First report of Fusarium sacchari causing sugarcane wilt in Bangladesh.

Wilt is one of the destructive fungal diseases of sugarcane (Saccharum officinarum L.) affecting sugarcane production in many sugarcane growing regions including Bangladesh. In July 2018, sugarcane plants showing wilting symptom including withered, dried leaves and shriveling stalks with brown to pinkish discoloration in internal tissues (Viswanathan, 2013) were collected from sugarcane field from Gazipur district (23°54'41″ N, 90°23'20″ E) of Bangladesh. Average disease incidence was 17% in 60 ha of sugarcane fields that were sampled for diseased plants with visible symptoms. Ten infected stalks (30cm) were collected from five severely infected fields. The stalks were surface sterilized with 70% ethanol for 1 minute followed by 0.2% sodium hypochlorite for 3 minutes and then rinsed three times with distilled water. Small pieces (2mm×2mm) of the inner discolored tissue were cut with a strile scalpel and placed on half strength Potato Dextrose Agar (PDA) medium followed by incubation for 7 days at 25 °C. A total of ten isolates were recovered from the infected sugarcane stalks and were purified by single spore culture method. Isolates produced floccose and cottony colony with white mycelia and dark violet pigmentation on the lower surface of the PDA plate. Microconidia were oval, generally, without septation measuring 5-9.6 µm length and 1.8-3.7 µm width in size (Appendix 1). Macro conidia were not found on the PDA medium. Conidiophores produced mono- or poly-phialides and conidia were arranged in a false head on conidiophore which is consistent with Fusarium sacchari (Duan et al. 2019). For molecular identification, DNA was extracted from two representative isolates BTFSS1 and BTFSS6. The internal transcribed spacer (ITS) region, translation elongation factor 1-α (TEF-1α) gene and RNA polymerase II (RPB2) genes were amplified using primers ITS1/ITS4, EF1α-F EF1α-R and 5F2/11aR, respectively (Appendix 2). Obtained ITS sequences (GenBank accession nos. MT176492.1 and MT177209.1) showed 99.8% similarity with the sequences from F. sacchari strain olowEKT1 (MK072727.1). Sequence identity of TEF-1α (GenBank accession nos. MW558264.1 and MW507839.1) was 99.36% with the sequence of F. sacchari strain FF031 (MK152501.1) and RPB2 (GenBank accession nos. MW558265.1 and MW507840.1) was 99.42% with the sequence of F. sacchari strain FF001 (MK152508.1), respectively. The isolates BTFSS1 and BTFSS6 were identified as F. sacchari based on the sequences alignment of ITS, TEF-1α and RPB2 and onolecular phylogenetic analyses by maximum likelihood tree method. To confirm the pathogen as a causal agent of wilt of sugarcane, pathogenicity tests were conducted in sugarcane plants by the plug method described by Viswanathan et al. (2011). Ten plants (variety BSRI Akh 42) were inoculated with 1 × 105 conidia mL-1 suspension of the isolate BTFSS1. A borehole was made on the third or fourth internode from the base of the sugarcane plant in where the conidial suspension was placed and the bore place was sealed with cane tissues. Control plants were inoculated with deionized distilled water. All canes were cuts longitudinally to evaluate the disease symptoms at fifteen and thirty days after inoculation. Inoculated stalks exhibited tissue discoloration that were similar to those observed in infected sugarcane field, whereas the control remained unaffected (Appendix 1). Pathogens were re-isolated from the artificially inoculated cane and characterized morphologically. Sugarcane is the second important cash crop and the only source of white sugar in Bangladesh (Rahman et al. 2016). Our report for the first time confirmed that F. sacchari is the causal pathogen of sugarcane wilt in Bangladesh. This report has significance to develop suitable management practices to control the pathogen in sugarcane field.