First Report of Bacterial Leaf Blight Disease of Rice Caused by <i>Xanthomonas oryzae</i> pv. <i>oryzae</i> in Kenya

Bacterial leaf blight (BLB) caused by Xanthomonas oryzae pv. oryzae (Xoo) is a devastating disease of rice crops which can lead to yield losses ranging from 20 to 30% and as high as 50% (Mew 1987). BLB was first reported in Japan in 1884 and is now prevalent in most of the rice-producing countries in Asia. In Africa, Xoo was first reported in Mali and Cameroon (Buddenhagen et al., 1979) and later in many West African countries. Recently, BLB also appears to be spreading in East Africa with reports in Tanzania (Schepler-Luu et al., 2023) and in Madagascar (Raveloson et al., 2023). In June 2023, typical symptoms of BLB such as water-soaked areas and yellowish lesions along the leaf margins and tips were observed on rice plants in the coastal region of Kenya. Five rice fields of the Taita Taveta County with an incidence of 5-10% were surveyed, and 10-15 symptomatic leaf samples were collected in each field following a W pattern. Rice varieties were either Saro 5 or Arize 6444, two high productivity rice. 4-5cm long leaf fragments of 45 samples were next processed for bacterial isolation and surface disinfected, rinsed with sterile water, and ground individually in tubes with steel beads using a TissueLyser (Qiagen). Thirty-four round, pale yellow pigmented, and mucoid Xanthomonas-like colonies were obtained upon plating of 30 µl of the leaf powder resuspended in 1mL of sterile water on a semi-selective medium (peptone, 10 g/L; sucrose, 10 g/L; purified bacteriological agar, 16 g/L; glutamic acid, 1g/L; cycloheximide, 25 mg/L; cephalexin, 20 mg/L and kasugamycin, 10 mg/L) 72h after incubation at 28°C. To confirm that the isolates were Xoo, multiplex PCR established for the identification of X. oryzae pathovars was used (Lang et al. 2010). Thirty-two strains resulted in the amplification of two bands characteristic of the Xoo pattern. In addition, 6 of these strains (namely CIX5498, CIX5504, CIX5513, CIX5514, CIX5516 and CIX5539) were subjected to gyrB sequencing using the universal primers XgyrB1F and XgyrB1R (Young et al., 2008). The resulting partial 800 bp gyrB sequences of these 6 strains showed 100% identity (767/767 nt) with that of the PX099A Asian reference Xoo strain (accession CP000967.2). The pathogenicity of these 6 strains and the reference strain PX099A used as positive control was tested on 1-month-old Oryza sativa cv. Komboka plants (n=8 per treatment, 1 repetition). Rice leaves were inoculated by leaf clipping using sterile scissors dipped in the bacterial suspension at an optical density of 0.2. Three weeks after inoculation, plants exhibited typical BLB lesions for each of the six tested strains similarly to that of the positive control, whereas Mock-inoculated plants remained symptomless. Bacteria re-isolated from these diseased leaves yielded colonies that were confirmed as Xoo by multiplex PCR, fulfilling Koch's postulate. All strains isolated in this study are referenced in the CIX collection of the French National Research Institute for Sustainable Development in Montpellier. To our knowledge, this study represents the first molecular confirmation of the presence of BLB in Kenya and complement our knowledge of this important disease in East Africa. This work will serve as a basis for future rice breeding for BLB resistance efforts in Kenya. Further surveys are needed to clarify the spatial distribution and prevalence of BLB in other regions of Kenya and further analysis will be needed to understand the origin of the outbreak and to identify strategies to control BLB in Kenya.