BIOCHEMICAL PHENOTYPES CORRESPONDING TO MOLECULAR PHYLOGENY OF THE RED ALGAE PLOCAMIUM (PLOCAMIALES,RHODOPHYTA): IMPLICATIONS OF INCONGRUENCE WITH THE CONVENTIONAL TAXONOMY1

Among five species of the genus Plocamium Lamouroux distributed around Japan, P. cartilagineum (Linnaeus) Dixon, P. recurvatum Okamura and P. telfairiae (Hooker and Harvey) Harvey are often difficult to distinguish morphologically from each other. Our previous study demonstrated that P. recurvatum and P. telfairiae were divided into two groups, A and C, based on RUBISCO spacer sequence and that the specimens belonging to group C had acidic cell saps. In this study, we inferred evolutionary relationships of these Plocamium species from internal transcribed spacer sequence of the ribosomal RNA genes and obtained a similar topology to the RUBISCO spacer tree. Color of the dried specimens in the acidic group C was darker red than that in the non‐acidic group A, although there was no difference in color in living thalli. The Br^? concentration in the cell sap of the acidic group C was 20 times higher than that of the non‐acidic group. We could not find any morphological differences to distinguish clearly between groups A and C despite exhaustive investigation of field‐collected and cultured thalli in both P. recurvatum and P. telfairiae. These results suggest that the color of dried specimens and the composition of intracellular inorganic ions are significant criteria for interpreting phylogenetic relationships in Japanese Plocamium spp.     

THE USE OF PLASTID DNA RESTRICTION ENDONUCLEASE PATTERNS IN DELINEATING RED ALGAL SPECIES AND POPULATIONS1

Plastid DNA band patterns generated by electrophoresis of endonuclease digests demonstrate remarkable conservation of DNA sequences at the species and subspecies level in flowering plants. Generally, patterns are identical or near-identical from different populations belonging to the same species. This methodology has now been applied to red algae to ascertain its value in systematic studies. Plastid DNA from nine bangiophycean and florideophycean red algae was isolated and cut with restriction endonucleases that recognize different 6-base pair sequences. The patterns generated upon the electrophoretic separation of digestion fragments show that within a species patterns are identical, but not within higher taxa. The proper identification of one Gracilaria population of uncertain taxonomic affinity was clearly established by this method of plastid DNA analysis. Differences between species in plastid DNA sequences were confirmed by probing blots of restriction fragments with known gene sequences. A number of heterologous plastid DNA probes were found to be sufficiently homologous to be useful in studying red algal DNA. Unexpectedly, supercoiled circular plasmids ranging in size from ca. 1.5–8 kb were found in some red algal species but not in others. The position of these plasmids in agarose gels following electrophoresis is uniform within a species but differs between different species of the same genus, contributing further patterns for taxonomic analysis.     

Identification and in vitro anti-esophageal cancer activity of a series of halogenated monoterpenes isolated from the South African seaweeds Plocamium suhrii and Plocamium cornutum

Five known (1, 2, 4, 6 and 7) halogenated monoterpenes together with 1Z,3R^∗,4S^∗,5E,7Z)-1-bromo-3,4,8-trichloro-7-(dichloromethyl)-3-methylocta-1,5,7-triene (3) and (3R^∗,4S^∗)-3,4,6,7-tetrachloro-3,7-dimethyl-octen-1-ene (5) were isolated from the red macroalga Plocamium suhrii and their structures deduced from their spectroscopic data. The seven compounds from P. suhrii together with five related compounds from Plocamium cornutum have been evaluated for their cytotoxic effects on an esophageal cancer cell line (WHCO1). Compounds 1-6 showed greater cytotoxicity in this assay as compared to the known anticancer drug cisplatin.     

Sinking movements of phytoplankton indicated by a simple trapping method

Recoveries of planktonic algae in a vertical series of small traps are analysed in relation to changes in the standing populations and to the physical and chemical characteristics of a shallow, stratified lake. Many motile or buoyant species were trapped predominantly within restricted depth ranges. The results are interpreted as representing the vertical ranges within which the species were directing their downward movements. Ceratium hirundinella, a species showing more wide-ranging movements, was trapped throughout the epilimnion. Non-motile diatoms were caught at all depths. Deduced activity ranges are similar to vertical distributions observed in earlier years and are invoked to support the view that, in a stratified lake, typical movements may play an important role in algal growth and succession.     

Mevalonosomes: specific vacuoles containing the mevalonate pathway in Plocamium brasiliense cortical cells (Rhodophyta)

This paper has identified, for the first time in a member of the Rhodophyta, a vacuolar organelle containing enzymes that are involved in the mevalonate pathway—an important step in red algal isoprenoid biosynthesis. These organelles were named mevalonosomes (Mev) and were found in the cortical cells (CC) of Plocamium brasiliense, a marine macroalgae that synthesizes several halogenated monoterpenes. P. brasiliense specimens were submitted to a cytochemical analysis of the activity of the 3‐hydroxy‐3‐methylglutaryl‐CoA synthase (HMGS). Using transmission electron microscopy (TEM), we confirmed the presence of HMGS activity within the Mev. Because HMGS is necessary for the biosynthesis of halogenated monoterpenes, we isolated a hexanic fraction (HF) rich in halogenated monoterpenes from P. brasiliense that contained a pentachlorinated monoterpene as a major metabolite. Because terpenes are often related to chemical defense, the antifouling (AF) activity of pentachlorinated monoterpene was tested. We found that the settlement of the mussel Perna perna was reduced by HF treatment (2.25 times less than control; 40% and 90% of fouled surface, respectively; P = 0.001; F_9,9 = 1.13). The HF (at 10 μg · mL^?1) also inhibited three species of fouling microalgae (Chlorarachnion reptans, Cylindrotheca cloisterium, and Exanthemachrysis gayraliae), while at a higher concentration (50 μg · mL^?1), it inhibited the bacteria Halomonas marina, Polaribacter irgensii, Pseudoalteromonas elyakovii, Shewanella putrefaciens, and Vibrio aestuarianus. The AF activity of P. brasiliense halogenated monoterpenes and the localization of HMGS activity inside Mev suggest that this cellular structure found in CC may play a role in thallus protection against biofouling.     

Evidence for positive selection in phycoerythrin genes of red algae and cyanobacteria <Emphasis Type="Italic">Prochlorococcus</Emphasis> and <Emphasis Type="Italic">Synechococcus</Emphasis>

Because of the shortage of phycoerythrin (PE) gene sequences from rhodophytes, peBA encoding - and -subunits of PE from three species of red algae (Ceramium boydenn, Halymenia sinensis, and Plocamium telfariae) were cloned and sequenced. Different selection forces have affected the evolution of PE lineages. 8.9 % of the codons were subject to positive selection within the PE lineages (excluding high-irradiance adapted Prochlorococcus). More than 40 % of the sites may be under positive selection, and nearly 20 % sites are weakly constraint sites in high-irradiance adapted Prochlorococcus. Sites most likely undergoing positive selection were found in the chromophore binding domains, suggesting that these sites have played important roles in environmental adaptation during PE diversification. Moreover, the heterogeneous distribution of positively selected sites along the PE gene was revealed from the comparison of low-irradiance adapted Prochlorococcus and marine Synechococcus, which firmly suggests that evolutionary patterns of PEs in these two lineages are significantly different.     

DIFFERENT CHEMICAL TYPES OF PLOCAMIUM VIOLACEUM (RHODOPHYTA) FROM THE MONTEREY BAY REGION,CALIFORNIA1

Differences in the halogenated natural products of Plocamium violaceum Farlow can be noted from different sites in the Monterey Bay region, California. Specimens from the northern region produce a set of alicyclic halogenated monoterpenes which are not synthesized by those from the southern region. Instead, the latter elaborates a set of acyclic halogenated monoterpenes not observed from the former. This situation is invariant regardless of life history forms or different seasons. Similarly, the composition of halogenated monoterpene structures from P. cartilagineum (L.) Dixon varies at different locations. From the data presented herein we conclude that at least two chemical types can be identified for P. violaceum based upon differences in their halogenated products. Thus we suggest that the north Monterey form can be designated as chemotype α and the southern one as chemotype β.     

Plocoralides A-C, polyhalogenated monoterpenes from the marine alga Plocamium corallorhiza

Three new polyhalogenated monoterpenes, plocoralides A-C (1-3) along with three known compounds (4-6) have been isolated from the organic extract of the red alga P. corallorhiza. Structures of the new compounds were characterized as 4,8-dibromo-1,1-dichloro-3,7-dimethyl-2E,6E-octadiene (1), 4,6-dibromo-1,1-dichloro-3,7-dimethyl-2E,7-octadiene (2) and 4,8-dibromo-1,1,7-trichloro-3,7-dimethyl-2E,5Z-octadiene (3) on the basis of one- and two-dimensional NMR spectroscopic data and MS analyses. Compounds 2-6 show moderate cytotoxicity toward esophageal cancer cells.     

Antiplasmodial halogenated monoterpenes from the marine red alga Plocamium cornutum

In our continuing search for antimalarial leads from South African marine organisms we have examined the antiplasmodial organic extracts of the endemic marine red alga Plocamium cornutum (Turner) Harvey. Two new and three known halogenated monoterpenes were isolated and their structures determined by standard spectroscopic techniques. The 3,7-dimethyl-3,4-dichloro-octa-1,5,7-triene skeleton is common to all five compounds. Interestingly, compounds bearing the 7-dichloromethyl substituent showed significantly higher antiplasmodial activity toward a chloroquine sensitive strain of Plasmodium falciparum.