Penicillium nalgiovense Laxa isolated from Antarctica is a new source of the antifungal metabolite amphotericin B

Abstract

Background: The need for new antibiotic drugs increases as pathogenic microorganisms continue to develop resistance against current antibiotics. We obtained samples from Antarctica as part of a search for new antimicrobial metabolites derived from filamentous fungi. This terrestrial environment near the South Pole is hostile and extreme due to a sparsely populated food web, low temperatures, and insufficient liquid water availability. We hypothesize that this environment could cause the development of fungal defense or survival mechanisms not found elsewhere.

Results: We isolated a strain of Penicillium nalgiovense Laxa from a soil sample obtained from an abandoned penguin's nest. Amphotericin B was the only metabolite secreted from Penicillium nalgiovense Laxa with noticeable antimicrobial activity, with minimum inhibitory concentration of 0.125 μg/mL against Candida albicans. This is the first time that amphotericin B has been isolated from an organism other than the bacterium Streptomyces nodosus. In terms of amphotericin B production, cultures on solid medium proved to be a more reliable and favorable choice compared to liquid medium.

Conclusions: These results encourage further investigation of the many unexplored sampling sites characterized by extreme conditions, and confirm filamentous fungi as potential sources of metabolites with antimicrobial activity.

Keywords: Amphotericin B; Antarctica; Penicillium nalgiovense Laxa.

Figure 1

Extraction procedure. Cultivation, extraction, and isolation of metabolites with antimicrobial activity from Penicillium nalgiovense Laxa. Fractions without antimicrobial activity are marked with thinner arrows under the HPLC boxes, and those with antimicrobial activity are marked with an asterisk (*).

Figure 2

Preparative work of fungal extracts. Chromatograms from preparative isolation recorded at 280 nm of (A) filtrate of liquid-based fungal culture, (B) methanol extracts of mycelial growth, and (C) methanol extract of fungal cultures on solid medium. The horizontal bars indicate fractionation, and the asterisks (*) indicate antimicrobial activity. Amphotericin B eluted at 21.50 min.

Figure 3

Identification of amphotericin B. The preparative and analytical results of the experiments identifying the only substance with antimicrobial activity in a culture extract of Penicillium nalgiovense Laxa. The methanol extracts of the mycelial growth and liquid filtrate were fractionated with HPLC (bottom two chromatograms recorded at 280 nm), analyzed with Q-TOF MS (middle spectra), and compared with the reference substance amphotericin B (top figure). Identical elution times (21 minutes 50 seconds), mother ions (924.4989 and 906.4837 m/z), and daughter ions (743.5, 761.5 m/z) between the extracted compound and reference amphotericin B, together with NMR analysis and antimicrobial assays, led to the identification of amphotericin B in the fungal extracts.

Figure 4

¹ H NMR spectra of extract (below) and reference amphotericin B (above) dissolved in dimethylsulfoxide- d6 .