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Current hypotheses on the distribution of freshwater microchlorophytes lead to predictions of low diversity and wide environmental tolerances. Thus, the same few species should be found worldwide in many different habitats. However, these hypotheses are based on a morphospecies concept, which precludes the possibility of numerous cryptic species among these organisms. In this study, we examined the diversity of coccoid green microalgae and chlamydomonads (Chlorophyta) isolated from sites in Minnesota and North Dakota (USA) using techniques of 18S rDNA sequence analysis. Of 93 distinct 18S rDNA sequences identified from among 273 isolates examined by molecular techniques, all but four are new to science. The spatial distribution of organisms represented by these 18S rDNA sequences was not uniform, because some lakes and ponds yielded distinct 18S rDNA types not found at other sites. In addition, organisms generally considered to be cosmopolitan, such as Chlamydomonas reinhardtii and Chlorella vulgaris, were not found. These results challenge predictions of low species number and wide environmental tolerances among these eukaryotic microorganisms.

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Isolates of the genera Monoraphidium Kom.‐Legn., Ankistrodesmus Corda and Raphidocelis Hindák emend. Marvan et al. were cultured from two areas in Minnesota and North Dakota, USA. These isolates were identified to species level (when possible), using light microscopy and standard monographs and then characterized by 18S rDNA sequence analysis. Phylogenetic analyses indicated that in some cases, 18S rDNA sequences from these isolates were very similar, but not identical to the sequences of other isolates of the same morphospecies from different parts of the world. However, some isolates that were identified as the same species actually belong to different lineages within the Selenastraceae, whereas other isolates with identical or nearly identical 18S rDNA sequences possessed rather different morphologies. Overall, our data suggest that the application of a broad morphospecies concept to the Selenastraceae has resulted in an underestimation of the species diversity of this family and probably erroneous conclusions about the distribution of species.

Strains of the genus Nannochloropsis (Eustigmatophyceae) from the Australian National Algae Culture Collection were evaluated by DNA sequence analysis. Strains were first assessed by sequence analysis of a 900-base-pair region of the plastid rbcL gene (bar-code analysis). This analysis allowed us to identify most of the strains at specific rank. However, the strains CS-416 and CS-759 produced a unique rbcL DNA sequence and were sister to N. oceanica in the phylogenetic analysis. Complete rbcL sequences and nuclear 18S ribosomal (r)DNA sequences were produced for these two strains. The results of phylogenetic analyses of an alignment of the concatenated 18S rDNA and rbcL sequences for Nannochloropsis taxa revealed that strains CS-416 and CS-759 represent a new species, N. australis. In addition, the analysis provided strong support for erecting a new genus, Microchloropsis, comprising the species M. salina and M. gaditana.

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A simple method for the purification of PCRquality DNA from microalgae is presented. This method uses the detergent dodecyltrimethylammonium bromide coupled with cell breakage by agitation in the presence of glass beads and chloroform. A final purification step involves a commercial cartridge system. The procedure requires only about 1-2 mL of algal culture and can be completed in about 20 min. DNA suitable for PCR has been obtained from several algal lineages using this method, including numerous green algae and stramenopiles.