Ralf Nauen

Overview of the Status and Global Strategy for Neonicotinoids

In recent years, neonicotinoid insecticides have been the fastest growing class of insecticides in modern crop protection, with widespread use against a broad spectrum of sucking and certain chewing pests. As potent agonists, they act selectively on insect nicotinic acetylcholine receptors (nAChRs), their molecular target site. The discovery of neonicotinoids can be considered as a milestone in insecticide research and greatly facilitates the understanding of functional properties of the insect nAChRs. In this context, the crystal structure of the acetylcholine-binding proteins provides the theoretical foundation for designing homology models of the corresponding receptor ligand binding domains within the nAChRs, a useful basis for virtual screening of chemical libraries and rational design of novel insecticides acting on these practically relevant channels. Because of the relatively low risk for nontarget organisms and the environment, the high target specificity of neonicotinoid insecticides, and their versatility in application methods, this important class has to be maintained globally for integrated pest management strategies and insect resistance management programs. Innovative concepts for life-cycle management, jointly with the introduction of generic products, have made neonicotinoids the most important chemical class for the insecticide market.

Applied aspects of neonicotinoid uses in crop protection.

Neonicotinoid insecticides comprise seven commercially marketed active ingredients: imidacloprid, acetamiprid, nitenpyram, thiamethoxam, thiacloprid, clothianidin and dinotefuran. The technical profiles and main differences between neonicotinoid insecticides, including their spectrum of efficacy, are described: use for vector control, systemic properties and versatile application forms, especially seed treatment. New formulations have been developed to optimize the bioavailability of neonicotinoids through improved rain fastness, better retention and spreading of the spray deposit on the leaf surface, combined with higher leaf penetration. Combined formulations with pyrethroids and other insecticides are also being developed with the aim of broadening the insecticidal spectrum of neonicotinoids and to replace WHO Class I products from older chemical classes. These innovative developments for life-cycle management, jointly with the introduction of generic products, will, within the next few years, turn neonicotinoids into the most important chemical class in crop protection.

Neonicotinoids—from zero to hero in insecticide chemistry

In recent years, neonicotinoids have been the fastest-growing class of insecticides in modern crop protection, with widespread use against a broad spectrum of sucking and certain chewing pests. As potent agonists, they act selectively on insect nicotinic acetylcholine receptors, their molecular target site. The discovery of neonicotinoids can be considered as a milestone in insecticide research and facilitates greatly the understanding of the functional properties of insect nicotinic acetylcholine receptors. Because of the relatively low risk for non-target organisms and environment, the high target specificity of neonicotinoid insecticides and their versatility in application methods, this important class has to be maintained globally for integrated pest management strategies and insect resistance management programmes. This review comprehensively describes particularly the origin, structure and bonding as well as associated properties of neonicotinoid insecticides.

Thiamethoxam is a neonicotinoid precursor converted to clothianidin in insects and plants

Neonicotinoid insecticides act agonistically on insect nicotinic acetylcholine receptors (nAChRs). Like imidacloprid (IMI), all neonicotinoids bind with high affinity (I 50 -values ∼ 1 nM) to [ 3 H]IMI binding sites on insect nAChRs. One notable omission is thiamethoxam (THIAM), showing binding affinities up to 10,000-fold less potent than other neonicotinoids, using housefly head membrane preparations. Clothianidin (CLOTHI) exhibits high activity as an agonist on isolated neurons at concentrations as low as 30 nM. Pharmacokinetic studies in different insect species revealed that THIAM was rapidly metabolized to CLOTHI, which shows high affinity to nAChRs in both binding assays and whole cell voltage clamp studies. When applied to cotton plants, THIAM was also quickly metabolized, with CLOTHI being the predominant neonicotinoid in planta briefly after application, as indicated by LC-MS/MS analyses. Our studies show that THIAM is likely to be a neonicotinoid precursor for CLOTHI and not active by itself.

Resistance of Bemisia tabaci (Homoptera: Aleyrodidae) to insecticides in southern Spain with special reference to neonicotinoids

The tobacco whitefly, Bemisia tabaci Gennadius (Homoptera: Aleyrodidae) which occurs in various parts of the world, has developed a high degree of resistance against several chemical classes of insecticide, including organophosphates, carbamates, pyrethroids, insect growth regulators and chlorinated hydrocarbons. The present studies were done in order to monitor the susceptibility of whitefly populations in southern Spain to insecticides commonly used there. Systemic bioassays using Spanish field populations of B tabaci collected in 1994, 1996 and 1998 indicated an increase, albeit a slow one, in resistance to imidacloprid over this period. Comparative studies of other neonicotinoids using the same bioassay revealed a high degree of cross-resistance to acetamiprid and thiamethoxam. Leaf-dip bioassays with adult females from these populations revealed a high level of resistance to cyfluthrin, endosulfan, monocrotophos, methamidophos, and pymetrozine, each at 200 mg litre−1. Buprofezin and pyriproxyfen were tested against second-instar nymphs and eggs, respectively. Buprofezin also showed a lower efficacy against ESP-98, a strain of B tabaci received from Almeria in 1998, but pyriproxyfen resistance was not obvious when tested against eggs of strain ESP-98. Field trials in 1998 revealed good efficacy of imidacloprid in one farm in the Almeria region and two greenhouses in Murcia and Sevilla, but a loss of activity by imidacloprid in another farm in the Almeria region. Cross-resistance between imidacloprid and thiamethoxam was also confirmed under field conditions. © 2000 Society of Chemical Industry

Over-expression of cytochrome P450 CYP6CM1 is associated with high resistance to imidacloprid in the B and Q biotypes of Bemisia tabaci (Hemiptera: Aleyrodidae).

The two most damaging biotypes of Bemisia tabaci, B and Q, have both evolved strong resistance to the neonicotinoid insecticide imidacloprid. The major mechanism in all samples investigated so far appeared to be enhanced detoxification by cytochrome P450s monooxygenases (P450s). In this study, a polymerase chain reaction (PCR) technology using degenerate primers based on conserved P450 helix I and heme-binding regions was employed to identify P450 cDNA sequences in B. tabaci that might be involved in imidacloprid resistance. Eleven distinct P450 cDNA sequences were isolated and classified as members of the CYP4 or CYP6 families. The mRNA expression levels of all 11 genes were compared by real-time quantitative RT-PCR across nine B and Q field-derived strains of B. tabaci showing strong resistance, moderate resistance or susceptibility to imidacloprid. We found that constitutive over-expression (up to approximately 17-fold) of a single P450 gene, CYP6CM1, was tightly related to imidacloprid resistance in both the B and Q biotypes. Next, we identified three single-nucleotide polymorphic (SNP) markers in the intron region of CYP6CM1 that discriminate between the resistant and susceptible Q-biotype CYP6CM1 alleles (r-Q and s-Q, respectively), and used a heterogeneous strain to test for association between r-Q and resistance. While survivors of a low imidacloprid dose carried both the r-Q and s-Q alleles, approximately 95% of the survivors of a high imidacloprid dose carried only the r-Q allele. Together with previous evidence, the results reported here identify enhanced activity of P450s as the major mechanism of imidacloprid resistance in B. tabaci, and the CYP6CM1 gene as a leading target for DNA-based screening for resistance to imidacloprid and possibly other neonicotinoids in field populations.

Baseline determination and detection of resistance to imidacloprid in Bemisia tabaci (Homoptera: Aleyrodidae)

Adult Bemisia tabaci (Gennadius) were tested with imidacloprid in a bioassay method using systemically treated cotton leaves. The method was simple, robust and repeatable and provided baseline data for a laboratory and a number of recently collected field strains. The LC 50 for imidacloprid susceptible strains was calculated to be 1.7 ppm and a concentration of 16 ppm determined as diagnostic for imidacloprid resistance. Ten strains of B. tabaci collected from the Almeria region of Spain showed significantly less mortality at the diagnostic dose than the susceptible strains. The intensive use of imidacloprid in the Almeria region is considered the reason for the occurrence of resistance in this locality. Resistance to organophosphates, pyrethroids and endosulfan in B-type and non-B type B. tabaci did not confer resistance to imidacloprid.

IRAC: Mode of action classification and insecticide resistance management

Insecticide resistance is a long standing and expanding problem for pest arthropod control. Effective insecticide resistance management (IRM) is essential if the utility of current and future insecticides is to be preserved. Established in 1984, the Insecticide Resistance Action Committee (IRAC) is an international association of crop protection companies. IRAC serves as the Specialist Technical Group within CropLife International focused on ensuring the long term efficacy of insect, mite and tick control products through effective resistance management for sustainable agriculture and improved public health. A key function of IRAC is the continued development of the Mode of Action (MoA) classification scheme, which provides up-to-date information on the modes of action of new and established insecticides and acaricides and which serves as the basis for developing appropriate IRM strategies for crop protection and vector control. The IRAC MoA classification scheme covers more than 25 different modes of action and at least 55 different chemical classes. Diversity is the spice of resistance management by chemical means and thus it provides an approach to IRM providing a straightforward means to identify potential rotation/alternation options.

Cross-Resistance, Inheritance, and Biochemistry of Mitochondrial Electron Transport Inhibitor-Acaricide Resistance in Tetranychus urticae (Acari: Tetranychidae)

Abstract Resistance of the twospotted spider mite, Tetranychus urticae Koch, to acaricides acting as mitochondrial electron transport inhibitors (METIs) is an increasing problem. Because of their high levels of cross-resistance to all commercially available METI-acaricides, a Japanese strain (AKITA) and an English strain (UK-99) of T. urticae were investigated in detail. Larvae of both strains, AKITA and UK-99, showed 1,100- and 480-fold resistance against pyridaben, 870- and 45-fold resistance against fenpyroximate, and 33- and 44-fold resistance against tebufenpyrad, respectively, in a foliar spray application bioassay compared with the susceptible strain GSS. These resistance factors remained stable even when maintained in the laboratory without further selection. Furthermore, strain AKITA showed cross-resistance to dicofol. The METI resistant strains AKITA and UK-99 showed 2.4- and 1.7-fold enhanced 7-ethoxycoumarin O-deethylation (cytochrome P450) activity. Increased oxidative metabolism of the METI-acaricides in the resistant strains could be partially suppressed in vivo by the monooxygenase-inhibitor piperonyl butoxide. Reciprocal crosses of homozygous, diploid females and hemizygous, haploid males of strains GSS (susceptible) and AKITA (resistant) revealed that resistance to pyridaben and fenpyroximate was inherited incompletely dominant with slight differences between maternal and paternal inheritance. This is the first attempt to mechanistically describe METI-acaricide resistance in T. urticae. The implications for resistance management strategies are discussed.

Efficacy of plant metabolites of imidacloprid against Myzus persicae and Aphis gossypii (Homoptera: Aphididae)

The metabolism of the chloronicotinyl insecticide imidacloprid is strongly influenced by the method of application. Whilst in foliar application most of the residues on the leaf surface display unchanged parent compound, most of the imidacloprid administered to plants by soil application or seed treatment is metabolized more or less completely, depending on plant species and time. The present study revealed that certain metabolites of imidacloprid which have been described in crop plants are highly active against aphid pests in different types of bioassays. Some of these metabolites showed a high oral activity against the green peach aphid (Myzus persicae), and the cotton aphid (Aphis gossypii). The aphicidal potency of the metabolites investigated was weaker in aphid dip tests than in oral ingestion bioassays using artificial double membranes. The most active plant metabolite was the imidazoline derivative of imidacloprid. The LC50 values of this metabolite for M. persicae and A. gossypii in oral ingestion bioassays were in the lower ppb-range, i.e. 0·0044 and 0·0068 mg litre-1, respectively. Most of the other reported metabolites showed much weaker activity. Compared to imidacloprid, the imidazoline derivative showed superior affinity to housefly (Musca domestica) head nicotinic acetylcholine receptors, while all other metabolites were less specific than imidacloprid. It seems possible that, after seed treatment or soil application, a few of the biologically active metabolites arising are acting in concert with remaining levels of the parent compound imidacloprid, thus providing good control and long-lasting residual activity against plant-sucking pests in certain crops.