Immunolocalization and ultrastructure shows ingestion of Cry protein expressed in Glycine max by Heterodera glycines and its mode-of-action.

Great interest exists in developing a transgenic trait that controls the economically important soybean (Glycine max) pest, soybean cyst nematode (SCN, Heterodera glycines), due to its adaptation to native resistance. Soybean plants expressing the Bacillus thuringiensis delta-endotoxin, Cry14Ab, were recently demonstrated to control SCN in both growth chamber and field testing. In that communication, ingestion of the Cry14Ab toxin by SCN second stage juveniles (J2) was demonstrated using fluorescently labeled Cry14Ab in an in vitro assay. Here, we show that consistent with expectations for a Cry toxin, Cry14Ab has a mode-of-action unique from the native resistance sources Peking and PI 88788. Further, we demonstrate in planta the ingestion and localization of the Cry14Ab toxin in the midgut of nematodes feeding on roots expressing Cry14Ab, using immunogold labeling and transmission electron microscopy. We observed immunolocalization of the toxin and resulting intestinal damage primarily in the microvillus-like (MvL)-containing region of the midgut intestine, but not in nematodes feeding on roots lacking toxin. This demonstrated that Cry14Ab was taken up by the J2 SCN, presumably through the feeding tube within the plant root cell that serves as its feeding site. This suggests that relatively large proteins can be taken up through the feeding tube. Electron microscopy showed that Cry14Ab caused lysis of the midgut MvL membrane, and eventual degradation of the MvL and the lysate, forming particulate aggregates. The accumulated electron dense aggregate in the posterior midgut intestine was not observed in SCN in non-Cry14Ab expressing plants.

Development and Validation of a Resistance Management Model for the Soybean Cyst Nematode, Heterodera glycines.

Plant-parasitic nematodes are a key yield limiting pest of crops around the world. Deployment of plant resistance genes are an important management tactic for many economically important plant-parasitic nematodes. The selection for virulence in nematode populations is a major threat to the effectiveness of resistance gene-based management. Little research has gone into resistance management modelling despite the importance of both plant-parasitic nematodes and resistance genes for their management. In this paper we report on a cyst nematode resistance management model created to explore the factors which are most important for determining the durability of resistance genes to this important family of plant-parasitic nematodes. The relative dominance of virulence expression, the level of inbreeding, and the number of generations per cropping season were the most important factors in predicting resistance gene durability. Aspects of cyst nematode biology that reduce the number of generations per season for a portion of the population had a much smaller effect on the durability of resistance genes. These factors included delayed hatching within a season and early dormancy. The accuracy and utility of the model was tested using the soybean cyst nematode (SCN) rhg1-mediated resistance system. The model accurately predicted the rate at which virulence to the rhg1b resistance gene developed in Iowa over a two-decade period. The model suggested resistance gene pyramids as the most durable management solution for SCN with multiple possible avenues to obtain acceptable efficacy and durability.

Soybean Cyst Nematode Management Is Improved by Combining Native and Transgenic Resistance.

Field trials were conducted to assess the benefit of combining a transgenic soybean cyst nematode (SCN) resistance trait, Cry14Ab-1 expressed by the event GMB151, with the native resistance allele rhg1b from PI 88788. The GMB151 event and rhg1b were crossed into common genetic backgrounds and segregated out to create four genetically related lines within each background. The lines created contained both native and transgenic resistance (rhg1b + GMB151), only native resistance (rhg1b alone), only transgenic resistance (GMB151 alone), or neither resistance type (susceptible). The benefit of GMB151 and rhg1b for SCN management was evaluated by measuring SCN control and yield protection. Soybean cyst nematode control was assessed by counting the number of females and cysts on roots early in the season and measuring the change in SCN egg population density over the entire season. The GMB151 transgenic event and the native resistance allele rhg1b both reduced early season SCN reproduction and contributed to significantly higher soybean yield. Compared to susceptible lines, the rhg1b allele improved yield by 33%, while GMB151 improved yield by 13%. Combining the GMB151 event and rhg1b allele resulted in greater SCN control and yield improvement than either provided alone. The combination of GMB151 and rhg1b reduced season-long SCN reproduction by 50% and resulted in 44% greater yield than the susceptible lines. Soybean cyst nematode virulence to rhg1b continues to increase due to the continuous planting of PI 88788-derived resistant cultivars. Pyramiding GMB151 with rhg1b provides a new management option to improve SCN control and soybean yield.

A Bacillus thuringiensis Cry protein controls soybean cyst nematode in transgenic soybean plants.

Plant-parasitic nematodes (PPNs) are economically important pests of agricultural crops, and soybean cyst nematode (SCN) in particular is responsible for a large amount of damage to soybean. The need for new solutions for controlling SCN is becoming increasingly urgent, due to the slow decline in effectiveness of the widely used native soybean resistance derived from genetic line PI 88788. Thus, developing transgenic traits for controlling SCN is of great interest. Here, we report a Bacillus thuringiensis delta-endotoxin, Cry14Ab, that controls SCN in transgenic soybean. Experiments in C. elegans suggest the mechanism by which the protein controls nematodes involves damaging the intestine, similar to the mechanism of Cry proteins used to control insects. Plants expressing Cry14Ab show a significant reduction in cyst numbers compared to control plants 30 days after infestation. Field trials also show a reduction in SCN egg counts compared with control plants, demonstrating that this protein has excellent potential to control PPNs in soybean.