Xanthomonas oryzae pv. oryzicola (Xoc) causes a destructive bacterial leaf streak disease in rice. Some of the gene products annotated as hypothetical proteins in the genome of Xoc may contribute to its virulence in rice. A mutant, Mxoc1679, screened from our previous Tn5-tagged mutant library for Xoc strain RS105, showed reduced virulence in rice. In this mutant, a gene named as Xoryp_08180 was disrupted by Tn5 insertion. Xoryp_08180 encodes a 1 306-aa hypothetical protein which is highly conserved in Xanthomonas spp. Non-polar mutation of Xoryp_08180 in RS105 strain led to a significant reduction in bacterial virulence and growth in rice, a delayed hypersensitive response (HR) in non-host tobacco, and a decrease in extracellular protease activity. The deficiencies above were restored to wild-type level in the complementary strain by expressing Xoryp_08180 in trans. In addition, the expression of Xoryp_08180 was repressed in hrpG and hrpX mutants in planta but not in a nutrient-rich condition. These results suggested that Xoryp_08180 is a virulence factor required for extracellular protease production, HR induction and full virulence of Xoc.

Xanthomonas oryzae pv. oryzicola (Xoc), the critical pathogen causing bacterial leaf streak in rice, possesses a hrp clusterthat is responsible for triggering hypersensitive response (HR) in non-host tobacco and pathogenicity in host rice, and isconsidered to be one of the model pathogens in the rice model plant. Here, we developed a high-throughput mutagenesissystem using a two-step integration mediated by a novel suicide vector pKMS1. It was used to generate single or poly-genemutants of hpa1, hpa2, hrcV, hrpE, hpaB, and hrpF gene for functional analysis. In total, five single, four double, and twotriple hrp gene mutants were constructed. The double and triple hrp gene deletion mutants triggered novel phenotypesin planta. Our data suggest that pKMS1 is a useful tool for non-marker mutagenesis of multiple genes in Xoc.

Two rice maintaining lines with different drought tolerances, viz., Jin 23B (tolerant) and Zhenshan 97B (sensitive), wereused to study the oxidative stress and soluble sugar in rice anthers and pollen viability under drought stress duringflowering stage. Higher antioxidant enzyme activities and lower malonaldehyde (MDA) content in rice anthers wereobserved in Jin 23B than in Zhenshan 97B under drought stress. Further, a great increase in the content of soluble sugarin rice anthers of Jin 23B was observed across the whole drought exposure, while Zhenshan 97B showed significantdecrease in soluble sugar during 9-12 d after drought stress (DADS). Accordingly, a marked decline of pollen fertility andactivity, pollen numbers in an anther and pollen numbers on a stigma was observed in Zhenshan 97B, whereas littledifference was found in Jin 23B. Thus, we suggest that pollen abortion caused by drought stress may be related with thereciprocity between oxidative stress and soluble sugar content in rice anthers.