On-board flow cytometric observation of picoplankton community structure in the East China Sea during the fall of different years

On-board flow cytometric determinations of picoplankton abundance (i.e. Synechococcus spp., Prochlorococcus spp., picoeukaryotes and also heterotrophic bacteria) were obtained in the East China Sea in fall of 2000 and 2003. The average abundances of Synechococcus, Prochlorococcus, picoeukaryotes and heterotrophic bacteria were 10(5), 10(5), 10(4) and 10(6) cells ml(-1), respectively. Synechococcus, picoeukaryotes and heterotrophic bacteria were abundant at all the stations and presented higher concentration in the inner shelf where influences from the Changjiang effluent plumes and the coastal upwelling were evident, while Prochlorococcus was absent from the near-shore stations and became the dominant picophytoplankton population in offshore waters, where its abundance was comparable to that for heterotrophic bacteria. All picoplankton groups showed a reduction in cell number with depth, and a positive correlation with water temperature were observed, which reflected the importance of light and temperature on picoplankton growth. A negative relationship with salinity was found for heterotrophic bacteria along two sections across the East China Sea Shelf, and distribution of picoplankton was dominated by different water masses. The fixation could lead to loss in Prochlorococcus cell numbers within one month, and all the picoplankton numbers decreased dramatically after three months.     

海洋微型和微微型浮游生物的区域分布与影响因素

微型和微微型浮游生物几乎存在于所有的海洋生态系统中,其群落遍布世界各大洋。在大部分海域,微型浮游生物的优势类群为鞭毛藻;原绿球藻是贫营养海域的优势类群,而聚球藻、微微型真核生物和异养细菌主要在营养盐丰富的海域出现,在热带、亚热带和温带的富营养区域占优势。温度、盐度、光照、营养盐可得性、水体稳定性和摄食压力是影响微型和微微型浮游生物的主要因素,各海域主要调控因子的不同造成了微型和微微型浮游生物类群和丰度分布的差异。本文主要综述海洋微型和微微型浮游生物类群检测方法、区域分布特点及其受环境影响的研究概况,并提出了今后的重点研究内容和发展方向。     

南黄海潮汐锋对浮游细菌生物量分布的影响

2 0 0 1年 5月 16～ 2 3日、6月 10～ 2 4日和 2 0 0 2年 6月 5～ 12日 ,利用“北斗号”船只对南黄海鱼产卵场进行了 3次专项调查。研究了潮汐锋断面叶绿素 a浓度、浮游细菌生物量的分布 ,目的是阐明潮汐锋的存在对浮游细菌生物量分布的影响。3个航次中的叶绿素 a浓度变化范围分别是 0 .0 6～ 2 .34mg/ m3(2 0 0 1- 0 5 )、0 .0 8～ 0 .9mg/ m3(2 0 0 1- 0 6 )、0 .14～ 3.0 4 mg/ m3(2 0 0 2 - 0 6 )。 3航次的聚球藻 (Synechococcus spp.)蓝细菌生物量变化范围分别为 7.6 2～ 2 2 .0 6 mg C/ m3(2 0 0 1- 0 5 )、8.5 3～2 7.5 2 mg C/ m3(2 0 0 1- 0 6 )、0 .6 9～ 5 5 .90 m g C/ m3(2 0 0 1- 0 6 )。异养细菌生物量变化范围分别为 7.5 6～ 5 1.82 mg C/ m3(2 0 0 1-0 5 )、8.5 4～ 2 4 .77mg C/ m3(2 0 0 1- 0 6 )、3.12～ 10 .0 5 mg C/ m3(2 0 0 2 - 0 6 )。而聚球藻蓝细菌对浮游植物总生物量的贡献 (CB:PB)平均值分别为 :5 8% (2 0 0 1- 0 5 )、77% (2 0 0 1- 0 6 )、31% (2 0 0 2 - 0 6 )。结果表明 :南黄海鱼产卵场在春末夏初 (5～ 6月份 ) ,叶绿素 a浓度最大值及次大值主要分布在锋区及其邻近的层化区2 0 m以浅位置 ;聚球藻蓝细菌生物量最大值主要分布于锋区及层化区表层和水体中层 ;异养     

Nanoflagellate predation on auto- and heterotrophic picoplankton in the oligotrophic Mediterranean Sea

Dynamics of autotrophic and heterotrophic prokaryotes and theirconsumption by nanoflagellates were studied in the euphoticzone at nine stations located from the Levantine Basin (34°E)to the Balearic sea (5°E) in June 1999. Bacterial biomassconstituted the largest proportion of living biomass at allstations. Integrated bacterial production at the furthest eaststation, was sixfold lower than integrated bacterial productionat the furthest west (13 and 75 mg C m^-2 d^-1 respectively).Estimated heterotrophic nanoflagellate bacterivory accountedfor 45–87% of bacterial production. Small protists (<3µm) dominated the bacterivore assemblage and accountedfor more than 90% of the heterotrophic bacterial consumption.Our results indicated that there was no negative selection againstSynechococcus and that both picoplankton groups were grazedaccording to their standing stocks. An estimated consumptionof Synechococcus derived from food vacuole content analysisof nanoflagellates revealed that they consumed from 0.5 to 45%(mean 13%) of Synechococcus stock per day. These data are amongthe first documenting the relative grazing impact of heterotrophicnanoflagellates on bacteria and Synechococcus in situ. Assumingthat there was no selection for or against Prochlorococcus,heterotrophic nanoflagellates could ingest from 1.4 to 21% (mean6%) of Prochlorococcus stock per day. The amount of organiccarbon obtained by heterotrophic nanoflagellates from photosyntheticprokaryotes represented 27% of the total amount of carbon obtainedfrom total prokaryotes     

Decrease in the autotrophic-to-heterotrophic biomass ratio of picoplankton in oligotrophic marine waters due to bottle enclosure

We investigated the effects of bottle enclosure on autotrophic and heterotrophic picoplankton in North and South subtropical Atlantic oligotrophic waters, where the biomass and metabolism of the microbial community are dominated by the picoplankton size class. We measured changes in both autotrophic (Prochlorococcus, Synechococcus, and picoeukaryotes) and heterotrophic picoplankton biomass during three time series experiments and in 16 endpoint experiments over 24 h in light and dark treatments. Our results showed a divergent effect of bottle incubation on the autotrophic and heterotrophic components of the picoplankton community. The biomass of picophytoplankton showed, on average, a >50% decrease, mostly affecting the picoeukaryotes and, to a lesser extent, Prochlorococcus. In contrast, the biomass of heterotrophic bacteria remained constant or increased during the incubations. We also sampled 10 stations during a Lagrangian study in the North Atlantic subtropical gyre, which enabled us to compare the observed changes in the auto- to heterotrophic picoplankton biomass ratio (AB:HB ratio) inside the incubation bottles with those taking place in situ. While the AB:HB ratio in situ remained fairly constant during the Lagrangian study, it decreased significantly during the 24 h of incubation experiments. Thus, the rapid biomass changes observed in the incubations are artifacts resulting from bottle confinement and do not take place in natural conditions. Our results suggest that short (<1 day) bottle incubations in oligotrophic waters may lead to biased estimates of the microbial metabolic balance by underestimating primary production and/or overestimating bacterial respiration.     

An immunological approach to detect phosphate stress in populations and single cells of photosynthetic picoplankton.

In the marine cyanobacterium Synechococcus sp. strain WH7803, PstS is a 32-kDa cell wall-associated phosphate-binding protein specifically synthesized under conditions of restricted inorganic phosphate (P1) availability (D. J. Scanlan, N. H. Mann, and N. G. Carr, Mol. Microbiol. 10:181-191, 1993). We have assessed its use as a potential diagnostic marker for the P status of photosynthetic picoplankton. Expression of PstS in Synechococcus sp. strain WH7803 was observed when the P1 concentration fell below 50 nM, demonstrating that the protein is induced at concentrations of P1 typical of oligotrophic conditions. PstS expression could be specifically detected by use of standard Western blotting (immunoblotting) techniques in natural mesocosm samples under conditions in which the N/P ratio was artificially manipulated to force P depletion. In addition, we have developed an immunofluorescence assay that can detect PstS expression in single Synechococcus cells both in laboratory cultures and natural samples. We show that antibodies raised against PstS cross-react with P-depleted Prochlorococcus cells, extending the use of these antibodies to both major groups of prokaryotic photosynthetic picoplankton. Furthermore, DNA sequencing of a Prochlorococcus pstS homolog demonstrated high amino acid sequence identity (77%) with the marine Synechococcus sp. strain WH7803 protein, including those residues in Escherichia coli PstS known to be directly involved in phosphate binding.     

Lake Superior supports novel clusters of cyanobacterial picoplankton

Very little is known about the biodiversity of freshwater autotrophic picoplankton (APP) in the Laurentian Great Lakes, a system comprising 20% of the world's lacustrine freshwater. In this study, the genetic diversity of Lake Superior APP was examined by analyzing 16S rRNA gene and cpcBA PCR amplicons from water samples. By neighbor joining, the majority of 16S rRNA gene sequences clustered within the "picocyanobacterial clade" consisting of freshwater and marine Synechococcus and Prochlorococcus. Two new groups of Synechococcus spp., the pelagic Lake Superior clusters I and II, do not group with any of the known freshwater picocyanobacterial clusters and were the most abundant species (50 to 90% of the sequences) in samples collected from offshore Lake Superior stations. Conversely, at station Portage Deep (PD), located in a nearshore urbanized area, only 4% of the sequences belonged to these clusters and the remaining clones reflected the freshwater Synechococcus diversity described previously at sites throughout the world. Supporting the 16S rRNA gene data, the cpcBA library from nearshore station PD revealed a cosmopolitan diversity, whereas the majority of the cpcBA sequences (97.6%) from pelagic station CD1 fell within a unique Lake Superior cluster. Thus far, these picocyanobacteria have not been cultured, although their phylogenetic assignment suggests that they are phycoerythrin (PE) rich, consistent with the observation that PE-rich APP dominate Lake Superior picoplankton. Lastly, flow cytometry revealed that the summertime APP can exceed 10(5) cells ml-1 and suggests that the APP shifts from a community of PE and phycocyanin-rich picocyanobacteria and picoeukaryotes in winter to a PE-rich community in summer.     

Aerobic anoxygenic phototrophic bacteria in the Mid-Atlantic Bight and the North Pacific Gyre

The abundance of aerobic anoxygenic phototrophic (AAP) bacteria, cyanobacteria, and heterotrophs was examined in the Mid-Atlantic Bight and the central North Pacific Gyre using infrared fluorescence microscopy coupled with image analysis and flow cytometry. AAP bacteria comprised 5% to 16% of total prokaryotes in the Atlantic Ocean but only 5% or less in the Pacific Ocean. In the Atlantic, AAP bacterial abundance was as much as 2-fold higher than that of Prochlorococcus spp. and 10-fold higher than that of Synechococcus spp. In contrast, Prochlorococcus spp. outnumbered AAP bacteria 5- to 50-fold in the Pacific. In both oceans, subsurface abundance maxima occurred within the photic zone, and AAP bacteria were least abundant below the 1% light depth. The abundance of AAP bacteria rivaled some groups of strictly heterotrophic bacteria and was often higher than the abundance of known AAP bacterial genera (Erythrobacter and Roseobacter spp.). Concentrations of bacteriochlorophyll a (BChl a) were low ( approximately 1%) compared to those of chlorophyll a in the North Atlantic. Although the BChl a content of AAP bacteria per cell was typically 20- to 250-fold lower than the divinyl-chlorophyll a content of Prochlorococcus, the pigment content of AAP bacteria approached that of Prochlorococcus in shelf break water. Our results suggest that AAP bacteria can be quite abundant in some oceanic regimes and that their distribution in the water column is consistent with phototrophy.     

Cyanobacterial photosynthesis in the oceans: the origins and significance of divergent light-harvesting strategies

Prochlorococcus and Synechococcus are abundant unicellular cyanobacteria and major participants in global carbon cycles. Although they are closely related and often coexist in the same ocean habitat, they possess very different photosynthetic light-harvesting antennas. Whereas Synechococcus and the majority of cyanobacteria use phycobilisomes, Prochlorococcus has evolved to use a chlorophyll a(2)/b(2) light-harvesting complex. Here, we present a scenario to explain how the Prochlorococcus antenna might have evolved in an ancestral cyanobacterium in iron-limited oceans, resulting in the diversification of the Prochlorococcus and marine Synechococcus lineages from a common phycobilisome-containing ancestor. Differences in the absorption properties and cellular costs between chlorophyll a(2)/b(2) and phycobilisome antennas in extant Prochlorococcus and Synechococcus appear to play a role in differentiating their ecological niches in the ocean environment.     

Spatial and Temporal Variation of Enterobacter Genotypes in Sediments and the Underlying Hyporheic Zone of an Agricultural Stream

Population studies of enteric bacteria in an agriculturally impacted stream (Ledbetter Creek, Murray, Kentucky, USA) were conducted over a period of 2 years. Total number of bacteria, cultivated heterotrophic aerobic bacteria, and enteric bacteria showed significant differences between winter and summer. The cultivated numbers of heterotrophic aerobic bacteria and enteric bacteria were significantly more abundant in summer than in winter. The abundance of enteric bacteria was 12.9% in an upwelling zone and 9.8% in a downwelling zone in summer. Most of the enteric bacterial strains isolated on MacConkey agar were assigned to Enterobacter cloacae and E. agglomerans by API 20E and an analysis of the restriction patterns produced by amplified DNA coding for 16S rRNA (ARDRA) with the enzyme Hpa II. E. cloacae and E. agglomerans genotypes isolated from three hyporheic and gravel bar depth intervals (0-10 cm, 15-25 cm, and 30-40 cm) in summer and fall showed significant spatial variation and were heterogeneously distributed along the stream. Temperature, inorganic nutrients, and occurrence of anoxic zones affected the distribution of enteric bacteria. These techniques can be used as a model to monitor shifts among different species in the stream ecosystem.     

Spatial and temporal distribution of nanoflagellates in the northern South China Sea

Seasonal variation, horizontal and vertical distribution, and cell size of nanoflagellates, together with physico-chemical and biological factors, were studied in the northern South China Sea (SCS). It was found that nanoflagellate abundance ranged from 0.157 × 10³ to 9.193 × 10³ cells/ml (with a mean of 0.891 × 10³) in winter (February, 2004), while it ranged from 0.107 × 10³ to 5.417 × 10³ cells/ml (with a mean of 0.599 × 10³) in summer (July, 2004). Nanoflagellates were more abundant in winter than summer in offshore regions, showing an unique seasonal pattern in this subtropical marginal sea. The abundance of nanoflagellates decreased from the estuary to the offshore region. Vertical distribution of nanoflagellates coupled well with that of bacteria and Chl a. The small size fraction of less than 5 μm dominated the nanoflagellate populations. Wind-driven mixing, eddies, availability of nutrients as well as Chl a and abundance of picoplankton seemed to be the major controlling factors for the spatial distribution and seasonal variation of nanoflagellates in the study area. Handling editor: P. Tasman     

Seasonal variation of phototrophic picoplankton in Lake Biwa (1994–1998)

Seasonal abundances of phototrophic picoplankton (PP) and heterotrophic nanoflagellates in Lake Biwa were studied from 1994 to 1998. Seasonal variation in cell volume and biomass of the phototrophic picoplankton were also studied. PP were counted using disposable glass microscopic plates, which gave superior accuracy to sample filtration onto membrane filters. Phycoerythrin-rich rod-shaped cyanobacteria (PEC), one of the major components of the picoplankton community, were sparse (about 10⁴ cells ml ^–1) in winter and began to increase in April. Several PEC peaks were observed during the period of thermal stratification, and a rapid fall took place after October or November. In the northern basin, PEC peaked during late June and early July in 3 of the 5 years, and in late summer in the remaining years. Phycocyanin-rich rod-shaped cyanobacteria (PCC) were abundant in the southern basin and were present in smaller numbers in the eutrophic nearshore area of the northern basin; they peaked several times during the period from July to October. Seasonal variations of these two kinds of picoplankton were correlated with seasonal changes in water temperature. Phycoerythrin-rich cylinder-shaped cyanobacteria exhibited narrow peaks in July, their abundance declining as the year progressed. The density of heterotrophic nanoflagellates was greatest in early spring. Average cell volume of PEC was largest in winter, then decreased gradually to a minimum in late summer; after the fall, it recovered to the winter cell volume. This change can likely be attributed to the depletion of nitrogen in the warmer seasons.     

OPTICAL BIOGEOGRAPHY OF PROCHLOROCOCCUS AND PHYCOERYTHRIN CONTAINING PICOCYANOBACTERIA ON THE WEST FLORIDA SHELF

Wood, A. M.¹, Li, W. K. W.², Arnone, R.³, Gould, R.⁴, and Lohrenz, S.^{4 1}Dept. of Biology, University of Oregon, Eugene, OR 97403, USA; ²Bedford Institute of Oceanography, Dartmouth, N.S., B2Y4A2, Canada; ³Naval Research Laboratory, Code 7333, Stennis Space Center, MS 39529 USA; ⁴Department of Marine Science, University of Southern Mississippi, Stennis Space Center, MS 39259 USA Prochlorococcus and phycoerythrin-containing picocyano-bacteria co-dominate oligotrophic waters in temperate and tropical areas. Less is known about their distribution in coastal areas where optical conditions lead to shallower euphotic zones and a different attenuation spectra for incident light. In this study we compare the distribution of Prochlorococcus and phycoerythrin-containing picocyano-bacteria (mostly Synechococcus) with data on inherent and apparent optical properties in a subtropical continental shelf environment. Abundance and pigment diversity for these two major groups of picoplankton was determined by flow cytometry and several fluorescence approaches at 22 stations on the continental shelf off of Tampa Bay, Florida, in the Gulf of Mexico. Hydrographic and optical data indicated that a wide range of optical environments were sampled, ranging from turbid, shallow water influenced by exchange with Tampa Bay, to very transparent oceanic water on the outer shelf. The abundance of the two major groups of picoplanktonic cyanobacteria, Synechococcus and Prochlorococcus, was negatively correlated with Prochlorococcus reaching peak concentrations of ~105 ml^-1 on the outer shelf and Synechococcus reaching peak concentrations of ~5 X 105 ml^-1 on the inner shelf. The high abundance of Synechococcus in turbid, green waters on the inner shelf indicates that this taxon includes a number of strains that are adapted to coastal environments and that, as a group, they do equally well in neritic waters as in oceanic regions. In contrast, Prochlorococcus appears to be an open-ocean specialist.     

Predation impact of ciliated and flagellated protozoa during a summer bloom of brown sulfur bacteria in a meromictic coastal lake

Anaerobic phagotrophic protozoa may play an important role in the carbon flux of chemically stratified environments, especially when phototrophic sulfur bacteria account for a high proportion of the primary production. To test this assumption, we investigated the vertical and temporal distribution of microbial heterotrophs and of autotrophic picoplankton throughout the water column of the meromictic coastal lake Faro (Sicily, Italy), in the summer of 2004, coinciding with a bloom of brown-colored green sulfur bacteria. We also assessed the grazing impact of ciliated and flagellated protozoa within the sulfur bacteria plate using a modification of the fluorescently labeled bacteria uptake approach, attempting to minimize the biases intrinsic to the technique and to preserve the in situ anoxic conditions. Significant correlations were observed between ciliate biomass and bacteriochlorophyll e concentration, and between heterotrophic nanoflagellate biomass and chlorophyll a concentration in the water column. The major predators of anaerobic picoplankton were pleuronematine ciliates and cryptomonad flagellates, with clearances of 26.6 and 9.5 nL per cell h⁻¹, respectively, and a cumulative impact on the picoplankton gross growth rate ranging between 36% and 72%. We concluded that protozoan grazing channels a large proportion of anaerobic picoplankton production to higher trophic levels without restraining photosynthetic bacteria productivity.     

青岛近海及其临近海域冬季微微型浮游植物的分布

微微型浮游植物（0.2～2.0 μm） 是海水中最小的自养浮游生物, 在世界各海域广泛分布, 并在海洋有机物质循环中起着非常重要的作用.利用荧光显微技术对青岛近海及其邻近海域冬季微微型浮游植物丰度进行了调查,研究了微微型浮游植物的空间变化和昼夜变化的特征, 并分析了微微型浮游植物丰度与环境因子的相关性. 结果显示, 冬季该海域以富含藻红素（Phycoerythrin-rich, PE）的聚球藻(Synechococcus, Syn)细胞占优势,微微型真核藻类（Picoeukaryote, Euk）次之,而富含藻蓝素（Phycocyanin-rich, PC）的聚球藻细胞数量很低, 未发现原绿球藻（Prochlorococcus, Pro）的存在.Syn 的变化范围为8.97×10³～1.95×10⁵ cells·ml^-1, 平均4.67×10⁴ cells·ml^-1; Euk的变化范围为1.95×10²～1.01×10⁴ cells·ml^-1, 平均2.39×10³ cells·ml^-1. Syn丰度在胶南以南海域出现高值区域, 在即墨海域和崂山东南海域出现低值区域. Euk丰度在日照海域出现高值区域; 崂山海域为低值区域; 各水层Syn和Euk丰度均无明显差异(P﹥0.05). 对胶州湾中部连续站4层水体的24 h昼夜连续变化进行观测发现, Syn、Euk丰度都有明显昼夜波动.相关性分析表明： Syn与温度、 电导率呈正相关, 与溶氧浓度呈显著负相关; Euk与盐度和溶氧浓度呈显著负相关. 微微型浮游植物对总浮游植物生物量的贡献约为20％.     

Autotrophic picoplankton in southern Lake Baikal: abundance, growth and grazing mortality during summer

Autotrophic picoplankton were highly abundant during the thermalstratification period in late July in the pelagic area (waterdepth 500–1300 m) of southern Lake Baikal; maximum numberswere 2 x 10⁶ cells ml^–1 in the euphotic zone ({small tilde}15m). Unicellular cyanobacteria generally dominated the picoplanktoncommunity, although unidentified picoplankton that fluorescedred under blue excitation were also abundant (maximum numbers4 x 10⁵ cells ml^–1) and contributed up to {small tilde}40%of the total autotrophic picoplankton on occasions. Carbon andnitrogen biomasses of autotrophic picoplankton estimated byconversion from biovolumes were 14–84 µg C l^–1and 3.6–21 µg N l^–1. These were comparableto or exceeded the biomass of heterotrophic bacteria. Autotropicpicoplankton and bacteria accounted for as much as 33% of paniculateorganic carbon and 81% of nitrogen in the euphotic zone. Measurementsof the photosynthetic uptake of [^l4C]bicarbonate and the growthof picoplankton in diluted or size-fractionated waters revealedthat 80% of total primary production was due to picoplankton,and that much of this production was consumed by grazers inthe <20 µ.m cell-size category. These results suggestthat picoplankton-protozoan trophic coupling is important inthe pelagic food web and biogeochemical cycling of Lake Baikalduring summer.     

Grazing impact of different heterotrophic nanoflagellates on eukaryotic (Ostreococcus tauri) and prokaryotic picoautotrophs (Prochlorococcus and Synechococcus)

Autotrophic picoplankton (<3 microm) composed of both prokaryotes and eukaryotes are the most abundant primary producers on Earth. In this study we examined the ingestion of the picoeukaryote Ostreococcus tauri by different marine heterotrophic nanoflagellates (HNF) with various morphologies, swimming and feeding behaviours. Cultures of specific bacterivorous nanoflagellates (Rhynchomonas nasuta, Jakoba libera, and a culture of Cafeteria sp./Monosiga sp.) and natural nanoflagellate populations were used as grazers. For comparison with Ostreococcus, we used similar-sized prokaryotes as prey, Prochlorococcus and Synechococcus. We observed large species-specific differences in terms of: use of picoautotrophs among nanoflagellates, time lag between prey addition and prey consumption (0-196 h), grazing rate (0-0.12 h(-1)), growth rate (0-0.3 h(-1)) and maximum abundance of HNF reached in experimental bottles (e.g. from 10(4) to 10(5) cells ml(-1), for a natural coastal population and a Cafeteria sp./Monosiga sp. culture feeding Ostreococcus respectively). Overall, this study shows that the nanoflagellate community composition is conclusive for picoautotrophic community structure and, vice versa, the picoautotrophic community structure favours or inhibits the growth of some nanoflagellate groups.     

In situ hybridization of Prochlorococcus and Synechococcus (marine cyanobacteria) spp. with RRNA-targeted peptide nucleic acid probes

A simple method for whole-cell hybridization using fluorescently labeled rRNA-targeted peptide nucleic acid (PNA) probes was developed for use in marine cyanobacterial picoplankton. In contrast to established protocols, this method is capable of detecting rRNA in Prochlorococcus, the most abundant unicellular marine cyanobacterium. Because the method avoids the use of alcohol fixation, the chlorophyll content of Prochlorococcus cells is preserved, facilitating the identification of these cells in natural samples. PNA probe-conferred fluorescence was measured flow cytometrically and was always significantly higher than that of the negative control probe, with positive/negative ratio varying between 4 and 10, depending on strain and culture growth conditions. Prochlorococcus cells from open ocean samples were detectable with this method. RNase treatment reduced probe-conferred fluorescence to background levels, demonstrating that this signal was in fact related to the presence of rRNA. In another marine cyanobacterium, Synechococcus, in which both PNA and oligonucleotide probes can be used in whole-cell hybridizations, the magnitude of fluorescence from the former was fivefold higher than that from the latter, although the positive/negative ratio was comparable for both probes. In Synechococcus cells growing at a range of growth rates (and thus having different rRNA concentrations per cell), the PNA- and oligonucleotide-derived signals were highly correlated (r = 0.99). The chemical nature of PNA, the sensitivity of PNA-RNA binding to single-base-pair mismatches, and the preservation of cellular integrity by this method suggest that it may be useful for phylogenetic probing of whole cells in the natural environment.     

The diversity of small eukaryotic phytoplankton (< or =3 microm) in marine ecosystems

Small cells dominate photosynthetic biomass and primary production in many marine ecosystems. Traditionally, picoplankton refers to cells ≤2 μm. Here we extend the size range of the organisms considered to 3 μm, a threshold often used operationally in field studies. While the prokaryotic component of picophytoplankton is dominated by two genera, Prochlorococcus and Synechococcus , the eukaryotic fraction is much more diverse. Since the discovery of the ubiquitous Micromonas pusilla in the early 1950s, just over 70 species that can be <3 μm have been described. In fact, most algal classes contain such species. Less than a decade ago, culture-independent approaches (in particular, cloning and sequencing, denaturing gradient gel electrophoresis, FISH) have demonstrated that the diversity of eukaryotic picoplankton is much more extensive than could be assumed from described taxa alone. These approaches revealed the importance of certain classes such as the Prasinophyceae but also unearthed novel divisions such as the recently described picobiliphytes. In the last couple of years, the first genomes of photosynthetic picoplankton have become available, providing key information on their physiological capabilities. In this paper, we discuss the range of methods that can be used to assess small phytoplankton diversity, present the species described to date, review the existing molecular data obtained on field populations, and end up by looking at the promises offered by genomics.     

Dynamic characteristics of Prochlorococcus and Synechococcus consumption by bacterivorous nanoflagellates

We compared the characteristics of ingestion of Prochlorococcus and Synechococcus by the marine heterotrophic nanoflagellate Pseudobodo sp. and by a mixed nanoflagellate culture (around 3 microm in size) obtained from an open sea oligotrophic area. Maximum ingestion rate on Synechococcus (2.7 Syn flagellate(-1) h(-1)) was reached at concentrations of 5 x 10(5) Syn mL(-1) and decreased between 6 x 10(5) and 1.5 x 10(6) Syn mL(-1). In order to validate laboratory data, one set of data on Synechococcus grazing was obtained during a field study in the oligotrophic northeastern Mediterranean Sea. Ingestion rates by heterotrophic nanoflagellates were related to Synechococcus abundance in the water, and the feeding rate showed a clear diel rhythm with consumption being highest during the night, declining during the day hours, and being lowest at dusk. Ingestion rates on Prochlorococcus increased linearly for the whole range of prey density used (i.e., from 1 x 10(3) to 3 x 10(6) Proc mL(-1)), with maximum ingestion of 6.7 Proc flagellate(-1) h(-1). However, for prey concentrations in the range of 10(3)-10(5), which are usually encountered in aquatic systems, ingestion rates were significantly less than on Synechococcus. In our experiments, both Prochlorococcus and Synechococcus proved to be poor food items for support of nanoflagellate growth.