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Research Project: Enhanced Alfalfa Germplasm and Genomic Resources for Yield, Quality, and Environmental Protection

Location: Plant Science Research

Title: First report of Mycoleptodiscus terrestris causing crown and root rot of alfalfa (Medicago sativa) in Minnesota

Author
item ZIVANOV, DALIBOR - Institute Of Field And Vegetable Crops
item ZIVANOV, SONJA TANCIC - Institute Of Field And Vegetable Crops
item Samac, Deborah - Debby

Submitted to: Plant Disease
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/10/2020
Publication Date: N/A
Citation: N/A

Interpretive Summary: In August 2019, alfalfa plants with stunted yellow foliage and rotted stems at the base of the crown were observed in a variety trial planted in spring 2018 at the University of Minnesota, St. Paul, Minnesota. Affected field plants were grouped in patches with approximately 20% of plants showing symptoms. Plants from patches had few lateral and fibrous roots and dark lesions appeared in the crown and root tissue. The fungus Mycoleptodiscus terrestris was isolated from infected plants and identified by morphological characteristics and DNA sequences from three genes. When inoculated onto alfalfa plants, the same disease symptoms were observed. The fungus is known to cause disease on several legume species including alfalfa, but it has not been identified previously causing disease on alfalfa in Minnesota. Information on occurrence of this pathogen will facilitate diagnosis of the disease and guide crop management strategies to reduce plant damage.

Technical Abstract: Mycoleptodiscus terrestris (Gerd.) Ostaz. has been studied extensively as a potential mycoherbicide against aquatic weeds since the early 1970s. However, it is also a pathogen on many legumes including alfalfa. In August 2019, alfalfa plants with stunted yellow foliage and rotted stems at the base of the crown were observed in a variety trial planted in spring 2018 at the University of Minnesota, St. Paul, Minnesota. Affected field plants were grouped in patches with approximately 20% of plants showing symptoms. Plants from patches had few lateral and fibrous roots and dark lesions appeared in the crown and root tissue. Samples (5 mm) from the symptomatic crown and root tissues were surface disinfected with 70% ethanol for 5 min, followed by 30 s in 10% NaOCl and then rinsed three times in sterile distilled water. Samples were air-dried and plated on water agar (WA) amended with 25 µg/ml rifampicin. Plates were incubated at room temperature for 3 days and hyphal tips of fungi growing from samples were transferred to potato dextrose agar (PDA). After 7 days, isolates were tentatively identified by morphological characteristics as M. terrestris. The hyaline mycelia turned from olive-gray to dark gray with age with abundant microsclerotia. The dark microsclerotia formed 5 days after incubation varied in size and shape and measured 300-860 x 270-600 µm. Molecular identification of two representative isolates (DAZ5, DAZ9) was done by sequencing regions of the rRNA internal transcribed spacer (ITS), the translation elongation factor 1a (TEF1), and the second largest subunit of RNA polymerase II (RPB2) genes. Based on a BLAST search of the NCBI nucleotide database, the ITS sequences (GenBank MN851265.1 and MN851266.1) had 100% identity with M. terrestris strain CBS 231.53 (MK487754.1). The TEF1 (MN873019, MN873020) and RPB2 sequences (MN873021, MN873022) had 100% identity with M. terrestris strain IMI 159038 (MK495977.1, MK492735.1). A pathogenicity test was performed by inoculating five 3-week-old alfalfa plants (cv. DKA44-16RR, Vernal, 53V52, and Agate) with two representative isolates (DAZ5 and DAZ9). Plants of each cultivar were inoculated around the exposed stem base with three PDA plugs from a culture of M. terrestris 5 mm in diameter covered with microsclerotia. Control plants were inoculated with sterile PDA plugs. After inoculation plants were incubated at 25°C with a 16-h photoperiod. The first symptoms appeared two months after inoculation on all cultivars as a dark lesion at the stem base followed with yellowing and rotting of stems. Inoculated plants had many more fibrous roots than controls. The control plants were symptomless with healthy root development. The pathogen was re-isolated from all infected alfalfa cultivars fulfilling Koch’s postulates. To our knowledge, this is the first report of M. terrestris causing crown and root rot disease of alfalfa in Minnesota.