2. Background & History
Bacillus thuringiensis
Bacillus thuringiensis (Bt) is a rod shaped, aerobic, spore-forming bacterium. It is related to other Bacillus species such as B. cereus, the causative agent of some types of food poisoning, and B. anthracis, the causative agent of anthrax. During sporulation Bt produces protein crystals which are often toxic to invertebrates. The species comprises diverse strains with different toxin profiles, and the range of toxins can affect several different types of invertebrates.
Bt was first isolated in 1901 by a Japanese biologist, S. Ishiwata. Ishiwata identified the bacterium as the causal agent of a disease of silkworms. In 1938, commercial production of Bt as a spray for insect control began in France, and the first commercial Bt formulations were made available for field testing in the USA in 1958. Until the 1970s it was generally accepted that lepidopteran insects (moths and butterflies) were the only targets of Bt.
New markets were opened by the discovery in 1976 of the israelensis subspecies, which is toxic to larval mosquitoes and black flies (known as sand flies in New Zealand), and the discovery of the tenebrionis subspecies which is toxic to several beetle species.
In the 1980s, commercial interest in Bt grew as alternatives to synthetic pesticides were sought. The use of Bt toxin genes in genetically modified plants for pest control became an established field of research in the mid-1980s. From the mid-1990s, plants genetically modified to express the Bt toxin have become increasingly common, and are now grown widely in the USA and other countries, though their use remains controversial.
Insecticide resistance & resistance management
Until the 1950s, relatively few people considered resistance to pesticides to be a serious threat to pest management. Although some insects had evolved resistance to DDT, the prevailing feeling was that resistance could be overcome by using ever newer pesticides. However, in the 1970s, it became clear that at least some pests were evolving resistance faster than new and environmentally acceptable pesticides could be developed and brought to market. Indeed, two of the first three genetically modified Bt crops registered in the USA, cotton and potatoes, were targeted at markets essentially created by the recurrent evolution of resistance to insecticides in certain pests (Roush, 1997).
The concept of resistance management began to evolve in the late 1950s. The aim of resistance management programmes is to "slow the evolution of resistance and thereby extend the useful life of valuable toxicants" (Roush, 1997). The historical lesson from insecticide resistance that is relevant to genetically modified crops is that resistance can most effectively be delayed by a combination of molecular biology, population genetics and careful management practices in the field.
Various Bt toxins (Cry proteins) can be inserted into target crops and this selection should minimise exposure of target pests to similar Cry proteins, for example by avoiding the use of the same Cry protein in sprays and genetically modified crops. Similarly, the use of the same Cry protein in multiple crops visited by the same pest species will favour the development of resistance and should also be avoided. Strategies for the application of these techniques in New Zealand have been proposed by Wigley et al. (1994) and Wearing and Hokkanen (1994), and are summarised by Madhusudhan et al. (2000).
Once genetically modified plants are deployed, there are a number of ways to slow selection in favour of resistance. Expert opinion around the world has mainly supported the use of a high dose / refuge approach where the genetically modified plants produce as much toxin as possible and refuges are provided for insects with genes for susceptibility. This strategy is currently used in commercial production in several countries.
High dose/refuge approach
In the pest control context, the word "refuge" is used to mean an area of habitat where susceptible pests can survive. For example, if a cabbage-eating caterpillar is susceptible to chemical X, a refuge for that caterpillar could be an area of cabbages that was not sprayed with chemical X. The refuge must contain host plants that make it possible for the susceptible insects to breed without coming into contact with chemical X and being killed. The host plants do not necessarily need to be of the same species as the crop that is being protected.
Allowing susceptible insects to survive in a refuge reduces the rate at which the entire population is likely to develop resistance to the pesticide (see Figure 1).
Refuges are aimed at maintaining susceptible populations in numbers that will sufficiently dilute any resistance that arises in the target populations. The approach assumes that mating will be random between insects living in the refuges and those in the crop being sprayed or the genetically modified crop. Growers of genetically modified plants are usually required to provide their own share of refuges, both for equity reasons and to keep refuge sites close to the genetically modified plants.
The amount and size of refuges that are necessary will differ depending on the mobility and ecology of the insect, the type of crop and geographical area. The necessary size of the refuge can also vary depending on whether the refuge is sprayed with any chemical control, and on the biology of the pest and the crop. If unsprayed, higher numbers of susceptible pests could be expected to build up, meaning that the refuge area can be smaller. Inclusion of a refuge may reduce the proportion of marketable produce for some crops, which will affect growers’ desire to use this strategy.
The ability of a pest species to move, its breeding biology, the crop biology and the pest’s feeding habits will all have impacts on the size and type of refuges that are necessary to preserve the effectiveness of Bt. For example, if a pest species feeds on clover but also on broccoli, and a Bt broccoli crop is introduced, the refuge requirement may simply be to plant the broccoli in fields that are close enough to clover pasture to allow a population of non-resistant pests to be maintained. However, if a pest species is highly specific to one plant (crop) type, the refuge requirement may need to be for an unsprayed refuge of the same crop. The potential impacts of alternate hosts, adult and larval movement, and refuge position are currently being investigated in model systems using brassicas and potatoes (Cameron et al. 2002, Madhusudhan et al. 2000).
Figure 1: Basic theory of refuges and resistance for a genetically modified Bt crop
Where the damaging stage of a pest comprises only the larvae, and they do not move from plant to plant (e.g. some stem boring insects), then seeds for crops can be pre-mixed to contain both conventional seeds and genetically modified Bt seeds. This ensures that every crop effectively contains a refuge. Interplanting of susceptible and resistant plant lines is a variation on the same idea. Evidence that some insects move away from plants expressing pesticides to conventional plants may reduce the value of interplanting or seed mixtures. This suggestion requires further research.
The success of the high dose strategy depends on resistance being a rare and recessive trait. Insects with resistance genes may have varying degrees of resistance depending on whether they carry one (RS in Figure 1) or two copies of the gene for resistance (RR). Controlling the dose is important. The dose of toxin should be sufficient to kill all homozygous susceptible individuals (SS) and all heterozygotes with genes for both resistance and susceptibility (RS). It is generally not feasible to deliver a high enough dose of toxin to kill individuals homozygous for resistance (RR). If the dose is allowed to degrade, heterozygotes may survive and increase the frequency of the resistance gene.
Current state of knowledge about Bt resistance
The most compelling evidence that Bt resistance can evolve comes from field populations of the diamondback moth (Plutella xylostella) where resistance has developed after frequent exposure to Bt sprays. This moth is a major pest of vegetables around the world, and receives frequent exposure to pesticides. This exposure and the moth’s biology have contributed to its extensive resistance to most insecticides in many growing areas. Laboratory selection in other species has shown that broad cross-resistance to many Bt toxins is possible. Four other lepidopterans in addition to diamondback moth have been suggested as showing resistance or potential for resistance to Bt under field selection (Glare and O’Callaghan, 2000).
Observations of diamondback moth have shown that resistance to Bt spray can evolve in the field in less than two years with sufficient selection pressure. The lack of field resistance to date in insects other than diamondback moth has been attributed to the relatively small amount of Bt currently applied in comparison to other pesticides.
With the increasing use of Bt genes in genetically modified plants, there is concern that resistance will develop more quickly and many strategies are under discussion to prevent this. After six years of commercialisation in the USA, no reported insect resistance has developed in response to the Bt toxins incorporated in Bt corn, Bt cotton, or Bt potato crops. A summary of research on the environmental impacts of Bt cotton in China claimed that insects would develop resistance to Bt cotton after 8-10 years of continuous planting (Xue 2002). However, one of the scientists whose work was quoted in the summary subsequently said that the research was incorrectly summarised and that no resistance was apparent after 4 years’ study of field populations (Wu 2002).
Scientists believe that it is too soon to conclude whether current resistance management strategies for Bt crops have been successful in preventing resistance in the long term. More information is still required on mechanisms of resistance, inheritance, pest behaviour and other aspects before a complete management plan can be developed for pest insects (Glare and O’Callaghan, 2000).
Contact for Enquiries
Dr Sharon Adamson
Manager,
Innovation Policy
Ministry of Agriculture and Forestry
PO Box 2526
Wellington
NEW ZEALAND
Phone: +64 4 894 0618
Fax: +64 4 4 894 0741
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