Phage sensitivity profiles of a nasopharyngeal opportunistic pathogen in Streptococcus pneumoniae carrier children with recurrent respiratory infections

Aim. To study the nature of microbiota and estimating the susceptibility to antibiotics and bacteriophages of conditionally pathogenic microflora of the nasopharynx in children-pneumococcal carriers with recurrent respiratory infections. Methods. Researching microflora was conducted in 182 pneumococcal carriers receiving help in Kazan Scientific and Research Institute of Epidemiology and Microbiology. Microbial identification, testing of susceptibility to antibiotics and bacteriophages was carried out following the regulatory documentation. Bacterial isolates were confirmed by mass spectrometry. The phage titer was determined by the method of agar layers according to Grazia. Results. Nasopharyngeal S. pneumoniae species was presented by Staphylococcus spp., Moraxella spp., Haemophilus spp., Corynebacterium spp., Klebsiella spp and Candida spp. The antimicrobial resistance profiles of Streptococcus pneumoniae: resistant to oxacillin was detected in 20.7% of strains, to erythromycin in 45.9%, to clindamycin in 20%, to trimethoprim-sulfamethoxazole in 18.4%. 19.6% of isolates were multidrug-resistant (MDR, resistant to 3 or more antimicrobial agents). Phage susceptibility test of S. pneumoniaе showed that 97.2% of isolates were resistant to streptococcal bacteriophage, 75% to pyobacteriophage. All antibiotic-resistant strains remained susceptible to Streptococcus phages. The phage titer of Klebsiella in agreement with Grazia method of Kl. pneumoniae ranged from 9106 to 5105 PFU/mL. The ranking results of activities of antistaphylococcal antibiotics (effectiveness descending): fusidic acid mupirocin chloramphenicol cyprofloxacin erythromycin. Conclusion. Nasopharyngeal microbiota of pneumococci carriers children is represented by a variable polymicrobial association; nasopharyngeal strains are effectively lysed by bacteriophages; mono- and polyvalent bacteriophages can be used as an alternative to antibacterial treatment in Streptococcus pneumoniae carriers children with recurrent respiratory infections.

Two Phages of the Genera Felixounavirus Subjected to 12 Hour Challenge on Salmonella Infantis Showed Distinct Genotypic and Phenotypic Changes

Salmonella Infantis is considered in recent years an emerging Salmonella serovar, as it has been associated with several outbreaks and multidrug resistance phenotypes. Phages appear as a possible alternative strategy to control Salmonella Infantis (SI). The aims of this work were to characterize two phages of the Felixounavirus genus, isolated using the same strain of SI, and to expose them to interact in challenge assays to identify genetic and phenotypic changes generated from these interactions. These two phages have a shared nucleotide identity of 97% and are differentiated by their host range: one phage has a wide host range (lysing 14 serovars), and the other has a narrow host range (lysing 6 serovars). During the 12 h challenge we compared: (1) optical density of SI, (2) proportion of SI survivors from phage-infected cultures, and (3) phage titer. Isolates obtained through the assays were evaluated by efficiency of plating (EOP) and by host-range characterization. Genomic modifications were characterized by evaluation of single nucleotide polymorphisms (SNPs). The optical density (600 nm) of phage-infected SI decreased, as compared to the uninfected control, by an average of 0.7 for SI infected with the wide-host-range (WHR) phage and by 0.3 for SI infected with the narrow-host-range (NHR) phage. WHR phage reached higher phage titer (7 × 1011 PFU/mL), and a lower proportion of SI survivor was obtained from the challenge assay. In SI that interacted with phages, we identified SNPs in two genes (rfaK and rfaB), which are both involved in lipopolysaccharide (LPS) polymerization. Therefore, mutations that could impact potential phage receptors on the host surface were selected by lytic phage exposure. This work demonstrates that the interaction of Salmonella phages (WHR and NHR) with SI for 12 h in vitro leads to emergence of new phenotypic and genotypic traits in both phage and host. This information is crucial for the rational design of phage-based control strategies.

Isolation and Characterization of Lytic Bacteriophages infecting Staphylococcus epidermidis

The Staphylococcus epidermidis isolates were obtained during a period extended between September 2014 and January 2015, depending on biochemical tests and VITEK 2 system. Several sewage water samples were assayed using a plaque assay of double agar overlay as a source of S. Epidermidis phages. The bacteriophages were described depending on plaques size and shapes. Phage 1 was the most predominant in the bacterial lawn and able to infect other S. species such as S. aureus. Therefore, it was decided to study the effect of temperature on its original titer. The results revealed a gradual decrease in the phage titer with increasing dilution number. Each temperature at several incubation periods, significantly vary depending on phage titer. The optimum temperature was 40 ° C, while the 80 ° C was represented the inhibitor temperature. L.S.D. at level (0.05) for interaction was 39.552. The pH 6.5 – 7.5 were represented the optimal pH for the best phage activity while the phage titer beginning to decline in above and below this range of optimal pH , L.S.D. at level 0.05 was 17.898. In conclusion, our study found that Phage1 was considered as predominant phage because of their ability to infect other Staphylococci species such as S. Aureus.

Use of Logistic Regression for Prediction of the Fate of Staphylococcus aureus in Pasteurized Milk in the Presence of Two Lytic Phages

ABSTRACT The use of bacteriophages provides an attractive approach to the fight against food-borne pathogenic bacteria, since they can be found in different environments and are unable to infect humans, both characteristics of which support their use as biocontrol agents. Two lytic bacteriophages, vB_SauS-phiIPLA35 (phiIPLA35) and vB_SauS-phiIPLA88 (phiIPLA88), previously isolated from the dairy environment inhibited the growth of Staphylococcus aureus. To facilitate the successful application of both bacteriophages as biocontrol agents, probabilistic models for predicting S. aureus inactivation by the phages in pasteurized milk were developed. A linear logistic regression procedure was used to describe the survival/death interface of S. aureus after 8 h of storage as a function of the initial phage titer (2 to 8 log10 PFU/ml), initial bacterial contamination (2 to 6 log10 CFU/ml), and temperature (15 to 37°C). Two successive models were built, with the first including only data from the experimental design and a global one in which results derived from the validation experiments were also included. The temperature, interaction temperature-initial level of bacterial contamination, and initial level of bacterial contamination-phage titer contributed significantly to the first model prediction. However, only the phage titer and temperature were significantly involved in the global model prediction. The predictions of both models were fail-safe and highly consistent with the observed S. aureus responses. Nevertheless, the global model, deduced from a higher number of experiments (with a higher degree of freedom), was dependent on a lower number of variables and had an apparent better fit. Therefore, it can be considered a convenient evolution of the first model. Besides, the global model provides the minimum phage concentration (about 2 × 108 PFU/ml) required to inactivate S. aureus in milk at different temperatures, irrespective of the bacterial contamination level.

Synthesis of Monomeric Acridine Derived Nucleic Acid Intercalators

A series of antiviral compounds consisting of an intercalating acridine derived part, a spacer region and a reactive EDTA-derived conjugate was synthesized in an easy sequence. In the presence of ascorbate a reduction of the phage-titer of MS2 phages by several logarithmic decades was achieved.

Synthesis of Dimeric Acridine Derived Nucleic Acid Intercalators

A series of antiviral compounds consisting of an intercalating acridine derived part, a spacer region and a reactive EDTA-derived conjugate was synthesized in an easy sequence. Suitably monoprotected 1,ω-alkyldiamines gave upon reaction with 6,9-dichloro-2-methoxyacridine (1) followed by deprotection and reaction with EDTA dianhydride the target molecules. In the presence of ascorbate a reduction of the phage-titer of the MS2 phages by > 8 logarithmic decades was achieved.

Effect of advanced oxidation processes on inactivation of coliphages

Chlorine and its derivatives are no longer regarded as acceptable disinfectants of water because of compounds they form with organic material in water. These compounds have been proved to be mutagenic and carcinogenic to man. Alternative disinfectants like UV radiation and ozonization are regarded as less harmful disinfectants of microorganisms in water. In the present study the effect of UV radiation alone and together with hydrogen peroxide, and ozone treatment on the inactivation of coliphages in tap water were studied. Two phage types, DNA- and RNA-phages were seeded into tap water and exposed to these disinfectants in batch experiments. The inactivation of phages was determined as a reduction of phage titer as a function of contact time. Disinfection with ozone proved to inactivate coliphages more rapidly than UV light or UV light together with hydrogen peroxide (H2O2). After two minutes exposure time the reduction in phage titer was from 6 to 8 log units with ozone while with UV light or UV with H2O2 the reduction was from three to four log units. According to these results ozonization seemed to be more efficient disinfectant than UV light radiation.