Functional and genetic characteristics of Chlorantraniliprole resistance in the diamondback moth,Plutella xylostella (Lepidoptera: Plutellidae)

The diamondback moth (Plutella xylostella) is a globally distributed and important economic pest, and it has developed resistance to all conventional insecticide classes used in the field. Chlorantraniliprole is a new chemical class of insecticide that acts as a conformation‐sensitive activator of the insect ryanodine receptor (RyR). In the present study, a field strain (16.3‐fold resistance to chlorantraniliprole) was collected in Korea and lab‐selected with chlorantraniliprole for more than one year. The resulting strain presented 2,157‐fold resistance to chlorantraniliprole. A point mutation (G4946E) in the RyR gene was observed at a high frequency in the resistant strain. Enzyme assays indicated that glutathione S‐transferase (GST) and P450 activity in the resistant strain were 2.4‐ and 1.96‐times higher than that of the susceptible strain, respectively. The expression of the RyR, GST (sigma, omega, and zeta) and CYP321E1 gene was higher in the resistant strain than in the susceptible strain. The F₁ progeny resulting from reciprocal crosses did not reveal maternal effects or a diamide‐susceptible phenotype, which suggests an autosomal nearly recessive mode of inheritance. In addition, we surveyed the susceptibility to 13 insecticides (3 diamides, 2 synthetic pyrethroids, 2 spinosyns, 1 organophosphate, 1 oxadiazine, 1 avermectin, and 3 others) in the chlorantraniliprole‐resistant strain. The resistant strain exhibited high cross‐resistance to flubendiamide (5,910 fold) and showed no cross‐resistance to spinetoram, spinosad, indoxacarb, and metaflumizone. These results can serve as an important basis for guiding the use of insecticides in the field.     

Limitations of RNAi of α6 nicotinic acetylcholine receptor subunits for assessing the in vivo sensitivity to spinosad

Abstract Spinosad is a widely used insecticide that exerts its toxic effect primarily through interactions with the nicotinic acetylcholine receptor. The α6 nicotinic acetylcholine receptor subunit is involved in spinosad toxicity as demonstrated by the high levels of resistance observed in strains lacking α6. RNAi was performed against the Dα6 nicotinic acetylcholine receptor subunit in Drosophila melanogaster using the Gal4‐UAS system to examine if RNAi would yield results similar to those of Dα6 null mutants. These Dα6‐deficient flies were subject to spinosad contact bioassays to evaluate the role of the Dα6 nicotinic acetylcholine receptor subunit on spinosad sensitivity. The expression of Dα6 was reduced 60%–75% as verified by quantitative polymerase chain reaction. However, there was no change in spinosad sensitivity in D. melanogaster. We repeated RNAi experiments in Tribolium castaneum using injection of dsRNA for Tcasα6. RNAi of Tcasα6 did not result in changes in spinosad sensitivity, similar to results obtained with D. melanogaster. The lack of change in spinosad sensitivity in both D. melanogaster and T. castaneum using two routes of dsRNA administration shows that RNAi may not provide adequate conditions to study the role of nicotinic acetylcholine receptor subunits on insecticide sensitivity due to the inability to completely eliminate expression of the α6 subunit in both species. Potential causes for the lack of change in spinosad sensitivity are discussed.     

A nicotinic acetylcholine receptor transmembrane point mutation (G275E) associated with resistance to spinosad in Frankliniella occidentalis

High levels of resistance to spinosad, a macrocyclic lactone insecticide, have been reported previously in western flower thrips, Frankliniella occidentalis, an economically important insect pest of vegetables, fruit and ornamental crops. We have cloned the nicotinic acetylcholine receptor (nAChR) α6 subunit from F. occidentalis (Foα6) and compared the nucleotide sequence of Foα6 from susceptible and spinosad‐resistant insect populations (MLFOM and R1S respectively). A single nucleotide change has been identified in Foα6, resulting in the replacement of a glycine (G) residue in susceptible insects with a glutamic acid (E) in resistant insects. The resistance‐associated mutation (G275E) is predicted to lie at the top of the third α‐helical transmembrane domain of Foα6. Although there is no direct evidence identifying the location of the spinosad binding site, the analogous amino acid in the C. elegans glutamate‐gated chloride channel lies in close proximity (4.4 Å) to the known binding site of ivermectin, another macrocyclic lactone pesticide. The functional consequences of the resistance‐associated mutation have been examined in the human nAChR α7 subunit. Introduction of an analogous (A272E) mutation in α7 abolishes the modulatory effects of spinosad whilst having no significant effect upon activation by acetylcholine, consistent with spinosad having an allosteric mechanism of action.     

Could resistance to insecticides in Plutella xylostella (Lep., Plutellidae) be overcome by insecticide mixtures?

Abstract: To investigate if synergism occurs between pyrethroids, organophosphates and new insecticides, we tested representatives of these compounds (bifenthrin, chlorpyrifos, spinosad, indoxacarb and emamectin) against the diamondback moth (Plutella xylostella). Larvicidal activity of these insecticides was assessed separately and together on a susceptible strain (Lab‐UK) of P. xylostella as well as a field population collected from Multan. The field population showed significant resistance to chlorpyrifos (331 100‐fold), bifenthrin (45 200‐fold), emamectin (1800‐fold), spinosad (11‐fold) and indoxacarb (5600‐fold) when compared with the Lab‐UK population. When insecticides were mixed based on LC₅₀ and tested at serial concentrations against Lab‐UK, significant synergy (CI < 1) occurred between bifenthrin, spinosad and emamectin. In contrast, the interaction between bifenthrin and indoxacarb was additive (CI = 1). The toxicity of bifenthrin against the field population increased significantly (P < 0.01) when combined with spinosad, emamectin and indoxacarb. Synergistic effects could be attributed to the complementary modes of action by these insecticide classes acting on different components of nerve impulse transmission (which are not identical for bifenthrin and indoxacarb either). However, chlorpyrifos/bifenthrin mixture was not significantly different either from bifenthrin or chlorpyrifos alone, indicating an additive affect. In combination with spinosad and emamectin, tested against the resistant field population, the toxicity of chlorpyrifos increased significantly and even more so with indoxacarb. Mixtures could also give rise to multiple resistance that may extend across other chemical classes and thus become difficult to manage. Therefore, alternative strategies such as mosaics or rotations should be considered. That is, though synergistic effects have been found, this should not be followed up as a strategy to manage resistant field populations.     

Development of resistance to spinosad in oriental fruit fly (Diptera: Tephritidae) in laboratory selection and cross-resistance

In this study, we assessed the potential for the development of resistance to the insecticide spinosad in a laboratory colony of the oriental fruit fly, Bactrocera dorsalis (Hendel) (Diptera: Tephritidae). Resistance was selected by using topical applications of spinosad. After eight generations of selection, the LD50 of the selected line was 408 times greater compared with that of the untreated parental colony. This spinosad-resistant line did not exhibit cross-resistance to 10 other insecticides tested, including six organophosphates (naled, trichlorfon, fenitrothion. fenthion, formothion, and malathion) one carbamate (methomyl), and three pyrethroids (cyfluthrin, cypermethrin, and fenvalerate). However, using lines previously selected for resistance to these same insecticides, two of the 10 lines tested (naled- and malathion-resistant) did show some cross-resistance to spinosad. Also, oriental fruit flies from different field collections where naled and malathion have been used for control purposes displayed some resistance to spinosad. In addition, the effects of direct ingestion of spinosad through dietary supplementation also were tested. Overall, the laboratory resistance and cross-resistance data developed in this study provide new information that will be useful for managing the development of resistance when spinosad is used to control B. dorsalis in the field.     

Stability of spinosad resistance in Frankliniella occidentalis (Pergande) under laboratory conditions

The stability of spinosad resistance in western flower thrips (WFT), Frankliniella occidentalis (Pergande), populations with differing initial frequencies of resistance was studied in laboratory conditions. The stability of resistance was assessed in bimonthly residual bioassays in five populations with initial frequencies of 100, 75, 50, 25 and 0% of resistant individuals. There were no consistent changes in susceptibility of the susceptible strain after eight months without insecticide pressure. In the resistant strain, very highly resistant to spinosad (RF50>23,000-fold), resistance was maintained up to eight months without further exposure to spinosad. In the absence of any immigration of susceptible genes into the population, resistance was stable. In the case of the population with different initial frequency of resistant thrips, spinosad resistance declined significantly two months later in the absence of selection pressure. With successive generations, these strains did not change significantly in sensitivity. Spinosad resistance in F. occidentalis declined significantly in the absence of selection pressure and the presence of susceptible WFT. These results suggest that spinosad resistance probably is unstable under field conditions, primarily due to the immigration of susceptible WFT. Factors influencing stability or reversion of spinosad resistance are discussed.     

Potential for insecticide resistance in populations of Bactrocera dorsalis in Hawaii: spinosad susceptibility and molecular characterization of a gene associated with organophosphate resistance

The potential for populations to become resistant to a particular insecticide treatment regimen is a major issue for all insect pest species. In Hawaii, for example, organophosphate (OP)‐based cover sprays have been the chemical treatment most commonly applied against oriental fruit fly, Bactrocera dorsalis (Hendel) (Diptera: Tephritidae), populations since the 1950s. Moreover, bait spray treatments using spinosad were adopted as a major control tactic in the Hawaii area‐wide fruit fly pest management program beginning in the year 2000. To determine the current level of spinosad and OP tolerance of wild B. dorsalis populations, bioassays were conducted on flies collected from a range of geographic localities within the Hawaiian islands. Adult B. dorsalis flies were tested (1) for the level of susceptibility to spinosad using LC₅₀ diagnostic criteria, and (2) for the presence of alleles of the ace gene previously shown to be associated with OP resistance. Regarding spinosad tolerance, only flies from Puna, the one area lacking prior exposure to spinosad, showed any significant difference compared to controls, and here the difference was only in terms of non‐overlap of 95% fiducial limit values. With respect to OP tolerance, specific mutations in the ace gene associated with resistance to these insecticides were found in only two populations, and in both cases, these alleles occurred at relatively low frequencies. These results suggest that at the present time, populations of B. dorsalis in Hawaii show no evidence for having acquired resistance to the insecticides widely used in control programs.     

Lack of fitness costs of insecticide resistance in the western flower thrips (Thysanoptera: Thripidae)

The fitness costs of spinosad and acrinathrin resistance was investigated in the western flower thrips, Frankliniella occidentalis (Pergande) (Thysanoptera: Thripidae). Fitness studies were conducted on susceptible and resistant strains of F. occidentalis. Resistant females were significantly more fecund (number of eggs per female) than susceptible females. The hatching rate (fertility) for both susceptible and acrinathrin-resistant strains was significantly lower than in the spinosad-resistant strain. Mean developmental time from egg to adult did not differ between thrips populations. Similarly, female longevity did not differ between populations. These data suggest that lack of fitness costs related to insecticide resistance may accelerate the development of insecticide resistance in populations of F. occidentalis from southeastern Spain.     

Monitoring and mechanisms of organophosphate resistance in San Jose scale,Diaspidiotus perniciosus (Hemiptera: Diaspididae)

San Jose scale, Diaspidiotus perniciosus (Comstock), is a serious pest in Chilean apple tree orchards, and a number of organophosphate insecticides were used to control them for decades. Recently, control failures with these insecticides were reported and linked to insecticide resistance development. In this study, 40 San Jose scale field populations were collected and their susceptibility to two commonly used organophosphate insecticides, that is chlorpyrifos and methidathion, was assessed. The obtained bioassay data suggest moderate levels of resistance to both insecticides when compared to a reference susceptible strain. The highest resistance ratio (RR) detected for chlorpyrifos and methidathion was 31‐fold and 11‐fold, respectively. The bioassay results suggest the occurrence of a significant cross‐resistance between both compounds. Biochemical measurements revealed a role for esterases in conferring resistance to organophosphates, but not modified acetylcholinesterase. The spatial spread and extend of insecticide resistance were also evaluated. Our result shows that no autocorrelation can be assumed, and then, insecticide resistance is caused by random factors.     

Insecticide resistance and control failure likelihood of the whitefly Bemisia tabaci (MEAM1; B biotype): a Neotropical scenario

Insecticide resistance is a standing concern for arthropod pest species, which may result in insecticide control failure. Nonetheless, while insecticide resistance has remained a focus of attention for decades, the incurring risk of insecticide control failure has been neglected. The recognition of both problems is paramount for arthropod pest management and particularly so when invasive species notoriously difficult to control and exhibiting frequent cases of insecticide resistance are considered. Such is the case of the putative whitefly species Middle East‐Asia Minor I (MEAM1) (Bemisia tabaci B‐biotype), for which little information is available in the Neotropics. Thus, the likely occurrence and levels of resistance to seven insecticides were surveyed among Brazilian populations of this species. The likelihood of control failure to the five insecticides registered for this species was also determined. Resistance was detected to all insecticides assessed reaching instances of high (i.e. >100×) to very high levels (>1000×) in all of them. Overall efficacy was particularly low (<60%) and the control failure likelihood was high (>25%) and frequent (70%) for the bioinsecticide azadirachtin, followed by spiromesifen and lambda‐cyhalothrin. In contrast, the likelihood of control failure was low for diafenthiuron, and mainly imidacloprid. As cartap and chlorantraniliprole are not used against whiteflies, but are frequently applied on the same host plants, inadvertent selection probably took place leading to high levels of resistance, particularly for the latter. The resistance levels of cartap and chlorantraniliprole correlated with imidacloprid resistance (r > 0.65, P < 0.001), suggesting that the latter use may have somewhat favoured inadvertent selection for resistance to both compounds not used against the whitefly. A further concern is that chlorantraniliprole use in the reported scenario may allow cross selection to cyantraniliprole, a related diamide with recent registration against whiteflies demanding attention in designing resistance management programmes.     

Temperature-related fitness costs of resistance to spinosad in the diamondback moth, Plutella xylostella (Lepidoptera: Plutelidae)

Fitness costs associated with resistance genes expressed in the absence of insecticides affect the evolution of insecticide resistance and the outcome of resistance management programmes. However, measurements of fitness costs may not be straightforward as they vary with environmental conditions. The diamondback moth, Plutella xylostella L. (Lepidoptera: Plutellidae), has developed resistance to spinosad, the first insecticide of the Naturalyte class, after only a few years of field application of this product. In this study, we compared the performance of two homogenous strains of P. xylostella, one susceptible (SS) and the other resistant (RR) to spinosad at an unfavourable, low natural temperature regime, a favourable median-fluctuating temperature regime and an unfavourable high-fluctuating temperature regime. The RR strain showed only marginal fitness cost at the median temperature regime. At the low temperature regime, the RR strain failed to produce any viable offspring, while the SS strain achieved positive population growth. At the high temperature regime, the RR strain showed a 33% decrease in intrinsic rate of increase compared to the SS strain. The results demonstrate that fitness costs of resistance to spinosad are temperature-dependent, increasing in scale at unfavourably low and high temperatures; costs were particularly high at low temperatures. Suggestions for designing effective management programmes are made to delay or avoid development of resistance to spinosad by P. xylostella under different temperature conditions.     

Impact of spinosad on ichneumonid‐parasitized Choristoneura rosaceana larvae and subsequent parasitoid emergence

The impact of low levels of spinosad on the obliquebanded leafroller, Choristoneura rosaceana Harris (Lepidoptera: Tortricidae), and the koinobiont endoparasitoid, Apophua simplicipes Cresson (Hymenoptera: Ichneumonidae), was assessed when the parasitoid was in the larval stage within second‐ and fourth‐instar hosts. These are developmental stages that would be exposed to spring orchard treatments of the insecticide. Oral spinosad LC₅₀ levels for unparasitized obliquebanded leafroller hosts were <1% of the recommended orchard treatment levels. Apophua simplicipes survival was significantly reduced within parasitized spinosad‐treated second‐ and fourth‐instar larval hosts. Both the leafroller host and parasitoid were much more susceptible (ca. 65‐fold) to spinosad when larval hosts fed on spinosad‐treated leaf material as opposed to being treated topically. When hosts were exposed to extremely low doses of spinosad, a small percentage of parasitoids was able to survive to emerge as adults. These laboratory trials predict that applications of spinosad may reduce biological control of C. rosaceana populations by ichneumonid endoparasitoids developing within treated hosts.     

Abamectin resistance in strains of vegetable leafminer,Liriomyza sativae (Diptera: Agromyzidae) is linked to elevated glutathione S‐transferase activity

Abamectin resistance was selected in the vegetable leafminer, Liriomyza sativae (Blanchard) (Diptera: Agromyzidae) under laboratory conditions, and cross‐resistance patterns and possible resistance mechanisms in the abamectin‐resistant strains (AL‐R, AF‐R) were investigated. Compared with the susceptible strain (SS), strain AL‐R displayed 39‐fold resistance to abamectin after 20 selection cycles during 25 generations, and strain AF‐R exhibited 59‐fold resistance to abamectin after 16 selection cycles during 22 generations. No cross‐resistance to cyromazine was found in both abamectin‐resistant strains. However, we failed to select for cyromazine resistance in L. sativae under laboratory conditions by conducting 17 selection cycles during 22 generations. However, moderate levels of cross‐resistance to abamectin (6–9 fold) were observed in strains which received cyromazine treatments. Biochemical analysis showed that glutathione S‐transferase (GST) activity in both abamectin‐resistant strains (AL‐R, AF‐R) was significantly higher than in the susceptible strain (SS), suggesting metabolically driven resistance to abamectinin L. sativae. Recommendations of mixtures or rotation of cyromazine and abamectin should be considered carefully, as consecutive cyromazine treatments may select for low‐level cross‐resistance to abamectin.     

Impact of phagostimulants on effectiveness of OMRI‐listed insecticides used for control of spotted‐wing drosophila (Drosophila suzukii Matsumura)

Spotted‐wing drosophila, Drosophila suzukii Matsumura, is an invasive pest in the United States that causes considerable damage to fruit crops. It is responsible for many millions of dollars of revenue loss. The female D. suzukii has a heavily sclerotized ovipositor and can lay eggs in ripening or ripe fruit. The arrival of this invasive species has disrupted existing integrated pest management programmes, and growers rely on repeated insecticide applications to protect fruit. Organic growers have few chemical control options, and their reliance on spinosad increases the risk of developing insecticide resistance. We hypothesized that combining phagostimulants with insecticides would increase insecticide efficacy by prompting flies to spend more time in contact with residues. Therefore, the objective of this study was to evaluate the effectiveness of sucrose and the yeast Saccharomyces cerevisiae as phagostimulants in combination with organic biopesticides against D. suzukii in blueberries. Adding sucrose with or without yeast did not improve insecticide efficacy in terms of adult fly mortality or fruit infestation. Spinosad was very effective in all experiments, and for this product, there is little room for improvement. The phagostimulants had no effect on residual activity of any insecticide. The addition of sucrose with or without yeast did not improve the effectiveness of organic insecticides for D. suzukii. Concentrations of these phagostimulants in our experiments (0.36%) may have been too low to elicit a response. Further research is recommended to test different types and concentrations of phagostimulants.     

The Wiggle Index: An Open Source Bioassay to Assess Sub-Lethal Insecticide Response in Drosophila melanogaster

Toxicological assays measuring mortality are routinely used to describe insecticide response, but sub-lethal exposures to insecticides can select for resistance and yield additional biological information describing the ways in which an insecticide impacts the insect. Here we present the Wiggle Index (WI), a high-throughput method to quantify insecticide response by measuring the reduction in motility during sub-lethal exposures in larvae of the vinegar fly Drosophila melanogaster. A susceptible wild type strain was exposed to the insecticides chlorantraniliprole, imidacloprid, spinosad, and ivermectin. Each insecticide reduced larval motility, but response times and profiles differed among insecticides. Two sets of target site mutants previously identified in mortality studies on the basis of imidacloprid or spinosad resistance phenotypes were tested. In each case the resistant mutant responded significantly less than the control. The WI was also able to detect a spinosad response in the absence of the primary spinosad target site. This response was not detected in mortality assays suggesting that spinosad, like many other insecticides, may have secondary targets affecting behaviour. The ability of the WI to detect changes in insecticide metabolism was confirmed by overexpressing the imidacloprid metabolizing Cyp6g1 gene in digestive tissues or the central nervous system. The data presented here validate the WI as an inexpensive, generic, sub-lethal assay that can complement information gained from mortality assays, extending our understanding of the genetic basis of insecticide response in D. melanogaster.     

Development and stability of insecticide resistance in the leafminer Liriomyza trifolii (Diptera: Agromyzidae) to cyromazine, abamectin, and spinosad

Three populations of the leafminer, Liriomyza trifolii (Burgess), were collected from commercial ornamental production greenhouses in the United States and tested for susceptibility to three commercial insecticides. A leaf dip bioassay of leaves containing young (1-2-d-old) larvae was used. Based on larval mortality and compared with a susceptible laboratory reference colony, the three strains varied in spectrum and level of resistance to the insecticides. CA-1, collected from Gerbera daisy, was moderately resistant to cyromazine (18.1-fold) and abamectin (22.0-fold), but highly resistant to spinosad (> 188-fold). CA-2, collected from chrysanthemums, was not resistant to abamectin, had a low level of resistance to cyromazine (8.2-fold), but was extremely resistant to spinosad (1,192-fold). GA-1, collected from chrysanthemums, had very low levels of resistance to cyromazine (5.4-fold) and spinosad (1.9-fold) but was moderately resistant to abamectin (30.6-fold). When reared in the absence of insecticide selection pressure, all three strains reverted to approximately the level of the reference strain. The CA-1 strain reverted in nine generations to cyromazine; however, the lowest levels of abamectin and spinosad resistance reverted to was 3.1-fold at F8 and 3.2 at the F10, respectively. The CA-2 strain reverted in five generations to both cyromazine and spinosad. GA-1 reverted in five generations to abamectin. Based on the results, resistance to these three insecticides was unstable. Additionally, there was no cross-resistance among these three insecticides.     

Evaluation of organic insecticides for management of spotted‐wing drosophila (Drosophila suzukii) in berry crops

Spotted‐wing drosophila, Drosophila suzukii (Matsumura), is an invasive pest affecting fruit production in many regions of the world. Insecticides are the primary tactic for controlling D. suzukii in organic as well as conventional production systems. Organic growers have a greater challenge because fewer insecticides are approved for use in organic agriculture. The most effective organically approved product is spinosad, but alternatives are needed because of label restrictions limiting the number of applications per year, toxicity to beneficial arthropods and the risk of developing resistance. We evaluated several organically approved insecticides against D. suzukii in laboratory assays and field trials conducted on organic blueberry and raspberry farms. Spinosad was consistently the most effective insecticide, but a few other insecticides such as azadirachtin + pyrethrins, Chromobacterium subtsugae and sabadilla alkaloids showed moderate activity. None of the treatments had long residual activity. Mortality started to decline by 3 days after treatment, and by 5 days after application, the treatments were not different from the controls. These products may be useful in rotation programmes, necessary for reducing reliance on spinosad and mitigating resistance. Cultural and biological control approaches are needed in fruit production for D. suzukii management, but insecticides will likely continue to be the dominant management tactic while these other approaches are being optimized and adopted.     

Antioxidant status and ultrastructural defects in the ovaries of red palm weevils (Rhynchophorus ferrugineus) intoxicated with spinosad

Red palm weevil, Rhynchophorus ferrugineus, destroys palms almost all over the world, especially in the Middle East, where dates are a strategic crop in arid places. In view of the urgent need to combat this destructive pest, effective pesticides with high environmental safety should be sought. Spinosad is a pesticide of bacterial origin that is presumed to have a high degree of environmental safety and is effective in combating a wide range of insect pests. In this study, the efficacy of spinosad was evaluated in females of R. ferrugineus. The lethal concentration for 50% of the treated females was calculated at 44.3 ppm. The effects of spinosad concentrations of 10, 50 and 200 ppm on the activity of catalase (CAT), glutathione S‐transferase (GST) and superoxide dismutase (SOD) in the ovaries were assessed. In addition, the pathological effects of these concentrations were documented on the ultrastructure of the follicle cells and ooplasm. The results showed a significant increase in CAT activity only in response to treatment with 200 ppm. Treating the females with different spinosad concentrations resulted in varying intensity changes in cell organelles, where the most pronounced sign of programmed cell death was at the concentration 200 ppm. This study demonstrates the possibility of using spinosad as an insecticide against females of R. ferrugineus.     

Spinosad and the Tomato Borer Tuta absoluta: A Bioinsecticide,an Invasive Pest Threat,and High Insecticide Resistance

The introduction of an agricultural pest species into a new environment is a potential threat to agroecosystems of the invaded area. The phytosanitary concern is even greater if the introduced pest’s phenotype expresses traits that will impair the management of that species. The invasive tomato borer, Tuta absoluta (Meyrick) (Lepidoptera: Gelechiidae), is one such species and the characterization of the insecticide resistance prevailing in the area of origin is important to guide management efforts in new areas of introduction. The spinosad is one the main insecticides currently used in Brazil for control of the tomato borer; Brazil is the likely source of the introduction of the tomato borer into Europe. For this reason, spinosad resistance in Brazilian populations of this species was characterized. Spinosad resistance has been reported in Brazilian field populations of this pest species, and one resistant population that was used in this study was subjected to an additional seven generations of selection for spinosad resistance reaching levels over 180,000-fold. Inheritance studies indicated that spinosad resistance is monogenic, incompletely recessive and autosomal with high heritability (h ² = 0.71). Spinosad resistance was unstable without selection pressure with a negative rate of change in the resistance level ( = −0.51) indicating an associated adaptive cost. Esterases and cytochrome P450-dependent monooxygenases titration decreased with spinosad selection, indicating that these detoxification enzymes are not the underlying resistance mechanism. Furthermore, the cross-resistance spectrum was restricted to the insecticide spinetoram, another spinosyn, suggesting that altered target site may be the mechanism involved. Therefore, the suspension of spinosyn use against the tomato borer would be a useful component in spinosad resistance management for this species. Spinosad use against this species in introduced areas should be carefully monitored to prevent rapid selection of high levels of resistance and the potential for its spread to new areas.