N-164. The Nitrogenase nifH Gene: Current Status of the Census

J. C. Gaby, D. H. Buckley;
Cornell Univ., Ithaca, NY.

The nifH gene is used widely for surveys of nitrogen-fixing capability in the environment. The current number of nifH genes available in Genbank stands at nearly 13 000 sequences. In this study we compiled an aligned database of these sequences with associated information on isolation source in the ARB software package. We used this database to examine the diversity and sampling extent of nifH genes with respect to phylogeny and environment of origin. Only 8.9% of these sequences are from cultured isolates. Of all nifH genes, 14% originate from open water marine environments, 57% terrestrial, 11% microbial mats, 4.1% termite gut, and 3.6% from freshwater. Analysis by country and environment reveals that sampling has neglected sources of potential biodiversity and focused heavily on sites within the developed world and on agricultural studies. Analysis was performed using the software DOTUR to assign OTUs and estimate richness. A richness of 6,482 (5,989, 7,048; lower, upper 95% C. I.) nifH genes is predicted from examining 7,201 overlapping sequences (Chao1 with OTU defined at 5% sequence dissimilarity), but accumulation curves indicate that the diversity of nitrogenase remains under-sampled and so this estimate should be considered a lower bound on global richness. In addition, analysis with respect to environment of origin reveals that nifH diversity remains under-sampled in a number of environments, including marine and soil systems. Current sampling efforts do not reveal a difference in richness between terrestrial and marine environments though the phylogenetic affiliation of diazotrophs in these systems differs greatly. For example, cluster 1A diazotrophs which have recently been revealed to contain relatives of the Geobacteraceae represent 8.0% of nifH sequences in the database, 79% of which were recovered from soil. The database was also used to evaluate current nifH primer sets for specificity and coverage. These results reveal the current status of our understanding of nitrogenase diversity and provide guidance for future research topics.