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Lygus spp. are polyphagous pests that overwinter in weedy vegetation. In the spring on the central coast of California, Lygus spp. emigrate from weeds into strawberry fields. Subsequent feeding on strawberry flowers causes fruit deformation that precludes sale on the fresh market. Use of alfalfa (Medicago sativa L.) (Fabales: Fabaceae) as a trap crop has been hypothesized to prevent Lygus spp. colonization in strawberries. We examined the movement of Lygus spp. and associated predators from weeds to strawberry fields with alfalfa trap crops using a protein mark–capture technique. Insects and spiders were collected from weeds, strawberry, and alfalfa 1 day, 2 days, and ∼2 wk after an albumin protein mark was applied to weeds bordering strawberry fields. For marked Lygus spp. that emigrated from weeds, the majority (79%) of adults were recovered from alfalfa trap crops; however, all nymphs immigrated to strawberry. Most protein-marked predators immigrated to strawberry, rather than trap crops, resulting in a marked predator-to-Lygus spp. ratio of 5:1. Trap cropping effectively reduced the colonization of Lygus adults in strawberry. Converting weedy areas to native perennial plantings could further mitigate the risk of pest migration, while simultaneously conserving beneficial insects. [ABSTRACT FROM AUTHOR]

In natural habitats, there is a strong evolutionary selection pressure on herbivorous insects to avoid danger and choose suitable host plants. Similar selection pressures may drive movement and choices of oviposition hosts by herbivorous insects living in agricultural cropping systems, in which insecticides are often used. In this study, we quantified movement responses and nymph emergence (collectively referred to as bio-responses) of western-tarnished plant bug (Lygus hesperus Knight (Hemiptera: Miridae)) individuals when exposed to environments associated with a perceived "risk" (experimental insecticide treatments and their corresponding controls). We introduce a novel analytical approach in which treatments (risk environments) are ranked in ascending order based on bio-responses (movement or nymph emergence). Consequently, linear regression coefficients were generated and used to interpret bio-responses of Lygus individuals in different life stages to risk environments. Initially, we predicted movement by Lygus individuals to be positively associated with environmental risk and nymph emergence to be negatively associated with environmental risk. Overall, based on a comprehensive combination of no- and two-choice bioassays, we found that: (1) In no-choice bioassays, movement parameters (both total distance moved and movement percentage) by all three life stages were lowest in low-risk environments and highest when Lygus individuals were exposed to either malathion or Grandevo. Accordingly, environments involving malathion or Grandevo were considered high-risk. (2) No-choice movement bioassays also revealed that Lygus males moved significantly more (based on comparison of regression intercepts) than other life stages, and that they responded significantly more (based on comparison of regression slopes) than conspecific females and nymphs. (3) In two-choice movement bioassays, neem elicited the most consistent movement responses by Lygus individuals, and adult life stages showed the strongest response. Two-choice movement bioassays also revealed that Lygus adults, compared to nymphs, were more likely to spend time in low-risk areas of the test arenas. (4) Nymph emergence was markedly lower in no-choice compared to two-choice bioassays, and in two-choice bioassays, Grandevo and malathion elicited especially biased nymph emergence from low-risk beans. To our knowledge, this is the first study in which movement bioassays have been used to quantify and characterize behavioral responses by Lygus life stages to environments associated with varying degrees of risk. The novel analytical approach presented in this study provides a high degree of complementarity to more traditional performance-testing methods used to evaluate responses to insecticides. Furthermore, we believe that this analytical approach can be of considerable relevance to studies of animal phenomics and behavioral studies of animals more broadly, in which adaptation and fitness parameters are examined in response to environmental risk and heterogeneity. [ABSTRACT FROM AUTHOR]

Based on multiple sequence alignments (Figures S1-S6), the I L. hesperus i melanin pathway proteins exhibit 60-92% similarity with homologs from other species ( I D. melanogaster i , I Bombyx mori i , I O. fasciatus i , and I Halyomorpha halys i ). Keywords: cuticular pigmentation; melanin pathway; Lygus hesperus; RNAi EN cuticular pigmentation melanin pathway Lygus hesperus RNAi 986 15 11/17/22 20221101 NES 221101 1. Because mutations to these pigmentation-associated genes can result in varied impacts on phenotype when comparing between stages and sexes, and across species, it is necessary to elucidate the role of each gene to determine which might be suitable as a distinctive marker of transgenesis in I L. hesperus i . Table S1: Top 5 tBLASTn hits of I Lygus hesperus i TSA datasets with defined insect melanin pathway genes; Table S2: Accession numbers for protein sequences used in phylogenetic analyses; Table S3: Oligonucleotide primers used in this study; Table S4: Top 5 BLASTx hits of Lygus hesperus melanin-associated transcripts; Table S5: Transcript metrics and protein domain analysis of melanin pathway associated proteins in I Lygus hesperus i and I Drosophila melanogaster i . RT-PCR Cloning and Expression Profiling Primers designed to amplify complete ORFs for the I L. hesperus i melanin pathway genes generated PCR products of the expected sizes (Figure 3). [Extracted from the article]

In natural habitats, there is a strong evolutionary selection pressure on herbivorous insects to avoid danger and choose suitable host plants. Similar selection pressures may drive movement and choices of oviposition hosts by herbivorous insects living in agricultural cropping systems, in which insecticides are often used. In this study, we quantified movement responses and nymph emergence (collectively referred to as bio-responses) of western-tarnished plant bug (Lygus hesperus Knight (Hemiptera: Miridae)) individuals when exposed to environments associated with a perceived “risk” (experimental insecticide treatments and their corresponding controls). We introduce a novel analytical approach in which treatments (risk environments) are ranked in ascending order based on bio-responses (movement or nymph emergence). Consequently, linear regression coefficients were generated and used to interpret bio-responses of Lygus individuals in different life stages to risk environments. Initially, we predicted movement by Lygus individuals to be positively associated with environmental risk and nymph emergence to be negatively associated with environmental risk. Overall, based on a comprehensive combination of no- and two-choice bioassays, we found that: (1) In no-choice bioassays, movement parameters (both total distance moved and movement percentage) by all three life stages were lowest in low-risk environments and highest when Lygus individuals were exposed to either malathion or Grandevo. Accordingly, environments involving malathion or Grandevo were considered high-risk. (2) No-choice movement bioassays also revealed that Lygus males moved significantly more (based on comparison of regression intercepts) than other life stages, and that they responded significantly more (based on comparison of regression slopes) than conspecific females and nymphs. (3) In two-choice movement bioassays, neem elicited the most consistent movement responses by Lygus individuals, and adult life stages showed the strongest response. Two-choice movement bioassays also revealed that Lygus adults, compared to nymphs, were more likely to spend time in low-risk areas of the test arenas. (4) Nymph emergence was markedly lower in no-choice compared to two-choice bioassays, and in two-choice bioassays, Grandevo and malathion elicited especially biased nymph emergence from low-risk beans. To our knowledge, this is the first study in which movement bioassays have been used to quantify and characterize behavioral responses by Lygus life stages to environments associated with varying degrees of risk. The novel analytical approach presented in this study provides a high degree of complementarity to more traditional performance-testing methods used to evaluate responses to insecticides. Furthermore, we believe that this analytical approach can be of considerable relevance to studies of animal phenomics and behavioral studies of animals more broadly, in which adaptation and fitness parameters are examined in response to environmental risk and heterogeneity.

Cuticle coloration in insects is a consequence of the accumulation of pigments in a species-specific pattern. Numerous genes are involved in regulating the underlying processes of melanization and sclerotization, and their manipulation can be used to create externally visible markers of successful gene editing. To clarify the roles for many of these genes and examine their suitability as phenotypic markers in Lygus hesperus Knight (western tarnished plant bug), transcriptomic data were screened for sequences exhibiting homology with the Drosophila melanogaster proteins. Complete open reading frames encoding putative homologs for six genes (aaNAT, black, ebony, pale, tan, and yellow) were identified, with two variants for black. Sequence and phylogenetic analyses supported preliminary annotations as cuticle pigmentation genes. In accord with observable difference in color patterning, expression varied for each gene by developmental stage, adult age, body part, and sex. Knockdown by injection of dsRNA for each gene produced varied effects in adults, ranging from the non-detectable (black 1, yellow), to moderate decreases (pale, tan) and increases (black 2, ebony) in darkness, to extreme melanization (aaNAT). Based solely on its expression profile and highly visible phenotype, aaNAT appears to be the best marker for tracking transgenic L. hesperus.

Each year, consultants and field scouts working in commercial agriculture undertake a massive, decentralized data collection effort as they monitor insect populations to make real-time pest management decisions. These data, if integrated into a database, offer rich opportunities for applying big data or ecoinformatics methods in agricultural entomology research. However, questions have been raised about whether or not the underlying quality of these data is sufficiently high to be a foundation for robust research. Here I suggest that repeatability analysis can be used to quantify the quality of data collected from commercial field scouting, without requiring any additional data gathering by researchers. In this context, repeatability quantifies the proportion of total variance across all insect density estimates that is explained by differences across populations and is thus a measure of the underlying reliability of observations. Repeatability was moderately high for cotton fields scouted commercially for total Lygus hesperus Knight densities (R = 0.631) and further improved by accounting for observer effects (R = 0.697). Repeatabilities appeared to be somewhat lower than those computed for a comparable, but much smaller, researcher-generated data set. In general, the much larger sizes of ecoinformatics data sets are likely to more than compensate for modest reductions in measurement precision. Tools for evaluating data quality are important for building confidence in the growing applications of ecoinformatics methods. [ABSTRACT FROM AUTHOR]

Although surveys of pest populations documenting evolved insecticide resistance often suggest abundant potential for insecticide control failures, studies documenting the actual occurrence of such failures in commercial agriculture are rare. If farmers currently practice adaptive management, abandoning the use of insecticides once resistance emerges, actual control failures could be rare. Here I use data gathered by independent pest management consultants to describe a case study of the realized efficacy of commercial field applications of insecticides, examining the control of Lygus hesperus Knight on cotton. On average, insecticides reduced target pest populations to 19% of their preapplication densities. Short-term efficacy of insecticides was variable, but only one severe control failure was observed (1 of 50, 2%). The rarity of severe control failures observed in this study is in agreement with the few other studies conducted in commercial settings, but additional research is needed to assess the generality of this result. Although pesticides can cause longer-term problems, including target pest resurgences and secondary pest outbreaks, risk-averse attitudes among farmers coupled with relatively consistent short-term insecticide efficacy may be potent forces propelling farmers toward the use of insecticides. [ABSTRACT FROM AUTHOR]

Conservation biological control is a fundamental tactic in integrated pest management (IPM). Greater biological control services can be achieved by enhancing agroecosystems to be more favorable to the presence, survival, and growth of natural enemy populations. One approach that has been tested in numerous agricultural systems is the deployment of synthetic chemicals that mimic those produced by the plant when under attack by pests. These signals may attract arthropod natural enemies to crop habitats and thus potentially improve biological control activity locally. A 2-yr field study was conducted in the cotton agroecosystem to evaluate the potential of synthetic methyl salicylate (MeSA) to attract native arthropod natural enemies and to enhance biological control services on two key pests. Slow-release packets of MeSA were deployed in replicated cotton plots season long. The abundance of multiple taxa of natural enemies and two major pests were monitored weekly by several sampling methods. The deployment of MeSA failed to increase natural enemy abundance and pest densities did not decline. Predator to prey ratios, used as a proxy to estimate biological control function, also largely failed to increase with MeSA deployment. One exception was a season-long increase in the ratio of Orius tristicolor (White) (Hemiptera: Anthocoridae) to Bemisia argentifolii Bellows and Perring (= Bemisia tabaci MEAM1) (Hemiptera: Aleyrodidae) adults within the context of biological control informed action thresholds. Overall results suggest that MeSA would not likely enhance conservation biological control by the natural enemy community typical of U.S. western cotton production systems. [ABSTRACT FROM AUTHOR]

The parasitoid Peristenus relictus was established in California to help regulate Lygus species. This study surveyed the San Joaquin Valley and South Coast of California to determine this parasitoid's current southern geographic range. Peristenus relictus recoveries were made in all sampled counties, including Santa Barbara and Ventura along the coast, and Tulare, Kings and Kern inland. Parasitism was notably high in Santa Barbara County, where 61.7% of Lygus nymphs collected from weedy plants were parasitised. However, as parasitism was scarce in the San Joaquin Valley (0–12.5%), future research should examine possible inhibitors (e.g. climate) of this biological control. [ABSTRACT FROM AUTHOR]

Alfalfa, Medicago sativa L. (Fabaceae), is a highly preferred host plant of Lygus spp. (Hemiptera: Miridae). As such, intercropping alfalfa trap‐crops in strawberry production can serve as a sink for both Lygus (primarily Lygus hesperus Knight) and its natural enemies. Here we investigated the population dynamics and dispersal characteristics of the generalist predator complex in strawberry fields with alfalfa trap‐crops spaced 50 rows (62 m) apart. Predator abundance was determined by counting six focal taxa collected from strawberry and alfalfa. The data revealed that Orius spp. (Hemiptera: Anthocoridae) were the numerically dominant predator taxa, comprising 84% of the focal predator population. In general, the population densities obtained for the various taxa throughout this agroecosystem were unexpectedly uniform. Predator movement from a central alfalfa trap‐crop row was determined using a protein mark–capture procedure. Most protein‐marked predator specimens were collected less than 2 m from the centrally marked alfalfa row, indicating that the trap‐crop often produces a predator sink. Results suggest that alfalfa is a useful cultural (trap‐cropping) and a biological (refuge for natural enemies) control tactic for managing Lygus spp. in strawberries. [ABSTRACT FROM AUTHOR]

[Display omitted] • Lygus hesperus has 7 β-tubulin genes. • β-tubulin2 is predominantly expressed in the testes. • Knockdown or knockout of β-tubulin2 reduces both spermatozoa abundance and quality, which limits fertilization and the subsequent egg hatch. Lygus hesperus Knight is an important insect pest of crops across western North America, with field management heavily reliant on the use of chemical insecticides. Because of the evolution of resistance to these insecticides, effective and environmentally benign pest management strategies are needed. Traditional sterile insect technique (SIT) has been successfully employed to manage or eradicate some insect pests but involves introducing irradiated insects with random mutations into field populations. New genetically-driven SIT techniques are a safer alternative, causing fixed mutations that manipulate individual genes in target pests to produce sterile individuals for release. Here, we identified seven β-tubulin coding genes from L. hesperus and show that Lhβtub2 is critical in male sperm production and fertility. Lhβtub2 is expressed primarily in the male testes and targeting of this gene by RNA interference or gene editing leads to male sterility. [ABSTRACT FROM AUTHOR]

The pyrokinin (PK) family of insect neuropeptides, characterized by C termini consisting of either WFGPRLamide (i.e., PK1) or FXPRLamide (i.e., PK2), are encoded on the capa and pk genes. Although implicated in diverse biological functions, characterization of PKs in hemipteran pests has been largely limited to genomic, transcriptomic, and/or peptidomic datasets. The Lygus hesperus (western tarnished plant bug) PK transcript encodes a prepropeptide predicted to yield three PK2 FXPRLamide-like peptides with C-terminal sequences characterized by FQPRSamide (LyghePKa), FAPRLamide (LyghePKb), and a non-amidated YSPRF. The transcript is expressed throughout L. hesperus development with greatest abundance in adult heads. PRXamide-like immunoreactivity, which recognizes both pk- and capa-derived peptides, is localized to cells in the cerebral ganglia, gnathal ganglia/suboesophageal ganglion, thoracic ganglia, and abdominal ganglia. Immunoreactivity in the abdominal ganglia is largely consistent with capa-derived peptide expression, whereas the atypical fourth pair of immunoreactive cells may reflect pk-based expression. In vitro activation of a PK receptor heterologously expressed in cultured insect cells was only observed in response to LyghePKb, while no effects were observed with LyghePKa. Similarly, in vivo pheromonotropic effects were only observed following LyghePKb injections. Comparison of PK2 prepropeptides from multiple hemipterans suggests mirid-specific diversification of the pk gene.

A prey immunomarking procedure (PIP) in combination with generic anti-rabbit and anti-chicken immunoglobulin G (IgG) enzyme-linked immunosorbent assays (ELISAs) are used frequently to study arthropod predation. This study was conducted to: (1) further standardize the PIP as a tool for predator gut analysis research, (2) investigate the most effective means for administering IgG marks to prey items, and (3) assess the possibility of the PIP yielding false positive reactions as a consequence of a predator obtaining a mark by incidental contact with, or by a failed predation attempt on, a protein-marked prey item. The pest Lygus hesperus Knight (Hemiptera: Miridae) was tagged with either an external rabbit IgG mark, an internal chicken IgG mark, or a double (external rabbit IgG and internal chicken IgG) mark treatment. Then, the variously marked prey items were fed to chewing and piercing-sucking type predators and their gut contents were examined for the presence of IgG remains. Data revealed that all three marking treatments were highly effective at tagging targeted prey. However, ELISA results showed that the prey items should only be marked internally to maximize the likelihood of detecting prey remains while minimizing the risk of obtaining false positive errors. The merits and limitations of using the generic PIP for predator gut analysis research are discussed. [ABSTRACT FROM AUTHOR]

Lygus hesperus Knight (Hemiptera: Miridae) is an important pest of strawberry in the central coast of California. Both adults and nymphs of L. hesperus feed on developing strawberry fruits causing cat-faced fruits (unmarketable). Although multiple tactics, such as vacuuming with tractor-mounted vacuum machine or the application of an insecticide, are used to manage L. hesperus , the combined effects of these tactics have not been thoroughly examined. Furthermore, the use of an electrostatic sprayer for L. hesperus management has not been tested. The objectives of the present study were to determine 1) the individual and combined effects of sulfoxaflor insecticide and vacuuming on L. hesperus and their feeding damage, and 2) the efficacy of sulfoxaflor when sprayed using an electrostatic sprayer at various rates of water volume against L. hesperus . The results showed that the vacuum alone did not reduce L. hesperus populations to untreated control (UTC). The treatments that involved sulfoxaflor, both individually and when combined with the vacuum reduced the number of L. hesperus and percentage cat-faced fruits compared to UTC. Neither the “bug vac” nor the sulfoxaflor had any impact on the capture of predaceous heteropterans and spiders compared to UTC. In various water volume treatments applied using electrostatic sprayer, the number of L. hesperus nymphs and percentage of cat-face fruits was significantly lower than in UTC treatment and was comparable to conventional sprayer. The numbers of predaceous heteropteran and spider captures were not significantly different between the treatments applied using electrostatic versus conventional sprayers. These results implies that effective insecticide will continue to the best tactic to reduce L. hesperus and damage. Use of electrostatic sprayer has a potential whereas, vacuum method will need further refinement for L. hesperus control. [ABSTRACT FROM AUTHOR]

Abstract: There is frequently a tradeoff between fecundity and longevity, but the relationship is inconsistent across species and influenced by various exogenous and endogenous factors. Previous studies of Lygus hesperus Knight (Hemiptera: Miridae) established that egg production is promoted by insemination, at least temporarily, but little is known about the long‐term effects of mating and nonsexual interactions with conspecifics on egg production and female lifespan. To elucidate these relationships, survivorship and oviposition rate were tracked daily in females that were isolated or paired with a fertile male or another female throughout their adult lives. Mating rates were determined by postmortem examination. Results indicate that male‐specific stimuli accelerate female reproductive maturation, and that mating elevates oviposition rate. However, females paired with either a female or male companion had shortened lifespans, suggesting that social contacts exact a significant cost in this solitary species. Despite the negative impact of conspecific interactions and the finding that a singly mated female has sufficient sperm to fertilize a lifetime supply of eggs, many females were found to have mated more than once. Multiply mated females had higher sustained oviposition rates, lived longer, and had greater lifetime fecundities. Collectively, no strong evidence was found of a direct physiological link between fecundity and longevity, but environmental factors and mating were found to significantly influence both traits. [ABSTRACT FROM AUTHOR]

The western tarnished plant bug, Lygus hesperus Knight, overwinters in an adult diapause. However, the ecological implications of diapause to L. hesperus population survival are poorly understood. Enhanced understanding of L. hesperus overwintering ecology might reveal opportunities to develop ecologically-based management tactics. One factor limiting efforts to understand the dynamics of overwintering is lack of a non-destructive method to distinguish diapause. We evaluated a suite of external characters for utility in distinguishing diapausing from non-diapausing L. hesperus. Abdomen coloration of female L. hesperus was a highly reliable indicator of diapause status. During these studies, a single female (0.6%) was misclassified using the abdomen color criterion. For male L. hesperus, corrected abdomen length (abdomen length / head capsule width) was used to predict diapause status with ?84% accuracy. Both criteria provided improved accuracy compared with earlier reports, in part because their application was limited to adults of a specific age (10 d) and reared at a specific temperature (26.7°C). Application of these criteria to individuals that were subsequently starved allowed us to unambiguously distinguish the survival functions of diapausing and non-diapausing males and females. Although neither criterion (abdomen color for females, corrected abdomen length for males) was error-free, both offer sufficient accuracy to justify their use in studies of the ecology, physiology, or molecular biology of L. hesperus diapause. These criteria provide the ability to non-destructively distinguish the diapause status of adult L. hesperus with reasonable accuracy, and should enhance efforts to better understand diapause and overwintering ecology in this important pest species. [ABSTRACT FROM AUTHOR]

Passive mechanisms of mate guarding are used by males to promote sperm precedence with little cost, but these tactics can be disadvantageous for their mates and other males. Mated females of the plant bug Lygus hesperus are rendered temporarily unattractive by seminal fluids containing myristyl acetate and geranylgeranyl acetate. These antiaphrodisiac pheromones are gradually released from the female’s gonopore, declining until they no longer suppress male courtship. Because starting quantities of these compounds can vary widely, the repellant signal becomes less reliable over time. Evidence was found of a complimentary mechanism that more accurately conveys female mating status. Once inside the female, geranylgeranyl acetate is progressively converted to geranylgeraniol then externalized. Geranylgeraniol counteracts the antiaphrodisiac effect despite having no inherent attractant properties of its own. This is the first evidence for such an anti-antiaphrodisiac pheromone, adding a new element to the communication mechanisms regulating reproductive behaviors.

The Gα subunits of heterotrimeric G proteins play critical roles in the activation of diverse signal transduction cascades. However, the role of these genes in chemosensation remains to be fully elucidated. To initiate a comprehensive survey of signal transduction genes, we used homology-based cloning methods and transcriptome data mining to identity Gα subunits in the western tarnished plant bug (Lygus hesperus Knight). Among the nine sequences identified were single variants of the Gαi, Gαo, Gαs, and Gα12 subfamilies and five alternative splice variants of the Gαq subfamily. Sequence alignment and phylogenetic analyses of the putative L. hesperus Gα subunits support initial classifications and are consistent with established evolutionary relationships. End-point PCR-based profiling of the transcripts indicated head specific expression for LhGαq4, and largely ubiquitous expression, albeit at varying levels, for the other LhGα transcripts. All subfamilies were amplified from L. hesperus chemosensory tissues, suggesting potential roles in olfaction and/or gustation. Immunohistochemical staining of cultured insect cells transiently expressing recombinant His-tagged LhGαi, LhGαs, and LhGαq1 revealed plasma membrane targeting, suggesting the respective sequences encode functional G protein subunits.

The western tarnished plant bug (Lygus hesperus Knight) (Hemiptera: Miridae) is a polyphagous pest of numerous western crops. This pest overwinters in a relatively short duration adult diapause, but many details regarding diapause induction and maintenance remain unstudied. Instar-specific responses to a switch from short (10-h) to long (14-h) days, long to short days, and a temporary (4-d) switch from short to long days were compared with responses of insects maintained under respective short- and long-day photoperiods. Influences of short days received during the nymphal stage on subsequent adult reproductive development under long days were also examined. Substantial diapause responses were observed only for insects switched to short days by 4th instar. Few insects that switched from short to long days exhibited diapause characters by Day 10 of adulthood. When 2nd instars were temporarily switched from short to long days no effect on the diapause response was observed, but the response was diminished when the switch occurred beginning at 4th instar. Reproductive development of adults reared under short days and switched to long days was modestly delayed compared with insects reared from egg to adult under long days. These results indicate the diapause response is elicited by short day cues received prior to the 5th instar, and that continued exposure to short days is necessary to maintain the adult diapause. These findings, combined with earlier reports of diapause termination in the field, suggest that southern populations of L. hesperus may express a more dynamic and complex overwintering strategy than was previously recognized. [ABSTRACT FROM AUTHOR]

Many strategies for controlling insect pests require an understanding of their hormonal signaling agents, peptides being the largest and most diverse single class of these molecules. Lygus hesperus is a pest species of particular concern, as it is responsible for significant damage to a wide variety of commercially important plant crops. At present, little is known about the peptide hormones of L. hesperus . Here, transcriptomic data were used to predict a peptidome for L. hesperus . Fifty-three L. hesperus transcripts encoding peptide precursors were identified, with a subset amplified by PCR for sequence verification. The proteins deduced from these transcripts allowed for the prediction of a 119-sequence peptidome for L. hesperus . The predicted peptides include isoforms of allatostatin A, allatostatin B (AST-B), allatostatin C, allatotropin, bursicon, CCHamide, corazonin, crustacean cardioactive peptide, crustacean hyperglycemic hormone/ion transport peptide, diuretic hormone 31, GSEFLamide, insulin-like peptide, myosuppressin, neuroparsin, neuropeptide F, orcokinin, orcomyotropin, pyrokinin, short neuropeptide F, SIFamide, sulfakinin and tachykinin-related peptide. Of note were several isoforms of AST-B that possess –W X 7 Wamide carboxyl-termini rather than the stereotypical –W X 6 Wamide ( e.g. , KWQDMQNPGWamide), an allatotropin ending in –SARGFamide rather than –TARGFamide (GLKNGPLNSARGFamide), a GSEFLamide ending in –GTEFLamide (TVGTEFLamide), several orcokinins with PMDEIDR– rather than NFDEIDR– amino-termini ( e.g. , PMDEIDRAGFTHFV), and an eight rather than 12 amino acid long isoform of SIFamide (PPFNGSIFamide). Collectively, the L. hesperus peptidome predicted here provides a resource for initiating physiological investigations of peptidergic signaling in this species, including studies directed at the biological control of this agricultural pest. [ABSTRACT FROM AUTHOR]

Understanding the population dynamics of herbivorous insects is critical to developing and implementing effective pest control protocols. In the context of inverse problems, we explore the dynamic effects of pesticide treatments onLygus hesperus, a common pest of cotton in the western United States. Fitting models to field data, we explore the topic of model selection for an appropriate mathematical model and corresponding statistical models, and use techniques including ANOVA-based model comparison tests and residual plot analysis to make the best selections. In addition we explore the topic of data information content: in this example, we are testing the question of whether data, as it is currently collected, can support time-dependent parameter estimation. Furthermore, we investigate the statistical assumptions often haphazardly made in the process of parameter estimation and consider the implications of unfounded assumptions. [ABSTRACT FROM AUTHOR]

Background: Departures from the standard genetic code in eukaryotic nuclear genomes are known for only a handful of lineages and only a few genetic code variants seem to exist outside the ciliates, the most creative group in this regard. Most frequent code modifications entail reassignment of the UAG and UAA codons, with evidence for at least 13 independent cases of a coordinated change in the meaning of both codons. However, no change affecting each of the two codons separately has been documented, suggesting the existence of underlying evolutionary or mechanistic constraints. Results: Here, we present the discovery of two new variants of the nuclear genetic code, in which UAG is translated as an amino acid while UAA is kept as a termination codon (along with UGA). The first variant occurs in an organism noticed in a (meta)transcriptome from the heteropteran Lygus hesperus and demonstrated to be a novel insectdwelling member of Rhizaria (specifically Sainouroidea). This first documented case of a rhizarian with a non-canonical genetic code employs UAG to encode leucine and represents an unprecedented change among nuclear codon reassignments. The second code variant was found in the recently described anaerobic flagellate Iotanema spirale (Metamonada: Fornicata). Analyses of transcriptomic data revealed that I. spirale uses UAG to encode glutamine, similarly to the most common variant of a non-canonical code known from several unrelated eukaryotic groups, including hexamitin diplomonads (also a lineage of fornicates). However, in these organisms, UAA also encodes glutamine, whereas it is the primary termination codon in I. spirale. Along with phylogenetic evidence for distant relationship of I. spirale and hexamitins, this indicates two independent genetic code changes in fornicates. Conclusions: Our study documents, for the first time, that evolutionary changes of the meaning of UAG and UAA codons in nuclear genomes can be decoupled and that the interpretation of the two codons by the cytoplasmic translation apparatus is mechanistically separable. The latter conclusion has interesting implications for possibilities of genetic code engineering in eukaryotes. We also present a newly developed generally applicable phylogeny-informed method for inferring the meaning of reassigned codons. [ABSTRACT FROM AUTHOR]

The western tarnished plant bug, Lygus hesperus Knight, is a key cotton (Gossypium spp.) pest, managed primarily by application of insecticides according to nominal thresholds. Efforts to reduce reliance on insecticide-based tactics will require a more astute understanding of the physiological ecology of L. hesperus than is currently available. A key biological parameter that may be manipulated through cultural or genetic means is reproduction. Estimates of L. hesperus oviposition are commonly obtained by visual inspection, but the accuracy of those estimates has been questioned. Because the eggs swell during development, we hypothesized that delaying counts of eggs to permit some development might improve sampling accuracy. Estimates of L. hesperus egg numbers were obtained immediately after oviposition and again at three days after oviposition, and counts at both times were regressed with total eggs determined by the sum of either hatched eggs or the number of nymphs, and unhatched eggs. Sampler experience was a major factor influencing fidelity of egg counts with total eggs, but a single repetition of the experiment accompanied by knowledge of the results was sufficient to optimize sampler effectiveness. Regressions relating total eggs to counts, whether the counts were taken immediately after oviposition or after three days, were significant. However, compared with counts of newly-laid eggs, the regression relating egg counts at three days with total eggs was more stable and showed better agreement. These results can be used to maximize statistical power and minimize sampling errors in future examinations of L. hesperus oviposition [ABSTRACT FROM AUTHOR]

BACKGROUND Landscape crop composition surrounding agricultural fields is known to affect the density of crop pests, but quantifying these effects, as well as measuring how they translate to changes in yield, is difficult. Using a large dataset consisting of 1498 records of commercial cotton production in California between 1997 and 2008, we explored the relationship between landscape composition and cotton yield, the density of Lygus hesperus (a key cotton pest) at field-level and within-field spatial scales and pesticide use. RESULTS We found that the crop composition immediately adjacent to a cotton field was associated with substantial differences in cotton yield, L. hesperus density and pesticide use. Furthermore, crops that tended to be associated with increased L. hesperus density also tended to be associated with increased pesticide use and decreased cotton yield. CONCLUSION Our results suggest a possible mechanism by which landscape composition can affect cotton yield: by increasing the density of pests which in turn damage cotton plants. Our quantification of how surrounding crops affect pest densities, and in turn yield, in cotton fields has significant impacts for cotton farmers, who can use this information to help optimize crop selection and ranch layout. © 2016 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

California strawberries have two major arthropod pests—the twospotted spider mite, Tetranychus urticae and the western tarnished plant bug, Lygus hesperus, which result in significant losses to the yield and quality of marketable berries. Other important insect pests that are frequently seen in strawberry include the greenhouse whitefly, Trialeurodes vaporariorum and the western flower thrips, Frankliniella occidentalis that cause varying levels of damage depending on the level of infestation. Chemical pesticides play a major role in managing these pests but not without the associated risk of pesticide resistance and environmental safety. Two field studies were conducted in commercial strawberry fields in Santa Maria, one of the strawberry growing areas in California Central Coast, to determine the efficacy of chemical, botanical and microbial pesticides in the integrated pest management (IPM) of strawberry. Chemical, botanical and microbial pesticides were evaluated against T. urticae in a small plot study in 2013 and against L. hesperus and other insect pests in a large plot study in 2015 in commercial strawberry fields. Bug vacuums were also used in the 2015 study. Results demonstrated that non-chemical alternatives can play an important role in strawberry IPM.

Conventional prey-specific gut content ELISA (enzyme-linked immunosorbent assay) and PCR (polymerase chain reaction) assays are useful for identifying predators of insect pests in nature. However, these assays are prone to yielding certain types of food chain errors. For instance, it is possible that prey remains can pass through the food chain as the result of a secondary predator (hyperpredator) consuming a primary predator that had previously consumed the pest. If so, the pest-specific assay will falsely identify the secondary predator as the organism providing the biological control services to the ecosystem. Recently, a generic gut content ELISA was designed to detect protein-marked prey remains. That assay proved to be less costly, more versatile, and more reliable at detecting primary predation events than a prey-specific PCR assay. This study examines the chances of obtaining a 'false positive' food chain error with the generic ELISA. Data revealed that the ELISA was 100% accurate at detecting protein-marked Lygus hesperus Knight ( Hemiptera: Miridae) remains in the guts of two (true) primary predators, Hippodamia convergens Guérin- Méneville ( Coleoptera: Coccinellidae) and Collops vittatus ( Say) ( Coleoptera: Melyridae). However, there was also a high frequency (70%) false positives associated with hyperpredators, Zelus renardii Kolenati ( Hemiptera: Reduviidae), that consumed a primary predator that possessed protein-marked L. hesperus in its gut. These findings serve to alert researchers that the generic ELISA, like the PCR assay, is susceptible to food chain errors. [ABSTRACT FROM AUTHOR]

Background: The plant bug Lygus hesperus Knight is a polyphagous pest of many economically important crops. Despite its pest status, little is known about the molecular mechanisms responsible for much of the biology of this species. Earlier Lygus transcriptome assemblies were limited by low read depth, or because they focused on specific conditions. To generate a more comprehensive transcriptome, we supplemented previous datasets with new reads corresponding to specific tissues (heads, antennae, and male reproductive tissues). This transcriptome augments current Lygus molecular resources and provides the foundational knowledge critical for future comparative studies. Findings: An expanded, Trinity-based de novo transcriptome assembly for L. hesperus was generated using previously published whole body Illumina data, supplemented with 293 million bp of new raw sequencing data corresponding to five tissue-specific cDNA libraries and 11 Illumina sequencing runs. The updated transcriptome consists of 22,022 transcripts (average length of 2075 nt), 62 % of which contain complete open reading frames. Significant coverage of the BUSCO (benchmarking universal single-copy orthologs) dataset and robust metrics indicate that the transcriptome is a quality assembly with a high degree of completeness. Initial assessment of the new assembly's utility revealed that the length and abundance of transcripts predicted to regulate insect physiology and chemosensation have improved, compared with previous L. hesperus assemblies. Conclusions: This transcriptome represents a significant expansion of Lygus transcriptome data, and improves foundational knowledge about the molecular mechanisms underlying L. hesperus biology. The dataset is publically available in NCBI and GigaDB as a resource for researchers. [ABSTRACT FROM AUTHOR]

The desert-adapted crops vernonia ( Centrapalus pauciflorus ), lesquerella ( Physaria fendleri ), and camelina ( Camelina sativa ) are being grown in the arid southwestern USA as potential feedstock for biofuel and/or other environmentally friendly products. A plant feeding choice test was conducted to determine the relative attractiveness of these three “new” crops to a possible insect pest, Lygus hesperus Knight. Adult L. hesperus were readily observed feeding or resting on the flowering structures of each plant type, but they were seen most often on vernonia and least often on camelina. Lygus hesperus readily deposited their eggs on each plant species, but again, the greatest amount of egg deposition was found on vernonia and the least on camelina. These studies indicate that L. hesperus might pose a threat to the production of these new crops. Moreover, the commercial expansion of these crops could significantly alter the population dynamics of the existing arthropod community. New challenges for managing this pest during regional crop production changes are discussed. [ABSTRACT FROM AUTHOR]

The western tarnished plant bug,Lygus hesperusKnight (Hemiptera: Miridae), is an important insect pest of strawberry (Fragaria ananassaDuchesne) in the Central Coast of California. Because little is known about efficacy of new insecticides, especially sulfoxaflor and flupyradifurone, againstL. hesperus, two replicated insecticide experiments were conducted in 2014 and 2015 in commercial strawberry fields naturally infested withL. hesperus. The insecticides, sulfoxaflor, flupyradifurone, flonicamid, thiamethoxam fenpropathrin, essential oils (rosemary and peppermint oils), and mineral oil, were compared with untreated check. Higher and lower rates of flupyradifurone and sulfoxaflor were tested. Higher rates of sulfoxaflor and flupyradifurone were effective in suppressingL. hesperus. The industry standard, thiamethoxam + fenpropathrin, was effective in 2014 but not in the 2015 experiment. Flonicamid provided a moderate level of efficacy againstL. hesperusin both years. The number of predacious bugs was significantly lower in the higher rate of flupyradifurone than in other treatments, although the number of spiders was not significantly different among treatments. [ABSTRACT FROM PUBLISHER]

Western tarnished bug or lygus bug (Lygus hesperus) is a major pest of California strawberries causing significant yield reduction. Other common insect pests include aphids (Chaetosiphon fragaefoliiandMyzus persicae), greenhouse whitefly (Trialeurodes vaporariorum), and western flower thrips (Frankliniella occidentalis). Growers mainly depend on chemical pesticides for managing these pests. Limited control with some of the available chemical pesticides and the risk of resistance development due to their repeated use warrant a strong integrated pest management program where multiple tools can be utilized. Field studies conducted in commercial strawberry fields from 2012–14 in Santa Maria with an emphasis on managingL. hesperusdemonstrated the potential of botanical and microbial pesticides. Replacing one or more applications of chemical pesticides with botanical and microbial alternatives and using them in different combinations can make a significant contribution to a sustainable management strategy. [ABSTRACT FROM PUBLISHER]

Ecdysteroids play a critical role in coordinating insect growth, development and reproduction. A suite of cytochrome P450 monooxygenases coded by what are collectively termed Halloween genes mediate ecdysteroid biosynthesis. In this study, we describe cloning and RNA interference (RNAi)-mediated knockdown of the CYP307B1 Halloween gene ( Spookiest) in the western tarnished plant bug, Lygus hesperus. Transcripts for Ly. hesperus Spookiest ( LhSpot) were amplified from all life stages and correlated well with timing of the pre-moult ecdysteroid pulse. In adults, LhSpot was amplified from heads of both genders as well as female reproductive tissues. Heterologous expression of a LhSpot fluorescent chimera in cultured insect cells co-localized with a fluorescent marker of the endoplasmic reticulum/secretory pathway. RNAi-mediated knockdown of LhSpot in fifth instars reduced expression of ecdysone-responsive genes E74 and E75, and prevented adult development. This developmental defect was rescued following application of exogenous 20-hydroxyecdysone but not exogenous 7-dehydrocholesterol. The unequivocal RNAi effects on Ly. hesperus development and the phenotypic rescue by 20-hydroxyecdysone are causal proof of the involvement of LhSpot in ecdysteroid biosynthesis and related developmental processes, and may provide an avenue for development of new control measures against Ly. hesperus. [ABSTRACT FROM AUTHOR]

Mating induces behavioral and physiological changes in the plant bug Lygus hesperus Knight (Hemiptera: Miridae). After receiving seminal products, which include the systemic regulator juvenile hormone (JH), females enter a post-mating period lasting several days during which they enhance their oviposition rate and lose interest in remating. To elucidate the regulation of these behavioral changes in L. hesperus, biogenic amines were quantified in the heads of females at 5 min, 1 h and 24 h after copulation and compared to levels in virgins using high-performance liquid chromatography coupled with electrochemical detection. Mating significantly increased dopamine (DA) after 1 and 24 h, and decreased octopamine (OA) after 5 min and 1 h. Serotonin did not change with mating, but tyramine was significantly reduced after 5 min. While injection of amines into virgin females did not influence sexual receptivity, OA caused a decrease in oviposition during the 24 h following injection. Topical application of the JH analog methoprene to virgins caused an increase in DA, and a decline in mating propensity, but did not influence other amines or the oviposition rate. The results suggest the decline in OA observed immediately after mating may promote egg laying, and that male-derived JH may induce an increase in DA that could account for the post-mating loss of sexual receptivity. [ABSTRACT FROM AUTHOR]

Background: The plant bug Lygus hesperus Knight is a polyphagous pest of many economically important crops. Despite its pest status, little is known about the molecular mechanisms responsible for much of the biology of this species. Earlier Lygus transcriptome assemblies were limited by low read depth, or because they focused on specific conditions. To generate a more comprehensive transcriptome, we supplemented previous datasets with new reads corresponding to specific tissues (heads, antennae, and male reproductive tissues). This transcriptome augments current Lygus molecular resources and provides the foundational knowledge critical for future comparative studies. Findings: An expanded, Trinity-based de novo transcriptome assembly for L. hesperus was generated using previously published whole body Illumina data, supplemented with 293 million bp of new raw sequencing data corresponding to five tissue-specific cDNA libraries and 11 Illumina sequencing runs. The updated transcriptome consists of 22,022 transcripts (average length of 2075 nt), 62 % of which contain complete open reading frames. Significant coverage of the BUSCO (benchmarking universal single-copy orthologs) dataset and robust metrics indicate that the transcriptome is a quality assembly with a high degree of completeness. Initial assessment of the new assembly's utility revealed that the length and abundance of transcripts predicted to regulate insect physiology and chemosensation have improved, compared with previous L. hesperus assemblies. Conclusions: This transcriptome represents a significant expansion of Lygus transcriptome data, and improves foundational knowledge about the molecular mechanisms underlying L. hesperus biology. The dataset is publically available in NCBI and GigaDB as a resource for researchers. [ABSTRACT FROM AUTHOR]

The transient receptor potential (TRP) family of cation channels are evolutionarily conserved proteins with critical roles in sensory physiology. Despite extensive studies in model species, knowledge of TRP channel functional diversity and physiological impact remains limited in many non-model insect species. To assess the TRP channel repertoire in a non-model agriculture pest species (Lygus hesperus), publicly available transcriptomic datasets were mined for potential homologs. Among the transcripts identified, 30 are predicted to encompass complete open reading frames that encode proteins representing each of the seven TRP channel subfamilies. Although no homologs were identified for the Pyrexia and Brivido channels, the TRP complement in L. hesperus exceeded the 13–16 channels reported in most insects. This diversity appears to be driven by a combination of alternative splicing, which impacted members of six subfamilies, and gene expansion of the TRPP subfamily. To validate the in silico data and provide more detailed analyses of L. hesperus TRP functionality, the putative Painless homolog was selected for more in depth analysis and its functional role in thermosensation examined in vitro. RT-PCR expression profiling revealed near ubiquitous expression of the Painless transcript throughout nymphal and adult development. Electrophysiological data generated using a Xenopus oocyte recombinant expression system indicated activation parameters for L. hesperus Painless homolog that are consistent with a role in noxious heat (40°–45 °C) thermosensation. [Display omitted] • Transcriptome data mining of Lygus hesperus datasets for TRP channel homologs • TRP channel diversity greater than that reported for other insects • Gene duplication and alternative splicing contribute to channel diversity. • Near ubiquitous and constitutive expression of a Painless homolog • Activation of Painless homolog in Xenopus oocytes suggests role in noxious heat thermosensation. [ABSTRACT FROM AUTHOR]

The western tarnished plant bug, Lygus hesperus Knight (Hemiptera: Miridae), is a key pest of many horticultural and agronomic crops in the western United States. Despite its well documented pest status, many aspects of the basic biology, including overwintering ecology, of L. hesperus are poorly understood. We examined the influence of eight constant temperatures from 10 to 35°C on survival of nondiapausing adult L. hesperus held with or without food, and the consequences of exposure to an extended period at 10°C on subsequent reproduction. Survival analyses indicated that, on average, fed insects tended to live longer than unfed insects, females lived longer than males, and the survival time decreased with increasing temperature. Nonlinear regressions indicated that median survival for insects grouped by gender and feeding status declined exponentially with increasing temperature. Survival functions for combinations of insect class (gender and feeding status) and temperature were adequately described by the respective two-parameter logistic functions. When adults were held for 9d at 27°C with food after a 33-d period at 10°C either with or without food, no deleterious effects of prior starvation on propensity to mate or fecundity were demonstrated. These findings indicate that when temperatures are low, nondiapausing L. hesperus adults are capable of extended host-free survival with little or no impact on subsequent reproduction. Our findings suggest the current understanding of L. hesperus overwintering dynamics is incomplete. In addition, our results provide quantitative baseline information to facilitate more comprehensive investigation of the ecology of L. hesperus overwintering. [ABSTRACT FROM AUTHOR]

In the context of inverse or parameter estimation problems we demonstrate the use of statistically based model comparison tests in several examples of practical interest. In these examples we are interested in questions related to information content of a particular given data set and whether the data will support a more complicated model to describe it. In the first example we compare fits for several different models to describe simple decay in a size histogram for aggregates in amyloid fibril formation. In a second example we investigate whether the information content in data sets for the pest Lygus hesperus in cotton fields as it is currently collected is sufficient to support a model in which one distinguishes between nymphs and adults. Finally in a third example with data for patients having undergone an organ transplant, we question whether the data content is sufficient to estimate more than 5 of the fundamental parameters in a particular dynamic model. [ABSTRACT FROM AUTHOR]

A segment of the nuclear 28S rRNA gene was compared among six species of Lygus [Lygus hesperus (Knight), Lygus keltoni (Schwartz), Lygus borealis (Kelton), Lygus elisus (Van Duzee), Lygus lineolaris (Palisot de Beauvois), and Lygus vanduzeei (Knight)]. The DNA sequences were separate into three main groups. The LL group contains L. lineolaris and L. vanduzeei. Group LBLE is composed of L. elisus and most of L. borealis. Group LH includes L. hesperus and most of L. keltoni. Some L. keltoni were part of the LBLE group and some L. borealis were part of the LH group. The 28S region does not contain sufficient genetic polymorphism to delineate species. The apparent polyphyly of L. borealis and L. keltoni could reflect historic interbreeding, recent development of a hybrid swarm, or highlight inadequacies of morphospecies identification. [ABSTRACT FROM PUBLISHER]

The Gα subunits of heterotrimeric G proteins play critical roles in the activation of diverse signal transduction cascades. However, the role of these genes in chemosensation remains to be fully elucidated. To initiate a comprehensive survey of signal transduction genes, we used homology-based cloning methods and transcriptome data mining to identity Gα subunits in the western tarnished plant bug (Lygus hesperus Knight). Among the nine sequences identified were single variants of the Gαi, Gαo, Gαs, and Gα12 subfamilies and five alternative splice variants of the Gαq subfamily. Sequence alignment and phylogenetic analyses of the putative L. hesperus Gα subunits support initial classifications and are consistent with established evolutionary relationships. End-point PCR-based profiling of the transcripts indicated head specific expression for LhGαq4, and largely ubiquitous expression, albeit at varying levels, for the other LhGα transcripts. All subfamilies were amplified from L. hesperus chemosensory tissues, suggesting potential roles in olfaction and/or gustation. Immunohistochemical staining of cultured insect cells transiently expressing recombinant His-tagged LhGαi, LhGαs, and LhGαq1 revealed plasma membrane targeting, suggesting the respective sequences encode functional G protein subunits. [ABSTRACT FROM AUTHOR]

ATP-binding cassette (ABC) transporters are a large superfamily of proteins that mediate diverse physiological functions by coupling ATP hydrolysis with substrate transport across lipid membranes. In insects, these proteins play roles in metabolism, development, eye pigmentation, and xenobiotic clearance. While ABC transporters have been extensively studied in vertebrates, less is known concerning this superfamily in insects, particularly hemipteran pests. We used RNA-Seq transcriptome sequencing to identify 65 putative ABC transporter sequences (including 36 full-length sequences) from the eight ABC subfamilies in the western tarnished plant bug (Lygus hesperus), a polyphagous agricultural pest. Phylogenetic analyses revealed clear orthologous relationships with ABC transporters linked to insecticide/xenobiotic clearance and indicated lineage specific expansion of the L. hesperus ABCG and ABCH subfamilies. The transcriptional profile of 13 LhABCs representative of the ABCA, ABCB, ABCC, ABCG, and ABCH subfamilies was examined across L. hesperus development and within sex-specific adult tissues. All of the transcripts were amplified from both reproductively immature and mature adults and all but LhABCA8 were expressed to some degree in eggs. Expression of LhABCA8 was spatially localized to the testis and temporally timed with male reproductive development, suggesting a potential role in sexual maturation and/or spermatozoa protection. Elevated expression of LhABCC5 in Malpighian tubules suggests a possible role in xenobiotic clearance. Our results provide the first transcriptome-wide analysis of ABC transporters in an agriculturally important hemipteran pest and, because ABC transporters are known to be important mediators of insecticidal resistance, will provide the basis for future biochemical and toxicological studies on the role of this protein family in insecticide resistance in Lygus species. [ABSTRACT FROM AUTHOR]

Lygus hesperus females exhibit a post-mating behavioural switch that triggers increased egg laying and decreased sexual interest. In Drosophila melanogaster, these changes are controlled by sex peptide ( SP) and the sex peptide receptor ( DmSPR). In Helicoverpa armigera, SPR ( HaSPR) also regulates some post-mating behaviour; however, myoinhibiting peptides ( MIPs) have been identified as the SPR ancestral ligand, indicating that SPR is a pleiotropic receptor. In the present study, we identified a transcript, designated L. hesperus SPR ( LhSPR), that is homologous to known SPRs and which is expressed throughout development and in most adult tissues. Lh SPR was most abundant in female seminal depositories and heads as well as the hindgut/midgut of both sexes. In vitro analyses revealed that fluorescent chimeras of LhSPR, DmSPR and HaSPR localized to the cell surface of cultured insect cells, but only DmSPR and HaSPR bound carboxytetramethylrhodamine-labelled analogues of Dm SP21-36 and DmMIP4. Injected DmSP21-36 also failed to have an effect on L. hesperus mating receptivity. Potential divergence in the LhSPR binding pocket may be linked to receptor-ligand co-evolution as 9 of 13 MIPs encoded by a putative L. hesperus MIP precursor exhibit an atypical W- X7- Wamide motif vs the W- X6- Wamide and W- X8- Wamide motifs of Drosophila MIPs and SP. [ABSTRACT FROM AUTHOR]