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Resuscitation of Viable But Not Culturable Sinorhizobium meliloti 41 pRP4-luc: Effects of Oxygen and Host Plant

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Abstract

A plasmid-borne, firefly-derived, luciferase gene (luc) was inserted and stably inherited in Sinorhizobium meliloti 41 as a reporter gene. The strain obtained, S. meliloti 41/pRP4-luc, and its parental strain served as a model system for viable but not culturable (VBNC) resuscitation experiments in both in vitro and soil samples. Incubation under oxygen (O2) concentrations varying from 1% to atmospheric levels did not result in resuscitation. A demonstration of recovery was attained through exposure to the appropriate concentrations of antibiotics, bacteriostatic chloramphenicol, and bactericidal ampicillin. The resuscitation ratio was 1 recovered VBNC cell in every 105 5-cyano-2,3-di-4-tolyl-tetrazolium chloride (CTC+) bacteria. Although isolated VBNC rhizobia were unable to nodulate Medicago sativa, which apparently did not enhance VBNC reversion, resuscitated bacteria maintained their symbiotic properties. Soil experiments showed that the lack of O2 leads to onset of VBNC status as in liquid microcosm, but the number of recoverable and culturable cells decreased more drastically in soil.

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Acknowledgments

This research was supported by an EC Contract and by MiPAF (Panda Project). Silvana Povolo was the recipient of “Assegno di Ricerca” from the University of Padova.

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  1. Dipartimento di Biotecnologie Agrarie, Agripolis–Università di Padova, Viale dell’Università 16, 35020, Legnaro, Padova, Italy

    Marina Basaglia, Silvana Povolo & Sergio Casella

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  1. Marina Basaglia
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  2. Silvana Povolo
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  3. Sergio Casella
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Correspondence to Sergio Casella.

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Basaglia, M., Povolo, S. & Casella, S. Resuscitation of Viable But Not Culturable Sinorhizobium meliloti 41 pRP4-luc: Effects of Oxygen and Host Plant. Curr Microbiol 54, 167–174 (2007). https://doi.org/10.1007/s00284-005-0482-3

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  • DOI: https://doi.org/10.1007/s00284-005-0482-3

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