Importance of the Host Phenotype on the Preservation of the Genetic Diversity in Codling Moth Granulovirus.

To test the importance of the host genotype in maintaining virus genetic diversity, five experimental populations were constructed by mixing two Cydiapomonella granulovirus isolates, the Mexican isolate CpGV-M and the CpGV-R5, in ratios of 99% M + 1% R, 95% M + 5% R, 90% M + 10% R, 50% M + 50% R, and 10% M + 90% R. CpGV-M and CpGV-R5 differ in their ability to replicate in codling moth larvae carrying the type I resistance. This ability is associated with a genetic marker located in the virus pe38 gene. Six successive cycles of replication were carried out with each virus population on a fully-permissive codling moth colony (CpNPP), as well as on a host colony (RGV) that carries the type I resistance, and thus blocks CpGV-M replication. The infectivity of offspring viruses was tested on both hosts. Replication on the CpNPP leads to virus lineages preserving the pe38 markers characteristic of both isolates, while replication on the RGV colony drastically reduces the frequency of the CpGV-M pe38 marker. Virus progeny obtained after replication on CpNPP show consistently higher pathogenicity than that of progeny viruses obtained by replication on RGV, independently of the host used for testing.

Variation in physiological host range in three strains of two species of the entomopathogenic fungus Beauveria.

Knowledge of the host range of a biocontrol agent (BCA) is fundamental. Host range determines the BCA's economic potential, as well as the possible risk for non-target organisms. Entomopathogenic fungal strains belonging to the genus Beauveria are widely used as BCA, but our knowledge of their physiological host range is only partial. The aim of this study was to improve our understanding of the physiological host range of three Beauveria strains belonging to two species, B. hoplocheli and B. bassiana. We performed laboratory mortality bioassays to assess their pathogenicity and virulence against nine insect pests, belonging to three orders: Lepidoptera, Coleoptera and Diptera. Mortality rate, mean survival time and mycosis rate were used to estimate virulence. Pathogenicity was assessed as the capacity to cause a disease and induce mortality. Virulence was assessed as the severity of the disease based on mortality rate, mean survival time and mycosis rate. The results of this study revealed significant differences in the physiological host range of the three Beauveria strains tested. The three strains were pathogenic to all Diptera and Lepidoptera species tested. In the case of the Coleoptera, only the B. hoplocheli strain was pathogenic to the white grub Hoplochelus marginalis and only the B. bassiana strains were pathogenic to Alphitobius diaperinus. The B. hoplocheli strain was less virulent on Lepidoptera and Diptera than the two B. bassiana strains. The latter both exhibited very similar virulence patterns. The fact that B. hoplocheli and B. bassiana strains have different host ranges means that they can be used as BCA to target different pests. Impacts on non-target insects across multiple orders cannot be ruled out in the absence of ecological host range studies.

Determination of Formulation Conditions Allowing Double Emulsions Stabilized by PGPR and Sodium Caseinate to Be Used as Capsules.

Water-in-oil-in-water (W1/O/W2) double emulsions stabilized by polyglycerol polyricinoleate (PGPR), a lipophilic food grade small polymer, and sodium caseinate, a hydrophilic milk protein, were developed to encapsulate vitamin B12, a model hydrophilic substance easy to titrate. Using rheology, sensitive to drop size evolution and water fluxes, static light scattering, and microscopy both giving the evolution of drops' size and vitamin B12 titration assessing the encapsulation, we were able to detect independently the double emulsion drop size, the encapsulation loss, and the flux of water as a function of time. By differentiating the PGPR required to cover the W1-droplets' surface from PGPR in excess in the oil phase, we built a PGPR-inner droplet volume fraction diagram highlighting the domains where the double emulsion is stable toward encapsulation and/or water fluxes. We demonstrated the key role played by nonadsorbed PGPR concentration in the intermediate sunflower oil phase on the emulsion stability while, surprisingly, the inner droplet volume fraction had no effect on the emulsion stability. At low PGPR concentration, a release of vitamin B12 was observed and the leakage mechanism of coalescence between droplets and oil-water interface of the oily drops (also called globules hereafter), was identified using confocal microscopy. For high enough PGPR content, the emulsions were stable and may therefore serve as efficient capsules without need of an additional gelling, thickening, complexion or interface rigidifying agent. We generalized these results with the encapsulation of an insecticide: Cydia pomonella granulovirus used in organic arboriculture.

Biological Characteristics of Experimental Genotype Mixtures of Cydia Pomonella Granulovirus (CpGV): Ability to Control Susceptible and Resistant Pest Populations.

The detection of resistance in codling moth (Cydia pomonella) populations against the Mexican isolate of its granulovirus (CpGV-M), raised questions on the sustainability of the use of this biological insecticide. In resistant host cells, CpGV-M is not able to complete its replication cycle because replication is blocked at an early step. Virus isolates able to overcome this resistance have been characterized-among them, the CpGV-R5 isolate. In mixed infections on resistant insects, both CpGV-M and CpGV-R5 viruses replicate, while CpGV-M alone does not induce mortality. Genetically heterogeneous virus populations, containing 50% of each CpGV-M and CpGV-R5 appear to control resistant host populations as well as CpGV-R5 alone at the same final concentration, even if the concentration of CpGV-R5 is only half in the former. The use of mixed genotype virus preparations instead of genotypically homogeneous populations may constitute a better approach than traditional methods for the development of baculovirus-based biological insecticides.

Water fluxes and encapsulation efficiency in double emulsions: impact of emulsification and osmotic pressure unbalance.

We study the influence of the emulsification process on encapsulation efficiency of drugs in double water-in-oil-in-water emulsions. Two drugs were used, first vitamin B12 which can be considered as a model drug and secondly a suspension of Cydia pomonella Granulovirus (CpGV), a virus used in organic agriculture to protect fruits against the Carpocapse insect. Encapsulation is measured by classical UV-Vis spectroscopy method. Additionally we show that rheology is a useful tool to determine water exchanges during emulsification. In a two-step emulsification process, using rotor-stator mixers, encapsulation reaches high levels, close to 100% whatever the flowing regime. This encapsulation decreases only if two conditions are fulfilled simultaneously: (i) during the second emulsification step the flow is turbulent and (ii) it leads to excessive fragmentation inducing formation of too small drops. We also investigate the effect of a deliberate loss of osmotic pressure balance on the encapsulation and characterize the induced water fluxes. We show that encapsulation of vitamin B12 is not affected by the osmotic pressure unbalance, while water exchanges, if they exist, are very fast and aim at restoring equilibrium. As a consequence, the emulsification efficiency is not very sensitive to osmotic stresses provided that the interfaces resist mechanically.

Resistance to bio-insecticides or how to enhance their sustainability: a review.

After more than 70 years of chemical pesticide use, modern agriculture is increasingly using biological control products. Resistances to conventional insecticides are wide spread, while those to bio-insecticides have raised less attention, and resistance management is frequently neglected. However, a good knowledge of the limitations of a new technique often provides greater sustainability. In this review, we compile cases of resistance to widely used bio-insecticides and describe the associated resistance mechanisms. This overview shows that all widely used bio-insecticides ultimately select resistant individuals. For example, at least 27 species of insects have been described as resistant to Bacillus thuringiensis toxins. The resistance mechanisms are at least as diverse as those that are involved in resistance to chemical insecticides, some of them being common to bio-insecticides and chemical insecticides. This analysis highlights the specific properties of bio-insecticides that the scientific community should use to provide a better sustainability of these products.

Progressive adaptation of a CpGV isolate to codling moth populations resistant to CpGV-M.

The NPP-R1 isolate of CpGV is able to replicate on CpGV-M-resistant codling moths. However, its efficacy is not sufficient to provide acceptable levels of control in natural (orchard) conditions. A laboratory colony derived from resistant codling moths was established, which exhibited a homogeneous genetic background and a resistance level more than 7000 fold. By successive cycles of replication of NPP-R1 in this colony, we observed a progressive increase in efficacy. After 16 cycles (isolate 2016-r16), the efficacy of the virus isolate was equivalent to that of CpGV-M on susceptible insects. This isolate was able to control both CpGV-M-susceptible and CpGV-M-resistant insects with similar efficacy. No reduction in the levels of occlusion body production in susceptible larvae was observed for 2016-r16 compared to CpGV-M.