High Variations in Endospore Numbers within Tidal Flat Sediments Revealed by Quantification of Dipicolinic Acid

Abstract

Bacterial endospores are resting stages without detectable metabolism that can remain viable for extended periods and hence might accumulate in sediments during burial. They can be stained by fluorescence dyes and are suggested to contribute to total cell counts, but cannot be distinguished from vegetative cells by epifluorescence microscopy. Viable counts obtained after pasteurization of sediment samples have suggested a significant contribution of spores to sediment microbiota. However, the actual contribution of endospores to total cell counts in marine sediments is largely unknown, because of the lack of methods for reliable quantification. In the present study, dipicolinic acid (DPA), which is accumulated in the endospore core, was used to quantify endospores in up to 5.5 m long sediment cores collected from tidal flats off the German North Sea coast (Wadden Sea). Dipicolinic acid contents were determined fluorimetrically using a highly sensitive post-column complexation HPLC approach and ranged from 0.02 to 4.4 nmol DPA g ⁻¹ sediment dry weight. Dipicolinic acid contents of spores of pure cultures, that were isolated from the sampling area, were used for the conversion into endospore numbers. Estimated numbers ranged from 1× 10 ⁵ to 2× 10 ⁷ spores g ⁻¹ sediment dry weight. In the uppermost 50 cm of the sediment section endospore numbers represented less than 1% of the total cell counts. However, in the layers beneath their contribution to total cell counts apparently increased with depth reaching up to 10% of total cell counts. The endospore depth profile was irregular, but reflected the vertical changes in lithology. The highest endospore numbers were found in thin black mud layers, significantly lower numbers in sandy sediments.

Keywords:

• endospores
• dormant cells
• tidal flat sediments
• lithology
• quantification
• dipicolinic acid
• terbium dipicolinate fluorescence
• post-column complexation
• HPLC

Acknowledgments

We thank Melanie Beck for providing the pore water samples, Jan Tebben for his assistance in preparing the sediment samples, and Barry Cragg for discussion. We are grateful that total cell counts were provided by Antje Gittel and Beate Köpke. This work was supported by the Deutsche Forschungsgemeinschaft (DFG) through a grant for the Research Group BioGeoChemistry of Tidal Flats (grant no. RU 458/24).