In vitro synergistic effect of minocycline combined with antifungals against Cryptococcus neoformans

Cryptococcus neoformans infections occur in immunocompromised patients, especially those with HIV infection, chemoradiotherapy after cancer, and organ transplantation. Infection can cause pneumonia and meningoencephalitis in severe cases with a high mortality rate if not treated. Although fluconazole and amphotericin B are the first-line treatments for cryptococcosis, the rate of fluconazole resistance has increased significantly due to long-term use. Minocycline is a derivative of tetracycline that exerts its antibacterial effect through inhibition of bacterial protein synthesis. It is also able to pass the blood-brain barrier to act on the central nervous system. The present study investigates the effects of minocycline in combination with antifungals in treating C. neoformans.

To determine in vitro interactions of minocycline combined with itraconazole, voriconazole, posaconazole, fluconazole and amphotericin B against C. neoformans.

The minimum inhibitory concentrations (MIC) of the antifungals were determined by the CLSI Clinical and Laboratory Standards Institute M27-A3 microdilution method. The in vitro synergistic effects of minocycline combined with itraconazole, voriconazole, posaconazole, fluconazole, and amphotericin B on C. neoformans were detected by the broth microdilution checkerboard technique and disk diffusion testing.

The working concentration ranges were 0.125-4 µg/mL for itraconazole, 0.03-0.125 µg/ml for voriconazole, 0.03-1 µg/ml for posaconazole, 0.25-16 µg/ml for fluconazole, and 0.125-2 µg/ml for amphotericin B. The synergistic rates of minocycline combinations against C. neoformans were 55% with itraconazole, 10% with voriconazole, 85% with posaconazole, 20% with fluconazole, and 70% with amphotericin B. The effective MIC value of minocycline in the synergistic combination decreased to 2-32 µg/ml, while the MIC of itraconazole decreased to 0.03-0.125 µg/ml, voriconazole 0.03-0.125 µg/ml, posaconazole 0.03-0.125 µg/ml, 0.125-4 µg/ml fluconazole, and 0.06-0.50 µg/ml amphotericin B. The disk diffusion assay showed that the plates containing minocycline and antifungal drugs produced inhibition zones with diameters larger than the single drug plates. Minocycline showed no antagonistic effect in the combinations. In conclusion, the combination of minocycline and azoles or amphotericin B has synergistic effects against C. neoformans in vitro.