Predation on Colorado Potato Beetle Eggs by Generalist Predators in Research and Commercial Potato Plantings

Field studies quantified predation on Colorado potato beetle [Leptinotarsa decemlineata(Say)] eggs and determined the relationship between predation and egg mass abundance in research and commercial potato plantings in eastern North Carolina. Predator exclusion experiments were conducted weekly in research plantings. In addition, egg mass density and predation on egg masses were monitored throughout the season in research plots and commercial potato fields. Predation was an important source of mortality for Colorado potato beetle eggs. Survivorship of eggs exposed to predators was consistently, significantly lower than survivorship of eggs protected from predation. Averaged over 2 years, the mean survivorship of eggs protected from predation was 69%, compared with 26% survivorship of eggs exposed to predation. Regression analysis failed to detect any relationship between egg mortality due to predation and egg abundance. These results imply that efforts to reduce Colorado potato beetle populations selectively will not be offset by an according decline in abundance of natural enemies and therefore should be fully compatible with naturally occurring biological control.     

Bacillus thuringiensis Delta-Endotoxin Cry1 Hybrid Proteins with Increased Activity against the Colorado Potato Beetle

Cry1 delta-endotoxins of Bacillus thuringiensis are generally active against lepidopteran insects, but Cry1Ba and Cry1Ia have additional, though low, levels of activity against coleopterans such as the Colorado potato beetle. Here we report the construction of Cry1Ba/Cry1Ia hybrid toxins which have increased activities against this insect species.     

Prey Preference by the StinkbugPerillus bioculatus,a Predator of the Colorado Potato Beetle

Perillus bioculatus(F.) is sometimes considered a generalist but has most often been recorded as predator of Colorado potato beetle (CPB),Leptinotarsa decemlineata.(Say). This study was designed to analyze prey selection inP. bioculatuswith respect to factors that may lead to specialization. To establish if parental prey determines preference in na?̈ve progeny, prey selection experiments were conducted with the CPB and two unusual prey, the yellow mealwormTenebrio molitor(L.) and the house cricketAcheta domesticus(L.). Na?̈ve nymphs reared from yellow mealworm-fed parents in the absence of contact with the CPB initiated feeding more frequently on CPB (81.4%) than on cricket prey (69.6%) (P = 0.038), suggesting genetically inheritable preference for CPB. Progeny from CPBfed parents initiated feeding more frequently on CPB prey (93.3%) than on yellow mealworm prey (76.6%) (P = 0.0001), but progeny from yellow mealwormfed parents initiated feeding on yellow mealworm prey as frequently (89.2%) as on CPB (95.0%) (P = 0.613). Similarly, progeny from CPB-fed parents established proboscis contact more frequently on CPB (39.2%) than on house cricket prey (25.0%) (P = 0.008), whereas those from cricket-fed parents probed house cricket prey as frequently (12.5%) as CPB prey (15.4%) (P = 0.416). Results confirm specialization ofP. bioculatustoward CPB or related prey and suggest genetically inheritable as well as maternally reinforcible affinity toward CPB prey. However, affinity of na?̈ve nymphs for the CPB can be lowered by rearing parents on alternative prey, increasing their chances of survival when alternative prey must be relied upon.     

Genetic Structure of Populations of the Colorado Potato Beetle Leptinotarsa decemlineata (Coleoptera: Chrysomelidae)

The genetic structure of a Colorado potato beetle population from Kiev oblast was examined by cluster analysis of individual RAPD patterns. The obtained clustering indicates that the population is structured. This may be explained by adaptation to pyrethroid insecticides used for controlling the population size of this pest. Microevolutionary factors affecting the genetic structure of local populations of Colorado potato beetle are discussed.     

Individual Polymorphism for RAPD Markers in Spring Generation of Colorado Potato Beetle Leptinotarsa decemlineata(Say)

Genetic diversity of Colorado potato beetle in a natural population from Fastovskii rayon, Kiev oblast was estimated using RAPD. The level of polymorphism for RAPD markers was far higher in the spring generation than in the sample obtained from the same locality in summer of the preceding year. The frequency of individual RAPD markers was found to differ in males and females of potato beetle.     

Expression of Poplar Chitinase in Tomato Leads to Inhibition of Development in Colorado Potato Beetle

The previously described poplar chitinase, WIN6, is induced during infestation by gypsy moth (Lymantria dispar L.) larvae, thus suggesting a role in defense against insect pests. To test this hypothesis, we produced tomato seedlings infected with a recombinant potato virus X (PVX), which produces WIN6, and tested its insecticidal properties on Colorado potato beetle [CPB; Leptinotarsa decemlineata (Say)], which is a serious pest of tomatoes and other crops. The advantage of PVX is that plant material is ready for insect bioassay within 3–4 weeks of constructing the recombinant virus. Considering that production of transgenic tomato seedlings using Agrobacterium takes at least 6 months, this hastens the rate at which genes can be examined. Upon insect bioassay, only 47% CPB neonates feeding on leaves containing >0.3% w/w WIN6 developed to 2nd instar while 93% of controls reached 2nd instar. To our knowledge this is the first plant chitinase that retards development of an insect pest. Revisions requested 12 December 2005; Revisions received 18 January 2006     

An In Vitro Inhibition Test That Predicts Toxicity of Bacterial Pathogen Combinations in the Colorado Potato Beetle

Many insect bacterial pathogens are not toxic enough for field control. Combinations of bacteria may increase toxicity. Bacteria toxic to Colorado potato beetle , Photorhabdus luminescens, Chromobacterium violaceum and Serratia marcescens , were tested in pair-wise combinations in an in vitro double streak test to determine bacterial compatibility. Only C. violaceum and S. marcescens grew to confluency. Their combined toxicity in vivo was additive. Other bacterial combinations had clear zones between bacterial streaks indicating inhibition. In the insect, the combined toxicity was less than the most toxic bacteria of the pair. For these strains, this in vitro test predicted compatibility in the insect.     

Influence of Pyriproxyfen on the Expression of Haemolymph Protein Genes in the Colorado Potato Beetle,Leptinotarsa decemlineata

Pyriproxyfen, a potent juvenile hormone analogue for the Colorado potato beetle, Leptinotarsa decemlineata, was applied topically to last-instar larvae and short-day adults at different times after moulting. The effect of the hormone analogue on concentration and composition of protein in the haemolymph was studied at different intervals after pyriproxyfen application. The hormone analogue had little effect on total protein concentration of the haemolymph, but affected protein composition. Diapause protein 1 was prevented from being synthesized if pyriproxyfen was applied before the gene was activated and disappeared from the haemolymph if applied after the gene had been expressed. It therefore inactivated the gene for diapause protein in both larvae and adults. Pyriproxyfen also induced appearance of vitellogenin at both stages, indicating induction of expression of the vitellogenin gene. It also affected the stability of mRNA for diapause protein. The analogue caused mRNA for diapause protein 1 to disappear untimely compared to controls in last-instar larvae and short-day adults. The response of adults to the JHA was much more pronounced than that of larvae, although the analogue had a strong biological effect on last-instar larvae because it prevented metamorphosis at low doses. Copyright 1997 Elsevier Science Ltd. All rights reserved     

Invasions on Large and Small Scales: Management of a Well-established Crop Pest, the Colorado Potato Beetle

Understanding the movement of invading organisms is critical to predicting invasion dynamics. The Colorado potato beetle, Leptinotarsa decemlineata (Say), is an invasive species on multiple spatial and temporal scales, and can serve as a model for studies of invasion dynamics. It is the major insect defoliator of potato in North America, and successful management requires an understanding of CPB invasions of individual fields. Its origin, spread, and biology, especially the cycle of annual invasions of agricultural potato fields, are described. Approaches to reducing the size of colonizing populations include rotation, delay of planting, and treatments of field margins. Rotation and sub-lethal insecticide treatments can slow the establishment of invasions within fields. These approaches interact with a late season diapause switch away from reproduction to reduce the impact of CPB. The refuge approach to delaying the fixation of resistance alleles is designed to encourage alleles for susceptibility to invade treated areas. We present data from an experimental refuge crop planted adjacent to a field treated with imidacloprid, an insecticide for which there is high variation in resistance. The treated field was four times as resistant as the untreated side, and a cline in resistance was formed from the untreated to the treated portion of the field. The cline width of about 100m provides an empirical basis for designing refuges to enhance the spread of alleles for susceptibility into treated areas and prevent fixation of resistance in the summer generation.     

Susceptibility of the Colorado Potato Beetle and the Sugarbeet Wireworm to Steinernema feltiae and S. glaseri

In laboratory tests, larvae of the Colorado potato beetle (CPB), Leptinotarsa decemlineata (Say), and the sugarbeet wireworm (SBW), Limonius californicus (Mannerheim), were exposed to the nematodes Steinernema feltiae Filipjev (Mexican strain) (= Neoaplectana carpocapsae) and S. glaseri Steiner in soil. S. feltiae caused significantly higher mortality in SBW larvae than did S. glaseri, but both nematode species were equally effective against CPB larvae. The minimum concentration of S. feltiae for 100% mortality of CPB larvae after 13 days was 157 nematodes/cm² of soil, and the LC₅₀ based on 6-day mortality was 47.5 nematodes/cm²; in contrast, 100% mortality of SBW larvae was not achieved with even the highest concentration tested, 393 nematodes/cm². CPB adults emerging from nematode-contaminated soil were not infected. In field cage tests, S. feltiae applied to the soil surface at the rates of 155 and 310 nematodes/cm² soil caused 59% and 71% mortality, respectively, of late-fourth-instar spring-generation CPB, and 28% and 29% mortality, respectively, of SBW. No infection was obtained when larvae of summer generation CPB and SBW were placed in the same cages approximately 6 weeks after nematodes were applied to the soil. Inundative soil applications of S. feltiae, though cost prohibitive at present, were effective in reducing caged CPB and SBW field populations.     

Impact of Host Plant Connectivity,Crop Border and Patch Size on Adult Colorado Potato Beetle Retention

Tagged Colorado potato beetles (CPB), Leptinotarsa decemlineata (Say), were released on potato plants, Solanum tuberosum L., and tracked using a portable harmonic radar system to determine the impact of host plant spatial distribution on the tendency of the pest to remain on the colonized host plant or patch. Results confirmed the long residency time on the host plant and showed that close connection of the plant to neighboring plants hastened dispersal between plants. Tracking walking CPB for over 6 h in small potato plots revealed that all types of mixed borders tested (potato/bare ground, potato/timothy and potato/woodland) acted as a strong barrier and retained beetles within the patch. In another experiment in potato patches surrounded by bare ground borders, tracked walking CPB displayed similar behaviour for up to four days. The distribution of turning angles in the CPB walking paths was not uniform and corresponded to beetles following the edge rows of potato patches in response to the crop border barrier or reversing their direction as they reached the end of a row and therefore a border. Patch size had no or little effect on beetle retention in the patch. The relative distribution of counts of tagged beetles detected among small (16 m²), medium (64 m²) and large size (256 m²) patches of potato four days after initial release remained similar to that of numbers released. Even though mixed crop borders were a strong barrier to walking CPB emigrating from potato patches, the departure rate of beetles over time was high. Results suggest that the effect of mixed borders is largely limited to dispersal by walking and does not apply to beetles leaving host patches by flight. The manipulation of crop borders and patch size seem to have limited potential for the management of CPB emigrating from potato fields.     

Laboratory Evaluation of Isaria fumosorosea CCM 8367 and Steinernema feltiae Ustinov against Immature Stages of the Colorado Potato Beetle

The Colorado potato beetle, Leptinotarsa decemlineata, has developed resistance to most registered pesticides and has become one of the most difficult insect pests to control. Development of new biopesticides targeting this pest might solve the resistance problem and contribute to sustainable crop production. Laboratory experiments were conducted to assess the efficacy of Isaria fumosorosea (syn. Paecilomyces fumosoroseus) strain CCM 8367 against L. decemlineata when applied alone or combined with the entomopathogenic nematode Steinernema feltiae. The last-instar larvae of the Colorado potato beetle showed the highest susceptibility to I. fumosorosea followed by pre-pupae and pupae. The median lethal concentration (LC₅₀) was estimated to be 1.03×10⁶ blastospores/ml. The strain CCM 8367 was more virulent, causing 92.6% mortality of larvae (LT₅₀ = 5.0 days) compared to the reference strain Apopka 97, which caused 54.5% mortality (LT₅₀ = 7.0 days). The combined application of the fungus with the nematodes increased the mortality up to 98.0%. The best results were obtained when S. feltiae was applied simultaneously with I. fumosorosea (LT₅₀ = 2.0 days); later application negatively affected both the penetration rate and the development of the nematodes. We can conclude that the strain CCM 8367 of I. fumosorosea is a prospective biocontrol agent against immature stages of L. decemlineata. For higher efficacy, application together with an entomopathogenic nematode is recommended.     

Botanical insecticides for controlling agricultural pests: piperamides and the Colorado Potato Beetle Leptinotarsa decemlineata say (Coleoptera: Chrysomelidae)

The efficacy of extracts from two Piperaceae species, Piper nigrum L. and P. tuberculatum Jacq. were evaluated using larvae and adults of the Colorado Potato Beetle Leptinotarsa decemlineata (Say). Young larvae and neonates were the most susceptible; a 24-h LD(50) of 0.064% extract of P. tuberculatum was determined for 4-day-old larvae, while 0.05% extract of P. nigrum reduced larval survival up to 70% within one week after treatment of potato Solanum tuberosum L. (Solanaceae) plants. When an insecticide resistant strain of L. decemlineata larvae was tested with the P. tuberculatum extract, there was less than a 2-fold tolerance ratio compared to the 22-fold tolerance ratio to cypermethrin, a pyrethroid. Older larvae, pre-pupal stage and adults, were less sensitive to the P. nigrum extracts; the 24-h LD(50) was 0.5% (95% C.I. = 0.36, 0.65). However, the same concentration was equally effective under field conditions. In the greenhouse, P. nigrum at 0.5% was as effective at reducing adult L. decemlineata feeding as combinations with 2 separate botanical mixtures, garlic and lemon grass oil. Under field conditions, the residual activity of the P. nigrum extracts was less than 3 h. When adult L. decemlineata were placed on treated plants exposed to full sunlight for 0, 1.5, and 3 h, leaf damage progressively increased as the main active compound, piperine, was found to degrade by 80% after 3 h. An in vitro polysubstrate monoxygenase (PSMO) enzyme assay, using the substrate methoxyresorufin O-demethylation (MROD), determined that the principal P. nigrum active compound, piperine, is responsible for inhibition of that specific enzyme. The results suggest that Piper extracts could be used effectively as contact botanical insect control agents to protect potato plants from developing L. decemlineata larvae at concentrations less than 0.1%. There is also potential for Piper extracts to control insecticide resistant populations in conjunction with other integrated pest management (IPM) strategies used in conventional and organic agriculture.     

Production of transgenic eggplant (Solanum melongena L.) resistant to Colorado Potato Beetle (Leptinotarsa decemlineata Say)

 A modified gene of Bacillus thuringiensis var. Tolworthi (Bt), encoding a coleopteran insect-specific CryIIIB toxin, was transferred via Agrobacterium tumefaciens to the female parent of the eggplant commercial F₁ hybrid ‘Rimina’. One-hundred and fifty eight transgenic plants were regenerated and tested by PCR and NPTII expression assays. The presence of the CryIIIB toxin in leaf extracts was demonstrated in 57 out of 93 transgenic plants tested by DAS-ELISA assay. High Bt-expressing plants contained a 74-kDa protein cross-reacting with serum anti-CryIIIB toxin. Twenty three out of 44 S. melongena plants tested by insect bioassay showed significant insecticidial activity on neonate larvae of Colorado Potato Beetle (CPB). The Bt transgene and the toxic effect on CPB larvae were transmitted to progenies derived by selfing. Thus, transgenic Bt eggplants represent a very effective means of CPB pest control. Received: 25 November 1996/Accepted: 31 January 1997     

Timing of the Initial Spray Application of Bacillus thuringiensis for Control of the Colorado Potato Beetle (Coleoptera: Chrysomelidae) in Potatoes

Field experiments were conducted in 1989 and 1990 to evaluate the timing of the initial spray application of Bacillus thuringiensis subsp. tenebrianis, relative to egg development in the field, for control of the Colorado potato beetle (CPB), Leptinotarsa decemlineata (Say) in potatoes. Effective potato beetle control was achieved with B. thuringiensis if the initial spray was applied during the period beginning when an average of 1-30% of marked egg masses had hatched, extending to 4 days after 30% egg hatch, of the initial flush of eggs. Significantly greater defoliation, numbers of third and fourth instar CPB and lower yields occurred in plots when the initial B. thuringiensis application was delayed until 6 days after 30% egg hatch, compared with plots treated at 30% egg hatch. In 1990, no improvement in CPB control was gained by application of the initial B. thuringiensis spray at first observation of CPB egg deposition or at first egg hatch, compared with application at 30% egg hatch. Recommendations for proper timing of B. thuringiensis sprays for effective CPB management are presented.     

Functional Response ofColeomegilla maculata(Coleoptera: Coccinellidae) to Colorado Potato Beetle Eggs (Coleoptera: Chrysomelidae)

A comparative study of the functional response ofColeomegilla maculataDeGeer (Coleoptera: Coccinellidae) fourth instars was conducted under laboratory, greenhouse, and field conditions. In the laboratory, individual larvae were placed in 9-cm petri dishes for 24 h, with 1, 3, 5, or 7 Colorado potato beetle (Leptinotarsa decemlineata[Say]) (Coleoptera: Chrysomelidae) egg masses. Each egg mass was standardized at 15 eggs. In the greenhouse and field,C. maculatalarvae were provided with an equivalent of 0.5 to 35L. decemlineataegg masses/m²of potato leaf. Fourth instars ofC. maculataexhibited a type II functional response toL. decemlineataeggs under laboratory, greenhouse, and field conditions. Predator search efficiency was inversely related with prey density. The maximum mean attack rate (8.7 eggs) byC. maculatalarvae in the field was about half the mean attack rate in the laboratory (17.6 eggs) and greenhouse (20.1 eggs). The difference in prey density between the laboratory and field seems to have been a major contributing factor in determining the rate of predation, whereas differences in environmental conditions (e.g., temperature and possible alternate food) may explain the differences observed in the predation rate in the greenhouse and field.     

Effects of Selected Fungicides and the Timing of Fungicide Application on Beauveria bassiana-Induced Mortality of the Colorado Potato Beetle (Coleoptera: Chrysomelidae)

Four fungicides used for controlling foliar diseases of potato (Solanum tuberosum) were evaluated under field and laboratory conditions for their effects on the infectivity and sporulation of Beauveria bassiana when used as a control for the Colorado potato beetle, Leptinotarsa decemlineata (CPB). We investigated the direct effects of fungicides on B. bassiana-induced CPB mortality and the effect of time between fungicide and B. bassiana application. Effects of fungicide on conidial survival in the soil and on foliage were examined in the field. Significantly more larval mortality was observed when larvae were sprayed with B. bassiana than with the water control. Fungicide had no significant effect on larval mortality in the field. In the laboratory, survival of larvae was significantly lower among larvae fed fungicide-treated foliage. B. bassiana-induced mortality in the laboratory was observed only when larvae were fed foliage treated with copper hydroxide or water. Larvae fed mancozeb- or chlorothalonil-treated foliage experienced high mortality regardless of B. bassiana treatment. While there was no significant effect of fungicide on B. bassiana sporulation on cadavers in the field, a pattern emerged that indicated higher proportions of cadavers producing conidia in plots sprayed with water or copper hydroxide than in plots sprayed with chlorothalonil or mancozeb. Survival of B. bassiana conidia in the soil and on foliage was significantly greater in plots treated with copper hydroxide or water than in plots treated with mancozeb or chlorothalonil. Fungicides such as copper hydroxide may be less deleterious to the fungus than mancozeb and chlorothalonil.     

Isolation and Culture of Corn Stunt Spiroplasma in Serum-Free Medium

A serum-free medium (medium A₆), supports the primary isolation and cultivation in vitro of corn stunt spiroplasma (CSS). The growth characteristics of CSS in A₆ medium are similar to those in the conventional serum dependent, ketoglutarate sorbitol medium (SMK).