The relationship of mixotrophic and autotrophic species to morphologically similar chemoorganotrophic bacteria (e.g. species, Eikelboom type 021N bacteria) has been a matter of debate for some years. These bacteria have alternatively been grouped together on the basis of shared morphological features or separated on the basis of their nutrition. Many of these bacteria are difficult to maintain in axenic culture and, until recently, few isolates were available to allow comprehensive phenotypic and genotypic characterization. Several isolates of spp. and Eikelboom type 021N strains were characterized by comparative 16S rRNA sequence analysis. This revealed that the spp. and Eikelboom type 021N isolates formed a monophyletic group. Furthermore, isolates of Eikelboom type 021N bacteria isolated independently from different continents were phylogenetically closely related. The 16S rRNA sequence-based phylogeny was congruent with the morphological similarities between and Eikelboom type 021N. However, one isolate examined in this study (Ben47) shared many morphological features with the spp. and Eikelboom type 021N isolates, but was not closely related to them phylogenetically. Consequently, morphology alone cannot be used to assign bacteria to the 021N group. Comparative 16S rRNA sequence analysis supports monophyly of the 021N group, and phenotypic differences between the spp. and Eikelboom type 021N bacteria are currently poorly defined. For example, both groups include heterotrophic organisms that deposit intracellular elemental sulfur. It is therefore proposed that the Eikelboom type 021N bacteria should be accommodated within the genus as a new species, sp. nov., and three further new species are described: sp. nov., sp. nov. and sp. nov.


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