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Flight mills are widely used to investigate insect flight behavior. As technology advances, the means to build a computerized control system for a flight mill has become more accessible in terms of both price and availability of components. However, the specialized electronics and programming knowledge required to build such a system can still present an obstacle to interested parties. Here, we describe a simple and inexpensive flight mill control system that can be easily assembled and operated without specialized experience. The hardware and software components are built around an Arduino single-board microcontroller, which outputs raw data in the form of timestamped detections of rotations of the flight mill arm. This control system is suitable both as the basis for new flight mills and for replacing outdated computer controls on existing flight mills. Additionally, it can be used with any rotary flight mill design that uses an electronic sensor to count rotations., (Published by Oxford University Press on behalf of Entomological Society of America 2023.)

We examined the feasibility of externally marking insects with the liquid fluorescent forensic theft deterrent, SmartWater (SmartWater CSI, LLC.). We sprayed captive Lygus hesperus (Knight) (Hemiptera: Miridae), Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae), and Hippodamia convergens Guérin-Méneville (Coleoptera: Coccinellidae) with SmartWater fluorophore, and then qualitatively examined them for fluorescence by visual inspection under ultraviolet (UV) light and quantitatively measured them with a multiwavelength microplate fluorometer. The results indicate that this product has enormous potential as a taggant for L. hesperus and B. tabaci. However, the marking efficiency for H. convergens was only adequate. The advantages and limitations of using SmartWater as a biological marker for arthropod mark-release-recapture research are discussed., (Published by Oxford University Press on behalf of Entomological Society of America 2021.)

Vernonia [Vernonia galamensis (Cass.) Less.] (Asterales: Asteraceae) was examined as a potential trap crop for the cotton (Gossypium hirsutum L., Malvales: Malvaceae) arthropod complex. Four rows of vernonia were embedded within a 96-row cotton field. The abundance of true bug pests, true bug predators, and spiders were determined by whole-plant and sweep net sampling procedures during the early, middle, and late phases of the cotton-growing season. The census data showed that the arthropods had a strong preference for the vernonia trap crop throughout the cotton-growing season. The movement of the arthropods from the trap crop into cotton was also measured using the protein immunomarking technique as a mark-capture procedure. The arthropods inhabiting the vernonia trap crop were marked directly in the field with a broadcast spray application of egg albumin (protein) during each phase of the study. In turn, the captured specimens were examined for the presence of the mark by an egg albumin-specific enzyme-linked immunosorbent assay. Very few marked specimens were captured beyond the vernonia trap crop 1, 3, and 6 d after each marking event. The arthropods' strong attraction and fidelity to vernonia indicate that it could serve as a trap crop for cotton pests and a refuge for natural enemies., (Published by Oxford University Press on behalf of Entomological Society of America 2021.)

The western tarnished plant bug, Lygus hesperus Knight, overwinters as a diapausing adult in response to short day lengths. Once environmental conditions are favorable, the bugs revert to an active reproductive state. To determine the impact on life-history traits of diverting resources toward diapause rather than oogenesis during early adulthood, diapausing and nondiapausing L. hesperus females were reared from the same cohorts. Body mass, ovarian maturation, ovipositional activity, and survivorship were monitored starting either at the time of release from diapause-inducing conditions or at adult eclosion for diapausers and nondiapausers, respectively. Females that had gone through 2 wk of diapause were larger and able to mobilize the resources necessary for oogenesis faster than nondiapausers, initiating oogenesis and ovipositing sooner and at a faster initial rate. However, lifetime egg production and average daily rates were similar for both groups. Postdiapausers lived longer than nondiapausers by an average of 19 d, which is five more than the 2-wk period when they were reproductively senescent. Overall, the results indicate that short-term diapause does not have a negative impact on life history. Furthermore, the extra endogenous resources stored during diapause may be able to enhance the alacrity with which the female can take advantage of improved environmental conditions and may prolong life by shielding the females against environmental stressors such as temperature extremes, oxidative agents, or food deficits., (Published by Oxford University Press on behalf of Entomological Society of America 2021.)

A molecular gut analysis technique is described to identify predators of Lygus hesperus (Knight), a significant pest of many crops. The technique is unique because it can pinpoint which life stage of the pest was consumed. Sentinel egg masses designed to mimic the endophytic egg-laying behavior of L. hesperus were marked with rabbit serum, while third instar and adult L. hesperus were marked with chicken and rat sera, respectively. Then, the variously labeled L. hesperus life stages were introduced into field cages that enclosed the native arthropod population inhabiting an individual cotton plant. After a 6-h exposure period, the predator assemblage, including the introduced and native L. hesperus population, in each cage were counted and had their gut contents examined for the presence of the variously marked L. hesperus life stages by a suite of serum-specific enzyme-linked immunosorbent assays (ELISA). The whole-plant sampling scheme revealed that Geocoris punticpes (Say) and Geocoris pallens Stal (Hemiptera: Geocoridae) and members of the spider complex were the numerically dominant predator taxa in the cotton field. The gut content analyses also showed that these two taxa appeared to be the most prolific predators of the L. hesperus nymph stage. Other key findings include that Collops vittatus (Say) (Coleoptera: Melyridae) and Solenopsis xyloni McCook (Hymenoptera: Formicidae) appear to be adept at finding and feeding on the cryptic L. hesperus egg stage, and that L. hesperus, albeit at low frequencies, engaged in cannibalism. The methods described here could be adapted for studying life stage-specific feeding preferences for a wide variety of arthropod taxa., (Published by Oxford University Press on behalf of Entomological Society of America 2020.)

A follow-up study was conducted to further evaluate the marking efficiency of broadcast spray applications of egg albumin (from chicken egg whites) on Hippodamia convergens Guérin-Méneville (Coleoptera: Coccinellidae) in alfalfa. A previous study recorded exceptional marking efficiency (e.g., >95% of the population) on H. convergens when using relatively high concentrations (10 to 50%) of chicken egg whites. The present study examines marking efficiency of egg whites using lower concentrations of 2.5, 5.0, and 10.0%. We used cadaver and free-roaming beetles to measure protein mark acquisition (and retention) of each protein concentration by direct contact with the spray application and incidental contact with protein residue on the plant tissue, respectively. The vertical distribution of the protein mark was also determined by sampling the upper and lower portions of the alfalfa canopy. The data indicate, regardless of the egg white treatment, that the backpack sprayer provided uniform coverage of egg albumin on the alfalfa plants and cadaver beetles. Also, almost every free-roaming beetle acquired a mark within 24 h after contact exposure to protein marked plants. This study shows that a very low concentration of egg albumin is sufficient for marking arthropods directly in the field., (Published by Oxford University Press on behalf of Entomological Society of America 2020.)

Acquisition and retention of two protein markers were tested on little fire ants, Wasmannia auropunctata Roger. Pure (100%) cow's milk and a dilution (10%) of chicken egg whites were applied to W. auropunctata directly by contact spray plus residue or indirectly via residual contact only with protein-marked plant debris. Protein-marked ants were held in plastic shoe-box-sized containers, collected at 0, 24, and 48 h after exposure to their respective marks, and then examined for the presence of the marks by a chicken egg albumin and milk casein-specific enzyme-linked immunosorbent assay. Cross-contamination rates were assessed by allowing ants marked with egg whites to interact with an equal number marked milk for 24 and 48 h, and then collected either individually or in bulk. Results indicated that the egg white biomarker was retained longer than milk and that more ants were successfully marked when the direct spray application method was employed. Cross-contamination rates were highest among bulk-collected ants and lowest among ants collected individually after 24 h. However, the rates of cross-contamination among individually collected ants increased and were similar to that of bulk-collected ants after 48 h. On the basis of our results, external protein marking may not be suitable if mass trapping is required or if the study extends beyond 24 h due to high cross-contamination rates among specimens collected in bulk and reduced marker detection rates., (© The Author(s) 2019. Published by Oxford University Press on behalf of Entomological Society of America.)

A universal food immunomarking technique (UFIT) is described for postmortem gut analysis detection of predation on the egg stage of Lygus hesperus Knight (Hemiptera: Miridae). Collops vittatus Say (Coleoptera: Melyridae) and Hippodamia convergens Guérin-Méneville (Coleoptera: Coccinellidae) were fed a single L. hesperus egg that was marked with rabbit and chicken sera proteins. The protein-marked egg remnants were detectable in the guts of the majority of the predators by each sera-specific enzyme-linked immunosorbent assay (ELISA) for 3 to 6 h after a feeding event. A novel technique was then developed to expose protein-marked eggs to predators that simulated the L. hesperus endophytic oviposition behavior. The procedure entailed embedding L. hesperus eggs in an artificial substrate that mimicked the stem of a plant. A predator feeding choice study was then conducted in cages that contained a cotton plant and artificial stems containing endophytic (concealed) and exophytic (exposed) egg patches. The endophytic and exophytic egg treatments were marked with chicken and rabbit protein, respectively. The gut analyses revealed that higher proportions of both predator populations contained remnants of the exophytic egg treatment and L. hesperus eggs were more vulnerable to C. vittatus than H. convergens. This study shows how the UFIT can be used to pinpoint stage-specific feeding activity on two distinct egg exposure treatments (endophytic and exophytic) of the same species., (Published by Oxford University Press on behalf of Entomological Society of America 2019.)

Alfalfa (Medicago sativa L.) (Fabales: Fabaceae) can be strategically planted as a trap crop for Lygus spp. in California's organic strawberry fields. Alfalfa has been shown to attract both Lygus spp. and, in turn, a Lygus-specific parasitoid, Peristenus relictus (Ruthe) (Hymenoptera: Braconidae). However, the impact of alfalfa trap-cropped strawberries on the Lygus spp. predator complex is unknown. Here we identify key predators of Lygus spp. found in organic strawberry. First, a general survey was conducted at an organic, non-trap cropped strawberry farm, to quantify predator abundance and to qualitatively assess their feeding activity on Lygus spp. We identified the 11 most abundant predator taxa present and, by using a Lygus-specific PCR assay, determined that about 18% of the insects and spiders contained Lygus spp. remains in their guts. We then conducted a study to examine alfalfa's role in conserving the most relevant predators in trap-cropped organic strawberries. Specifically, we quantified predator abundance and qualitatively measured predator feeding activity (by gut analysis) on Lygus spp. collected in strawberry plots either lacking or containing an alfalfa trap crop. Data revealed that some predator taxa, including the numerically dominant predator, Orius tristicolor (White) (Hemiptera: Anthocoridae), aggregated in alfalfa trap crops. The gut content analyses revealed that insect and spider predators collected from the alfalfa trap crop had a significantly higher proportion of their population containing Lygus spp. remains than those collected from nearby rows of strawberries. These results suggest that alfalfa trap cropping might be a useful tactic for conserving the biological control services of generalist predators in organically grown strawberries in California.

Studies of bee movement and activities across a landscape are important for developing an understanding of their behavior and their ability to withstand environmental stress. Recent research has shown that proteins, such as egg albumin, are effective for mass-marking bees. However, current protein mass-marking techniques require sacrificing individual bees during the data collection process. A nonlethal sampling method for protein mark-capture research is sorely needed, particularly for vulnerable, sensitive, or economically valuable species. This study describes a nonlethal sampling method, in which three non-Apis bee species (Bombus bifarius Cresson [Hymenoptera: Apidae], Osmia lignaria Say [Hymenoptera: Megachilidae], and Megachile rotundata Fabricius [Hymenoptera: Megachilidae]) were tested for a unique protein marker by immersing them momentarily in saline buffer and releasing them. Results showed that an egg albumin-specific enzyme-linked immunosorbent assay was 100% effective at detecting the protein on bees that were sampled nonlethally. Furthermore, this sampling method did not have an impact on bee survivorship, suggesting that immersing bees in buffer is a reliable and valid surrogate to traditional, destructive sampling methods for mark-capture bee studies.

A study was conducted that compared the effectiveness of a sweepnet versus a vacuum suction device for collecting arthropods in cotton. The study differs from previous research in that body-mounted action cameras (B-MACs) were used to record the activity of the person conducting the arthropod collections. The videos produced by the B-MACs were then analyzed with behavioral event recording software to quantify various aspects of the sampling process. The sampler's speed and the number of sampling sweeps or vacuum suctions taken over a fixed distance (12.2 m) of cotton were two of the more significant sampling characteristics quantified for each method. The arthropod counts obtained, combined with the analyses of the videos, enabled us to estimate arthropod sampling efficiency for each technique based on fixed distance, time, and sample unit measurements. Data revealed that the vacuuming was the most precise method for collecting arthropods in the relatively small cotton research plots. However, data also indicates that the sweepnet method would be more efficient for collecting most of the cotton-dwelling arthropod taxa, especially if the sampler could continuously sweep for at least 1 min or ≥80 m (e.g., in larger research plots). The B-MACs are inexpensive and non-cumbersome, the video images generated are outstanding, and they can be archived to provide permanent documentation of a research project. The methods described here could be useful for other types of field-based research to enhance data collection efficiency.

Recent studies on adult diapause in the western tarnished plant bug, Lygus hesperus Knight (Heteroptera: Miridae), have highlighted the need to identify a reliable external marker for the internal changes that differentiate a normal animal from one that is overwintering. To test the efficacy of a color based discrimination system, L. hesperus of both genders were reared from eggs through day 10 of adulthood under a 10 hour photophase at a constant temperature. They were separated into three color groups (yellow, pale green, dark green), then dissected for diapause categorization based on internal development. Most yellow individuals were in diapause, dark green individuals were not, and pale green ones were mixed. A group of 25 assessors, naïve with regard to L. hesperus development, were then asked to use a simplified color criteria (yellow = diapause, green = non-diapause) to estimate the status of a mixture of diapausing and non-diapausing adults of both genders aged two to seven days post-eclosion. After dissection to verify diapause status, assessor accuracy was found to be ineffective for assessing adults of both sexes younger than four days because color differences, which increased with age, were subtle or non-existent at this stage. For four to seven-day-old bugs, 84% of females and 67% of males were correctly categorized, on average. Incorrect assessments in all but the youngest males over-identified diapause, but for females there was no trend in miscategorizations. Overall, the results indicate that diapause status can be adequately discriminated by color assessment, and with greatest accuracy when sampling older females. However, factors other than photoperiod appear to also influence coloration.

A study was conducted in 2006 and 2007 designed to examine the foraging range of honey bees, Apis mellifera (Hymenoptera: Apidae), in a 15.2 km(2) area dominated by a 128.9 ha glyphosate-resistant Roundup Ready® alfalfa seed production field and several non-Roundup Ready alfalfa seed production fields (totaling 120.2 ha). Each year, honey bee self-marking devices were placed on 112 selected honey bee colonies originating from nine different apiary locations. The foraging bees exiting each apiary location were uniquely marked so that the apiary of origin and the distance traveled by the marked (field-collected) bees into each of the alfalfa fields could be pinpointed. Honey bee self-marking devices were installed on 14.4 and 11.2% of the total hives located within the research area in 2006 and 2007, respectively. The frequency of field-collected bees possessing a distinct mark was similar, averaging 14.0% in 2006 and 12.6% in 2007. A grand total of 12,266 bees were collected from the various alfalfa fields on seven sampling dates over the course of the study. The distances traveled by marked bees ranged from a minimum of 45 m to a maximum of 5983 m. On average, marked bees were recovered ~ 800 m from their apiary of origin and the recovery rate of marked bees decreased exponentially as the distance from the apiary of origin increased. Ultimately, these data will be used to identify the extent of pollen-mediated gene flow from Roundup Ready to conventional alfalfa.

Inexpensive and non-intrusive marking methods are essential to track natural behavior of insects for biological experiments. An inexpensive, easy to construct, and easy to install bee marking device is described in this paper. The device is mounted at the entrance of a standard honey bee Apis mellifera L. (Hymenoptera: Apidae) hive and is fitted with a removable tube that dispenses a powdered marker. Marking devices were installed on 80 honey bee colonies distributed in nine separate apiaries. Each device held a tube containing one of five colored fluorescent powders, or a combination of a fluorescent powder (either green or magenta) plus one of two protein powders, resulting in nine unique marks. The powdered protein markers included egg albumin from dry chicken egg whites and casein from dry powdered milk. The efficacy of the marking procedure for each of the unique markers was assessed on honey bees exiting each apiary. Each bee was examined, first by visual inspection for the presence of colored fluorescent powder and then by egg albumin and milk casein specific enzyme-linked immunosorbent assays (ELISA). Data indicated that all five of the colored fluorescent powders and both of the protein powders were effective honey bee markers. However, the fluorescent powders consistently yielded more reliable marks than the protein powders. In general, there was less than a 1% chance of obtaining a false positive colored or protein-marked bee, but the chance of obtaining a false negative marked bee was higher for "protein-marked" bees.

Lygus hesperus Knight and Lygus lineolaris (Palisot de Beauvois) (Heteroptera: Miridae) are economically important plant bugs on many crops worldwide. However, these omnivores are also facultative predators on a wide variety of insects. This study was conducted to quantify and compare herbivory and carnivory exhibited among different lifestages of these two insect pests. The feeding activity of a total of 422 individuals was observed for 1 h each in feeding arenas containing a cotton leaf disk and copious amounts of the sweetpotato whitefly, Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) eggs, nymphs and adults. The L. hesperus and L. lineolaris lifestages examined included adults and 3rd, 4th and 5th instar nymphs. Plant feeding occupied the majority of both species' time budget, regardless of the species or lifestage examined. There was a tendency for L. lineolaris lifestages to feed more often and for longer duration on plant tissue than L. hesperus. All lifestages of both species rarely fed on B. tabaci, but when they did, they preferred nymphs > adults > eggs. There were only a few cases where there were significant differences in predation rates and prey handling times exhibited among lifestages and between species, but juvenile L. hesperus tended to be more predaceous than juvenile L. lineolaris on whitefly nymphs and adults and 5th instar and adult L. lineolaris were significantly more herbaceous than their L. hesperus counterparts. In addition, the younger individuals of both species tended to have greater prey handling times than their older counterparts. The significance of these findings is discussed.

The western tarnished plant bug Lygus hesperus Knight (Heteroptera: Miridae), a major pest of cotton and other key economic crops, was tested for its sensitivity to population density during nymph and adult stages. Nymphs reared to adulthood under increasing densities in laboratory conditions exhibited incremental delays in maturation, heightened mortality rates, and reductions in body mass and various size parameters. In contrast, gonadal activity in both males and females rose with initial density increases. Supplemental nutrients provided to the nymphs failed to offset the negative effects of high density, suggesting that contact frequency, rather than resource partitioning, may be the primary stress. Unlike nymphs, newly enclosed adults exposed to increasing population densities did not suffer negative physiological effects; body mass, mortality rates and patterns of ovipositional activity were unchanged. Collectively, these results indicate that population density can dramatically influence Lygus development, but the specific effects are stage-dependent.

Bt cotton plants are genetically engineered to produce insecticidal toxins from the Bacillus thuringiensis (Bt) Berliner (Bacillales: Bacillaceae) bacterium and target key lepidopteran pests. In all previous strains of pink bollworm, Pectinophora gossypiella (Saunders) (Lepidoptera: Gelechiidae) selected in the laboratory for resistance to insecticidal Cry1Ac toxin using an artificial diet containing the toxin, resistance to Cry1Ac and to Bt cotton is linked to three cadherin alleles (r1, r2, and r3). In contrast, the BG(4) pink bollworm strain was selected for resistance to Bt cotton by feeding larvae for four days in each of 42 generations on bolls of 'NuCOTN33B(R)' that expressed Cry1Ac toxin. After additional selection for eleven generations on Cry1Ac-incorporated diet, the susceptibility to Cry1Ac, fecundity, egg viability, and mating of this strain (Bt4R) was compared with the unselected Cry1Ac-susceptible parent strain. Some larvae of the Bt4R strain survived on diet containing >or= 10 microg Cry1Ac per milliliter artificial diet, but none survived on transgenic cotton bolls. In contrast to strains selected exclusively on Cry1Ac diet, some survival of progeny of reciprocal moth crosses of Bt4R resistant and Bt-susceptible strains occurred on Cry1Ac-treated diet, suggesting differences in levels of dominance. The Bt4R resistant strain does not have the r1, r2, or r3 mutant cadherin genes as do all previous strains of pink bollworm selected on Cry1Ac-treated artificial diet. The combined results suggest a mechanism of resistance to Cry1Ac that is different from previously described cadherin mutations.