In vitro and in vivo study for antibacterial activity of endolysin-HEC gel and mixture gel on acne vulgaris caused by multidrug-resistant Staphylococcus aureus bacteria

Main Article Content

Haneen Emad Hussain
Ahmed Sahib Abdulamir
Iqbal Ghalib Farhood
Ahmed Rahma Ali
Muhannad Abdullah Al-azzawy

Keywords

Chronic renal failure, Renin , Aspartate Amino Transferase Alanine Amino Transferase , Albumin, Globulin, Calcium , Sodium , Potassium .

Abstract

Background: Acne is named “acne vulgaris” medically, and it is classified as the eighth most frequent illness in the world. Acne is derived from the Greek word “acme,” which means “prime of life.” It is predominantly a condition of adolescents and may continue into adulthood.
Aim: This study aimed to extract, purify, and prepare endolysin-hydroxyethyl cellulose (HEC) gel from Staphylococcus aureus–specific bacteriophages, evaluate its effectiveness via in vitro antibacterial validation tests, and clinically investigate its ability to target facial multidrug-resistant (MDR) S. aureus when applied topically to participants with moderate-to-severe acne lesions.
Methods: Twenty-three isolates of MDR S. aureus bacteria were obtained from inflammatory acne lesions of human skin by using disposable cotton swabs. Three bacteriophages with specific lytic activity against MDR S. aureus bacteria were isolated by “conventional microbiological methods” and prepared as bacteriophage cocktail, and endolysin was extracted and purified from them. Gel-based formulations, endolysin-HEC and mixture gel from bacteriophage cocktail, were prepared. In vitro validation tests was performed using spot lysis and top layer plaque assays, and antibacterial activities were also assessed. In vivo tests included the topical application of the gel three times daily for 1 week on facial skin of volunteers aged above 25 years with moderate-to-severe acne lesions caused by MDR S. aureus bacteria.


Results: Gel-based formulations exhibited 100% in vitro lytic spectrum analysis against MDR S. aureus isolates. No allergic reaction was displayed against the therapy when applied topically on the skin (in vivo) of the subjects with moderate-to-severe facial acne lesions caused by MDR S. aureus. It showed excellent results of clinical improvement response, with decrease in inflammatory signs, size, number of comedones, and presence of decolonized MDR S. aureus viable bacterial growth as compared to the untreated acne lesions “control” when cultured microbiologically.
Conclusion: Endolysin-HEC gel and mixture gel therapy have highly significant potential as an alternative strategy for MDR S. aureus infections of acne vulgaris. It offers a real chance for those suffering from chronic acne lesions caused by MDR bacterial infections, and protects teenagers from the overuse of antibiotics, anxiety, and depression.

Abstract 491 | pdf Downloads 359

References

1. Lee HJ, Jang YJ. Recent understandings of biology, prophylaxis and treatment strategies for hypertrophic scars and keloids. Int J Mol Sci.
2018;19(3):711. https://doi.org/10.3390/ijms 19030711
2. Barbieri JS, Spaccrelli N, Margolis DJ, James WD. Approaches to limit systemic antibiotic use in acne: Systemic alternatives, emerging topical therapies, dietary modification, and laser and light- based treatments. J Am Acad Dermatol. 2019;8(2):538–49. https://doi.org/10.1016/j.jaad.2018.09.055
3. Lobanovska M, Pilla G. Focus: Drug development: Penicillin’s discovery and antibiotic resistance: Lessons for the future. Yale J Biol Med.
2017;90(1):135.
4. Harkins CP, Pichon B, Doumith M, Parkhill J, Westh H, Tomasz A, et al. Methicillin-resistant Staphylococcus aureus emerged long before the
introduction of methicillin into clinical practice. Genome Biol. 2017;18(1):130. https://doi.org/10.1186/s13059-017-1252-9
5. Tenover FC, Tickler IA, Le VM, Dewell S, Mendes RE, Goering RV. Updating molecular diagnostics for detecting methicillin-susceptible and
methicillin-resistant Staphylococcus aureus isolates in blood culture bottles. J Clin Microbiol. 2019;57(11):e01195–19.
https://doi.org/10.1128/JCM.01195-19
6. Abd El-Aziz NK, Abd El-Hamid MI, Bendary MM, El-Azazy AA, Ammar AM. Existence of vancomycin resistance among methicillin resistant
S. aureus recovered from animal and human sources in Egypt. Slov Vet Res. 2018;55(Suppl 20):221–30. https://doi.org/10.26873/SVR-649-
2018
7. Silva V, Almeida F, Carvalho JA, Castro AP, Ferreira E, Manageiro V, et al. Emergence of community-acquired methicillin-resistant
Staphylococcus aureus EMRSA-15 clone as the predominant cause of diabetic foot ulcer infections in Portugal. Eur J Clin Microbiol Infect Dis.
2020;39(1):179–86. https://doi.org/10.1007/s10096-019-03709-6
8. The AMR innovation challenge. The AMR Action Fund. 2022. [cited 2022 Jul 27]. Available from: https://www.amractionfund.con/
9. Lehman SM. Mearns G, Rankin D, Cole RA, Smrekar F, Branston SD, et al. Design and preclinical development of a phage product for the
treatment of antibiotic-resistant Staphylococcus aureus infections. Viruses. 2019;11(1):88. https://doi.org/10.3390/v11010088.
10. Huang Y, Wang W, Zhang Z, Gu Y, Huang A, Wang J, Hao H. Phage products for fighting antimicrobial resistance. Microorganisms.
2022;10(7):1324. https://doi.org/10.3390/microorganisms10071324
11. Haddad H, Schmelcher M, Sabzalipoor H, Hosseini ES, Moniri R. Recombination endolysins as potential therapeutic against antibiotic-resistant Staphylococcus aureus: Current status of research and a novel delivery strategies. Clin Microbiol Rev. 2018;31(1):300071–17.
https://doi.org/101128/CMR.0071-17
12. Vander Elst N, Linden SB, Lavigne R, Meyer E, Briers Y, Nelson DC. Characterization of the bacteriophage-derived endolysins PlySs2 and
PlySs9 with in vitro lytic activity against bovine mastitis Streptococcus uberis. Antibiotics. 2020;9(9):621. https://doi.org/103390/antibiotics9090621
13. Rashedul H, Mrityunjoy A, Rashed N. Prevalence of vancomycin resistant Staphylococcus aureus (VRSA) in methicillin resistant S. aureus (MRSA) strains isolated from burn wound infections. Tzu Chi Med J. 2016;28(2):4953. https://doi.org/10.1016/j.tcmj.2016.03.002
14. Hyman P. Phages for phage therapy: Isolation, characterization, and host range breadth. Pharmaceuticals (Basel). 2019;12(1):35.
https://doi.org/10.3390/ph/12010035
15. Missiakas DM, Schneewind O. Growth and laboratory maintenance of Staphylococcus aureus. Curr Protoc Microbiol. 2013;28(1):91.
https://doi.org/10.1002/9780471729259.mcoqcols28.
16. Pallavali RR, Degati VL, Lomada D, Reddy MC, Durbaka VRP. Isolation and in vitro evaluation of bacteriophages against MDR bacterial isolates from septic wound infections. PLoS One. 2017;12(7):179–245. https://doi.org/10.1371/journal.pone.0179245
17. Gonzalez-Menendez E, Fernandez L, Gutierrez D, Rodríguez A, Martínez B, García P. Comparative analysis of different preservation techniques for the storage of Staphylococcus phages aimed for the industrial development of phage-based antimicrobial products development of phagebased antimicrobial products. PLoS One. 2018;13(10):0205728. https://doi.org/10.1371/journal.pone.0205728
18. Rodrguez-Melconc C, Alonso-Callejac C, GarciaFernandez C, Carballo J, Capita R. Minimum inhibitory concentration (MIC) and minimum
bactericidal concentration (MBC) for twelve antibiotics (Biocides and antibiotics) in right strains of Listeria monocytogenes. Biology.
2021;11(1):46. https://doi.org/10.3390/biology/1010046
19. Mohiuddin AK. A comprehensive review of acne vulgaris. J Clin Pharm. 2019;1(1):17–45. https://doi.org/10.15226/2378-1726/6/2/00186
20. Dobler D, Schmidts T, Zinecker C, Schlupp P, Schäfer J, Runkel F. Hydrophilic ionic liquids as ingredients of gel-based dermal formulations.
AAPS Pharm Sci Tech. 2016;17(4):923–31. https://doi.org/10.1208/s12249-015-042-y
21. Jonczyk-Matysiak E, Weber-Dabrowska B, Zaczek M, Międzybrodzki R, Letkiewicz S, ŁusiakSzelchowska M, et al. Prospects of phage
application in the treatment of acne caused by Propionibacterium acnes. Front Microbiol. 2017;8:164.
https://doi.org/10.3389/fmicb.2017.00164
22. Principi N, Silvestri E, Esposito S. Advantages and limitations of bacteriophages for the treatment of bacterial infections. Front Pharmacol. 2019;10:1–9. https://doi.org/10.3389/fphar.2019.00513
23. McCallin S, Sacher JC, Zheng J, Benjamin K Chan. Current state of compassionate phage therapy. Viruses. 2019;11(4):343.
https://doi.org/10.3390/v11040343
24. Kazmierczak Z, Gorski A, and Dabrowska D. Facing antibiotic resistance: Staphylococcus aureus phages as a medical tool. Viruses. 2015;7(4):1667. https://doi.org/10.3390/v7041667
25. Chan BK, Abedon ST, Loc-Carrillo C. Phage cocktails and the future of phage therapy. Future Microbiol. 2013;8:769–83. https://doi.org/
10.2217/fmb.13.47
26. Wittebole X, De Roock S, Opal S. Ahistorical overview of bacteriophage therapy as an alternative to antibiotics for the treatment of bacterial pathogens. Virulence. 2014;5(1):226–35. https://doi.org/10.4161/viru.25991
27. Wan X, Hendrix H, Skurnik M, Lavigne R. Phagebased target discovery and its exploitation towards novel antibacterial molecules. Curr Opin Biotechnol. 2021;68:1–7. https://doi.org/10.1016/j.copbio.2020.08.015
28. Gutiérrez D, Fernández L, Rodríguez A, García P. Are phage lytic proteins the secret weapon to kill Staphylococcus aureus? mBio. 2018;9(1):e01923–17. https://doi.org/10.1128/mBio.01923-17
29. Villa TG, Crespo PV, editors. Enzybiotics: Antibiotic enzymes as drugs and therapeutics. Hoboken, NJ: John Wiley & Sons; 2010. 284 p.
https://doi.org/10.1002/9780470570548.oth1
30. Ramachandran G. Gram-positive and gram-negative bacterial toxins in sepsis: A brief review. Virulence. 2014;5(1):213–8. https://doi.org/10.4161/viru.27024.
31. Zhang H, Buttaro BA, Fouts DE, Sanjari S, Evans BS, Stevens RH. Bacteriophage φEf11 ORF28 endolysin, a multifunctional lytic enzyme with
properties distinct from all other identified Enterococcus faecalis phage endolysins. Appl Environ Microbiol. 2019;85(13):e00555–19. https://doi.org/10.1128/AEM.00555-19.
32. Love MJ, Bhandari D, Dobson RCJ, Billington C. Potential for bacteriophage endolysins to supplement or replace antibiotics in food
production and clinical care. Antibiotics (Basel). 2018;7(1):17. https://doi.org/10.3390/antibiotics7010017.
33. Kwon SS, Kong BJ, Park SN. Physicochemical properties of PH—Sensitive hydrogels based on hydroxyethyl cellulose-hyaluronic acid and for
applications as transdermal delivery systems for skin lesions. Eur J Pharm Biopharm. 2015;92:146–54. https://doi.org/10.1016/ejpb.2015.02.025
34. Maciejewska B, Olszak T, Drulis-Kawa Z. Applications of bacteriophages versus phage enzymes to combat and cure bacterial infections:
An ambitious and also a realistic application. Appl Microbiol Biotechnol. 2018;102(6):2563–81. https://doi.org/10.1007/s00253-018-8811-1
35. Hamed ZO, Abdulamir AS. Therapeutic effect of cloned lysin derived from bacteriophage on infected wound with multidrug-resistant
Staphylococcus aureus. PhD Thesis. College Med. AL-Nahrain University, 2021.
36. Seah C, Alexander DC, Louie L, Simor A, Low DE, Longtin J, et al. MupB, a new high-level mupirocin resistance mechanism in Staphylococcus aureus.
Antimicrob Agents
Chemother. 2012;56(4):1916–20.
https://doi.org/10.1128/AAC.05325-11
37. Jault P, Leclerc T, Jennes S, Pirnay JP, Que YA,
Resch G, et al. Efficacy and tolerability of a
cocktail of bacteriophages to treat burn wounds
infected by Pseudomonas aeruginosa (PhagoBurn):
Arandomised, controlled, double-blind phase1/2
trial. Lancet Infect Dis. 2019;19(1):35–45.
https://doi.org/10.1016/s1473-3099(18)30482-1
38. Pirnay JP, Ferry T, Resch G. Recent progress toward
the implementation of phage therapy in western
medicine. FEMS Microbiol Rev. 2022;46(1):40.
https://doi.org/10.1093/femsre/fuab040
39. Khan A, Subba R, Joshi H. Phage therapy in the
Covid-19 era: Advantages over antibiotics. Curr
Res Microb Sci. 2022;3:100115.
https://doi.org/10.1016/j.crmicr.2022.100115
40. Lebax D, Merabishvili M, Caudron E, Lannoy D,
Van Simaey L, Duyvejonck H, et al. A case of
phage therapy against pandrug-resistant
Achromobacter xylosoxidans in a 12 year-old lung
transplanted cystic fibrosis patient. Viruses.
2021;13(1):60. https://doi.org/10.3390/v13010060