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Twenty-two isolates ofAnabaena azollae derived from sevenAzolla species from various geographic and ecological sources were characterized by DNA-DNA hybridization. Cloned DNA fragments derived from the genomic sequences of three differentA. azollae isolates were used to detect restriction fragment length polymorphism among all symbiotic anabaenas. DNA clones were radiolabeled and hybridized against southern blot transfers of genomic DNAs of different isolates ofA. azollae digested with restriction endonucleases. Eight DNA probes were selected to identify theAnabaena strains tested. Two were strain specific and hybridized only toA. azollae strains isolated fromAzolla microphylla orAzolla caroliniana. One DNA probe was section specific (hybridized only to anabaenas isolated fromAzolla ferns representing the sectionEuazolla ), and five other probes gave finer discrimination among anabaenas representing various ecotypes ofAzolla species. These cloned genomic DNA probes identified 11 different genotypes ofA. azollae isolates. These included three endosymbiotic genotypes withinAzolla filiculoides species and two genotypes within bothA. caroliniana andAzolla pinnata endosymbionts. Although we were not able to discriminate among anabaenas extracted from different ecotypes ofAzolla nilotica, Azolla mexicina, Azolla rubra andAzolla microphylla species, each of the endosymbionts was easily identified as a unique genotype. When total DNA isolated from free-livingAnabaena sp. strain PCC7120 was screened, none of the genomic DNA probes gave detectable positive hybridization. Total DNA ofNostoc cycas PCC7422 hybridized with six of eight genomic DNA fragments. These data imply that the dominant symbiotic organism in association withAzolla spp. is more closely related toNostoc spp. than to free-livingAnabaena spp.

applied and environmental microbiology

DNA Probes Show Genetic Variation in Cyanobacterial Symbionts of the Azolla Fern and a Closer Relationship to Free-Living Nostoc Strains than to Free-Living Anabaena Strains