georgia's health care system. METHODOLOGY: We conducted a qualitative study exploring patient needs and wants by performing in-depth, contextual interviews with patients using CAM. We recruited participants at CAM clinics and collected data until we reached saturation. A thematic analysis involving line by line coding explored perspectives and allowed us to formulate recommendations of CAM integration in Georgia. RESULTS: Study participants voiced that they pursued cure beyond symptom relief; their disappointment in that regard with conventional medicine in Georgia directed them towards CAM as a safe and effective care setting. Most sought natural approaches as a sustained approach to their health and perceived CAM as empathetic therapeutic and preventive space. Participants were in favor of integration of CAM with conventional health services through government support and regulation. They saw coverage of payments for CAM visits and treatments as important aspect for an effective and sustainable integration. CONCLUSION: Patients favor an integration of CAM into Georgia's current healthcare systems to ensure CAM's broad availability as well as effective regulation and financing, including coverage by health insurance.

Selection of Potential Therapeutic Bacteriophages that Lyse a CTX-M-15 Extended Spectrum ß-Lactamase Producing Salmonella enterica Serovar Typhi Strain from the Democratic Republic of the Congo.

Recently, a Salmonella Typhi isolate producing CTX-M-15 extended spectrum ß-lactamase (ESBL) and with decreased ciprofloxacin susceptibility was isolated in the Democratic Republic of the Congo. We have selected bacteriophages that show strong lytic activity against this isolate and have potential for phage-based treatment of S. Typhi, and Salmonella in general.