Host-delivered RNAi: An effective strategy to silence genes in plant parasitic nematodes

Root-knot nematodes (Meloidogyne spp.) are obligate, sedentary endoparasites that infect many plant species causing large economic losses worldwide. Available nematicides are being banned due to their toxicity or ozone-depleting properties and alternative control strategies are urgently required. We have produced transgenic tobacco (Nicotiana tabacum) plants expressing different dsRNA hairpin structures targeting a root-knot nematode (Meloidogyne javanica) putative transcription factor, MjTis11. We provide evidence that MjTis11 was consistently silenced in nematodes feeding on the roots of transgenic plants. The observed silencing was specific for MjTis11, with other sequence-unrelated genes being unaffected in the nematodes. Those transgenic plants able to induce silencing of MjTis11, also showed the presence of small interfering RNAs. Even though down-regulation of MjTis11 did not result in a lethal phenotype, this study demonstrates the feasibility of silencing root-knot nematode genes by expressing dsRNA in the host plant. Host-delivered RNA interference-triggered (HD-RNAi) silencing of parasite genes provides a novel disease resistance strategy with wide biotechnological applications. The potential of HD-RNAi is not restricted to parasitic nematodes but could be adapted to control other plant-feeding pests.

Hanging drop: An in vitro air toxic exposure model using human lung cells in 2D and 3D structures

Using benzene as a candidate air toxicant and A549 cells as an in vitro cell model, we have developed and validated a hanging drop (HD) air exposure system that mimics an air liquid interface exposure to the lung for periods of 1 h to over 20 days. Dose response curves were highly reproducible for 2D cultures but more variable for 3D cultures. By comparing the HD exposure method with other classically used air exposure systems, we found that the HD exposure method is more sensitive, more reliable and cheaper to run than medium diffusion methods and the CULTEX (R) system. The concentration causing 50% of reduction of cell viability (EC50) for benzene, toluene, p-xylene, m-xylene and o-xylene to A549 cells for 1 h exposure in the HD system were similar to previous in vitro static air exposure. Not only cell viability could be assessed but also sub lethal biological endpoints such as DNA damage and interleukin expressions. An advantage of the HD exposure system is that bioavailability and cell concentrations can be derived from published physicochemical properties using a four compartment mass balance model. The modelled cellular effect concentrations EC50(cell) for 1 h exposure were very similar for benzene, toluene and three xylenes and ranged from 5 to 15 mmol/kg(dry weight) which corresponds to the intracellular concentration of narcotic chemicals in many aquatic species, confirming the high sensitivity of this exposure method. (C) 2013 Elsevier B.V. All rights reserved.