Inventory of the Seaweeds and Seagrasses of the Hawaiian Islands

Simple Summary This inventory represents the first complete inventory of seaweeds (benthic brown, red, and green algae) and seagrasses from the Hawaiian Islands. We present taxonomic records compiled from the literature which include many recent descriptions of new species. Taxonomic records are accompanied by additional information allowing an assessment of the degree of molecular confirmation: whether the record has been verified by a match to DNA sequences from a type specimen, topotype sequences, or regional DNA sequences. In addition, taxa that have been identified solely based on morphology are indicated. In the face of numerous threats to biodiversity in the coming decades, it is hoped that this inventory will provide baseline data sets against which future changes may be compared. Abstract This updated list is composed of a total of 661 records, which includes 71 brown algae, 450 red algae, 137 green algae, and three seagrasses, with an overall rate of endemism of 13.2%. Almost half (46.7%) of the Hawaiian records presented here are represented by at least one DNA sequence, while 16.3% are confirmed through a DNA sequence match to a topotype, and 6.7% are confirmed through a DNA sequence match to a type specimen. The data are presented in the context of the natural history of the Hawaiian Islands, which is heavily influenced by the volcanic hotspot origin of the archipelago in the middle of the Pacific Ocean, as well as the important cultural role of seaweeds and other marine plants in Hawai‘i, and the current threats to marine ecosystems, which include the introduction and proliferation of a number of invasive marine macroalgae.


Introduction
The Hawaiian Archipelago consists of eight major islands, together with numerous atolls and islets, positioned in the central North Pacific Ocean [1]. The islands were formed sequentially through hotspot volcanic activity on the Pacific Plate, such that islands to the northwest end of the archipelago are the oldest (i.e., Hōlanikū, or Kure Atoll, at over 28 million years [Ma]), and those at the southeastern end of the chain are the youngest notably Hawai'i Island, at 0-0.6 Ma, with volcanic activity still producing new land [2]. The Archipelago extends for approximately 2400 km, and is extremely isolated, with the nearest continent (North America) being approximately 3900 km away [1].
The youngest and most southeastern islands of the archipelago, known as the Main Hawaiian Islands, are home to Hawai'i's permanent human population. The largest marine wildlife reserve in the world was established in 2006 to encompass the northwestern islands/atolls and surrounding waters of Hawai'i (1,510,000 km 2 -approximately the size of Germany), named the Papahānaumokuākea Marine National Monument (PMNM). The largely uninhabited PMNM has deep cultural significance to Native Hawaiians, and research on the marine macroalgae from PMNM has indicated substantial undescribed biodiversity-e.g., [3][4][5][6][7][8]. Shorelines in Hawai'i are largely determined by the geological processes that shaped the islands. Thus, initial volcanic activity yielded basaltic lava islands, and this feature still dominates the shorelines of Hawai'i Island and nearby east The list includes taxonomic information for each record including major lineage (e.g., phylum), order, family, genus, and species (and, where appropriate, subspecific taxon) ( Table 1). The table column "regionally reported as" is used to indicate instances where a different name is or has been used to refer to the taxon in the Hawaiian Islands and includes taxonomic synonyms as well as common and widespread misidentifications. Whether a taxon is endemic to the Hawaiian Islands is indicated (as yes, no, or maybe). Four subsequent columns give information about the degree of molecular confirmation (where present), and whether the taxon has been verified based on morphological comparisons: "Morphology" (identification is based on morphological examination), "Regional DNA Sequence" (identification based on one or more DNA sequences generated from Hawaiian material and compared to sequences from elsewhere), "Topotype Sequence Match" (sequences of Hawaiian material have been shown to reasonably match those from topotype material), or "Type Sequence Match" (sequences of Hawaiian material have been shown to reasonably match that of a type specimen for that taxon). Hawaiian sequences were considered to represent the topotype locality if they were from the same island. Finally, one or more representative references for each species record are listed (not a comprehensive list of all records), and a column for taxonomic notes is included. Taxa listed as doubtful records are still included in the totals presented, until such time as they are discounted as members of the Hawaiian flora.
As for any inventory, the current listing represents a point-in-time and will change as more new species and genera are described (we are aware of a number of these in progress at the time of writing but are not yet accepted for publication), and other kinds of taxonomic proposals (e.g., synonymies or removal of records) are put forward.

Results
A total of 661 marine algal and seagrass taxa are included in this Hawaiian inventory (Table 1, Figure 2). These records include 71 Phaeophyceae, or brown algae (representing 11 orders, 16 families, and 33 genera), 450 Rhodophyta, or red algae (23 orders, 58 families, and 197 genera), 137 Chlorophyta/Prasinodermatophyta, or green algae (7 orders, 23 families, and 41 genera), and 3 Tracheophyta, or seagrasses (1 order, 2 families, and 2 genera). The list includes taxonomic information for each record including major lineage (e.g., phylum), order, family, genus, and species (and, where appropriate, subspecific taxon) ( Table 1). The table column "regionally reported as" is used to indicate instances where a different name is or has been used to refer to the taxon in the Hawaiian Islands and includes taxonomic synonyms as well as common and widespread misidentifications. Whether a taxon is endemic to the Hawaiian Islands is indicated (as yes, no, or maybe). Four subsequent columns give information about the degree of molecular confirmation (where present), and whether the taxon has been verified based on morphological comparisons: "Morphology" (identification is based on morphological examination), "Regional DNA Sequence" (identification based on one or more DNA sequences generated from Hawaiian material and compared to sequences from elsewhere), "Topotype Sequence Match" (sequences of Hawaiian material have been shown to reasonably match those from topotype material), or "Type Sequence Match" (sequences of Hawaiian material have been shown to reasonably match that of a type specimen for that taxon). Hawaiian sequences were considered to represent the topotype locality if they were from the same island. Finally, one or more representative references for each species record are listed (not a comprehensive list of all records), and a column for taxonomic notes is included. Taxa listed as doubtful records are still included in the totals presented, until such time as they are discounted as members of the Hawaiian flora.
As for any inventory, the current listing represents a point-in-time and will change as more new species and genera are described (we are aware of a number of these in progress at the time of writing but are not yet accepted for publication), and other kinds of taxonomic proposals (e.g., synonymies or removal of records) are put forward.
be characterized using molecular tools, at any level. Seagrasses have the highest percentage of the flora represented by at least one DNA sequence (66.7%, albeit only three taxa comprise this component of the flora), followed by red algae (53.8%), brown algae (32.4%), and green algae (30.7%). A total of 16.3% of the records in the inventory has been shown to have a reasonably close match to a topotype, with values ranging from 0% (seagrasses), to 8.8% (green algae), 16.9% (brown algae), and 18.7% (red algae). A relatively small percentage of records (6.7%) has been confirmed through a reasonably close match to a sequence derived from type material (which includes recently described species for which DNA sequence data have been provided as part of the description); these values range from 0% (seagrasses), to 4.4% (green algae), 5.6% (brown algae), and 7.6% (red algae).  The vast majority (99.5%) of these records are based, at least in part, on morphological identifications. Approximately 46.7% of the flora is represented by at least one DNA sequence generated from Hawaiian material, meaning that 53.3% of the flora remains to be characterized using molecular tools, at any level. Seagrasses have the highest percentage of the flora represented by at least one DNA sequence (66.7%, albeit only three taxa comprise this component of the flora), followed by red algae (53.8%), brown algae (32.4%), and Biology 2023, 12, 215 6 of 71 green algae (30.7%). A total of 16.3% of the records in the inventory has been shown to have a reasonably close match to a topotype, with values ranging from 0% (seagrasses), to 8.8% (green algae), 16.9% (brown algae), and 18.7% (red algae). A relatively small percentage of records (6.7%) has been confirmed through a reasonably close match to a sequence derived from type material (which includes recently described species for which DNA sequence data have been provided as part of the description); these values range from 0% (seagrasses), to 4.4% (green algae), 5.6% (brown algae), and 7.6% (red algae).

Discussion
We present the first comprehensive inventory of Hawaiian marine algae and seagrasses combining both morphological and molecular data and provide data on the degree of endemism of the flora, as well as degree of certainty in identification for each record. Despite having several very thorough morphological compendia available for various lineages of the Hawaiian flora, no single compilation of species existed. The most recent compendium of the red, green, and brown marine algae of Hawaii reported a total of 512 species (343 red algae, 107 green algae, and 62 brown algae [12,14]), which compares with the 661 subgeneric taxa (450 red algae, 137 green algae, and 71 brown algae, plus 3 seagrasses) in our current inventory. With 47% of records represented by at least one DNA sequence generated from Hawaiian material, a large amount of survey work and inventory curation awaits additional sequencing efforts. However, much recent taxonomic and systematic revision has occurred for the flora, which further emphasizes the need for the present inventory. Future updates are certain as this work continues, particularly with respect to new species descriptions from the mesophotic flora.
Degree of confidence in identification is becoming increasingly recognized as an important part of reporting in taxonomic inventories, lists and checklists-e.g., [23]. Variation in confidence of identification is an unavoidable side-effect of combining records from historical sources, which largely relied on a strictly morphological identification approach, with more recent works that include DNA sequence comparisons. Moreover, interpretation of DNA sequence comparisons is being undertaken with a higher level of scrutiny than when the tool first become available to phycologists, when sequences deposited in public databases may or may not have represented the labeled taxon due to sequence quality issues or misidentification (which could be due to several factors, further described below). For example, highest confidence can be placed on an identification when one or more DNA sequences matches or nearly matches a sequence derived from type material of that taxon. These instances are rare given that most taxa are not represented by DNA sequences generated from their type specimens, but 6.7% of the records in the current inventory have indeed been verified at that level, primarily as a result of new species descriptions from the Hawaiian flora that include molecular characterization. The next highest level of confidence in identification stems from an often-used proxy for type sequences-topotype sequence data, or sequences derived from specimens collected from (or relatively close to) the type locality. Topotype sequences, however, may in fact be representative of other taxa that cohabit that locality, or other taxa found in nearby areas, if the topotype locality is interpreted too broadly. The third highest level of confidence in identification stems from sequences generated from regionally collected material that was morphologically identified as belonging to a specific taxon or had one or more molecular markers reasonably matching a labeled sequence from a public database of that taxon. Strictly morphological identifications may be reasonably accurate for some groups of marine algae and seagrasses but are known to be rife with potential for misidentification due to several factors.
Marine algal taxonomy traditionally relied upon comparison of morphological characters, which remains a critical component of marine macroalgal identification, but can have several disadvantages [24]. For example, features associated with sexual reproduction are often necessary to discern species. Yet, these can be missing from specimens.

References 3 Notes
Sonderophycus Denizot Pihiellales Pihiellaceae      According to O'Kelly et al. [133], this species is likely not found in Hawai'i.  [14,22] According to O'Kelly et al. [133], this species is likely not found in Hawai'i.

U. reticulata
Forsskål N Y N N N [9,14,22] According to O'Kelly et al. [133], this species is likely not found in Hawai'i.
U. rigida C.Agardh N Y N N N [14,22] According to O'Kelly et al. [133], this species is likely not found in Hawai'i. According to O'Kelly et al. [133], this species is likely not found in Hawai'i.  [9,135] Due to seasonality of reproduction, or simply the infrequent nature of these events, many marine algae have heteromorphic life histories, with morphologically distinct gametophytic and sporophytic phases, and the full suite of relevant morphological characters may not be available in the specimens at hand for identification. Moreover, in recent years there have been many demonstrated instances of cryptic and pseudocryptic speciation, e.g., [136,137], as well as suspected incipient speciation-e.g., [52,138], which can provide other scenarios where reliance on morphological features can yield misidentification. Given these pitfalls of employing a strictly morphological approach to identification, greater emphasis has been placed on incorporating molecular comparisons into taxonomic identifications over the past several decades (e.g., as exemplified by recent systematic work on the brown algal genus Lobophora; [139][140][141]), and this is reflected in the degree of confidence in taxonomic identification that can be discerned in the current list.
The Hawaiian marine algal flora is presently comprised of 661 subgeneric taxa (652 species), which compares to the approximately 515 taxa reported for the red, green, and brown Hawaiian marine algal flora by Abbott [12] and Abbott and Huisman [14] during the most recent comprehensive compilations. Relative to other regions, the Hawaiian flora is moderately rich: it compares to the 442 species recorded from Madagascar [23], 425 from French Polynesia [142], 522 from north-western Australia [143,144], 900 from New Zealand [145], and 850 from South Africa [146]. Endemism in the Hawaiian flora is relatively low (13.1%) when compared to other organismal groups, for example: Hawaiian flowering plants (90%) and ferns (about 70% [147]). Vieira et al. [23] reported that the Malagasy seaweeds are 6.5% endemic based on their analyses to date; this is much lower compared to the reported endemism rates for Malagasy terrestrial flora and fauna (37-100%). They also noted that more comprehensive molecular surveys and cross-referencing of records are needed to clarify these figures. Indeed, the Hawaiian inventory includes several records that may be excluded or modified in the future as molecular frameworks (bolstered by intensive systematic study of smaller groups of taxa) clarify the names included here (e.g., for many of the Corallinales).
The Hawaiian archipelago is well known as a center of high endemism and biological uniqueness [2], and an urgent need exists to document the biodiversity of this unique island chain in the face of threats from habitat loss, on-going alien species introductions, invasive species and land-derived pollutants [148], changing coastlines with shoreline development, bloom-forming algae encroaching on shoreline habitat, climate change, corresponding sea-level rise and loss of coral reefs due to depth, and temperature and salinity changes [148,149]. Documentation of the baseline diversity of Hawai'i's marine algae and seagrasses is necessary to monitor and assess new arrivals to the State. The Hawaiian Islands are an extremely isolated island chain, clearly the most isolated on the planet, and the relatively dense human population relies heavily on imported material goods, which primarily arrive via shipping. Although research in this area has not been exhaustive, the results from a single survey of hull-fouled ships raise cause for concern. Godwin [150] surveyed eight maritime vessels on the island of O'ahu for hull-fouling organisms and reported 14 species of red algae, nine green algae and three brown algae (a total of 26 species), of which only nine (35%) were native to the Hawaiian Islands. If these results are extrapolated to the number of vessels traveling to the Hawaiian Islands, then the potential for new algal introductions is truly staggering. At present, new records are often reported without knowledge of the vector of introduction (e.g., Ulva ohnoi, a "green tide" species, was reported from the coastlines of Hawai'i for the first time in 2010 [133]), but building knowledge of the current macroalgal diversity, especially within a molecular context, will allow future introductions to be assessed more easily and accurately.

Conclusions
This compilation of 661 Hawaiian marine algae and seagrasses provides a point-intime summary that includes a 27% increase over the last compilations from approximately 20 years ago. The fields of taxonomy and systematics have revolutionized during that interval due to the near-ubiquitous incorporation of molecular analyses, which have supported numerous new descriptions at almost all taxonomic levels, confirmation of previous morphology-based identifications, and new taxonomic combinations. With the inclusion of information about "degree of confidence" in identification derived from the use of molecular data and type specimen comparisons, we aim to bring a new level of utility to the taxonomic inventory and enable future researchers to have a solid understanding of the basis for application of each taxonomic name to the Hawaiian flora. In the face of numerous threats to biodiversity in the coming decades, it is hoped that critical inventories, such as these, will provide baseline data sets against which future changes may be compared.