Development of a 16S rRNA-targeted fluorescence in situhybridization probe for quantification of the ammonia-oxidizer Nitrosotalea devanaterra and its relatives

Restrepo-Ortiz, Claudia X.Merbt, S.Barrero-Canossa, J.Fuchs, B. M.Casamayor, Emilio O.  Systematic and Applied Microbiology : doi:10.1016/j.syapm.2018.04.003 (2018)  DIGITAL CSIC

The Thaumarchaeota SAGMCG-1 group and, in particular, members of the genus Nitrosotalea have highoccurrence in acidic soils, the rhizosphere, groundwater and oligotrophic lakes, and play a potential rolein nitrogen cycling. In this study, the specific oligonucleotide fluorescence in situ hybridization probeSAG357 was designed for this Thaumarchaeota group based on the available 16S rRNA gene sequencesin databases, and included the ammonia-oxidizing species Nitrosotalea devanaterra. Cell permeabiliza-tion for catalyzed reporter deposition fluorescence in situ detection and the hybridization conditionswere optimized on enrichment cultures of the target species N. devanaterra, as well as the non-targetammonia-oxidizing archaeon Nitrosopumilus maritimus. Probe specificity was improved with a competi-tor oligonucleotide, and fluorescence intensity and cell visualization were enhanced by the design andapplication of two adjacent helpers. Probe performance was tested in soil samples along a pH gradient,and counting results matched the expected in situ distributions. Probe SAG357 and the CARD-FISH pro-tocol developed in the present study will help to improve the current understanding of the ecology andphysiology of N. devanaterra and its relatives in natural environments.