Selection for and Analysis of UV-Resistant Cryptophlebia Leucotreta Granulovirus-SA as a Biopesticide for Thaumatotibia leucotreta.

Cryptophlebia leucotreta granulovirus-SA (CrleGV-SA) is used as a commercial biopesticide for the false codling moth, Thaumatotibia leucotreta, in citrus and other crops. The virus is sensitive to UV irradiation from sunlight, which reduces its efficacy as a biopesticide in the field. We selected a UV-resistant CrleGV-SA isolate, with more than a thousand-fold improved virulence compared to the wild-type isolate, measured by comparing LC50 values. CrleGV-SA purified from infected T. leucotreta larvae was exposed to UV irradiation under controlled laboratory conditions in a climate chamber mimicking field conditions. Five cycles of UV exposure, followed by propagating the virus that retained infectivity in vivo with re-exposure to UV, were conducted to isolate and select for UV-resistant virus. Serial dilution bioassays were conducted against neonates after each UV exposure cycle. The concentration-responses of the infectious UV-exposed virus populations were compared by probit analysis with those from previous cycles and from the original CrleGV-SA virus population. NGS sequences of CrleGV-SA samples from UV exposure cycle 1 and cycle 5 were compared with the GenBank CrleGV-SA sequence. Changes in the genomes of infective virus from cycles 1 and 5 generated SNPs thought to be responsible for establishing UV tolerance. Additional SNPs, detected only in the cycle 5 sequence, may enhance UV tolerance and improve the virulence of the UV-tolerant population.

Evaluation of spray-dried lignin-based formulations and adjuvants as solar protectants for the granulovirus of the codling moth, Cydia pomonella (L).

Commercial formulations of the codling moth, Cydia pomonella L., granulovirus (CpGV) are limited by their short residual activity under orchard conditions in the Pacific Northwest. We evaluated spray-dried lignin-encapsulated formulations of CpGV for improved solar stability based on laboratory bioassays with a solar simulator and in field tests in an infested apple orchard. In laboratory tests, aqueous lignin formulations containing a high dosage of 3 x 10(10) occlusion bodies (OB)/L, with and without the additives titanium dioxide (TiO(2)) and sugar, provided significant solar protection of virus, i.e., mortality of codling moth exposed to lignin formulations that had been irradiated with 9.36 x 10(6) joules/m(2) was 92-94%, compared with 66-67% from a glycerin-stabilized product (Cyd-X) or suspension of pure unformulated virus at the same rates. By comparison, a lower dosage of the lignin formulation (3 x 10(8)OB/L) did not provide significant solar protection. Equivalent dosage-dependent patterns in solar protection were observed in further tests with the lignin formulation, when an intermediate (3 x 10(9)OB/L) as well as the low dosage provided no solar protection. Equivalent rates of a blank lignin formulation (containing no virus) did not affect larval mortality, suggesting a protective effect of the lignin on the virus at the high rate. The use of several spray adjuvants, 'NuFilm-17' and 'Organic Biolink' (sticker-spreaders at 0.06% v/v), 'Raynox' (sunburn protectant at 5% v/v), and 'Trilogy'(neem oil at 1% v/v) did not provide solar protection of a commercial CpGV preparation in laboratory tests. In season long orchard tests (Golden Delicious), the lignin formulation of CpGV applied at 6.57 x 10(12)OB/ha did not significantly improve control of codling moth or protection of fruit compared with Cyd-X at equivalent rates. Our studies show that lignin-based CpGV formulations provided solar protection at relatively high virus dosages. The testing of lignin formulations containing reduced virus concentrations may allow virus solar protection to be achieved at more economical rates.

New method for testing solar sensitivity of commercial formulations of the granulovirus of codling moth (Cydia pomonella, Tortricidae: Lepidoptera).

A method for screening codling moth granulovirus (CpGV) formulation sensitivity to sunlight using specially prepared half apples and a solar simulator is described. The half apple preparation allows an even coverage of virus over the surface of the fruit that would not be possible using whole apples. Leaves and artificial medium were not usable for extended periods of exposure in the solar simulator due to excess drying. Fruit was sprayed with 10(-3) and 10(-5) dilutions of three commercial formulations of CpGV (Carpovirusine, Cyd-X, and Virosoft) and infested with codling moth neonates. Half of the sprayed fruit was exposed to 650 W/m2 for 4 h in an Atlas Suntest CPS solar simulator resulting in an accumulated radiant energy of 9.36x10(6) J/m2 before they were infested with neonate codling moth larvae. Spraying non-irradiated fruit with the 10(-3) dilution of Cyd-X and Virosoft resulted in nearly 100% mortality of neonate larvae. Irradiation reduced viral activity by 71-98% at the 10(-3) dilution and by up to 32% at the 10(-5) dilution relative to non-irradiated fruit. The procedures utilized enabled good preservation of the fruit throughout the incubation period and minimized invasion of the fruit by plant pathogens and saprophytic organisms. This laboratory method for screening candidate formulations and potential UV protectants could conserve time and resources by eliminating adjuvants with less potential in laboratory tests and field testing only the most promising candidates. It also enables year-round testing.