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\begin{document}

\title{Range Expansion and First Observation of Tridacna noae in American S\selectlanguage{polish}ā\selectlanguage{english}moa\selectlanguage{english}}



\author[1]{Paolo Marra-Biggs}%
\author[2]{James Fatherree}%
\author[3]{Alison Green}%
\author[1]{Rob Toonen}%
\affil[1]{University of Hawai'i at Mānoa Hawaiʻi Institute of Marine Biology}%
\affil[2]{Hillsborough Community College}%
\affil[3]{Alison Green Marine, Gold Coast Australia}%


\vspace{-1em}



  
  \date{May 24, 2024}


\begingroup
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\selectlanguage{english}
\begin{abstract}
Giant clams are ecologically important, benefitting species of all
trophic levels. But numerous populations have declined drastically in
numbers due to past intensive exploitation that led to their listing in
both CITES Appendix II and IUCN Red List of Threatened Species..
However, giant clams are notoriously difficult to identify, and recent
molecular work has revealed that morphological misidentification of
giant clams have confounded current population assessments and
extinction risk. ~The most recent study of the status of giant clams in
the Samoan Archipelago was published in a journal over 20 years ago,
without molecular corroboration of visual identifications. Using
morphologic characteristics and ezRAD genetic techniques, we identify
the existence of \emph{Tridacna noae} in the Samoan Archipelago,
presenting the first observation and a resulting range expansion.
Accurately identifying the extant species in the archipelago is the
first step towards a much-needed population status assessment to
effectively manage these long-lived species.%
\end{abstract}%



\sloppy


\selectlanguage{polish}\textbf{Range Expansion and First Observation of \emph{Tridacna noae} in
American Sāmoa}\selectlanguage{english}

Paolo Marra-Biggs \textsuperscript{1}, James
Fatherree\textsuperscript{2}, Alison Green \textsuperscript{3}, and
Robert J. Toonen \textsuperscript{1}

{[}1{]} Hawai`i Institute of Marine Biology, University of Hawai`i at
M\selectlanguage{polish}ā\selectlanguage{english}noa, K\selectlanguage{polish}ā\selectlanguage{english}ne`ohe, HI, USA

{[}2{]} Hillsborough Community College (Tampa, Florida)

{[}3{]} Alison Green Marine, Gold Coast Australia

\emph{Corresponding Author:}

Paolo Marra-Biggs

Email address: PaoloMB@Hawaii.edu

Phone Contact: (831) 254 -- 3780

\textbf{Keywords:}

Tridacninae, giant clams, conservation, mitogenome,

\textbf{Abstract:}

Giant clams are ecologically important, benefitting species of all
trophic levels. But numerous populations have declined drastically in
numbers due to past intensive exploitation that led to their listing in
both CITES Appendix II and IUCN Red List of Threatened Species..
However, giant clams are notoriously difficult to identify, and recent
molecular work has revealed that morphological misidentification of
giant clams have confounded current population assessments and
extinction risk. ~The most recent study of the status of giant clams in
the Samoan Archipelago was published in a journal over 20 years ago,
without molecular corroboration of visual identifications. Using
morphologic characteristics and ezRAD genetic techniques, we identify
the existence of \emph{Tridacna noae} in the Samoan Archipelago,
presenting the first observation and a resulting range expansion.
Accurately identifying the extant species in the archipelago is the
first step towards a much-needed population status assessment to
effectively manage these long-lived species.

Giant clams of the subfamily Tridacninae are conspicuous members of
Pacific coral reef communities that play such important ecological roles
that some authors have considered them as keystone species (Guibert et
al., 2020; Mamat et al., 2021). All species of Tridacninae are of
widespread conservation concern and are listed on both the Convention on
International Trade in Endangered Species of Wild Fauna and Flora
(CITES) and the International Union for Conservation of Nature (IUCN).
However, giant clams are notoriously difficult to identify, and recent
molecular work has revealed that morphological misidentification of
giant clams have confounded current population assessments and
extinction risk (Neo et al., 2017; Tisdell, 1989; Wells, 1997). As a
case in point, Tridacna noae that was first described by C. R\selectlanguage{ngerman}öding in
1798 became synonymized to T. maxima as it appeared to be a variant of
the latter species based on shell morphology (Rosewater, 1965). Only
recently has \emph{T. noae} been resurrected as a valid species distinct
from its congener \emph{T. maxima} based on both morphological and
molecular data (Su, 2014).

Neo et al. (2017) present the most comprehensive survey of giant clam
species distributions and status, but no samples from the Samoan
archipelago were included in that study. In previous studies by Fauvelot
et al., 2019, a team sampled Upolu, Independent S\selectlanguage{polish}ā\selectlanguage{english}moa finding
individuals belonging to \emph{T. noae}. However, the most recent
published studies conducted on tridacnine clams in the territory of
American S\selectlanguage{polish}ā\selectlanguage{english}moa were published nearly 20 years ago (Green \& Craig,
1999). This study was conducted prior to recent genetic recognition of
\emph{T. noae} (Su et al., 2014), so the authors assumed all visually
similar samples were \emph{T. maxima} (A. Green, pers. obs.), which
would confound population density estimates if \emph{T. noae} were also
present in American S\selectlanguage{polish}ā\selectlanguage{english}moa. As genetic techniques are more commonly used
to confirm species, more cases of \emph{T. noae} are being identified,
as seen in Militz et al. (2015).

Previously, American S\selectlanguage{polish}ā\selectlanguage{english}moa hosted two species of giant clams in the
territorial waters: \emph{Tridacna squamosa} and \emph{T. maxima}.
Fossilized shells indicate a third species, \emph{Hippopus hippopus},
used to occur in S\selectlanguage{polish}ā\selectlanguage{english}moa, but is now locally extinct (Nagaoka, 1993;
Newman \& Gomez, 2000). Additionally, an aquaculture program was started
for juvenile \emph{H. hippopus} \emph{T. gigas,} and \emph{T. derasa},
but stocks were harvested prior to reproduction and appear to be
functionally extirpated. Here, we use next generation sequencing to
confirm identification of giant clams and expand the species range of
\emph{T. noae} to include American S\selectlanguage{polish}ā\selectlanguage{english}moa. Additionally, this discovery
highlights the need to revisit historical population assessments to
determine the distribution and abundance of each \emph{T. noae} and
\emph{T. maxima} across the Samoan Archipelago.

Tissue biopsies were collected from four clams, two putative \emph{T.
noae} and two putative \emph{T. maxima,} found during surveys on
Tutuila, American S\selectlanguage{polish}ā\selectlanguage{english}moa, with \emph{in situ} photos taken of each
individual sampled, seen in figure 1. Clams were sampled from depths
between 27-37 ft and ranged from 12 to 20 cm in antero-posterior shell
length. Clams were identified morphologically as either \emph{T. noae}
or \emph{T. maxima} (following Militz et al. 2015; Fatherree 2016) and
then sequenced using reduced representation genomic sequencing (ezRAD,
Toonen et al., 2013). Reads were mapped to reference mitogenomes from
Tan et al. (2022) using Minimap2 (Li, 2018) in Geneious Prime v2021.1.1
(www.geneious.com).

The two individuals identified morphologically as \emph{T. noae} (Figure
1) were confirmed genetically with nearly complete mitogenomes 15,240 \selectlanguage{ngerman}±
4 base pairs in length (86.5 ± 1.2\%), and with high coverage depth
(106.2 ± 75X), that matched the Tan et al. (2022) reference sequence
provided by Danwei Huang at 99.7 ± 0.05\% pairwise identity.~ Another
individual identified morphologically as \emph{T. maxima} was also
confirmed genetically with a nearly complete mitogenome of 14,978 base
pairs (80.7\%), with relatively high coverage depth (53.5X), matching at
99.6\% pairwise identity to the \emph{T. maxima} reference sequence (E.
Y. Tan et al., 2022).~ Given this clear distinction among the
morphologically identified samples, the final \emph{T. maxima} sample
was not sequenced here.

~

There is considerable morphological variation within American Samoan
\emph{T. maxima} (Figure 1B and C) and \emph{T. noae}. The morphological
characteristics that are typically used for identification (such as the
size of byssal orifice, number of radial folds in the rib interstices,
elaborate vs. simple incurrent siphon tentacles, size of scutes) are
variable within the species which can lead to confusion or
misidentification between \emph{T. maxima} and \emph{T. noae}.
Additionally, the mantle can vary dramatically in color, pattern and
texture, and are unreliable as proxies for species identification (Su et
al., 2014). As with the many phenotypic morphs of \emph{Tridacna
maxima}, \emph{T. noae} can exhibit two color morphs, brown and blue
(Militz et al. 2015).

Many of these characteristics also fluctuate based on individual health
(Mies, 2019). However, certain morphologic characteristics can aid in
distinguishing between \emph{T. maxima} and \emph{T. noae}. Examples of
features more commonly observed in \emph{T. noae} are ``tear-drop'' oval
patterns on the mantle edge and less pronounced hyaline organs (Su et
al., 2014).

Based on these findings, we can update the catalog of extant species in
American S\selectlanguage{polish}ā\selectlanguage{english}moa to include: \emph{Tridacna maxima}, \emph{Tridacna
squamosa}, and now \emph{Tridacna noae}. By confirming the
identification of these two sometimes difficult to distinguish clam
species genetically along with photo-documentation to highlight
morphological distinctions, we hope to improve the efficacy and accuracy
of identification of \emph{T. noae} and \emph{T. maxima} during future
field surveys. Tridacnid clams are listed as a priority species under
the territory's Local Action Strategy, and accurately identifying
species is critical to understanding the distribution and abundance of
each and can help ensure the appropriate and most effective management
of these long-lived species.\selectlanguage{english}
\begin{figure}[H]
\begin{center}
\includegraphics[width=0.98\columnwidth]{figures/Figure-1-Clam-comparison/Figure-1-Clam-comparison}
\caption{\selectlanguage{polish}{Tridacna noae (A, B) from Tutuila, American Sāmoa, showing the
teardrop-shaped spots on the edge of the mantle with the typical golden
ring, and the lack of a distinct row of eyes (or hyaline organs).
Tridacna maxima (C and D) shows considerable variation in the color on
the mantle, but a row of eyes along the periphery of the mantle is a
distinctive character. (C) T. maxima and (D) a blue variant of T. maxima
from Tutuila, American Sāmoa for comparison highlighting the row of eyes
at the upper mantle's margin and lacking the ringed spots.
{\label{423902}}
Photos: Paolo Marra-Biggs%
}\selectlanguage{english}}
\end{center}
\end{figure}\selectlanguage{english}
\begin{figure}[H]
\begin{center}
\includegraphics[width=0.98\columnwidth]{figures/Fig-2---TNoae-Range/Fig-2---TNoae-Range}

\end{center}
\end{figure}

\textbf{Acknowledgements:}

Many thanks to the numerous collaborators on this project including: the
National Park of American S\selectlanguage{polish}ā\selectlanguage{english}moa, the National Marine Sanctuary of
American S\selectlanguage{polish}ā\selectlanguage{english}moa, USFWS, the Department of Marine and Wildlife Resources,
and the Coral Reef Advisory Group. Thank you to members of the ToBo lab
for expertise, guidance, and advice: ~Brian Bowen, Ingrid Knapp, Mareike
Sudek, Chris Suchocki, Sean Canfield, Emily Conklin, Evan Barba, and
Xander Velkeneers. Additional thanks to Danwei Huang for sharing
additional sequence data, used in the reference mapping of these
samples.

This project was funded by the Hawai'\selectlanguage{english}i Sea Grant and the Pacific Islands
Climate Adaptations Science Center, and the Colonel Willys E. Lord and
Sandina L. Lord Scholarship, Ruth D. Gates Scholarship, Robinson
Fellowship, and Watson T. Yoshimoto Fellowship.

\par\null

\textbf{Compliance:}

Sampling of giant clams were conducted under guidance of the National
Park of American S\selectlanguage{polish}ā\selectlanguage{english}moa and under the scientific permit of American
Samoan Government: Department of Marine and Wildlife, permit NO.
DMWR-2016/046.

\par\null

\textbf{Genetic Sequences:}

The sequences are submitted to the Genbank Short Read Archives, under
the Bioproject ID PRJNA867692 and the SRA biosamples accession codes
are: SAMN30219961, SAMN30219962, and SAMN30219963.

\textbf{}

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\par\null

\selectlanguage{english}
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