Alternative production strategies of active bioinsecticides with plant cell cultures and endophytes from Azadirachta indica
14 December 2016 (online)
All parts of neem (Azadirachta indica A. Juss) show a broad spectrum efficacy against insect pests including insecticidal, anti-feedant or insect repellent activities. The commercial products are obtained only in low concentrations via complex extraction procedure. Several studies have shown that plant cell cultures of A. indica can produce bioinsecticides like Azadirachtin A. In light of the increasing reports on endophytes it can further be hypothesized that some of these compounds are either directly produced by endophytic microorganisms or their production is linked to the plant metabolism. Both strategies allow to by-pass the cost intensive transport of plant material and extraction of plant metabolites. The overall aim of a BMBF funded project is to develop a competitive process to produce high concentrations of bioinsecticidal compounds with neem plant cell cultures and endophytes. Here, we present data on isolation of endophytes and induction of callus as well as first results of our screening approach. First, we induced plant cell cultures from various plant tissues with a medium that allows a callus proliferation from more than 90% of the explants. Besides, in total 346 endophytes were isolated from plant material of different origins. Furthermore we developed an in situ product removal method to bind and stabilize more than 85% of the produced Azadirachtin A in liquid medium. We report first cultivation data obtained with the prototype of a novel high-throughput cultivation system (RoboLector® with an integrated BioLector® Pro from m2 p-labs GmbH) that enables 48 parallel batch or 32 fed-batch cultivations in a volume under 1 ml. In addition to that, we will present results on characterization of selected endophytes as well as induced cell lines, submerged cultivation and classification of metabolites via UHPLC-DAD-MS/MS. Finally, we will show effects of extracted metabolites on the development of Sf9 cell cultures.
Keywords: Azadirachtin-A, plant cell culture, endophytes, high-throughput cultivation.