High sequence diversity of Alteromonas macleodii-related cloned and cellular 16S rDNAs from a Mediterranean seawater mesocosm experiment

Small subunit rDNA clone libraries were generated from amplified DNA of bacterioplankton taken at different time points from a mesocosm containing eutrophied Mediterranean seawater and made eutrophic by the addition of N and P. Analysis of 96 partial sequences indicated that 22% of the clones formed four clusters which showed the highest sequence sirnilarity with the 16S rDNA sequence of Alteromonas macleodii DSM 6062(T). A fifth cluster, comprising 31% of the clone sequences is moderately related to the A. macleodiisequence. Similarity between the almost complete sequences of two representatives of clone clusters 1, 2 and 3 and A. macleodii ranged between 97.7 and 98.1%. Four oligonucleotide probes, representing four clone clusters, were developed on the basis of partial clone sequences. Dot blot hybridization with PCR-amplified 16S rDNA from 739 clones revealed that 24% of clones belong to one of these clusters. Dot blot hybridization between the four probes and PCR-amplified 16S rDNA from 128 strains isolated from the mesocosm identified 21% of the isolates possessing the probe target region. While probes GP-1 and GP-4 unambiguously identified 0.8 and 4.0% of the strains, respectively, probes GP-2 and GP-3 showed cross hybridization with 16% of the strains. Analysis of the probe target region of the 16S rDNA of one of the isolates indeed demonstrated the presence of double peaks in the relevant region of the sequence which is indicative of microheterogeneity at the rrn operon level. Although some of the diversity can be attributed to intra-strain variation, the data indicate that the phylogenetic diversity of A. macleodii is higher than represented by the type strain of this species. (C) 1999 Federation of European Microbiological Societies. Published by Elsevier Science B.V. All rights reserved.