First report of <i>Stenotrophomonas maltophilia</i> causing Bacterial Leaf blight of Rice in West Bengal, India

A survey was conducted in August 2024 to assess the occurrence of bacterial leaf blight disease in the major rice-producing areas of Dakshin Dinajpur District, West Bengal, India. During the survey, bacterial leaf blight symptoms were observed over approximately 1 ha at Badkhorna (25.3285° N, 88.7847° E) with a disease incidence of 46.7% at the active tillering stage. Infected leaves with white to straw-colored streaks and wavy margins were collected and cut into small fragments (5 mm) from the junction between infected and healthy area. The segments were surface-sterilized with 70% ethanol for 1 min, rinsed thrice with sterile distilled water, and dried on sterilized filter paper. The leaf fragments were then placed in sterilized test tubes containing 2 ml distilled water for 20 min followed by fourfold serial dilutions and streaking onto Peptone Sucrose Agar (Peptone 5 g, Sucrose 20 g, K₂HPO₄ 0.5 g, MgSO₄·7H₂O 0.25 g, and Bacto Agar 15 g per litre of distilled water). After 48 h of incubation at 28±1°C, colonies appeared light yellow, convex, opaque, mucoid, circular, and smooth-edged. Purification was done taking single colonies in nutrient agar plates and stored at 4°C for future use. For pathogenicity testing, a bacterial suspension from a 48-hour-old nutrient broth culture was adjusted to 10⁷ CFU/ml using 2000V Vis Spectrophotometer at OD 600 nm and used to inoculate 45-day-old rice cv. Samba Mahsuri (BPT 5204) using the clip inoculation method (Ke et al. 2017). Seeds were pretreated with 500 ppm streptocycline (90% streptomycin sulfate and 10% tetracycline hydrochloride, w/w) to avoid seed-borne infection, while control plants received sterile water. Symptoms appeared within 5 days as greyish-white lesions progressing downward with wavy margins, and complete leaf blighting occurred with light brown necrosis within 15 d post-inoculation. No symptoms were observed in control plants. The pathogen was further reisolated for molecular identification. The 16S rRNA gene (~1500 bp) and two housekeeping genes (atpD and gapA) were amplified and sequenced (Kaiser et al. 2009). The sequences obtained were deposited in GenBank under accession numbers PX136222 (16S rRNA), PZ098256 (atpD) and PZ098257 (gapA). Phylogenetic analysis conducted using the Neighbor-Joining method in MEGA 11 confirmed that the isolate closely clustered with Stenotrophomonas maltophilia (KT970985.1), S. maltophilia (KT970983.1), S. maltophilia (KT970982.1) and S. maltophilia (PV992366.1) confirming its identity as S. maltophilia (Fig 2). To our knowledge, this is the first study to report S. maltophilia as a causative agent of bacterial leaf blight of rice in West Bengal, India. Previously, S. maltophilia species have been isolated from rice, causing Sanqi root soft rot disease (Zheng et al. 2022) and bacterial white stripe disease (Devi et al. 2002; Azmat et al. 2024). Identifying S. maltophilia as the causal agent of bacterial leaf blight in rice is significant for future prevention and control of this disease.