Anti-Pseudomonas aeruginosa activity of a C16-terpene dilactone isolated from the endophytic fungus Neofusicoccum luteum of Kigelia africana (Lam.)

Abstract

Fungal endophytes have the capacity to biosynthesize secondary metabolites that are produced by their host plants. In this study, a dilactone terpenoid of C₁₆ architecture was isolated from the fungal endophytes of Kigelia africana, in our attempt to identify anti-Pseudomonas aeruginosa metabolites. Thirty-eight fungal isolates were cultured for biomolecule production over a period of thirty days. Extracts from three (ZF 34, ZF 52 and ZF 91) of the fungi showed good anti-P. aeruginosa activity, with ZF 52 presenting the best MIC of 19.53 µg/mL and was accordingly subjected to chromatographic separation. Based on nuclear magnetic resonance (NMR) spectroscopy, high resolution mass spectrometry and single crystal X-ray diffraction (XRD) analyses, the isolated compound was identified as a C₁₆-terpene dilactone, with a structure consistent with that of the known diterpene, CJ-14445. The isolated dilactone showed anti-P. aeruginosa activity with MIC of 0.61 µg/mL, signifying the antibacterial potential of the biomolecule. The bioactive fungal isolate (ZF 52) was identified as Neofusicoccum luteum based on genomic DNA sequencing. This is the first report of the endophyte N. luteum from K. africana and the first reported occurrence of CJ-14445 in the fungus.

Figure 1

Pure fungal cultures identified from Kigella africana leaves.

Figure 2

Growth of endophytic fungi upon resuscitation on potato dextrose agar media for four days. Top view of ZF 52, ZF 53, KZ 81 & KZ 44 are represented by (A–D) respectively while bottom views are represented by (A’–D’), respectively.

Figure 3

Kirby Bauer disc diffusion assay demonstrating anti-Pseudomonas aeruginosa activity of ZF 52 extracts after varying fermentation periods (T1, T2, T3, and T4 represent 1, 2, 3 and 4 weeks, respectively).

Figure 4

Structure of compound 1.

Figure 5

UPLC-ESI–MS fingerprint of ZF 52 EtOAc extract.

Figure 6

Proposed structures for compounds identified in ZF 52 ethyl acetate extract by UPLC-ESI–MS.

Figure 7

Gel electrophoresis image of ZF 52 genomic DNA. A1—Mid-range molecular weight marker, A2—ZF52 DNA, A3—negative control, B1—Mid-range molecular weight marker, B2—PCR amplification product, B3—negative control.