Isolation and detection of bean yellow mosaic, clover yellow vein and peanut stunt viruses from Trifolium L. species

Abstract

Trifolium L. (clover) are annual or perennial species established in pasturelands to improve forage productivity and quality. In the southeastern United States, Trifolium repens L. (white clover) and Trifolium pratense L. (red clover) are important species, susceptible to virus infection. Objectives of this research were to isolate bean yellow mosaic (BYMV), clover yellow vein (CYVV) and peanut stunt (PSV) viruses from naturally infected white and red clovers from different locations in Virginia; and, to compare Indirect Enzyme-Linked ImmunoSorbent Assay (i-ELISA) and tissue immunoblot assay (TIBA) as methods for virus detection. A total of five white clover samples from Augusta, Richmond and Washington Counties were positive against CYVV antiserum and four white clover samples from Augusta County were positive against PSV antiserum. Single red clover samples from Frederick and Montgomery Counties were positive against BYMV antiserum. There were notable differences in host range with samples that tested positive for CYVV and BYMV, indicating they may be different strains. PSV was evenly distributed in the plant, whereas CYVV was higher in older plant parts. Viruses were successfully detected by blotting leaf samples directly onto membranes, thereby simplifying the sample preparation step. A number of membranes, such as nitrocellulose, nylon, chromatography paper, filter paper and writing pad could be used to detect viruses. In terms of specificity, immunoblots were equal or superior to i-ELISA. The TIBA should be useful in support of breeding and plant pathology studies as it is simple and rapid, and is less laborious and less expensive than i-ELISA.