Broadening the host range of lytic phage against Methicillin-resistant Staphylococcus aureus
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Date
2018
Authors
Ní Mhaoldomhnaigh, Eimear Sinead
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Publisher
University College Cork
Published Version
Abstract
Introduction
In recent years the rise in antibiotic resistance in pathogenic bacteria has meant that significant research has gone into finding alternative treatments. The higher mortality and morbidity associated with multidrug resistant bacteria suggest that solutions are urgently needed. The World Health Organisation (WHO) recently published a list of 12 bacteria which are considered the greatest threat in this regard (a list which includes drug resistant Staphylococcus aureus). Unless a viable alternative to antibiotics is found global human health will suffer and we may see a return to mortality and morbidity rates similar to those seen in the pre-antibiotic era. Despite the development of vaccines, and improved hygiene and living standards, bacterial infections remain a very real treat. In particular the developing world is most at risk of unchecked antibiotic resistance disease due to insufficient access to health care, overcrowding and decreased water sanitation.
Objectives
Bacteriophage targeting of specific pathogens may provide an alternative to antibiotic therapy in certain clinical settings. However specific issues may limit their use. The high specificity of phage for their host makes them potentially impractical for therapeutic use, without first identifying the causative agent/strain and then having access to a phage that infects it. The current study therefore investigated the factors that affect phage specificity for S. aureus isolates which vary at the strain level. The study also investigated whether bacteriophage populations can evolve to overcome bacterial defence mechanisms and whether it is possible to use this ability to increase or shift host ranges. We hypothesize that p cocktails of phage targeting different hosts may be the solution to overcoming narrow host ranges of individual phage. In numerous previous studies phage cocktails have been shown to have success in treating infections (O'Flynn et al., 2004; Gu et al., 2012; Chan & Abedon, 2013; Örmälä & Jalasvuori, 2013; Niu et al., 2014; Chadha et al., 2016;). Methods
In this study the host range of the phage was characterized by plaque assay, the anti-biotic resistance profile of the bacterial sample set examined by agar disc diffusion method and a method was used whereby MRSA phage were co-cultured with susceptible and un-susceptible strains of MRSA of clinical importance. Following the generation of mutants in Phage B1 and phage K with favourable adaptations to host range, genetic sequencing was carried out and hypothesis proposed as to what changes lead to the altered host range.
Results
It was found that after prolonged exposure phage evolved to be able to infect formerly non-susceptible strains. We analysed the host range of a novel staphylococcal phage, Phage B1 in comparison with reference phage commonly used in the literature. The study established key parameters such as the exposure time required and the ratio of target strains: permissive strains that provided for optimal phage adaptation. Some of the mutants had the ability to infect previously insensitive strains and others showed increased efficacy. In practical therapeutic terms, this would mean that it is possible to rapidly select adapted phage isolates to target previously non-permissive pathogenic bacterial strains. Sequencing of the original phage, phage B1 as well as the mutated/adapted phage derivatives, phage B1 0.0066 and phage B1 3488 revealed insights into genomic variation of phage during adaptation to previously non-permissive hosts. Acquisition of specific genes (including genes encoding alternative phage tail proteins) could explain altered host infection potential in the mutated/adapted phage, although further work is necessary to associate altered genes with functional properties of the phage.
Conclusion
The current study established the host range of a novel staphylococcal phage, phage B1 which has potential to be used in phage therapy. Whilst some S. aureus isolates were resistant to targeting by phage B1 we established conditions under which we could rapidly select for phage variants to target such strains. Genomic analysis of both phage B1 and phage variants (phage B1 0.0066 and phage B1 3488) provided molecular insights into the process of phage adaptation. Overall, the results suggest the potential for phage adaptation and rapid isolation of new phage variants in the clinical setting.
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Keywords
Bacteriophage , Antibiotic resistance , MRSA , Methicillin-resistant Staphylococcus aureus , Phage , Phage cocktail , Pathogens , Phage B1 , Phage K , Co-culturing
Citation
Ní Mhaoldomhnaigh, E. S. 2018. Broadening the host range of lytic phage against Methicillin-resistant Staphylococcus aureus. MSc Thesis, University College Cork.