Contents - Faperta

496 Biotechnological Approaches for Pest Management and Ecological Sustainability
and A. gambiae are in the same order, only a small number of the members of the three
gene families have been identifi ed as orthologs between the two species (Ranson et al.,
2002). Most clades of genes within each of the three families were represented in both
species, but the origins of most genes within each clade were explained by independent
duplication events within each species. Therefore, fi nding similar mutations in orthologous
genes may be an exception rather than the rule.
Microarrays and Regulatory Mutations in Cytochrome P450s
Mutation in a P450 gene leading to insecticide resistance has been elucidated at the molecular
level (Daborn et al., 2002). Expression profi ling with microarrays has shown that high
levels of DDT resistance in strains of D. melanogaster is due to 100-fold upregulation of a
specifi c P450 enzyme (Cyp6g1), owing to insertion of a transposable element into its promoter.
Expression profi ling identifi ed a specifi c causal change in a specifi c member of a
large gene family without any prior knowledge or assumption regarding the identity of that
gene within the family. Several enzymes that metabolize pesticides but do not belong to the
three major detoxifi cation gene families (P450s, carboxylesterases, and GSTs) have been
found in soil bacteria, and several of these have homologs of unknown function in insects
(Claudianos et al., 2002), suggesting that additional resistance mechanisms may be discovered
as the power of genomic technologies is applied to understand the resistance mechanisms.
Proteomics along with positional cloning using quantitative trait loci (QTLs) will be
quite useful for this purpose. There may be some mechanisms that will be diffi cult to
resolve even with genomic technologies, for example, upregulation mediated by changes to
trans-acting factors, a mechanism that appears to underlie some cases of resistance involving
P450s, carboxylesterases, and GSTs (Feyereisen, 1999; Hemingway, 2000).
Quantitative Trait Loci, Positional Cloning, and Multiple Resistance to Bt Toxins
The Bt toxins differ from most other insecticides in that they are proteins and are not
neurotoxins. In fact, their mode of action is quite complex and not properly understood,
involving binding to sites on at least four different protein and carbohydrate targets in the
insect midgut (Tabashnik et al., 1997; Marroquin et al., 2000; Ferre and van Rie, 2002).
Resistance to Bt toxins has become a critical concern with the expression of Bt toxins in
transgenic crops (Ferre and van Rie, 2002). Resistance has already been reported in natural
populations of the diamondback moth, Plutella xylostella (L.), and it has been quite easy
to select for resistance in laboratory populations of several insect species. Several Bt toxin
resistance genes have been reported from P. xylostella, most of which probably encode the
proteins that act as toxin-binding sites. Similarly, most of the laboratory-selected strains
of insects with resistance to Bt toxins involve multiple genes (Ferre and van Rie, 2002).
Some individual genes underlying Bt toxin resistance have now been mapped onto highdensity
linkage maps using QTL mapping (Heckel et al., 1999; Marroquin et al., 2000).
Two laboratory-selected Bt toxin resistance genes have been identifi ed using genomic
technologies. The fi rst involved positional cloning in Heliothis virescens (F.) (Gahan, Gould,
and Heckel, 2001). The coding region of this gene was apparently disrupted in resistant
individuals by insertion of a transposable element. The mutant target site appears to be
insensitive to Bt, although the native function of the target molecule has probably been lost,
leading to a signifi cant fi tness penalty in the absence of insecticide—a phenomenon that is
also characteristic of the resistant P. xylostella found in the fi eld (Tabashnik et al., 1997).