Could Bt transgenic crops have nutritionally favourable effects on resistant insects? Ecology Letters v.4, i.5 May03

IDEAS AND PERSPECTIVES 

Ali H. Sayyed 1, Hugo Cerda 1,2* and Denis J. Wright1

1 Department of Biological Sciences, Imperial College London, Silwood Park campus Ascot SL5 7PY, UK 2 Universidad Simon Rodriguez, IDECYT, Apartado Postal 47.925, Caracas, Venezuela *Correspondence: E-mail: h.cerda@ic.ac.uk

Abstract
We present an idea that larvae of some Bacillus thuringiensis (Bt ) resistant populations of the diamondback moth, Plutella xylostella (L.), may be able to use Cry1Ac toxin derived from Bt as a supplementary food protein. Bt transgenic crops could therefore have unanticipated nutritionally favourable effects, increasing the fitness of resistant populations. This idea is discussed in the context of the evolution of resistance to Bt transgenic crops.

Keywords Bacillus thuringiensis, fitness, insect nutrition, resistance, transgenic. Ecology Letters (2003) 6: 167-169

Bacillus thuringiensis (Bt) is a Gram-positive soil bacterium distinguished from other bacilli by its production of parasporal crystal protein endotoxins (Yamamoto & Powell 1993). By the action of midgut proteases, ingested crystal (Cry) protoxins are processed into active, toxic fragments (c. 55-70 kDa) (Schnepf et al. 1998), which then bind to toxin-specific receptors on the midgut membrane.

The planting of Bt transgenic crops increased from 1.7 million ha in 1996 to 44.2 million ha in 2000 and is expected to increase further to give a global market of US$25 billion in 2010 (Shelton et al. 2002). With the widespread use of Bt crops, many more insect species are subjected to selection pressure for resistance. In the majority of the studies to date, Bt resistance appears to be related to loss of membrane binding by Cry toxins (Ferre´ & Van Rie 2002). However, there are several other possible mechanisms by which resistance could occur and a protease-mediated resistance mechanism has been reported in laboratory-selected populations of the Indian meal moth, Plodia interpunctella and the tobacco budworm, Heliothis virescens (Oppert et al. 1997; Forcada et al. 1999).

Fitness costs associated with resistance alleles are common.

Studies with some strains of Plutella xylostella (Groeters et al. 1994; Tang et al. 1997) and other lepidopteran species (Liu et al. 1999; Alyokhin & Ferro 1999; Carriere et al. 2001) have shown strong fitness costs associated with Bt resistance. In contrast, resistance to Bt in two populations of the diamond-back moth, P. xylostella strains from Malaysia (SERD4 and SERD5) had no apparent pleiotropic effects on development time, pupal weight, fecundity or the intrinsic rate of population increase (Sayyed & Wright, 2001).

The evolution of resistance to transgenic insecticidal crops involves the interaction of many genetic and biological factors (Tabashnik 1994), including fitness costs. Here we look at two possible fitness costs, the larval development time and fresh pupal weight, of a highly Cry1Ac-resistant re-selected sub-population of P. xylostella SERD4, an unselected (UNSEL) sub-population of SERD4 and a susceptible ROTH popula-tion of this species. All populations were reared in the presence or absence of Bt toxin Cry1Ac. The aim was to conduct a preliminary experiment to test the idea that such a toxin could have nutritionally favourable effects on insects that, because of their resistance, could consume large amounts of Cry protein with impunity.

Batches of five randomly-selected third-instar larvae from the re-selected, UNSEL and ROTH populations were weighed, placed on leaf discs treated with Cry1Ac or on control leaves (Sayyed et al. 2000) and their developmental time to pupation recorded (Sayyed & Wright, 2001). The concentrations of Cry1Ac used, 0.0001, 0.1 and 10 lg ml )1 were chosen according to the LC50 values for Cry1Ac against the ROTH (0.001 lg ml )1), UNSEL (1.6 lg ml )1) and re-selected (209 lg ml )1) populations, respectively. The leaf discs were then transferred to individual plastic Petri dishes (9 cm diameter) containing a single moistened Whatman No 1 filter paper and maintained at 20 C. Each treatment was replicated 10 times. The leaf discs were replaced ad libitum and pupation was recorded daily. Each batch of five larvae and pupae were weighed together and each batch was treated as a separate replicate. An ANCOVA in GLIM (GLIM 3.77, Numerical Algorithms Group, Oxford, UK; Crawley, 1993) was used to compare the average weight gained by pupae.

The observed mortality was 4, 6 and 0% for ROTH, UNSEL and re-selected populations, respectively. Larvae from the re-selected population reached a significantly greater mean pupal weight (t .2.20; d.f. 18; P < 0.05) on Cry1Ac-treated cabbage leaf discs compared with untreated leaves (Table 1). The development time from egg hatch to eclosion was also 1-2 days shorter on Cry1Ac-treated leaf discs compared with treated leaf discs, resulting in a 56% greater growth rate (pupal weight days )1 to pupal stage)

(Table 1). There was no significant (t .1.21, t .0.20; d.f.18; P > 0.05) difference on pupal weight or developmental time on Cry1Ac-treated cabbage leaf discs compared with untreated leaves for ROTH and UNSEL, which were treated with much lower doses of toxin compared with the re-selected population.

The present results and previous work on re-selected SERD4 populations (Sayyed & Wright 2001) suggest that resistant larvae may be using Cry1Ac as a supplementary food protein, and that this may account for the observed faster development rate of Bt resistant insects in the presence of the Bt toxin. This could either be a pleiotropic effect linked to Bt resistance or, more simply, because of resistant insects exploiting their ability to survive high doses of the toxin. In both cases, the insects are able to digest and utilize the Cry protein. Plant insect nutrition is the result of a complex balance between beneficial and toxic components in the plant. The presence of Cry1Ac toxin could have modified the nutritional balance in the plants for resistant larvae.

The suggestion that an insect toxin can support the growth of an insect is not unique. The tree locust, Anacridium melanohodon uses the phenol gallic acid in sclerotization of the cuticle (Panzuto et al. 2002). Dauphin-ais (1998) observed that tannic acid-reared larvae of the oblique-banded leaf roller, Choristoneura rosaceana developed faster compared with control insects. Because of the potential significance of Bt resistant insects being able to utlize Bt toxins as food, ecological and biochemical studies are required to establish the nutritional effects of Cry1Ac on resistant populations of P. xylostella.

ACKNOWLEDGMENTS

HC was supported by PhD fellowship from FONACIT Venezuela.

REFERENCES

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Oppert, B., Kramer, K.J., Beeman, R.W., Johnson, D. & McGaughey, W.H. (1997). Proteinase-mediated insect resistance to Bacillus thuringiensis toxins. J. Biochem. 272, 23473-23476. ... Table 1 Larval development time and fresh pupal weight of Plutella xylostella SERD4 and UNSEL populations reared in the presence or absence of Cry1Ac toxin

Strain Cry1Ac concentration (lg ml )1) Mortality (%) Pupal weight (mg) Mean ± SE Development time (days) Mean ± SE Pupal weight/ development time +Cry1Ac )Cry1Ac +Cry1Ac )Cry1Ac +Cry1Ac )Cry1Ac ROTH 0.0001 4 7.93 ± 0.27 a 7.8 ± 0.3 a 22 ± 0.84 a 23 ± 0.9 a 0.36 0.34 UNSEL * 0.1 6 6.10 ± 0.2 a 5.63 ± 0.19 a 21 ± 0.53 a 22 ± 0.79 a 0.29 0.26 SERD4

10 0 10.89 ± 0.37 a 8.89 ± 0.15 b 19 ± 0.73 a 20 ± 0.90 a 0.57 0.44

Within rows, figures sharing a common letter are not significantly different (P > 0.05); n.10.

*Unselected SERD4 population from Malaysia (LC50 .1.61 lg Cry1Ac ml )1 ). SERD4 selected with Cry1Ac to give high level of resistance (LC50 .209 lg Cry1Ac ml )1). ROTH (LC50 .0.001 lg Cry1Ac ml )1) n .number of replicates per treatment (five insects per replicate). 168 A. H. Sayyed, H. Cerda and D. J. Wright 2003 Blackwell Publishing Ltd/CNRS