First report of brown leaf spot of rice caused by Curvularia spicifera in Pakistan
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First report of brown leaf spot of rice caused by Curvularia spicifera in Pakistan Hafiz Muhammad Usman Aslam 1 & Safdar Ali 1 & Muhammad Atiq 1 & Muhammad Zeeshan Mansha 2 & Hafiz Muhammad Aatif 2 & Hafiz Arslan Anwaar 3 & Khalid Naveed 4 Received: 25 August 2019 / Accepted: 30 January 2020 # Società Italiana di Patologia Vegetale (S.I.Pa.V.) 2020
Keywords Pathogen detection . Fungal diversity . Field crops . Molecular analysis Brown leaf spot of rice is the most important disease of rice that decreases the rice production up to 52% (Barnwal et al. 2013). In 2018, a survey of five major rice growing areas of Punjab (Nankana, Narowal, Hafizabad, Sialkot and Gujranwala) was conducted to investigate this disease. Brown-black, 6–9 mm in diam. Necrotic lesions surrounded by yellow halos were observed on the infected leaves. To isolate the pathogen, infected portion of the leaves was cut into 5 mm2 pieces which were disinfected in 0.1% HgCl2 followed by 70% ethanol, rinsed with distilled sterile water, put onto potato dextrose agar (PDA) and incubated for one week at 25 °C. The resulting colonies displayed velvety mycelium that turned dark green-grayish black after one week. Conidiophores were dispersed or grouped, light-medium brown, 102 to 115 μm in length and 4.7 to 7.4 μm in width (Jeon et al. 2015). Conidia were light brown, ellipsoidal or oblong, thinner towards the ends, 3-septate, 18.2–30.7 × 7– 11.6 μm. Two representative isolates (US-1 and US-4) were submitted in the Fungal Culture Bank of Plant Pathology, UAF. For molecular identification, GAPDH, TEF and ITS region of the isolate (US-1) were amplified using primers GPD1F/GPD2R (F-CAACGGCTTCGGTCGCATTG/R-
GCCAAGCAGTTGGTTGTGC), EF1-983F/EF1-2218R (FGCYCCYGGHCAYCGTGAYTTYAT/R-ATGACACC RACRGCRACRGTYTG) and ITS1F/ITS4R (FTCCGTAGGTGAACCTGCGG/R-TCCTCCGCTTATTG ATATGC), respectively. The sequences of GAPDH (MN173003), TEF (MN173002) and ITS (MN153760), were deposited in GenBank. A BLASTn queries revealed that all the amplifications were 99% identical to reference sequences of Curvularia spicifera (Bainier) Boedijn (MH809680, KM062878, KC999911). Pathogenicity experiment was performed by inoculating 10 seedlings of cv. Basmati-385 at twoleaf stage with a spore inoculum (105 spores/ml) by using an atomizer. Ten control plants were sprayed with distilled, sterile water and incubated at 25 °C for one week. The pathogen was re-isolated and its taxonomy was confirmed morphologically. To our knowledge, this is the first report of C. spicifera causing brown leaf spot of rice in Pakistan. Funding information This study was funded by Higher Education Commission of Pakistan grant number 2789.
Compliance with ethical standards Conflict of interest The authors do not have any conflict of interest.
* Hafiz Muhammad Usman Aslam [email protected] 1
Department of Plant Pathology, University of Agriculture, Faisalabad, Pakistan
2
Department of Plant Pathology, College of Agriculture BZU, Bahadur Sub-campus, Layyah, Pakistan
3
Department of Plant Pathology,
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