Culture-dependent and culture-independent diversity surveys target different bacteria: a case study in a freshwater sample Ivone Vaz-Moreira • Conceic ¸a ˜o Egas • Olga C. Nunes • Ce ´lia M. Manaia Abstract Compared with culture-independent appr- oaches, traditionally used culture-dependent methods have a limited capacity to characterize water microbiota. Nevertheless, for almost a century the latter have been optimized to detect and quantify relevant bacteria. A pertinent question is if culture-independent diversity surveys give merely an extended perspective of the bacterial diversity or if, even with a higher coverage, focus on a different set of organisms. We compared the diversity and phylogeny of bacteria in a freshwater sample recovered by currently used culture-dependent and culture-independent methods (DGGE and 454 pyrosequencing). The culture-dependent diversity survey presented lower coverage than the other meth- ods. However, it allowed bacterial identifications to the species level, in contrast with the other procedures that rarely produced identifications below the order. Although the predominant bacterial phyla detected by both approaches were the same (Proteobacteria, Actinobacteria, Bacteroidetes), sequence similarity analysis showed that, in general, different operational taxonomical units were targeted by each method. The observation that culture-dependent and independent approaches target different organisms has implications for the use of the latter for studies in which taxonomic identification has a predictive value. In comparison to DGGE, 454 pyrosequencing method had a higher capacity to explore the bacterial richness and to detect cultured organisms, being also less laborious. Keywords Culture-dependent Á Culture- independent Á DGGE Á 454 Pyrosequencing Á Freshwater Á Bacterial diversity Introduction Bacterial diversity surveys of natural waters are important approaches to assess the ecology and evolution of bacteria, to support management poli- cies or to sustain risk assessment studies. For almost a century, the microbiological quality of waters was I. Vaz-Moreira Á C. M. Manaia (&) CBQF/Escola Superior de Biotecnologia, Universidade Cato ´lica Portuguesa, R. Dr. Anto ´nio Bernardino de Almeida, 4200-072 Porto, Portugal e-mail: cmmanaia@esb.ucp.pt I. Vaz-Moreira Á O. C. Nunes LEPAE—Departamento de Engenharia Quı ´mica, Faculdade de Engenharia, Universidade do Porto, 4200-465 Porto, Portugal C. Egas Advanced Services Unit, Biocant, 3060-197 Cantanhede, Portugal C. Egas Center for Neurosciences and Cellular Biology, University of Coimbra, 3004-517 Coimbra, Portugal