Full survival of Galleria mellonella infected with Staphylococcus aureus after treatment with Nisin Z

Abstract

Diabetes mellitus affects nearly 6.4% of the worldwide population, and this number may double by 2030. Up to 25% of diabetic patients may develop diabetic foot ulcers (DFUs). Among DFU patients, 80% will suffer lower-limb amputations due to diabetic foot infections (DFIs), which are generally colonized by polymicrobial biofilms. Staphylococcus aureus is the DFIs’ predominant pathogen, frequently found together with Pseudomonas aeruginosa in chronic and severe infections. Due to their high virulence and antibiotic resistant profile, it is crucial to find alternatives to conventional antibiotics for DFI treatment. Previous studies showed that Nisin Z supplemented with EDTA (0.4%) had higher antibacterial, antibacteriostatic, and antibiofilm efficiency towards S. aureus and P. aeruginosa DFI isolates. Therefore, we aimed to confirm these data in a Galleria mellonella model. G. mellonella wax moth larvae were reared at 25 °C in the dark, and worms of the final- instar larval stage were selected (10 larvae for each experiment). The larvae were injected with a lethal dose of each bacterium via the hindmost left proleg. After approximately 1 hour, the larvae were injected with Nisin Z (200 μg/ml) in the penultimate right proleg. Then, they were kept in Petri dishes and maintained in the dark at 37 °C for 120 hours. Each larva was scored daily on the G. mellonella health index: survival, melanization, mobility, and cocoon formation. Experiments were performed with three independent replicates. Nisin Z treatment led to 100% survival of the larvae infected with S. aureus but had no antibacterial activity against P. aeruginosa. Unexpectedly, EDTA supplementation did not increase antipseudomonal activity. Nisin Z was not cytotoxic to the larvae. Nisin Z may be used as a complement to conventional antibiotic therapy against S. aureus in DFI. G. mellonella is a valuable model before proceeding to preclinical studies in mammals. Congress of Microbiology and Biotechnology 2023 385 MicroBiotec 2023 - Covilhã, Portugal Acknowledgements: Authors would like to acknowledge: CIISA—Centro de Investigação Interdisciplinar em Sanidade Animal, Faculdade de Medicina Veterinária, Universidade de Lisboa, Lisbon, Portugal (Project UIDB/00276/2020); AL4AnimalS-Laboratório Associado para a Ciência Animal e Veterinária (LA/P/0059/2020); iBB Institute for Bioengineering and Biosciences (UIDB/04565/2020 and UIDP/04565/2020), i4HB Associate Laboratory Institute for Health and Bioeconomy (LA/P/0140/2020); and GHTM - (UID/04413/2020 and LA-REAL – LA/P/0117/2020).