Diversity Of Nitrogen-fixing Microorganisms Research Articles

Abundance and diversity of diazotrophs in the surface sediments of Kongsfjorden, an Arctic fjord

Diazotrophy in the Arctic environment is poorly understood compared to tropical and subtropical regions. Hence in this study, we report the abundance and diversity of diazotrophs in Arctic fjord sediments and elucidate the role of environmental factors on the distribution of diazotrophs. The study was conducted during the boreal summer in the Kongsfjorden, an Arctic fjord situated in the western coast of Spitsbergen. The abundance of nifH gene was measured through quantitative real-time PCR and the diversity of diazotrophs was assessed by nifH targeted clone library and next generation sequence analysis. Results revealed that the abundance of nifH gene in the surface sediments ranged from 2.3 × 106 to 3.7 × 107 copies g- 1. The δ-proteobacterial diazotrophs (71% of total sequence) were the dominant class observed in this study. Major genera retrieved from the sequence analysis were Desulfovibrionaceae (25% of total sequence), Desulfuromonadaceae (18% of total sequence) and Desulfobacteriaceae (10% of total sequence); these are important diazotrophic iron and sulfur-reducing bacterial clade in the Kongsfjorden sediments. The abundance of nifH gene showed a significant positive correlation TOC/TN ratio (r2 = 0.96, p ≤ 0.05) and total organic carbon (p ≤ 0.05) content in the fjord sediments. The higher TOC/TN ratio (4.24-14.5) indicated low nitrogen content organic matter in the fjord sediments through glacier runoff, which enhances the abundance and diversity of nitrogen fixing microorganisms.

Variation in diazotrophic community structure in forest soils reflects land use history

Microbial nitrogen fixation is an important source of nitrogen in soils of both old and secondary-growth forests. Considering that many soils, which today support secondary-growth forests were once under cultivation, no studies have examined how this kind of disturbance history affects contemporary populations of nitrogen-fixing microbes in forest soils. In the work presented here, we compare secondary-growth forest sites, which were under cultivation more than 150 years ago, to old-growth forest sites in Cadwell Forest, Massachusetts. For each site, nitrogenase activity was measured and the diversity of the nifH gene pools was examined. Three sites with prior agricultural history exhibited higher nitrogenase activity and were dominated by diazotrophs closely related to the α- and γ-Proteobacteria. In contrast, lower nitrogenase activity and the dominance of the anaerobic Cluster III diazotrophs characterized the three old forest sites. Further analyses of species overlap among all six sites revealed that the diazotrophic composition was closely related to previous management history, with agricultural sites clustering together and separate from old forest sites, independent of the proximity between sites. By specifically targeting one of the main functions of microbial communities in soils, the activity and diversity of nitrogen-fixing microorganisms, this work points to a long-lasting effect of former agricultural activities on secondary-growth forest soils, more than one hundred years after succession.

Analysis of the diversity of diazotrophic bacteria in peat soil by cloning of the nifH gene

The diversity of nitrogen-fixing microorganisms in the soil of an oligotrophic Sphagnum peat bog was studied by molecular cloning of fragments of the nifH gene encoding one of the main components of the nitrogenase complex. The fragments were amplified from the DNA isolated from the peat samples collected at the same site in January (library I) and November (library II), 2005. Analysis of the nifH sequence libraries revealed high diversity of diazotrophic bacteria in peat soil: the first library consisted of 237 clones and 55 unique sequence types, the second one included 171 clones and 52 sequence types. Comparison of the two clone libraries showed that the composition and population structure of the nitrogen-fixing community depended greatly on the sampling time; they shared only 11 phylotypes. The sequences of representatives of the class Alphaproteobacteria prevailed in both libraries (27% and 57% of clones in libraries I and II, respectively). Representatives of the classes Deltaproteobacteria and Chlorobea were minor components of library I (6% and 7% of clones, respectively), whereas they prevailed in library II (18% and 24% of clones, respectively). Members of the class Chloroflexi were present only in library I, while members of the classes Bacilli, Clostridia, and Methanomicrobia were present only in library II. Our studies demonstrated that, for complete evaluation of the diversity of natural nitrogen-fixing communities, nifH libraries should consist of at least 200–300 clones.

Fingerprinting diazotroph communities in the Chesapeake Bay by using a DNA macroarray.

Investigations of the distribution and diversity of nitrogen-fixing microorganisms in natural environments have often relied on PCR amplification and sequence analysis of a portion of one of the key enzymes in nitrogen fixation, dinitrogenase reductase, encoded by nifH. Recent work has suggested that DNA macroarrays provide semiquantitative fingerprints of diversity within mixtures of nifH amplicons (G. F. Steward, B. D. Jenkins, B. B. Ward, and J. P. Zehr, Appl. Environ. Microbiol. 70:1455-1465, 2004). Here we report the application of macroarrays for a study in the Chesapeake Bay. Samples from different locations in the bay yielded distinct fingerprints. Analysis of replicates and samples from different locations by cluster analysis showed that replicates clustered together, whereas different samples formed distinct clusters. There was a correspondence between the hybridization pattern observed and that predicted from the distribution of sequence types in a corresponding clone library. Some discrepancies between the methods were observed which are likely a result of the high nifH sequence diversity in the Chesapeake Bay and the limited number of sequences represented on this version of the array. Analyses of sequences in the clone library indicate that the Chesapeake Bay harbors unique, phylogenetically diverse diazotrophs. The macroarray hybridization patterns suggest that there are spatially variable communities of diazotrophs, which have been confirmed by quantitative PCR methods (S. M. Short, B. D. Jenkins, and J. P. Zehr, Appl. Environ. Microbiol., in press). The results show that DNA macroarrays have great potential for mapping the spatial and temporal variability of functional gene diversity in the environment.

Comparison of diazotroph community structure in Lyngbya sp. and Microcoleus chthonoplastes dominated microbial mats from Guerrero Negro, Baja, Mexico

The nitrogenase activity and phylogenetic diversity of nitrogen fixing microorganisms in several different cyanobacterial mat types from Guerrero Negro, Baja California, Mexico were investigated by acetylene reduction assay, and by amplification and sequencing of the nitrogenase nifH gene. Acetylene reduction assays performed on a Lyngbya sp. and two Microcoleus chthonoplastes dominated microbial mats showed a typical diel pattern of nitrogenase activity in these mats. The highest rates of activity were found at night, with 40 and 37 μmol C 2H 4 m −2 h −1 measured in the Microcoleus mats, and 9 μmol C 2H 4 m −2 h −1 in the Lyngbya mat. Nitrogenase sequences were obtained that clustered with sequences from cyanobacteria, γ-Proteobacteria, and cluster 3 of nifH. In addition, novel and divergent sequences were also recovered. The composition of nifH sequence types recovered differed between the Lyngbya and Microcoleus mats. Interestingly, nifH sequences belonging to filamentous cyanobacteria were absent in most mat samples even though both mats were dominated by filamentous cyanobacteria. nifH sequences clustering with those of unicellular cyanobacteria were found, some of which were virtually identical to the nifH sequence from Halothece sp. MPI96P605, which had previously been isolated from the mat. In manipulation experiments, the Lyngbya and Microcoleus mats were allowed to re-colonize a cleared surface. In these developing mats, nifH sequences not previously observed in the mats were discovered. Our results showed that organisms capable of N 2 fixation were present in N 2 fixing mats, that the composition of the N 2 fixing communities differs between mats, and that filamentous cyanobacterial diazotrophs may not have a large role in the early stages of mat development.