Catalysed reporter deposition–fluorescence in situ hybridization (CARD-FISH) is a powerful approach to quantify bacterial taxa. In this study, we compare the performance of the widely used Bacteroidetes CF319a probe with the new CF968 probe. In silico analyses and tests with isolates demonstrate that CF319a hybridizes with non-Bacteroidetes sequences from the Rhodobacteraceae and Alteromonadaceae families. We test the probes’ accuracy in 37 globally distributed marine samples and over two consecutive years at the Blanes Bay Microbial Observatory (NW Mediterranean). We also compared the CARD-FISH data with the Bacteroidetes 16S rRNA gene sequences retrieved from 27 marine metagenomes from the TARA Oceans expedition. We find no significant differences in abundances between both approaches, although CF319a targeted some unspecific sequences and both probes displayed different abundances of specific Bacteroidetes phylotypes. Our results demonstrate that quantitative estimations by using both probes are significantly different in certain oceanographic regions (Mediterranean Sea, Red Sea and Arabian Sea) and that CF968 shows seasonality within marine Bacteroidetes, notably large differences between summer and winter that is overlooked by CF319a. We propose CF968 as an alternative to CF319a for targeting the whole Bacteroidetes phylum since it has better coverage, greater specificity and overall better quantifies marine Bacteroidetes

This work was supported by Grants BACTERIOMICS (CTM2010-12317-E) and MicroOcean PANGENOMICS (CGL2011-26848/BOS) to SGA and HotMix (CTM2011-30010/MAR) to JMG from the Spanish Ministry of Science and Innovation (MICINN). SGA was supported by FP7-OCEAN-2011 (MicroB3) from the EU. Financial support was provided by Ph.D. JAE-Predoc fellowships from CSIC to CD-V, CR-G and GS. FMC-C was supported by a FPI fellowship (MICINN). Postdoctoral Juan de la Cierva research contracts also from MICINN were held by IF and HS. HS also benefited from a fellowship from the Brazilian ‘Ciências sem Fronteiras’ Program from CAPES (BJT 013/ 2012)

Special issue: Marine Microbes.-- This is Contribution No. 16 of the TARA Oceans expedition 2009–2012.-- 13 pages, 5 figures, additional supporting information http://onlinelibrary.wiley.com/doi/10.1111/1462-2920.12517/suppinfo

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