Confirmation of Bacterial Leaf Streak of Rice Caused by Xanthomonas oryzae pv. oryzicola in Vietnam
Résumé
Xanthomonas oryzae is one of the most important pathogens of rice (Oryza sativa) (Niño-Liu et al. 2006). In early October of 2013, Vietnamese colleagues reported a high prevalence of leaf bacterial diseases on irrigated rice fields of the Red River delta (western coast of the Gulf of Tonkin). We performed an exploratory survey in provinces of North Vietnam. In the Hanoi, Hung Yen, and Nam Dinh provinces, we observed symptoms reminiscent of bacteria leaf streak (BLS) with variable (<5 to 90%) prevalence. While the presence of BLS disease in Vietnam has been recorded in international databases (Saddler 2002), to our knowledge, the role of X. oryzae pv. oryzicola as the causal agent has never been unambiguously established. Field samples exhibited typical BLS symptoms, such as water-soaked, linear lesions running along the veins and developing into yellow to brown streaks with yellow droplets of exudates at the surface. Bacteria were isolated by grinding 4-cm-long leaf pieces in sterile water. Tissue lysates were plated on semiselective PSA medium (Poulin et al. 2014) and incubated for 3 to 4 days at 28°C. Single, yellow-pigmented, mucoid colonies were further purified on PSA medium. In parallel, diagnostic multiplex PCR was performed on these colonies for microbial identification (Lang et al. 2010). In total, isolates from 15 independent samples yielded a PCR profile characteristic of X. oryzae pv. oryzicola. Six of these strains from the Nam Dinh and Hanoi provinces were further analyzed. First, a portion of their gyrB gene was sequenced following PCR amplification (Young et al. 2008). All six 780-bp, quality-trimmed gyrB sequences were deposited in GenBank under Accession Nos. KP872841 to KP872846. They are identical to the gyrB sequence of strain BLS256 (Accession No. CP003057), a reference strain for the oryzicola pathovar, and exhibited 10 polymorphic positions relative to the one from PXO99A (Accession No. CP000967), a oryzae pathovar reference strain. These strains were also tested for pathogenicity by infiltrating suspensions at 1 × 108 CFU/ml in water into 4-week-old O. sativa cv. Nipponbare leaves with a needleless syringe (3 infiltrations × 5 plants per strain). Leaves inoculated with Vietnamese isolates were indistinguishable from positive controls challenged with strain BLS256. In contrast to water-infiltrated negative controls, typical water-soaked lesions appeared after five days in the greenhouse (27°C, 12-h photoperiod, ∼80% humidity) and ultimately produced yellow exudates resembling those initially observed in rice fields. For bacterial reisolation, symptomatic tissue lysates were streaked on PSA medium. Bacterial colonies showed a typical Xanthomonas morphology and were again assigned to the oryzicola pathovar by multiplex PCR typing, thus fulfilling Koch’s postulates. These data confirm the status of X. oryzae pv. oryzicola as the causal agent of BLS in two provinces of Vietnam using modern molecular identification tools. To facilitate inquiries on the phylogeny and virulence of Vietnamese X. oryzae pv. oryzicola, strains VXO32 and VXO39 have been deposited in the Collection Française de Bactéries Phytopathogènes under Accession Nos. CFBP8314 and CFBP8315, respectively.