VIEW ARTICLE

Techniques

An Elutriation Method for Quantitative Isolation of Cylindrocladium crotalariae Microsclerotia from Peanut Field Soil. P. M. Phipps, Research Associate, Department of Plant Pathology, North Carolina State University, Raleigh 27607; M. K. Beute(2), and K. R. Barker(3). (2)(3)Associate Professor, and Professor, respectively, Department of Plant Pathology, North Carolina State University, Raleigh 27607. Phytopathology 66:1255-1259. Accepted for publication 12 April 1976. Copyright © 1976 The American Phytopathological Society, 3340 Pilot Knob Road, St. Paul, MN 55121. All rights reserved.. DOI: 10.1094/Phyto-66-1255.

Plant debris and microsclerotia of Cylindrocladium crotalariae were elutriated from soil using a semi-automatic elutriator which was designed for separating nematodes from soil. Plant debris (larger than 425 μm) and microsclerotia-size particles (38-425 μm) in soil were collected on 425- and 38-μm sieves, respectively. Plant debris from the 425-μm sieve was blended for 2 minutes in water, then concentrated on a 38-μm sieve. Each sieve residue was treated for 1 minute in 0.25% NaClO. After rinsing, the sieve residues were suspended in water and 5-ml subsamples were pipetted into 100 ml of an isolation medium at 45 C. The medium then was mixed and poured into 10 petri dishes. The isolation medium contained glucose, 15 g; KNO₃, 0.5 g; yeast extract, 0.5 g; KH₂PO₄, 1 g; MgSO₄·7H₂O, 0.5 g; Tergitol (NPX), 1 ml; thiabendazole, 1 mg; chloramphenicol, 100 mg; chlortetracycline, 40 mg; and 20 g agar per liter of water. After 5 days of incubation, the presence of brown microsclerotia and asexual sporulation permitted recognition and counting of C. crotalariae colonies in assay plates. The elutriation and enumeration procedure was effective in recovery of at least 91% of laboratory-grown microsclerotia in artificially infested soil. Numbers of microsclerotia in 23 naturally-infested, peanut field soils ranged from 0.2 to 72/g soil.