Purification and Partial Characterization of Isometric Virus-like Particles in Kalanchoe Species

Isometric virus-like particles (IVLP) were detected in crude sap from Kalanchoe pinnata, K. daigremontiana and K. tubiflora plants showing a mild mosaic on the leaves. These particles of 35 nm in diameter were transmitted mechanically to several test plants but not to healthy Kalanchoe. Air temperatures above 30 °C hindered the infection process. The IVLP were purified from systematically infected Nicotiana benthamiana using Triton X-100 as clarifying agent followed byP, EG precipitation. IVLP were degraded by organic solvents and formed aggregates in the presence of 2 mmol/1 CaCl₂. The particles occurred in relatively low concentration in plant sap and lost infectivity in leaves frozen at -70 °C for one week and in purified preparations kept at 4 °C. In buffer crude sap of N. benthamiana IVLP had a thermal inactivation point between 45 and 50 °C on a longevity in vitro of 20 h at 25 °C. Particles contained one nucleoprotein component witha molecular weight of 46,000 daltons and a ssRNA species which, when denatured, had a molecular weight of 1.2 × 10⁶. IVLP purified preparations exhibited a typical nucleoprotein absorption spectrum with a maximum at 254–260 nm and a minimum at 240,–243 nm and a A 260/280 ratio of 1.56. The buoyant density of the IVLP was 1.32 g/ml calculated by isopycnic centrifugation on CsCl. Ultrastructural studies in infected leaves of K. pinnata indicated that IVLP caused an increase in chloroplast volume, distortion of the grana and reduced the number of thylakiods per grana. IVLP infection also impared the diurnal pattern of synthesis and hydrolysis of starch, characteristic of CAM plants. The non-serological reaction of the IVLP with antisera specific to members of 7 different groups of spherical viruses as well as the combination of physicochemical properties and host range, exhibited by these particles impeded their taxonomic location. In nature, young Kalanchoe plantlets acquire the IVLP through their physical connections with the infected mature leaves.