DNA hybridization to compare species compositions of natural bacterioplankton assemblages

Little is known about the species composition and variability of natural bacterial communities, mostly because conventional identification requires pure cultures, but less than 1% of active natural bacteria are cultivable. This problem was circumvented by comparing species compositions via hybridization of total DNA of natural bacterioplankton communities for the estimation of the fraction of DNA in common between two samples (similarity). DNA probes that were labeled with 35S by nick translation were hybridized to filter-bound DNA in a reciprocal fashion; similarities (in percent) were calculated by normalizing the values to self-hybridizations. In tests with DNA mixtures of pure cultures, the experimentally observed similarities agreed with expectations. However, reciprocal similarities (probe and target reversed) were often asymmetric, unlike those of DNA from single strains. This was due to the relative complexity and G + C content of DNA, which provided a means to interpret the asymmetry that was occasionally observed in natural samples. Natural bacteria were collected by filtration from Long Island Sound (LIS), N.Y., the Caribbean and Sargasso seas, and a coral reef lagoon near Bermuda. The samples showed similarities of less than 10 to 95%. The LIS and Sargasso and Caribbean sea samples were 20 to 50% similar to each other. The coral reef sample was less than 10% similar to the others, indicating its unique composition. Seasonality was also observed; an LIS sample obtained in the autumn was 40% similar to two LIS samples obtained in the summer; these latter two samples were 95% similar. We concluded that total DNA hybridization is a rapid, simple, and unbiased method for investigating the variation of bacterioplankton species composition over time and space, avoiding the need of culturing.