Nmol C2H4 Research Articles

Xylophagous termites: A potential sink for atmospheric nitrous oxide

To provide a better understanding of soil–atmosphere gas exchange processes, this study describes the atmospheric nitrous oxide (N2O) uptake by xylophagous termites and the biological process involved. The N2O consumption rates of three xylophagous termite species (Hodotermes mossambicus, Nasutitermes voeltzkowi and Hodotermopsis sjoestedti) were determined in incubation vials with ambient, artificially enhanced N2O concentrations in the headspace. Live individuals of the three termite species significantly decreased N2O concentrations (88%) in the headspace of the vials after 24 h incubation in the dark. The acetylene reduction assay method applied to N. voeltzkowi, a xylophagous termite species, showed a decrease in N2O uptake in acetylene-treated individuals, indicating the potential involvement of termite gut denitrifying microbes. The N2 formed is potentially subjected to assimilation via nitrogenase reductase into termite biomass through biological fixation as demonstrated by the reduction of acetylene to ethylene at an average rate of 18.21 ± 1.34 nmol C2H4 g−1 dw d−1. Further studies should focus on measurements of N2O-reductase (nosZ) gene activity in termite guts to gain a better understanding of the N2O reduction process in xylophagous termite species.

Physiological and molecular characterization of locally adapted Rhizobium strains of lentil (Lens culinaris Medik.) having restricted phage sensitivity

Indigenous rhizobia were isolated from root nodules of lentil plants collected from various agro-climatic regions of India. These isolates together with four standard lentil Rhizobium strains were screened for sensitivity against eight phages. Four strains, USDA 2431, BHULR 104, BHULR 113 and BHULR 115 having restricted sensitivity to lytic phages LRP-1, LRP-4, LRP-13 and LRP-15 respectively, were characterized for both physiological and molecular characters. All strains had a generation time of between 3.8 and 5.6 h in yeast extract–mannitol (YM) broth, and colonies on YM agar plates showed an acidic reaction. Compared to other strains, strain USDA 2431 grew poorly when sucrose was the sole carbon source and showed maximum growth in arabinose-containing medium. The intrinsic antibiotic resistance level in all strains was tested against seven antibiotics and found to be high with ampicillin and kanamycin (50–60 μg ml-1) but very low with neomycin (0.03 μg ml-1). With the exception of strain BHULR 113, all strains expressed ex planta nitrogenase activity, with strain USDA 2431 showing the maximum activity (26.8 nmol C2H4 h-1 mg-1 protein) after 6 h of incubation. Genomic and phylogenetic relationships among the strains were examined by randomly amplified polymorphic DNA and 16S rRNA sequence analysis. Genetic distance varied from 0.09 to 0.23 among the strains, and the primer OPL-11 was found to be suitable for the discrimination of these strains. The 16S rRNA sequence analysis revealed 99–100% similarity with Rhizobium leguminosarum bv. viciae. These results clearly indicate that phage sensitivity is a useful marker for discriminating locally soil-adapted rhizobial strains forming effective nodules in lentil.

Cyanobacterial blooms in Lake Atitlan, Guatemala

Lake Atitlan, one of the most important lakes not only in Central America but in the whole world, is facing serious problems with increasing water pollution. Over the last several decades, the uncontrolled nutrient input into the lake has lead to high P levels and low N:P ratios, initiating cyanobacterial blooms. The first bloom occurred in December of 2008, followed by more extensive bloom in October 2009. The blooms are formed by cyanobacteria from the rare planktic Lyngbya hieronymusii/birgei/robusta complex. Based on the species morphology, the Atitlan population corresponds to L. robusta and this is the first case of reported bloom of this species worldwide. Remote sensing images documented that at the maximum bloom development, 40% of the 137 km2 of the lake area were covered by dense patches of Lyngbya, with the chlorophyll a concentration reaching over 100 μg L−1. The only toxins detected in the 2009 bloom were trace levels of cylindrospermopsin and saxitoxin with 12 and 58 ng g−1, respectively. The nitrogen fixation followed a pattern expected in non-heterocytous cyanobacteria, i.e., the nitrogenase activity was minimal during the day, while during the night the activity reached 2.2 nmol C2H4 μg Ch a−1 h−1. Delta 15N of −0.86‰ was well in the range given for nitrogen fixing organisms. The cell C, N and P content was 36.7%, 5.9% and 0.9%, respectively, resulting in the molar ratio of 105:14.4:1. A well designed and executed lake monitoring program, strict control of nutrient input into the lake, and public education are the necessary prerequisites for potential prevention of even more severe blooms than the one from 2009.

In Situ Nitrogen Fixation by Cyanobacteria at the Andragulla Cave, Spain

2 Abstract: Andragulla Cave is 2 m high, 6 m wide, and 2 m long (deep). Its lack of depth means that it is not isolated from external influences, and the microclimate is very similar to that experienced externally. The common stress factors on the growth of cyanobacterial communities on walls inside of Andragulla Cave include excess light, dryness, lack of nutrients, and cold temperatures. Nitrogen fixation, photon flux, relative humidity, and temperature in Andragulla Cave were measured hourly over 24 hours in winter. Nitrogen fixation by the reddish-brown mat formed by both cyanobacteria Scytonema mirabile and Gloeocapsa sanguinea in cave-like environments was measured in situ for the first time by acetylene reduction. The mat-specific rates (1.6 to 7.5 nmol C2H4 m 22 s 21 ) were lower compared to published values from laboratory measurements of rehydrated samples from different environments. Daytime fixation was slightly higher than nighttime fixation, where nitrogen fixation by Gloeocapsa sanguinea played an important role. The most influential parameters for environmental nitrogen fixation in Andragulla Cave were photon flux for daytime fixation, temperature for nighttime fixation, and relative humidity for both. Nitrogen fixation by cyanobacteria may be significant in these N-poor ecosystems.

Effects of cyanobacteria strains selected for their bioconditioning and biofertilization potential on maize dry matter and soil nitrogen status in a South African soil

Some cyanobacteria strains have biofertilization and/or bioconditioning effects in soils as a result of their ability to fix dinitrogen or produce exocellular polysaccharides. The objective of the present study was to screen indigenous cyanobacteria strains with the potential to improve the N fertility and structural stability of degraded soils, and evaluate their ameliorative effectiveness in semiarid soils of the Eastern Cape, South Africa. Soils from Guquka, Hertzog and Qunu villages, and Fort Cox College were used in the screening study. The results showed that only three cyanobacteria strains (3g, 3v and 7e) out of 97 isolated strains were heterocystous, with appreciable nitrogenase activity and the ability to produce exocellular polysaccharides. Nostoc strains 3g and 3v had a greater ability to produce exocellular polysaccharides, but low potential to fix dinitrogen (4.7 and 1.3 nmol C2H4 μg−1 chl h−1, respectively). Strain 7e had the greatest ability to fix dinitrogen (16.1 nmol C2H4 μg−1 chl h−1), but produced fewer exocellular polysaccharides. The ability of strains 3g and 7e to influence maize dry matter (DM) and soil C and N contents was tested in a nitrogen-poor soil with Nostoc strain 9v as a reference strain. Potted soils with and without growing maize plants were inoculated with the different cyanobacteria strains in a glasshouse at a rate of 6 g m−2 soon after maize emergence. Harvesting and soil sampling were done 6 weeks after inoculation. Inoculation with strains 3g and 7e increased maize DM and N uptake significantly, on par with the reference strain. These increases were consistent with increases in nitrate-N observed at harvest time in inoculated cropped and non-cropped soils. Strain 7e resulted in greater increases in soil nitrate-N, tissue N and uptake than strain 3g, perhaps because of its greater ability to fix dinitrogen. Cropping with maize reduced soil total C and N, possibly owing to its negative effects on cyanobacteria establishment. These results suggest that indigenous cyanobacteria strains screened for greater N2-fixing ability have the potential to improve the productivity of N-poor soils in semiarid regions in South Africa.

Burrowing deeper into benthic nitrogen cycling: the impact of bioturbation on nitrogen fixation coupled to sulfate reduction

MEPS Marine Ecology Progress Series Contact the journal Facebook Twitter RSS Mailing List Subscribe to our mailing list via Mailchimp HomeLatest VolumeAbout the JournalEditorsTheme Sections MEPS 409:1-15 (2010) - DOI: https://doi.org/10.3354/meps08639 FEATURE ARTICLE Burrowing deeper into benthic nitrogen cycling: the impact of bioturbation on nitrogen fixation coupled to sulfate reduction Victoria J. Bertics1,2,*, Jill A. Sohm1,*, Tina Treude1,2, Cheryl-Emiliane T. Chow1, Douglas G. Capone1, Jed A. Fuhrman1, Wiebke Ziebis1,** 1Dept. of Biological Sciences and Wrigley Institute for Environmental Studies, University of Southern California, 3616 Trousdale Parkway, Allan Hancock Foundation Building, Los Angeles, California 90089-0371, USA 2Leibniz Institute of Marine Sciences, Wischhofstr. 1–3, GEB. 4, Kiel 24148, Germany *These authors contributed equally to this manuscript**Corresponding author. Email: wziebis@usc.edu ABSTRACT: Biological dinitrogen (N2) fixation is the primary input of fixed nitrogen (N) into the marine biosphere, making it an essential process contributing to the biological functions of all organisms. Because biologically available N often limits marine productivity, microbial processes leading to its loss and gain (e.g. denitrification and N2 fixation, respectively) play an important role in global biogeochemical cycles. Bioturbation is known to influence benthic N cycling, most often reported as enhancement of denitrification and a subsequent loss of N2 from the system. N2 fixation has rarely been addressed in bioturbation studies. Instead, sedimentary N2 fixation typically has been considered important in relatively rare, localized habitats such as rhizosphere and phototrophic microbial mat environments. However, the potential for N2 fixation in marine sediments may be more widespread. We show here that nitrogenase activity can be very high (up to 5 nmol C2H4 cm–3 h–1) in coastal sediments bioturbated by the ghost shrimp Neotrypaea californiensis and at depths below 5 cm. Integrated subsurface N2-fixation rates were greater than those previously found for un-vegetated estuarine sediments and were comparable to rates from photosynthetic microbial mats and rhizospheres. Inhibition experiments and genetic analysis showed that this activity was mainly linked to sulfate reduction. Sulfate-reducing bacteria (SRB) are widespread and abundant in marine sediments, with many possessing the genetic capacity to fix N2. Our results show that N2 fixation by SRB in bioturbated sediments may be an important process leading to new N input into marine sediments. Given the ubiquity of bioturbation and of SRB in marine sediments, this overlooked benthic N2 fixation may play an important role in marine N and carbon (C) cycles. KEY WORDS: Bioturbation · Crustaceans · Marine sediment · Microniche · Nitrogen fixation · Sulfate reduction Full text in pdf format Information about this Feature Article NextCite this article as: Bertics VJ, Sohm JA, Treude T, Chow CET, Capone DG, Fuhrman JA, Ziebis W (2010) Burrowing deeper into benthic nitrogen cycling: the impact of bioturbation on nitrogen fixation coupled to sulfate reduction. Mar Ecol Prog Ser 409:1-15. https://doi.org/10.3354/meps08639 Export citation RSS - Facebook - Tweet - linkedIn Cited by Published in MEPS Vol. 409. Online publication date: June 23, 2010 Print ISSN: 0171-8630; Online ISSN: 1616-1599 Copyright © 2010 Inter-Research.

Biological Nitrogen Fixation in Two Tropical Forests: Ecosystem-Level Patterns and Effects of Nitrogen Fertilization

Humid tropical forests are often characterized by large nitrogen (N) pools, and are known to have large potential N losses. Although rarely measured, tropical forests likely maintain considerable biological N fixation (BNF) to balance N losses. We estimated inputs of N via BNF by free-living microbes for two tropical forests in Puerto Rico, and assessed the response to increased N availability using an on-going N fertilization experiment. Nitrogenase activity was measured across forest strata, including the soil, forest floor, mosses, canopy epiphylls, and lichens using acetylene (C2H2) reduction assays. BNF varied significantly among ecosystem compartments in both forests. Mosses had the highest rates of nitrogenase activity per gram of sample, with 11 ± 6 nmol C2H2 reduced/g dry weight/h (mean ± SE) in a lower elevation forest, and 6 ± 1 nmol C2H2/g/h in an upper elevation forest. We calculated potential N fluxes via BNF to each forest compartment using surveys of standing stocks. Soils and mosses provided the largest potential inputs of N via BNF to these ecosystems. Summing all components, total background BNF inputs were 120 ± 29 μg N/m2/h in the lower elevation forest, and 95 ± 15 μg N/m2/h in the upper elevation forest, with added N significantly suppressing BNF in soils and forest floor. Moisture content was significantly positively correlated with BNF rates for soils and the forest floor. We conclude that BNF is an active biological process across forest strata for these tropical forests, and is likely to be sensitive to increases in N deposition in tropical regions.

Nitrogen fixation by epiphytic and epibenthic diazotrophs associated with seagrass meadows along the Tanzanian coast, Western Indian Ocean

Tropical seagrass ecosystems are highly productive and important for sustaining marine life and associated coastal societies. In this study, the diversity and role of nitrogen-fixing cyanobacteria associated with five common seagrass genera in coastal regions of the western Indian Ocean (WIO; Tanzania) were examined, as well as the impact of anthropogenic activities. Cyanobacteria were characterized morphologically and genetically (16S rRNA and nifH gene phylogeny), as were diel variations in nifH gene expression, NifH protein levels and nitrogenase activity. The results revealed that WIO seagrass beds supported rich cyanobacterial diversity and that these represented approx. 83% of total clones obtained (DNA and RNA nifH clone libraries). Non-heterocystous genera, such as Oscillatoria, Lyngbya, Leptolyngbya, Phormidium and Microcoleus dominated, while heterocystous morphotypes such as Calothrix were less frequent and unicellular morphotypes (e.g. Gloeocapsa, Chroococcus and Chroococcidiopsis) were few. Additionally, the phylogenetic analysis revealed several novel uncharacterized cyanobacterial clades. Cyanobacterial composition and nitrogenase activity varied over seasons and between the seagrass species. Day time nitrogenase activity originated primarily from heterocystous phylotypes, while non-heterocystous filamentous phylotypes fixed nitrogen at night. The highest activity in the diel cycle was 358 ± 232 nmol C2H4 g-1 h-1at 09.00 associated with epiphytes of the seagrass Cymodocea. Nitrogenase activity was consistently lower in anthropogenically disturbed (eutrophication) seagrass sites. Such data suggest that diazotrophic cyanobacteria may be a significant source of ‘new’ nitrogen in the often oligotrophic coastal regions of tropical oceans. It is also proposed that the rapid shifts in the cyanobacterial population and function found may also be used as early disturbance indicator in coastal management practices.

Cross‐basin comparison of phosphorus stress and nitrogen fixation in Trichodesmium

We investigated the phosphorus (P) status and N2 fixation rates of Trichodesmium populations from the North Pacific, western South Pacific, and western North Atlantic. Colonies of Trichodesmium were collected and analyzed for endogenous alkaline phosphatase (AP) activity using enzyme‐labeled fluorescence (ELF) and for nitrogenase activity using acetylene reduction. AP hydrolyzes dissolved inorganic phosphate (DIP) from dissolved organic phosphorus and is active in Trichodesmium colonies experiencing P stress. Across multiple stations in the subtropical North and South Pacific, there was low to moderate ELF labeling in Trichodesmium, although labeling was present in other taxa. In contrast, Trichodesmium ELF labeling in the North Atlantic ranged from low to high. Low ELF labeling corresponded with high DIP concentrations while high ELF labeling occurred only at North Atlantic stations with DIP concentrations #40 nmol L21, indicating that Trichodesmium was not experiencing dramatic P stress in the Pacific Ocean while P stress was evident in the western North Atlantic. However, nitrogenase activity was significantly higher in the P‐stressed western North Atlantic than in the Pacific Ocean (0.40‐1.30 compared to 0.01‐0.46 nmol C2H4 h−1 colony−1). These data underscore the differential basinlevel importance of P availability to Trichodesmium and suggest that factors other than P are constraining their N2 fixation rates in the Pacific.

Lichenized unicellular cyanobacteria fix nitrogen in the light

Diurnal variation in N2-fixation (acetylene reduction) rate was measured in Thyrea girardii (Durieu & Mont.) Bagl. & Carestia and Thyrea confusa Henssen, lichens containing a unicellular cyanobacterial photobiont. In field assays, mean acetylene-reduction rates in the light were 25.8 ± 8.9 (n = 11) and 21.0 ± 5.6 nmol C2H4·g–1·h–1(n = 13) for T. girardii and T. confusa, respectively, and the respective mean rates in the dark were 8.2 ± 1.8 (n = 26) and 13.5 ± 5.4 (n = 8) nmol C2H4·g–1·h–1. In laboratory assays under relatively isothermal conditions (ca. 19–22 °C), the maximum acetylene reduction rate (52.0 ± 6.0 nmol C2H4·g–1·h–1) was recorded in the light and the minimum rate (20.2 ± 6.0 nmol C2H4·g–1·h–1) in the dark. This diurnal pattern is contrary to expectations for unicellular cyanobacteria. We suggest carbon flow to the fungal symbiont reduces the potential for nitrogenase activity in the dark.

Nitrogen fixation associated with Suillus tomentosus tuberculate ectomycorrhizae on Pinus contorta var. latifolia.

Tuberculate ectomycorrhizae are a unique form of ectomycorrhiza where densely packed clusters of mycorrhizal root tips are enveloped by a thick hyphal sheath to form a tubercle. The functional significance of such a unique structure has not previously been established. The purpose of the present study was to investigate and measure the potential nitrogenase activity associated with Suillus tomentosus/Pinus contorta tuberculate ectomycorrhizae in two stand ages, young and old, and across a range of nitrogen-poor soil conditions. Short roots were compared with other mycorrhizae and non-mycorrhizal secondary roots using tuberculate ectomycorrhizae. Assessment of nitrogenase activity was determined and quantitative measurements were taken on tuberculate ectomycorrhizae in situ in a variety of different circumstances, by using an adaptation of the acetylene reduction assay. Significant nitrogenase activity was measured associated with S. tomentosus/P. contorta tuberculate ectomycorrhizae whereas no nitrogenase activity was measured with non-tuberculate mycorrhizae or secondary roots without mycorrhizae. Average nitrogenase activity ranged from undetectable to 5696.7 nmol C2H4 g(-1) tubercle 24 h(-1). Maximum nitrogenase activity was 25,098.8 nmol C2H4 g(-1) tubercle 24 h(-1). Nitrogenase activity was significantly higher in young stands than in old stands of P. contorta. Season or some covariate also seemed to affect nitrogenase activity and there was suggestion of a site effect. Suillus tomentosus/P. contorta tuberculate ectomycorrhizae are sites of significant nitrogenase activity. The nitrogenase activity measured could be an important contribution to the nitrogen budget of P. contorta stands. Season and stand age affect levels of nitrogenase activity.

Enhancement of Nitrogen-fixing Activity of Enterobacteriaceae Strains Isolated from Sago Palm (Metroxylon sagu) by Microbial Interaction with Non-nitrogen Fixers

This study was done to determine microbial interactions between nitrogen-fixing bacteria (NFB) and different indigenous bacteria, both of which were isolated from the same samples of sago palm. Co-culture of NFB and indigenous bacteria, isolated using nutrient agar medium, into a nitrogen-free Rennie medium showed significantly higher nitrogen-fixing activity (NFA) than single inoculations of NFB did in almost all combinations. A reduced oxygen status also enhanced the NFA of NFB. All NFB preferred simple sugars as their substrates for NFA and showed extremely low levels of NFA in starch, hemicellulose, and pectin-containing media. NFA was markedly stimulated in the consortium of starch-degrading Bacillus sp. strain B1 and NFB, ranging from 0-1.5 to 150-270 nmol C2H4 culture-1 h-1. The consortium of hemicellulose-degrading Agrobacterium sp. strain HMC1 or Flexibacter sp. strain HMC2 and NFB also showed enhanced NFA, ranging from 0-0.1 to 16-38 nmol. In contrast, no enhanced NFA was detected in the consortium of pectin-degrading Burkholderia sp. strain BT1 or Paenibacillus sp. strain P1 and NFB. These results may indicate that beneficial microbial interactions occur in sago palm to enhance nitrogen-fixing activity through collaborative utilizations of starch, hemicellulose and their degradation products.

Seasonal changes in periphyton nitrogen fixation in a protected tropical wetland

Cyanobacteria are important for global nitrogen cycle and often form complex associations referred to as cyanobacterial mats or periphyton that are common in tropical, limestone-based wetlands. The objective of this study was to monitor the nitrogen fixation rate using the acetylene reduction assay of these cyanobacterial mats in a tropical, unfertilized, and protected wetland. To account for temporal and spatial variation of nitrogenase activity, we were interested in seasons in a hydrological cycle (dry, rains, and end of rains), sites with different vascular vegetation, and rates of nitrogenase activity in a 24-h cycle. The annual average of nitrogenase activity was 22 nmol C2H4 cm−2 h−1, with a range of <6 to 35 nmol C2H4 cm−2 h−1, and the annual nitrogen fixation rate of our study site (9.0 g N m−2 year−1) is higher than similar estimates from other freshwater wetlands. There was a clear temporal pattern in nitrogenase activity with a maximum rate occurring during the rainy season (August) and a maximum nitrogenase activity occurring between 0600 and 1200 hours. We found spatial differences in nitrogenase activity among the four sites that could be attributed to variations in species composition within the periphyton.

Diversity and activity of epiphytic nitrogen-fixers on standing dead stems of the salt marsh grass Spartina alterniflora

AME Aquatic Microbial Ecology Contact the journal Facebook Twitter RSS Mailing List Subscribe to our mailing list via Mailchimp HomeLatest VolumeAbout the JournalEditorsSpecials AME 39:271-279 (2005) - doi:10.3354/ame039271 Diversity and activity of epiphytic nitrogen-fixers on standing dead stems of the salt marsh grass Spartina alterniflora Pia H. Moisander1,2,*, Michael F. Piehler1, Hans W. Paerl1 1University of North Carolina at Chapel Hill, Institute of Marine Sciences, 3431 Arendell Street, Morehead City, North Carolina 28557, USA2Present address: Ocean Sciences Department, University of California Santa Cruz, 1156 High Street, Santa Cruz, California 95064, USA *Email: pia_moisander@alumni.unc.edu ABSTRACT: Microbial communities growing on the surfaces of standing dead stems of salt marsh grasses contribute to fixation of atmospheric nitrogen (N2) in these N-limited intertidal systems. Salt marshes in North Carolina, USA, have been shownto exhibit N2 fixation throughout the growth season, but the composition of these diazotrophic communities is poorly known. This study investigated the diversity of the epiphytic N2-fixing microbial community on the surfaces of dead stems (shoots) of the cord grass Spartina alterniflora in a salt marsh in North Carolina. Sequencing of the nifH gene and microscopy showed that a diverse diazotrophic community consisting of at least 8 diazotrophic cyanobacterial taxa, as well as an assemblage of α-, γ-, and δ-proteobacteria, was present. Half of the recovered unique nifH sequences fell into the cluster with anaerobes. N2-fixation rates under natural irradiance ranged from 0.001to 2.58 nmol C2H4 µg chl a–1 h–1 (0.07 to 206 nmol C2H4 cm–2 h–1) and decreased from spring tofall. Nitrogen-fixation rates had a positive relationship with chlorophyll a, suggesting phototrophs contributed to N2 fixation. The dominant diazotrophs in this study differed from epiphytic communities described elsewhere in the southeast United States, suggesting regional differences are present in the diversity of these communities. KEY WORDS: NifH · Nitrogen fixation · Epiphytes · Salt marsh · Spartina alterniflora · Cyanobacteria · Anaerobes · Diversity Full text in pdf format PreviousNextExport citation RSS - Facebook - Tweet - linkedIn Cited by Published in AME Vol. 39, No. 3. Online publication date: June 20, 2005 Print ISSN: 0948-3055; Online ISSN: 1616-1564 Copyright © 2005 Inter-Research.

15N as an indicator of N2-fixation by cyanobacterial mats in tropical marshes

Cyanobacterial mats (CBM) are important components of wetland ecosystems in limestone-based regions of the Caribbean. During two sampling periods (July 1999 and January 2000) we measured N2-fixation in samples from 23 different marshes simultaneously with measurements of relevant environmental factors. Samples were evaluated for abundance of five groups of cyanobacteria: (1) Leptolyngbya, (2) Oscillatoria, (3) Chroococcales, (4) Nostoc-& Stigonematales, and (5) dead sheaths. Differences in nitrogen fixation, expressed as nitrogenase activity in nmol C2H4 cm−2 h−1, were best explained by the proportion of heterocyst-forming cyanobacteria. The samples were analyzed for the natural abundance of δ15N. δ15N values ranged from −1.99 to 11.44 ‰ and were strongly negatively correlated with N2-fixation. With all data included, δ15N was also strongly correlated with nitrates in water. With the samples from Little Belize (high nitrate content marshes) excluded, the effect of nitrate became insignificant. N2-fixation predicted from δ15N measured on an independent data set from September 2000 was moderately accurate (r2 = 0.68, 0.52 and 0.54 for predictions based on July 1999, January 2000 and combined data sets, respectively). When individual sample sets were divided into two groups with δ15N 2, the two groups were always highly significantly different in terms of their N2-fixation. The presented evidence suggests that δ15N can be used as a reliable indicator of N2-fixation by CBM.

Periphyton chemistry and nitrogenase activity in a northern Everglades ecosystem

Cyanobacterial peri­phyton communities are a dominant feature of oligotrophic Everglades marshes, however, little is known regarding the biogeochemical aspects of this ecosystem component. This study was undertaken to investigate the potential for N2 fixation in the peri­phyton communities of a hydrologically-controlled portion of the northern Everglades marsh (Water Conservation Area 2A, WCA-2A). The objectives of this research were to characterize the temporal patterns of nutrient composition and N2 fixation of the natural WCA-2A peri­phyton communities and to compare fixation rates of peri­phyton with those of other ecosystem components in both natural and nutrient-impacted WCA-2A areas. In general, N2 fixation (measured by the acetylene reduction (AR) method) of natural WCA-2A peri­phyton was enhanced under light conditions showing a nitrogenase pattern characteristic of autotrophic cyanobacteria. Winter (November–March) rates of AR expressed per gram organic carbon (gOC) ranged from 147–240 nmol C2H2 g OC−1 h−1, while summer rates were elevated with an observed peak of 1148 nmol C2H2 g OC−1 h−1 in July 1998. This translates into an estimated yearly contribution of approximately 10 g N m−2 to an unimpacted WCA-2A slough ecosystem. Nitrogenase activity did not correlate seasonally with nutrients (Ca, Mg, Fe, N, P, Mn), but closely followed measured N stable isotopic ratios (δ15N) in floating peri­phyton. In oligotrophic marsh areas, AR (on a weight basis) decreased in the order floating peri­phyton > benthic peri­phyton floc > soil > water > detrital plant biomass, while highest AR rates were observed for detrital biomass in areas impacted by agricultural discharges.

Nitrogen fixation in coarse woody debris of Thuja plicata and Tsuga heterophylla forests on northern Vancouver Island

Asymbiotic nitrogenase activity in coarse woody debris was measured using the acetylene reduction assay under ambient conditions in three different stand ages (5, 53, and 88 years old) of an unmanaged second-growth Tsuga heterophylla (Raf.) Sarg. – Abies amabilis (Dougl. ex Loud.) Dougl. ex J. Forbes forest type and a Thuja plicata Donn. ex D. Don – Tsuga heterophylla old-growth forest on northern Vancouver Island, British Columbia, Canada. Four different decay classes of coarse woody debris, different species in the early decay stages, and sapwood and heart wood were sampled separately. Mean nitrogenase activity ranged between 1.3 and 19.5 nmol C2H4·d–1·(g dry mass)–1, with an overall mean of 5.7. High variability of the activity rates between logs and within logs was observed in all four stands. Mean activity rates were, in most cases, significantly different between decay classes, with generally increasing nitrogenase activity with the progress of decay. Moisture content of the samples was a good predictor of nitrogenase activity and could explain differences between decay classes. Only minor differences in nitrogenase activity were found between the different stands. Estimates of nitrogen fixation ranged from 1.0 to 2.1 kg N·ha–1·year–1, the magnitude of these values depending more on the mass of coarse woody debris substrate available for asymbiotic nitrogen-fixing bacteria (103–158 t·ha–1 in this study) than on differences in nitrogenase activity rates. The measured nitrogenase activity and the resultant estimates of nitrogen fixation are among the highest values reported in the literature.

Cultivable heterotrophic N2-fixing bacterial diversity in rice fields in the Yangtze River Plain

Heterotrophic N2-fixing bacteria are a potentially important source of N2 fixation in rice fields due to the moist soil conditions. This study was conducted at eight sites along a geographic gradient of the Yangtze River Plain in central China. A nitrogen-free solid malate-sucrose medium was used to isolate heterotrophic N2-fixing bacteria. Numbers of the culturable N2-fixing bacteria expressed as CFU (colony forming units) ranged between 1.41±0.42×106 and 1.24±0.23×108 in the sampled paddy field sites along the plain. Thirty strains with high ARA (acetylene reduction activity) were isolated and purified; ARA of the strains varied from 0.9 to 537.8 nmol C2H4 culture-1 h-1, and amounts of 15N fixed ranged between 0.008 and 0.4866 mg·culture-1·day-1. According to morphological and biochemical characteristics, 14 strains were identified as the genus Bacillus, 2 as Burkholderia, 1 as Agrobacterium, 4 as Pseudomonas, 2 as Derxia, 1 as Alcaligenes, 1 as Aeromonas, 2 as Citrobacter, and 3 strains belonged to the corynebacter-form group.

N2-dependent growth and nitrogenase activity in the metal-metabolizing bacteria, Geobacter and Magnetospirillum species.

Cells of Geobacter metallireducens, Magnetospirillum strain AMB-1, Magnetospirillum magnetotacticum and Magnetospirillum gryphiswaldense showed N2-dependent growth, the first anaerobically with Fe(III) as the electron acceptor, and the latter three species microaerobically in semi-solid oxygen gradient cultures. Cells of the Magnetospirillum species grown with N2 under microaerobic conditions were magnetotactic and therefore produced magnetosomes. Cells of Geobacter metallireducens reduced acetylene to ethylene (11.5+/-5.9 nmol C2H4 produced min(-1) mg(-1) cell protein) while growing with Fe(III) as the electron acceptor in anaerobic growth medium lacking a fixed nitrogen source. Cells of the Magnetospirillum species, grown in a semi-solid oxygen gradient medium, also reduced acetylene at comparable rates. Uncut chromosomal and fragments from endonuclease-digested chromosomal DNA from these species, as well as Geobacter sulphurreducens organisms, hybridized with a nifHDK probe from Rhodospirillum rubrum, indicating the presence of these nitrogenase structural genes in these organisms. The evidence presented here shows that members of the metal-metabolizing genera, Geobacter and Magnetospirillum, fix atmospheric dinitrogen.

A function of cyanobacterial mats in phosphorus-limited tropical wetlands.

Cyanobacterial mats are important components of oligotrophic wetland ecosystems in the limestone-based regions of the Caribbean. Our goals were to: (1) Estimate the biomass and primary production of cyanobacterial mats, quantify the extent of nitrogen fixation and measure the activity of alkaline phosphatase (APA) in representative marshes of northern Belize; (2) Record changes in these variables following nutrient additions. The mat biomass ranged from 200 to 700 g m−2 AFDM, with the epipelon contributing up to 87% of the total. Tissue nitrogen was similar in all marshes (1.1–1.5%), while tissue phosphorus was extremely low (0.0055–0.0129%) and well correlated with the N:P ratio in water. Nitrogen fixation expressed as nitrogenase activity was high in some marshes (17.5 nmol C2H4 cm−2 h−1) and low (< 5 nmol C2H4 cm−2 h−1) in others depending mainly on the proportion of heterocyst-forming cyanobacteria (Nostocales, Stigonematales) in the mat. Alkaline phosphatase activity was positively correlated with the N:P ratio of the mat. Experimental addition of phosphorus resulted in significant increase in primary production and nitrogen fixation while it suppressed the APA activity. The presented data clearly showed that oligotrophic marshes of northern Belize are strongly P limited. Increased input of phosphorus would profoundly change their structure and functions.