A molecular investigation of the genus Ecklonia (Phaeophyceae,Laminariales) with special focus on the Southern Hemisphere

Brown algae of the order Laminariales, commonly referred to as kelps, are the largest and most productive primary producers in the coastal inshore environment. The genus Ecklonia (Lessoniaceae, Phaeophyceae) consists of seven species with four species in the Northern Hemisphere and three in the Southern Hemisphere. It was recently transferred to the family Lessoniaceae based on phylogenetic analyses of nuclear and chloroplastic markers, though the type of the genus was not included and its relationship with allied genera Eckloniopsis and Eisenia remained unresolved. The present study is the first to produce a phylogeny focussed on the genus Ecklonia. It included sequences from nuclear, mitochondrial, and chloroplastic DNA, for most of the distribution range of the three current Southern Hemisphere species (Ecklonia radiata, Ecklonia maxima, and a sample of a putative Ecklonia brevipes specimen), sequences for East Asiatic species (Ecklonia cava, Ecklonia kurome, and Ecklonia stolonifera), as well as the closely related genera Eckloniopsis and Eisenia. Results confirmed E. radiata and E. maxima as two distinct species in South Africa, E. radiata as a single species throughout the Southern Hemisphere (in South Africa, Australia, and New Zealand) and East Asiatic species as a distinct lineage from the Southern Hemisphere clade. Results further pointed out a close sister relationship between Eckloniopsis radicosa and two Eisenia species (including the type species: Eisenia arborea) to the genus Ecklonia suggesting that the genera Eckloniopsis and Eisenia are superfluous.     

Temperature acclimation and heat tolerance of photosynthesis in Norwegian Saccharina latissima (Laminariales,Phaeophyceae)

Kelps, seaweeds and seagrasses provide important ecosystem services in coastal areas, and loss of these macrophytes is a global concern. Recent surveys have documented severe declines in populations of the dominant kelp species, Saccharina latissima, along the south coast of Norway. S. latissima is a cold‐temperate species, and increasing seawater temperature has been suggested as one of the major causes of the decline. Several studies have shown that S. latissima can acclimate to a wide range of temperatures. However, local adaptations may render the extrapolation of existing results inappropriate. We investigated the potential for thermal acclimation and heat tolerance in S. latissima collected from three locations along the south coast of Norway. Plants were kept in laboratory cultures at three different growth temperatures (10, 15, and 20°C) for 4–6 weeks, after which their photosynthetic performance, fluorescence parameters, and pigment concentrations were measured. S. latissima obtained almost identical photosynthetic characteristics when grown at 10 and 15°C, indicating thermal acclimation at these temperatures. In contrast, plants grown at 20°C suffered substantial tissue deterioration, and showed reduced net photosynthetic capacity caused by a combination of elevated respiration and reduced gross photosynthesis due to lowered pigment concentrations, altered pigment composition, and reduced functionality of Photo‐system II. Our results support the hypothesis that extraordinarily high temperatures, as observed in 1997, 2002, and 2006, may have initiated the declines in S. latissima populations along the south coast of Norway. However, observations of high mortality in years with low summer temperatures suggest that reduced population resilience or other factors may have contributed to the losses.     

Examining the reproductive success of bull kelp (Nereocystis luetkeana,Phaeophyceae, Laminariales) in climate change conditions

Climate change is affecting marine ecosystems in many ways, including raising temperatures and leading to ocean acidification. From 2014 to 2016, an extensive marine heat wave extended along the west coast of North America and had devastating effects on numerous species, including bull kelp (Nereocystis luetkeana). Bull kelp is an important foundation species in coastal ecosystems and can be affected by marine heat waves and ocean acidification; however, the impacts have not been investigated on sensitive early life stages. To determine the effects of changing temperatures and carbonate levels on Northern California's bull kelp populations, we collected sporophylls from mature bull kelp individuals in Point Arena, CA. At the Bodega Marine Laboratory, we released spores from field-collected bull kelp, and cultured microscopic gametophytes in a common garden experiment with a fully factorial design crossing modern conditions (11.63 ± 0.54°C and pH 7.93 ± 0.26) with observed extreme climate conditions (15.56 ± 0.83°C and 7.64 ± 0.32 pH). Our results indicated that both increased temperature and decreased pH influenced growth and egg production of bull kelp microscopic stages. Increased temperature resulted in decreased gametophyte survival and offspring production. In contrast, decreased pH had less of an effect but resulted in increased gametophyte survival and offspring production. Additionally, increased temperature significantly impacted reproductive timing by causing female gametophytes to produce offspring earlier than under ambient temperature conditions. Our findings can inform better predictions of the impacts of climate change on coastal ecosystems and provide key insights into environmental dynamics regulating the bull kelp lifecycle.     

Taxonomy of the southwestern Atlantic endemic kelp: Laminaria abyssalis and Laminaria brasiliensis (Phaeophyceae,Laminariales) are not different species

Two endemic species of Laminaria, Laminaria abyssalis Joly & Oliveira Filho and L. brasiliensis Joly & Oliveira Filho, from the tropical southwestern Atlantic coast have been described. The aim of this work was to determine the conspecificity of these species based on morphological and molecular analyses (ribulose 1,5‐bisphosphate carboxylase/oxgenase, large subunit (rbcL), internal transcribed spacer (ITS) and cytochrome c oxidase subunit I (coxI)). We found an overlap between the morphological characters that are considered taxonomically important for distinguishing these two species; these characters included a differing pattern of blade splitting. In the three molecular analyses, the Brazilian Laminaria specimens were grouped into one clade with maximum support. These data support the hypothesis that the individuals analyzed represent only one species, L. abyssalis. The molecular analysis also showed L. abyssalis to be sister group to L. digitata.     

Variability in iodine in temperate seaweeds and iodine accumulation kinetics of Fucus vesiculosus and Laminaria digitata (Phaeophyceae,Ochrophyta)

The biogeochemistry of iodine in temperate coastal ecosystems is largely mediated by macroalgae, which act as a major biological sink and source of iodine. Their capacity to accumulate, retain and release iodine has been associated with abiotic and biotic stressors, but quantitative information is limited. We evaluated the seasonal iodine retention capacity of eleven macroalgal species belonging to different systematic groups, collected from two sites in Ireland. Iodine accumulation and retention were then further quantified in Fucus vesiculosus and Laminaria digitata in relation to I^? concentrations in seawater and temperature. In general, iodine contents were ~10¹–10² μmol · (g dw)^?1 for Laminariales, 10⁰–10¹ μmol · (g dw)^?1 for Fucales, 10^?1–10⁰ μmol · (g dw)^?1 for Rhodophyta, and 10^?1 μmol · (g dw)^?1 for Chlorophyta. Typically, algal iodine contents were above average in winter and below average in summer. Iodine accumulation in F. vesiculosus and L. digitata depended on I^? availability and followed the Michaelis‐Menten kinetic. The ratio of maximum accumulation rate to half accumulation coefficient (ρ_max: K _t) was 2.4 times higher for F. vesiculosus than for L. digitata , suggesting that F. vesiculosus was more efficient in iodine accumulation. Both species exhibited a temperature‐dependent net loss of iodine, and only an exposure to sufficient external I^? concentrations compensated for this loss. This study revealed that both environmental (e.g., I^? in seawater, temperature) and organismal (e.g., the status of the iodine storage pool) variables determine retention and variability in iodine in temperate seaweeds.     

Diversification and biogeographic history of the Western Palearctic freshwater flatworm genus Schmidtea (Tricladida: Dugesiidae), with a redescription of Schmidtea nova

The freshwater flatworm genus Schmidtea is endemic in the Western Palearctic region, where it is represented by only four species, thus contrasting with the high species diversity of the closely related genus Dugesia within Europe. Although containing an important model species in developmental and regeneration research, viz. Schmidtea mediterranea, no evolutionary studies on the genus Schmidtea have been undertaken. For the first time, we present a well‐resolved molecular phylogenetic tree of the four species of the genus, inferred on the basis of two molecular markers, and provide also the first detailed morphological account of Schmidtea nova. The phylogenetic tree generated corroborates an earlier speciation hypothesis based on karyological data and points to chromosomal rearrangements as the main drivers of speciation in this genus. The high genetic divergence between the four species, in combination with previous dating studies and their current geographic distribution, suggests that Schmidtea could have originated in Laurasia but lost most of its diversity during the Oligocene. Thus, its present distribution pattern may be the result of the expansion of three of its four relictual species over Europe, probably after the Pleistocene glaciations. Our detailed morphological study of S. nova revealed that it shows a number of remarkable features: interconnected testis follicles, parovaria, an ejaculatory duct exiting into the primary as well as the secondary seminal vesicle by means of a nipple, and the wall of the distal section of the ejaculatory duct being sclerotic or chitinized.     

Shading by giant kelp canopy can restrict the invasiveness of Undaria pinnatifida (Laminariales,Phaeophyceae)

The spread of non-indigenous and invasive seaweeds has increased worldwide, and their potential effects on native seaweeds have raised concern. Undaria pinnatifida is considered among the most prolific non-indigenous species. This species has expanded rapidly in the Northeast Pacific, overlapping with native communities such as the iconic giant kelp forests (Macrocystis pyrifera). Canopy shading by giant kelp has been argued to be a limiting factor for the presence of U. pinnatifida in the understory, thus its invasiveness capacity. However, its physiological plasticity under light limitation remains unclear. In this work, we compared the physiology and growth of juvenile U. pinnatifida and M. pyrifera sporophytes transplanted to the understory of a giant kelp forest, to juveniles growing outside of the forest. Extreme low light availability compared to that outside (~0.2 and ~4.4 mol photon ⋅ m⁻² ⋅ d⁻¹, respectively) likely caused a “metabolic energy crisis” in U. pinnatifida, thus restricting its photoacclimation plasticity and nitrogen acquisition, ultimately reducing its growth. Despite M. pyrifera juveniles showing photoacclimatory responses (e.g., increases in photosynthetic efficiency and lower compensation irradiance, E_c), their physiological/vegetative status deteriorated similarly to U. pinnatifida, which explains the low recruitment inside the forest. Generally, our results revealed the ecophysiological basis behind the limited growth and survival of juvenile U. pinnatifida sporophytes in the understory.     

Short‐ and long‐term acclimation patterns of the giant kelp Macrocystis pyrifera (Laminariales,Phaeophyceae) along a depth gradient

The giant kelp, Macrocystis pyrifera, is exposed to highly variable irradiance and temperature regimes across its geographic and vertical depth gradients. The objective of this study was to extend our understanding of algal acclimation strategies on different temporal scales to those varying abiotic conditions at various water depths. Different acclimation strategies to various water depths (0.2 and 4 m) between different sampling times (Jan/Feb and Aug/Sept 2012; long‐term acclimation) and more rapid adjustments to different depths (0.2, 2 and 4 m; short‐term acclimation) during 14 d of transplantation were found. Adjustments of variable Chl a fluorescence, pigment composition (Chl c, fucoxanthin), and the de‐epoxidation state of the xanthophyll cycle pigments were responsible for the development of different physiological states with respect to various solar radiation and temperature climates. Interestingly, the results indicated that phlorotannins are important during long‐term acclimation while antioxidants have a crucial role during short‐term acclimation. Furthermore, the results suggested that modifications in total lipids and fatty acid compositions apparently also might play a role in depth acclimation. In Aug/Sept (austral winter), M. pyrifera responded to the transplantation from 4 m to 0.2 m depth with a rise in the degree of saturation and a switch from shorter‐ to longer‐chain fatty acids. These changes seem to be essential for the readjustment of thylakoid membranes and might, thus, facilitate efficient photosynthesis under changing irradiances and temperatures. Further experiments are needed to disentangle the relative contribution of solar radiation, temperature and also other abiotic parameters in the observed physiological changes.     

Seasonal changes in taxon richness and abundance of mobile invertebrates inhabiting holdfast of annual kelp Ecklonia radicosa (Phaeophyceae,Lessoniaceae) at the central Pacific coast of Japan

To investigate seasonal changes in the taxon richness and abundance of mobile invertebrates inhabiting holdfasts of the warm temperate annual kelp Ecklonia radicosa, five holdfasts were collected monthly at the central Pacific coast of Japan from April to November 2014. During the study period, there was little variation in holdfast height and diameter, which ranged from 5.9 to 8.5 cm and from 7.1 to 10.8 cm, respectively. In total 7087 animals were collected from 40 holdfasts (177.2 individuals inds./holdfast, on average). The number of mobile invertebrates gradually increased from May (15 ± 9.9 inds./holdfast) to August (346 ± 152.5 inds./holdfast), with over 300 inds./holdfast until October before rapidly decreasing in November (110 ± 85.6 inds./holdfast). Similarly, taxon richness increased gradually from April (4.3 ± 1.0 taxa/holdfast) to August (11.0 ± 3.7 taxa/holdfast), and decreased in November (8.6 ± 2.3 taxa/holdfast). Interestingly, hundreds of mobile invertebrates inhabited holdfasts of kelp plants that had shed their blade in October and November. Taxon composition from August to October and the number of invertebrates from July to November were comparable to data previously reported for perennial kelps. In this study, the importance of annual Ecklonian species as biogenic habitats was demonstrated for the first time.     

Linking Physiology To Ecological Function: Environmental Conditions Affect Performance And Size Of The Intertidal Kelp Hedophyllum Sessile (Laminariales,Phaeophyceae)

For autogenic ecosystem engineers, body size is an aspect of individual performance that has direct connections to community structure; yet the complex morphology of these species can make it difficult to draw clear connections between the environment and performance. We combined laboratory experiments and field surveys to test the hypothesis that individual body size was determined by disparate localized physiological responses to environmental conditions across the complex thallus of the intertidal kelp Hedophyllum sessile, a canopy‐forming physical ecosystem engineer. We documented substantial (> 40%) declines in whole‐thallus photosynthetic potential (as Maximum Quantum Yield, MQY) as a consequence of emersion, which were related to greater than 10‐fold increases in intra‐thallus MQY variability (as Coefficient of Variation). In laboratory experiments, desiccation and high light levels during emersion led to lasting impairment of photosynthetic potential and an immediate > 25% reduction in area due to tissue contraction, which was followed by complete loss of structural integrity after three days of submersion. Tissue exposed to desiccation and high light during emersion had higher nitrogen concentrations and lower phlorotannin concentrations than tissue in control treatments (on average 1.36 and 0.1x controls, respectively), suggesting that conditions during emersion have the potential to affect food quality for consumers. Our data indicate that the complex thallus morphology of H. sessile may be critical to this kelp’s ability to persist in the intertidal zone despite the physiological challenges of emersion and encourage a more nuanced view of the concept of “sub‐lethal stress” on the scale of the whole individual.     

Species Diversity,Phylogeny and Large Scale Biogeographic Patterns of the Genus Padina (Phaeophyceae,Dictyotales)

The brown algal genus Padina (Dictyotales, Phaeophyceae) is distributed worldwide in tropical and temperate seas. Global species diversity and distribution ranges, however, remain largely unknown. Species‐level diversity was reassessed using DNA‐based, algorithmic species delineation techniques based on cox3 and rbcL sequence data from 221 specimens collected worldwide. This resulted in estimates ranging from 39 to 61 putative species (ESUs), depending on the technique as well as the locus. We discuss the merits, potential pitfalls, and evolutionary and biogeographic significance of algorithmic species delineation. We unveil patterns whereby ESUs are in all but one case restricted to either the Atlantic or Indo‐Pacific Ocean. Within ocean basins we find evidence for the vast majority of ESUs to be confined to a single marine realm. Exceptions, whereby ESUs span up to three realms, are located in the Indo‐Pacific Ocean. Patterns of range‐restricted species likely arise by repeated founder events and subsequent peripatric speciation, hypothesized to dominate speciation mechanisms for coastal marine organisms in the Indo‐Pacific. Using a three‐gene (cox3, psaA and rbcL), relaxed molecular clock phylogenetic analysis we estimated divergence times, providing a historical framework to interpret biogeographic patterns.     

Species delimitation and phylogeographic analyses in the Ectocarpus subgroup siliculosi (Ectocarpales,Phaeophyceae)

The genus Ectocarpus (Ectocarpales, Phaeophyceae) contains filamentous algae widely distributed in marine and estuarine habitats of temperate regions in both hemispheres. While E. siliculosus has become a model organism for genomics and genetics of the brown macroalgae, accurate species delineation, distribution patterns and diversity for the genus Ectocarpus remain problematic. In this study, we used three independent species delimitation approaches to generate a robust species hypothesis for 729 Ectocarpus specimens collected mainly along the European and Chilean coasts. These approaches comprised phylogenetic reconstructions and two bioinformatics tools developed to objectively define species boundaries (General Mixed Yule Coalescence Method and Automatic Barcode Gap Discovery). Our analyses were based on DNA sequences of two loci: the mitochondrial cytochrome oxidase subunit 1 and the nuclear internal transcribed spacer 1 of the ribosomal DNA. Our analyses showed the presence of at least 15 cryptic species and suggest the existence of incomplete lineage sorting or introgression between five of them. These results suggested the possible existence of different levels of reproductive barriers within this species complex. We also detected differences among species in their phylogeographic patterns, range and depth distributions, which may suggest different biogeographic histories (e.g., endemic species or recent introductions).     

Restriction enzyme analysis of variation and taxonomy in the kelp genus Laminaria (Laminariales,Phaeophyta)

High levels of phenotypic variation in kelp species necessitate the use of taxonomic markers that are independent of morphology. Restriction fragment length polymorphism (RFLP) analysis of nuclear DNA can provide such markers. In this paper we present the results of an RFLP analysis of cytoplasmic ribosomal DNA (rDNA) in three Laminaria species (L. agardhii, L. digitata, L. groenlandica). Comparison of the restriction maps of the nontranscribed spacer (NTS) in the rDNAs suggests that this method should be useful for the differentiation of these taxa. These results are discussed, as are the applications of RFLP mapping to the identification of field-collected, morphologically variable plants.     

Pathogenic bacteria associated with lesions and thallus bleaching symptoms in the Japanese kelp Laminaria religiosa Miyabe (Laminariales,Phaeophyceae)

During early Spring (April–May) when the seawater salinity drops suddenly and the seawater temperature increases drastically, severe lesions and thallus bleaching were observed in the Laminaria religiosa population at Oshoro Bay, Otaru, Hokkaido, Japan. The healthy and diseased kelp blades were collected and subjected to enumeration of total number of culturable bacteria and bacterial species. Bacterial enumerations were done using 3 different media formulations; high-nutrient media (Media 1), low-nutrient media (Media 2) and modified low-nutrient media with 5% kelp extract (Media 3). Seven bacterial species were isolated from the healthy kelp. These were Alcaligenes aquamarinas, Alteromonas sp., Azomonas agilis, Azotobacter beijerinckii, Escherichia coli, Halobacterium sp. and Halococcus sp. All 7 bacterial species were isolated on Media 2 and Media 3, but only 5 species were isolated using Media 1 with the absence of Halobacterium sp. and Halococcus sp. Highest total number of culturable bacteria was 2050 CFU/cm² on Media 3. Eight species of bacteria were isolated from the diseased kelp thallus with the addition of Erwinia amylovora. All 8 bacteria grew on Media 2 and Media 3, but only 6 species were isolated using Media 1 with the absence of Halobacterium sp. and Halococcus sp. Highest total number of culturable bacteria was 5830 CFU/cm² on Media 3. However, only 3 species were isolated from the lesioned area. The most abundant species was Alteromonas sp. followed by Halococcus sp. and Alcaligenes aquamarinas. The surface bacteria showed best growth on Media 3. Scanning Electron Microscopic images of the healthy and diseased thallus gave distinctive evidence of the severity of the lesions as well as the relative abundance in the bacterial population. In an effort to identify the symptoms causative organism, the isolated bacterial species were cultured and used to test Koch's postulates. Out of the 8 species, only Alteromonas sp. induced lesions on the axenic kelp blades. The inoculated bacteria were also re-isolated without any significant contamination. Hence, Alteromonas sp. is suggested as the possible disease causing organism.     

Phylogenetic relationships within the genus Sargassum (Fucales, Phaeophyceae), inferred from ITS-2 nrDNA, with an emphasis on the taxonomic subdivision of the genus

Sequences from the ribosomal DNA internal transcribed spacer‐2 (ITS‐2) were compared among species of Sargassaceae including the genera Sargassum and Hizikia. Species of different subgenera and sections of Sargassum were used to assess the taxonomic relationships within the genus, especially the subdivisions of the subgenus Bactrophycus. Sequences were aligned in accordance with their common secondary structure. Phylogenetic trees were constructed using neighbor‐joining, maximum likelihood and maximum parsimony methods with three species of Turbinaria as outgroups. The resulting phylogenetic trees showed that the genus Sargassum is divided into three clades corresponding to the subgenera Phyllotrichia, Sargassum and Bactrophycus. This last subgenus is further divided into four distinct groups: a Spongocarpus clade, a Teretia clade, a Hizikia clade, and a Halochloa/ Repentia clade. The position of the section Phyllo‐cystae, excluded from the subgenus Bactrophycus and included within the subgenus Sargassum is once again confirmed by the present study. Current results strongly support the assignation of Hizikia fusiformis to the genus Sargassum. Based on morphological differences and a distinct position in the molecular trees, Hizikia should be recognized as a section in the subgenus Bactrophycus so that Hizikia (Okamura) Yoshida, stat. nov. is proposed. A remarkably low divergence of ITS‐2 sequences was observed for the species in the sections Repentia and Halochloa, suggesting very recent radiation of these species. The subgenus Sargassum is divided into three clades corresponding to the three known sections: Acanthocarpicae, Malacocarpicae and Zygocarpicae, previously recognized by the morphology of receptacles. The position of Sargassum duplicatum, S. carpophyllum, S.yendoi, S. piluliferum and S. patens within the subgenus Sargassum is discussed.     

Adaptive radiations in butterflies: evolutionary history of the genus Erebia (Nymphalidae: Satyrinae)

We studied the speciose butterfly genus Erebia by reconstructing its phylogenetic relationships using parsimony and Bayesian approaches. We estimated times and rates of diversification for its lineages and employed a biogeographical analysis in order to reconstruct its evolutionary history. DNA sequence data from one mitochondrial gene and three nuclear genes were analyzed for a total of 74 species in Erebia. The estimated dates of origin and diversification for clades, in combination with a biogeographical analysis, suggest that the genus originated in Asian Russia and started its diversification process around 23 Myr. An important event was the dispersal of a lineage from Asia to Western Europe between 23 and 17 Myr, which allowed the radiation of most of species in the genus. The diversification pattern is consistent with a model of diversity limited by clade richness, which implies an early rapid diversification followed by deceleration due to a decrease in speciation. We argue that these characteristics of the evolutionary history of Erebia are consistent with a density‐dependent scenario, with species radiation limited by filling of niche space and reduced resources. We found that the Boeberia parmenio appears strongly supported in the genus Erebia and therefore we place Boeberia Prout, 1901 as a junior synonym of Erebia Dalman, 1816 ( syn. nov. ).     

Ottia meiospora (Ottiaceae,Rhodophyta), a new genus and family endophytic within the thallus of Nothocladus (Batrachospermales,Rhodophyta)

A new genus, Ottia, and family, Ottiaceae, are proposed within the Acrochaetiales to accommodate the uniseriate red algal endophyte of batrachspermalean taxa previously named Balbiania meiospora. Prior to this study, Balbiania investiens was transferred to its own family and order (Balbianiales) based on comparative DNA sequence data and a distinctive reproductive morphology. However, the second species described in this genus, B. meiospora, continued to be treated as a species of Audouinella (A. meiospora) pending further investigation. Phylogenetic analyses of sequence data confirmed only a distant relationship between the two endophytes, and a closer alliance of B. meiospora to Acrochaetiales. The data also showed that Ottia meiospora was the deepest diverging lineage in the Acrochaetiales, sister to all of the currently recognized genera and families. In this study, we review the classification of what we now call O. meiospora – reported from Australia, New Zealand and Brazil – based on sequence and morphological data. Morphological observations provided little clarity around the reproductive morphology or the life cycle of this endophyte of Nothocladus s. lat. found commonly in mainland Australia but, to date, less so in New Zealand.