An antimicrobial peptide from endophytic Fusarium tricinctum of Rhododendron tomentosum Harmaja
Endophytic microbes have attracted considerable attention as a completely new source of pharmaceuticals in recent years. An endophytic Fusarium tricinctum was isolated from Rhododendron tomentosum, a shrub that is found widely across the northern hemisphere. The endophytic F. tricinctum produced antimicrobial compounds that were active against Staphylococcus carnosus, Candida albicans and C. utilis. The transcriptome of the F. tricinctum was sequenced, and a total of 12,006 contigs were assembled, having an N50 value of 1390 bp. Analysis of the transcriptomic PD library of F. tricinctum yielded an antimicrobial peptide named Trtesin. The expression of Trtesin transcripts was >1000 fold in the mRNA library originating from PDB-grown fungi exhibiting high antimicrobial activity compared to FG4-grown fungi with no antimicrobial activity. Trtesin was cloned, expressed, and purified in pET32(a), and it consisted of 52 amino acids with 6 cysteine molecules. The molecular weight of Trtesin is 6138.92 Da. An additional N-terminal sequencing was done to confirm the intact peptide, as well as to check the correct amino acid sequence. The MIC of Trtesin was determined against several bacteria as 64 μg/mL, and peptide showed a mild activity against F. oxysporum in agar diffusion assay, as a zone of inhibition of 10 mm was formed at 100 μg of peptide.
KeywordsFungal defensin Antimicrobial Endophyte Transcriptome
We thankfully acknowledge the funding received from the European Union projects (PIAP-GA-2008-218191 and PIIF-GA-2008-220253). We thank Steen Buskov (Novozymes A/S) for expert assistance with MS work and Margaretha Hasselbalch (Novozymes) for technical assistance on total RNA extraction. We also thank Ida Ellingsgaard and Jeanette Theil for technical assistance with regard to the expression and purification of Trtesin.
- Altier DJ, Ellanskaya I, Gilliam JT, Hunter-Cevera J, Presnail JK, Schepers E, Simmons CR, Torok T, Yalpani N (2009) Antifungal polypeptides. US patent: US7589176 1–47.Google Scholar
- Hyde KD, Soytong K (2008) The fungal endophyte dilemma. Fungal Diversity 33:163–173Google Scholar
- Kesting JR, Olsen L, Staerk D, Tejesvi MV, Kini KR, Prakash HS, Jaroszewski JW (2011) Production of unusual dispiro metabolites in Pestalotiopsis virgatula endophyte cultures: HPLC-SPE-NMR, electronic circular dichroism, and time-dependent density-functional computation study. J Nat Prod 74(10):2206–2215PubMedCrossRefGoogle Scholar
- Mygind PH, Fischer RL, Schnorr KM, Hansen MT, Sonksen CP, Ludvigsen S, Raventos D, Buskov S, Christensen B, De Maria L, Taboureau O, Yaver D, Elvig-Jørgensen SG, Sørensen MV, Christensen BE, Kjærulff S, Frimodt-Moller N, Lehrer RI, Zasloff M, Kristensen H-H (2005) Plectasin is a peptide antibiotic with therapeutic potential from a saprophytic fungus. Nature 437:975–980PubMedCrossRefGoogle Scholar
- National Committee for Clinical Laboratory Standards (2002) Performance Standards for Antimicrobial Susceptibility Testing: Twelfth Informational Supplement M100-S12. NCCLS, Wayne, PA, USA.Google Scholar
- Smith SA, Tank DC, Boulanger L-A, Bascom-Slack CA, Eisenman K, Kingery D, Babbs B, Fenn K, Greene JS, Hann BD, Keehner J, Kelley-Swift EG, Kembaiyan V, Lee SJ, Li P, Light DY, Lin EH, Ma C, Moore E, Schorn MA, Vekhter D, Nunez PV, Strobel GA, Donoghue MJ, Strobel SA (2008) Bioactive Endophytes Warrant Intensified Exploration and Conservation (HA El-Shemy, Ed.). PLoS One 3:e3052PubMedCrossRefGoogle Scholar
- Tejesvi MV, Kini KR, Prakash HS, Subbiah V, Shetty HS (2007) Genetic diversity and antifungal activity of species of Pestalotiopsis isolated as endophytes from medicinal plants. Fungal Divers 24:37–54Google Scholar