Action of a novel nonsteroidal ecdysteroid mimic ... - Insects.ugent.be

86 Guy Smagghe, Danny Degheele
Consequently, high concentrations of natural ecdysteroids
possess chemosterilizing properties in several
insect species.’8-20
The present study evaluates the toxicity and biological
activity of tebufenozide on insects of different orders,
families and genera. The compound was applied topically
or orally to various larval stages of several important
pest insects, namely; Lepidoptera: Spodoptera exigua
(Hiibner), Spodoptera exempta (Walker), Spodoptera
littoralis (Boisd.), Mamestra hrassicae L., and Galleria
mellonella L.: Coleoptera: Leptinotarsa decemlineata
(Say) and Diabrotica uirgijera uirgifera (LeConte); and
Orthoptera: L. migratoria migratorioides. Likewise,
tebufenozide was applied to nymphs of the predatory
Heteroptera, P. sagitta and Orius insidiosus (Say). This
research aimed to provide evidence for the novel mode
of action of tebufenozide, and its relatively high selectivity
for Lepidoptera compared to the prototype analogue
RH-5849. In addition, it evaluates the chemosterilizing
properties of tebufenozide in some species.
2.1 Chemicals
2 EXPERIMENTAL METHODS
Technical tebufenozide was obtained from Rohm and
Haas Company (Spring House, Pennsylvania, USA). All
solvents were of analytical grade.
2.2 Insects
S. exempta, the African armyworm, was reared on freshly
cut maize seedlings.’, Larvae of the beet armyworm, S.
exigua, and the Egyptian cotton leafworm, S. littoralis,
were fed on fresh castor-bean leaves, Ricinus communis
L.” The cabbage moth, M. brassicae, and the greater
wax moth, G. mellonella, were cultured as previously
de~cribed.’~ All stages of L. decemlineata, the Colorado
potato beetle, were fed on freshly cut potato foliage
(Solanum tuberosum L. cv ‘Bit~tJe’)~~ and D. uirgifera
uirgijera, the Western corn rootworm, on roots of live
maize seedings (Janssen, S., 1993, pers. comm.). Likewise,
the laboratory rearing of P. sagitta,2s 0. insidiosus26 and
L. migratoria rnigratorioide~~~ was carried out under
standard conditions.
2.3 Toxicity and insecticidal activity assay
For dipping experiments, different concentrations of
technical tebufenozide were prepared in water containing
02 ml litre-’ ‘Triton’ X-100. Freshly cut leaves were
dipped for 10 s, dried for 30-40 min at room temperature
in a fume hood and fed to the larvae ad libitum. For S.
exempta, D. uirgifera and L. migratoria larvae, freshly cut
maize seedlings were used; for S. exigua and S. littoralis,
fresh R. communis leaves and, for L. decemlineata, fresh
potato foliage. Leaves treated with water containing
0.2 ml litre-’ Triton X-100, were provided to the
controls.
For topical treatments, doses ranging from 001 to
40,000 ng per larva were applied by a dorsal administration
of 1 pl of methanolic solutions of technical
tebufenozide. Control specimens were treated with
methanol alone.
In another series of experiments, a high concentration
of 100 g litre-’ was prepared in dimethyl sulfoxide
(DMSO) and 1-2 p1 was topically applied on last-instars
of S. exempta and S. exigua. Control larvae were treated
with DMSO.
For P. sagitta nymphs, tebufenozide was topically
applied to the dorsal side, and, in a separate experiment,
to the ventral side because of the heavy dorsal sclerotization.
In another assay, P. sagitta nymphs were treated
orally by providing live last-instar S. exigua larvae as
prey. The latter were topically treated with 20 pg per
larva and allowed to take up the compound for 0.5 to 1 h.
In a preliminary experiment, the susceptibility of 0.
insidiosus nymphs to tebufenozide was also tested. As
food, fresh S. exigua eggs previously dipped for 10 min
in 1 g litre-’ (in methanol) and dried for 15 min, were
provided ad libitum to all nymphal stages (N,-N,) of
one generation. Controls were treated with methanol.
For both the leaf-dipping technique and topical
application, mortality percentages included both dead
and affected individuals. Mortality was monitored after
a time-period equal to the duration of the specific instar
plus 48 h for the last instar or 24 h for the other instars.
Mortality data were corrected for mortalities in the
controls and subjected to probit analysis using the
computer program POLO-PC.28 For each concentration,
20 to 40 insects were used and at least six different
concentrations were applied in order to calculate LC,,
and LD,, values.
Data on weight gain of treated specimens were also
determined and tested by one-way analysis of variance
(ANOVA) and mean values separated by a least significant
difference (LSD) multiple range test (P < 0.05).29
For internal abnormalities, control and treated specimens
were collected and fixed in 4% formaldehyde.
Subsequently, they were rinsed in distilled water and
dehydrated by passage through ethanol and xylol and
then embedded in paraplast. Sections were made with a
Historange microtome prior to examination with a light
microscope.
2.4 Effects on adult survival, fecundity and egg
viability of Spodoprera exigua
Different concentrations of tebufenozide were made in a
10% honeywater solution and supplied to the adults ad
libitum. Eight newly emerged S. exigua adults (sex ratio
1: 1) were placed together in a plastic box (1 1 x 1 1 x 16 cm)
the inside walls of which were covered with paper