Biological Control Efficacy and Action Mechanism of Klebsiella pneumoniae JCK-2201 Producing Meso-2,3-Butanediol Against Tomato Bacterial Wilt

Abstract

Bacterial wilt caused by <i>Ralstonia solanacearum</i> is a fatal disease that affects the production of tomatoes and many other crops worldwide. As an effective strategy to manage bacterial wilt, biological control agents using plant growth-promoting rhizobacteria (PGPR) are being developed. In this study, we screened 2,3-butanediol (BDO)-producing PGPR to control tomato bacterial wilt and investigated the action mechanism of the disease control agent. Of the 943 strains isolated from soil, <i>Klebsiella pneumoniae</i> strain JCK-2201 produced the highest concentration of 2,3-BDO. The culture broth of <i>K. pneumoniae</i> JCK-2201 did not show any direct activity on <i>R. solanacearum in vitro</i>, but a 100-fold dilution effectively controlled tomato bacterial wilt with a control value of 77% <i>in vivo</i>. Fermentation utilizing <i>K. pneumoniae</i> JCK-2201 was optimized to produce 48 g/L of meso-2,3-BDO, which is 50% of the sucrose conversion efficiency. In addition, the control efficacy and mechanism of meso-2,3-BDO produced by JCK-2201 in tomato bacterial wilt were determined by comparative analysis with <i>Bacillus licheniformis</i> DSM13 producing meso-2,3-BDO and <i>B. licheniformis</i> DSM13 <i>ΔalsS</i> that did not produce 2,3-BDO, as the step of converting pyruvate to α-acetolactate was omitted. Tomato seedlings treated with the <i>K. pneumoniae</i> JCK-2201 (500-fold dilution) and <i>B. licheniformis</i> DSM13 (100-fold dilution) culture broth produced meso-2,3-BDO that significantly reduced <i>R. solanacearum</i>-induced disease severity with control values of 55% and 63%, respectively. The formulated meso-2,3-BDO 9% soluble concentrate (SL; 1,000-fold dilution) showed 87% control against tomato bacterial wilt in the field condition. <i>Klebsiella pneumoniae</i> JCK-2201 and <i>B. licheniformis</i> DSM13 treatment induced the expression of plant defense marker genes, such as <i>LePR1</i>, <i>LePR2</i>, <i>LePR5</i>, <i>LePR3</i>, and <i>PI-II</i>, in the salicylic acid and jasmonic acid signaling pathways at 4 days after inoculation. These results show that 2,3-BDO-producing bacteria and 2,3-BDO are potential biological control agents that act through induction of resistance for controlling tomato bacterial wilt.