Research Article |
Corresponding author: Cecili B. Mendes ( cecilimendes@gmail.com ) Academic editor: Jon Norenburg
© 2021 Cecili B. Mendes, Paul Delaney, James M. Turbeville, Terra Hiebert, Svetlana Maslakova.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation:
Mendes CB, Delaney P, Turbeville JM, Hiebert T, Maslakova S (2021) Redescription of Emplectonema viride – a ubiquitous intertidal hoplonemertean found along the West Coast of North America. ZooKeys 1031: 1-17. https://doi.org/10.3897/zookeys.1031.59361
|
Emplectonema viride Stimpson, 1857, a barnacle predator, is one of the most common and conspicuous intertidal nemerteans found along the West Coast of North America from Alaska to California, but it is currently referred to by the wrong name. Briefly described without designation of type material or illustrations, the species was synonymized with the Atlantic look-alike, Emplectonema gracile (Johnston, 1837) by Coe. Here we present morphological and molecular evidence that E. viride is distinct from E. gracile. The two species exhibit differences in color of live specimens and egg size and are clearly differentiated with species delimitation analyses based on sequences of the partial regions of the 16S rRNA and cytochrome c oxidase subunit I genes. In order to improve nomenclatural stability, we re-describe E. viride based on specimens from the southern coast of Oregon and discuss which species should be the type species of the genus. Emplectonema viride was one of the two species originally included in the genus Emplectonema Stimpson, 1857, but subsequent synonymization of E. viride with E. gracile resulted in acceptance of the Atlantic species, E. gracile, as the type species of the genus. We resurrect E. viride Stimpson, 1857 and following Corrêa’s designation, this should be the type species of the genus Emplectonema.
Cryptic species, marine diversity, Nemertea, species delimitation
The genus Emplectonema was established by
The original description of E. viride is but a few lines in Latin: “Corpus depressum, lineare v. proteum, supra viride, subtis album. Caput subdiscretum, marginibus albis; foveis elongatis bipartitis; fronte emarginata. Ocellorum acervi quattuor; posteriores distincti, rotundati, ocellis confertis; anteriores marginales juxta foveas, ocellis sparsis. Long. 11; lat. 0.05 poll. Hab. In portu ‘San Francisco;’ littoralis inter lapillos” (
Emplectonema gracile was first described as Nemertes gracile by
The species currently recognized as E. gracile is listed as having a wide geographic distribution in the Northern Hemisphere, including Japan (Hokkaido), Russia (Kamchatka Peninsula), the Aleutian Islands, the Atlantic and Pacific coast of North America, northern coasts of Europe, Mediterranean, the Romanian coast of the Black Sea, and Madeira (
Here we present molecular and morphological evidence that E. viride is a separate species from E. gracile. We compare the two cryptic species and re-describe E. viride, the type species of the genus Emplectonema, as designated by
Clusters of acorn barnacles, typically Balanus glandula Darwin, 1854, were collected from intertidal zones at two locations in southern Oregon (Oregon Scientific Take Permits #22780 and 23609) in 2019 and 2020 by C. Mendes and S. Maslakova (Table
Live worms were photographed with external flash using a Canon Eos 5D Mark III. For close ups, worms were anesthetized with a mixture of 1:1 MgCl2 and seawater. Anterior end and proboscis were removed, gently compressed between a glass slide and a cover slip, and photographed using a Spot 5.2 camera mounted on an Olympus BX51 equipped with DIC optics. Eggs, sperm, and larval stages were photographed similarly. The anterior region of each morphological voucher was fixed in 10% formalin, post-fixed in Bouin’s solution, and stored in 70% ethanol. The posterior region was preserved in 95% ethanol and kept at -20 °C until DNA extraction.
Genomic DNA was extracted with DNEasy Blood and Tissue kit (Qiagen) following the manufacturer’s protocol. Partial regions of cytochrome c oxidase subunit I (COI) and 16S ribosomal DNA (16S rRNA) were amplified using the primer pairs in Table
Consensus sequences were aligned in the online version of Mafft software v. 7 (
Independent phylogenetic analyses from each gene region apportioned the specimens into two main clades with high support, corresponding to sampling location. Specimens from the Pacific Ocean (Emplectonema viride) form one clade, and specimens from the Atlantic Ocean and North Sea (Emplectonema gracile) form another (Fig.
Resulting trees from the Maximum Likelihood analysis with RAxML. A: COI phylogeny (lnL = -1520.573862). B: 16S rRNA phylogeny (lnL = -909.477668). Support values above 50 presented in each branch. Branch in purple comprises specimens of Emplectonema gracile. Branch in blue comprises specimens of Emplectonema viride.
The haplotype networks show a low diversity with many mutational steps (85 for COI and 33 for 16S rRNA) between specimens from the Pacific and Atlantic Oceans (Fig.
Order Monostilifera Brinkmann, 1917
Suborder Eumonostilifera Chernyshev, 2003
Family Emplectonematidae Bürger, 1904
Emplectonema viride Stimpson, 1857: 163;
Emplectonema gracile
Seven adults from Charleston Marina, OR (43°20.63'N, 124°19.38'W); 27 Nov. 2019; collected from wooden pilings among acorn barnacles, Balanus glandula (Table
Three females and four non-sexed adults of E. gracile from Pawleys Island, SC (33°24.63'N, 79°7.88'W); 29 Nov. 2019; among scorched mussels, Brachidontes exustus on granite rocks; GenBank and NMNH accession numbers in Table
Sampling locations, specimen ID, and accession numbers. Morphological vouchers listed in bold. † Sequences previously available in GenBank.
Species | Abbreviation | Sampling location | NMNH # | GenBank accession COI | GenBank accession 16S rRNA |
---|---|---|---|---|---|
Emplectonema viride | CH_OR_1_E_viride | OIMB Boathouse dock, OR | – | MT649099 | MT647808 |
CH_OR_2_E_viride | – | MT649110 | MT647809 | ||
CH_OR_3_E_viride | – | MT649101 | MT647812 | ||
CH_OR_4_E_viride | – | MT649102 | MT647811 | ||
CH_OR_5_E_viride | – | MT649109 | MT647814 | ||
CH_OR_6_E_viride | – | MT649100 | MT647815 | ||
CH_OR_7_E_viride | – | MT649103 | MT647816 | ||
CH_OR_8_E_viride | – | MT649104 | MT647807 | ||
CH_OR_9_E_viride | – | MT649105 | MT647818 | ||
CH_OR_10_E_viride | – | MT649106 | MT647817 | ||
CH_OR_11_E_viride | OIMB Boathouse dock, OR |
|
MT649107 | MT647813 | |
CH_OR_12_E_viride | Charleston Marina, OR |
|
MT649114 | MT647820 | |
CH_OR_13_E_viride |
|
MT649115 | MT647810 | ||
CH_OR_14_E_viride |
|
MT649108 | MT647821 | ||
CH_OR_15_E_viride |
|
MT649111 | MT647824 | ||
CH_OR_16_E_viride |
|
MT649116 | MT647823 | ||
CH_OR_17_E_viride |
|
MT649112 | MT647819 | ||
CH_OR_18_E_viride |
|
MT649113 | MT647822 | ||
E4H2 | Charleston, OR | – | KU197596† | KU197260† | |
E5B5 | – | KU197597† | KU197261† | ||
E5B6 | – | KU197598† | KU197262† | ||
E5B7 | – | KU197599† | KU197263† | ||
Emplectonema gracile | PI_SC_1_E_gracile | Pawleys Island, SC | – | MT649119 | MT647832 |
PI_SC_2_E_gracile | – | MT649121 | MT647827 | ||
PI_SC_3_E_gracile | – | MT649127 | MT647825 | ||
PI_SC_4_E_gracile | – | MT649117 | MT647830 | ||
PI_SC_5_E_gracile | – | MT649124 | MT647829 | ||
PI_SC_6_E_gracile | – | MT649125 | MT647834 | ||
PI_SC_7_E_gracile | Pawleys Island, SC |
|
MT649126 | MT647828 | |
PI_SC_8_E_gracile |
|
MT649122 | MT647835 | ||
PI_SC_9_E_gracile |
|
MT649123 | MT647826 | ||
PI_SC_10_E_gracile |
|
MT649118 | MT647831 | ||
PI_SC_11_E_gracile |
|
MT649120 | MT647833 | ||
PI_SC_12_E_gracile |
|
– | – | ||
PI_SC_13_E_gracile |
|
– | – | ||
PI_SC_14_E_gracile |
|
– | – | ||
– | Salcombe, UK | – | AJ436903† | AJ436793† | |
DNA10615 | Crosby, UK | – | HQ848620† | JF277621† | |
NemBar0378 | Sweden | – | KU839979† | – | |
NemBar0400 | – | KU839991† | – | ||
NemBar0401 | – | KU839992† | – | ||
NemBar0402 | – | KU839993† | – | ||
NemBar0403 | – | KU839994† | – | ||
NemBar0404 | – | KU839995† | – | ||
K21 | Spain | – | KU697656† |
Gene | Forward primer | Reverse primer | Reference |
---|---|---|---|
COI | HCO1490 – GGTCAACAAATCATAAAGATATTGG | LCO2198 – AAACTTCAGGGTGACCAAAAAATCA |
|
16S rRNA | 16SARL – CGCCTGTTTATCAAAAACAT | 16SBRH – CCGGTCTGAACTCAGATCACGT |
|
16S rRNA | 16SKr – AATAGATAGAAACCAACCTGGC | Jon Norenburg unpublished |
Based on specimens from Oregon, body long and thread-like, 35–103 mm long, 0.6–1.0 mm wide. However,
Rhynchocoel is short, approximately 1/3 of body length. Central stylet slightly curved, 170–326 μm long (n = 7), smooth. Basis is slender, 480–815 μm long (n = 7), its distal end abruptly widening into a truncated bulb (Fig.
Reproductive individuals of E. viride were collected in Charleston, OR, in September 2009, October 2019, January 2020, and June 2020. When ripe, males and females free-spawn gametes into the water, with no known reliable spawning cue. Swimming larvae hatch from the egg chorion after ~30 h and begin feeding on small planktonic crustaceans after developing a functional proboscis and stylet (~4 d). Planktonic period lasts several months (Mendes unpublished observations). Wild-caught larvae of E. viride were found in the plankton samples taken with 50–153 μm net at the Charleston Marina, OR, in October 2013, March 2019, February 2020, and June 2020. Emplectonema viride larvae are easily recognized by their distinctive green color (Fig.
Larva of Emplectonema viride collected from plankton in Charleston, OR, on 17 Oct 2013. Same individual is shown in two focal planes to highlight apical tuft (upper left, A) and posterior cirrus (lower right, A) and green epidermal pigment (B). Note paired subepidermal eyes, which are anterior to cerebral organs. Scale bar: 100 μm.
Northeastern Pacific Ocean from Alaska to California. Type locality is San Francisco Bay, California, USA.
As has been pointed out by
The simple morphology of nemertean worms makes it notoriously difficult to identify species, and the use of DNA sequence data as well as gamete morphology can help differentiate between morphologically cryptic nemertean species (e.g.,
The combination of molecular and morphological data presented here confirms the existence of two cryptic species of North American Emplectonema, one from the Pacific and another from the Atlantic coast. Our results support the validity of E. viride described from the Pacific coast (
The genus Emplectonema was established by
DNA-based phylogenies (18S rRNA, COI) suggest that Emplectonema neesii (Örsted, 1844) is not closely related to E. gracile (
This study was partially funded by São Paulo Research Foundation (FAPESP) grant 2019/10375-8 to CM. TCH was partly supported by the NSF grant OCE-1030453 to Dr Craig Young and SM while carrying out work related to this manuscript. The authors are also thankful to Dr Hiroshi Kajihara, Dr Irina Cherneva, and Dr Jon Norenburg for the helpful comments during the revision of this manuscript.
Table S1
Data type: occurrence
Explanation note: Linked data table of specimens and sequences analyzed in this study. Abbreviations. COI: Cytochrome oxidase I gene fragment; 16S rRNA: 16S ribosomal RNA; †: Sequences previously available in GenBank. Specimens morphologically analyzed marked in bold.
Figure S2
Data type: phylogenetic
Explanation note: Species delimitation results from PTP for COI (A) and 16S rRNA (B) gene regions. Values on branches are posterior probabilities of those taxa form one species under the PTP model and a flat prior. Red branches indicate taxa that should be considered as part of the same lineage.