Effects Of Endophytic Bacteria Colonization On The Degradation Of Thiamethoxam In Rice

Thiamethoxam(TMX),a second-generation neonicotinoid insecticide,has been widely used to control pests such as aphids and planthoppers.The residue of thiamethoxam in the environment brings great dangers to the safety of agricultural products,and also has a great impact on non-target organisms.In this paper,an endophyte with thiamethoxam degradation activity was selected and successfully colonized in rice.The effect of endophyte-plant interaction on the degradation of thiamethoxam and root exudates was studied.The main results are as follows:1.Methods for extraction and detection of TMX in minimal salt medium(MSM)and rice plants,amino acids and organic acids in root exudates were established.2.An endophytic bacterium belonging to the genus Enterobacter spp.was isolated from wild Ophiopedilum spp.and we named it TMX-6.This bacterium can degedrate the thiamethoxam in minimal salt medium with an initial concentration of 10 ppm TMX more than 20% within 10 days.The optimal degradation conditions of TMX of the bacterium TMX-6 were 8% inoculation amount,culture temperature 30 ° C,and pH 7.0.The three-parent hybridization method was successfully used to introduce stably expressed the plasmid containing the green fluorescent protein gene into the strain.Observe the root,stem and leaf tissues at different times after colonizing rice with endophyte through fluorescence confocal microscopy to confirm that endophyte can stably colonization in the rice plant,and the approximate distribution dynamics of endophyte on the ground and underground parts of rice plants were obtained by counting the colony in plate.3.Rice seedlings with hydroponic cultivation experiments were used to explore the effects of strains,un-inoculated rice and inoculated rice on the degradation of thiamethoxam,and to determine the changes of the metabolites of thiamethoxam in non-inoculated and inoculated rice plants at different periods.The results showed that both strains and rice could promote the degradation of thiamethoxam,and the ability to accelerate the degradation of thiamethoxam was further amplified in the endophyte-rice symbiotic system and the residue of thiamethoxam in the inoculated rice plants was lower than that of non-inoculated rice.The half-life of thiamethoxam for strain,rice and inoculated rice was 38.5 days,9.9 days and 4.7 days respectively,and the control group had a half-life of 46.2 days.The determination of thiamethoxam metabolites in rice plants was performed by time-of-flight mass spectrometry.The types and concentrations of thiamethoxam metabolites in inoculated rice plants were increased.4.Collect the root exudates of un-inoculated rice,inoculated rice,uninoculated rice treat with TMX,inoculated rice treat with TMX to determine the contents of 17 amino acids and 8 organic acids.The results showed that a variety of amino acids and organic acids in the root exudates changed significantly after endophytic colonization,and the difference between un-inoculated and inoculated rice might be more significant after the rice was exposed to TMX.Result shows that this significant difference mainly manifested in the early stage of the experiment,and the gap became very little in the end.