E. P. Lampert

Aphid carboxylesterases: Biochemical aspects and importance in the diagnosis of insecticide resistance☆

Abstract The activity of general carboxylesterases from susceptible and malathion-resistant strans of the green peach aphid, Myzus persicae , and the tobacco aphid, M. nicotianae , was examined using 10 1-naphtholic esters. The activity showed a parabolic relationship with the number of carbon atoms in the substrate acyl moiety. The maximal activity was obtained with 1-naphthyl propionate and 1-naphthyl butyrate, respectively, for the above species. No interstrain differences in the general profile of the structure-activity relationship were observed. However, an elevated carboxylesterase activity in the resistant strains when compared to the reference-susceptible strain was observed with each of the tested substrates, and the bestowed activity correlated with the degree of malathion resistance. The zymograms and intensity of the carboxylesterase bands when resolved by wide range isoelectric focusing (pH 3.5–9.5) and stained with Fast Blue B salt using 1-naphthyl acetate, 1-naphthyl propionate, and 1-naphthyl caproate as substrates correlated favorably with the general activity. From the above results a filter paper esterase test was developed and used for the diagnosis of the level of resistance in individual aphids. This test showed an excellent agreement with the susceptibility bioassay test of malathion against the green peach aphid. However, the test did not make possible an unambiguous discrimination between susceptible and resistant tobacco aphids. The possible impact of this and other more specific tests on the management of aphid populations is discussed.

Diagnostic esterases and insecticide resistance in the tobacco aphid, Myzus nicotianae Blackman (Homoptera: Aphididae)

Abstract Insecticide resistance in the tobacco aphid, Myzus nicotianae Blackman, from different localities in the southeastern United States has been found to exist mainly in aphids with red body coloration. Frequency distribution of esterase activity in individual tobacco aphids collected from a tobacco field and a greenhouse indicated that organophosphate resistance was always linked to high carboxylesterase activity toward 1-naphthyl acetate. Two approaches were taken to speed up the detection methodology of the frequency of resistant phenotypes. First, an abbreviated bioassay test, using malathion as a reporter molecule, was developed to rapidly distinguish between OP-resistant and susceptible phenotypes of the tobacco aphid. A time threshold of 50 min following aphid exposure to 50 ppm malathion in a water suspension accurately discriminated between susceptible and resistant phenotypes. Second, assays of aphid carboxylesterases, using the acetates and propionates of 1-naphthol, 2-naphthol, and 4-nitrophenol, were performed to optimize and better understand the esterase discriminating activity between resistant and susceptible phenotypes. Only activity toward 1-naphtholic esters unambiguously discriminated between susceptible and resistant aphids. In general, resistance to malathion appeared to be esterase-mediated and some electrofocusing-detectable esterase isozymes were associated with resistance.

Effect of tobacco etch virus on the seasonal growth of flue-cured tobacco

Abstract The effect of tobacco etch virus (TEV) infection on flue-cured tobacco (Nicotiana tabacum L.) plant growth was invisaged under field conditions in 1985 and 1986. Natural field infection of TEV was allowed to develop a patchy distribution of healthy plants and plants infected at various times of the season. Plants showing symptoms of TEV infection early in the season had significantly less leaf area by seasons end (2 weeks after topping) than healthy plants or plants infected late in the season. Significant stunting was evident in plants first showing symptoms as late as 7 weeks after transplantation. Harvestable green leaf weight was not correlated with time of first TEV symptom expression. However, leaf weight per unit area was significantly correlated with time of plant infections, with earlier-infected plants having greater weight per unit area than later-infected plants or healthy plants. Yield, quality and value of the harvested tobacco were not correlated with number of infected plants per plot, however, probably because of compensation by neighbouring plants.

Novel substrates for the kinetic assay of esterases associated with insecticide resistance

Naphthalene thioesters were synthesized as substrates for a continuous, non- disruptive kinetic assay of general carboxylesterase activity. The continuous nature of the assay is based on the production of a soluble dianion chromophore from the reaction of naphthalene thio with 5,5′-dithiobis (2-nitrobenzoic acid). Applications with 1- and 2-naphthalene thioacetates demonstrated their use in a fast accurate kinetic microassay of esterase activity, using porcine carboxylesterase as a model. These novel esters proved to be useful as substrates for the spectrophotometric assay of insecticide-resistance in two aphid species and may be applicable to other esterasebased diagnostic procedures.

Biology and physiology of the white mutant of the Tobacco Hornworm, Manduca sexta (L.)

Abstract A white mutant of the tobacco hornworm, Manduca sexta , was discovered among wild-type, blue-green larvae in a laboratory colony routinely supplemented with field-collected insects. The mutant is pinkish-white when reared on artificial diet and maintains this coloration from egg hatch through pupation. Spectral analyses showed that the blue chromophore, biliverdin, which is present in the integument and plasma of wild-type, diet and tobacco-reared larvae, was lacking in the white mutant reared on the same diets. Insecticyanin levels determined by radial immunodiffusion in the integument and plasma of the white mutant were half that of normal blue-green larvae. The topical application of increasing amounts of the juvenoid, 2-(1-methyl-2-(4-phenoxyphenoxy)ethoxy)pyridine, to wild-type, day 1 first stadium larvae of M. sexta was positively correlated with the intensity of white coloration, which first appeared in newly molted second instars and was retained until pupation. White-mutant larvae developed slower and were slightly larger than wild-type larvae. The corpora allata from last stadium, day 0 larvae of the white mutant demonstrated a higher rate of JH/JH acid biosynthesis and the hemolymph had a higher titer of JH than the wild type of the same age. These studies suggested that JH may play a role in the regulation of the white phenotype. No differences were noted in the level of JH esterase activity between the two strains during the last stadium.