Ecological genomics of marine picocyanobacteria.

Marine picocyanobacteria of the genera Prochlorococcus and Synechococcus numerically dominate the picophytoplankton of the world ocean, making a key contribution to global primary production. Prochlorococcus was isolated around 20 years ago and is probably the most abundant photosynthetic organism on Earth. The genus comprises specific ecotypes which are phylogenetically distinct and differ markedly in their photophysiology, allowing growth over a broad range of light and nutrient conditions within the 45 degrees N to 40 degrees S latitudinal belt that they occupy. Synechococcus and Prochlorococcus are closely related, together forming a discrete picophytoplankton clade, but are distinguishable by their possession of dissimilar light-harvesting apparatuses and differences in cell size and elemental composition. Synechococcus strains have a ubiquitous oceanic distribution compared to that of Prochlorococcus strains and are characterized by phylogenetically discrete lineages with a wide range of pigmentation. In this review, we put our current knowledge of marine picocyanobacterial genomics into an environmental context and present previously unpublished genomic information arising from extensive genomic comparisons in order to provide insights into the adaptations of these marine microbes to their environment and how they are reflected at the genomic level.

Diel patterns of UVBR-induced DNA damage in picoplankton size fractions from the Gulf of Aqaba, Red Sea.

This study focuses on the impact of natural levels of UVBR (ultraviolet-B radiation: 280 to 315 nm) on bacterio- and phytoplankton (<10 microm) from the Gulf of Aqaba, Red Sea. Incident biologically effective doses (BEDs) and attenuation of biologically effective radiation in the water column were measured using a DNA biodosimeter. UVBR-induced DNA damage was measured as cyclobutane pyrimidine dimers (CPDs), using an antibody directed to CPDs followed by chemiluminescent detection. Depth profiles of DNA damage were determined in two plankton size fractions (0.2 to 0.8 microm and 0.8 to 10 microm) collected down to 50 m depth. Furthermore, accumulation and removal of CPDs were monitored in surface plankton samples during several daily cycles. Small plankton (plankton <10 microm) composition was determined by flow cytometry. The plankton community in the Gulf of Aqaba was dominated by nonphototrophic bacteria and the free-living prochlorophyte Prochlorococcus spp. (<0.8 microm). In general, no DNA damage could be detected in dosimeter DNA below 15 m. In contrast, DNA damage (up to 124 CPD Mnucl-1) could be detected in all bacterio- and phytoplankton samples. DNA damage accumulated throughout the day, indicating that plankton in the Gulf of Aqaba undergo UVBR stress via CPD induction. Although the numbers of CPDs decreased during darkness, both size fractions showed some residual DNA damage at the end of the night. This suggests that dark repair processes did not remove all CPDs, or that part of the plankton community was incapable of repair at all. CPD levels in the two size fractions showed no significant differences in situ. During full solar radiation exposures (samples incubated in bags), more CPDs were detected in the smaller (0.2 to 0.8 microm) size fraction as compared to the larger (0.8 to 10 microm) size fraction. In these experiments, initial plankton composition was significantly different from the field samples. This implies that a shift in the population structure or irradiance conditions can lead to a significant change in UVBR sensitivity. In conclusion, the results show that the picoplankton-dominated phyto- and bacterioplankton communities in the clear surface waters from the Gulf of Aqaba undergo UVBR stress. Repair pathways are not sufficient to eliminate damage during or after UVBR exposure hours, suggesting photomortality as a potential loss parameter of the plankton community.

Ecological aspects of ntcA gene expression and its use as an indicator of the nitrogen status of marine Synechococcus spp.

Nitrogen nutrition in cyanobacteria is regulated by NtcA, a transcriptional activator that is subject to negative control by ammonium. Using Synechococcus sp. strain WH7803 as a model organism, we show that ntcA expression was induced when cells were exposed to nitrogen stress but not when they were subjected to phosphorus or iron deprivation. Transcript levels accumulated in cells grown on a variety of inorganic and organic nitrogen sources, with the sole exception of ammonium. ntcA transcription was induced when ammonium levels dropped below 1 microM and reached maximal levels within 2 h. Furthermore, the addition of more than 1 microM ammonium led to a rapid decline in ntcA mRNA. The negative effect of ammonium was prevented by the addition of L-methionine-D,L-sulfoximine (MSX) and azaserine, inhibitors of ammonium assimilation. Thus, basal ntcA transcript levels are indicative of ammonium utilization. Conversely, the highest ntcA transcript levels were found in cells lacking a nitrogen source capable of supporting growth. Therefore, maximal ntcA expression would indicate nitrogen deprivation. This state of nitrogen deprivation was induced by a 1-h incubation with MSX. The rapid response of ntcA gene expression to the addition of ammonium and MSX was used to design a protocol for assessing relative ntcA transcript levels in field populations of cyanobacteria, from which their nitrogen status can be inferred. ntcA was basally expressed in Synechococcus at a nutrient-enriched site at the northern tip of the Gulf of Aqaba, Red Sea. Therefore, these cyanobacteria were not nitrogen stressed, and their nitrogen requirements were met by regenerated nitrogen in the form of ammonium.

Occipital neuralgia secondary to hypermobile posterior arch of atlas. Case report.

The authors report on the management of occipital neuralgia secondary to an abnormality of the atlas in which the posterior arch was separated by a fibrous band from the lateral masses, resulting in C-2 nerve root compression. The causes and treatments of occipital neuralgia as well as the development of the atlas are reviewed.

Nitrate assimilation genes of the marine diazotrophic, filamentous cyanobacterium Trichodesmium sp. strain WH9601.

A 4.0-kb DNA fragment of Trichodesmium sp. strain WH9601 contained gene sequences encoding the nitrate reduction enzymes, nirA and narB. A third gene positioned between nirA and narB encodes a putative membrane protein with similarity to the nitrate permeases of Bacillus subtilis (NasA) and Emericella nidulans (CrnA). The gene was shown to functionally complement a DeltanasA mutant of B. subtilis and was assigned the name napA (nitrate permease). NapA was involved in both nitrate and nitrite uptake by the complemented B. subtilis cells. napA is distinct from the nrt genes that encode the nitrate transporter of freshwater cyanobacteria.

Regulation of ntcA expression and nitrite uptake in the marine Synechococcus sp. strain WH 7803.

NtcA is a transcriptional activator involved in global nitrogen control in cyanobacteria. In the absence of ammonium it regulates the transcription of a series of genes encoding proteins required for the uptake and assimilation of alternative nitrogen sources (I. Luque, E. Flores, and A. Herrero, EMBO J. 13:2862-2869, 1994). ntcA, present in a single copy in the marine Synechococcus sp. strain WH 7803, was cloned and sequenced. The putative amino acid sequence shows a high degree of identity to NtcA from freshwater cyanobacteria in two functional domains. The expression of ntcA was negatively regulated by ammonium from a putative transcription start point located downstream of an NtcA consensus recognition sequence. Addition of either rifampin or ammonium led to a rapid decline in ntcA transcript levels with half-lives of less than 2 min in both cases. Nitrate-grown cells showed high ntcA transcript levels, as well as the capacity for active nitrite uptake. However, ammonium-grown cells showed low levels of the ntcA transcript and did not utilize nitrite. The addition of ammonium to nitrite uptake-active cells resulted in a gradual decline in the rate of uptake over a 24-h period. Active nitrite uptake was not induced in cells transferred to medium lacking a nitrogen source despite evidence of elevated expression of ntcA, indicating that ntcA expression is not sufficient for uptake capacity to develop. Nitrate and nitrite addition led to the development of nitrite uptake, whereas the addition of leucine did not. Furthermore, nitrite addition triggered the de novo protein synthesis required for uptake capacity to develop. These data suggest that nitrite and nitrate act as specific inducers for the synthesis of proteins required for nitrite uptake.

Photosynthesis of Prochlorothrix hollandica under Sulfide-Rich Anoxic Conditions.

The photosynthetic activity and photosystem II fluorescence of Prochlorothrix hollandica were studied under anoxic, sulfide-rich conditions. Oxygenic photosynthetic activity with water as the electron donor was highly resistant to inhibition by sulfide. Cells still retained 50% of their oxygenic photosynthetic activity at >1 mM sulfide. In the presence of DCMU [N-(3,4-dichlorophenyl)-N(prm1)-dimethylurea], an inhibitor of photosystem II activity, P. hollandica cells exhibited a low but significant anoxygenic photosynthetic activity when sulfide was present. This activity increased with higher sulfide concentrations and reached maximal rates at concentrations exceeding 1 mM sulfide. The effects of hydroxylamine on both oxygen evolution and fluorescence induction kinetics were similar to those observed for sulfide. It was concluded that the oxidizing site of photosystem II was the site of sulfide action leading to reduced or even fully inhibited electron donation to photosystem II. These observations bear similarity to the situation in some cyanobacteria in which both hydroxylamine and sulfide inhibit electron donation from H(inf2)O to P(inf680). The high resistance of photosystem II to sulfide is related to the hydrophobic nature of the manganese-stabilizing protein in P. hollandica (T. S. Mor, A. F. Post, and I. Ohad, Biochim. Biophys. Acta 1141:206-212, 1993). The observed sulfide tolerance of P. hollandica may confer a competitive advantage in its natural environment, where it forms a dominant fraction of phytoplankton in waters in which sulfide presence is a recurring phenomenon.

Cloning and functional analysis of the pmmA gene encoding phosphomannomutase from the photosynthetic prokaryote Prochlorothrix hollandica.

The pmmA gene encoding a bifunctional phosphomannomutase/phosphoglucomutase (PMM/PGM) from the photosynthetic prokaryote, Prochlorothrix hollandica has been cloned and sequenced. The gene encodes a 51827 Da polypeptide 48% identical to the PMM of Azospirillum brasilense, 37% identical to the PGMs of pathogenic Neisseria sp. and 37% identical to the bifunctional AlgC PGM/PMM of Pseudomonas aeruginosa. The pmmA gene encodes an enzyme having both PGM and PMM activities as judged by both enzyme assays and complementation analysis in which the cloned gene partially corrected the pgm-1 mutation of Escherichia coli.

The prochlorophytes: are they more than just chlorophyll a/b-containing cyanobacteria?

The prochlorophytes are a diverse group of photosynthetic prokaryotes that fall within the cyanobacterial lineage, yet lack phycobilisomes as light harvesting structures. Instead, the prochlorophytes have a light-harvesting apparatus composed of the higher plant pigments chlorophylls a and b. This review discusses the evolutionary relationships among these bacteria, with focus on the structure and function of the photosynthetic apparatus. This analysis yields a consensus from studies both on Prochloron sp. and Prochlorothrix hollandica as to how the thylakoid membrane is organized. Overall, we propose that the structure of the light-harvesting complexes (LHC) from prochlorophytes is very different from those of chloroplast systems, and is evolutionarily very ancient. The functional association of the light-harvesting apparatus with photosystem I (PSI) in both Prochlorothrix and Prochloron, as well as a demonstrated capacity for PSI-dependent anoxygenic photosynthesis in Prochlorothrix, may indicate that there is an increased dependence on cyclic photophosphorylation in these organisms. Finally, the structure of the prochlorophyte thylakoid membrane is discussed with respect to the forces that drive thylakoid membrane stacking in prochlorophytes and chloroplasts. We suggest that the light-harvesting structures in prochlorophytes play little, if any, role in this process.

Functional Analysis of the Photosynthetic Apparatus of Prochlorothrix hollandica (Prochlorales), a Chlorophyll b Containing Procaryote.

Light-shade adaptation of the chlorophyll a/b containing procaryote Prochlorothrix hollandica was studied in semicontinuous cultures adapted to 8, 80 and 200 mumole quanta per square meter per second. Chlorophyll a contents based on dry weight differed by a factor of 6 and chlorophyll b by a factor of 2.5 between the two extreme light conditions. Light utilization efficiencies determined from photosynthesis response curves were found to decrease in low light grown cultures due to lower light harvesting efficiencies; quantum requirements were constant at limiting and saturating irradiances for growth. At saturating growth irradiances, changes in light saturated oxygen evolution rate originated from changes in chlorophyll a antenna relative to the number of reaction centers II. At light-limiting conditions both the number of reaction centers II and the antenna size changed. The amount of chlorophyll b relative to reaction center II remained constant. As in cyanobacteria, the ratio of reaction center I to reaction center II was modulated during light-shade adaptation. On the other hand, time constants for photosynthetic electron transport (4 milliseconds) were low as observed in green algae and diatoms. The occurrence of state one to two and state two to one transitions is reported here. Another feature linking photosynthetic electron transport in P. hollandica to that in the eucaryotic photosynthetic apparatus was blockage of the state one to two transition by 3-(3,4-dichlorophenyl)-1,1-dimethylurea. Although chlorophyll b was reported in association with photosystem I, the 630 nanometer light effect does not exclude that chlorophyll b is the photoreceptor for the state one to two transition.