Cry3Bb1 protein from Bacillus thuringiensis in root exudates and biomass of transgenic corn does not persist in soil

Abstract

The Cry3Bb1 protein, insecticidal to the corn rootworm complex (Diabrotica spp.), of Bacillus thuringiensis (Bt) subsp. kumamotoensis was released in root exudates of transgenic Bt corn (event MON863) in sterile hydroponic culture (7.5 +/- 1.12 ng/ml after 28 days of growth) and in nonsterile soil throughout growth of the plants (2.2 +/- 0.62 ng/g after 63 days of growth). Kitchawan soil, which contains predominantly kaolinite (K) but not montmorillonite (M), was amended to 3 or 6% (vol./vol.) with K (3K and 6K soils) or M (3M and 6M soils) and with 1, 3, 5, or 10% (wt./wt.) of ground biomass of Bt corn expressing the Cry3Bb1 protein and incubated at 25 +/- 2 degrees C at the -33-kPa water tension for 60 days. Soils were analyzed for the presence of the protein every 7 to 10 days with a western blot assay (ImmunoStrip) and verified by ELISA. Persistence of the protein varied with the type and amount of clay mineral and the pH of the soils and increased as the concentration of K was increased but decreased as the concentration of M was increased. Persistence decreased when the pH of the K-amended soils was increased from ca. 5 to ca. 7 with CaCO(3): the protein was not detected after 14 and 21 days in the pH-adjusted 3K and 6K soils, respectively, whereas it was detected after 40 days in the 3K and 6K soils not adjusted to pH 7. The protein was detected for only 21 days in the 3M soil and for 14 days in the 6M soil, which were not adjusted in pH. These results indicate that the Cry3Bb1 protein does not persist or accumulate in soil and is degraded rapidly.

Fate and Effects in Soil of Cry Proteins from Bacillus thuringiensis: Influence of Physicochemical and Biological Characteristics of Soil

“…To determine whether the Cry3Bb1 protein is released in root exudates of Bt corn, two hybrids of corn, DeKalk DKC46-23 (event MON863) containing a truncated synthetic version of a gene from Btku coding for the expression of the anticoleopteran -endotoxin, Cry3Bb1, and its near-isogenic nongenetically modified variety, DKC46-26, were grown in sterile hydroponic culture and nonsterile soil [117]. The Cry3Bb1 protein was released in root exudates in sterile hydroponic culture (7.5 ± 1.12 ng/ml after 28 days of growth) and in nonsterile rhizosphere soil throughout growth of the plants (2.2 ± 0.62 ng/g after 63 days of growth), as determined by Western blots and ELISA (ImmunoStrip; Agdia, Elkhart, IN), similar to the Cry1Ab (in Bt corn and rice) and Cry3A (in Bt potato) proteins [26,30,31,[82][83][84], albeit at lower levels.…”

“…Other studies have shown no apparent deleterious effects of Cry proteins released by Bt plants in root exudates or from biomass of Bt plants incorporated into soil on microbial communities or representative enzymes [29,70,72,81,115,117,126,[164][165][166][167][168][169]. Studies using culture-independent methods reported minor or no Bt-specific effects on soil microorganisms, and the age and type of plant and the type and texture of soil seemed to be the major factors affecting bacterial diversity [159,[170][171][172].…”

Soil organisms as an essential element of a monitoring plan to identify the effects of GMO cultivation. Requirements – Methodology – Standardisation

“…soil type, pH, temperature, year, season, region etc.) makes it difficult to demonstrate the effects of GMOs [Pagel-Wieder et al 2004, Icozand andStotzky 2008]. These factors contribute to the variability of biological systems and have an impact on the potential disturbance value of the GMO (e.g.…”

Scientific Opinion on an application from Pioneer Hi‐Bred International and Dow AgroSciences LLC (EFSA‐GMO‐NL‐2005‐23) for placing on the market of genetically modified maize 59122 for food and feed uses, import, processing and cultivation under Regulation (EC) No 1829/2003

“…The data submitted by the applicant and the review of published literature did not indicate any interactions in the expression of the proteins or their biological activity compared with GM crops expressing similar single proteins. The newly expressed proteins in maize 59122 can be introduced into the soil via physical damage to plant tissues, via decomposition of shed root cells during plant growth, via decomposing plant residues remaining in fields after harvest, which might be incorporated into the soil during tillage operations (Stotzky, 2004), and via root exudation (e.g., Saxena et al, 2002Saxena et al, , 2004Icoz and Stotzky, 2007;, resulting in exposure of non-target soil organisms to the Cry34Ab1/Cry35Ab1 proteins. Indirect exposure through manure and faeces from animals fed maize 59122 was also considered, although most of the Cry34Ab1/Cry35Ab1 proteins would be degraded by enzymatic activity in the intestinal tract and subsequently by microbial processes in the manure.…”