The influence of endophytic Penicillium oxalicum B4 on growth and artemisinin biosynthesis of in vitro propagated plantlets of Artemisia annua L.

Artemisia annua L. is presently the sole natural source of antimalarial drug artemisinin. We established dual cultures of A. annua callus or regenerated plantlets with endophytic Penicillium oxalicum B4 to explore endophyte-mediated effects on artemisinin biosynthesis. Although A. annua callus could not produce artemisinin with or without the endophyte, simultaneous growth stimulation of the endophyte and inhibition of A. annua callus were observed in dual cultures. In an in vitro dual culture of endophyte-regenerated plantlets, the endophyte enhanced growth and artemisinin content of host plant. The endophyte could simultaneously induce oxidative stress in regenerated plantlets through the generation of reactive oxygen species (ROS) including O-2(center dot-) and H2O2, which was then accompanied by the activation of antioxidant enzymes such as peroxidase, catalase and superoxide dismutase during the later stages. There was a significant increase in amorphadiene synthase (ADS) and amorpha-4,11-diene monooxygenase (CYP71AV1) transcripts in dual culture of endophyte-plantlets. The induced ROS could modulate the expression of those key genes for artemisinin biosynthesis and might be responsible for conversion of artemisinin acid into artemisinin production. Our results demonstrated that endophytic P. oxalicum B4 could be applied as a promising means to enhance artemisinin production in plants.