Infectivity of Entomopathogenic Fungal Isolates Against Tarnished Plant Bug Lygus lineolaris (Hemiptera: Miridae).

Twelve isolates of entomopathogenic fungi belonging to Metarhizium robertsii, M. pinghaense, M. brunneum, Beauveria bassiana, and Isaria fumosorosea were screened against tarnished plant bug. All isolates were pathogenic, causing mortality from 28.8 ± 3.4 to 96.3 ± 2.7%. The LT50 values ranged from 2.7 to 6.0 d while the LT90 values varied between 6.6 and 15.0 d. Metarhizium robertsii isolate CPD6 (will be under the trade name NoVil) was among the isolates that caused high mortality within shorter times and was selected for study on developmental stages and greenhouse trial. The third-, fourth-, and fifth-instar nymphs, and adults were inoculated with 106, 107, and 108 conidia per ml of NoVil. All the stages were susceptible to fungal infection. However, third and fourth instars were the most susceptible with no significant differences in mortality across the three concentrations. On the other hand, mortality was dose-dependent with fifth-instar nymph and adult stages. The LT50 and LT90 values were also dose-dependent, with higher concentrations having shorter lethal-time values as compared to the lower concentrations. In the greenhouse, pepper plants were sprayed with NoVil and chemical insecticide Flonicamid (as industrial standard), before releasing adult tarnished plant bug. Mortality of 37.3, 75.5, and 76.3% was recorded in the control, NoVil, and Flonicamid, respectively. This study has identified NoVil as a potential mycoinsecticide candidate for the control of tarnished plant bug under greenhouse conditions. Further field testing on juvenile and adults is needed to evaluate the potential for in-field control.

Development of a Method for Rearing Nezara viridula (Heteroptera: Pentatomidae) on a Semi-solid Artificial Diet.

A method for rearing the southern green stinkbug, (Nezara viridula L.) (Heteroptera: Pentatomidae), using a modified lygus semi-solid artificial diet was developed. First to second-instar nymph were reared in a density of 631.5 ± 125.05 eggs per Petri-dish (4 cm deep × 15 cm diam). Second instar to adult were reared in a density of 535.0 ± 112.46 s instar nymphs per rearing cage (43 × 28 × 9 cm). Mating and oviposition occurred in popup rearing cages (30 × 30 cm), each holding 60-90 mixed sex adults of similar age. Adults emerged 35.88 ± 2.13 d after oviposition and survived for an average of 43.09 ± 9.53 d. On average, adults laid 223.95 ± 69.88 eggs in their lifetime, for a total production of 8,099 ± 1,277 fertile eggs/oviposition cage. Egg fertility was 77.93% ± 16.28. Egg masses held in petri-dishes had a total hatchability of 79.38% ± 20.03. Mortality of early nymphs in petri-dishes was 0.64% ± 0.12 for the first instar and 1.37% ± 0.45 for second instar. Late nymphal mortality in rearing cages was 1.41% ± 0.10, 3.47% ± 1.27, and 4.72% ± 1.29 for the third, fourth, and fifth instars, respectively. Survivorship from nymphs to adults was 88.48% ± 2.76. Using artificial diet for rearing N. viridula could reduce cost by avoiding time-consuming issues with daily feeding fresh natural hosts and insect manipulation. It could increase reliability and simplicity of bug production, which should facilitate mass rearing of its biological control agents.

A Laboratory Diet-Overlay Bioassay to Monitor Resistance in Lygus lineolaris (Hemiptera: Miridae) to Insecticides Commonly Used in the Mississippi Delta.

A laboratory, diet-overlay pesticide bioassay was developed using a susceptible population of the tarnished plant bug, Lygus lineolaris (Palisot de Beauvois), to study its susceptibility to neonicotinoid, sulfoxamine, organophosphate, and pyrethroid insecticides (thiamethoxam, sulfoxaflor, acephate, and permethrin, respectively). The diet-overlay bioassay was compared to the traditional glass-vial surface residue bioassay. We measured LC50 values by feeding tarnished plant bug adults known doses of insecticides dispensed on top of diet in a 10% solution of honey water for thiamethoxam and 10% acetone in water solutions for permethrin, acephate, and sulfoxaflor. Both the diet-overlay and glass-vial bioassays used dose-response (mortality) regression lines to calculate LC50 values for each insecticide at 6-, 24-, 48-, and 72-h post-exposure. Data variability from the glass-vial bioassay was higher for permethrin, sulfoxaflor, and thiamethoxam than the diet-overlay bioassay, for all evaluation times. In contrast, there was lower variability among replicates to acephate in the glass-vial assay compared to the diet-overlay assay. Control mortalities observed on diet-overlay bioassay were lower (0-5%) than those observed on the glass-vial bioassay (4-27%). The use of green beans, floral-foam, rolling glass vials, and insect handling made the existing standard method tedious to manipulate and difficult to handle large numbers of individuals. The nonautoclaved solid diet provides an opportunity to significantly reduce cost and variability associated with procedures of other bioassay methods. In general, the baseline data provide a basis for future comparison to determine changes in resistance over time.

A Novel Method to Evaluate the Reproductive Potential of Phymastichus coffea (Hymenoptera: Eulophidae) in Hypothenemus hampei (Coleoptera: Curculionidae: Scolytinae) Under Laboratory Conditions.

Life fertility tables were constructed for Phymastichus coffea LaSalle, the only known endoparasitoid wasp that attacks the adult female of the coffee berry borer (CBB) Hypothenemus hampei Ferrari. No preoviposition period was observed, and the parasitoid female attacked the CBB females immediately after emergence. The reproductive period was 20.33 (±0.87 SE) h and the postreproductive period of 14.78 (±0.99 SE) h. Mean generation time was 37 d. Median longevity for female and male fed on honey-water solution (50:50) was 35.22 (±1.18 SE) and 17.16 (±0.96 SE) h, respectively. The highest oviposition values were observed during the first 4 h after emergence. The sex ratio (F:M) was 1.11 (±0.31 SE): 1.04 (±0.32 SE) with a finite rate of increase [λ] of 1.06 (±0.002 SE) wasp per days and a rm value of 0.067 (±0.002 SE). Gross fecundity [Mx] and net fecundity [mx] were 19.87 (±1.57 SE) males and females per female and 10.9 (±0.88 SE) females per female, respectively. The net reproductive rate [R0] was 9.49 (±0.75 SE) female per wasp. This study was designed to quantify reproductive rates of P. coffea in CBB females reared on artificial diets and used to improve a mass rearing system for this parasitoid.

Abridged Life Tables for Cephalonomia stephanoderis and Prorops nasuta (Hymenoptera: Bethylidae) Parasitoids of Hypothenemus hampei (Coleoptera: Curculionidae: Scolytinae) Reared on Artificial Diet.

Biological aspects and demographic parameters of Cephalonomia stephanoderis Betrem (Hymenoptera: Bethylidae) and Prorops nasuta Waterston (Hymenoptera: Bethylidae) parasitoids of the coffee berry borer (CBB), Hypothenemus hampei (Ferrari) (Coleoptera: Curculionidae: Scolytinae) were investigated using diet-reared CBB hosts. Developmental time from eggs to adults, oviposition, and postoviposition period were comparable for both parasitoids. However, P. nasuta had a considerably longer preoviposition and longevity period averaging 17.3 and 63.1 d, respectively. The reproductive rate for C. stephanoderis was 46.1 daughters per female with a mean generation time of 47.4 d, whereas P. nasuta had a reproductive rate of 18.3 daughters per female in a mean time of 58.6 d. Oviposition behavior was also different with C. stephanoderis typically ovipositing on CBB prepupae and pupae, while P. nasuta preferred prepupae and second-instar CBB larvae. An abridged cohort life table for both parasitoids was constructed for growth rates estimations.

Patterns of Tarnished Plant Bug (Hemiptera: Miridae) Resistance to Pyrethroid Insecticides in the Lower Mississippi Delta for 2008-2015: Linkage to Pyrethroid Use and Cotton Insect Management.

Populations of tarnished plant bug, Lygus lineolaris (Palisot de Beauvois) (Hemiptera: Miridae), from the Lower Mississippi Delta regions of Arkansas, Louisiana, and Mississippi were evaluated from 2008 through 2015 for susceptibility to pyrethroid insecticides using a diagnostic-dose assay with permethrin. Resulting data add to the compilation of pyrethroid susceptibility data carefully tracked in this pest since 1994 and provide continuing evidence of high frequencies of pyrethroid resistance in field populations of the tarnished plant bug. Resistance levels are variable, and some populations remain susceptible suggesting practical value in the continued use of the diagnostic-dose assays prior to pyrethroid treatments. Recent studies with dose-response models suggest that levels of pyrethroid resistance in some populations may still be evolving, with some populations requiring higher doses to reach levels of control comparable to those observed 10 yr ago. Concerns for frequent use of multiple classes of insecticides and possible selection for tarnished plant bugs with metabolic resistance mechanisms capable of detoxifying available insecticide chemistries warrant continued efforts to manage resistance in this important crop pest. Associations among measured pyrethroid resistance levels, published data on annual use of pyrethroid insecticides, and annual estimates of cotton insect losses and control costs were explored and summarized for the 8 yr of this investigation. Mortality of tarnished plant bugs at the diagnostic-dose of permethrin was negatively correlated with kilograms of pyrethroids applied per acre of harvested cropland.

Evidence of multiple/cross resistance to Bt and organophosphate insecticides in Puerto Rico population of the fall armyworm, Spodoptera frugiperda.

Fall armyworm (FAW) is a damaging pest of many economic crops. Long-term use of chemical control prompted resistance development to many insecticide classes. Many populations were found to be significantly less susceptible to major Bt toxins expressed in transgenic crops. In this study, a FAW strain collected from Puerto Rico (PR) with 7717-fold Cry1F-resistance was examined to determine if it had also developed multiple/cross resistance to non-Bt insecticides. Dose response assays showed that the PR strain developed 19-fold resistance to acephate. Besides having a slightly smaller larval body weight and length, PR also evolved a deep (2.8%) molecular divergence in mitochondrial oxidase subunit II. Further examination of enzyme activities in the midgut of PR larvae exhibited substantial decreases of alkaline phosphatase (ALP), aminopeptidase (APN), 1-NA- and 2-NA-specific esterase, trypsin, and chymotrypsin activities, and significant increases of PNPA-specific esterase and glutathione S-transferase (GST) activities. When enzyme preparations from the whole larval body were examined, all three esterase, GST, trypsin, and chymotrypsin activities were significantly elevated in the PR strain, while ALP and APN activities were not significantly different from those of susceptible strain. Data indicated that multiple/cross resistances may have developed in the PR strain to both Bt toxins and conventional insecticides. Consistently reduced ALP provided evidence to support an ALP-mediated Bt resistance mechanism. Esterases and GSTs may be associated with acephate resistance through elevated metabolic detoxification. Further studies are needed to clarify whether and how esterases, GSTs, and other enzymes (such as P450s) are involved in cross resistance development to Bt and other insecticide classes.

A novel bioassay to evaluate the potential of Beauveria bassiana strain NI8 and the insect growth regulator novaluron against Lygus lineolaris on a non-autoclaved solid artificial diet.

A non-autoclaved solid diet was used to evaluate the entomopathogenic fungus Beauveria bassiana (Balsamo) Vuillemin (Hypocreales: Clavicipitaceae) strain NI8 and the insect growth regulator novaluron (Diamond® 0.83EC insecticide) for control of the tarnished plant bug, Lygus lineolaris (Palisot de Beauvois) (Hemiptera: Miridae). The diet was composed of toasted wheat germ, ground lima bean meal, soy flour, yolk of chicken eggs, inhibitor, and agar. It was prepared in one step by blending the ingredients in boiling water. The diet was used to bioassay L. lineolaris from the second instar to the adult stage. Fourth and fifth instars and adults of L. lineolaris were more susceptible than second and third instars to infection by B. bassiana, whereas second, third, and fourth instars had higher mortality than fifth instars 10 days after exposure to novaluron. No effects on longevity were observed in adults treated with novaluron when compared with the control, but longevity was significantly different from that of adults exposed to B. bassiana. Adults of L. lineolaris were maintained for over a month without changing the diet. The non-autoclaved diet is semi-liquid before it cools, which facilitates the mechanics of diet packaging similar to food packaging or lepidopteran diet preparation. This solid artificial diet for Lygus bugs provides improved research capacity for studying the ecology and susceptibility of Lygus spp. to a number of different control agents, including beneficial organisms, insect pathogens, and insecticidal toxins being developed for transgenic technologies.

Effectiveness and residual activity of four common insecticides used in the Mississippi Delta to control tarnished plant bugs in cotton.

The tarnished plant bug, (TPB) Lygus lineolaris Palisot de Beauvois (Hemiptera: Miridae) is a key pest of cotton in the midsouth region and some areas of the eastern United States. Its control methods have been solely based on chemical insecticides which has contributed to insecticidal resistance and shortened residual periods for control of this insect pest. This study was conducted over a two-year period and examined the efficacy and residual effect of four commercial insecticides including lambda-cyhalothrin (pyrethroid), acephate (organophosphate), imidacloprid (neonicotinoid), and sulfoxaflor (sulfoxamine). The effectiveness and residual effects of these insecticides were determined by application on cotton field plots on four different dates during each season using three different concentrations (high: highest labeled commercial dose (CD), medium: 1/10 of the CD, low: 1/100 of the CD) on field cotton plots. Four groups of cotton leaves were randomly pulled from each treated plot and control 0-, 2-, 4-, 7-, and 9-days post treatment (DPT) and exposed to a lab colony of TPB adults. One extra leaf sample/ plot/ spray /DPT interval (0-2-4-7-9-11) during 2016 was randomly collected from the high concentration plots and sent to Mississippi State Chemical Laboratory for residual analysis. Mortality of TPB adults was greatest for those placed on leaves sprayed with the organophosphate insecticide with mortalities (%) of 81.7±23.4 and 63.3±28.8 (SE) 1-day after exposure (DAE) on leaves 0-DPT with the high concentration for 2016 and 2017, respectively, reaching 94.5±9.5 and 95.4±7.6 6-DAE each year. Mortality to all insecticides continued until 9 and 4-DPT for high and medium concentrations, respectively. However, organophosphate (39.4±28.6) and pyrethroid (24.4±9.9) exhibited higher mortality than sulfoxamine (10.6±6.6) and the neonicotinoid (4.0±1.5) 7-DAE on 9-DPT leaves with the high concentration. Based on our results using the current assay procedure, TPB adults were significantly more susceptible to contact than systemic insecticides and due to its residual effect, organophosphate could kill over 80% of the TPB population 7-DPT.

Biochemical and molecular characterization of neonicotinoids resistance in the tarnished plant bug, Lygus lineolaris.

In the southern United States, neonicotinoids are commonly applied as foliar insecticides to control sucking insect pests, such as the tarnished plant bug (TPB, Lygus lineolaris). In this study, spraying bioassays were conducted to determine the toxicity of five neonicotinoids and sulfoxaflor to susceptible and late fall field-collected TPB adults from Mississippi Delta region. Compared to a susceptible population, the field-collected TPBs exhibited the highest resistance to imidacloprid (up to 19.5-fold), a moderate resistance to acetamiprid (9.43-fold), clothianidin (13.68-fold), thiamethoxam (7.88-fold) and the least resistance to thiacloprid (4.61-fold) and sulfoxaflor (1.82-fold), respectively. A synergist study demonstrated that piperonyl butoxide (PBO) significantly increased the toxicity of imidacloprid and thiamethoxam by 22.2- and 15.3-fold, respectively, while triphenyl phosphate (TPP) and diethyl maleate (DEM) only showed 2-3-fold synergism to both neonicotinoids. In the field-collected TPBs, activities of the three detoxification enzymes esterase, glutathione S-transferase (GST) and CYP450 monooxygenase (P450) were significantly increased by 3.43-, 1.48- and 2.70-fold, respectively, when compared to the susceptible population. Additionally, after 48 h exposure to imidacloprid or thiamethoxam, resistant TPB adults exhibited elevated esterase activities, decreased GST activities, and no significant changes in P450 activities. Further examinations revealed that the expression of certain esterase and P450 detoxification genes were significantly elevated in resistant TPBs. Overall, these results suggest that elevated esterase and P450s expression and enzyme activity are key mechanisms for metabolic resistance in TPBs to neonicotinoids. Our findings also provide valuable information for selection and adoption of neonicotinoid insecticides for resistance management of TPBs and minimizing toxic risk to foraging bees.

Microarray and Functional Pathway Analyses Revealed Significantly Elevated Gene Expressions Associated with Metabolic Resistance to Oxamyl (Vydate) in Lygus lineolaris.

The tarnished plant bug (TPB, Lygus lineolaris) remains a major pest for a variety of crops. Frequent sprays on row crops, especially cotton, prompted resistance development in field populations. To maintain chemical control as an effective tool against the pest, knowledge of global gene regulations is desirable for better understanding and managing the resistance. Novel microarray expressions of 6688 genes showed 685 significantly upregulated and 1382 significantly downregulated genes in oxamyl-selected TPBs (Vyd1515FF[R]) from a cotton field. Among the 685 upregulated genes (participated in 470 pathways), 176 genes code 30 different enzymes, and 7 of the 30 participate in 24 metabolic pathways. Six important detoxification pathways were controlled by 20 genes, coding 11 esterases, two P450s, two oxidases, and three pathway-associated enzymes (synthases, reductase, and dehydrogenase). Functional analyses showed substantially enhanced biological processes and molecular functions, with hydrolase activity as the most upregulated molecular function (controlled by 166 genes). Eleven esterases belong to the acting on ester bond subclass of the 166 hydrolases. Surprisingly, only one GST showed significant upregulation, but it was not involved in any detoxification pathway. Therefore, this research reports a set of 20 genes coding 6 enzyme classes to detoxify a carbamate insecticide oxamyl in Vyd1515FF. Together with three previous reports, we have obtained the best knowledge of resistance mechanisms to all four conventional insecticide classes in the economically important crop pest. This valuable finding will greatly facilitate the development of molecular tools to monitor and manage the resistance and to minimize risk to environment.

Sublethal effects of a commercial Bt product and Bt cotton flowers on the bollworm (Helicoverpa zea) with impacts to predation from a lady beetle (Hippodamia convergens).

Insecticidal Bacillus thuringiensis Berliner (Bt) toxins produced by transgenic cotton (Gossypium hirsutum L.) plants have become an essential component of cotton pest management. Bt toxins are the primary management tool in transgenic cotton for lepidopteran pests, the most important of which is the bollworm (Helicoverpa zea Boddie) (Lepidoptera: Noctuidae) in the United States (U.S.). However, bollworm larvae that survive after consuming Bt toxins may experience sublethal effects, which could alter interactions with other organisms, such as natural enemies. Experiments were conducted to evaluate how sublethal effects of a commercial Bt product (Dipel) incorporated into artificial diet and from Bt cotton flowers impact predation from the convergent lady beetle (Hippodamia convergens Guérin-Méneville) (Coleoptera: Coccinellidae), common in cotton fields of the mid-southern U.S. Sublethal effects were detected through reduced weight and slower development in bollworm larvae which fed on Dipel incorporated into artificial diet, Bollgard II, and Bollgard 3 cotton flowers. Sublethal effects from proteins incorporated into artificial diet were found to significantly alter predation from third instar lady beetle larvae. Predation of bollworm larvae also increased significantly after feeding for three days on a diet incorporated with Bt proteins. These results suggest that the changes in larval weight and development induced by Bt can be used to help predict consumption of bollworm larvae by the convergent lady beetle. These findings are essential to understanding the potential level of biological control in Bt cotton where lepidopteran larvae experience sublethal effects.

Toxicity Assessment of Four Formulated Pyrethroid-Containing Binary Insecticides in Two Resistant Adult Tarnished Plant Bug (Lygus lineolaris) Populations.

Over the past several decades, the extensive use of pyrethroids has led to the development of resistance in many insect populations, including the economically damaging pest tarnished plant bug (TPB), Lygus lineolaris, on cotton. To manage TPB resistance, several commercially formulated pyrethroid-containing binary mixtures, in combination with neonicotinoids or avermectin are recommended for TPB control and resistance management in the mid-South USA. This study aimed to evaluate the toxicity and resistance risks of four formulated pyrethroid-containing binary mixtures (Endigo, Leverage, Athena, and Hero) on one susceptible and two resistant TPB populations, which were field-collected in July (Field-R1) and October (Field-R2), respectively. Based on LC50 values, both resistant TPB populations displayed variable tolerance to the four binary mixtures, with Hero showing the highest resistance and Athena the lowest. Notably, the Field-R2 exhibited 1.5-3-fold higher resistance compared to the Field-R1 for all four binary insecticides. Moreover, both resistant TPB populations demonstrated significantly higher resistance ratios towards Hero and Leverage compared to their corresponding individual pyrethroid, while Endigo and Athena showed similar or lower resistance. This study also utilized the calculated additive index (AI) and co-toxicity coefficient (CTC) analysis, which revealed that the two individual components in Leverage exhibited antagonist effects against the two resistant TPB populations. In contrast, the two individual components in Endigo, Hero, and Athena displayed synergistic interactions. Considering that Hero is a mixture of two pyrethroids that can enhance the development of TPB resistance, our findings suggest that Endigo and Athena are likely superior products for slowing down resistance development in TPB populations. This study provides valuable insight for selecting the most effective mixtures to achieve better TPB control through synergistic toxicity analysis, while simultaneously reducing economic and environmental risks associated with resistance development in the insect pest.

Resistance risk assessment of six pyrethroids and acephate toward the resistant adult tarnished plant bug, Lygus lineolaris.

Due to rapidly developed resistance, pest management relies less on pyrethroids to control economically damaging infestations of the tarnished plant bug (TPB), Lygus lineolaris (Palisot de Beauvois) in cotton fields of Mississippi. Yet, pyrethroid resistance remains prevalent in TPB populations. This study assessed the resistance levels in adult TPB to six common pyrethroids and acephate. Resistant TBPs were collected from wild host plants in late October after harvest in the Mississippi Delta region of the United States. Based on LC50 values, the field-resistant TPBs displayed higher resistance to permethrin, esfenvalerate, and bifenthrin (approximately 30 fold) and moderate resistance to λ-cyhalothrin, ß-cyfluthrin, ζ-cypermethrin, and acephate (approximately 15 fold). Further investigations showed that the inhibitors of three detoxification enzyme, triphenyl phosphate (TPP), diethyl maleate (DEM), and piperonyl butoxide (PBO) had synergistic effects on permethrin, λ-cyhalothrin, and bifenthrin in resistant TPBs. Furthermore, elevated esterase, GST, and P450 activities were significantly expressed in field-resistant TPBs. Additionally, GST and esterase were reduced after 48 h exposure to certain pyrethroids at LC50 dose. The synergistic and biochemical assays consistently indicated that P450 and esterase were involved in pyrethroid detoxification in TPBs. This study provides valuable information for the continued use of pyrethroids and acephate in controlling TPBs in cotton fields in the Mississippi Delta region of the United States.

Entomopathogenicity of Ascomycete Fungus Cordyceps militaris on the Cotton Bollworm, Helicoverpa zea (Boddie) (Lepidoptera: Noctuidae).

This study investigated the exposure of the cotton bollworm, Helicoverpa zea (Boddie) (Lepidoptera: Noctuidae) to a novel pathogenic fungal agent historically associated with human medicinal value, a commercial strain of Cordyceps militaris ((L.) Fr.) Vuill. (Hypocreales). A series of comparative studies were conducted to evaluate the efficacy of two different exposure methods using four concentrations (n × 109, n × 108, n × 107, n × 106) of C. militaris, where n × 109 provided a concentration of approximately 420 ± 37 spores per mm2 with 398 ± 28 viable spores. Survival of cotton bollworms of all stages was not affected by C. militaris at any concentration 1 d post-exposure. The greatest reduction in survival and highest sporulation rates were observed primarily on or after 7 d post-exposure for early instars (first and second). Significant declines in the survival of early instars were observed for all concentrations at 7 d, and 95% mortality by 10 d, with the exception of the fifth instars that experienced a less severe reduction in survival (35%) when exposed to any concentrations used in the study. Survival of late instars (third to fifth) ranged from 44% to 68% on day 10, while adult survival was near 99% across the duration of the experiment. The relatively narrow range observed for both the lethal concentration and sporulation of second, third, and fifth instar cotton bollworms exposed to the C. militaris strain may demonstrate potential field application for control of larval populations of cotton bollworms.

The mechanisms of metabolic resistance to pyrethroids and neonicotinoids fade away without selection pressure in the tarnished plant bug Lygus lineolaris.

BACKGROUND: Heavy selection pressure prompted the development of resistance in a serious cotton pest tarnished plant bug (TPB), Lygus Lineolaris in the mid-southern United States. Conversely, a laboratory resistant TPB strain lost its resistance to five pyrethroids and two neonicotinoids after 36 generations without exposure to any insecticide. It is worthwhile to examine why the resistance diminished in this population and determine whether the resistance fade away has practical value for insecticide resistance management in TPB populations. RESULTS: A field-collected resistant TPB population in July (Field-R1) exhibited 3.90-14.37-fold resistance to five pyrethroids and two neonicotinoids, while another field-collected TPB population in April (Field-R2) showed much lower levels of resistance (0.84-3.78-fold) due to the absence of selection pressure. Interestingly, after 36 generations without exposure to insecticide, the resistance levels in the same population [laboratory resistant strain (Lab-R)] significantly decreased to 0.80-2.09-fold. The use of detoxification enzyme inhibitors had synergistic effects on permethrin, bifenthrin and imidacloprid in resistant populations of Lygus lineolaris. The synergism was more pronounced in Field-R2 than laboratory susceptible (Lab-S) and Lab-R TPB population. Moreover, esterase, glutathione S-transferase (GST), and cytochrome P450-monooxygenases (P450) enzyme activities increased significantly by approximately 1.92-, 1.43-, and 1.44-fold in Field-R1, respectively, and 1.38-fold increased P450 enzyme activities in Field-R2 TPB population, compared to the Lab-S TPB. In contrast, the three enzyme activities in the Lab-R strain were not significantly elevated anymore relative to the Lab-S population. Additionally, Field-R1 TPB showed elevated expression levels of certain esterase, GST and P450 genes, respectively, while Field-R2 TPB overexpressed only P450 genes. The elevation of these gene expression levels in Lab-R expectedly diminished to levels close to those of the Lab-S TPB populations. CONCLUSION: Our results indicated that the major mechanism of resistance in TPB populations was metabolic detoxification, and the resistance development was likely conferred by increased gene expressions of esterase, GST, and P450 genes, the fadeaway of the resistance may be caused by reversing the overexpression of esterase, GST and P450. Without pesticide selection, resistant gene (esterase, GST, P450s) frequencies declined, and detoxification enzyme activities returned to Lab-S level, which resulted in the recovery of the susceptibility in the resistant TPB populations. Therefore, pest's self-purging of insecticide resistance becomes strategically desirable for managing resistance in pest populations. Published 2023. This article is a U.S. Government work and is in the public domain in the USA.

Lethal Concentration and Sporulation by Contact and Direct Spray of the Entomopathogenic Fungus Beauveria bassiana on Different Stages of Nezara viridula (Heteroptera: Pentatomidae).

The southern green stink bug, Nezara viridula (L.) (Heteroptera: Pentatomidae) is the most significant pest of soybean worldwide. The present study was conducted to compare the effectiveness of a Delta native strain NI8 of Beauveria bassiana by contact and direct spray on nymphs (2nd to 5th instar) and adults of N. viridula. Water control and four concentrations of B. bassiana were used to evaluate the survival, mortality, and molting percentage and to estimate median lethal concentration (LC50), median lethal sporulation (LS50), and resistance ratio (RR50). Direct spray at all concentrations observed the greatest reduction in survival on all life stages. Mortality and sporulation were positively correlated by concentration, while molting was highly variable with a significantly lower negative correlation on insects that were directly sprayed. Pathogenicity exhibited reduction as young stages developed and emerged to adult. The LC50 (Contact: 612 spores/mm2; Direct spray: 179 spores/mm2) and LS50 (Contact: 1960 spores/mm2 Spray: 3.3 × 106) values showed that adults of N. viridula were highly resistant than any other life stage when exposed to either contact or direct spray. Fourth instar was the most susceptible (LC50: Contact: 18 spores/mm2; Direct spray: 23 spores/mm2) (LS50: Contact: 53 spores/mm2; Direct spray: 26 spores/mm2) followed by second, third, and fifth instars.

Susceptibility of Different Life Stages of Kudzu Bug Megacopta cribraria (F.) (Hemiptera: Plataspidae) to Two Different Native Strains of Beauveria bassiana.

This is the first study that examined and compared the survival, LC50, and RR50 estimates of Megacopta cribraria F. (Hemiptera: Plataspidae) nymphs and adults that were exposed to two native Beauveria bassiana isolates (Previously codified as NI8 and KUDSC strains) at four concentrations. The greatest reduction in survival and mortality was observed primarily on or after 10 d post-exposure to B. bassiana isolates. Survival of early instars (2nd, 3rd) were not affected by either strains or concentration at 3 d and 5 d post-exposure. Survival of later instars (5th) and adults was significantly reduced when exposed to the KUDSC strain at all concentrations. Comparison of dose−mortality values (LC50) using resistance ratios (RR50) were significantly different between life stages of the kudzu bug for both strains of B. bassiana. The LC50 values showed that kudzu bug adults are more susceptible than any other life stage when exposed to either strain. The KUDSC strain was more pathogenic than NI8 10 d after exposure, but NI8 exhibited significantly higher pathogenicity than KUDSC 20 d after exposure. Our results suggest potential field application of B. bassiana for kudzu bug control and their integration into pest management strategies to suppress them before they cause economic damage to soybean crops.

Effect of modification of the NI artificial diet on the biological fitness parameters of mass reared western tarnished plant bug, Lygus hesperus.

The NI artificial diet is the only known successful diet for mass rearing the western tarnished plant bug, Lygus hesperus Knight (Hemiptera: Miridae). This diet has been used for more than a decade. However, because it contains cooked chicken egg, and thus requires laborious preparation (Cohen 2000), this diet is difficult to use. Three modifications (D1, D2, D3) of the NI diet were investigated in hopes of developing a more easily prepared diet that avoids the cooked egg and improves mass fitness parameters of L. hesperus. The modified D3 diet, containing autoclaved chicken egg yolk based component, had the highest egg/cage/day production (13120 ± 812 SE). This was significantly greater than diets D1, containing autoclaved dry chicken egg yolk based component (9027 ± 811 SE), D2, containing autoclaved chicken egg white based component (8311 ± 628 SE), and NI, which contained autoclaved chicken egg yolk + cooked egg diet (7890 ± 761 SE). Significant differences were observed in the weights of all developmental stages except for eggs and first instar nymphs. Higher rates of fertility, hatchability, and low mortality in nymphs during the first instar were also obtained in the modified D3 diet. The results clearly indicated that the D3 diet provided an opportunity to significantly reduce rearing cost by avoiding time-consuming issues with preparation of a cooked egg diet. This should result in an increase in production capacity and a reduction in production costs.

Herbicide treatment alters the effects of water hyacinth on larval mosquito abundance.

Invasive aquatic weeds are managed with herbicides to reduce their negative impacts on waterways in many areas, including the California Delta Region. Herbicides create a dynamic environment of living and decomposing plant matter that could affect larval mosquitoes and other invertebrates, such as their predators and competitors. Our objective was to compare the number of larval mosquitoes in water or water hyacinth, before and after an herbicide treatment. We created replicated pond mesocosms with water hyacinth, water hyacinth treated with glyphosate and an oil adjuvant, open water, and water with glyphosate plus adjuvant. We sampled for larval mosquitoes and other aquatic invertebrates. Before herbicide addition, there was a trend for more larval mosquitoes in open water tanks than in tanks with water hyacinth. Herbicide application resulted in an immediate decrease of larval mosquitoes. As decay progressed, larval mosquitoes became most abundant in mesocosms with herbicide-treated hyacinth and very few larval mosquitoes were found in other habitat treatments. Although the numbers of predatory and competitor insects had some variation between treatments, no clear pattern emerged. This information on how invasive weed management with herbicides affects larval mosquitoes will allow control practices for larval mosquitoes and invasive weeds to be better integrated.