Chapter 11 Resistance to Plant Allelochemicals in Heliothis virescens (Fabricius) 1
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Randy L. Rose , Fred Gould , Patricia Levi , Takimichi Konno , and Ernest Hodgson 1
Downloaded by UNIV OF ARIZONA on August 2, 2012 | http://pubs.acs.org Publication Date: September 22, 1992 | doi: 10.1021/bk-1992-0505.ch011
1Department of Toxicology and2Departmentof Entomology, North Carolina State University, Raleigh, NC 27695 Potential metabolic routes for the resistance of tobacco budworm (TBW) larvae to host plant allelochemicals nicotine, 2-tridecanone and quercetin were explored. Midgut preparations from larvae resistant to nicotine and 2-tridecanone had elevated levels of cytochrome P450 which were associated with significant increases in metabolism for five of six monooxygenase substrates. In quercetin tolerant larvae, metabolism of two monooxygenase substrates was significantly enhanced although no increase in P450 content was observed. Glutathione transferases and esterases did not appear to be involved in the resistance of any of the strains examined. Patterns of substrate oxidations varied between strains and inducing agents, suggesting that different isozymes of P450 are associated with resistance and induction.
The tobacco budworm (TBW), Heliothis virescens (Fabricius), is a polyphagous insect which has been observed feeding on 31 different plant species in as many as 14 plant families (7). These various host plants produce fitness reducing and/or antifeedant chemicals as defenses (2). Some of the toxic allelochemicals encountered by the TBW in its natural host plants include gossypol and related terpenoids, condensed tannins, quercetin, rutin, anthocyanin, nicotine and flavonoids (5). Mechanisms for Tolerance to Allelochemicals and Pesticides At least three different mechanisms for dealing with host plant allelochemicals have been demonstrated in TBW. Nicotine tolerance was postulated to be the result of an efficient excretory system, like that previously demonstrated in the tobacco hornworm (4). This idea was supported by the lack of discernible 3
Current address: Nihon Nohyaku Company, Ltd., Osaka, Japan
0097-6156/92/0505-0137S06.00/0 © 1992 American Chemical Society
In Molecular Mechanisms of Insecticide Resistance; Mullin, C., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1992.
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MOLECULAR MECHANISMS OF INSECTICIDE RESISTANCE
metabolites in TBW larvae and their feces ten hours after treatment with 200 ug free base nicotine (5). Behavioral adaptations have also been observed in TBW. Young larvae feeding in cotton avoid gossypol-producing glands (6). As they become older and less susceptible to gossypol (7, 8), presumably as a result of allelochemically-based induction of detoxifying enzymes (9, 70), non-selective consumption of gossypol-containing glands occurs (77). Recent studies suggest that the tobacco budworm utilizes its inducible metabolic system for the detoxication of a variety of allelochemicals (9, 70, 72, 13). Studies of enzyme induction by host plants and/or allelochemicals derived from host plants demonstrate that induced insects can detoxify pesticides faster than non-induced insects (see ref. 14 for review). For example, sixth instar larvae of the variegated cutworm, Peridroma saucia (Hubner), had greater tolerance to acephate, methomyl and malathion when reared on peppermint leaves versus bean leaves. The increase in tolerance was associated with a significant increase in monooxygenase activity (75). Host plant induction of enzyme systems other than monooxygenases, such as glutathione S-transferases, are also associated with insecticide tolerance (16,17). Results of these studies suggest that changes in the chemistry of the host plants consumed by a particular insect species (e.g. after the introduction of a resistant host genotype) may influence the susceptibility of the consumer to insecticides. Hence, tolerance of host plant allelochemicals and insecticides are related. Relatively few insects with intraspecific genetic variation with respect to allelochemically-based resistance are in culture. Yet, an understanding of such genetically based resistance is necessary in order to predict which pesticides might best be utilized in combination with host plant resistance. In addition, little information is available concerning mechanisms of genetic resistance to allelochemicals. After over 30 generations of laboratory selection, several strains of tobacco budworm have been developed with resistance to allelochemicals (18, Gould, unpublished data). For our studies, we selected strains possessing heritable resistance to nicotine (WAKE-N), 2-tridecanone (WAKE-T), and quercetin (BCQ). The control strains from which the resistant strains were selected included NC-1 (from which BC-Q was selected) and WAKE (from which WAKE-T and WAKE-N were selected). These studies utilized midgut homogenates from larvae reared on diets in the absence of the selecting agent for the preparation of microsomes (100,000 g pellet) and cytosolic (100,000 g supernatants) fractions. Substrates utilized for esterase and glutathione transferase enzymes included a-naphthyl acetate and 1chloro, 2,4-dintrobenzene, respectively. For monooxygenase determinations, several substrates were selected in an effort to represent a variety of metabolic possibilities. These included £-nitroanisole and methoxyresorufin (0demethylation), benzphetamine (N-demethylation), benzo(a)pyrene (aryl hydroxylation), lauric acid (alkyl hydroxylation), and phorate (sulfoxidation). Resistance Development to Pesticides vs Allelochemicals Prior to the introduction of the pyrethroids, the tobacco budworm had developed resistance to nearly every insecticide used against it, including DDT, carbaryl,
In Molecular Mechanisms of Insecticide Resistance; Mullin, C., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1992.
Downloaded by UNIV OF ARIZONA on August 2, 2012 | http://pubs.acs.org Publication Date: September 22, 1992 | doi: 10.1021/bk-1992-0505.ch011
11. ROSE ET AL.
Resistance in Heliothis virescens
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endrin, parathion, EPN, and monocrotophos (79). Tobacco budworm resistance to pyrethroids was first diagnosed in populations collected from western Texas where field control failures had occurred (20). Since that time, pyrethroid resistance has been documented in several southeastern states (20-25) however, pyrethroid management plans adopted in these areas have likely curtailed the development of extremely high resistance levels (24). The likelihood that pyrethroid resistance would eventually develop in TBW was suggested by a laboratory study in which selection pressure was exerted at a level near 80% mortality. At this rate, significant levels of resistance (37-fold) were attained within 11 generations (25). By 36 generations, this strain had attained resistance levels of > 1000-fold (26). Although pyrethroid resistance in field populations have not yet approached either of these levels, with intense population pressure the possibility for such dramatic increases does exist (27). Contrasting with the high resistance levels attained by selection with synthetic insecticides, selection of TBW larvae with various allelochemicals has not resulted in high levels of resistance. After more than 30 generations of selection pressure for nicotine and 2-tridecanone resistance using dietary concentrations resulting in acute toxicity for up to 80% of the population, resistance levels at the L C do not exceed 2.5-fold (Gould, unpublished data). Selection for tolerance to quercetin was based upon the ability to grow on quercetin containing diets, rather than on mortality. Concentrations of 0.5% quercetin were non-lethal, but severely limited growth of non-adapted larvae (Table I). Larvae selected for tolerance to quercetin, however, attain normal body weight within the sametimeperiod as larvae reared in absence of 0.5% quercetin. 50
Resistance to 2-Tridecanone. Dimock and Kennedy (28) demonstrated that first instar cotton bollworm, H. zea, placed on leaves of accession PI 134417 of the wild tomato, Lycopersicon hirsutum f glabratum, C.H. Mull, were quickly paralyzed, even in absence of foliar consumption. This paralytic response was postulated to be due to the fumigant action of 2-tridecanone emanating from glandular trichomes of the tomato leaf. The quick recovery of exposed larvae suggested involvement of an inducible detoxication system. Exposure of neonates (29) and/or eggs (30) to 2-tridecanone resulted in greater tolerance to subsequent exposures of both 2-tridecanone and the insecticide carbaryl. Table I. Effects of Quercetin on the Larval Growth of Tobacco Budworm" Body Weight (mg)* Diet Regular + Quercetin
Control 252.6 + 62.9 17.4+ 5.3*
BC-Q 222.8 + 71.3 232.6 + 75.2**
'Body weights taken 10 days following inoculation of first instar larvae. Mean + standard deviation for 20 larvae. *p
