Effect of storage temperatures on Rhizobium meliloti survival in peat- and clay-based inoculants

Abstract

Legume growers and extension agents require advice on optimum conditions under which inoculants may be stored without deterioration of rhizobia or the effectiveness of the inoculant. This study was conducted to determine effects of storage duration and temperature on survival of Rhizobium meliloti in two inoculant carriers, one with a peat base and the other with a clay base. Inoculant subsamples were placed in polyethylene bags, then packaged into plastic containers and stored in wooden boxes at either −23 °C (freezer), 4 °C (refrigerator), or room temperature (21–33 °C). Before and after storage for 0.5, 3, 6, 9, 12, 15 and 20 mo, samples were analyzed for surviving rhizobia by most probable number (MPN) plant-infection technique using plastic pouches with seedlings of alfalfa (Medicago sativa L. ’Algonquin’). These counts showed no significant reduction of rhizobial viability in either inoculant at any temperature over the 20-mo study. The surprising lack of population decreases was attributed partly to the wide span between the confidence limits of MPN estimates (at P = 0.05) and partly to the effectiveness of the protective packaging in preventing any desiccation or acidification of the inoculants. Throughout the experiment pH remained unchanged at 6.8 and 8.5 and moisture content at 12.3 and 3.0% in the peat-base and clay-base inoculants, respectively. Nodule color and size indicated that symbiotic effectiveness was not affected by storage temperature. Counts, when averaged across temperatures, showed rhizobial populations remained constant around 1.3 × 109 cells g−1 peat and 1.1 × 108 cells g−1 clay. Re-testing of R. meliloti inoculants stored for up to 10 yr at 4 °C, after earlier regulatory testing, showed no significant viability reductions in any product stored < 7 yr and in some products stored for as long as 10 yr. The results indicated that rhizobial populations could be maintained at levels much above the regulatory minimum MPN (103 seed−1), which would be of critical importance when forage legumes are grown under adverse soil conditions and inoculation is required. Key words: Rhizobium meliloti, inoculant storage, temperature effects, carrier effects, MPN plant-infection technique, inoculant quality control