Applications of Personalised Phage Therapy highlighting the importance of Bacteriophage Banks against Emerging Antimicrobial Resistance

Emerging antibiotic resistance is one of the most important microbiological issues of the 21st century. This poses a query regarding the future use of antibiotics and availability of other promising therapeutic alternatives. The awareness about antibiotic misuse has improved insufficiently and is evident by the increased incidences of multidrug resistant infections globally. Amongst different antibacterial therapeutic approaches phage therapy has created a niche of its own due to continuous use for treatment of human infections in Eastern Europe. Synergistic compounds along with phages have also been proposed as a better alternative compared to antibiotics or phage alone for treatment of chronic cases and seriously debilitating diseases. As such, why not allow custom made phage therapy for treatment of chronic infections? However, the success of phage therapy will depend upon instant availability of characterised bacteriophages from bacteriophage banks which may serve as the major catalyst in bringing Phage Therapy to main stream treatment alternatives or in combination therapy at least. In the current article we present a glimpse of comprehensive approach about utility of bacteriophage banks and further present personalised phage therapy in a synergistic role with antibiotics to overcome emerging antimicrobial resistance

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Not AvailableBacteriophages play an important role in bacterial control in natural niche however a little is known about Bacillus sp. phages prevailing in cadaver affected soils. In the current study, the Bacillus sp. phage was isolated from the equine cadaver disposal site and characterised to gain an insight into the issue of role of phages in biological dynamics of manure thus formed over years. Firstly, the host bacterium was isolated and identified as Bacillus cereus group member as assessed by phylogenetic analysis and secondly it’s corresponding phage from same soil sample was also enriched and characterised. The phage (VTCCBPA38) was found to belong to family Myoviridae and was active within the temperature range of 4 °C - 45 °C. As assessed by biological sensitivity by spot test, the phage was active against 6/19 (31.6 %) Bacilli tested including Bacillus cereus from goat mastitis. Thus the phage may find potential use in biocontrol of diseases caused by Bacillus sp. Furthermore, this report is valuable as the first study for investigation of Bacillus sp. phage in carcass burial sites.Not Availabl

Indian Veterinary Research Institute,

Not AvailableA bacteriophage (VTCCBPA6) against a pathogenic strain of Aeromonas hydrophila was isolated from the sewage of an organized equine breeding farm. On the basis of TEM analysis, phage belonged to family Myoviridae. PCR amplification and sequence analysis of gp23 gene (encoding for major capsid protein) revealed phylogenetic resemblance to T4 like virus genus. Protein profiling by SDS-PAGE also indicated its resemblance to T4 like phage group. However, the comparison of its gp23 gene sequence with previously reported phages showed similarity with T4-like phages infecting Enterobacteriaceae instead of Aeromonas spp. Thus, to our knowledge, this report points toward the fact that a novel/evolved phage might exist in equine environment against A. hydrophila, which can be potentially used as a biocontrol agent.ICA

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Not AvailableKlebsiella pneumoniae is an important emerging pathogen of humans and animals which may lead to major clinical implications including mortality. Moreover the increased use of antibiotics has promoted emergence of carbapenem resistant strains and extended spectrum β-lactamase producers (ESBLs) of K. pneumoniae. Recently, phage therapy has gained momentum as a conceivable alternative against emerging antibiotic resistance. OBJECTIVE: With the aim to explore the efficacy of phage therapy against virulent K. pneumoniae infection, the current study was undertaken to assess the therapeutic effects of a novel lytic phage-VTCCBPA43 in pneumonic mouse model. METHODS: The tailed phage - VTCCBPA43 was assessed for it's growth kinetics, in vitro host range analysis, temperature and pH sensitivity. The protein constituents were analysed by SDS-PAGE and Lc MS/MS and the therapeutic efficacy was observed 2 hr post challenge with virulent K. pneumoniae in BALB/c mouse model. RESULTS: The phage-VTCCBPA43 was found to exhibit high temperature (upto 80 °C) tolerant property. It was most active at pH 5, had a burst size of 172 PFU/ml and exhibited a narrow host range. It was identified as KP36 like phage by shotgun proteomics. Following intranasal application of a single dose of 2 × 109 PFU/mouse post challenge, presence of biologically active phage in vivo and a significant reduction of bacterial load in lungs at all time points was observed which was even more impressive at 96hpi, 6dpi and 10dpi. The loss of severity of lesions suggested overall beneficial effects of phage therapy using BPA43 in the pneumonic mouse model. CONCLUSION: The current research represents first in vivo evidences for effective phage therapy against K. pneumoniae infection by using intranasal route.Not Availabl

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Not AvailablePhage therapy has been previously tried for treatment of diarrhoea in calves, pigs and lambs but those trials were conducted without any detailed information of used phages. Here, we report isolation of a broad-spectrum phage which showed bactericidal activity against 47.3 % of calf diarrhoeal isolates of Escherichia coli, in vitro. The isolated phage resembled the characteristics of Myoviridae family and showed *97 % similarity with earlier reported bacteriophages of sub family-Tevenvirinae, genus-T4-like virus, based on nucleotide sequence of major head protein— gp23 gene. The phage exhibits the potential to be used as drug substitute tool against E. coli causing diarrhoea in cattle in farm environments.ICA

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Not AvailableThe ecosystem is continuously exposed to a wide variety of antimicrobials through waste effluents, agricultural run - offs and animal - related and anthropogenic activities, which contribute to the spread of antibiotic resistance genes (ARGs). The contamination of ecosystems with ARGs may create increased opportunities for their transfer to naive microbes and eventually lead to entry into the human food chain. Transduction is a significant mechanism of horizontal gene transfer in natural environments, which has traditionally been underestimated as compared to transformation. We explored the presence of ARGs in environmental bacteriophages in order to recognize their contribution in the spread of ARGs in environmental settings. Bacteriophages were isolated against environmental bacterial isolates, purified and bulk cultured. They were characterized, and detection of ARG and intI genes including blaTEM, blaOXA - 2, intI1, intI2, intI3, tetA and tetW was carried out by PCR. This study revealed the presence of various genes [tetA (12.7 %), intI1 (10.9 %), intI2 (10.9 %), intI3 (9.1 %), tetW (9.1 %) and blaOXA - 2 (3.6 %)] and blaTEM in a significantly higher proportion (30.9 %). blaSHV, blaOXA - 1, tetO, tetB, tetG, tetM and tetS were not detected in any of the phages. Soil phages were the most versatile in terms of ARG carriage. Also, the relative abundance of tetA differed significantly vis - a - vis source. The phages from organized farms showed varied ARGs as compared to the unorganized sector, although blaTEM ARG incidences did not differ significantly. The study reflects on the role of phages in dissemination of ARGs in environmental reservoirs, which may provide an early warning system for future clinically relevant resistance mechanisms.ICA

<span style="font-size:11.0pt;font-family: "Times New Roman";mso-fareast-font-family:"Times New Roman";mso-bidi-font-family: Mangal;mso-ansi-language:EN-GB;mso-fareast-language:EN-US;mso-bidi-language: HI" lang="EN-GB">Molecular characterization of virulence-associated protein (Vap) family genes of pathogenic <i style="mso-bidi-font-style:normal">Rhodococcus equi</i> isolates from clinical cases of Indian equines</span>

195-202<span style="font-size:9.0pt;mso-ansi-language:EN-IN;mso-bidi-font-weight: bold">Virulence-associated plasmid (Vap) genes of clinical isolates of Rhodococcus equi were characterized. Isolates were identified by 16S rRNA, choE and <i style="mso-bidi-font-style: normal">traA gene PCR, followed by amplification, cloning and sequencing of 7 Vap<span style="mso-bidi-font-weight: bold"> genes. <span style="font-size:9.0pt;mso-ansi-language:EN-IN;mso-bidi-font-weight: bold">The isolates were found positive for <i style="mso-bidi-font-style: normal">vapA gene family. The comparative sequence analysis of vap genes revealed 99-100% similarity at both nt and aa levels with all sequences. The aa sequences of the predicted Vap proteins<span style="font-size: 9.0pt;mso-ansi-language:EN-IN;mso-bidi-font-weight:bold"> exhibited a high degree of similarity to each other, especially at the carboxy terminal ends. Invariable point mutations were observed in Vap proteins<span style="font-size:9.0pt;mso-bidi-font-weight: bold" lang="EN-GB">.<span style="font-size:9.0pt;mso-ansi-language:EN-IN;mso-bidi-font-weight: bold"> The changes did not cause alteration in hydropathicity and secondary structures in VapA & H proteins. Few major changes in polarity and structures were observed in VapD, E and G proteins. Phylogenetic analysis revealed that all <i style="mso-bidi-font-style: normal">Vap genes followed similar branching pattern except isolate from Bahadurgarh (BBG163). <span style="font-size:9.0pt;mso-ansi-language: EN-IN;mso-bidi-font-weight:bold">We conclude that VapA is highly conserved and plays a major role in pathogenesis<span style="font-size:9.0pt;mso-ansi-language: EN-IN;mso-bidi-font-weight:bold">. This is the first report of sequence analysis and phylogenetic studies of Vap gene family of clinical virulent isolates of <i style="mso-bidi-font-style: normal">R. equi from India. </span