Isolation, characterization, and antibacterial activity of lytic bacteriophage against methicillin-resistant Staphylococcus aureus causing bedsore and diabetic wounds
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Abstract
Background and Objectives: Phage therapy has gained interest as an alternative treatment for methicillin-resistant Staphylococcus aureus (MRSA) infections. The purpose of this study was to isolate and characterize an effective bacteriophage against isolates of MRSA.
Materials and Methods: Bacteriophage was isolated from hospital sewage. Lytic activity and the titers of phage lysates were measured using spot test and double-layer plaque assay. The phage characterization was determined through transmission electron microscopy. Adsorption rate, host range and stability tests were investigated. The latent period and burst size were estimated from a one-step growth curve. The effect of bacteriophage against MRSA biofilms was determined and Real-time PCR was used to assess the effects of the bacteriophage on the expression of the biofilm-associated genes.
Results: TEM results showed that the phage resembled the Cystoviridae family. Its latent period was 30 min, corresponding to about 71/43 phage particles per infected cell. The phage had a broad host range and it was most stable at 37°C and pH 7. It was sensitive to NaCl concentrations. The expressions of the biofilm-associated genes were significantly reduced in the presence of the phage.
Conclusion: The isolated phage was effective against MRSA strains and it can be an optional strategy of controlling biofilm development.
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8. Wang Z, Zheng P, Ji W, Fu Q, Wang H, Yan Y, et al. SLPW: A Virulent Bacteriophage Targeting Methicillin-Resistant Staphylococcus aureus In vitro and In vivo. Front Microbiol 2016; 7: 934.
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10. Van Twest R, Kropinski AM. Bacteriophage enrichment from water and soil. Methods Mol Biol 2009; 501: 15-21.
11. Sillankorva S. Isolation of bacteriophages for clinically relevant bacteria. Methods Mol Biol 2018; 1693: 23-30.
12. Ji J, Liu Q, Wang R, Luo T, Guo X, Xu M, et al. Identification of a novel phage targeting methicillin-resistant Staphylococcus aureus In vitro and In vivo. Microb Pathog 2020; 149: 104317.
13. Ackermann H-W. Basic phage electron microscopy. Methods Mol Biol 2009; 501: 113-126.
14. Abedon ST. Phage therapy dosing: The problem(s) with multiplicity of infection (MOI). Bacteriophage 2016; 6(3): e1220348.
15. Drulis-Kawa Z, Mackiewicz P, Kęsik-Szeloch A, Maciaszczyk-Dziubinska E, Weber-Dąbrowska B, Dorotkiewicz-Jach A, et al. Isolation and characterisation of KP34--a novel φKMV-like bacteriophage for Klebsiella pneumoniae. Appl Microbiol Biotechnol 2011; 90: 1333-1345.
16. Nabergoj D, Modic P, Podgornik A. Effect of bacterial growth rate on bacteriophage population growth rate. Microbiologyopen 2018; 7(2): e00558.
17. Zafari M, Adibi M, Chiani M, Bolourchi N, Barzi SM, Shams Nosrati MS, et al. Effects of cefazolin-containing niosome nanoparticles against methicillin-resistant Staphylococcus aureus biofilm formed on chronic wounds. Biomed Mater 2021; 16: 035001.
18. Seaman PF, Day MJ. Isolation and characterization of a bacteriophage with an unusually large genome from the Great Salt Plains National Wildlife Refuge, Oklahoma, USA. FEMS Microbiol Ecol 2007; 60: 1-13.
19. Dvořáčková M, Růžička F, Benešík M, Pantůček R, Dvořáková-Heroldová M. Antimicrobial effect of commercial phage preparation Stafal® on biofilm and planktonic forms of methicillin-resistant Staphylococcus aureus. Folia Microbiol (Praha) 2019; 64: 121-126.
20. Cha Y, Son B, Ryu S. Effective removal of staphylococcal biofilms on various food contact surfaces by Staphylococcus aureus phage endolysin LysCSA13. Food Microbiol 2019; 84: 103245.
21. Costa GA, Rossatto FCP, Medeiros AW, Correa APF, Brandelli A, Frazzon APG, et al. Evaluation antibacterial and antibiofilm activity of the antimicrobial peptide P34 against Staphylococcus aureus and Enterococcus faecalis. An Acad Bras Cienc 2018; 90: 73-84.
22. Benamara H, Rihouey C, Abbes I, Ben Mlouka MA, Hardouin J, Jouenne T, et al. Characterization of membrane lipidome changes in Pseudomonas aeruginosa during biofilm growth on glass wool. PLoS One 2014; 9(9): e108478.
23. Oosthuizen MC, Steyn B, Lindsay D, Brözel VS, Von Holy A. Novel method for the proteomic investigation of a dairy-associated Bacillus cereus biofilm. FEMS Microbiol Lett 2001; 194: 47-51.
24. Atshan SS, Shamsudin MN, Karunanidhi A, Van Belkum A, Lung LT, Sekawi Z, et al. Quantitative PCR analysis of genes expressed during biofilm development of methicillin resistant Staphylococcus aureus (MRSA). Infect Genet Evol 2013; 18: 106-112.
25. Rahimzadeh G, Saeedi M, Moosazadeh M, Hashemi SMH, Babaei A, Rezai MS, et al. Encapsulation of bacteriophage cocktail into chitosan for the treatment of bacterial diarrhea. Sci Rep 2021; 11: 15603.
26. Rahimzadeh G, Saeedi M, Nokhodchi A, Moosazadeh M, Ghasemi M, Rostamkalaei SS, et al. Evaluation of in-situ gel-forming eye drop containing bacteriophage against Pseudomonas aeruginosa keratoconjunctivitis in vivo. Bioimpacts 2021; 11: 281-287.
27. Ackermann HW. Phage classification and characterization. Methods Mol Biol 2009; 501: 127-140.
28. Peng C, Hanawa T, Azam AH, LeBlanc C, Ung P, Matsuda T, et al. Silviavirus phage ɸMR003 displays a broad host range against methicillin-resistant Staphylococcus aureus of human origin. Appl Microbiol Biotechnol 2019; 103: 7751-7765.
29. Cui Z, Feng T, Gu F, Li Q, Dong K, Zhang Y, et al. Characterization and complete genome of the virulent Myoviridae phage JD007 active against a variety of Staphylococcus aureus isolates from different hospitals in Shanghai, China. Virol J 2017; 14: 26.
30. Fierheller M, Sibbald RG. A clinical investigation into the relationship between increased periwound skin temperature and local wound infection in patients with chronic leg ulcers. Adv Skin Wound Care 2010; 23: 369- 379.
31. Jones EM, Cochrane CA, Percival SL. The effect of pH on the extracellular matrix and biofilms. Adv Wound Care (New Rochelle) 2015; 4: 431-439.
32. Yüksel EB, Yıldırım AM, Bal A, Kuloglu T. The effect of different topical agents (silver sulfadiazine, povidone-iodine, and sodium chloride 0.9%) on burn injuries in rats. Plast Surg Int 2014; 2014: 907082.
2. Borysowski J, Lobocka M, Międzybrodzki R, Weber-Dabrowska B, Górski A. Potential of bacteriophages and their lysins in the treatment of MRSA: current status and future perspectives. BioDrugs 2011; 25: 347-355.
3. Noori Goodarzi N, Bolourchi N, Fereshteh S, Soltani Shirazi A, Pourmand MR, Badmasti F. Investigation of novel putative immunogenic targets against Staphylococcus aureus using a reverse vaccinology strategy. Infect Genet Evol 2021; 96: 105149.
4. Turner NA, Sharma-Kuinkel BK, Maskarinec SA, Eichenberger EM, Shah PP, Carugati M, et al. Methicillin-resistant Staphylococcus aureus: an overview of basic and clinical research. Nat Rev Microbiol 2019; 17: 203-218.
5. Ganesh PS, Veena K, Senthil R, Iswamy K, Ponmalar EM, Mariappan V, et al. Biofilm-associated Agr and Sar Quorum sensing systems of Staphylococcus aureus are inhibited by 3-Hydroxybenzoic acid derived from Illicium verum. ACS Omega 2022; 7: 14653-14665.
6. Abedon ST, Kuhl SJ, Blasdel BG, Kutter EM. Phage treatment of human infections. Bacteriophage 2011; 1: 66-85.
7. Loc-Carrillo C, Abedon ST. Pros and cons of phage therapy. Bacteriophage 2011; 1: 111-114.
8. Wang Z, Zheng P, Ji W, Fu Q, Wang H, Yan Y, et al. SLPW: A Virulent Bacteriophage Targeting Methicillin-Resistant Staphylococcus aureus In vitro and In vivo. Front Microbiol 2016; 7: 934.
9. Nasser A, Azizian R, Tabasi M, Khezerloo JK, Heravi FS, Kalani MT, et al. Specification of bacteriophage isolated against clinical methicillin-resistant Staphylococcus aureus. Osong Public Health Res Perspect 2019; 10: 20-24.
10. Van Twest R, Kropinski AM. Bacteriophage enrichment from water and soil. Methods Mol Biol 2009; 501: 15-21.
11. Sillankorva S. Isolation of bacteriophages for clinically relevant bacteria. Methods Mol Biol 2018; 1693: 23-30.
12. Ji J, Liu Q, Wang R, Luo T, Guo X, Xu M, et al. Identification of a novel phage targeting methicillin-resistant Staphylococcus aureus In vitro and In vivo. Microb Pathog 2020; 149: 104317.
13. Ackermann H-W. Basic phage electron microscopy. Methods Mol Biol 2009; 501: 113-126.
14. Abedon ST. Phage therapy dosing: The problem(s) with multiplicity of infection (MOI). Bacteriophage 2016; 6(3): e1220348.
15. Drulis-Kawa Z, Mackiewicz P, Kęsik-Szeloch A, Maciaszczyk-Dziubinska E, Weber-Dąbrowska B, Dorotkiewicz-Jach A, et al. Isolation and characterisation of KP34--a novel φKMV-like bacteriophage for Klebsiella pneumoniae. Appl Microbiol Biotechnol 2011; 90: 1333-1345.
16. Nabergoj D, Modic P, Podgornik A. Effect of bacterial growth rate on bacteriophage population growth rate. Microbiologyopen 2018; 7(2): e00558.
17. Zafari M, Adibi M, Chiani M, Bolourchi N, Barzi SM, Shams Nosrati MS, et al. Effects of cefazolin-containing niosome nanoparticles against methicillin-resistant Staphylococcus aureus biofilm formed on chronic wounds. Biomed Mater 2021; 16: 035001.
18. Seaman PF, Day MJ. Isolation and characterization of a bacteriophage with an unusually large genome from the Great Salt Plains National Wildlife Refuge, Oklahoma, USA. FEMS Microbiol Ecol 2007; 60: 1-13.
19. Dvořáčková M, Růžička F, Benešík M, Pantůček R, Dvořáková-Heroldová M. Antimicrobial effect of commercial phage preparation Stafal® on biofilm and planktonic forms of methicillin-resistant Staphylococcus aureus. Folia Microbiol (Praha) 2019; 64: 121-126.
20. Cha Y, Son B, Ryu S. Effective removal of staphylococcal biofilms on various food contact surfaces by Staphylococcus aureus phage endolysin LysCSA13. Food Microbiol 2019; 84: 103245.
21. Costa GA, Rossatto FCP, Medeiros AW, Correa APF, Brandelli A, Frazzon APG, et al. Evaluation antibacterial and antibiofilm activity of the antimicrobial peptide P34 against Staphylococcus aureus and Enterococcus faecalis. An Acad Bras Cienc 2018; 90: 73-84.
22. Benamara H, Rihouey C, Abbes I, Ben Mlouka MA, Hardouin J, Jouenne T, et al. Characterization of membrane lipidome changes in Pseudomonas aeruginosa during biofilm growth on glass wool. PLoS One 2014; 9(9): e108478.
23. Oosthuizen MC, Steyn B, Lindsay D, Brözel VS, Von Holy A. Novel method for the proteomic investigation of a dairy-associated Bacillus cereus biofilm. FEMS Microbiol Lett 2001; 194: 47-51.
24. Atshan SS, Shamsudin MN, Karunanidhi A, Van Belkum A, Lung LT, Sekawi Z, et al. Quantitative PCR analysis of genes expressed during biofilm development of methicillin resistant Staphylococcus aureus (MRSA). Infect Genet Evol 2013; 18: 106-112.
25. Rahimzadeh G, Saeedi M, Moosazadeh M, Hashemi SMH, Babaei A, Rezai MS, et al. Encapsulation of bacteriophage cocktail into chitosan for the treatment of bacterial diarrhea. Sci Rep 2021; 11: 15603.
26. Rahimzadeh G, Saeedi M, Nokhodchi A, Moosazadeh M, Ghasemi M, Rostamkalaei SS, et al. Evaluation of in-situ gel-forming eye drop containing bacteriophage against Pseudomonas aeruginosa keratoconjunctivitis in vivo. Bioimpacts 2021; 11: 281-287.
27. Ackermann HW. Phage classification and characterization. Methods Mol Biol 2009; 501: 127-140.
28. Peng C, Hanawa T, Azam AH, LeBlanc C, Ung P, Matsuda T, et al. Silviavirus phage ɸMR003 displays a broad host range against methicillin-resistant Staphylococcus aureus of human origin. Appl Microbiol Biotechnol 2019; 103: 7751-7765.
29. Cui Z, Feng T, Gu F, Li Q, Dong K, Zhang Y, et al. Characterization and complete genome of the virulent Myoviridae phage JD007 active against a variety of Staphylococcus aureus isolates from different hospitals in Shanghai, China. Virol J 2017; 14: 26.
30. Fierheller M, Sibbald RG. A clinical investigation into the relationship between increased periwound skin temperature and local wound infection in patients with chronic leg ulcers. Adv Skin Wound Care 2010; 23: 369- 379.
31. Jones EM, Cochrane CA, Percival SL. The effect of pH on the extracellular matrix and biofilms. Adv Wound Care (New Rochelle) 2015; 4: 431-439.
32. Yüksel EB, Yıldırım AM, Bal A, Kuloglu T. The effect of different topical agents (silver sulfadiazine, povidone-iodine, and sodium chloride 0.9%) on burn injuries in rats. Plast Surg Int 2014; 2014: 907082.
| Files | ||
| Issue | Vol 14 No 5 (2022) | |
| Section | Original Article(s) | |
| DOI | https://doi.org/10.18502/ijm.v14i5.10967 | |
| Keywords | ||
| Bacteriophage; Methicillin-resistant Staphylococcus aureus; Phage therapy; Wounds | ||
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This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License. |
How to Cite
1.
Rezaei Z, Elikaei A, Barzi SM, Shafiei M. Isolation, characterization, and antibacterial activity of lytic bacteriophage against methicillin-resistant Staphylococcus aureus causing bedsore and diabetic wounds. Iran J Microbiol. 2022;14(5):712-720.
