Lack of relationship between gliding cyanobacteria and filamentous gliding heterotrophic eubacteria: comparison of 16S rRNA catalogues of Spirulina,Saprospira, Vitreoscilla,Leucothrix, and Herpetosiphon

Spirulina platensis and Spirulina maxima were characterized by 16S ribosomal RNA oligonucleotide cataloguing. Both species are highly related. They fall into the main group of the cyanobacteria, showing a remote relationship to Nostoc, Fischerella, Aphanocapsa, and also to Prochloron. Low similarity coefficients were found between the Spirulina species and certain organotrophic and sulfide oxidizing bacteria, viz. Saprospira grandis, Vitreoscilla stercoraria, Leucothrix mucor, Herpetosiphon aurantiacus, and Beggiatoa leptomitiformis, respectively. This result does not support the classical hypothesis that certain filamentous gliding bacteria are apochlorotic cyanobacteria.     

Intracellular coagulation inhibits the extraction of proteins from Prochloron

Protein extraction from the prokaryotic alga Prochloron LP (isolated from the ascidian host Lissoclinum patella) was complicated by an irrevers     

Assessment of Photosynthetic Performance of Prochloron in Lissoclinum patella in hospite by Chlorophyll Fluorescence Measurements

Two new PAM fluorometers (pulse amplitude modulated) were usedin an investigation of photosynthetic performance of Prochloronresident as a symbiont in the ascidian Lissoclinum patella,growing in a coral reef of Heron Island on the Great BarrierReef. With a new DIVING-PAM in situ measurements of effectivePSII quantum yield (     

The α-1,4-glucans of Prochloron, a prokaryotic green marine alga

Prochloron, a symbiont found associated with Lissoclinum patella (a marine colonial ascidian), was lyophilized and its glucans extracted. The glucans were complexed with s-triazine reactive dyes and separated by electrophoresis on cellulose acetate membranes. A highly branched glucan similar to phytoglycogen, and a linear unbranched glucan resembling a short-chain amylose were both detected. This unusual polysaccharide mixture suggests a possible mode of starch biosynthesis in algae in general.     

Photoadaptation and Protection against Active Forms of Oxygen in the Symbiotic Procaryote Prochloron sp. and Its Ascidian Host

Superoxide dismutase, ascorbate peroxidase, and catalase activities were studied in the symbiotic photosynthetic procaryote Prochloron sp. and its ascidian host Lissoclinum patella. The protein-specific activities of these antioxidant enzymes in the Prochloron sp. and L. patella collected at different depths from the Great Barrier Reef, Australia, were directly proportional to irradiance, whereas the pigment concentrations in the Prochloron sp. were inversely proportional to irradiance. The presence of a cyanide-sensitive superoxide dismutase, presumably a Cu-Zn metalloprotein, in the Prochloron sp. extends the possible phylogenetic distribution of this protein. The concentration of UV-absorbing mycosporine-like amino acids is inversely proportional to irradiance in both the host and symbiont, suggesting that these compounds may not provide sufficient protection against UV radiation in high-irradiance environments. The significant differences in the specific activities of these antioxidant enzymes, cellular photosynthetic pigment concentrations, and UV-absorbing compounds from high- and low-irradiance habitats constitute an adaptive response to different photic environments. These photoadaptive responses are essential to prevent inhibition of photosynthesis by high fluxes of visible and UV radiation.     

Prevalence,complete genome,and metabolic potentials of a phylogenetically novel cyanobacterial symbiont in the coral-killing sponge,Terpios hoshinota

Terpios hoshinota is an aggressive, space-competing sponge that kills various stony corals. Outbreaks of this species have led to intense damage to coral reefs in many locations. Here, the first large-scale 16S rRNA gene survey across three oceans revealed that bacteria related to the taxa Prochloron, Endozoicomonas, SAR116, Ruegeria, and unclassified Proteobacteria were prevalent in T. hoshinota. A Prochloron-related bacterium was the most dominant and prevalent cyanobacterium in T. hoshinota. The complete genome of this uncultivated cyanobacterium and pigment analysis demonstrated that it has phycobiliproteins and lacks chlorophyll b, which is inconsistent with the definition of Prochloron. Furthermore, the cyanobacterium was phylogenetically distinct from Prochloron, strongly suggesting that it should be a sister taxon to Prochloron. Therefore, we proposed this symbiotic cyanobacterium as a novel species under the new genus Candidatus Paraprochloron terpiosi. Comparative genomic analyses revealed that ‘Paraprochloron’ and Prochloron exhibit distinct genomic features and DNA replication machinery. We also characterized the metabolic potentials of ‘Paraprochloron terpiosi’ in carbon and nitrogen cycling and propose a model for interactions between it and T. hoshinota. This study builds a foundation for the study of the T. hoshinota microbiome and paves the way for better understanding of ecosystems involving this coral-killing sponge.     

CYTOCHEMICAL STUDIES ON PROCHLOROPHYTES: LOCALIZATION OF DNA AND RIBULOSE 1,5-BISPHOSPHATE CARBOXYLASE-OXYGENASE1

We studied the distribution of the DNA-containing region and the ribulose 1,5-bisphosphate carboxylase-oxygenase (RuBisCo) content of polyhedral bodies in three different prochlorophyte cell types in a search for broad evolutionary affinities of these chlorophyll b-containing prokaryotes. DNA was localized by DAPI staining and electron microscopy utilizing monoclonal anti-DNA antibody 2C-10 plus a secondary antibody labeled with colloidal gold. Antibodies against the large RuBisCo subunit from a higher plant raised in rabbits were used to localize RuBisCo in polyhedral bodies. We studied Prochloron Lewin cells from two different didemnid ascidian hosts (Lissoclinum patella and Didemnum molle) collected in Palau, West Caroline Islands, and cells of Prochlorothrix hollandica Burger-Wiersma, Stal, and Mur grown in laboratory culture. Cells of the blue-green alga Anabaena 7120 were studied for comparison. The DNA distribution was markedly different in the two Prochloron cell types. The thylakoids in cells from L. patella were concentrically arranged around a large central vacuole; the DNA-containing stromal areas appeared in thin sections as a concentric arcs between the thylakoid stacks. The central vacuole was lacking in cells from D. molle, and the thylakoid stacks and strands of DNA-containing stroma showed a more haphazard arrangement. In the filamentous Prochlorothrix the DNA-containing stroma was largely limited to a central nucleoid structure running the length of the cell. Although the DNA arrangements in Prochloron might be considered “chloroplast-like” since DNA-containing stroma is distributed, as in chloroplasts, in scattered sites among photosynthetic membranes, this is not so in Prochlorothrix, where there is an axial nucleoid, as in many filamentous cyanobacteria. Our anti-RuBisCo antibodies were selectively bound to the polyhedral bodies of all three cell types, indicating that Prochloron and Prochlorothrix, like many other autotrophic prokaryotes, possess typical carboxysomes.     

Genotypic relationships between Prochloron samples from different localities and hosts as determined by DNA-DNA reassociations

Genotypic relationships between seven Prochloron samples isolated from different didemnid ascidian hosts collected at the Palau archipelago and Munda (Solomon Islands) and one cyanobacterial (Synechocystis) strain were determined by DNA-DNA reassociations. Thermal stability values of DNA-DNA hybrids indicate that all Prochloron samples involved are mutually very closely related and only slightly related with the Synechocystis strain. It is concluded that the Prochloron samples tested are representatives of one and the same species.     

Photosynthesis of Prochloron as Affected by Environmental Factors

The effects of light intensity, pH, temperature, and UV irradiation on the photosynthetic rate of Prochloron isolated from the ascidian host Lissoclinum patella, collected from Palau, were examined. Photosynthesis increased with light intensity with saturation at 500 μmol/m² per second. It was maximum at pH 8 to 9 but almost completely suppressed below pH 7. The optimum temperature was 35° to 40°C, but the photosynthesis was absent at ≤20°C and at 45°C. It was recovered when the symbiont was transferred from 1 hour of incubation at ≤20°C to 35°C but not when transferred from incubation at 45°C. Ultraviolet irradiation severely inhibited the photosynthesis of Prochloron in isolation but not in vivo. This protection was brought about by the tunic covering the ascidian colony, which contains UV-absorbing mycosporine-like amino acids. These results indicate that the characteristic condition of the tropical marine environment largely determines the ecological distribution of Prochloron, and the ascidian tunic protects the organism from UV radiation. Received February 17, 2000; accepted August 8, 2000.     

Nutritional diversity of symbiotic ascidians in a Fijian seagrass meadow

In a coastal lagoon of Dravuni Island, Fiji, at least six species of compound ascidians, some of them harboringProchloron as symbiotic algae, were found in aSyringodium-dominated seagrass meadow. Based on their heterotrophic (filrer feeding rates) and autotrophic (photosynthetic) activities, carbon gain of the ascidians was categorized into two groups: (i) supported by heterotrophic metabolism; and (ii) supported by both heterotrophic and autotrophic metabolisms.Didemnum molle, Lissoclinum bistratum andLissolinum voeltzkowi belong to the latter group, and the relative contribution of the autotrophic process was a significant portion of their carbon gain (52–74%). These symbiotic ascidians were found in light microhabitats, while the heterotrophic species occupied shady environments rich in suspended organic materials, such as the sheath surface of the seagrass.     

Deoxyribonucleic acid base composition and genome size of Prochloron

The DNA base composition of the photosynthetic prokaryote Prochloron was determined (on samples collected from the natural environment) to be 40.8 mol% GC. The sharp differential melting curve indicated the absence of significant quantities of contaminating DNA from other organisms. The genome size, estimated from the renaturation kinetics of thermally denatured DNA, was 3.59×10⁹ daltons mol. wt, similar to that of many other prokaryotes. The fact that Prochloron has not yet been cultured in the laboratory cannot, therefore, be attributed to a reduced genetic information content.     

Cladistic analysis of blue-green procaryote interrelationships and chloroplast origin based on 16S rRNA oligonucleotide catalogues

Oligonucleotide catalogues from 16S rRNA have been a major source of information for phylogenetic reconstruction among procaryotes. Several large procaryote groups have been analyzed and phylogenies presented. Catalogues are also available for many chloroplasts. The hypotheses of phylogeny are derived mainly from similarity (phenetic) comparisons of the catalogues and the extent of the homoplasy (parallelisms and reversals) involved has not been estimated properly. Although catalogues are currently being superseded by complete sequence data, an evaluation of the strength of catalogue data, and hence of the strength of the extensive phylogenetic hypotheses derived from them, is in order. Cladistic analysis of 16S rRNA oligonucleotide catalogues from three blue-green procaryotes, Prochloron, and chloroplasts of a red alga, Euglena, a green alga and two flowering plants shows that there is extensive homoplasy in the catalogues and several phylogenetic trees are possible. The corresponding consensus trees indicate that little or nothing can be said about interrelationships and chloroplast origin on the basis of these particular catalogues, except that Prochloron may be more closely related to the blue-greens than to chloroplasts.     

Partial sequence of ribulose-1,5-bisphosphate carboxylase/oxygenase and the phylogeny of Prochloron and Prochlorococcus (Prochlorales)

The prochlorophytes, oxygenic photosynthetic prokaryotes having no phycobiliprotein but possessing chlorophylls a and b, have been proposed to have a common ancestry with green chloroplasts, yet this is still controversal. We report here that partial sequence comparisons of the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase, including sequence data from two prochlorophytes, Prochlorococcus and Prochloron, indicate that Prochlorococcus is more closely related to a photosynthetic bacterium, Chromatium vinosum (-purple bacteria), than to cyanobacteria, while Prochloron is closely related to the prochlorophyte Prochlorothrix and to cyanobacteria. The molecular phylogenetic tree indicates that a common ancestor of Prochlorococcus and -purple bacteria branched off from the land plant lineage earlier than Prochloron, Prochlorothrix, and cyanobacteria.Correspondence to: A. Shimada     

Contents of ultraviolet-absorbing substances in two color morphs of the photosymbiotic ascidian Didemnum molle

The contents of mycosporine-like amino acids (MAAs) were compared in the two color morphs (dark-gray and brown colonies) of the tropical ascidian Didemnum molle (Herdman, 1886), which harbors the photosymbiotic prokaryote Prochloron. The colonies of each color morph were exclusively distributed in shallow reef lagoons at the different sites. Spectroscopic and chromatographic analyses showed that the Prochloron cell density and MAA concentration in the dark-gray colonies were an estimated 1.4 and 2.4 times higher, respectively, than in the brown colonies. The significant difference in MAA contents between the color morphs was primarily due to the difference in shinorine contents (p < 0.01, Mann–Whitney U-test). The high concentration of MAAs in the dark-gray colonies may provide better conditions for Prochloron cells, compared to the brown colonies with lower MAA concentrations.     

Compounds produced from potential tunicate-blue-green algal symbiosis: A review

Tunicates of the family Didemnidae can be host to two genera of blue-green algae:Synechocystis andProchloron. The presence of symbiotic algae raises questions as to the exact origin of the biologically important metabolites which have been isolated from tunicates in recent years. Is the compound produced by the tunicate, the alga, or through a combined effort of both organisms? Although this question cannot be fully addressed at the present time, there is evidence which supports the argument that the metabolic origin of certain ascidian metabolites resides in the alga, or is due to a collaborative effort of both organisms. The purpose of this review is to present compounds isolated from tunicates that possess a likely symbiotic relationship with eitherSynechocystis orProchloron. Attention will also be given to the ecology of the organisms and the biological activities of metabolites isolated.     

DNA-DNA REASSOCIATION STUDIES WITH DNA FROM PROCHLORON (PROCHLOROPHYTA) SAMPLES OF INDO-WEST PACIFIC ORIGIN1

Seven Indo-West Pacific Prochloron Lewin samples and one Caribbean cyanobacterial sample were compared by DNA-DNA reassociation experiments. The quality and quantity of extracted DNA were affected by several cell preservation methods and DNA extraction and purification procedures. We conclude that Prochloron samples from six different hosts and three different geographical locations spanning ca. 9000 km belong to a single species but are not closely related to the strain of cyanohacteria used. The intraspecific genotypic variation within Prochloron is discussed with respect to host specificity and geographical distribution.     

Muramic acid in the cell walls of Prochloron

Muramic acid has been detected in Prochloron with the aid of two different techniques. It was assayed by cleaving D-lactate from muramic acid and then reducing NAD with D-lactate dehydrogenase and measuring the NADH with bacterial luciferase. Gas-liquid chromatography of trimethylsilyl derivatives of cell extracts confirmed that muramic acid was present in about the quantity given by the D-lactate assay. The amount of muramic acid present was 1.7±0.2 g/mg dry weight or 1.3fg/m² of cell surface. This suggests that the thickness of the peptidoglycan layer in Prochloron is similar to that in blue-green algae.Abbreviations D-LDH d-lactate dehydrogenase - MA muramic acid - TMS trimethylsilyl - TLE thin layer electrophoresis - GLC gas-liquid chromatography     

16S rRNA及rpoC1基因用于螺旋藻、节旋藻系统发育的比较北大核心CSCD

【目的】利用16S rRNA和rpoC1基因分子标记研究螺旋藻、节旋藻的系统发育关系,并对其区分能力进行比较。【方法】以84株螺旋藻、节旋藻为研究对象,对其进行16S rRNA、rpoC1基因序列的扩增、测序及分析,并对构建的系统发育树进行对比。【结果】rpoC1基因序列保守位点所占比例49.7%、平均G+C百分含量47.7%和序列相似度76%–100%明显低于16S rRNA基因序列的79.4%、55.6%和91%–100%,其变异程度高于16S rRNA基因;基于16S rRNA、rpoC1基因构建的系统发育NJ树拓扑结构基本一致,84株实验藻株分为2个属3个类群,其中仅F-351、F-904-2、F-1070和TJBC14-1藻株为螺旋藻,其余均为节旋藻;虽然2个基因都不能区分形态种和地理种,但rpoC1基因NJ树的置信度(100%)高于16S rRNA基因(99%),属内分群效果也明显优于16S rRNA基因。【结论】支持了螺旋藻、节旋藻为两个不同属的结论,且在属内分类时rpoC1基因比16S rRNA基因具有更高的区分度。     

Microbial diversity of biofilm communities in microniches associated with the didemnid ascidian Lissoclinum patella

We assessed the microbial diversity and microenvironmental niche characteristics in the didemnid ascidian Lissoclinum patella using 16S rRNA gene sequencing, microsensor and imaging techniques. L. patella harbors three distinct microbial communities spatially separated by few millimeters of tunic tissue: (i) a biofilm on its upper surface exposed to high irradiance and O₂ levels, (ii) a cloacal cavity dominated by the prochlorophyte Prochloron spp. characterized by strong depletion of visible light and a dynamic chemical microenvironment ranging from hyperoxia in light to anoxia in darkness and (iii) a biofilm covering the underside of the animal, where light is depleted of visible wavelengths and enriched in near-infrared radiation (NIR). Variable chlorophyll fluorescence imaging demonstrated photosynthetic activity, and hyperspectral imaging revealed a diversity of photopigments in all microhabitats. Amplicon sequencing revealed the dominance of cyanobacteria in all three layers. Sequences representing the chlorophyll d containing cyanobacterium Acaryochloris marina and anoxygenic phototrophs were abundant on the underside of the ascidian in shallow waters but declined in deeper waters. This depth dependency was supported by a negative correlation between A. marina abundance and collection depth, explained by the increased attenuation of NIR as a function of water depth. The combination of microenvironmental analysis and fine-scale sampling techniques used in this investigation gives valuable first insights into the distribution, abundance and diversity of bacterial communities associated with tropical ascidians. In particular, we show that microenvironments and microbial diversity can vary significantly over scales of a few millimeters in such habitats; which is information easily lost by bulk sampling.     

Prochlorophyta – a matter of class distinctions

Prochloron (a marine symbiont) and Prochlorothrix (from freshwater plankton) contain chlorophylls a and b; Prochlorococcus (common in marine picoplankton) contains divinyl-chlorophylls a and b. Like cyanophytes they are all clearly photosynthetic prokaryotes, but since they contain no blue or red bilin pigment they were assigned to a new algal sub-class, the Prochlorophyta. However, since their possible phylogenetic relationships to ancestral green-plant chloroplasts have not received support from molecular biology, it now seems expedient to consider them as aberrant cyanophytes. This revised version was published online in June 2006 with corrections to the Cover Date.