Presentation

1. ASSESSMENT OF THE LYTIC PROPERTY OF BACTERIOPHAGES
AGAINST SOME NOSOCOMIAL MULTIDRUG RESISTANT
BACTERIAL ISOLATES
MSc. RESEARCH PROPOSAL
By
ALABI, Emmanuel Dayo
(PG8/SCI/2020/0887)
Department of Microbiology, Federal University Dutsin-Ma,
Katsina State, Nigeria
Supervisors:
DR. I. MZUNGU
DR. A.T. ADESOJI

2. OUTLINES
 Introduction
 Statement of Research problem
 Research question(s)
 Justification
 Aim and Objectives
 Materials and methods
 Statistical Analysis
 Expected Contribution to Knowledge
 References
Alabi, E. D.
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3. INTRODUCTION
 Bacteriophages are simple, yet incredibly diverse, non-living biological entities
consisting of DNA or RNA enclosed within a protein capsid (Lin et.al., 2017).
 They are classified into two: Temperate phages and Virulent phages (Willey et
al., 2008; Ryan et al., 2011).
 They replicate by injecting their genetic material into the host cell, which either
integrate into the bacterial genome (Temperate phage) or usurp the bacterial
replication machinery to produce the next generation of phage progeny and lyse
the cell (Lytic phage).
 They are the most common biological entities in nature, and have been shown to
effectively fight and destroy multi-drug resistant bacteria(Lin et.al. 2017).
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4. INTRODUCTION CON’TD
 Health care-related infections (HCAIs), previously referred to as
Nosocomial infections (NIs), are infections acquired by patients on
admission in health care settings (Haque et al., 2018).
 Nosocomial infections (NIs) are reported to be one of the 10 most important
causes of death in the United States. The US Center for Disease Control and
Prevention reported that approximately 1.7 million hospitalized patients
annually acquire HCAIs or NIs while being treated due to other health
problems, and that more than 98,000 of these patients (approximately 1 in
17) die due to Nosocomial infections.
 The British Government in May 2016 predicted that by 2050,
approximately 10 million people would die every year globally as a result of
antimicrobial resistance, than currently die from cancer ( O’Neill, 2016).
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5. INTRODUCTION CON’TD
 Almost a decade before the discovery of penicillin, the controversial practice
of phage therapy was being developed as a treatment for bacterial infections
such as bacillary dysentery (Chanishvili, 2012).
 In fact, the nature of their existence was a topic of contention until they were
visualized in the 1940’s after the invention of electron microscopy
(Ackermann, 2011).
 A number of logistical and technical obstacles in developing phage therapy
led to its widespread abandonment after the discovery of antibiotics (Lin
et.al, 2017).
 The universal decline in the effectiveness of antibiotics and alarming public
health crisis that antimicrobial resistance (AMR) pose has generated
renewed interest in revisiting this practice (Lin et. al., 2017).
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6. STATEMENT OF RESEARCH PROBLEM
 Clinical conditions caused by Staphylococcus aureus and
Pseudomonas aeruginosa are main causes of concern in the
health care setting.
 Diminished ability of antibiotics alone to control infectious
disease due to increasing resistance in pathogens and
decreasing number of new antibiotics approved for use.
 The appearance of multidrug-resistant (MDR), extensively
drug-resistant (XDR) and Pan drug-resistant (PDR)
bacteria (Majid et.al, 2020).
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7. RESEARCH QUESTIONS
 Can antibiotics alone be able to control infectious diseases
due to increasing resistance in pathogens and decreasing
number of new antibiotics approved for use?
 Will cocktail of Bacteriophages be effective in the control of
multidrug resistant bacterial isolates?
 Will synergistic effect of combined antibiotic therapy and
Bacteriophage activities against multidrug resistant isolates
be more effective than cocktail of Bacteriophage therapy
alone?
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8. JUSTIFICATION
 Bacteriophages are very specific, targeting only one or a few strains of
bacteria. Traditional antibiotics have more broad-spectrum effect,
killing both harmful and useful bacteria (Cerveny, 2002).
 The species and strain specificity of bacteriophages makes it unlikely
that harmless or useful bacteria will be killed when fighting an
infection (Cerveny et al., 2002).
 Encouraging reports of successful phage treatment indicates high
efficacy. There is renewed interest in revisiting the use of
bacteriophage to treat bacterial infections in the developed countries
(Majid et al., 2020).
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9. JUSTIFICATION CONT’D
 The need to locally explore and exploit the activities
and properties of bacteriophages to improve clinical
and public health which is threatened daily by
pathogenic microorganisms.
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10. AIM
 To evaluate the lytic property of bacteriophages against
some multidrug resistant Bacteria of nosocomial origin
as a biological control /treatment for antibiotic resistance
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11. OBJECTIVES
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 To isolate Bacteria from clinical samples of patients that overstayed
in the hospital due to complications such as; Infected surgical
incision, patients with prolonged indwelling catheters, Skin Burns
and infected trauma wounds.
 To identify bacterial isolates by conventional methods (Gram stain
and Biochemical tests) and Screen pure bacterial isolates using
Antibiotic susceptibility Disc diffusion Method.
 To confirm the identity and antibiogram of multi-drug resistant
bacterial isolates using Vitek Technology.
 To characterize multi-drug resistant genes (whole genome sequence
analysis).

12. OBJECTIVES CONT’D
 Isolate and Concentrate Bacteriophage from waste water
(sewage), Dam water and wet soil and examine their lytic
activity against multidrug resistant Bacterial isolates of
nosocomial origin.
 To evaluate combined synergistic effect of antibiotic therapy
and lytic activities of Bacteriophages as well determine the
combined activities of Bacteriophage cocktail against the
multidrug resistant isolates.
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Alabi, E. D.

13. MATERIALS AND METHODS
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Flow chart for Bacterial Isolation and Identification
Isolation of pure cultures
Screening for MDR Strains using Antibiotic
Disk diffusion method
Identification of isolates using biochemical
method
Carryout Molecular characterization / identification of MDR bacterial isolates
Collection of clinical samples and isolation
Of bacterial pathogens

14. MATERIALS AND METHODS CONT’D
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- Flow Chart for phage isolation and lytic activity determination
Collection of waste water and soil samples
Phage isolation by centrifugation
and filtration of water and soil
Phage enrichment by incubation with bacterial isolate in double
strength media and purification through centrifugation and
filtration
Phage identification by spot assay
and agar plaque assay
Determination of lytic property
using tube dilution assay
Determination of lytic activity/
infectivity using agar plaque assay
Determination of lytic property
using lysis in tube
Statistical analysis comparing phage, antibiotics
and phage combined with standard antibiotic using chi-square test < 0.5 level of confidence

15. Alabi, E. D.
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REFERENCES
Ackermann, H.W. (2011). The first phage electron micrographs. Bacteriophage 2011; 1: 225-
227 [PMID: 23050215 http://doi.org/ 10.4161/bact.1.4.17280].
Bala, D. T. (2016). Determining The Lytic Property of Bacteriophage on Salmonella species.
Zaria: ABU PRESS.
Cerveny, K. E.; Depaola, A.; Duckworth, D. H.; Gulig, P. A.(2002) Phage therapy of local
and systemic disease caused by Vibrio vulnificus in iron-dextran-treated mice.
Infect. Immun., 2002, 70 (11), 6251-6262.
Chanishvili, N. (2012). Phage therapy--history from Twort and d’Herelle through Soviet
experience to current approaches. Adv Virus Res 2012; 83: 3-40 [PMID:
22748807. http://doi.org/ 10.1016/B978-0-12-394438-2.00001-3].
.

16. REFERENCES CONT’D
Haque, M., Sartelli, M., McKimm, J. and Bakar, M. A. (2018). Health care-associated
infections–an overview. Infectious Drug Resistance. 2018; 11; 2321–33
Lin, D.M., Koskella, B. and Lin, H.C. (2017). Phage therapy: An alternative to antibiotics
in the age of multi-drug resistance. World Journal of Gastrointestinal
Pharmacology Therapeutics,(3);162-173. http://doi.org/10.4292/wjgpt.v8.i3.162
Majid, T.M., Nour, A., Nour, S., Masoumeh, H., Shiva, M., Amin, K. and Faramarz, M.J.
(2020). How Phages Overcome the Challenges of Drug Resistant Bacteria in
Clinical Infections. Journal of Infection and Drug Resistance, 13, 45-61.
http://doi.org/10.2147/IDR.S234353
O’Neil, J. (2016). The Review on Antimicrobial Resistance. London, UK: Internet.
Retrieved from: https://amr-review.org.
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17. REFERENCES CONT’D
Ryan, E.M., Gorman, S.P., Donnelly, R.F. and Gilmore, B.F. (2011). Recent
Advances in Bacteriophage Therapy; how delivery routes, Formulation,
concentration and timing influence the success of phage therapy.
Journal of Pharmacy and Pharmacology. 63; 1253-1264
Willey, J.M., Sherwood, L.M. and Woolverton C.J. (2008). Prescott, Harvey and
Klein's Microbiology (7th Ed.). New York, NY 10020: McGraw-Hill
companies Inc. pp. 997-1100
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18. THANKS
THANK U 4LISTENING
Alabi, E. D.