Ramon Varela

Protistan microbial observatory in the Cariaco Basin, Caribbean. I. Pyrosequencing vs Sanger insights into species richness

Microbial diversity and distribution are topics of intensive research. In two companion papers in this issue, we describe the results of the Cariaco Microbial Observatory (Caribbean Sea, Venezuela). The Basin contains the largest body of marine anoxic water, and presents an opportunity to study protistan communities across biogeochemical gradients. In the first paper, we survey 18S ribosomal RNA (rRNA) gene sequence diversity using both Sanger- and pyrosequencing-based approaches, employing multiple PCR primers, and state-of-the-art statistical analyses to estimate microbial richness missed by the survey. Sampling the Basin at three stations, in two seasons, and at four depths with distinct biogeochemical regimes, we obtained the largest, and arguably the least biased collection of over 6000 nearly full-length protistan rRNA gene sequences from a given oceanographic regime to date, and over 80 000 pyrosequencing tags. These represent all major and many minor protistan taxa, at frequencies globally similar between the two sequence collections. This large data set provided, via the recently developed parametric modeling, the first statistically sound prediction of the total size of protistan richness in a large and varied environment, such as the Cariaco Basin: over 36 000 species, defined as almost full-length 18S rRNA gene sequence clusters sharing over 99% sequence homology. This richness is a small fraction of the grand total of known protists (over 100 000–500 000 species), suggesting a degree of protistan endemism.

Si cycle in the Cariaco Basin, Venezuela: Seasonal variability in silicate availability and the Si:C:N composition of sinking particles

result of rapid utilization. In most years, the upper water column during winter and spring is marked by Si(OH)4 :NO3 and Si* values of less than 1. This indicates that silicate limitation in Cariaco Basin is most severe during upwelling and may restrict diatom production. Conversely, during the summer and fall when upwelling is reduced, Si(OH)4 :NO3 ratios in the upper 50 m of the water column exceed 10, implying that nitrate rather than silicate is acting to limit production during this time of year. On average, sinking particles collected at 150-m depth in the Cariaco Basin have Si:C and Si:N values of 0.17 ± 0.01 and 1.14 ± 0.10, respectively. These ratios increase with depth to 400 m and then remain relatively constant, suggesting minimal selective removal of elements with remineralization in the anoxic portion of the water column. Similar depth-dependent changes in these ratios are seen in surface sediments from the basin. Seasonally, particulate Si:C and Si:N are highest during the early part of the year when upwelling is most intense, while both ratios decrease to their lowest values during summer and fall. The observed seasonal variability in these ratios is due to changes in both nutrient utilization by diatoms and the contribution of diatoms to the total phytoplankton. The high ratios during upwelling suggest enhanced export of Si relative to C and N during this time of year.

Ecosystem responses in the southern Caribbean Sea to global climate change

Over the last few decades, rising greenhouse gas emissions have promoted poleward expansion of the large-scale atmospheric Hadley circulation that dominates the Tropics, thereby affecting behavior of the Intertropical Convergence Zone (ITCZ) and North Atlantic Oscillation (NAO). Expression of these changes in tropical marine ecosystems is poorly understood because of sparse observational datasets. We link contemporary ecological changes in the southern Caribbean Sea to global climate change indices. Monthly observations from the CARIACO Ocean Time-Series between 1996 and 2010 document significant decadal scale trends, including a net sea surface temperature (SST) rise of ∼1.0 ± 0.14 °C (±SE), intensified stratification, reduced delivery of upwelled nutrients to surface waters, and diminished phytoplankton bloom intensities evident as overall declines in chlorophyll a concentrations (ΔChla = −2.8 ± 0.5%⋅y−1) and net primary production (ΔNPP = −1.5 ± 0.3%⋅y−1). Additionally, phytoplankton taxon dominance shifted from diatoms, dinoflagellates, and coccolithophorids to smaller taxa after 2004, whereas mesozooplankton biomass increased and commercial landings of planktivorous sardines collapsed. Collectively, our results reveal an ecological state change in this planktonic system. The weakening trend in Trade Winds (−1.9 ± 0.3%⋅y−1) and dependent local variables are largely explained by trends in two climatic indices, namely the northward migration of the Azores High pressure center (descending branch of Hadley cell) by 1.12 ± 0.42°N latitude and the northeasterly progression of the ITCZ Atlantic centroid (ascending branch of Hadley cell), the March position of which shifted by about 800 km between 1996 and 2009.

Simulation of carbon‐nitrogen cycling during spring upwelling in the Cariaco Basin

Coupled biological-physical models of carbon-nitrogen cycling by phytoplankton, zooplankton, and bacteria assess the impacts of nitrogen fixation and upwelled nitrate during new production within the shelf environs of the Cariaco Basin. During spring upwelling in response to a mean wind forcing of 8 m s−1, the physical model matches remote-sensing and hydrographic estimates of surface temperature. Within the three-dimensional flow field, the steady solutions of the biological model of a simple food web of diatoms, adult calanoid copepods, and ammonifying/nitrifying bacteria approximate within ∼9% the mean spring observations of settling fluxes caught by a sediment trap at ∼240 m, moored at our time series site in the basin. The models also estimate within ∼11% the average 14C net primary production and mimic the sparse observations of the spatial fields of nitrate and light penetration during the same time period of February-April. Stocks of colored dissolved organic matter are evidently small and diazotrophy is minimal during spring. In one summer case of the model with weaker wind forcing, however, the simulated net primary production is 14% of that measured in August-September, while the predicted detrital flux is then 30% of the observed. Addition of a cyanophyte state variable, with another source of new nitrogen, would remedy the seasonal deficiencies of the biological model, attributed to use of a single phytoplankton group.

Protistan microbial observatory in the Cariaco Basin, Caribbean. II. Habitat specialization

This is the second paper in a series of three that investigates eukaryotic microbial diversity and taxon distribution in the Cariaco Basin, Venezuela, the oceans largest anoxic marine basin. Here, we use phylogenetic information, multivariate community analyses and statistical richness predictions to test whether protists exhibit habitat specialization within defined geochemical layers of the water column. We also analyze spatio-temporal distributions of protists across two seasons and two geographic sites within the basin. Non-metric multidimensional scaling indicates that these two basin sites are inhabited by distinct protistan assemblages, an observation that is supported by the minimal overlap in observed and predicted richness of sampled sites. A comparison of parametric richness estimations indicates that protistan communities in closely spaced—but geochemically different—habitats are very dissimilar, and may share as few as 5% of total operational taxonomic units (OTUs). This is supported by a canonical correspondence analysis, indicating that the empirically observed OTUs are organized along opposing gradients in oxidants and reductants. Our phylogenetic analyses identify many new clades at species to class levels, some of which appear restricted to specific layers of the water column and have a significantly nonrandom distribution. These findings suggest many pelagic protists are restricted to specific habitats, and likely diversify, at least in part due to separation by geochemical barriers.

Increased marine sediment suspension and fluxes following an earthquake

Earthquakes are commonly cited as one possible triggering mechanism for turbidity flows—dense sediment–water plumes that can transport large volumes of sediment great distances down slope—in both marine and lacustrine settings. Heezen and Ewing were the first to make such a suggestion, attributing breaks in a sea-floor telephone cable in the North Atlantic Ocean to turbidity flows generated by the 1929 Grand Banks earthquake. Anumber of workers have consequently used sedimentary turbidite records to reconstruct the earthquake histories of various regions. Here we present direct observations of a seismically induced turbidity flow. Measurements of light scattering and sediment fluxes in the Cariaco basin indicate that the earthquake that occurred along the coast of northern Venezuela on 9 July 1997 resulted in considerable downslope displacement of sediments—probably >105 tonnes into the deep part of the basin. In such a seismically active region, this mechanism of sediment transport may be responsible for a significant component of the long-term sediment accumulation in the basin. Furthermore, this process may result in the sequestration in deep sea sediments of large amounts of carbon initially deposited at shallow depths.

Fossil methane source dominates Cariaco Basin water column methane geochemistry

Natural radiocarbon measurements on methane (14C-CH4) extracted from the Cariaco Basin water column show that 98% of the methane in Cariaco Basin waters is derived from fossil (radiocarbon-free) sources. Previous work on Cariaco Basin methane (CH4) considered only a diagenetic sediment source. Similar measurements of sediment 14C-CH4 indicate that sediment CH4 is produced from modern particulate material; thus the sediment and water column CH4 have distinct sources. Using time-dependent CH4 geochemical box models which include a fossil seep CH4 source term, we estimate 1) 0.024-0.028 Tg y-1 of seep CH4 are added to the Cariaco Basin water column, 2) the water column CH4 will reach a steady-state concentration by the year 2065, and 3) the seep CH4 inputs possibly began in 1967, following the July 30 Caracas earthquake. Oxidizing this CH4 to dissolved inorganic carbon does not appear to affect Cariaco Basin 14C chronologies. Copyright 2005 by the American Geophysical Union.

Local and regional geochemical signatures of surface sediments from the Cariaco Basin and Orinoco Delta, Venezuela

We have analyzed the chemical compositions of 87 samples from the Cariaco Basin and 20 samples from the Orinoco Delta, Venezuela, in order to better characterize the fluvial sources of material to the Cariaco Basin. We observe distinctive regional patterns in composition; shelf sediments found near the Tuy River, Manzanares River, the Araya-Margarita region, and in the northern portion of the Unare platform record reproducible compositional differences. However, linear mixing models using only the composition from these rivers do not satisfy the variability observed in modern sediment trap samples collected from the basin. Average trap values are best approximated by mixing upper crust (30%–50%) with local river sources (50%–70% of either the Tuy, Unare, or Neveri Rivers) and eolian dust (

MICROBIAL ECOLOGY OF THE CARIACO BASIN'S REDOXCLINE: THE U.S.-VENEZUELA CARIACO TIMES SERIES PROGRAM

The cooperative U.S.-Venezuela CARIACO program (CArbon Retention I nA Colored Ocean) has begun to elucidate the microbial ecology of the Cariaco Basins redoxcline. This anoxic water column supports highly stratified microbial assemblages of prokaryotes, protozoa and viruses, exhibiting abundance and activity maxima near the O2/H2S interface. In the oxic layer, abundance and activity of microheterotrophs vary annually to the same extent (16 to 20-fold) as primary producers in the upper 75-100 m, but out of phase. In the redoxcline and anoxic layer, relationships of these same variables to surface production are not readily apparent. Heterotrophic carbon demands within the redoxcline exceed delivery of sinking organic matter from the mixed layer. The Cariacos redoxcline appears to be inhabited by microaerophilic and anaerobic chemoautotrophs, such as e-proteobacteria, whose metabolism is controlled by inorganic chemical gradients and transport. Time series data demonstrate that distribution and activity profiles of prokaryotes, protozoa and viruses vary in response to one another and to fluctuations in the interfaces position. Rapid turnover of prokaryotic biomass in the redoxcline is deduced from the perennial presence of bacterivorous protozoan and viral communities. Chemoautotrophic production is sufficient to support heterotrophic demand for reduced carbon within the redoxcline and yields reasonable specific growth rates for total prokaryotic communities, averaging between 0.4 and 0.6 d −1 . However, reconciliation of microbial demand for energy and oxidants with supply is not possible applying the classic 1-D vertical model to the Cariaco and remains one of the greatest challenges to understanding the microbial ecology of anoxic water columns in general.

Identification of bacteria in enrichment cultures of sulfate reducers in the Cariaco Basin water column employing Denaturing Gradient Gel Electrophoresis of 16S ribosomal RNA gene fragments

BackgroundThe Cariaco Basin is characterized by pronounced and predictable vertical layering of microbial communities dominated by reduced sulfur species at and below the redox transition zone. Marine water samples were collected in May, 2005 and 2006, at the sampling stations A (10°30′ N, 64°40′ W), B (10°40′ N, 64°45′ W) and D (10°43’N, 64°32’W) from different depths, including surface, redox interface, and anoxic zones. In order to enrich for sulfate reducing bacteria (SRB), water samples were inoculated into anaerobic media amended with lactate or acetate as carbon source. To analyze the composition of enrichment cultures, we performed DNA extraction, PCR-DGGE, and sequencing of selected bands.ResultsDGGE results indicate that many bacterial genera were present that are associated with the sulfur cycle, including Desulfovibrio spp., as well as heterotrophs belonging to Vibrio, Enterobacter, Shewanella, Fusobacterium, Marinifilum, Mariniliabilia, and Spirochaeta. These bacterial populations are related to sulfur coupling and carbon cycles in an environment of variable redox conditions and oxygen availability.ConclusionsIn our studies, we found an association of SRB-like Desulfovibrio with Vibrio species and other genera that have a previously defined relevant role in sulfur transformation and coupling of carbon and sulfur cycles in an environment where there are variable redox conditions and oxygen availability. This study provides new information about microbial species that were culturable on media for SRB at anaerobic conditions at several locations and water depths in the Cariaco Basin.