Mauricio Rodriguez-Lanetty

Transcriptome analysis of a cnidarian-dinoflagellate mutualism reveals complex modulation of host gene expression

BackgroundCnidarian – dinoflagellate intracellular symbioses are one of the most important mutualisms in the marine environment. They form the trophic and structural foundation of coral reef ecosystems, and have played a key role in the evolutionary radiation and biodiversity of cnidarian species. Despite the prevalence of these symbioses, we still know very little about the molecular modulators that initiate, regulate, and maintain the interaction between these two different biological entities. In this study, we conducted a comparative host anemone transcriptome analysis using a cDNA microarray platform to identify genes involved in cnidarian – algal symbiosis.ResultsWe detected statistically significant differences in host gene expression profiles between sea anemones (Anthopleura elegantissima) in a symbiotic and non-symbiotic state. The group of genes, whose expression is altered, is diverse, suggesting that the molecular regulation of the symbiosis is governed by changes in multiple cellular processes. In the context of cnidarian – dinoflagellate symbioses, we discuss pivotal host gene expression changes involved in lipid metabolism, cell adhesion, cell proliferation, apoptosis, and oxidative stress.ConclusionOur data do not support the existence of symbiosis-specific genes involved in controlling and regulating the symbiosis. Instead, it appears that the symbiosis is maintained by altering expression of existing genes involved in vital cellular processes. Specifically, the finding of key genes involved in cell cycle progression and apoptosis have led us to hypothesize that a suppression of apoptosis, together with a deregulation of the host cell cycle, create a platform that might be necessary for symbiont and/or symbiont-containing host cell survival. This first comprehensive molecular examination of the cnidarian – dinoflagellate associations provides critical insights into the maintenance and regulation of the symbiosis.

Resistance to thermal stress in corals without changes in symbiont composition

Discovering how corals can adjust their thermal sensitivity in the context of global climate change is important in understanding the long-term persistence of coral reefs. In this study, we showed that short-term preconditioning to higher temperatures, 3°C below the experimentally determined bleaching threshold, for a period of 10 days provides thermal tolerance for the symbiosis stability between the scleractinian coral, Acropora millepora and Symbiodinium. Based on genotypic analysis, our results indicate that the acclimatization of this coral species to thermal stress does not come down to simple changes in Symbiodinium and/or the bacterial communities that associate with reef-building corals. This suggests that the physiological plasticity of the host and/or symbiotic components appears to play an important role in responding to ocean warming. The further study of host and symbiont physiology, both of Symbiodinium and prokaryotes, is of paramount importance in the context of global climate change, as mechanisms for rapid holobiont acclimatization will become increasingly important to the long-standing persistence of coral reefs.

The phylogeography and connectivity of the latitudinally widespread scleractinian coral Plesiastrea versipora in the Western Pacific

Whereas terrestrial animal populations might show genetic connectivity within a continent, marine species, such as hermatypic corals, may have connectivity stretching to all corners of the planet. We quantified the genetic variability within and among populations of the widespread scleractinian coral, Plesiastrea versipora along the eastern Australian seaboard (4145 km) and the Ryukyu Archipelago (Japan, 681 km) using sequences of internal transcribed spacers (ITS1‐2) from ribosomal DNA. Geographic patterns in genetic variability were deduced from a nested clade analysis (NCA) performed on a parsimony network haplotype. This analysis allowed the establishment of geographical associations in the distribution of haplotypes within the network cladogram, therefore allowing us to deduce phylogeographical patterns based under models of restricted gene flow, fragmentation and range expansion. No significant structure was found among Ryukyu Archipelago populations. The lack of an association between the positions of haplotypes in the cladogram with geographical location of these populations may be accounted for by a high level of gene flow of P. versipora within this region, probably due to the strong Kuroshio Current. In contrast, strong geographical associations were apparent among populations of P. versipora along the south‐east coast of Australia. This pattern of restricted genetic connectivity among populations of P. versipora on the eastern seaboard of Australia seems to be associated with the present surface ocean current (the East Australian Current) on this side of the south‐western Pacific Ocean.

Evolving lineages of Symbiodinium-like dinoflagellates based on ITS1 rDNA

Symbiodinium-like dinoflagellates have been shown to be a diverse group of endosymbionts that associate mutualistically with many kinds of coral reef dwellers, including cnidarians, molluscs, and protists. A high number of genetically ITS types of symbionts have been reported to date. However, whether these recently identified Symbiodinium ITS types indeed represent independent evolutionary lineages is still unsettled. Here I tested the null hypothesis that certain group of symbionts sampled from different geographical locations are derived from a single evolutionary lineage using a nested clade analysis (NCA). I analyzed a total of 174 ITS1 sequences from GenBank and pooled them into 74 ITS1 distinct haplotypes. Using these haplotypes, the statistical parsimony criterion produced 23 independent network trees, each one corresponding to a genetically independent evolving lineage. Some of these lineages revealed certain degree of specificity with some host groups at least at the phylum level. Within the previously described 28S-rDNA phylotype A, five ITS1 lineages were resolved. Phylotypes B and C resolved each in two ITS1 lineages. The highest ITS1 symbiont diversity was observed within the phylotype F, in which 11 lineages were resolved. Moreover, most of these lineages were associated uniquely with protist hosts from the group of foraminiferans. Here it is suggested that this high genetic diversity of endosymbionts associated with foraminiferans is linked with the evolution of soritacean foraminifera, which seems to have been driven by endosymbiosis. Lastly, the absence of genetic recombination presented in this study, suggest a lack of hybridisation at least among the major 28S-rDNA phylotypes within Symbiodinium-like dinoflagellates. This supports highly the idea that these phylotypes are indeed independent evolutionary units, which should be considered at least as different species. Whether they belong to the same genus or to different higher taxa still needs to be revised.

Comparative analysis of superfamilies of NBS-encoding disease resistance gene analogs in cultivated and wild apple species

Eleven distinct families of resistance gene analogs (RGAs) with the characteristic nucleotide-binding sequence (NBS) were identified in two wild apple species, Malus prunifolia and M. baccata, and two cultivated apple cultivars, M. domestica cv. Fuji and M. domestica cv. Hong-ok, using PCR approaches with degenerate primers based on two conserved motifs of known NBS-LRR resistance genes. These RGA families were found to be represented in all the apple species tested, including wild and cultivated species. However, their sequences are very divergent from each other. Furthermore, the low level of recombination detected within their RGA families supports the idea that the evolution of NBS-encoding sequences in apple species involves the gradual accumulation of mutations. Despite the high diversity of the RGA families found in all apple species, the apparent lack of differentiation between wild and cultivated forms suggests that other factors, such as the capacity to tolerate pathogens, might play an important role in the survival of wild-type species.

Emergent effects of heavy metal pollution at a population level: Littorina brevicula a study case

Changes in genetic variability and allele frequency can be responses from natural populations when encountering a novel contaminated environment. The genetic diversity and population structuring of natural populations of the gastropod Littorina brevicula from heavy-metal polluted and unpolluted environments along the southeast coast of Korea were examined using two mtDNA markers, cyt b and ND6. This study applied a nested clade analysis to test the existence of structuring association of haplotype distribution with environments (polluted and unpolluted). No genetic differences within cyt b mtDNA were detected between environments. On the other hand, differences in population haplotype diversity and structuring were found within ND6 mtDNA between polluted and unpolluted environments. The ND6-mtDNA haplotype (=genetic) diversity was significant lower in polluted environments. This decreased genetic diversity along with differences in the haplotype distribution within heavy-metal polluted environments compared to those unpolluted ones stand out as emergent effects from pollution at a population level. In this study, we propose the use of different approaches, such as the NCA, that takes into account the rare haplotypes, when assessing the effects of pollution on population genetic structuring.

Isolation of Symbiosomes and The Symbiosome Membrane Complex from The Zoanthid Zoanthus Robustus

A. Kazandjian, V.A. Shepherd, M. Rodriguez-Lanetty, W. Nordemeier, A.W.D. Larkum and R.G. Quinnell. 2008. Isolation of symbiosomes and the symbiosome membrane complex from the zoanthid Zoanthus robustus. Phycologia 47: 294–306. DOI: 10.2216/07-23.1 The zoanthid Zoanthus robustus was used as a model organism to develop procedures for isolating pure symbiosomes and symbiosome membranes. The symbiosome is comprised of a zooxanthella (Symbiodinium sp.) cell that divides rarely and is separated from the host gastrodermal cytoplasm by a symbiosome multimembrane complex. Devising a method to isolate membranes at the interface between the symbiotic partners is a critical first step in characterising the molecular components involved in the metabolic trafficking necessary to sustain an effective symbiosis. After zoanthid gastrodermal cells were extracted, symbiosomes were released by mechanical disruption, recovered by centrifugation, and then purified using discontinuous sucrose gradient centrifugation. The material forming the membrane complex around symbiosomes proved highly resistant to disruption. Methods used to dissociate this interface from symbionts included (1) Triton X-100 detergent solubilisation, (2) osmotic shock with mechanical disruption, and (3) vigorous mechanical disruptions, where powerful shearing forces were used, combined with a series of sucrose density gradient centrifugation steps. The lipophilic styryl fluorochrome FM 1-43, at a concentration of 30 µM, selectively labelled the symbiosome membrane complex, both for isolated symbiosomes and those in hospite. Other cell membranes, including plasma membranes, endoplasmic reticulum, tonoplast, and organelle membranes, were not visibly labelled at this concentration. The selective labelling of the symbiosome membrane complex remained stable even after long exposure times (3 h). At 30 µM concentration, FM 1-43 also labelled symbiosome membrane fragments isolated using methods (1), (2) and (3). Method (3) proved to be the most effective in producing a fraction enriched in FM-143-labelled membrane material, which we call a symbiosome membrane complex. Transmission electron microscopy, together with confocal and conventional epifluorescence microscopy of the FM 1-43-stained preparations, was used to validate the purity of symbiosome preparations and to infer the complexity of the symbiosome membrane complex. This membrane complex has regions where the membranes contributed by the alga are appressed, and punctate regions whose function remains unclear.

Genetic population structure of Littorina brevicula around Korean waters

Because Korean waters are influenced by different minor current systems, this may lead us to expect certain genetic differentiation among populations of marine invertebrates around Korean coastlines. To test this idea, we examined the genetic differentiation of 14 populations of a widely distributed marine gastropod, Littorina brevicula, using mtDNA markers, cyt b and ND6. Sequencing analyses of 500 bp fragment of the cyt b and 501 bp of the ND6 mtDNA revealed the existence of 14 and 24 haplotypes, respectively. Geographic association with population genetic variability was tested using a nested clade analysis (NCA). However, no significant association between geography and haplotype distribution was found, which suggests the existence of a high level of gene flow of L. brevicula around Korean waters. The lack of genetic structure and likely high gene flow among these populations, even under influences of different minor current systems, may be accounted for long periods of dispersal of the planktonic larvae from this gastropod species within major currents on the north western Pacific Ocean.

Transport of symbiotic zooxanthellae in mesogleal canals of Zoanthus robustus

In symbiotic dinoflagellate–cnidarian associations, regulation of algal numbers is an essential feature during steady state conditions (Reimer 1971; Falkowski et al. 1993). During mass bleaching events there is massive loss of zooxanthellae due to processes that are poorly understood (Gates et al. 1992; Jones and Yellowlees 1997). Under normal environmental conditions, the symbiotic algal population is maintained by the lowlevel expulsion of algal cells from the host coelenteron (=gastrovascular cavity); this low-level expulsion is thought to balance increases in endosymbiont numbers that occur from algal division, thereby maintaining a constant algal density (Hoegh-Guldberg et al. 1987). Although symbiotic algae normally exist only in endodermal cells, here we describe the presence of zooxanthellae within canals which are distributed within the host’s mesoglea and which connect to mesenteries. This finding implies that some zooxanthellae discharged from host endodermal cells are transported through mesogleal ducts to the inner cavity of mesenteries, from where they are finally degraded and/or expelled into the coelenteron. Most zooxanthellae reside within the endodermal host cells but in certain cnidarian species zooxanthellae are also found within the ectodermal and mesogleal layers (Hyman 1940), which, always, at least in theory, are separated from the coelenteron. After zooxanthellae are released, either by exocytosis or through whole host cell detachment (Gates et al. 1992) they become submerged in a three-dimensional matrix of host tissue, which is relatively distant from the coelenteron, which is the site from where they will be ejected finally through the host pharynx and oral aperture. Some studies have suggested that zooxanthellae from scleractinian hosts are normally extruded from the gastrodermis (endodermal cells) and accumulated directly into the coelenteron (Titlyanov et al. 1996). However, this direct algal cell discharge from the coelenteron might happen only in endodermal cells lining the coelenteron, but not from those located in the inner cell layers of the host tissue (Fig. 1a). There is some other work (Yonge 1966; Reimer 1971; Trench 1974) that has shown that some released zooxanthellae are ejected into the coelenteron from mesenterial cavities, in which partial algal degradation seems to take place (Trench 1974; Titlyanov et al. 1996). Algae then amass on the lateral lobes of the mesenterial filaments, which are rolled and finally extruded from the body as a mass called a ‘‘zooxanthellae body or pellet’’ (Yonge 1966; Reimer 1971). However, how the zooxanthellae, that are released from the inner host tissue, reach the mesenteries was unclear.