Efficacy of liposomal amphotericin B eye drops at two concentrations in experimental aspergillus keratomycosis in rabbits
Abstract
Background: Keratomycosis, a fungal corneal infection prevalent in tropical regions, frequently leads to visual impairment and blindness. Liposomal amphotericin B (L-AmB) offers improved efficacy and reduced toxicity compared to conventional amphotericin B deoxycholate and represents a promising option for topical ophthalmic use.
Objective: To evaluate and compare the therapeutic efficacy of L-AmB eye drops at 0.15% and 0.5% concentrations in experimental Aspergillus keratomycosis in New Zealand White rabbits.
Methods: Four rabbits received midstromal corneal inoculation with a mixed suspension of Aspergillus flavus, A. fumigatus, A. niger, and A. terreus (2 x 10⁴ CFU/0.05 mL). L-AmB eye drops were applied hourly from day 8 post-inoculation until corneal cultures became negative. Antifungal susceptibility was assessed by disc diffusion. Clinical response was monitored by slit lamp examination using a modified scoring system.
Results: Negative corneal cultures were achieved in all treated eyes within 10 to 18 days (mean 15 days, L-AmB 0.15%) and 11 to 15 days (mean 14 days, L-AmB 0.5%). No toxic effects were observed. Complete corneal transparency without cicatrix was achieved in one eye treated with L-AmB 0.15%.
Conclusion: Both concentrations were effective and safe. Liposomal AmB 0.15% appears as efficacious as 0.5% for experimental Aspergillus keratomycosis. Clinical studies in humans are warranted.
References
Khorashad AS, Kashani M, Parandin F, et al. Prevalence of Fungal Keratitis in Suspected Keratomycosis Cases Over Five Years in Southeastern Iran. Arch Clin Infect Dis. 2025;20(6):e161256. https://doi.org/10.5812/archcid-161256
Brown L, Leck AK, Gichangi M, Burton MJ, Denning DW. The global incidence and diagnosis of fungal keratitis. Lancet Infect Dis. 2021;21(3):e49-e57. https://doi.org/10.1016/S1473-3099(20)30448-5
Hoffman JJ, Burton MJ, Leck A. Mycotic Keratitis: A Global Threat from the Filamentous Fungi. J Fungi (Basel). 2021;7(4):273. https://doi.org/10.3390/jof7040273
Ahmadikia K, Aghaei Gharehbolagh S, Fallah B, et al. Distribution, Prevalence, and Causative Agents of Fungal Keratitis: A Systematic Review and Meta-Analysis (1990 to 2020). Front Cell Infect Microbiol. 2021;11:698780. https://doi.org/10.3389/fcimb.2021.698780
Wahyuningsih R, Adawiyah R, Sjam R, et al. Serious fungal disease incidence and prevalence in Indonesia. Mycoses. 2021;64(10):1203-1212. https://doi.org/10.1111/myc.13304
Mahmoud YAG. Fungal keratitis efficient treatments using surface active agents (cetrimide): an overview. J Bacteriol Mycol Open Access. 2016;2(3):69-73. https://doi.org/10.15406/jbmoa.2016.02.00026
Sharma N, Bagga B, Singhal D, Nagpal R, Kate A, Saluja G, Maharana PK. Fungal keratitis: A review of clinical presentations, treatment strategies and outcomes. Ocul Surf. 2022;24:22-30. https://doi.org/10.1016/j.jtos.2021.12.001
Qasim M, Baipaywad P, Udomluck N, et al. Enhanced Therapeutic Efficacy of Lipophilic Amphotericin B Against Candida albicans with Amphiphilic Poly(N-isopropylacrylamide) Nanogels. Macromolecular Research. 2014;22(10):1125-1131. https://doi.org/10.1007/s13233-014-2162-2
Rodrigues CF, Henriques M. Liposomal and Deoxycholate Amphotericin B Formulations: Effectiveness against Biofilm Infections of Candida spp. Pathogens. 2017;6(4):62. https://doi.org/10.3390/pathogens6040062
Hamill RJ. Amphotericin B formulations: a comparative review of efficacy and toxicity. Drugs. 2013;73(9):919-934. https://doi.org/10.1007/s40265-013-0069-4
Lim C-b, Abuzar SM, Karn PR, et al. Preparation, Characterization, and In Vivo Pharmacokinetic Study of the Supercritical Fluid-Processed Liposomal Amphotericin B. Pharmaceutics. 2019;11(11):589. https://doi.org/10.3390/pharmaceutics11110589
Stone NRH, Bicanic T, Salim R, Hope W. Liposomal Amphotericin B (AmBisome®): A review of the pharmacokinetics, pharmacodynamics, clinical experience and future directions. Drugs. 2016;76(4):485-500. https://doi.org/10.1007/s40265-016-0538-7
Cornely OA, Sprute R, Bassetti M, et al. Global guideline for the diagnosis and management of candidiasis: an initiative of the ECMM in cooperation with ISHAM and ASM. Lancet Infect Dis. 2025;Feb 13:S1473-3099(24)00749-7. https://doi.org/10.1016/S1473-3099(24)00749-7
Agarwal R, Iezhitsa I, Agarwal P, et al. Liposomes in topical ophthalmic drug delivery: an update. Drug Deliv. 2014:1-17. https://doi.org/10.3109/10717544.2014.943336
Yavas GF, Ozturk F, Kusbeci T, et al. Antifungal efficacy of voriconazole, itraconazole and amphotericin B in experimental Fusarium solani keratitis. Graefes Arch Clin Exp Ophthalmol. 2008;246:275-279. https://doi.org/10.1007/s00417-007-0687-1
Ghosh AK, Rudramurthy SM, Gupta A, et al. Evaluation of Liposomal and Conventional Amphotericin B in Experimental Fungal Keratitis Rabbit Model. Transl Vis Sci Technol. 2019;8(3):35. https://doi.org/10.1167/tvst.8.3.35
Touvron G, Denis D, Doat M, et al. Efficacité de l'amphotéricine B liposomale dans le traitement d'une kératite à Fusarium solani résistante. J Fr Ophtalmol. 2009;32:721-726. https://doi.org/10.1016/j.jfo.2009.10.011
Tandon R, Lomi N, Vanathi M, Gupta N, et al. Evaluation of liposomal amphotericin B for the treatment of fungal keratitis in a tertiary eye care hospital. Indian J Ophthalmol. 2023;71(2):518-523. https://doi.org/10.4103/ijo.IJO_1474_22
Amalia H, Estuningtyas A, Ridwan R, Sitompul R, Adawiyah R, Freisleben HJ. Pharmacokinetic and Toxicological Studies of Liposomal Amphotericin B Eye Drops on the Rabbit Eye. Int J Pharm Res. 2020;12(4):1828-1834. https://doi.org/10.31838/ijpr/2020.12.04.261
Clinical and Laboratory Standards Institute (CLSI). Performance Standards for Antimicrobial Disk Diffusion Susceptibility Tests. 19th ed. Wayne, PA: CLSI; 2009. Document M100-S19.
Fuentes GC, Newgren J. Physiology and Clinical Pathology of Laboratory New Zealand White Rabbits Housed Individually and in Groups. J Am Assoc Lab Anim Sci. 2008;47(2):35-38.
Mähler M, Berard M, Feinstein R, et al. FELASA recommendations for the health monitoring of mouse, rat, hamster, guinea pig and rabbit colonies in breeding and experimental units. Lab Anim. 2014;48(3):178-192. https://doi.org/10.1177/0023677213516312
Warnke GR, Kluge R. Feeding concepts and methods in the laboratory animal facility and in animal experiments: Rabbits (Oryctolagus cuniculus L.) at the example of the breed New Zealand White. GV-SOLAS, Committee for Laboratory Animal Nutrition. 2015, translated 2021:1-15.
Akter MA, Yesmin N, Talukder MBA, et al. Evaluation of anaesthesia with xylazine-ketamine and xylazine-fentanyl-ketamine in rabbit: A comparative study. J Adv VetBio Sc Tech. 2023;8(1):38-46. https://doi.org/10.31797/vetbio.1129402
Henke JA, Astner S, Brill T, et al. Comparative study of three intramuscular anaesthetic combinations (medetomidine/ketamine, medetomidine/fentanyl/midazolam and xylazine/ketamine) in rabbits. Vet Anaesth Analg. 2005;32(5):261-271. https://doi.org/10.1111/j.1467-2995.2005.00242.x
Leary S, Underwood W, Anthony R, et al. AVMA Guidelines for the Euthanasia of Animals. 2013 ed. Schaumburg, IL: American Veterinary Medical Association; 2013.
Eaton JS, Miller PE, Bentley E, Thomasy SM, Murphy CJ. Slit Lamp-Based Ocular Scoring Systems in Toxicology and Drug Development: A Literature Survey. J Ocul Pharmacol Ther. 2017;33(10):707-717. https://doi.org/10.1089/jop.2017.0021
Kaji Y, Yamamoto E, Hiraoka T, Oshika T. Toxicities and Pharmacokinetics of subconjunctival injection of liposomal amphotericin B. Graefes Arch Clin Exp Ophthalmol. 2009;247:549-553. https://doi.org/10.1007/s00417-008-1003-4
Arianti A, Setiabudy R, Rozaliyani A, Siregar NC, Susiyant M. Comparison of intravitreal antifungal 100 µg voriconazole and 5 µg amphotericin B in experimental Aspergillus flavus endophthalmitis model in rabbits. Med J Indones. 2019;28:215-222. https://doi.org/10.13181/mji.v28i3.1794
Kumar G, Kalita AH, Ahmed J, et al. Advancing fungal keratitis treatment: transitioning from conventional amphotericin B therapy to nanocarrier-based delivery systems. Naunyn-Schmiedeb Arch Pharmacol. 2025; May 17. https://doi.org/10.1007/s00210-025-04268-8
FlorCruz NV, Evans JR. Medical interventions for fungal keratitis. Cochrane Database of Systematic Reviews. 2015;Issue 4:CD004241. https://doi.org/10.1002/14651858.CD004241.pub4
Christy JS, Mathews P, Ravisankar R, Akpek EK. Long-term outcomes of therapeutic corneal transplants performed in a tertiary eye care hospital in South India. Indian J Ophthalmol. 2023;71:3171-3177. https://doi.org/10.4103/IJO.IJO_1344_23
Mundra J, Dhakal R, Mohamed A, et al. Outcomes of therapeutic penetrating keratoplasty in 198 eyes with fungal keratitis. Indian J Ophthalmol. 2019;67:1599-1605. https://doi.org/10.4103/ijo.IJO_1952_18
Albadr AA, Tekko IA, Vora LK, et al. Rapidly dissolving microneedle patch of amphotericin B for intracorneal fungal infections. Drug Deliv Transl Res. 2022;12:931-943. https://doi.org/10.1007/s13346-021-01032-2
Wang YC, Wang JS, Wang B, Peng X, Xie HT, Zhang MC. Two-step strategy: conjunctival flap covering surgery combined with secondary deep anterior lamellar keratoplasty for the treatment of high-risk fungal keratitis. Int J Ophthalmol. 2023;16(7):1065-1070. https://doi.org/10.18240/ijo.2023.07.10
Copyright (c) 2026 Husnun Amalia, Rahmawati Ridwan, Ari Estuningtyas, Ratna Sitompul, Robiatul Adawiyah, Hans-Joachim Freisleben
This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.
