The costs and benefits of basal infection resistance vs immune priming responses in an insect.

In insects, basal pathogen resistance and immune priming can evolve as mutually exclusive strategies, with distinct infection outcomes. However, the evolutionary drivers of such diverse immune functions remain poorly understood. Here, we addressed this key issue by systematically analyzing the differential fitness costs and benefits of priming vs resistance evolution in Tribolium beetle populations infected with Bacillus thuringiensis. Surprisingly, resistant beetles had increased post-infection reproduction and a longer lifespan under both starving as well as fed conditions, with no other measurable costs. In contrast, priming reduced offspring early survival, development rate and reproduction. Priming did improve post-infection survival of offspring, but this added trans-generational benefit of immune priming might not compensate for its pervasive costs. Resistance was thus consistently more beneficial. Overall, our work demonstrates the evolutionary change in trans-generational priming response, and provides a detailed comparison of the complex fitness consequences of evolved priming vs resistance.

Bacterial larvicides used for malaria vector control in sub-Saharan Africa: review of their effectiveness and operational feasibility.

Several trials and reviews have outlined the potential role of larviciding for malaria control in sub-Saharan Africa (SSA) to supplement the core indoor insecticide-based interventions. It has been argued that widespread use of long-lasting insecticide-treated nets (LLINs) and indoor residual spraying (IRS) interventions in many parts of Africa result in many new areas with low and focal malaria transmission that can be targeted with larvicides. As some countries in SSA are making good progress in malaria control, larval source management, particularly with bacterial larvicides, could be included in the list of viable options to maintain the gains achieved while paving the way to malaria elimination. We conducted a review of published literature that investigated the application of bacterial larvicides, Bacillus thuringiensis var. israelensis (Bti) and/or Bacillus sphaericus (Bs) for malaria vector control in SSA. Data for the review were identified through PubMed, the extensive files of the authors and reference lists of relevant articles retrieved. A total of 56 relevant studies were identified and included in the review. The findings indicated that, at low application rates, bacterial larvicide products based on Bti and/or Bs were effective in controlling malaria vectors. The larvicide interventions were found to be feasible, accepted by the general community, safe to the non-target organisms and the costs compared fairly well with those of other vector control measures practiced in SSA. Our review suggests that larviciding should gain more ground as a tool for integrated malaria vector control due to the decline in malaria which creates more appropriate conditions for the intervention and to the recognition of limitations of insecticide-based vector control tools. The advancement of new technology for mapping landscapes and environments could moreover facilitate identification and targeting of the numerous larval habitats preferred by the African malaria vectors. To build sustainable anti-larval measures in SSA, there is a great need to build capacity in relevant specialties and develop organizational structures for governance and management of larval source management programmes.

Bt eggplant (Solanum melongena L.) in Bangladesh: Fruit production and control of eggplant fruit and shoot borer (Leucinodes orbonalis Guenee), effects on non-target arthropods and economic returns.

Eggplant or brinjal (Solanum melongena) is a popular vegetable grown throughout Asia where it is attacked by brinjal fruit and shoot borer (BFSB) (Leucinodes orbonalis). Yield losses in Bangladesh have been reported up to 86% and farmers rely primarily on frequent insecticide applications to reduce injury. Bangladesh has developed and released four brinjal varieties producing Cry1Ac (Bt brinjal) and is the first country to do so. We report on the first replicated field trials comparing four Bt brinjal varieties to their non-Bt isolines, with and without standard insecticide spray regimes. Results of the two-year study (2016-17) indicated Bt varieties had increased fruit production and minimal BFSB fruit infestation compared with their respective non-Bt isolines. Fruit infestation for Bt varieties varied from 0-2.27% in 2016, 0% in 2017, and was not significantly affected by the spray regime in either year. In contrast, fruit infestation in non-Bt lines reached 36.70% in 2016 and 45.51% in 2017, even with weekly spraying. An economic analysis revealed that all Bt lines had higher gross returns than their non-Bt isolines. The non-sprayed non-Bt isolines resulted in negative returns in most cases. Maximum fruit yield was obtained from sprayed plots compared to non-sprayed plots, indicating that other insects such as whiteflies, thrips and mites can reduce plant vigor and subsequent fruit weight. Statistically similar densities of non-target arthropods, including beneficial arthropods, were generally observed in both Bt and non-Bt varieties. An additional trial that focused on a single Bt variety and its isoline provided similar results on infestation levels, with and without sprays, and similarly demonstrated higher gross returns and no significant effects on non-target arthropods. Together, these studies indicate that the four Bt brinjal lines are extremely effective at controlling BFSB in Bangladesh without affecting other arthropods, and provide greater economic returns than their non-Bt isolines.

Novel larvicide tablets of Bacillus thuringiensis var. israelensis: Assessment of larvicidal effect on Aedes aegypti (Diptera: Culicidae) in Colombia.

INTRODUCTION: Aedes (Stegomyia) aegypti is the vector for dengue, chikungunya, and Zika arboviruses. Bti-CECIF is a bioinsecticide designed and developed in the form of a solid tablet for the control of this vector. It contains Bacillus thuringiensis var. israelensis (Bti) serotype H-14. OBJECTIVE: To evaluate under semi-field and field conditions the efficacy and residual activity of Bti-CECIF tablets on Aedes aegypti larvae in two Colombian municipalities. MATERIALS AND METHODS: We tested under semi-field conditions in plastic tanks (Rotoplast™) four different Bti doses (0.13, 0.40, 0.66 and 0.93 mg/L) in the municipality of Apartadó, department of Antioquia, to assess Bti-CECIF efficacy (percentage of reduction of larval density) and the residual activity in water tanks containing A. aegypti third-instar larvae. The efficacy and residuality of the most lethal dose were subsequently evaluated under field conditions in cement tanks in the municipality of San Carlos, department of Córdoba. RESULTS: Under semi-field conditions, the highest tested dose exhibited the greatest residual activity (15 days) after which larval mortality was 80%. Under field conditions, the highest tested Bti-CECIF doses showed 100% mortality and exhibited a residual activity of seven days in 90% of the tanks. CONCLUSION: Bti-CECIF tablets effectively controlled A. aegypti larvae under field conditions for up to seven days post-treatment.

Application of a dense genetic map for assessment of genomic responses to selection and inbreeding in Heliothis virescens.

Adaptation of pest species to laboratory conditions and selection for resistance to toxins in the laboratory are expected to cause inbreeding and genetic bottlenecks that reduce genetic variation. Heliothis virescens, a major cotton pest, has been colonized in the laboratory many times, and a few laboratory colonies have been selected for Bacillus thuringiensis (Bt) resistance. We developed 350-bp double-digest restriction-site associated DNA-sequencing (ddRAD-seq) molecular markers to examine and compare changes in genetic variation associated with laboratory adaptation, artificial selection and inbreeding in this nonmodel insect species. We found that allelic and nucleotide diversity declined dramatically in laboratory-reared H. virescens as compared with field-collected populations. The declines were primarily a result of the loss of low frequency alleles present in field-collected H. virescens. A further, albeit modest decline in genetic diversity was observed in a Bt-selected population. The greatest decline was seen in H. virescens that were sib-mated for 10 generations, in which more than 80% of loci were fixed for a single allele. To determine which regions of the genome were resistant to fixation in our sib-mated line, we generated a dense intraspecific linkage map containing three PCR-based and 659 ddRAD-seq markers. Markers that retained polymorphism were observed in small clusters spread over multiple linkage groups, but this clustering was not statistically significant. Overall, we have confirmed and extended the general expectations for reduced genetic diversity in laboratory colonies, provided tools for further genomic analyses and produced highly homozygous genomic DNA for future whole genome sequencing of H. virescens.

Lack of fitness costs and inheritance of resistance to Bacillus thuringiensis Cry1Ac toxin in a near-isogenic strain of Plutella xylostella (Lepidoptera: Plutellidae).

BACKGROUND: Resistance to Bacillus thuringiensis (Bt) formulations in insects may be associated with fitness costs. A lack of costs enables resistance alleles to persist, which may contribute to the rapid development and spread of resistance in populations. RESULTS: To assess the fitness costs associated with Bt Cry1Ac resistance in Plutella xylostella, life tables were constructed for a near-isogenic resistant strain (NIL-R) and a susceptible strain in this study. No fitness costs associated with Cry1Ac resistance in NIL-R were detected, based on the duration of egg and larval stages, the survival of eggs and larvae, adult longevity, fecundity, net reproductive rate, gross reproduction rate, finite rate of increase and mean generation time. Based on log dose-probit lines, resistance in NIL-R is incompletely recessive and results from a single, autosomal, recessive locus; the degree of dominance was estimated to be -0.74 and -0.71 for F1 (resistant ♀ × susceptible ♂) and F1 ' (susceptible ♀ × resistant ♂) progeny respectively. CONCLUSION: Assessment of near-isogenic Cry1Ac-resistant and Cry1Ac-susceptible strains of P. xylostella indicated that resistance is not accompanied with fitness costs, and that resistance is incompletely recessive. These findings should be useful in managing the development of Bt Cry1Ac resistance.

Assessment of Reactive Catch Basin Larvicide Treatments Toward Improved Water Quality Using FourStar(®) Briquets and CocoBear™ Larvicide Oil.

Because it is often logistically impossible to monitor all catch basins within an operational area, local mosquito control programs will preemptively treat catch basins with larvicides each season. However, these larvicides can, ostensibly, be considered water quality pollutants. To experimentally reduce the use of larvicides toward improving water quality, 30 basins within a small 0.7-km(2) residential area were monitored weekly for the presence of larvae and pupae for 14 wk in the summer of 2013. Once a basin was found to reach a threshold of 12 mosquitoes per dip sample, it received a FourStar® Briquet (a 180-day briquet formulation of 6% Bacillus sphaericus and 1% B. thuringiensis israelensis). Each week a FourStar-treated basin surpassed this threshold, it was treated with an application of CocoBear™ oil (10% mineral oil). By the end of trials, all but one basin received a briquet and 13 required at least 4 treatments of CocoBear, suggesting that preemptive treatment is appropriate for the study area.

Bacterial Cooperation Causes Systematic Errors in Pathogen Risk Assessment due to the Failure of the Independent Action Hypothesis.

The Independent Action Hypothesis (IAH) states that pathogenic individuals (cells, spores, virus particles etc.) behave independently of each other, so that each has an independent probability of causing systemic infection or death. The IAH is not just of basic scientific interest; it forms the basis of our current estimates of infectious disease risk in humans. Despite the important role of the IAH in managing disease interventions for food and water-borne pathogens, experimental support for the IAH in bacterial pathogens is indirect at best. Moreover since the IAH was first proposed, cooperative behaviors have been discovered in a wide range of microorganisms, including many pathogens. A fundamental principle of cooperation is that the fitness of individuals is affected by the presence and behaviors of others, which is contrary to the assumption of independent action. In this paper, we test the IAH in Bacillus thuringiensis (B.t), a widely occurring insect pathogen that releases toxins that benefit others in the inoculum, infecting the diamondback moth, Plutella xylostella. By experimentally separating B.t. spores from their toxins, we demonstrate that the IAH fails because there is an interaction between toxin and spore effects on mortality, where the toxin effect is synergistic and cannot be accommodated by independence assumptions. Finally, we show that applying recommended IAH dose-response models to high dose data leads to systematic overestimation of mortality risks at low doses, due to the presence of synergistic pathogen interactions. Our results show that cooperative secretions can easily invalidate the IAH, and that such mechanistic details should be incorporated into pathogen risk analysis.

Dietary mechanism behind the costs associated with resistance to Bacillus thuringiensis in the cabbage looper, Trichoplusia ni.

Beneficial alleles that spread rapidly as an adaptation to a new environment are often associated with costs that reduce the fitness of the population in the original environment. Several species of insect pests have evolved resistance to Bacillus thuringiensis (Bt) toxins in the field, jeopardizing its future use. This has most commonly occurred through the alteration of insect midgut binding sites specific for Bt toxins. While fitness costs related to Bt resistance alleles have often been recorded, the mechanisms behind them have remained obscure. We asked whether evolved resistance to Bt alters dietary nutrient intake, and if reduced efficiency of converting ingested nutrients to body growth are associated with fitness costs and variation in susceptibility to Bt. We fed the cabbage looper Trichoplusia ni artificial diets differing in levels of dietary imbalance in two major macronutrients, protein and digestible carbohydrate. By comparing a Bt-resistant T. ni strain with a susceptible strain we found that the mechanism behind reduced pupal weights and growth rates associated with Bt-resistance in T. ni was reduced consumption rather than impaired conversion of ingested nutrients to growth. In fact, Bt-resistant T. ni showed more efficient conversion of nutrients than the susceptible strain under certain dietary conditions. Although increasing levels of dietary protein prior to Bt challenge had a positive effect on larval survival, the LC50 of the resistant strain decreased when fed high levels of excess protein, whereas the LC50 of the susceptible strain continued to rise. Our study demonstrates that examining the nutritional basis of fitness costs may help elucidate the mechanisms underpinning them.

PROPOSED MEASURES OF CONTROL MANAGEMENT OF THE GRAPE MOTH, LOBESIA BOTRANA DEN AND SCHIFF (LEPIDOPTERA: TORTRICIDAE), IN REFERENCE TO INFESTATION PERCENTAGES, YIELD LOSS AND ECONOMICS OF CONTROL IN EGYPT.

This study aims to evaluate the efficacy of five measures; including the release of two strains of Trichogramma, Bacillus thuringiensis (B.t.). Sweetened B.t. and the environmentally friend insecticide (Tracer®), to manage the grape moth, Lobesia botrana that infested the grape vines c.v. "Thompson seedless " in Egypt severely during the two successive years of 2011 and 2012. In concern to the release of the worldwide, Trichogramma evanescens Westwood., the mean total grape yield ranged from 8.00 to 14.00 ton/fed and from 6.90 to 11.9, in respect to the two seasons and the mean infestation percentages fluctuated between 50.00 and 16.14; 43.48 and 21.01 in sequence for the same years. The corresponding grape bunches loss percentages averaged from 21.43 to 6.91; 18.63 to 9.00,respectively for the same periods of this study. Whereas, it was noticed that in releasing the indigenous T. bourarchae was more effective than T. evanescens in reducing all the above mentioned grape yield particularly. Applying Bacillus thuringiensis (B.t.) and/or adding sugar as a stimulant to improve yields reduced the infestation and the crop loss percentage. However, Bacillus thuringiensis mixed with sugar which overweighed the use of Bacillus thuringiensis alone. Although Tracer® provided good harvest in both years, but unfortunately it affected the grape berries components rather than the other treatments. Control plots indicated 48 and 50% infestation, with very low yield compared with the other five trials to manage the assigned insect pest. For the first time worldwide economics and profits were computed taking into account the whole market sale, the total control costs and the gross income to decide the most profitable treatment for the table grape growers under the Egyptian conditions. It was found that the mean (L.E.) profits/fed were arranged in a descending order as follows: 470.95, 513.28, 504.78, 500.82, 397.25 and 0.00 for Bacillus thuringiensis + sugar, Trichogramma bourarchae, Trichogramma evanescens Westwood, Dipel-2x®, Tracer® and control in respect in 2011. The corresponding values in the year 2012 were 453.85, 445.58, 427.18, 377.00, 349.18 and 0.00 for Trichogramma bourarchae, Bacillus thuringiensis + sugar, Dipel-2x®, Trichogramma evanescens Westwood, Tracer® and control in respect. The obtained results were analysed and discussed in the text.

Improvement of bioinsecticides production by sporeless Bacillus thuringiensis strains in response to various stresses in low cost medium.

The use of bioinsecticides, particularly those produced by sporeless Bacillus thuringiensis strains, has been shown to be a good alternative in pest management. Two types of sporeless mutants were distinguished. The asporogenic mutants which completely lack spores produce a regular bipyramidal crystal inclusion. The oligosporogenic mutants kept the ability to produce insecticidal crystal proteins. However, sporulation in such mutants was not totally blocked and very few of them could still produce spores. In order to improve bioinsecticides production, adaptation of sporeless strains to heat shock and osmotic stress was investigated. Delta-endotoxin production by 78% of sporeless mutants was significantly improved by osmotic stress with an overproduction of about 17%, compared to the wild strain BNS3. However, toxin production was improved by only 21% of mutants after heat shock, in low cost medium. The statistical analysis proved that delta-endotoxin production, cell growth, and spore formation of asporogenic and oligosporogenic mutants depended on the type of applied stress. Each strain has an important potential when applying the adequate stress. Moreover, adaptation of sporeless mutants to NaCl may allow the substitution of all minerals of the medium by diluted sea water which appeared to be a good alternative for the economic production of bioinsecticides at industrial scale which is of great importance from the practical point of view.

Wood losses and economical threshold of Btk aerial spray operation against spruce budworm.

BACKGROUND: Spruce budworm, Choristoneura fumiferana (Clem.), causes cumulative defoliation and hence annual growth loss of the balsam fir, Abies balsamea (L.) Mill, host tree. Annual growth increments of mixed balsam fir stands were measured by stem analysis over a 9 year period (1994-2002), when Bacillus thuringiensis ssp. kurstaki (Btk) was applied to control spruce budworm defoliation. With this approach, it was possible to quantify the change in stand volume growth after aerial spray applications of Btk. RESULTS: Differences between the periodic volume increment of protected and unprotected plots were statistically significant, while differences between protected and budworm-free plots were not significant. After 9 years, the difference in periodic increment between protected and unprotected plots was 20 m(3) ha(-1), and the difference in periodic mortality was 20.5 m(3) ha(-1). CONCLUSION: An economic assessment of Btk treatments indicates that biopesticide aerial spraying operations are justified, as they prevented substantial balsam fir mortality and growth losses over the 9 year study.

Comparison of Bacillus thuringiensis corn hybrids and insecticide-treated isolines exposed to bivoltine European corn borer (Lepidoptera: Crambidae) in South Dakota.

The performances of Bacillus thuringiensis (Bt) corn hybrids and permethrin-treated isolines were evaluated in South Dakota from 1997 to 1999. The corn hybrids were exposed to natural seasonal fluctuations of a bivoltine ecotype European corn borer population. Larval injury parameters, grain yields, and gross incomes were quantified during the 3-yr study. The use of either Bt hybrids or whorl-applied permethrin granules resulted in significant yield advantages in only 1 out of 3 yr of performance testing. Under high corn borer pressure in 1997, the performances of the Bt corn hybrids and permethrin-treated isolines were similar, and significantly better than the untreated isolines. However, injury-free corn did not necessarily translate into higher yields in some of the hybrid groups. Grain moisture at harvest, which can result in moisture penalty or dockage, was significantly higher in some Bt corn hybrids. Neither the Bt hybrids nor the permethrin-treated isolines produced significant advantages in yields in 1998 and 1999 when infestations of first-generation corn borer were almost nonexistent and infestations of second-generation European corn borers were moderate.

Risk assessment of etofenprox (Vectron) on non-target aquatic fauna compared with other pesticides used as Simulium larvicide in a tropical environment.

Within the rotational scheme developed by the Programme to fight the resistance of Simulium damnosum to chemical larvicides, there was an operational gap at discharges between 5 and 70 m3 s(-1) for the treatment of rivers where resistance to organophosphates was present. The use of permethrin and carbosulfan was precluded because of risk of environmental impact and, Bacillus thuringiensis ser. H-14 treatments were not envisageable due to cost and logistics constraints. Among the possible complementary groups of larvicides tested, the pseudo-pyrethroids, held promise, because of a mode of action similar to that of pyrethroids, but along with a usually lower toxicity for fish. Etofenprox, one of the pseudo-pyrethroids tested, shows a global detachment of non-target insects in 24 h close to that of pyraclofos, an organo-phosphorus compound (27 against 23%). In laboratory conditions, six times the operational dose which is 0.03 mg l(-1) 10 min, is needed to cause 50% mortality of Caridina sp. (a small shrimps species) and 30 times this same dose for 95% mortality. For fish species, a safety margin of 400-800 times the operational dose is observed for Oreochromis niloticus and 200-400 times for Tilapia zillii.

First steps to improve cotton crop management in Thailand.

Thai cotton growers yearly face serious pest problems consisting of heavy infestation associated with resistance to pesticides leading to expensive and harmful pest control practices. Thanks to a multi discipline research work carried out from 1991 to 1996, a decision-oriented model for cotton crop management was developed and compared to farmers' practices during the 1988 cotton growing season 1998 in Klang Hat district, Thailand. This model, based on plant monitoring, involved a new cotton cultivar (DORA 11 as a replacement for Sri Samrong 60), seed treatment with imidacloprid (at 3.5 g of a.i./kg of seeds), a threshold-based spray of mepiquat chloride, a cotton growth regulator (Pix at 1.5 l/ha) and a set of rules to selectively control with chemical and biological insecticides the major pests the American bollworm (Helicoverpa armigera Hübner) and the cotton leafhopper (Amrasca biguttula biguttula Ishida). This package was more friendly towards environment than farmers' practices since seed treatment does not harm beneficials at the beginning of cotton growing season while 1 and 5 sprays were saved against respectively the American bollworm and sucking pests and more than 50% of the sprays directed against American bollworms involved biological insecticides (Bacillus thuringiensis toxins potentialized by a low dosage of endosulfan). Threshold-based sprays against the main pests, the use of a cotton growth regulator, earliness of cotton cultivar and seed treatment were accountable for savings in pesticide sprays. Involving a biological insecticide and endosulfan at the beginning of cotton growing season and using the same quantity of active ingredient at each spray during the cotton growing season also improved the management of pyrethroid resistance in American bollworm populations was also. There was no significant difference between yields inside farmers' fields and innovation plots. However, thanks to cheaper pest control practices economic results were better. Lastly few improvements of this package are proposed.

Evaluating the environmental effects of the GM revolution.

In the US, cradle of genetically modified crops, agricultural scientists and regulators are already tackling one environmental issue, and rushing to determine the magnitude of another.

Bacillus thuringiensis: fermentation process and risk assessment. A short review.

Several factors make the local production of Bacillus thuringiensis (Bt) highly appropriate for pest control in developing nations. Bt can be cheaply produced on a wide variety of low cost, organic substrates. Local production results in considerable savings in hard currency which otherwise would be spent on importation of chemical and biological insecticides. The use of Bt in Brazil has been limited in comparison with chemical insecticides. Although Bt is imported, some Brazilian researchers have been working on its development and production. Fermentation processes (submerged and semi-solid) were applied, using by-products from agro-industries. As the semi-solid fermentation process demonstrated to be interesting for Bt endotoxins production, it could be adopted for small scale local production. Although promising results had been achieved, national products have not been registered due to the absence of a specific legislation for biological products. Effective actions are being developed in order to solve this gap. Regardless of the biocontrol agents being considered atoxic and harmless to the environment, information related to direct and indirect effects of microbials are still insufficient in many cases. The risk analysis of the use of microbial control agents is of upmost importance nowadays, and is also discussed.