gms | German Medical Science

Objectives: Fungal periprosthetic joint infections are difficult to treat and often associated with a limited outcome for patients. Candida species account for approximately 90% of all fungal infections. In vivo biofilm models play major role to study biofilm development, morphology, and regulatory molecules for bacteria. However, in vivo modeling of biofilm-associated fungi models are very rare. Furthermore, due to ethical restrictions, mammalian models are replaced with other alternative models in basic research. Recently, we have developed insect infection model G. mellonella larvae to study implant associated biofilm infections with bacteria. This model organism is easy to handle, cheap and ethical restriction free and could be used for the high through put screening of anti microbial compounds to treat implant associated bacterial infections but was not used for fungi biofilm infection yet. Thus, we aimed to establish G. mellonella as in vivo model to study fungal implant infections using Candida albicans as model organism and to test anti-fungal medication.

Methods: G. mellonella larvae are maintained at 30^oC on artificial diet in an incubator. Titanium and Stainless steel K-wires were cut into small pieces with size of 4mm.For the infection process, implants were pre-incubated in specified fungal growth culture Candida albicans at 1x10⁷ CFU/ml for 30 min at 150 rpm shaking conditions. Later, these implants were washed with 10ml PBS and implanted in the larvae. To analyze the susceptibility of the implant-associated fungal infections towards anti fungal compounds, the larvae were treated with amphotericin B, fluconazole and voriconazole after 24h of implantation. The effect of anti-fungal compounds was measured in terms of survival observation for 5 days and fungal load in larvae on 2nd day. To reveal the fungal biofilm formation on implant, the implants were removed on day 3 and processed for SEM analysis.

Results and conclusion: The results are very promising to use of G. mellonella as in vivo model to study fungal infections on implanted materials. Pre-incubated K-wire caused the Candida infection and observed the death of the larvae. The treatment with antifungal compounds recovered the larvae from the implant-infection, except in case of Voriconazole. However, the recovery of the larvae from implant infection with treatment of anti fungal compounds was not effective as the case of planktonic infection treatment with anti fungal compounds in G. mellonella, which highlights resistance of biofilm to anti-fungal compounds. Further, the treatment Amphotericin B and Fluconazole reduced the C. albicansload in larvae tissue. The SEM analysis revealed the formation fungal biofilm with hyphae and spores associated with larvae tissue on implant surface. Thus, our study highlights the use of G. mellonella larvae as alternative in vivo model to study implant-associated infections that reduces the use of the higher mammals.