Intra-genomic variation in symbiotic dinoflagellates: Recent divergence or natural hybridization?

<p>The perpetuity of coral reefs will ultimately depend on the ability of corals to adapt to changing conditions. Inter-specific hybridization can provide the raw genetic material necessary for adaptation, and stimulate macro-evolutionary leaps during periods of environmental upheaval. Though well-documented in corals, hybridization has yet to be identified in their dinoflagellate symbionts (genus Symbiodinium), despite growing evidence of sexual reproduction in this genus. The integral roles that these symbiotic algae play in coral productivity, reef accretion and ‘coral bleaching’ emphasize the need to better understand their short-term evolutionary potential. In this thesis, I develop new molecular and statistical methodology, and combine lab- and field-based analysis to explore the potential for hybridization between divergent Symbiodinium taxa. To screen for putative Symbiodinium hybrids, intra-genomic variation was examined within individual symbionts isolated from the reef-building coral Pocillopora damicornis at Lord Howe Island (Australia). A nested quantitative PCR (qPCR) assay was developed to quantify polymorphic internal transcribed spacer 2 (ITS2) sequences within the genome of each symbiont cell. Three genetically distinct Symbiodinium populations were detected co-existing within the symbiont consortium of P. damicornis. Mixed populations of ‘pure’ Symbiodinium types C100 and C109 coexisted with a population of cells hosting co-dominant C100 and C109 ITS2 repeats. Genetically heterogeneous Symbiodinium cells were more common than homogeneous symbionts in four of the six colonies analysed, with a maximum proportional abundance of 89%. Morphological, functional and ecological attributes of heterogeneous Symbiodinium cells were characterized to assess their candidacy as putative hybrids. The proportional abundance of genetically heterogeneous symbionts was spatially and temporally conserved within colonies, indicating a lack of competition between Symbiodinium populations. However, this abundance ratio varied considerably between colonies separated by metres to tens of metres, and to a greater extent between sites isolated by hundreds to thousands of metres. The local thermal maximum emerged as a significant predictor of the proportional abundance of genetically heterogeneous Symbiodinium cells, suggesting that the distribution of these ‘putative hybrids’ is influenced by a reduced affinity for thermal stress. Genetically heterogeneous Symbiodinium cells were around 50% larger (by volume) than homogeneous cells, occupied tissue of the coral host at reduced densities, and showed relatively poor light-harvesting efficiency. Colonies hosting a higher proportion of these symbionts suffered a reduction in overall photosynthetic performance (maximum gross photosynthesis normalised to respiration; P:R) at the ambient temperature of 25 °C. This disparity was maintained when the temperature was elevated to simulate the maximum experienced within the LHI lagoon (29 °C). Under these stressful conditions, colonies dominated by putative Symbiodinium hybrids were only marginally capable of net oxygen production. The influence of putative Symbiodinium hybrids on the growth and survival of P. damicornis was tested by reciprocally transplanting coral colonies between reef sites featuring distinct temperature regimes. Neither calcification nor mortality was influenced by the proportional abundance of genetically heterogeneous cells in the symbiont consortium. This uncoupling of symbiont performance and host fitness may be explained by stochastic events such as predation and disease, which substantially increase variation in growth and mortality in field experiments. Alternatively, it may represent some unknown benefit associated with hosting hybrid symbionts, belying their relatively poor photosynthetic performance, and explaining the widespread abundance of these heterogeneous Symbiodinium cells on the Lord Howe Island reef. Our inability to maintain many clade C Symbiodinium types in culture prevents direct observations of hybridization between C100 and C109. Unequivocal evidence of this phenomenon will therefore likely remain elusive until high-resolution, single-copy nuclear markers can be developed, since the incomplete displacement of ancestral polymorphisms can leave a similar genomic signature to that of hybridization. However, this study serves to provide an initial proof-of-principle for hybridization between divergent Symbiodinium taxa. In doing so, it highlights the need to better understand the evolutionary processes underpinning coral- and symbiont-adaptation in a changing climate.</p>

Living on the Edge: Protective Mechanisms Underlying Thermal Tolerance in High Latitude Symbiodinium spp.

<p>The association between symbiotic dinoflagellates (Symbiodinium spp.) and corals extends to subtropical and temperate regions, where sea surface temperatures (SSTs) are generally lower than in the tropics and can vary substantially over the course of the year due to seasonal changes. These high latitude coral-dinoflagellate symbioses might be better able to withstand thermal variability and might be particularly well equipped to cope with lower SSTs compared to their tropical relatives. The aim of this thesis was to analyze the cellular mechanisms that underlie heat and/or cold tolerance in a range of reef-building corals (Acropora yongei, Acropora solitariensis, Isopora palifera, Pocillopora damicornis, Porites heronensis and Stylophora sp.), as well as the symbiotic sea anemone Entacmaea quadricolor. In particular, the study focussed on protective mechanisms in their dinoflagellate symbionts as a potential determinant of thermal sensitivity (i.e. bleaching) or resistance of the intact symbiosis. High latitude reef-building corals were analyzed at the world’s southernmost coral reef at Lord Howe Island, while E. quadricolor was sampled at the subtropical coral community at North Solitary Island; both sites are located in New South Wales, Australia. The specific objectives were to assess the roles of: (1) xanthophyll deepoxidation; (2) thylakoid fatty acid composition; (3) Symbiodinium superoxide dismutase (SOD) and ascorbate peroxidase (APX) activity; and (4) D1 repair on the photophysiology, bleaching susceptibility and survivorship of a range of high-latitude coral-Symbiodinium associations from Lord Howe Island when exposed to elevated or decreased temperature. Furthermore, I aimed to: (5) characterise Symbiodinium diversity in the anemone E. quadricolor on the west coast of Australia; and (6) measure the dynamics of Symbiodinium ITS2 populations and SOD activity in two E. quadricolor phenotypes (green and pink colour phenotypes) in response to elevated temperature. I showed that thermal responses in high latitude corals and their dinoflagellate symbionts are highly variable, depending on host species (or phenotype) and Symbiodinium genotype, and that the activation of protective mechanisms in Symbiodinium was not necessarily correlated with sub-lethal bleaching susceptibility or survivorship of their coral hosts. More specifically: (1) In response to short-term heat stress and cold stress, xanthophyll de-epoxidation increased in some but not all bleaching susceptible (e.g. P. damicornis) and bleaching tolerant (P. heronensis) corals; (2) overall unsaturated thylakoid fatty acids increased in symbionts of a bleaching tolerant coral association, yet was not correlated with PSII photochemical efficiency; and (3) SOD and APX activity remained unchanged in the majority of Symbiodinium types regardless of bleaching susceptibility of the coral host, but decreased in bleaching susceptible Pocillopora damicornis when exposed to short-term heat stress. Elevated temperatures resulted in enhanced D1 turnover in two warm-water bleaching susceptible Symbiodinium-host combinations; however a direct link between increased dependence on D1 turnover and bleaching susceptibility was not demonstrated. From the results obtained it seems unlikely that the specific cellular adaptations in Symbiodinium alone determine the tolerance of Lord Howe corals to thermal variations. In contrast, the results highlight the significance of the particular host-symbiont combination and it appears that the host is important in determining, at least in part, the thermal response of the coral. Additionally, this study revealed a high diversity of Symbiodinium ITS2 (internal transcribed spacer 2) types in E. quadricolor from five locations on the west coast of Australia. E. quadricolor predominantly associated with six types of clade C (four of which were novel) and most anemones harboured multiple types simultaneously. At North Solitary Island, anemones simultaneously harboured Symbiodinium C25 and C3.25 (a novel variant of C3). Experimentally, I showed that anemones shuffled the relative proportions of C25 and C3.25 in response to elevated temperature, but not in both anemone colour phenotypes analyzed. Furthermore, baseline photobiological characteristics were distinct in the two different anemone colour morphs but were not correlated with the ratio of Symbiodinium C25 to C3.25, suggesting that host mechanisms such as pigmentation were involved in regulating light utilization by the symbionts. My hypothesis that symbiont shuffling was related to SOD activity, as such that those symbionts with enhanced SOD activity and increased capability to scavenge superoxide anion would increase in relative abundance in response to short-term heat stress, could not be proved. In summary, this thesis provides detailed information on some key cellular mechanisms that could underpin thermal sensitivity and resistance in high latitude Symbiodinium, and most importantly highlights the significance of the host-symbiont combination in determining the response to thermal stress. The various mechanistic findings described here further our understanding of the coral bleaching process in general and particularly give insight into physiological and cellular responses to coldwater stress in reef-building corals at high-latitude sites. The results of this thesis indicate that in light of ongoing climate change, as episodes of cold-water and warm-water anomalies will become more frequent, branching corals such as Acropora yongei or Pocillopora damicornis and their symbionts will experience physiological stress more frequently than massive species such as Porites heronensis. This might have profound impacts on the long-term stability and species composition of high latitude coral reefs.</p>

Living on the Edge: Protective Mechanisms Underlying Thermal Tolerance in High Latitude Symbiodinium spp.

<p>The association between symbiotic dinoflagellates (Symbiodinium spp.) and corals extends to subtropical and temperate regions, where sea surface temperatures (SSTs) are generally lower than in the tropics and can vary substantially over the course of the year due to seasonal changes. These high latitude coral-dinoflagellate symbioses might be better able to withstand thermal variability and might be particularly well equipped to cope with lower SSTs compared to their tropical relatives. The aim of this thesis was to analyze the cellular mechanisms that underlie heat and/or cold tolerance in a range of reef-building corals (Acropora yongei, Acropora solitariensis, Isopora palifera, Pocillopora damicornis, Porites heronensis and Stylophora sp.), as well as the symbiotic sea anemone Entacmaea quadricolor. In particular, the study focussed on protective mechanisms in their dinoflagellate symbionts as a potential determinant of thermal sensitivity (i.e. bleaching) or resistance of the intact symbiosis. High latitude reef-building corals were analyzed at the world’s southernmost coral reef at Lord Howe Island, while E. quadricolor was sampled at the subtropical coral community at North Solitary Island; both sites are located in New South Wales, Australia. The specific objectives were to assess the roles of: (1) xanthophyll deepoxidation; (2) thylakoid fatty acid composition; (3) Symbiodinium superoxide dismutase (SOD) and ascorbate peroxidase (APX) activity; and (4) D1 repair on the photophysiology, bleaching susceptibility and survivorship of a range of high-latitude coral-Symbiodinium associations from Lord Howe Island when exposed to elevated or decreased temperature. Furthermore, I aimed to: (5) characterise Symbiodinium diversity in the anemone E. quadricolor on the west coast of Australia; and (6) measure the dynamics of Symbiodinium ITS2 populations and SOD activity in two E. quadricolor phenotypes (green and pink colour phenotypes) in response to elevated temperature. I showed that thermal responses in high latitude corals and their dinoflagellate symbionts are highly variable, depending on host species (or phenotype) and Symbiodinium genotype, and that the activation of protective mechanisms in Symbiodinium was not necessarily correlated with sub-lethal bleaching susceptibility or survivorship of their coral hosts. More specifically: (1) In response to short-term heat stress and cold stress, xanthophyll de-epoxidation increased in some but not all bleaching susceptible (e.g. P. damicornis) and bleaching tolerant (P. heronensis) corals; (2) overall unsaturated thylakoid fatty acids increased in symbionts of a bleaching tolerant coral association, yet was not correlated with PSII photochemical efficiency; and (3) SOD and APX activity remained unchanged in the majority of Symbiodinium types regardless of bleaching susceptibility of the coral host, but decreased in bleaching susceptible Pocillopora damicornis when exposed to short-term heat stress. Elevated temperatures resulted in enhanced D1 turnover in two warm-water bleaching susceptible Symbiodinium-host combinations; however a direct link between increased dependence on D1 turnover and bleaching susceptibility was not demonstrated. From the results obtained it seems unlikely that the specific cellular adaptations in Symbiodinium alone determine the tolerance of Lord Howe corals to thermal variations. In contrast, the results highlight the significance of the particular host-symbiont combination and it appears that the host is important in determining, at least in part, the thermal response of the coral. Additionally, this study revealed a high diversity of Symbiodinium ITS2 (internal transcribed spacer 2) types in E. quadricolor from five locations on the west coast of Australia. E. quadricolor predominantly associated with six types of clade C (four of which were novel) and most anemones harboured multiple types simultaneously. At North Solitary Island, anemones simultaneously harboured Symbiodinium C25 and C3.25 (a novel variant of C3). Experimentally, I showed that anemones shuffled the relative proportions of C25 and C3.25 in response to elevated temperature, but not in both anemone colour phenotypes analyzed. Furthermore, baseline photobiological characteristics were distinct in the two different anemone colour morphs but were not correlated with the ratio of Symbiodinium C25 to C3.25, suggesting that host mechanisms such as pigmentation were involved in regulating light utilization by the symbionts. My hypothesis that symbiont shuffling was related to SOD activity, as such that those symbionts with enhanced SOD activity and increased capability to scavenge superoxide anion would increase in relative abundance in response to short-term heat stress, could not be proved. In summary, this thesis provides detailed information on some key cellular mechanisms that could underpin thermal sensitivity and resistance in high latitude Symbiodinium, and most importantly highlights the significance of the host-symbiont combination in determining the response to thermal stress. The various mechanistic findings described here further our understanding of the coral bleaching process in general and particularly give insight into physiological and cellular responses to coldwater stress in reef-building corals at high-latitude sites. The results of this thesis indicate that in light of ongoing climate change, as episodes of cold-water and warm-water anomalies will become more frequent, branching corals such as Acropora yongei or Pocillopora damicornis and their symbionts will experience physiological stress more frequently than massive species such as Porites heronensis. This might have profound impacts on the long-term stability and species composition of high latitude coral reefs.</p>

Intra-genomic variation in symbiotic dinoflagellates: Recent divergence or natural hybridization?

<p>The perpetuity of coral reefs will ultimately depend on the ability of corals to adapt to changing conditions. Inter-specific hybridization can provide the raw genetic material necessary for adaptation, and stimulate macro-evolutionary leaps during periods of environmental upheaval. Though well-documented in corals, hybridization has yet to be identified in their dinoflagellate symbionts (genus Symbiodinium), despite growing evidence of sexual reproduction in this genus. The integral roles that these symbiotic algae play in coral productivity, reef accretion and ‘coral bleaching’ emphasize the need to better understand their short-term evolutionary potential. In this thesis, I develop new molecular and statistical methodology, and combine lab- and field-based analysis to explore the potential for hybridization between divergent Symbiodinium taxa. To screen for putative Symbiodinium hybrids, intra-genomic variation was examined within individual symbionts isolated from the reef-building coral Pocillopora damicornis at Lord Howe Island (Australia). A nested quantitative PCR (qPCR) assay was developed to quantify polymorphic internal transcribed spacer 2 (ITS2) sequences within the genome of each symbiont cell. Three genetically distinct Symbiodinium populations were detected co-existing within the symbiont consortium of P. damicornis. Mixed populations of ‘pure’ Symbiodinium types C100 and C109 coexisted with a population of cells hosting co-dominant C100 and C109 ITS2 repeats. Genetically heterogeneous Symbiodinium cells were more common than homogeneous symbionts in four of the six colonies analysed, with a maximum proportional abundance of 89%. Morphological, functional and ecological attributes of heterogeneous Symbiodinium cells were characterized to assess their candidacy as putative hybrids. The proportional abundance of genetically heterogeneous symbionts was spatially and temporally conserved within colonies, indicating a lack of competition between Symbiodinium populations. However, this abundance ratio varied considerably between colonies separated by metres to tens of metres, and to a greater extent between sites isolated by hundreds to thousands of metres. The local thermal maximum emerged as a significant predictor of the proportional abundance of genetically heterogeneous Symbiodinium cells, suggesting that the distribution of these ‘putative hybrids’ is influenced by a reduced affinity for thermal stress. Genetically heterogeneous Symbiodinium cells were around 50% larger (by volume) than homogeneous cells, occupied tissue of the coral host at reduced densities, and showed relatively poor light-harvesting efficiency. Colonies hosting a higher proportion of these symbionts suffered a reduction in overall photosynthetic performance (maximum gross photosynthesis normalised to respiration; P:R) at the ambient temperature of 25 °C. This disparity was maintained when the temperature was elevated to simulate the maximum experienced within the LHI lagoon (29 °C). Under these stressful conditions, colonies dominated by putative Symbiodinium hybrids were only marginally capable of net oxygen production. The influence of putative Symbiodinium hybrids on the growth and survival of P. damicornis was tested by reciprocally transplanting coral colonies between reef sites featuring distinct temperature regimes. Neither calcification nor mortality was influenced by the proportional abundance of genetically heterogeneous cells in the symbiont consortium. This uncoupling of symbiont performance and host fitness may be explained by stochastic events such as predation and disease, which substantially increase variation in growth and mortality in field experiments. Alternatively, it may represent some unknown benefit associated with hosting hybrid symbionts, belying their relatively poor photosynthetic performance, and explaining the widespread abundance of these heterogeneous Symbiodinium cells on the Lord Howe Island reef. Our inability to maintain many clade C Symbiodinium types in culture prevents direct observations of hybridization between C100 and C109. Unequivocal evidence of this phenomenon will therefore likely remain elusive until high-resolution, single-copy nuclear markers can be developed, since the incomplete displacement of ancestral polymorphisms can leave a similar genomic signature to that of hybridization. However, this study serves to provide an initial proof-of-principle for hybridization between divergent Symbiodinium taxa. In doing so, it highlights the need to better understand the evolutionary processes underpinning coral- and symbiont-adaptation in a changing climate.</p>

Reports of Cubiceps baxteri McCulloch 1923 from Indian Ocean are probably misidentifications of Cubiceps whiteleggii (Waite 1894)

Cubiceps baxteri McCulloch 1923 was described based on a single, imperfect (devoid of a tail) stranded specimen collected from a beach in Lord Howe Island, Tasman Sea. Though C. baxteri was reported as a widely distributed tropical species (Butler 1979), it was mainly a result of its incorrect identification (see Agafonova 1994; Stewart and Last 2015). The distribution of C. baxteri is reported to be restricted to the Pacific Ocean, from Japan and eastwards to Baja California (Mexico), southwards to the Hawaiian Islands, New South Wales (Australia), and Lord Howe Island (Tasman Sea) to the Southern parts of Chile (Eschmeyer et al. 2017; Mundy 2005; Agafonova 1994).

First record of the Kermadec Clingfish, Flexor incus Conway, Stewart & Summers, 2018 (Gobiesocidae), from New Caledonia and Australia

Two specimens (17.1 and 29.1 mm standard length) of Flexor incus Conway, Stewart &amp; Summers, 2018 (Gobiesocidae) were collected from New Caledonia and Lord Howe Island, Australia. The species and genus were originally described on the basis of 15 specimens from the Kermadec Islands, New Zealand, where the genus has been considered endemic. The two specimens reported herein represent the first records of F. incus from New Caledonia and Australia.

Hypsipyrgias joseliae, a new species of lace bugs (Heteroptera: Tingidae: Tinginae) from New Guinea with a key to species of the genus Hypsipyrgias, and comments on three allied genera Hypsipyrgias Kirkaldy, 1908, Diplocysta Horváth, 1925, and Hypsotingis Drake, 1960

Hypsipyrgias joseliae sp. n. (Heteroptera: Tingidae: Tinginae) from New Guinea is described, illustrated and compared with its two relatives, namely H. telamonides Kirkaldy, 1908 from Australia, and H. euphues Drake and Ruhoff, 1962 from Lord Howe Island. Key to species of the genus Hypsipyrgias is also provided. Two genera very closely related to Hypsipyrgias Kirkaldy, 1908, namely Hypsotingis Drake, 1960 and Diplocysta Horváth, 1925 are re-diagnosed. Diplocysta globuliformis Hacker, 1928, D. papuana Drake, 1960, D. rustica, Drake, 1960 and D. thaleia Drake and Ruhoff, 1965 are transferred from Diplocysta to Hypsotingis.