Pest control and resistance management through release of insects carrying a male-selecting transgene

Abstract

Background: Development and evaluation of new insect pest management tools is critical for overcoming over-reliance upon, and growing resistance to, synthetic, biological and plant-expressed insecticides. For transgenic crops expressing insecticidal proteins from the bacterium Bacillus thuringiensis ('Bt crops') emergence of resistance is slowed by maintaining a proportion of the crop as non-Bt varieties, which produce pest insects unselected for resistance. While this strategy has been largely successful, multiple cases of Bt resistance have now been reported. One new approach to pest management is the use of genetically engineered insects to suppress populations of their own species. Models suggest that released insects carrying male-selecting (MS) transgenes would be effective agents of direct, species-specific pest management by preventing survival of female progeny, and simultaneously provide an alternative insecticide resistance management strategy by introgression of susceptibility alleles into target populations. We developed a MS strain of the diamondback moth, Plutella xylostella, a serious global pest of crucifers. MS-strain larvae are reared as normal with dietary tetracycline, but, when reared without tetracycline or on host plants, only males will survive to adulthood. We used this strain in glasshouse-cages to study the effect of MS male P. xylostella releases on target pest population size and spread of Bt resistance in these populations.

Results: Introductions of MS-engineered P. xylostella males into wild-type populations led to rapid pest population decline, and then elimination. In separate experiments on broccoli plants, relatively low-level releases of MS males in combination with broccoli expressing Cry1Ac (Bt broccoli) suppressed population growth and delayed the spread of Bt resistance. Higher rates of MS male releases in the absence of Bt broccoli were also able to suppress P. xylostella populations, whereas either low-level MS male releases or Bt broccoli alone did not.

Conclusions: These results support theoretical modeling, indicating that MS-engineered insects can provide a powerful pest population suppressing effect, and could effectively augment current Bt resistance management strategies. We conclude that, subject to field confirmation, MS insects offer an effective and versatile control option against P. xylostella and potentially other pests, and may reduce reliance on and protect insecticide-based approaches, including Bt crops.

Fig. 1

Suppression of caged populations of Plutella xylostella by weekly introduction of OX4319L males. Graphs showing (a) number of eggs collected, (b) number of dead adult females collected weekly from cages, and (c) proportion of cage progeny that were transgenic re-entering the cages (fluorescence proportions), over the experimental period. With female moths present in the cages being only wild-type, transgenic progeny (including those re-introduced) were heterozygotes (restrictive conditions). Fluorescence proportions thus equate to twice the MS transgene allele frequency in the cage population at that time point. Solid lines represent OX4319L-treated populations (Cages 1 and 2, circular and square data-points, respectively). Dashed lines represent untreated control populations (Cages 3 and 4, circular and square data-points, respectively). In week 9, return of pupae into treatment and control cages was made proportional and release of OX4319L males into treatment cages began (marked with asterisk)

Fig. 2

Effects of Bt broccoli and OX4319L releases on caged Plutella xylostella populations over multiple generations. Caged populations were established with hybrid Bt-resistant/wild-type insects – the founder strain – with a low estimated frequency of homozygous-resistant individuals. a Graph shows the mean peak population size per plant, per generation, in four experimental treatments over the experimental period: Treatment 1, Bt broccoli, no OX4319L releases; Treatment 2, Bt broccoli, low-rate weekly OX4319L releases (release rate of 3:1 in Generation 1, increased to 5:1 in subsequent generations); Treatment 3, non-Bt broccoli, low-rate weekly OX4319L releases (identical release rates to Treatment 2); and Treatment 4, non-Bt broccoli, high-rate weekly OX4319L releases (release rate of 20:1 in Generation 1, increased to 40:1 in subsequent generations). Means were calculated from three experimental cage replicates, with the exception of Treatments 2 and 4, which were reduced to two and one cage replicates in Generation 3, respectively. Treatment 3 cages were terminated in Generation 3 as the insect populations had reached maximum capacity. Error bars represent standard error of the mean. b Bt survival assays. Mean survival of third-instar larvae from three experimental cage treatments and the founder strain used to begin these experimental treatments when exposed to a discriminating dose of Bt in artificial diet assays. Bt dose in this assay is high enough to ensure that only homozygous Bt-resistant individuals will survive (as in the high-dose/refuge strategy). This assay therefore indicates the proportion of each population remaining Bt-resistant (homozygous) and Bt-susceptible (heterozygous or homozygous-susceptible). For each cage, two Bt assays and one no-Bt control assay were performed. Bt assays in each cage were summed and means represent averages of each set of treatment cages corrected for control mortality. The assays took place using individuals from the final generation in which each treatment was run or, in the case of the founder strain, in the generation prior to the start of the experiment. Survival was corrected for control mortality prior to analysis and error bars represent Pearson’s exact confidence intervals. Survival of insects from the low OX4319L release cages (Treatment 2) was not significantly different from the founder strain; other pairwise comparisons are significantly different (Table 1)

Fig. 3

Schematic showing design of the population suppression and insecticide resistance management experiments