Desiccation induces viable but Non-Culturable cells in Sinorhizobium meliloti 1021
Jan Ac Vriezen1, Frans J de Bruijn, Klaus R Nüsslein
1Department of Microbiology, University of Massachusetts, Amherst, Massachusetts, USA, MA 01003. nusslein@microbio.umass.edu.
Abstract:
Sinorhizobium meliloti is a microorganism commercially used in the production of e.g. Medicago sativa seed inocula. Many inocula are powder-based and production includes a drying step. Although S. meliloti survives drying well, the quality of the inocula is reduced during this process. In this study we determined survival during desiccation of the commercial strains 102F84 and 102F85 as well as the model strain USDA1021.The survival of S. meliloti 1021 was estimated during nine weeks at 22% relative humidity. We found that after an initial rapid decline of colony forming units, the decline slowed to a steady 10-fold reduction in colony forming units every 22 days. In spite of the reduction in colony forming units, the fraction of the population identified as viable (42-54%) based on the Baclight live/dead stain did not change significantly over time. This change in the ability of viable cells to form colonies shows (i) an underestimation of the survival of rhizobial cells using plating methods, and that (ii) in a part of the population desiccation induces a Viable But Non Culturable (VBNC)-like state, which has not been reported before. Resuscitation attempts did not lead to a higher recovery of colony forming units indicating the VBNC state is stable under the conditions tested. This observation has important consequences for the use of rhizobia. Finding methods to resuscitate this fraction may increase the quality of powder-based seed inocula.
Insights
Desiccation induces a Viable But Non Culturable (VBNC)-like state in Sinorhizobium meliloti, impacting seed inocula quality. This state may be stable, suggesting improved methods are needed for rhizobial recovery and inocula enhancement.
Area of Science:
- Microbiology
- Agricultural Science
- Biotechnology
Background:
- Sinorhizobium meliloti is crucial for Medicago sativa seed inocula production.
- Powder-based inocula involve a drying step that reduces quality.
- Understanding desiccation survival is key to improving inocula.
Purpose of the Study:
- To determine the survival of Sinorhizobium meliloti strains during desiccation.
- To investigate the viability and culturability of S. meliloti under low humidity.
- To explore the potential for a Viable But Non Culturable (VBNC)-like state induced by desiccation.
Main Methods:
- Assessing survival of S. meliloti strains (102F84, 102F85, USDA1021) during nine weeks at 22% relative humidity.
- Estimating colony-forming units (CFUs) over time.
- Utilizing Baclight live/dead stain to assess cell viability.
- Conducting resuscitation attempts to recover non-culturable cells.
Main Results:
- A steady 10-fold CFU reduction every 22 days was observed after initial decline.
- Viability (42-54%) assessed by Baclight stain remained constant, despite reduced CFU counts.
- Desiccation induced a stable Viable But Non Culturable (VBNC)-like state in a portion of the S. meliloti population.
- Resuscitation attempts did not significantly increase CFU recovery.
Conclusions:
- Plating methods may underestimate rhizobial survival due to the VBNC-like state.
- Desiccation induces a stable VBNC-like state in Sinorhizobium meliloti, previously unreported.
- Developing resuscitation methods could enhance the quality of powder-based seed inocula.
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