Multidrug-resistant Klebsiella pneumoniae is a nosocomial pathogen, produces septicemia, pneumonia and UTI. Excessive use of antibiotics contributes towards emergence of multidrug-resistance. Bacteriophage-therapy is a potential substitute of antibiotics with many advantages. In this investigation, microbiological and genome characterization of TSK1 bacteriophage and its biofilm elimination capability are presented. TSK1 showed narrow host range and highest stability at pH 7 and 37 °C. TSK1 reduced the growth of K. pneumoniae during the initial 14 hours of infection. Post-treatment with TSK1 against different age K. pneumoniae biofilms reduced 85–100% biomass. Pre-treatment of TSK1 bacteriophage against the biofilm of Klebsiella pneumoniae reduced > 99% biomass in initial 24 hr of incubation. The genome of TSK1 phage comprised 49,836 base pairs with GC composition of 50.44%. Total seventy-five open reading frames (ORFs) were predicted, 25 showed homology with known functional proteins, while 50 were called hypothetical, as no homologs with proved function exists in the genome databases. Blast and phylogenetic analysis put it in the Kp36 virus genus of family Siphoviridae. Proposed packaging strategy of TSK1 bacteriophage genome is headful packaging using the pac sites. The potential of TSK1 bacteriophage could be used to reduce the bacterial load and biofilm in clinical and non-clinical settings.

Methicillin-resistant Staphylococcus aureus (MRSA) is a prevailing nosocomial pathogen that causes a large number of diseases in healthcare and community settings. The MRSA causes infections in different tissues of immunocompromised individuals leading to increased morbidity and mortality. It possess various virulence mechanisms to show resistance against to a lot of beta-lactam antibiotics. To tackle this emerging issue of MRSA, there is an urgent need of antibiotic alternatives and utilizing lytic bacteriophages is one of the best promising therapeutic approach. In the present study, a lytic bacteriophage TSP was isolated from hospital wastewater against MRSA. Its morphology, physiology, host specificity, burst size and lytic spectrum were determined and complete genome sequence was analyzed. TSP phage efficiently inhibit bacterial growth for up to 12 hours. TSP phage showed broad lytic spectrum against clinical isolates of MRSA (78%) and MSSA (37%). It showed stability at varying temperatures (25ºC, 37ºC) and pH (5–9), while its maximum storage stability was observed at 4ºC. It had short latent period (20min) and high burst size (103 PFU/ infected cell). TSP genome sequence and restriction analysis revealed that its genome is linear having 17,987 bp in length with an average GC content of 29.7%. The TSP genome showed 98% similarity to S aureus phages SCH1, SCH11 and vB SauP-436A1. According to comparative genomic analysis and phylogenetic tree analysis, TSP phage can be considered as a member of genus “P68viruses”. The strong lytic activity, broad host range and short latent period along with absence of any lysogenic and toxic genes make TSP a very good candidate for phage therapy against MRSA infections if prove safe during in vivo studies.

Methicillin-resistant Staphylococcus aureus (MRSA) is a prevailing nosocomial pathogen that is increasingly isolated in community settings. It shows resistance against all beta-lactam drugs and has acquired mechanisms to resist other groups of antibiotics. To tackle this emerging issue of MRSA, there is an urgent need for antibiotic alternatives, and utilizing lytic bacteriophages is one of the most promising therapeutic approaches. In the present study, a lytic bacteriophage TSP was isolated from hospital wastewater against MRSA. The phage efficiently inhibited bacterial growth for up to 12 h at MOI of 1 and 10. TSP phage showed activity against various isolates of MRSA and MSSA, isolated from different clinical samples, with variable antibiotic susceptibility patterns. The bacteriophage TSP showed stability at varying temperatures (25 °C, 37 °C) and pH values (5–9), while its maximum storage stability was observed at 4 °C. It had a short latent period (20 min) and burst size of 103 ± 5pfu/infected cells. TSP genome sequence and restriction analysis revealed that its genome has a linear confirmation and length of 17,987 bp with an average GC content of 29.7%. According to comparative genomic analysis and phylogenetic tree,TSP phage can be considered a member of genus “P68viruses”. The strong lytic activity and short latent period in addition to its lytic nature makes it a good candidate for phage therapy against MRSA infections, if it proves to be effective in in-vivo studies.