Impact of Endochitinase-Transformed White Spruce on Soil Fungal Biomass and Ectendomycorrhizal Symbiosis

Abstract

The impact of transgenic white spruce [Picea glauca (Moench) Voss] containing the endochitinase gene (ech42) on soil fungal biomass and on the ectendomycorrhizal fungi Wilcoxina spp. was tested using a greenhouse trial. The measured level of endochitinase in roots of transgenic white spruce was up to 10 times higher than that in roots of nontransformed white spruce. The level of endochitinase in root exudates of three of four ech42-transformed lines was significantly greater than that in controls. Analysis soil ergosterol showed that the amount of fungal biomass in soil samples from control white spruce was slightly larger than that in soil samples from ech42-transformed white spruce. Nevertheless, the difference was not statistically significant. The rates of mycorrhizal colonization of transformed lines and controls were similar. Sequencing the internal transcribed spacer rRNA region revealed that the root tips were colonized by the ectendomycorrhizal fungi Wilcoxina spp. and the dark septate endophyte Phialocephala fortinii. Colonization of root tips by Wilcoxina spp. was monitored by real-time PCR to quantify the fungus present during the development of ectendomycorrhizal symbiosis in ech42-transformed and control lines. The numbers of Wilcoxina molecules in the transformed lines and the controls were not significantly different (P > 0.05, as determined by analysis of covariance), indicating that in spite of higher levels of endochitinase expression, mycorrhization was not inhibited. Our results indicate that the higher levels of chitinolytic activity in root exudates and root tissues from ech42-transformed lines did not alter the soil fungal biomass or the development of ectendomycorrhizal symbiosis involving Wilcoxina spp.

The effects of high-tannin leaf litter from transgenic poplars on microbial communities in microcosm soils

“…Hampp et al (1996) found that transformation of poplars with hygromycin marker or indoleacetic acid biosynthetic genes did not affect the ability of Amanita muscaria to form mycorrhizae. Results similar to these examples were obtained in work with endochitinase-transformed white spruce; Stefani et al (2010) did not detect any interference with ectendomycorrhizal Wilcoxina spp. and Lamarche et al (2011) did not detect any significant impacts of transformation on fungal biomass or community structure in such trees.…”

“…Laboratory, greenhouse, and field studies have largely reported little to no impact of transformation on microbial communities of soils (Hampp et al, 1996; Kaldorf et al, 2002; Pasonen et al, 2005; Lamarche and Hamelin, 2007; Newhouse et al, 2007; Seppänen et al, 2007; Oliver et al, 2008; Stefani et al, 2009, 2010; Lottman et al, 2010; Lamarche et al, 2011), although some have found significant changes in particular microbial groups (LeBlanc et al, 2007; Andreote et al, 2009). These responses to genetic modifications occur within a wider context of variable natural genotypic influences on microbial communities.…”

Impact of endochitinase-transformed white spruce on soil fungal communities under greenhouse conditions

“…Vauramo et al (2006) showed no negative effect on fungal biomass associated with decaying leaf litter by contrasting the ergosterol content of litters made of control and chitinase-expressing silver birch leaves after 8 months of decomposition in the field. Stefani et al (2010) showed that an increase of up to 10 times in endochitinase levels within root tissues of transformed white spruce lines did not prevent the colonization and development of ectendomycorrhizal symbiosis by Wilcoxina sp. They did not find evidence of a difference in the fungal structure community associated with transgenic and wild-type genotypes clearly related to the expression of sugar beet chitinase IV.…”

“…This growth reduction appeared to be associated with high lignin content and overexpression of peroxidase and glucanase activity (Faize et al, 2003), which are both involved in plant disease response to fungal pathogens. However, Stefani et al (2010) showed that ech42-transformed white spruce released endochitinase into soil through root exudates and the level of endochitinase in roots of transgenic white spruce was up to 10 times higher than that in roots of untransformed white spruce. These metabolism modifications could, in turn, have unexpected effects on nontarget fungal communities.…”

“…Also, similar transgenic apple trees, transformed with endochitinase and exochitinase genes from Trichoderma atroviride, showed a negative correlation between growth and endochitinase activity. Until now, investigation of the impact of chitinase-transformed plants on nontarget fungi has mainly focused on mycorrhizal fungi (Vierheilig et al, 1993;Pasonen et al, 2005;Schäfer et al, 2009;Stefani et al, 2010), and conclusions from those studies are conflicting (Vierheilig et al, 1993;Pasonen et al, 2005;Stefani et al, 2010). These examples show that the insertion and expression of the chitinase genes not only increase plant antifungal activities, but also affect apple tree metabolism.…”