First Report of Root Rot on Mungbean Caused by Pythium myriotylum in China

During August and September 2020, symptoms of leaf chlorosis, stunting, and wilting were observed on indoor hemp plants (Cannabis sativa L. cv. ‘Wedding Cake’) in a commercial indoor facility located in Coolidge, Arizona. Plants were grown in soilless coconut coir growing medium (Worm Factory COIR250G10), watered with 1.5 to 2.1 liters every 24 h through drip irrigation, and supplemented with 18 h of lighting. About 35% of plants displayed symptoms as described above and many symptomatic plants collapsed. To identify the causal agent, crown and root tissues from four symptomatic plants were harvested and rinsed with tap water. Tissue fragments (approx. 2 to 4 mm in size) were excised from the margins of the stem and root lesions, surface sterilized in 0.6% sodium hypochlorite for 1 min, rinsed well in sterile distilled water, blotted dry, and plated on potato dextrose agar (PDA) and on oomycete-selective clarified V8 media containing pimaricin, ampicillin, rifampicin, and pentachloronitrobenzene (PARP). Plates were incubated at room temperature (21-24 oC). Five isolates resembling Pythium were transferred after 3 days and maintained on clarified V8 media. Morphological characteristics were observed on grass blade cultures (Waterhouse 1967). Grass blades were placed on CV8 inoculated with the isolate. After a 1-day incubation at 25°C, the colonized blades were transferred to 8 ml of soil water extract in a Petri dish. Ten sporangia and oogonia were selected randomly and their diameters were measured under the microscope. Sporangia were mostly filamentous, undifferentiated or inflated lobulate, ranging from 7 to 17 µm in diameter. Knob-like appressoria were observed on branching clusters. Bulbous-like antheridia were formed on branched stalk with 1-8 antheridia per oogonium. Globose oogonia were terminal or intercalary and ranged from 21 to 33 µm in diameter. Globose oospores were mostly aplerotic and ranged from 15 to 21 μm in diameter. Based on these morphological characteristics, isolates were tentatively identified as Pythium myriotylum (Watanabe, 2002). Genomic DNA was extracted from mycelial mats of two isolates using DNeasy Plant Pro Kit (Qiagen Inc., Valencia, CA) according to the manufacturer’s instructions. The internal transcribed spacer (ITS) region of rDNA was amplified with primers ITS1/ITS4 and two identical nucleotide sequences were obtained and deposited under accession number MW380925. A BLASTn search revealed ≥ 98% query coverage and 100% match with sequences HQ237488.1, KY019264.1, and KM434129, which were isolates of P. myriotylum from palm, tobacco, and ginger, respectively. To fulfill Koch’s postulates, pathogenicity tests were conducted with 2 isolates using plants of ‘Wedding Cake’ grown in 12 1.9-liter pots filled with a steam-disinfested potting mix (Sungro Professional Growing Mix). Pots were placed in a plastic container and watered to flooding three times a week. Plants were maintained in a greenhouse with 18 h/10 h day/night supplemental light cycle (15-28 oC). Plants were fertilized weekly with Peters Professional fertilizer at 1mg/ml. Four plants were inoculated with each isolate at three weeks after seed sowing by placing two 5-mm mycelial plugs from active growing 4 days-old cultures on PDA media adjacent to the main root mass at an approximately 3 cm depth. Four plants were inoculated with blank PDA plugs as controls. Symptoms of leaf chlorosis, crown rot and wilting were observed after four weeks while control plants remained symptomless. P. myriotylum was re-isolated from necrotic roots of inoculated plants after surface-sterilization, but not from control plants. The pathogenicity test was repeated once. While P. myriotylum often occurs in warmer regions and has a wide host range of >100 host plant species including numerous economically important crops (Wang et al., 2003), there are only two reports of this pathogen on indoor hemp plants in a greenhouse in Connecticut (McGehee et al., 2019) and in Canada (Punja et al., 2019). This is the first report of P. myriotylum causing root and crown rot of indoor hemp in Arizona. A more careful water management in soilless growth medium to reduce periods of saturation would minimize the risk of Pythium root rot in indoor hemp production.

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First Report of Root Rot of Cocoyam Caused by Pythium myriotylum in Sri Lanka

Plant Disease ◽

10.1094/pd-89-1132c ◽

2005 ◽

Vol 89 (10) ◽

pp. 1132-1132 ◽

Cited By ~ 2

Author(s):

M. Tojo ◽

H. Ono ◽

C. Nakashima ◽

S. Yoneyama ◽

J. A. S. Jayakody

Keyword(s):

Sri Lanka ◽

Root Rot ◽

Continuous Light ◽

Rdna Sequence ◽

Its Rdna ◽

Molecular Characteristics ◽

Root Tips ◽

Pythium Myriotylum ◽

First Report ◽

Initial Symptoms

Root rot of cocoyam (Xanthosoma sagittifolium L. Schott) caused by Pythium myriotylum Drechsler is a major disease of this crop in Africa (1,2) but is unreported from other regions of the world. During September 1999, commercially grown cocoyam (cv. Ratu-kiri-ala) in Gampaha (7°05′N, 80°00′E), Sri Lanka suffered from severe root rot. Initial symptoms were water-soaked lesions at the root tips that gradually enlarged to rot the entire root system and tuber. Wilting and yellowing of leaves were observed in advanced stages of disease. A Pythium sp. was regularly isolated from the affected roots and an isolate, SC5, was identified as P. myriotylum on the basis of morphology and the internal transcribed spacer (ITS) rDNA sequence. Characteristics of isolate SC5, grown on a grass-leaf water culture (3) were main hyphae up to 8.5 μm wide, oogonia terminal or intercalary (22.5 to 33.8 μm in diameter), antheridia diclinous occasionally monoclinous, one to eight per oogonium, stalks branched, often more or less loosely enveloping the oogonium, antheridium clavate or crook-necked, making apical contact with the oogonium, breadth of antheridium 2.5 to 7.0 μm, oospores aplerotic (17.0 to 22.5 μm in diameter), oospore wall 0.8 to 2.0 μm in thickness, sporangia terminal or intercalary, filamentous, inflated lobulate, and digitate, of variable length, breadth of sporangia 7.0 to 17.5 μm, formed in water; zoospores formed at 25°C, and diameter of encysted zoospores 10.0 to 12.5 μm. Cardinal temperatures on potato carrot agar 8°C minimum, 34°C optimum, and 37°C maximum with daily radial growth rate for 34°C at 32.8 mm. The ITS rDNA sequence of the isolate matched the sequences of P. myriotylum in GenBank (Accession Nos. AB095051 and AF452156) and isolate CBS254.70 used for the species description by van der Plaats-Niterink (3). The sequence of SC5 has been deposited in GenBank, Accession No. DQ102701. Pathogenicity tests used potted cocoyam plants (20 cm high), planted in an autoclaved potting mix. Four agar disks (8 mm in diameter) of isolate SC5 grown at 25°C for 48 h on potato dextrose agar was mashed and injected at a depth of 2 to 3 cm in the soil around the roots. Inoculated plants were placed in transparent plastic bags and kept for 7 days in a growth chamber maintained at 24 to 26°C with continuous light (52 to 98 μmol m-2·s-1). The experiment was carried out twice with three replications for each test. Dark brown rotting on roots and wilting of leaves were observed in 7 days after the inoculation. P. myriotylum was reisolated from diseased tissues and found to be morphologically identical to the original isolate SC5. Noninoculated control plants remained healthy. On the basis of the symptoms, morphological and molecular characteristics and confirmation of pathogenicity, P. myriotylum is the causal agent of root rot of cocoyam. To our knowledge, this is the first report of P. myriotylum causing root rot of cocoyam in Sri Lanka. References: (1) S. Nzietchueng. L'agronomie Tropicale 38:321, 1983. (2) R. P. Pacumbaba et al. J. Phytopathol. 135:265, 1992. (3) A. J. Van Der Plaats-Niterink. Stud. Mycol. 21:1, 1981.

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