Cross-shelf investigation of coral reef cryptic benthic organisms reveals diversity patterns of the hidden majority.

Coral reefs harbor diverse assemblages of organisms yet the majority of this diversity is hidden within the three dimensional structure of the reef and neglected using standard visual surveys. This study uses Autonomous Reef Monitoring Structures (ARMS) and amplicon sequencing methodologies, targeting mitochondrial cytochrome oxidase I and 18S rRNA genes, to investigate changes in the cryptic reef biodiversity. ARMS, deployed at 11 sites across a near- to off-shore gradient in the Red Sea were dominated by Porifera (sessile fraction), Arthropoda and Annelida (mobile fractions). The two primer sets detected different taxa lists, but patterns in community composition and structure were similar. While the microhabitat of the ARMS deployment affected the community structure, a clear cross-shelf gradient was observed for all fractions investigated. The partitioning of beta-diversity revealed that replacement (i.e. the substitution of species) made the highest contribution with richness playing a smaller role. Hence, different reef habitats across the shelf are relevant to regional diversity, as they harbor different communities, a result with clear implications for the design of Marine Protected Areas. ARMS can be vital tools to assess biodiversity patterns in the generally neglected but species-rich cryptic benthos, providing invaluable information for the management and conservation of hard-bottomed habitats over local and global scales.

Molecular characterization reveals the complexity of previously overlooked coral-exosymbiont interactions and the implications for coral-guild ecology.

Several obligate associate crabs and shrimps species may co-occur and interact within a single coral host, leading to patterns of associations that can provide essential ecological services. However, knowledge of the dynamics of interactions in this system is limited, partly because identifying species involved in the network remains challenging. In this study, we assessed the diversity of the decapods involved in exosymbiotic assemblages for juvenile and adult Pocillopora damicornis types α and ß on reefs of New Caledonia and Reunion Island. This approach revealed complex patterns of association at regional and local scales with a prevalence of assemblages involving crab-shrimp partnerships. Furthermore, the distinction of two lineages in the snapping shrimp Alpheus lottini complex, rarely recognized in ecological studies, reveals a key role for cryptic diversity in structuring communities of mutualists. The existence of partnerships between species that occurred more commonly than expected by chance suggests an increased advantage for the host or a better adaptation of associated species to local environmental conditions. The consideration of cryptic diversity helps to accurately describe the complexity of interaction webs for diverse systems such as coral reefs, as well as the functional roles of dominant associated species for the persistence of coral populations.

Ecological speciation in anemone-associated snapping shrimps (Alpheus armatus species complex).

Divergent natural selection driven by competition for limited resources can promote speciation, even in the presence of gene flow. Reproductive isolation is more likely to result from divergent selection when the partitioned resource is closely linked to mating. Obligate symbiosis and host fidelity (mating on or near the host) can provide this link, creating ideal conditions for speciation in the absence of physical barriers to dispersal. Symbiotic organisms often experience competition for hosts, and host fidelity ensures that divergent selection for a specific host or host habitat can lead to speciation and strengthen pre-existing reproductive barriers. Here, we present evidence that diversification of a sympatric species complex occurred despite the potential for gene flow and that partitioning of host resources (both by species and by host habitat) has contributed to this diversification. Four species of snapping shrimps (Alpheus armatus, A. immaculatus, A. polystictus and A. roquensis) are distributed mainly sympatrically in the Caribbean, while the fifth species (A. rudolphi) is restricted to Brazil. All five species are obligate commensals of sea anemones with a high degree of fidelity and ecological specificity for host species and habitat. We analysed sequence data from 10 nuclear genes and the mitochondrial COI gene in 11-16 individuals from each of the Caribbean taxa and from the only available specimen of the Brazilian taxon. Phylogenetic analyses support morphology-based species assignments and a well-supported Caribbean clade. The Brazilian A. rudolphi is recovered as an outgroup to the Caribbean taxa. Isolation-migration coalescent analysis provides evidence for historical gene flow among sympatric sister species. Our data suggest that both selection for a novel host and selection for host microhabitat may have promoted diversification of this complex despite gene flow.

Asymmetric conspecific sperm precedence in relation to spawning times in the Montastraea annularis species complex (Cnidaria: Scleractinia).

In broadcast spawners, prezygotic reproductive isolation depends on differences in the spatial and temporal patterns of gamete release and gametic incompatibility. Typically, gametic incompatibility is measured in no-choice crosses, but conspecific sperm precedence (CSP) can prevent hybridization in gametes that are compatible in the absence of sperm competition. Broadcast spawning corals in the Montastraea annularis species complex spawn annually on the same few evenings. Montastraea franksi spawns an average of 110 min before M. annularis, with a minimum gap of approximately 40 min. Gametes are compatible in no-choice heterospecific assays, but it is unknown whether eggs exhibit choice when in competition. Hybridization depends on either M. franksi eggs remaining unfertilized and in proximity to M. annularis when the latter species spawns or M. franksi sperm remaining in sufficient viable concentrations when M. annularis spawns. We found that the eggs of the early spawning M. franksi demonstrate strong CSP, whereas CSP appears to be lacking for M. annularis eggs. This study provides evidence of diverging gamete affinities between these recently separated species and suggests for the first time that selection may favour CSP in earlier spawning species when conspecific sperm is diluted and aged and is otherwise at a numeric and viability disadvantage with heterospecific sperm.

Apolipoprotein-defined lipoprotein abnormalities in rheumatoid arthritis patients and their potential impact on cardiovascular disease.

OBJECTIVES: The aim of this study was to explore apolipoprotein-defined lipoproteins for abnormalities when comparing non-rheumatological controls to rheumatoid arthritis (RA) patients. METHODS: Apolipoprotein and lipoprotein profiles were measured on 94 RA patients and 79 controls by immunoturbidimetric procedures, electroimmunoassays, and immunoprecipitation. Differences between means were tested with a two-sided Student t test with Satterthwaite adjustment. p-values were adjusted for multiple comparisons using the Bonferroni procedure. RESULTS: RA patients had significantly higher levels of total cholesterol (TC), triglycerides (TG), and very low density lipoprotein cholesterol (VLDL-C) than controls, but no significant differences in the levels of high density lipoprotein cholesterol (HDL-C) and LDL-C. RA patients had significantly lower levels of apolipoprotein (apo)A-I and lipoprotein (Lp)A-I:A-II, but no difference in levels of LpA-I than normal controls. There was a significant difference in the levels of LpB:C but not in LpB:C:E between RA patients and controls. The main abnormality among apoB lipoproteins was the significantly increased concentration of the LpA-II:B:C:D:E subclass in RA patients in comparison with controls. The high levels of LpA-II:B:C:D:E are also reflected in significantly increased levels of apoC-III, and apoC-III bound to apoB lipoproteins. CONCLUSION: The LpA-II:B:C:D:E subclass has potential as a new marker for cardiovascular disease (CVD) in RA patients.

Apolipoprotein B-containing lipoprotein subclasses as risk factors for cardiovascular disease in patients with rheumatoid arthritis.

OBJECTIVE: The purpose of this study was to explore whether nontraditional risk factors, such as apolipoprotein C-III (Apo C-III) and its corresponding Apo B lipoprotein (Lp) subclasses, contribute to the risk of cardiovascular disease in rheumatoid arthritis (RA) patients. METHODS: Apolipoprotein and lipoproteins were measured in 152 RA patients by immunoturbidimetric procedures, electroimmunoassay, and immunoprecipitation. Patients had a coronary artery calcium (CAC) score assessed at baseline and at year 3. Differences in the CAC scores between baseline and year 3 were calculated and dichotomized at 0, where patients with a difference score >0 were denoted as progressors and the rest were denoted as nonprogressors. Differences between means were tested with a 2-sided independent Student's t-test with Satterthwaite's adjustment. Proportion differences were tested with a chi-square test. Multiple logistic regression was performed to assess the relationship between apolipoprotein and lipoprotein levels and the dichotomized CAC score. RESULTS: Progressors accounted for almost 60% of the cohort. Progressors had significantly higher levels of triglycerides, very low-density lipoprotein (VLDL) cholesterol, total cholesterol/high-density lipoprotein (HDL), triglycerides/HDL, Apo B, LpA-II:B:C:D:E, LpB:C, Apo B/Apo A-I, Apo C-III, and Apo C-III-heparin precipitate than the nonprogressors. After adjusting for age, sex, statin use (yes/no), and hypertension (yes/no), significant risk factors of progressors were total cholesterol, triglycerides, VLDL cholesterol, LDL cholesterol, Apo B, LpB:C, Apo C-III, and Apo B/Apo A-I. CONCLUSION: Apo C-III-containing Apo B lipoprotein subclasses were found to be significantly elevated in progressors compared to nonprogressors. Many of these same lipoproteins were found to be associated with an increase in CAC scores among progressors. These lipoproteins may be considered new risk factors for progression of atherosclerosis in RA patients.

Implications of isolation and low genetic diversity in peripheral populations of an amphi-Atlantic coral.

Limited dispersal and connectivity in marine organisms can have negative fitness effects in populations that are small and isolated, but reduced genetic exchange may also promote the potential for local adaptation. Here, we compare the levels of genetic diversity and connectivity in the coral Montastraea cavernosa among both central and peripheral populations throughout its range in the Atlantic. Genetic data from one mitochondrial and two nuclear loci in 191 individuals show that M. cavernosa is subdivided into three genetically distinct regions in the Atlantic: Caribbean-North Atlantic, Western South Atlantic (Brazil) and Eastern Tropical Atlantic (West Africa). Within each region, populations have similar allele frequencies and levels of genetic diversity; indeed, no significant differentiation was found between populations separated by as much as 3000 km, suggesting that this coral species has the ability to disperse over large distances. Gene flow within regions does not, however, translate into connectivity across the entire Atlantic. Instead, substantial differences in allele frequencies across regions suggest that genetic exchange is infrequent between the Caribbean, Brazil and West Africa. Furthermore, markedly lower levels of genetic diversity are observed in the Brazilian and West African populations. Genetic diversity and connectivity may contribute to the resilience of a coral population to disturbance. Isolated peripheral populations may be more vulnerable to human impacts, disease or climate change relative to those in the genetically diverse Caribbean-North Atlantic region.

Ablation of Stat3 by siRNA alters gene expression profiles in JEG-3 cells: a systems biology approach.

Control of inflammation at the maternal-fetal interface is a critical element in mammalian pregnancy. Previous work from our laboratory has shown that Stat3 may be a placental mediator involved in maintaining immunologic homeostasis at the maternal-fetal interface. The aim of the current study is to further elucidate the role of Stat3 in response to inflammation. As ablation of Stat3 in mice results in embryonic lethality, we evaluated the role of Stat3 in vitro using an siRNA approach. Trophoblast-like JEG-3 cells were transfected with an siRNA construct specific to Stat3. Experimental and control cells were exposed to conditioned medium from PHA-activated peripheral blood mononuclear cells and incubated for 45 min. Cells were then collected and RNA isolated for transcriptional profiling using human Affymetrix U133 plus 2.0 GeneChips. Differences in gene expression between control and Stat3-ablated cells were evaluated using conventional statistical methods. Fifty-two genes were detected as up-regulated in conditioned medium in both mock transfected and in Stat3 siRNA transfected JEG-3 cells. Two genes (EPAS1 and RASGEF1B) were up-regulated only in cells transfected with negative control siRNA, while 36 genes were up-regulated only in cells transfected with Stat3 siRNA. Sixty genes were differentially expressed between Stat3 siRNA transfected cells relative to mock transfected cells both in basal and conditioned medium. These included 31 genes up-regulated with Stat3 siRNA transfected cells and 29 genes down-regulated with Stat3 siRNA. Eleven genes were differentially expressed only in basal medium. Seven of these were up-regulated in the presence of Stat3 siRNA and four were down-regulated. Nine genes were differentially expressed only in conditioned medium. Six of these were up-regulated and three down-regulated in the presence of Stat3 siRNA. Off-target effects were excluded in a second set of experiments in which Stat3 mRNA was targeted at a different site and quantitative real-time PCR performed on selected genes derived from the microarray analysis. While some of the genes that showed differential expression between Stat3-ablated cells and mock transfected controls were genes typically associated with immune response (e.g., CCR7 and IRAK1), in silico modeling of the microarray data also revealed complex networks of signaling molecules and molecules associated with cellular metabolism previously seen in transcription factor ablation in model organisms. We conclude thus: Stat3 controls a specific gene set in trophoblast-like JEG-3 cells. While some differentially expressed genes and in silico models of their functions are consistent with the hypothesis that Stat3 plays a role in regulating inflammation, Stat3-mediated response to inflammation appears to also involve complex homeostatic adaptations of a non-immunologic nature.

Defective T-cell ERK signaling induces interferon-regulated gene expression and overexpression of methylation-sensitive genes similar to lupus patients.

Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by the production of autoantibodies against a host of nuclear antigens. The pathogenesis of lupus is incompletely understood. Environmental factors may play a role via altering DNA methylation, a mechanism regulating gene expression. In lupus, genes including CD11a and CD70 are overexpressed in T cells as a result of promoter hypomethylation. T-cell DNA methyltransferase expression is regulated in part by the extracellular signal-regulated kinase (ERK) signaling pathway. In this study, we investigate the effects of decreased ERK pathway signaling in T cells using transgenic animals. We generated a transgenic mouse that inducibly expresses a dominant-negative MEK in T cells in the presence of doxycycline. We show that decreased ERK pathway signaling in T cells results in decreased expression of DNA methyltransferase 1 and overexpression of the methylation-sensitive genes CD11a and CD70, similar to T cells in human lupus. Our transgenic animal model also develops anti-dsDNA antibodies. Interestingly, microarray expression assays revealed overexpression of several interferon-regulated genes in the spleen similar to peripheral blood cells of lupus patients. This model supports the contention that ERK pathway signaling defects in T cells contribute to the development of autoimmunity.

Coral reefs under rapid climate change and ocean acidification.

Atmospheric carbon dioxide concentration is expected to exceed 500 parts per million and global temperatures to rise by at least 2 degrees C by 2050 to 2100, values that significantly exceed those of at least the past 420,000 years during which most extant marine organisms evolved. Under conditions expected in the 21st century, global warming and ocean acidification will compromise carbonate accretion, with corals becoming increasingly rare on reef systems. The result will be less diverse reef communities and carbonate reef structures that fail to be maintained. Climate change also exacerbates local stresses from declining water quality and overexploitation of key species, driving reefs increasingly toward the tipping point for functional collapse. This review presents future scenarios for coral reefs that predict increasingly serious consequences for reef-associated fisheries, tourism, coastal protection, and people. As the International Year of the Reef 2008 begins, scaled-up management intervention and decisive action on global emissions are required if the loss of coral-dominated ecosystems is to be avoided.

Multiplex serum cytokine monitoring as a prognostic tool in rheumatoid arthritis.

OBJECTIVE: Early optimized therapy of rheumatoid arthritis (RA) results in improved outcomes. The initiation of optimized therapy is hindered by the difficulty of early diagnosis and the limitations of current disease activity and therapeutic response assessment tools. Identifying patients requiring early combination DMARD/biologic therapy is currently a significant clinical challenge given the lack of definitive prognostic criteria. Since cytokines are soluble intracellular signaling molecules that modulate disease pathology in RA, we tested the recent conjecture that en mass serum cyto-kine measurement and monitoring will provide a useful tool for effective therapeutic management in RA. METHODS: We assayed the levels of 16 serum cytokines in 18 RA patients treated prospectively with methotrexate and from 18 unaffected controls. Specific mechanistic aspects of inflammatory pathology in the periphery could be discerned on a patient-specific basis from patients' serum cytokine profiles, information that may aid in the design of anti-cytokine biologic therapy. A serum Cytokine Activity Index (CAI) was also created using multi-variant analysis methods. RESULTS: Distinct cytokines were significantly elevated in RA patients relative to controls, and three distinct clusters with correlations to disease activity were identified. The Cytokine Activity Index correlated well with the therapeutic res-ponse; responders and non-responders in this cohort were distinguishable as early as one month post initiation of methotrexate therapy, well before clinical assessments of response are commonly completed. CONCLUSION: Clinical assessment tools could be derived from this approach that may provide a means to continually track patients, allowing intervention strategies to be better evaluated on a patient-specific basis and to identify residual cytokine activity that could be used to guide combination therapy.

Circulating cytokines in Norwegian patients with psoriatic arthritis determined by a multiplex cytokine array system.

OBJECTIVES: Serum cytokines play an important role in the pathogenesis of psoriatic arthritis (PsA) by initiating and perpetuating various cellular and humoral autoimmune processes. The aim of this study was to describe a broad spectrum of T- and B-cell cytokines, growth factors and chemokines in patients with PsA and healthy individuals. METHODS: A novel protein array system, denoted as multiplex cytokine assay was utilized to measure simultaneously the levels of 23 circulating cytokines of patients with PsA and healthy individuals. Additionally, correlational clustering and discriminant function analysis (DFA), two multivariate, supervised analysis methods were employed to identify a subset of biomarkers in order to describe potential functional inter-relationships among these pathological cytokines and identify biomarkers with prognostic and diagnostic utility. RESULTS: Univariate analysis demonstrated that serum levels of a complex set of immune and inflammatory modulating cytokines are significantly up-regulated in patients with PsA relative to unaffected controls including interleukin (IL)-10, IL-13, interferen (IFN)-alpha, epidermal growth factor (EGF), vascular endothelial growth factor (VEGF), fibroblast growth factor [CCL3 macrophage inflammatory protein (MIP)-1alpha], CCL4 (MIP-1beta) and CCL11 (Eotaxin), while granulocyte-colony stimulating factor was significantly reduced in PsA patients. Correlational clustering was able to discriminate among, and hence subclassify, patients with varying levels of disease activity, which may prove useful in guiding therapy in these apparently phenotypically distinct disease subsets. DFA identified EGF, IFN-alpha, VEGF, CCL3 (MIP-1alpha) and IL-12p40 as analytes with the strongest discriminatory power among various PsA patients and controls. CONCLUSIONS: Our findings suggest that these factors modulate PsA pathology and the articular involvement in a synergistic manner. Identifying factors could be used in the development of clinical diagnostic tests, which are valuable to guide evidence-based diagnosis and disease management of PsA.

Template-driven gene selection procedure.

The hierarchical clustering and statistical techniques usually used to analyse microarray data do not inherently represent the underlying biology. Herein, a hybrid approach involving characteristics of both supervised and unsupervised learning is presented. This approach is based on template matching in which the interaction of the variables of inherent malignancy and the ability to express the malignant phenotype are modelled. Immortalised normal urothelial cells and bladder cancer cells of different malignancy were grown in conventional two-dimensional tissue culture and in three dimensions on extracellular matrices (ECMs) that were either permissive or restrictive for expression of the malignant phenotype. The transcriptome represents the effects of two variables--inherent malignancy and the modulatory effect of ECM. By assigning values to each of the biological variables of inherent malignancy and the ability to express the malignant phenotype, a template was constructed, which encapsulated the interaction between them. Gene expression correlating both positively and negatively with the template was observed, but when iterative correlations were carried out, the different models for the template converged on the same actual template. A subset of 21 genes was identified, which correlated with two a priori models or an optimised model above the 95% confidence limits identified in a bootstrap resampling with 5000 permutations of the data set. The correlation coefficients of expression of several genes were > 0.8. Analysis of upstream transcriptional regulatory elements (TREs) confirmed that these genes were not a randomly selected set of genes. Several TREs were identified as significantly over-expressed in the sample of 20 genes for which TREs were identified, and the high correlations of several genes were consistent with transcriptional co-regulation. The authors suggest that the template method can be used to identify a unique set of genes for further investigation.

A genome-scale assessment of peripheral blood B-cell molecular homeostasis in patients with rheumatoid arthritis.

OBJECTIVE: While rheumatoid arthritis (RA) is considered a prototypical autoimmune disease, the specific roles of B-cells in RA pathogenesis is not fully delineated. METHODS: We performed microarray expression profiling of peripheral blood B-cells from RA patients and controls. Data were analysed using differential gene expression analysis and 'gene networking' analysis (characterizing clusters of functionally inter-relelated genes) to identify both regulatory genes and the pathways in which they participate. Results were confirmed by quantitative real-time polymerase chain reaction and by measuring the levels of 10 serum cytokines involved in the pathways identified. RESULTS: Genes regulating and effecting the cell-cycle, proliferation, apoptosis, autoimmunity, cytokine networks, angiogenesis and neuro-immune regulation were differentially expressed in RA B-cells. Moreover, the serum levels of several soluble factors that modulate these pathways, including IL-1beta, IL-5, IL-6, IL-10, IL-12p40, IL-17 and VEGF were significantly increased in this cohort of RA patients. CONCLUSIONS: These results outline aspects of the multifaceted role B-cells play in RA pathogenesis in which immune dysregulation in RA modulates B-cell biology and thereby contributes to the induction and perpetuation of a pathogenic humoral immune response.

Mobile classification in microarray experiments.

In a homogeneous group of samples, there are genes whose expression variations can be attributed to factors other than experimental errors. These factors can include natural biological oscillations or metabolic processes. These genes are rarely classified as 'interesting' based on their variability profile. However, their dynamic behaviour can tease out important clues about naturally occurring biological processes in the organism under study and can be used for group classification. Dynamical discriminate function analysis was developed on the concept that stable classification parameters (roots) can be derived from highly variable gene-expression data. Stability of these combinations implies a strongly compensatory relationship that may divulge functional interconnections.

The marine indo-west pacific break: contrasting the resolving power of mitochondrial and nuclear genes.

Simultaneous studies of both nuclear and mitochondrial markers were undertaken in two widespread Indo-West Pacific (IWP) marine invertebrates to compare and contrast the ability of these markers to resolve genetic structure. In particular, we were interested in the resolution of a genetic break between the Indian and Pacific Oceans due to historical isolation. Sequence variation from the nuclear gene encoding myosin heavy chain (MyHC) and the mitochondrial gene cytochrome oxidase I (COI) were examined for the snapping shrimp Alpheus lottini from wide-ranging populations throughout the Indian and Pacific Oceans. A previously identified genetic break between oceans based on COI sequences appears to have been an artifact caused by the inadvertent inclusion of pseudogene sequences; our new COI data provide evidence only of a break between IWP and East Pacific populations. Distribution of a single nucleotide polymorphism in MyHC, on the other hand, shows evidence of a cline between Indian and Pacific Oceans. New allozyme and mtDNA sequence data were also obtained for the starfish Linckia laevigata. Allozyme data show a clear genetic break between Indian Ocean populations and Pacific (including western Australian) populations, whereas the distribution of mtDNA haplotypes shows a region of overlap in the central IWP. Comparisons of our data for both Alpheus and Linckia with data from other population genetic studies in the IWP suggest that nuclear markers (allozymes, sequence data and morphological characters) may in some instances reveal historical patterns of genetic population structure whereas mtDNA variation better reflects present day patterns of gene flow.

Zooxanthellae of the Montastraea annularis species complex: patterns of distribution of four taxa of Symbiodinium on different reefs and across depths.

Corals of the Montastraea annularis complex host several different dinoflagellates in the genus Symbiodinium. Here we address two questions arising from our previous studies of these associations on an offshore reef. First, do the same taxa and patterns of association (Symbiodinium A and B found in higher irradiance habitats than Symbiodinium C) occur on an inshore reef? Second, does M. franksi at the limits of its depth range host only Symbiodinium C, as it does at intermediate depths? In both surveys, a new Symbiodinium taxon and different patterns of distribution (assayed by analyses of small ribosomal subunit RNA genes [srDNA]) were observed. Inshore, a taxon we name Symbiodinium E predominated in higher irradiance habitats in M. franksi and its two sibling species; the only other zooxanthella observed was Symbiodinium C. Offshore, M. franksi mainly hosted Symbiodinium C, but hosted Symbiodinium A, B, C, and E in shallow water and Symbiodinium E and C in very deep water. Symbiodinium E may be stress-tolerant. Observed srDNA heterogeneity within samples of Symbiodinium B, C, and E is interpreted as variation across copies within this multigene family. Experimental bleaching of Symbiodinium C supported this interpretation. Thus sequences from natural samples should be interpreted cautiously.

Repopulation of Zooxanthellae in the Caribbean corals Montastraea annularis and M. faveolata following experimental and disease-associated bleaching.

Caribbean corals of the Montastraea annularis species complex associate with four taxa of symbiotic dinoflagellates (zooxanthellae; genus Symbiodinium) in ecologically predictable patterns. To investigate the resilience of these host-zooxanthella associations, we conducted field experiments in which we experimentally reduced the numbers of zooxanthellae (by transplanting to shallow water or by shading) and then allowed treated corals to recover. When depletion was not extreme, recovering corals generally contained the same types of zooxanthellae as they did prior to treatment. After severe depletion, however, recovering corals were always repopulated by zooxanthellae atypical for their habitat (and in some cases atypical for the coral species). These unusual zooxanthellar associations were often (but not always) established in experimentally bleached tissues even when adjacent tissues were untreated. Atypical zooxanthellae were also observed in bleached tissues of unmanipulated Montastraea with yellow-blotch disease. In colonies where unusual associations were established, the original taxa of zooxanthellae were not detected even 9 months after the end of treatment. These observations suggest that zooxanthellae in Montastraea range from fugitive opportunists and stress-tolerant generalists (Symbiodinium A and E) to narrowly adapted specialists (Symbiodinium B and C), and may undergo succession.

Evidence for three major clades within the snapping shrimp genus Alpheus inferred from nuclear and mitochondrial gene sequence data.

The snapping shrimp genus Alpheus is among the most diverse of caridean shrimps, and analyses of taxa separated by the Isthmus of Panama have been used to estimate rates of molecular evolution. Although seven morphological groups have been informally suggested, no formal phylogenetic analysis of the genus has been previously attempted. Here we infer the phylogenetic relationships within Alpheus using sequence data from two nuclear genes, glucose-6-phosphate isomerase and elongation factor-1alpha, and from the mitochondrial gene cytochrome oxidase I. Three major clades corresponding to previously noted morphological features were identified. Discrepancies between earlier informal morphological groupings and molecular analyses largely consisted of species whose morphologies were not entirely typical of the group to which they had been assigned. The traditional placements of shrimp with highly sessile lifestyles and consequently simplified morphologies were also not supported by molecular analyses. Phylogenies for Alpheus suggest that specialized ecological requirements (e.g., symbiotic associations and estuarine habitats) and modified claw morphologies have evolved independently several times. These new analyses also support the sister species status of transisthmian pairs analyzed previously, although very similar pairs were not always resolved with the more slowly evolving nuclear loci. In addition, six new cryptic species were identified in the course of these studies plus a seventh whose status remains to be determined.