Using a sequence characterized amplified region (SCAR) marker for detection of Bacillus strain TS02 sprayed on strawberry plants to bio-control powdery mildew in fields

Abstract

This work used sequence characterized amplified region (SCAR) marker to detect the Bacillus cereus strain in strawberry fields. The purpose was to develop an effective molecular method for detecting the functional target microorganisms applied in agricultural fields. A 3×10⁹ CFU/ml vegetative cell suspension based on the functional B. cereus strain TS02 was sprayed on strawberry plants to control powdery mildew. Primer pair LS400: 5’-TCC AAC TAC TTC TCC AT-3' and 3’-TTT GCC ATT ACA TAG AGT-5’ was used to amplify the 400 bp SCAR marker TSS₁. TSS₁ could detect TS02 specifically, in relation to other 7 Bacillus species and 6 B. cereus strains. Sensitivity detection of TSS₁ to the lowest DNA concentration of TS02 was 78 pg/μl, which corresponded to a density of 8×10⁵ CFU/ml cells of strain TS02. TS02 was specifically detected by TSS1 from the collected mixture of microorganisms on leaves during the days 1 - 12 after TS02 was sprayed in strawberry fields. TS02 DNA concentration reached about 2500 pg/μl during the days 5 - 7 after spraying and 156 pg/μl in the final day 12, which corresponded to a density of about 2.5×10⁷ and 1.6×10⁶ CFU/ml cells of TS02 respectively. These results showed that SCAR marker TSS1 is an effective method for the detection of strain TS02 after applied in fields. This research provided a simple and convenient method of detection of target microorganisms in fields, It could especially help monitor strain TS02 more effectively when widely been applied in agriculture.

Key words: Bacillus cereus strain, biocontrol, sequence characterized amplified region marker, strawberry, Sphaerotheca macularis.