THE NAUTILUS

Volume 128, Number 4

December 31, 2014

ISSN 0028-1344

A quarterly devoted

to malacology.

Ql

801

.03/8

rz

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Dr. Jose H. Leal

The Bailey- Matthews Shell Museum

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National Museum of

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Department of I nvertebrates

Field Museum of

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THE0NAUTILUS

Volume 128, Number 4

December 31, 2014

ISSN 0028-1344

CONTENTS

Anabela Lopes Functional studies on the shell soluble matrix of Anodonta cygnea

Mariana Hinzmann (Bivalvia: Unionidae) 105

Iulius Bobos

Manuel Lopes- Lima

Jose Fernando Gonsalves

Sergio Ferreira

Bernardo Domingues

Jorge Machado

David W. Behrens Two new species of Velutinidae Gray, 1840 (Gastropoda) from the

Elysse Ornelas North Pacific with a preliminary molecular phytogeny of the family 114

Angel Valdes

Kazutaka Amano A new genus of Buccinoidea (Gastropoda) from Paleocene deposits

Anton Oleinik in eastern Hokaido, Japan 122

Bonald G. Noseworthy First report of Cryptoplax proprior Is. and Iw. Taki, 1930

Hee-Jung Lee (Polyplacophora: Cryptoplacidae) in Korea 129

Young-Chul Kang

Sukgeun Jung

Hyung-Kee Cha

Kwang-Sik Choi

Notice

135

Author Index

137

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16. Electronic Copy Circulation

THE NAUTILUS 128(4): 105-1 13, 2014

Page 105

Functional studies on the shell soluble matrix of Anodonta cijgnea

(Bivalvia: Unionidae)

Anabela Lopes

Mariana, Hinzmann

ICBAS-UP - Institute de Ciencias Biomedicas Abel Salazar

Universidade do Porto

Rua de Jorge Viterbo Ferreira, 228

4050-313 Porto, PORTUGAL

and

CIIMAR-LA - Centro Interdisciplinar de Investigate Marinha

e Ambiental

Universidade do Porto

Rua dos Bragas, 289

4050-123, Porto, PORTUGAL

Manuel Lopes-Lima

Jose Fernando Gonsalves

Sergio Ferreira

Bernardo Domingues

ICBAS-UP - Institute de Ciencias Biomedicas Abel Salazar

Universidade do Porto

Rua de Jorge Viterbo Ferreira, 228

4050-313 Porto, PORTUGAL

and

CIIMAR-LA - Centro Interdisciplinar de Investigate Marinha

e Ambiental

Universidade do Porto

Rua dos Bragas, 289

4050-123, Porto, PORTUGAL

Iulius Bobos

FCUP - Faculdade de Ciencias da Universidade do Porto

Rua Campo Alegre, 687

4169-007 Porto, Portugal

Jorge Machado1

ICBAS-UP - Institute de Ciencias Biomedicas Abel Salazar

Universidade do Porto

Rua de Jorge Viterbo Ferreira, 228

4050-313 Porto, PORTUGAL

and

CIIMAR-LA - Centro Interdisciplinar de Investigate Marinha

e Ambiental

Universidade do Porto

Rua dos Bragas, 289

4050-123, Porto, PORTUGAL

and

LABIOMEP - Biomechanics Laboratory

University of Porto

Rua Doutor Placido da Costa

4200-450 Porto, PORTUGAL

[email protected]

ABSTRACT

The biomineralization process in molluscan shells is controlled

by an extracellular organic matrix, embedded in a fluid, pro-

duced by the calcifying outer mantle epithelium (OME) and

secreted within the extrapallial compartment.

In the present work, the study subject is the nacreous layer of

the freshwater bivalve Anodonta cijgnea and the functional role

of its organic matrix, which is still a large field to explore. From

the organic matrix it was possible to extract two fractions, but

only the soluble fraction was studied. Different techniques

were used, including biochemical protein characterization by

electrophoresis for the extrapallial fluid and the shell, quantifi-

cation and detection of the matrix proteins and glycosamy-

noglycans (GAGs) directly in the shell, through immunogold

techniques, using SEM and ATR-IR observations. Seven pro-

tein fractions in both extrapallial fluid and shell were detected

by electrophoretic analysis with molecular weights of approxi-

mately 102/106, 76/74, 66/66, 60/52, 45/43, 35/35 and 31/29 KDa,

respectively. This may suggest a narrow functional correlation

1 Author for correspondence

between specific proteins from the extrapallial fluid and the

shell. Despite the low percentage of the organic matrix relative

to the whole nacreous shell, it was observed that it is mainly

composed of proteins (13.40-23.32 mg/ml) and GAGs (2.50-

3.12 mg/ml), which appear to be very relevant on the micro-

structure and polymorphism organization of the major calcium

carbonate fraction. In agreement, the immunogold technique

showed that the shell organic matrix is mainly intercrystalline.

Additionally, the common detection by infrared spectroscopy

of amide groups on both soluble shell matrix and solid shell

crystal fraction suggests that this molecule is one of the intra-

crystalline inductors of the aragonite crystals formation in the

nacreous layer of A. cijgnea.

Additional Keywords: biomineralization, extrapallial fluids,

organic matrix

INTRODUCTION

The shell structure in the molluscan shell carries a good

historical record that helps explain the evolution of this

Page 106

THE NAUTILUS, Vol. 128, No. 4

group of animals since the Cambrian period. Shells are

secreted by a vast majority of the estimated 70,000-

76,000 named molluscan species (estimate in Rosenberg,

2014) and their construction begins in the early stages of

development and almost continuously during their entire

life (Boggikl, 1930; Marin and Luquet, 2004). The shell

crystals in bivalves show a great variety of morphologies

and organization levels originating different microstruc-

tures and polymorphisms (Checa et ah, 2007; Lopes-

Lima et ah, 2010). However, all of them are constituted

by calcium carbonate representing 95-99% of the shell

and by 1-5% corresponding to organic matrix (Duplat

et ah, 2006). Adult shells are highly variable being built

up of one or more shell layers, each of which may have

a different microstructure (Boggild, 1930; Weiss et ah

2002). In a longitudinal section, the Unionidae shell reveals

generally two calcified layers, prismatic and nacreous

layers, and one outer organic layer, the periostracum,

which protects the calcified layers from water dissolution

(Boggild, 1930; Moura et ah 2003; Marin and Luquet,

2004). The most common mineral polymorphs identified

in the calcified layers of calcium carbonate are aragonite

and/or calcite (Weiner, 1983; Checa et ah, 2007). In the

families Pinnidae and Pteriidae the shell has one internal

layer with calcium carbonate in the aragonite form and

another external in the calcite form whereas in the

Unionidae family the two calcified layers are both arago-

nitic (Boggild, 1930; Taylor et ah, 1969; Caiping et ah,

2005; Marie et ah, 2007).

Although the shell calcification process occurs outside

the living tissues, it is neither in contact nor directly

dependent on the external environment. The process is

mainly dependent on three components: a closed com-

partment where the calcification occurs, an ionic mem-

brane transport and an extracellular organic matrix

(Moura et ah, 2003). The closed compartment filled with

extrapallial fluid is bounded by the shell, the periostracum,

and the calcifying outer mantle epithelium (OME). This

isolation is critical to provide a supersaturated environ-

ment which is essential for the formation of crystals

(Marin and Luquet, 2004). The extracellular organic

matrix, secreted by the calcifying epithelium towards the

extrapallial fluid, consists of a complex mixture of pro-

teins, glycoproteins, proteoglycans and chitin (Moura, 2000;

Pereira-Mouries et ah, 2002). This matrix has important

roles in the physical-chemical interactions involved in

crystal nucleation, polymorphic selection, growth, and inhi-

bition (Marxen and Becker, 1997; Levi-Kalisman et ah,

2001; Pereira-Mouries et ah, 2002).

Little is known about the extracellular organic matrix

of the unionid freshwater mussel Anodonta cygnea. The

shell exhibits the three layers already mentioned and,

though the two calcified layers present different micro-

structures, both correspond to the same calcium carbon-

ate polymorph, aragonite. According to Moura et ah

(2000) there are 4—6 protein fractions in the calcifying

fluids of A. cygnea.

Nacreous

layer

Prismatic

layer

Figure 1. SEM images of the

shell of Anodonta cygnea in back-scattered electron inode. (A) The three layers are perfectly distinct

in transversal sections; magnification of prismatic (B) and nacreous (C) layers.

A. Lopes et al., 2014

Page 107

Prismatic

layer

Nacreous

layer

Figure 2. SEM images of the external shell surfaces of

Anodonta cijgnea in hack-scattered electron mode (A). Magni-

fication of prismatic (B) and nacreous (C) layers.

The objective of this work is to study the soluble fraction

of the extracellular organic matrix from the extrapallial

fluid and nacreous layer of the freshwater mussel

Anodonta cijgnea, in order to gain further knowledge

about its components. The main goal was to extract and

purify the organic matrix from the shell in order to quan-

tify and analyze it by electrophoresis and Attenuated Total

Reflectance-Infrared Spectroscopy (ATR-IR). The direct

visualization of the organic matrix presence in the shell

was also an aim of the study, through the use of imimino-

histochemical techniques.

MATERIALS AND METHODS

deoxycholate), followed by the addition of 1/10 of trichlo-

roacetic acid (TCA) 100% and centrifuged for 15 min at

4 °C in a mierofuge at maximum speed (15000 g). For the

SDS-PAGE, the protein pellet was re-suspended in a

minimal volume in PBS buffer.

In this work only the soluble fraction was analyzed.

Two independent replicas of the extraction were accom-

plished. Total protein and glycosaminoglycans concentra-

tions in the shell were determined respectively according

to the methods of Bradford ( 1979) and Whiteman ( 1973).

Protein fractions from the extrapallial fluid and shell

samples and molecular weight standard were separated

and analyzed using a mini-SDS-PAGE system in 8%

polyacrylamide gels at 130V during 60 min and stained

with silver nitrate (Gromova and Celis, 2(X)6).

Peptide Mass Mapping from the Extrapallial Fluid

and Shell Protein Band

The observed protein band from the SDS-PAGE gel was

cut and transferred to the Eppendorf tubes before being

sent to Alphalyse, Inc. (USA) for peptide mass mapping.

From the seven protein bands detected, only the five

marked bands with highest expression were sent for

analysis (Figure 4). The protein samples were reduced

and alkylated with iodoacetamide, i.e., earbamidometh-

ylated, and subsequently digested with trypsin and ehy-

motrypsin. Trypsin cleaves after lysine and arginine

residues. The resulting peptides were spotted directly

onto an anchorchip target or were concentrated on a

CIS ZipTip micropurification column and eluted onto

an anchorchip target for analysis on a Bruker Autoflex

Speed MALDI TOF/TOF instrument. The peptide mix-

ture was analyzed in positive reflector mode for accurate

peptide mass determination (MALDI-MS).

Organic Matrix Extraction, Quantification and

SDS-PAGE Analysis

Freshwater bivalves, Anodonta cijgnea , were collected from

the bottom of Mira Lagoon (40°26.712N, 8°47.817W) in

the end of July 2012. The nacreous layer organic matrix

was extracted as described by Caiping et al. (2005) with

the following exceptions: the initial amount of nacreous

layer sample was higher (90 g), the dialysis was performed

against ultrapure water and, at the end, the sample powder

was re-dissolved in PBS buffer.

The TCA-DOC protein precipitation technique was

used for the extraction of very low contents of soluble

protein from the extrapallial fluid. For this, to the extra-

pallial fluid, was added 1/100 vol. of 2% DOC (sodium

Polyclonal Antibodies Production and Visualization of

the Shell Organic Matrix by Immunocold Technique

The polyclonal antibodies were produced in two rabbits.

For each immunization 100 pg of organic matrix were

injected in the intradermic neck region. The immuniza-

tion procedures were reinforced after 30 and 42 days.

Bleedings were carried at 0 (pre-immune serum), 30, 42,

and 54 days. The sera were then titrated by standard

ELISA assays, for evaluation of the more appropriate

antibody concentration. The third bleeding was then

chosen and used in the following procedures. IgGs were

purified in a protein G column (GE Healthcare) and

used in the immunogold assays. This assay was per-

formed as described by Marin et al. (2007). Briefly, the

Table 1 Quantitative results of organic matrix extraction from the nacreous layer of A. cijgnea.

Page 108

THE NAUTILUS, Vol. 128, No. 4

nacreous layer was broken in small pieces and etched

with EDTA 1% (w/v), pH 7.5 during 2-3 min with agita-

tion. The pieces were then incubated overnight with a

1:3000 dilution of the IgGs produced against the organic

matrix. The secondary antibody used (Anti-rabbit IgG -

Gold antibody produced in goat, affinity isolated anti-

body, aqueous glycerol suspension, 5 nm. Sigma) was

diluted 1:400 and incubated during 2 h. The silver

enhancement was performed with a Silver Enhancer Kit

by Sigma. As a negative control the pieces were first

incubated with pre-immune serum. The results were

observed through scanning electron microscopy (SEM).

Scanning Electron Microscopy Imaging (SEM) of

the Shell and Attenuated Total Reflectance-Infrared

Spectroscopy ( ATR-IR ) of Aqueous Shell Matrix.

Untreated shell pieces were gold-coated (FINE-COAT

Ion sputter JFC-1100) and glued to aluminum stubs for

SEM observations using JEOL [SM-35C scanning elec-

tron microscope operated at 15 keV in Centro de

Materials da Universidade do Porto (CEMUP). Shell

pieces treated with immunogold technique were carbon-

coated and analyzed in back-scattered electron mode

at 15 keV.

Two aqueous samples of soluble organic matrix were

analyzed by ATR-IR using a Bruker Tensor-27 spec-

trometer equipped with a DTGS (deuterated triglycine

sulfate) single detector plate and a horizontal ATR

unit, where a horizontal ZnSe ATR crystal was mounted

at 45° in a 30 ml rectangular cell made of polypropyl-

ene. Samples were run in the frequency range 800-

4000 cm '. Additionally, few milligrams of nacreous

shell layer were extracted from the freshwater mussel

A. cygnea and then were analyzed by infrared spectros-

copy in absorbance mode using a BRUKER Tensor-45

spectrometer. The pellet disks of 1.5 cm diameter were

prepared by mixing I mg of sample with 200 mg KBr and

pressing at 10 Kg/cnrT.

RESULTS AND DISCUSSION

Characterization of the Natural Microstructure

of Anodonta cygnea

SEM observations (Figures 1 and 2) highlight the nature

of the shell microstructure in the freshwater mussel

A. cygnea. The three different layers are evident: one

organic (periostracum) and two calcareous layers (pris-

matic and nacreous). In the prismatic layer the aragonitic

crystals are organized in prisms covered with organic

matrix surrounding them, whereas in the nacreous layer

the crystals are organized with the shape of tablets with

organic matrix between them.

In this work, we have mainly focused on the nacre-

ous layer, specifically in its organic matrix. Regarding

the shell nacreous layer, the quantitative results of its

main organic components are presented in Table 1. As

described in the literature for A. cygnea by Moura et al.

MW E S

KDa

200

1 ^3

Figure 3. S DS-PAGE of the soluble organic matrix extracted

from the extrapallial fluid (E) and shell nacreous layer (S) of

Anodont a cygnea. MW: molecular weight standards; E: Protein

fraction from the extrapallial fluid; S: Protein fraction from

the shell.

A. Lopes et al., 2014

Page 109

(2000) and for other species of bivalves (Marie et al.,

2007), it was also stated that the organic matrix repre-

sents a small amount of the total shell weight. The

majority of this matrix is represented by proteins (13.40-

23.32 mg.ml '), while the glycosaminoglycans (GAGs)

were found in lesser amounts (2.50-3.12 mg.ml ').

Although present in smaller amounts, GAGs are always

found in the organic matrix, denoting their importance in

most biomineralizing systems (Pereira-Mouries et al.,

2002; Moura et al., 2000, 2003; Lopes- lima et al, 2005,

2010). Naturally, the total protein and GAGs contents

were higher in the shell matrix compared to the organic

fluids as reported by Moura et al. (2000) for the same

period. In fact, these results confirm that the shell matrix

structure act as a sponge. Furthermore, while results

of Moura et al. (2000) reported 6 protein fractions on

the haemolymph and extrapallial fluids along the year,

the present study adds complementary data based on the

detection of seven different protein bands in extrapallial

fluid which are similar to odiers in the shell matrix of

A. cygnea (Figure 3).

The SDS-PAGE technique recorded seven protein

fractions with molecular weights bands of approximately

102-106, 76-74, 66-66, 60-52, 45-43, 35-35, and 31-29 KDa,

Extrapallial Samples (E)

Molecular

Weight

Standard

Shelf Samples (S)

Molecular Shell

weight

Standard

3 8 sM| S |

i i 9 a s S a ?

SI

i ? % *

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s ?

- S s

V 'AiMoFMn Spe«xtDlGESTM)GEST0700^T9aDCESTO7«no,enniSffcl

S2

• e ’'O =>’’ j| , o ^ j

a ii^ ** 5 * a s - i 2 8

a IS I ^ 7 5 La s£ "I .253 j £

S3 l

s 2 =

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« Spood\DlGeSTVOIGESTO7O(WJ79»DGEST0757\0.G

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S4

< 'Auto Flex Sp«xM)lGESl\D*3eSTOra0-0n»»OGEST0787N0 K?

S5

Figure 4. Diagram of a spectra of peptides mass mapping of soluble organic matrix extracted from the extrapallial fluid (E1-E5)

and shell nacreous layer (S1-S5) of A. cygnea by MALDI-MS determination.

Page 110

THE NAUTILUS, Vol. 128, No. 4

respectively in the extrapallial fluids and in the shell

matrix (Figure 3). However, from these seven hands, only

five were submitted for peptide mass analysis, since the

two heaviest (74-106 KDa) showed poor resolution.

According to Moura et al. (2000), the protein bands in

the fluids can show different expression levels along the

year, which can be correlated with its own functional role

in the shell. Similar studies by electrophoresis have been

previously performed in at least three families of mol-

lusks, two of them in the Bivalvia, and the number of

protein fractions found was also low (Marxen and Becker,

1997; Pereira-Mouries et al., 2002; Caiping et al., 2005;

Marie et al., 2007). Yet, regarding the number of bands,

these results revealed great conformity with previous

studies, which state between 4 to 6 bands, depending on

the mussel species (Misogianes and Chasteen, 1979;

Keith et al., 1993; Moura et al., 2000). Curiously, the

new data showed that the seven protein fractions pres-

ent close molecular weight both on the shell matrix and

extrapallial fluid samples. This finding may point out

that specific proteins in the extrapallial fluid are involved

on the shell biomineralization process in A. cygnea.

Additionally, the MALDI-MS analysis revealed a large

number of peptides per protein band in all extrapallial

and shell fractions. There are similarities in the peptide

mass from equivalent protein bands among different

extrapallial samples and the same occurs in the shell

samples. Furthermore, similarities were also found when

comparing equivalent fractions from fluid and shell

samples (Figure 4). These statements may eventually

predict the presence of similar proteins in fluid and

shell samples. All these aspects, in general, lead us to

propose a specific functional role of fluid proteins on the

shell formation.

The results of the immunogold assay in A. cygnea to

detect organic matrix protein proved to be effective and

useful. In the back-scattered electron mode, the gold

particles (covalently bound to the secondary antibody)

appeared as tiny bright spots. As shown in Figure 5,

these tiny bright spots were mainly found in the spaces

between the calcareous crystals, though some were also

observed within the crystals, denoting the presence of

organic matrix as a fundamental component on the bio-

mineral phase. Actually, these spots were distributed

either around the columnar structure of aragonite crys-

tals or filling the spaces between nacreous layers of

aragonite crystals. This confirms that the organic matrix

may play an essential role on the mineral formation and

organization in both vertical and horizontal axes in

bivalve shell (Krampitz et al., 1983; Cheea, 2000).

Molecular Vibrations of the Orcanic Matrix

of the Nacreous Layer

Infrared spectroscopy is a resource for a characterization

at a molecular level of the structure and bonding of

surface functional groups and adsorbed species. In this

study, ATR-IB spectra of aqueous organic matrix shows

two molecular vibrations at 3300 cm 1 and 1640 cm 1

(Figure 6) corresponding to amide group (amide-A). The

band observed at 3300 cm 1 is very broad because the

O-H stretching band appears as a typical polymeric

hydrogen bonded envelope near 3300 cm 1 . This means

that the polymeric hydrogen bonding donors are part

of the amide-A (C-H) group which displays strong

and broad C-H asymmetric stretching absorptions

at 3300 cm '. The molecular vibration at 1640 cm-1

Figure 5. Immunogold assay results. Topographical view of (A) negative control, (B) and (C) test with positive results; (D) positive

result in a transversal view.

A. Lopes et al., 2014

Page 1 1 1

Figure 6. ATR-IR spectra of aqueous organic matrix from nacreous layer A. cijgnea.

represents a symmetric C-O stretching. A hand may be

attributed to the presence of some C-O (amide l°-band).

On the other hand, nacreous solid samples analysed

(Figure 7) show a strong stretching vibration of CO.r

at 1470 cm " 1 corresponding to v3 vibration mode, which

is relevant for aragonite structure. The v2 asymmetric

bending vibration at 860 cm-1 is also observed, which

suggests a higher Ca“+ contribution than Sr"! or Mg1'"'

in aragonite structure. The vibration at 710 cm 1 also

corresponds to the v4 vibration mode (O-C-O) in plane

bending of C032 . The vibration band at 1016 cm 1

corresponds to the V] vibration mode of CO.3 -. The

shoulder at about 1600 cm 1 is related with amide- 1

functional group.

CONCLUSIONS

In general, this study provides new information on the

organic shell matrix of Anodonta cijgnea and confirmed

similarities with other studied families. Despite its small

Figure 7. FT-IR spectrum of the internal nacreous layer (with aragonitic structure) collected from the freshwater mussel Anodonta

cijgnea shell.

Page 112

THE NAUTILUS, Vol. 128, No. 4

percentage, the organic matrix, constituted mainly by

proteins and sulfated GAGs, has a relevant influence

on microstructure (proteins) and on the crystal nucle-

ation (sulfated GAGs) during the formation of the calcium

carbonate layers (Moura et ah, 2000; Lopes-Lima et al.

2005). Additionally, it has been suggested (Tong et ah,

2002) that while the intra-crystalline organic matrix,

mainly composed of small negatively charged molecules,

provides nucleating points and induces nucleation process

(Nudelman et al., 2006), the inter-crystalline organic

framework possesses a more complex organic composition

and is responsible for supporting, limiting size and shape,

and determining crystal growth spatial orientation.

In the present study, fluid and nacreous protein frac-

tions, composed by similar weight molecular peptides as

found by electrophoretic and MALDI-MS analysis as

well as the protein matrix observed in the nacreous by

SEM hystochemical techniques, seem to play mainly an

inter-crystalline role in the shell biomineralization. On

the other hand, the results obtained from soluble matrix

and solid nacreous samples by infrared spectroscopy anal-

yses, showing the similar occurrence of an amide group,

probably points out an intra-crystalline factor contributing

for the aragonite crystal formation in the nacreous layer

of A. cygnea. In fact, according to Choi and Kim (2000),

Xiao et al. (2005) and Kasat et al. (2006), the great elec-

tronegativity of oxygen allows amides to act as H-bond

acceptors changing the H -bonding states of C=0 groups

and consequently may define secondary structure and

polymer crystallinity. Possibly, the amide-I group is

involved in the calcium carbonate intra-crystalline struc-

ture acting as an inductor of aragonite polymorph.

AC K N OVVLE DC, M E NTS

Tl ie authors were supported by Fundayao para a Ciencia

e Tecnologia (FCT) under projects PTDC/MAR/098066/

2008, PTDC/AAC-AMB/1 17688/2010, scholarship

SFRH/BI/33 128/2007 and COST Action TD0903.

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THE NAUTILUS 128(4): 1 14— 121, 2014

Page 114

Two new species of Velutinidae Gray, 1840 (Gastropoda) from the

North Pacific with a preliminary molecular phylogeny of the family

David W. Behrens

1 124 250th Avenue NE

Sammamish, WA 98074 USA

Elysse Ornelas

Angel Valdes

Deparment of Biological Sciences

California State Polytechnic University

3801 West Temple Avenue

Pomona, CA 91768 USA

ABSTRACT

Two recent collections of shelled gastropods in the temperate

North Pacific have identified two undescribed species of the

family Velutinindae. Marsenina zadei new species is described

from Port Townsend, Washington. Onchidiopsis clarki new

species is described from Pribilof Island, Alaska, Bering Sea.

Both species are barcoded with sequences of the mitochondrial

COI and 16S genes. Additionally, a preliminary phylogeny for

the Velutinidae based on these two genes is provided.

Additional Keywords: Velutinidae, Lamellariidae, Marsenina,

Onchidiopsis

INTRODUCTION

Current classifications place lamellarid gastropods in the

family Velutinidae Gray, LS40 (Bouchet and Rocroi, 2005).

These are caenogastropods with an internal shell that have

been traditionally neglected. The species-level taxonomy

of this family has been in disarray. It is not the intention of

this paper to review the higher taxonomic nomenclatural

problems, so we follow the most recent review of species

presented in Gulbin and Golikov (2001, but see 1997,

1998, 1999, 2000). Prior to that, the most comprehensive

treatment of species from the North Pacific Ocean was

given by Behrens ( 1980), which, while focusing on species

known from the eastern Pacific (Alaska to Mexico), differ-

entiated members of the genera Lamellaria, Marsenina,

and Marseniopsis.

The main objective of this paper is to describe two new

species of the genera Marsenina and Onchidiopsis (sub-

family Velutininae Gray, 1842). Species of Marsenina are

distinguished by a radula having a formula of 2. 1.1. 1.2

(two outer teeth are present on each side), being her-

maphroditic, having a fissure or pore in the mantle expos-

ing the shell and permitting the retraction of the mantle,

and having a small, smooth foot that remains hidden

under the mantle. Species of Onchidiopsis has the same

radular formula as Marsenina , and are hermaphroditic,

but are distinguished in that they have an internal shell

fully enveloped by the mantle that is not retractile, and a

long foot with a distinctive rugose or nodular edge.

In order to allocate these two species within the phy-

logeny of the group, preliminary molecular phylogenetic

analyses (based on two mitochondrial genes) were con-

ducted, comprising the two new species and other mem-

bers of the Velutinidae for which sequences are available

in Gen Bank.

MATERIALS AND METHODS

Collection and Preservation: Specimens were col-

lected by scuba and trawl, respectively. All collected spec-

imens were fixed and preserved in 95% ethanol to

facilitate genetic analyses. All specimens were catalogued

and deposited in the Invertebrate collection of the Natu-

ral History Museum of Los Angeles County (LACM).

Morphological Examination: Preserved specimens

were dissected and the internal features were examined

using a dissecting microscope. The buccal mass of one

individual of each species was removed and dissolved in

10% sodium hydroxide until the radula and jaw were

isolated from the surrounding tissue. The radula and

jaw were then rinsed in water, dried, mounted, and

sputter-coated for examination under a scanning elec-

tron microscope (SEM) Hitachi S-3000N at the LACM.

The anterior end of the body including the head and

the penis, where dissected and chemically dried with

hexamethyldisilazane for SEM examination.

DNA Extraction, PCR, and Analyses: DNA extrac-

tion was performed using a hot Chelex1" protocol with

approximately 1-3 mg of the foot cut into fine pieces.

The tissue was rinsed and rehydrated using 1.0 mLTE

buffer (10 inM Tris, 1 mM EDTA, pH 8.0) for 20 min-

utes. A 10% (w/v) Chelex® 100 (100-200 mesh, sodium

form, Bio-Rad) solution was prepared using TE buffer.

After rehydration, the tissue mixture was then centri-

fuged, 975.00 pL of the supernatant was removed, and

175.00 pL of the Chelex® solution was added. Samples

D.W. Behrens et al., 2014

Page 115

were then heated in a 56°C water bath for 20 minutes,

heated in a 100°C heating block for 8 minutes, and the

supernatant was used for PCR. UniversallGS rRNA

primers (16S ar-L 5TCGCCTGTTTATCAAAAACAT-3',

168 br-H 5'- CCGGTCTGAACTCAGATCACGT-3' devel-

oped by Palumbi, 1996) and universal COl primers

(LCO1490 5'-GGTCAACAAATCATAAAGATATTGG-3',

HC02198, S'-TAAACTTCAGGGTGACCAAAAAATCA-

3' developed by Folmer et al., 1994) were used to amplify

the regions of interest for all specimens.

The master mix was prepared using 34.75 pL H20,

5.00 pL Buffer B (ExACTGene, Fisher Scientific), 5.00 pL

25 mM MgCl2, LOO pL 40m M dNTPs, 1.00 pL lOmM

primer 1, 1.00 pL primer 2, 0.25 pL 5 mg/mL Taq, and

2.00 pL extracted DNA. Reaction conditions for 16S were

as follows: an initial denaturation for 2 min at 94°C,

30 cycles of 1) denaturation for 30 sec at 94° C, 2) anneal-

ing for 30 sec at 50°C, and 3) elongation for 1 min at

72°C, and a final elongation for 7 min at 72°C. Reaction

conditions for COl an initial denaturation for 3 min at

95° C, 35 cycles of 1) denaturation for 45 sec at 94°C,

2) annealing for 45 sec at 45° C, and 3) elongation for

2 min at 72°C, and a final elongation for 10 min at 72°C.

PCR products yielding bands of appropriate size

(approximately 475 bp for 16S and 700 bp for COl) were

purified using the Genejet PCR Purification Kit (Thermo

Scientific). Cleaned PCR samples were quantified using a

Nano Drop 1000 Spectrophotometer (Thermo Scientific).

Each primer was diluted to 2.0 pmol/pL to send out for

sequencing with the PCR products. PCR products were

diluted to 7.5 and 11.5 ng/pL for 16S, and COl, respec-

tively. Samples were sequenced at Source Bioscience

(Santa Fe Springs, CA).

For the phylogenetic analyses, sequences of the fol-

lowing species were obtained from Gen Bank: Coriocella

nigra Blainville, 1824 (16S: AY161381, COl: AY161614),

Lamellaria sp. 1 (16S: AY161382, COl: AY161615),

Lamellaria sp. 2 (16S: AY 16 1383, COl: AY161616), and

Marsenurpsis mollis (COl: GU227110). The triviid species

Triveilla millardi (Cate, 1979) (16S: AY 16 1389, COl:

AY161622) was selected as the outgroup based on recent

phylogenetic analyses (Meyer, 2003). Sequences for each

gene were assembled and edited using Geneious Pro 4.7.4

(Drummond et al., 2010). Geneious was also used to

extract the consensus sequence between the primer

regions, to construct the alignment for each gene using

the default parameters and to concatenate the alignments.

The sequences were trimmed after alignment. A total of

473 bp for 16S, and 614 bp for COl were used for the

phylogenetic analyses.

The phylogenetic analyses were conducted for both

genes concatenated. The Akaike information criterion

(Akaike, 1974) was executed in MrModeltest (Nylander,

2004) to determine the best-fit models of evolution for

each gene (GTR+I+G for COl and GTR+I for 16S).

The Bayesian analysis was executed in Mr Bayes 3.2.1

(Huelsenbeek and Ronquist, 2001), partitioned by gene

(unlinked). The Markov chain Monte Carlo analysis was

run with two runs of six chains for ten million genera-

tions, with sampling every 100 generations. The default

25% bum-in was applied before constructing majority-

rule consensus tree/s. The maximum likelihood analysis

was conducted with the program GARLI v0.96b8 (Zwickl,

2006). Default parameters were used to run three differ-

ent GARLI searches of 10 replicates each, and a total of

2,000 bootstrap replicates were performed to assess the

robustness of each clade (Felsenstein, 1985).

SYSTEMATICS

Family Velutinidae Gray, 1840

Subfamily Velutininae Gray, 1840

Genus Marsenina Gray, 1850

Type Species: Lamellaria prodita Loven, 1846

(= Oxynoe? glabra Couthouy, 1838)

Marsenina zadei new species

(Figures 1-4, 7-9)

Description: External Morphology: Mantle color

makes this species difficult to find on its substrate (Figure 1).

Mantle color dusky-white to tan to orange, with a sprin-

kling of black specks. A dark brown horseshoe mark pres-

ent posterior-medially in some specimens. Mantle with a

dorsal slit or fissure, which may lie retracted exposing

shell. Mantle with an anterior and right lateral fold creat-

ing incurrent and excurrent siphons, respectively, circulat-

ing water over gills. Surface of the mantle covered with a

pattern of spots resembling atrial siphons of aseidian host.

Some specimens with radiating ridges on mantle (Figure 2).

A most distinctive mantle feature is a series of tubercles

within the dark horseshoe (Figures 2^4).

Shell: Shell oval, translucent-white, with a number of

growth lines. Protoconch situated on posterior right side

of shell, partially engulfed by teleoconch. Protoconch

large, elongate, about 600 pm x 1 mm with 1.1 whorls

(Figure 8).

Radula and Jaws: The radular formula is 98 x

2. 1.1. 1.2. The rachidian tooth bears 1 to 2 strong denti-

cles to each side of the central cusp (Figure 7). The inner

lateral teeth bear a single, strong denticle to each side of

the central cusp. The outer lateral teeth are smooth and

hamate. Masticatory border of the jaw (Figure 9) with a

series of uniform denticles.

Penis: The penis (Figure 10) is flat, branching into

three blunt apices distally.

Molecular Data: Sequences of this species are

available in Gen Bank. Molecular phylogenetic analy-

ses place this species as sister to Onchidiopsis clarki ,

but with limited support in the Bayesian analysis

(Figure 14).

Biology: All specimens were collected on an encrusting

compound aseidian, tentatively identified as Trididemnum

Page 116

THE NAUTILUS, Vol. 128, No. 4

Figures 1-6. Living animals of Marsenina and Onchidiopsis species. 1-4. Marsenina zadei new species. Port Townsend,

Washington, black arrows indicate the salmon-orange egg capsules. Photos by Rick Zade. 5. Marsenina steamsii (Dali, 1871).

Keysone Jetty, Whidbey Island, Washington. Photo by Jan Kocian. 6. Onchidiopsis clarki new species. Bering Sea, NNE of Pribilof

Island, Alaska. Photo by Roger Clark.

opacum (Ritter, 1907), at depths from 3 to 15 m. Col-

lected with the specimens, and buried in the tunic of the

ascidian, were salmon-orange egg capsules (Figure 1).

Upon dissection the orange color within some of the

capsules was determined to be the color of the develop-

ing larvae.

Type Material: Holotype, LAGM 3280, 15 mm pre-

served length; Paratypes, LACM 3281, 11 specimens,

6-15 mm preserved length, all Richard Zade coll., 8 Feb-

ruary 2010, from type locality.

Type Locality: Hudson’s Point (48°6.949N,

122°44.999W), Port Townsend, Washington State, USA,

15 m depth.

Distribution: Port Townsend, Washington (present

study); Ten Mile Point, Greater Victoria, British Columbia,

Canada (photo by James Hester); Pigeon Point, San Mateo

County, California (photos by Gary McDonald and Doug

Mason); Carmel Point, Monterey County, California

(photo by Gary' McDonald).

Etymology: This species is named after Richard Zade,

the collector of the type specimens.

Remarks: Marsenina zadei differs significantly from

the two other described species from the North Pacific,

both internally and externally. As in M. zadei, the mantle

of Marsenina steamsii resembles the encrusting tuni-

cate, Trididemnum opacum, but M. steamsii lacks any

dark markings or black specks, and has a smooth mantle

D.W. Behrens et si., 2014

Page 117

Figures 7-10. Marsenina zadei new species, Port Townsend, Washington. 7. SEM of a section of the radula, showing rachidian,

lateral, and marginal teeth. 8. SEM of protoconch. 9. SEM of masticator)- border of jaw. 10. SEM of penis (pe) and oral tentacle (ot).

lacking tubercles (Figure 5). Dali in Orcutt (1885) and

Smith (1948) state that Marsenina steamsii var.

orhiculata is not a valid taxonomic entity. Ghiselin

(1964) and Behrens (1980; 1984) describe the morphol-

ogy of the mantle and are the only known published

photographs of living specimens of M. steamsii. In

M. steamsii, the mantle coloration is white to creamy

white with slightly elevated darker cream colored

spots resembling the atrial siphons of its host ascidian

(Figure 5). Numerous photos were submitted by col-

leagues and more found on the web that are attributable

to M. zadei, but those had been idntified as M. steamsii.

Bsed on these photos, it is possible extend the geograph-

ical range of M. zadei south to Carmel Pt. California,

December 2, 1971 (photo by G. McDonald).

Ol the internal anatomy, only the radula and shell have

been described for M. steamsii and M. rhomb ica. Dali

(1871, 1885) reported the distinguishing characteristics

of the shell surface of M. steamsii to be microscopic fine

revolving striulae. However, such striulae were not

observed on the shell a specimen of M. steamsii from

Marin County, California (LACM) examined for this

study. The main differences between the shells of

M. steamsii and M. zadei is the protoconch morphology;

in M. steamsii the protoconch is more circular in shape

and has nearly 2 whorls, whereas the protoconch of

M. zadei is more oval and has 1.1 whorls.

In most cases, as in other genera of this family, only the

penis has been described for Marsenina. The penis of

Marsenisn zadei is not similar to any of those species for

which descriptions are available. The penis of co-occuring

Marsenina rhornbica is figured and described by Gulbin

and Golikov (2000) as horn-shaped, not flattened and

slightly bifurcate as described for M. zadei herein.

Behrens (1980) described the radulae of M. steamsii

and M. rhornbica. While the number of rows of teeth in

THE NAUTILUS, Vol. 128, No. 4

Page 1 18

the radula was not given for either of the north Pacific

Marsenina species, the morphology of the teeth was found

to he identical between the two and differs slightly from

that of M. zadei. Marsenina steamsii is reported to have a

rachidian tooth with zero or one denticle flanking the

central cusp, while the rachidian of M. zadei bears 1 to 2

strong denticles to each side of the central cusp. The inner

and outer lateral teeth are similar in all three species.

Genus Onchidiopsis Bergh, 1853

Type Species: Onchidiopsis groenlandica Bergh, 1853

Onchidiopsis clarki new species

(Figures 6, 11-13)

Description: External morphology: Mantle com-

pletely covers shell. Mantle covered with randomly

spaced, large triangular tubercles (Figure 6). An anterior

fold gives origin to incurrent siphon. Oral tentacles pro-

truding from under mantle. Oral tentacles long, slender,

and tapering (Figure 13). Specimens brown with uni-

formly distributed white specks. Brown ground color

fades to off-white on tubercles and siphon.

Shell: The shell is an un-caleifled, soft plate without

any recognizable characteristics in the specimens examined.

Radula and Jaws: Radular formula is 74 x 2. 1.1. 1.2

(Figure 11). Rachidian bears 5-7 irregularly sized denti-

cles on each side of central cusp. Inner lateral teeth bear

3-5 dissimilarly sized denticles on each side of the cen-

tral cusp. Pair of outer lateral teeth simple, hooked, lack-

ing denticles. Masticatory margin of jaws with a series of

nearly uniform denticles, posteriorly with a large dentic-

ulate flange (Figure 12).

Penis: The reproductive system is typical of members

of the genus, Onchiopsis , from what little information we

could find. The penis (Figure 13) is thickened, highly

twisted, with a blunt truncated end.

Figures 1 1-13. Ochidiopsis clarki new species, Alaska, Bering Sea. 1 1. SEM of a section of the radula, showing rachidian, lateral

and marginal teeth. 12. SEM of masticatory border of jaw. 13. SEM of penis (pe) and oral tentacle (ot).

D.W. Behrens et a!., 2014

Page 119

Molecular Data: Sequences of this species are avail-

able in Gen Bank. Molecular phylogenetic analyses place

this species as sister to Marsenina zadei, but with limited

support in the Bayesian analysis (Figure 14).

Biology: Both specimens were collected in the same

trawl over a mud bottom at a depth of 87 m. There were

no indications of which of the other organisms collected

in the trawl might be this species prey.

Type Material: Holotype, LACM 3282, specimen

28 mm preserved length; Paratype, specimen 26 mm

preserved length, LACM 3283, all trawled by the R/V

Artur us, leg. Roger Clark, from type locality.

Type Locality: NNE of Pribilof Island (58°00.79N,

170°58.18W), Bering Sea, Alaska, Bering Sea, USA,

87 m depth on mud, bottom temperature 4.0°C (NMFS

88-2003-1-138).

Etymology: This species is named after Roger Clark,

the collector of the type specimens.

Remarks: The genus Onchidiopsis is found in both

the cold temperate Atlantic and Pacific Oceans. Both

faunas are poorly known. Depending on which database

we examined, numbers of species varied around 15 for

the North Atlantic, while 10 are reported from the North

Pacific (Gulbin and Golikov, 2001). Several of these are

reported from throughout the Arctic Sea, reaching into

both oceans.

Although Balch’s (1910) original description of

Onchidiopsis conjs from Newfoundland and Labrador,

North Atlantic, describes the notum as smooth on the

top and sides, with wrinkles and folds elsewhere, photos

on the web (http://eol.org/pages/72611/overview and

http://eol.org/pages/593815/overview) of recent speci-

mens “thought to be” O. conjs bear some external simi-

larity with O. clarki, having triangular tubercles on the

mantle. These specimens have lighter color and their

mantle surface is knobby and granular between the

tubercles. Balch’s description of the internal anatomy is

vague and could apply to any species in the genus.

The most obvious Circumboreal species to be consid-

ered here are Onchidiopsis gladalis (Sars, 1850) and

Onchidiopsis groenlandica Bergh, 1853. These species

seem to have been maintained as separate in the literature

(Bergh, 1886; MacGinitie, 1959; Gulbin and Golikov,

2001) even though there appears to be adequate argu-

ments to synonymize the two (Balch, 1910; Thorson,

1944; Macpherson, 1971). Where the mantles of these

two very' similar species are discussed, descriptions vary

from rugose and wrinkled (Gulbin and Golikov, 2001) to

convoluted (brain-like) (Macpherson, 1971). We have

found no mention of specimens with triangular tubercles.

Nowhere in the literature can we find a North Pacific

species with large triangular tubercles seen in O. clarki.

Marseniopsis mollis

i

Too"

0 92

90

Marsenina zadei

Onchidiopsis clarki

Lamellaria sp. 1 Australia

0 79

65

Lamellaria sp 2 Tanzania

Coriocella nigra

Gaeatrivia millardi

0 07

Figure 14. Bayesian consensus tree of the concatenated analysis including posterior probabilities and bootstrap values from the

maximum likelihood analysis.

Page 120

THE NAUTILUS, Vol. 128, No. 4

All of the species figured in Gulbin and Golikov (2001)

appear to have smooth or slightly granular mantle sur-

faces, lacking tubercles.

We were unable to find any published description of

the internal reproductive system, only that of the penis,

which in the genus seems to be diagnostic. Of those

species described by Gulbin and Golikov (2001), none

are comparable to the penis of O. clarki described here

as thickened, highly twisted, with a blunt truncated end.

In O. groenlandica, they described the penis as having

an ancillary appendage hanging over the penis. In

O. variegatci, they described the penis as long, cylindri-

cal, its distal part thickened and curved, terminating in a

swelling with a thin fold. In O. zuchsi, they describe the

penis as highly characteristic with a large divided lobe.

On the side it bears an extending papilla, rimmed with

a thin fold. All members of the subgenus Rostroonch-

idiopsis have a long tapering, hook shaped penis with

a crest at the bend, while members of the genus

Bulloonchidiopsis have a flattened, hammer-shaped

penis, the distal end of which is recurved upward.

DISCUSSION

In this paper we include the first, albeit very preliminary,

molecular phylogeny of the Velutinidae, based on COI

and 16S sequence data. The resulting tree lacks sup-

port for most branches, suggesting that the two genes

sequenced are not adequate to recover the phylogeny

of the Velutinidae. However, the general structure of

the tree appears to partially support the classification

scheme proposed by Bouchet and Roeroi (2005). The gen-

era Lamellaria and Coriocella (subfamily Lamellariinae)

are placed in the same clade. However, the other subfam-

ily recognized by Bouchet and Roeroi (2005), Velutininae,

is paraphyletic in the present analysis, as Marseniopsis

does not cluster with Marsenina and Onchidiopsis, which

form a monophyletic group well supported in the maxi-

mum likelihood analysis.

In order to reconstruct the phylogeny of this group it

will be necessary to sequence additional genes, includ-

ing nuclear markers and substantially expand the taxo-

nomic coverage.

ACKNOWLEDGMENTS

Tl le authors would like to thank the two gentlemen who

made us aware of these two interesting species. Richard

Zade, avid diver and master underwater photographer

from Spanaway, Washington, somehow noticed this very

cryptic species hiding on its host tunicate. Thanks also to

Roger Clark, contract biologist to the National Marine

Fisheries Service, who was responsible for invertebrate

identification from trawl samples aboard the R/V

Arcturus. Ironically, he is specialist in crabs! The

authors would also like to thank James Hester, Gary

McDonald, and Doug Mason for providing photo-

graphic evidence of Marsenina zadei occurring in

Canada and California, and [an Kocian for the photo

of M. steamsii from Whidbey Is. Washington. Lindsey

Groves (LACM) curated the specimens studied and

provided us with access to the collection. The SEM work

was conducted at the LACM SEM facility sponsored by

the NSF (MRI grant DB1-0216506) with the assistance

of Giar-Ann Kung.

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the Northern Hemisphere. V. Onchidiopsinae. Ophelia

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THE NAUTILUS 128(4):122-128, 2014

Page 122

A new genus of Buceinoidea (Gastropoda) from Paleocene

deposits in eastern Hokkaido, Japan

Kazutaka Amano

Department of Geoscience

Joetsu University of Education

1 Yam ay as hi ki

Joetsu 943-8512, JAPAN

Anton Oleinik

Department of Geosciences

Florida Atlantic University

777 Glades Hoad

Boca Raton, FL 33431 USA

ABSTRACT

A new genus and a new species of the gastropod superfamily

Buceinoidea, Urahorosphaera kanekoi , is described from the

Paleocene Katsuhira Formation in eastern Hokkaido, Japan.

This species comprises the first Paleocene record of buccinoid

gastropods in Japan and originates from relatively deep-water

deposits, which makes it very unique among all Paleocene

buccinoidean gastropods in the North Pacific. This occurrence

may suggest a trace of bucciniform gastropods diversification

similar to the pattern found for the Southern Hemisphere.

Additional Keywords: New species, Brachysphingus,

Austrosphaera, Seymou rosphaera

INTRODUCTION

Late Cretaceous and Paleocene buccinoidean gastropods

of uncertain affinity are known from the North Pacific,

Antarctica, and Soudi America. The genus Brachysphingus

Gabb, 1869 is known from the Paleocene (Danian-

Thanetian) of Kamchatka (Gladenkov et ah, 1997) and

California (Squires, 1997), genus Austrosphaera

Camacho, 1949 from the Late Cretaceous and Paleocene

strata of Tierra del Fuego, and genus Seymourosphaera

Oleinik and Zinsmeister, 1996 from the Paleocene

(Danian) of Seymour Island, Antarctica (Oleinik and

Zinsmeister, 1996; Stilwell et ah, 2004). The last two

genera have once been tentatively included in the sub-

family Pseudolivinae Cossmann, 1901 (Oleinik and

Zinsmeister, 1996). Vermeij (1998) excluded them from

Pseudolivinae based on the lack of a pseudolivid groove

and a labral tooth. Based on their occurrence in the

Southern Hemisphere, the genera Austrosphaera and

Seymourosphaera most probably belong to the austral

family Buceinulidae Finlay, 1928, rather than to the

North Pacific family Buccinidae Rafinesque, 1815. All

these buccinoid genera share a semi-ovate outline of

the shell, predominantly smooth surface, low to moder-

ately elevated spire, short siphonal canal and a poorly

developed faseiole.

No Paleocene buccinoidean gastropods of similar aff in-

ities were previously reported from Japan. Two specimens

of smooth-surfaced, subovate buccinids have recently

been collected from the Piileocene part of the Katsuhira

Formation (Amano and Jenkins, 2014) in eastern

Hokkaido. We herein propose a new genus and a new

species for these unusual buccinids.

MATERIALS AND METHODS

Two buccinoidean specimens were collected from two

separate carbonate concretions, 20 to 40 cm in diameter,

found as floats. Concretions were originally embedded

within the mudstone of the Katsuhira Formation, eastern

Hokkaido (Figure 1; Loe. 1 and 2). Although most con-

cretions at these localities occurred as floats, they must

have been derived from localities nearby. For example,

some autochthonous concretions yielding fossils were

cropped out 50 m upstream of Loc. 1.

Many calcareous concretions with plant debris are

found in the upper part of this formation near the tvpe

locality. As discussed by Amano and Jenkins (2014), the

age of the upper part of this formation has been assigned

to the Paleocene, based on planktonic foraminifers. The

age of the upper part of the Katsuhira Formation was

assigned to early Selandian, based on planktonic fora-

minifers and calcareous nannofossils (Kaiho, 1984).

One well-preserved gastropod specimen was obtained

from a float calcareous concretion (30 cm in diameter) at

900m upstream of the small river, 1.5 km south to

Ponkatsuhira-zawa (Loc. 1). Another, rather poorly pre-

served specimen was collected from a float calcareous

concretion (about 30cm in size) at approximately 1 km

upstream of Ponsetarai River (Loc. 2). Multiple fragments

of plant material, protobranch bivalves as Acila , Leionucula,

and malletiids, aporrhaid gastropod Kangilioptera inouei

Amano and Jenkins, 2014 and scleractinian corals are

associated with the buccinoidean specimens in both

localities. One of the corals was found near the aperture

of the poorly preserved specimen at Loc. 2. From other

localities of the Katsuhira Formation, deep-water arcoid

K. Amano and A. Oleinik, 2014

Page 123

Figure 1. Locality map of Urahorosphaera kanekoi new

genus and new species (base map is from “Tokomuro”, scale 1:

50,000 topographical map published by the Geospacial Infor-

mation Authority of Japan).

Bentharca was also found. The paleoenvironments of

the Katsuhira Formation has not been studied in

any detail. Fossils assemblage is indicative of deep-

water accumulation.

Abbreviations used are: |UE: Joetsu University of

Education, Joetsu, Niigata Prefecture; FAU: Florida

Atlantic University, Boca Raton, Florida; PRI: Paleonto-

logical Research Institution, Ithaca, New York.

SYSTEMATIC PALEONTOLOGY

Class Gastropoda Cuvier, 1797

Order Neogastropoda Wenz, 1938

Superfamily Bucci noidea Rafinesque, 1815

Family (P)Buccinidae Rafinesque, 1815

Remarks: The Buccinidae is one of the most diverse

families of neogastropods. Members of the family are

distributed from the Equator to the poles, and inhabit

exclusively marine environments. The current under-

standing is to place the northern hemisphere genera in the

family Buccinidae, and the southern hemisphere genera

in the family Buccinulidae ( Bouchet and Waren, 1986;

Schnetler, 1997; Harasewyeh and Kantor, 1999, 2004;

Squires and Saul, 2000; Kantor and Harasewyeh, 2013).

Problems of Buccinidae origin, appearance in the fossil

record, and relationship to families Fasciolariidae,

Nassariidae, and Melogenidae, have been debated in

the literature for some time (Ponder, 1974; Tracey et al.,

1993; Bandel, 1993; Kantor, 1996; Ponder and Lindberg,

1997). The phylogenetic relationships of the recent

Buccinidae remain unclear. There is no agreement on

the exact limits of the family, as well as relationships

among it’s over than 200 genera and subgenera

(Harasewyeh, 1998). The molecular phylogenetic data

for buccinids remain insufficient. Whatever phylogenetic-

data exist mostly point on the paraphyly for the Buccinidae,

but also suggest the limited resolving power of current

molecular phylogenetic analyses (Hayashi, 2005; Oliverio

and Modica, 2010; Kantor et al., 2012).

The fossil record indicates the diversification in

bucciniform gastropods during the Late Cretaceous and

early Cenozoic. A leading tendency in multiple publica-

tions is to place these genera in the present-day families

Buccinidae or Nassariidae. Allmon (1990) had commented

that placing late Mesozoic and early Cenozoic bucciniform

gastropods into a few traditionally recognized living fami-

lies obscures the phylogeny and leads to underestimation

of family-level diversity during this time interval. Squires

(1997) had commented that in all probability. Late Cre-

taceous and earlv Cenozoic bucciniform gastropods most

probably belong to several new undescribed families that

are waiting to be properly erected.

It is most likely that the new genus described in this

manuscript belongs to a separate, undescribed family of

bucciniform gastropods. Limitations in quantity and

quality of our material, however, prevent us from desig-

nating a new family with confidence at this point.

Urahorosphaera new genus

Type Species: Urahorosphaera kanekoi new species,

Paleocene (early Selandian), upper part of the Katsuhira

Formation, Urahoro Town, eastern Hokkaido, Japan.

Diagnosis: Shell subovate, inflated; spire low;

protoconch bulbous; surface smooth and glossy, except

for weak axial riblets near aperture; thick peristomate

outer lip; siphonal canal short with weakly developed

siphonal notch.

Description: Shell medium-sized, thick, with glossy

surface, subovate. Last whorl large and globose; spire

very low; protoconch smooth and bulbous. Surface of

last whorl sculptured by thin and low axial riblets; shallow,

but distinct subsutural groove; aperture pear-shaped;

outer lip thick, forming peristome; thin callus covering

body whorl, spire and protoconch; weakly developed

siphonal notch and parietal canal; anterior end of colu-

mella abruptly tapered.

Remarks: Species in Urahorosphaera have broad thin

callus covering whole surface. Such character is usually

seen in members of the Olivoidea. However, Landau and

Marquet (1999) described buccinoidean gastropod,

Cijllene ( Ci/llenina ) lucenensis from the Pliocene deposit

Page 124

THE NAUTILUS, Vol. 128, No. 4

in Spain which has a callus covering teleoconch whorls.

This genus is also considered to be an aberrant group-

ing within Buccinoidea.

Urahorosphaera resembles Brachysphingus Gabb,

1869, from the Paleoeene to Eocene formations of

California in having a subovate shell form, low spire,

and short siphonal canal. However, Urahorosphaera

differs from Brachysphingus by having overall larger

size, a smooth glossy shell surface, and a thick outer lip

and lacking of spiral cords on the base.

The austral genus Austrosphaera Camacho, 1949 (in

Furque and Camacho, 1949) from the Late Cretaceous

to Paleoeene in Argentina is another similar to the

Urahorosphaera in having overall subovate shell and

low spire. Urahorosphaera differs from Austrosphaera

by having a glossy shell surface, wider callus, thick outer

Figures 2-10. Urahorosphaera kanekoi new genus and new species. 2-8. Holotype, JUE no. 15924; 2, apertural view;

3, adapertural view; 4, side view; 5, enlargement of base; 6, apical view; 7, apical view of protoconch; 8, side view of protoconch.

9-10. Paratype, JUE no. 15925; 9, apertural view, cr, coral; 10, oblique view of aperture.

K. Amano and A. Oleinik, 2014

Page 125

lip, and by lacking multiple columellar plications toward

the anterior end of the columella.

The Antarctic Paleogene genus Seymou ro.sphaera

Oleinik and Zinsmeister, 1996 differs from Urahorosphaera

by having multiple fine spiral threads and poorly devel-

oped siphonal canal, and by lacking a siphonal notch.

Urahorosphaera differs from Pangoa Marwick, 1931,

from the Miocene (Lillburnian) of New Zealand by hav-

ing a more compressed shell with lower spire, shorter

siphonal canal, broader callus, and glossy shell surface.

Urahorosphaera differs from Sycostoma Cox, 1931

from the Eocene of Europe and North America by having

a more rounded semi-ovate shell, lower spire, wider

callus, shorter siphonal and weakly developed parietal

canals, and larger bulbiform protoconch.

The genus Liochlamys Dali, 1889 (family Fasciolariidae)

from the Neogene of the southeastern United States,

although lias a glossy shell surface, overall globose shape

of the shell, and bulbiform protoconch, differs from the

Urahorosphaera by having two to three columellar folds,

or plications, wider and more elongated siphonal canal,

higher spire, and presence of apertural ribs in the interior

of the aperture.

Etymology: The new genus is named after the town

of Urahoro, site of the type locality in Hokkaido.

Age and Occurrence: Paleocene Katsuhira Formation

in Urahoro, eastern Hokkaido.

Urahorosphaera kanekoi new species

Japanese name: Urahoro-migaki-bora

(Figures 2-10)

Diagnosis: Same as that of the new genus.

Description: Shell medium-sized, attaining 43.0 mm

in height, thick, polished, subovate with four whorls.

Last whorl large and globose; spire very low, covered by

thin glaze, comprising approximately 1/6 of the total

shell height; protoconch smooth, bulbous, low-domed,

consisting of 1.5 whorls. Surface of last whorl sculptured

by twelve thin and low axial riblets near aperture; riblets

becoming obsolete anteriorly. Subsutural groove very

shallow, but distinct. Aperture pear-shaped; outer lip thick,

forming distinct peristome. Outer lip forming blunt angle

near boundary between base and posterior end; columella

concave and smooth; inner lip broadly covered by thin

calcareous callus; callus extending over front of shell and

over parietal sinus, covering suture and protoconch;

siphonal canal short and slightly oblique with weakly

developed siphonal notch; parietal canal weakly developed;

anterior portion of columella tapering abruptly.

Type Material: Holotype, JUE no. 15924 (shell height,

43.0 mm; diameter, 29.5 mm); Paratype, JUE no. 15925

(diameter, 28.2 mm).

Type Locality: 900 m upstream of the small river, 1.5 km

south to Ponkatsuhira-zawa, Urahoro, eastern Hokkaido.

Remarks: Urahorosphaera kanekoi new species shares

a subovate shell with low spire and generally smooth

surface with Brachysphingus mammilatus Clark and

Woodford, 1927 (especially paratype UCMP 31235; see

Squires, 1997, figs. 5-10, 11) from the upper Paleocene

and lower Eocene in California, and with Brachysphingus

gibbosus Nelson, 1925 from the early Paleocene of

Kamchatka (Figures 11-12). However, Urahorosphaera

kanekoi new species differs from these species by the

presence of a thick outer lip, suddenly tapering anterior

end and lack of spiral cords on the basal part.

Urahorosphaera kanekoi is similar to Seymourosphaera

bulloides Oleinik and Zinsmeister, 1996 (Figures 13-14,)

in general shape and smooth surface, but differs by a

higher spire, thicker peristome, suddenly tapering ante-

rior end, and broader callus.

Figures 11-14. Paleocene smooth-surfaced subovate gastropods. 11, 12. Brachysphingus gibbosus Nelson, 1925. PRI 49420.

1 1, aperural view; 12, abapertural view. Paleocene, Danian, Getkilninskaya Formation, Northwestern Kamchatka.

13, 14. Seymourosphaera bulloides Oleinik and Zinsmeister, 1996. FAU 18384-11. 13, apertural view; 14, abapertural view.

Paleocene, Danian, Sobral Formation, Seymour Island, Antarctica.

Page 126

THE NAUTILUS, Vol. 128, No. 4

Etymology: The new species is named for Mr. Atsushi

Kaneko who collected the holotype.

Distribution: Known from the type locality and 1 km

upstream of Ponsetarai River, Katsuhira Formation,

Urahoro, eastern Hokkaido.

DISCUSSION

Most of the North Pacific buccinids, other than the

Paleocene through middle Eocene genera Siphonalia

and Snatolia (Family Siphonaliidae, according to

Goryachev, 1987, Oleinik, 1988, and Gladenkov et ah,

1988), appeared in the late Eocene and their diversity

gradually increased from the late Eocene to recent

following the general trend of climatic cooling in the

region (Titova, 1994; Gladenkov et ah, 1997). The Late

Cretaceous and Paleocene record of the North Pacific

Buccinidae remains rather poorly known. Apart from

apparently buccinoidean, highly sculptured gastropods,

such as genera Omopsis Wade, 1916 and Deussenia

Stephenson, 1941 from the Late Cretaceous and Paleo-

cene of California and northern Mexico, the only early

Paleocene buccinoidean gastropod, morphologically

similar to Urahorosphaera , in the northern circum-

Paeifie, is the genus Brachijsphingus. Three species of

Brachysphigus {B. gibbo.sus Nelson, 1925, B. sinuatus

Gabb, 1869, and B. mammilatus Clark and Woodford,

1927) are known from the uppermost Cretaceous (?)-

lower Paleocene strata of California and Baja California

(Squires, 1997). Two species {B. sinuatus and B. gibbosus)

are known from the Paleocene Getkilninskaya Formation

of northwestern Kamchatka (Gladenkov et ah, 1997),

which comprises the northernmost record of this genus

in the northern eireum-Pacific. The distribution of the

genus Brachijsphingus in the Paleocene of the northern

circum-Pacific follows the general pattern outlined by

Oleinik (2001). At that time, marine isotopic records do

not indicate a significant departure in temperature

values from the Late Cretaceous and do not indicate a

significant thermocline of depth (Bralower et ah, 2002;

Dutton et ah, 2005), with some indications of warmer

temperatures (Adatte et ah, 2002) during the early Paleo-

cene. These paleoceanographic conditions would promote

the dispersal of molluscan faunas across the northern rim

of the Pacific Ocean which explains similarity of the east-

ern North Pacific and northwestern Pacific Paleocene

molluscan faunas. Paleocene molluscan faunas from Japan

remained virtually unknown until recently (Amano and

Jenkins, 2014). The new genus Urahorosphaera is mor-

phologically distinct from the Brachijsphingus, showing

more similarity with the austral genera Austrosphaera

and Seymourosphaera that diversified in the southern

hemisphere. Discovery of the Urahorosphaera in the

lowermost upper Paleocene of Hokkaido may suggest a

trace of similar early Paleocene diversification in the

North Pacific. The genus Brachijsphingus in the Paleo-

cene of the North Pacific is also known from a shallow-

marine deposits represented by lithic sandstone and

associated with shallow-water bivalves. On the other

hand, the lithology and assemblage of the Katsuhira

Formation, from which Urahorosphaera was found, is

indicative of relatively deep-water deposits, which

makes it unique among other North Pacific Paleocene

localities. Somewhat similar deep-water assemblages

are found in the lower portion of the Getkilninskaya

Formation of northwestern Kamchatka, but those do

not contain Brachijsphingus, Urahorosphaera, or any

apporhaid gastropods.

ACKNOWLEDGMENTS

We thank Steffen Kiel (Georg- August University Gottingen)

for his insights. We also thank Yuri I. Kantor (A.N.Severtzov

Institute of Ecology and Evolution, Russian Academy of

Sciences) and an anonymous reviewer for their review

and useful comments. This study was partly supported by

a Grant-in-aid for Scientific Research from the Japan

Society for Promotion of Science (C, 23540456, 201 1-2013;

C, 26400500, 2014-2016) to KA.

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THE NAUTILUS 128(4):129-134, 2014

Page 129

First report of Cryptoplax propior Is. and Iw. Taki, 1930

(Polyplacophora: Cryptoplacidae) in Korea

Ronald G. Noseworthy

Hee-Jinig Lee

School of Marine Biomedical Science

(BK21 PLUS)

Jeju National University

102 Jejndaehankno

Jeju, 690-756, REPUBLIC OF KOREA

Hyung-Kee Cha

Dokdo Fisheries Research Center

East Sea Fisheries Research Institute

National Fisheries Research and

Development Institute (NFRDI)

Pohang, REPUBLIC OF KOREA

Young-Chul Kang

Benthos Research Institute

Room 416, Smart Building, Jeju Science Park

Jeju Ocean Inc.

Jeju, REPUBLIC OF KOREA

Kwang-Sik Choi1

School of Marine Biomedical Science

(BK21 PLUS)

Jeju National University

102 Jejudaehankno

Jeju, 690-756, REPUBLIC OF KOREA

[email protected]

Sukgeun Jung

School of Marine Biomedical Science

Jeju National University

102 Jejudaehankno

Jeju, 690-756, REPUBLIC OF KOREA

ABSTRACT

We report the first occurrence of the chiton Cryptoplax

propior Is. and Iw. Taki, 1930, a species of Polyplacophora

newly reported for the Korean molluscan fauna. Two speci-

mens of C. propior were obtained during faunal surveys from

two localities on both the north and south coasts of Jeju Island,

off the southern coast of Korea. Although reference has been

made to C. propior in several faunal and taxonomic studies,

little is known about the ecology of this species, and only a few

specimens have been collected since it was first described in

1930. Scanning Electron Microscopy (SEM) revealed the char-

acteristics of C. propior: the granular rows on the median and

tail valves, and the short, thick blunt spicules on the perinotum.

Due to the present study, the distribution range of C. propior

now extends from south-central Japan to |eju Island, south-

western Korea.

Additional Keywords: Jeju Island

INTRODUCTION

Located in the northern East China Sea, Jeju Island has

a warm humid temperate climate with an average yearly

temperature of 16°C (Korean Meteorological Adminis-

tration, 2013). The ocean around the island is a complex

region, where three major water masses from the north

and northwest are mixed. The warm Tsushima current

washes the southern coastal area of the island, giving this

1 Author lor correspondence

area somewhat warmer winter sea surface temperatures

than the rest of the country, ranging from 14-16 °C

(Limpanont et ah, 2010, 2011). During the summer, the

sea surface temperature in Jeju often reaches a maxi-

mum temperature of 28 °C (Global Sea Temperature,

2013). Since Jeju Island is part of the Warm Temperate

Northwest Pacific Province (Spalding et ah, 2007), there

is a strong affinity with the fauna of southern Japan and

eastern China, and also northeast Taiwan (Noseworthy

et ah, 2007). Furthe rmore, many warmer water species

of the Indo-West Pacific region also occur in Jeju Island.

For the past 12 years surveys have been conducted to

enumerate the mollusk fauna, with special attention to

its biodiversity, biogeography, and ecology. According

to Noseworthy et ah (2007), there are 1,072 mollusk spe-

cies in Jeju Island, and 755 species are exclusively distrib-

uted in marine environments. Due to the extensive surveys,

several species newly reported for the island fauna have

been reported (Noseworthy and Choi, 2010; Noseworthy

et ah, 2012).

Currently, 12 species of Polyplacophora are known to

occur in Jeju Island (Min et ah, 2004; Noseworthy et ah,

2007), and several other specimens which may represent

new species records have been obtained. Cryptoplax

occurs mainly in the warm Indo-West Pacific region as well

as in cooler regions further south, such as South Africa,

Tasmania, and Western Australia (Malacos.com, 2006).

In Japan four species of Cryptoplax have been reported

(Saito, 2000), but only C. japonica Pilsbry, 1895 has been

identified in Korea. This species ranges from Sakhalin

Island southward to the southern Korean peninsula and

Jeju Island. Here we report the occurrence of Cryptoplax

Page 130

THE NAUTILUS, Vol. 128, No. 4

propior Is. and Iw. Tald, 1930, a species newly reported

for the Korean molluscan fauna.

MATERIALS AND METHODS

Two chiton specimens were obtained by SCUBA diving

in tbe autumn of 2012 at Munseom, a small island about

one kilometer south of Seogwipo harbor on the south

coast of Jeju Island, and at Bukcheon-ri on the north

coast (Figure 1). Both specimens were obtained at a

depth of 5 m. These specimens were identified as

belonging to Cryptoplacidae but did not resemble any

members of this family found in the Korean fauna litera-

ture (for instance, Kwon et ah, 1993, 2001; Min et al,

2004). An examination of the Polyplaeophora in Saito

(2000) and other Japanese literature revealed that it was

a specimen of Cryptoplax propior Is and Iw. Tald, 1930.

The species has not been previously reported from the

Korean Peninsula.

The specimens were measured, examined with a ste-

reo microscope, then fixed in 70% ethanol. They were

compared with the original description of C. propior and

with specimens of C. japonica, which most closely

resembles C. propior. Valves and girdle were dissected

from one of the specimens for further study, and a scan-

ning electron microscope (JSM-6700F, JEOL Korea

Ltd.) was used for the examination of each valve and the

perinotum spicules. The material is stored at the School

of Marine Biomedical Science, Jeju National University

(#L145S1005).

As the Jeju Island material is separated from the orig-

inal material in both space and time, it was deemed

appropriate to redescribe the valves and girdle, the most

distinctive features of this species. An examination of the

morphology of the radula would have been useful but

such a study could not be conducted because the speci-

mens had been dried before being placed in ethanol.

116°E 120°E 124°E 128°E 132°E 136°E

Figure 1. Sampling sites of specimens of Cryptoplax propior : ■

This created difficulties in extracting the radula and ren-

dered it unusable. Radula studies can be done if more

material becomes available in the future. The taxonomy

used follows that of Sirenko (2006), and the descriptive

nomenclature is based on that of Schwabe (2010).

An appendix to this work, by the same authors,

Appendix 1. A Catalogue of Cryptoplax Species, is

posted online at http://nautilus.shelhnuseum.org.

SYSTEMATICS

Order Chitonida Thiele, 1909

Superfamily Cryptoplaeoidea H. and A. Adams, 1858

Family Cryptoplacidae H. and A. Adams, 1858

Genus Cryptoplax Blainville, 1818

Type Species: Cryptoplax larmefonnis (Burrow, 1815),

by subsequent designation.

Diagnosis: Mainly moderately-large to large vermiform

chitons with reduced tegmentum and wide, rather fleshy

girdle covered with spicules of varying shape and size.

Cryptoplax propior Is. and Iw. Taki, 1930

(Figures 1-19)

Type Locality: “Prov. Shima” (Shima Peninsula, now

part of Mie Prefecture, southeastern Japan).

Material Examined: Jeju Island, Seogwipo-shi,

Munseom (one specimen); Jeju Island, Bukcheon-ri

(one specimen.)

Dimensions: Length, 22 mm, width, 7 mm (extended)

(Munseom); Length, 8 mm, width, 4 mm (curled)

(Bukcheon-ri.)

Bukcheon-ri; •: Munseom

R.G. Noseworthy et al., 2014

Page 131

Figures 2-1 1. Cryptoplax propior. 2, 3. Munseom, Jeju Island. 2. Dorsal view. 3. Ventral view. Scale bar = 1 cm. 4-1 1. SEM view

of ultrastructure of individual valves. 4. Head valve. 5. Second valve. 6-10. Median valves. 1 1. Tail valve. Scale bar = 1 cm.

Description: Tegmentum strongly reduced. Anterior

four valves slightly overlapping; posterior four valves more

widely separated (Figure 2). Color golden-brown; grooves

between granulose rows somewhat lighter. Head valve

rounded with moderately straight posterior margin, and

possessing several indistinct granulated radial ridges and

growth lines (Figure 4). Second valve round with wide,

smooth jugum, tapering posteriorly, with low, longitudinal,

lightly-beaded ridges. Apophyses moderately long, well-

developed, with rather wide, shallow jugal lamina (Fig-

ure 5). Third to seventh valves (Figures 6-10) with about

eight well-defined, slightly radiating, rather granulose rows

on pleurolateral areas; jugum distinct, nearly parallel-sided.

Valves narrow with slightly-beaked posterior ends; apophy-

ses and jugal laminae similar to those of second valve but

with narrower laminae. Tail valve (Figure 11) possesses

posteriorly-terminal mucro directed backwards, apex

overhanging terminal margin; antemucronal area

exhibiting well-developed jugum. Radiating granular

rows, apophyses, and jugal laminae similar to those of

other valves. Length, height, and jugum length of valves

provided in Table 1. Perinotum with dense, short, rather

blunt spicules of unequal size, light purplish-brown mot-

tled with grayish-tan, with light tan base. Girdle fringe

grayish-white with irregular, indistinct reddish-brown

areas, possessing rather short, rounded spicules ( Figure 2).

Foot grayish-brown with posterior area somewhat pointed

(Figure 3), hyponotum light tan.

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THE NAUTILUS, Vol. 128, No. 4

Table 1. Cryptoplax propior. Length, height, and jugum

length of individual valves, from anterior to posterior

(Munseom specimen). Measurements in mm.

Remarks: The shape of the second, smaller specimen

is more uniform, with parallel sides. The valves are sim-

ilar in morphology to the dissected specimen, but are

somewhat closer, suggesting a subadult specimen. How-

ever, there is no clear division between the head valve

and the perinotum, the head valve appearing to blend

into the orange-brown perinotum, which has dense,

short, blunt spicules, somewhat coarser at both ends.

The distinct girdle fringe has longer and coarser spicules

which are whitish with small orange-brown patches. The

foot, somewhat narrow and yellowish-white, is in the

center of the ventral area, the hypnotum being a light

grayish-tan.

Both specimens compare quite well with the original

description and valve illustrations (Figures 14-17). The

short, rather blunt spicules exhibit weak, vertical stria-

tions, as also mentioned in the description (Figure 18).

However, the holotype (Figure 19) is larger with the

extended length estimated at 33 mm, and has an elongate-

oval shape with roundly-pointed extremities. Although the

anterior of the dissected specimen is narrower, gradually

widening to the posterior end, the shape of this specimen

may be an artefact of preservation. Also, the tail valve of

the Korean specimens is somewhat less pointed (Fig, 17).

The color is somewhat different, the girdle of the holotype

being reddish brown, while the dissected specimen has a

purplish tinge and the second specimen is orange-brown.

Additionally, the original description mentions a

unicolored girdle, while the girdles of both Jeju specimens

exhibit some mottling. Coloration in chiton species, often

in the girdle, can be quite variable. Of the two specimens

obtained, the smaller one resembles more closely the

specimen figured in Saito (2000); however, the head valve

in the illustration is more prominent.

Distribution: South-central Japan to Jeju Island,

southwestern South. Korea.

DISCUSSION

Van Belle (1983; see also Kaas and Van Belle, 1998) and

others have placed Cryptoplax in the subfamily

Cryptoplacinae within Aeanthochitonidae, but Sirenko

(2006; see also Sehwabe, 2014) has more recently

accepted the same grouping as a family, Cryptoplacidae.

Chitons belonging to C. propior are apparently much

smaller than other adult members of this genus, and

previously reported only from Izu-Oshima Island, south

of Tokyo, eastern Japan, and the type locality, the Shima

Peninsula, in east-central Japan, (Is. and Iw. Taki, 1930).

Although the species has been known for the Japanese

fauna for more than 80 years, surprisingly little is known

about its distribution, ecology, and life cycle. Is. Taki, in

various studies on Japanese Polyplacophora, made refer-

ence to it (1938; 1961; 1962). Saito (2000) included an

illustration of a curled specimen, and Higo and Goto

(1993) and Higo et al. (1999) included it in their listing

of the Japanese Polyplacophora. Kaas and Van Belle

(1998) also included this species in their catalogue of

Figures 12-13. Cryptoplax propior. SEM views of girdle and valves. 12. Median area. 13. Posterior area. Abbreviations:

s: spicules; f: girdle fringe; g: granular rows.

R.G. Noseworthy et al, 2014

Page 133

Figures 14-19. Cnjptoplax propior. Original illustrations. 14. Head valve. 15. Second valve. 16. Median valve. 17. Tail valve. 18.

Girdle spicule. 19. Holotype (dorsal view). No scale provided. (From Is. and Iw. Taki, 1930, Venus, The Japanese Journal of

Malacology; used with permission.)

Recent chitons, as well as Saito in his discussion of the

taxonomy of the genus Cnjptoplax (Saito, 1994).

The original detailed description made no mention of

the habitat of C. propior or the depth at which it was

obtained; however, Saito (2000), stated that it is found

“on the rhizoids of laminarian brown algae in the

subtidal zone”. Bergenhayn (1933) examined specimens

from the intertidal zone at Misaki (Miura), Saga mi Bay,

which resembled those in the original description, but

were smaller and could possibly be subadults. According

to Bergenhayn, his specimens agreed in the moqihology

of the tegmentum and radula with the original descrip-

tion but the tegmentum color was yellowish-white instead

of brown (Bergenhayn, 1933). Leloup (1940) commented

on Bergenhayns specimens but provided no new details.

Saito (2006) mentioned C. propior in his list of chitons

from the Sagami Sea and questioned Bergenhayns

record, asserting that this species is usually found in the

subtidal zone among rhizoids of laminarian algae. How-

ever, Higo et al. (1999), while including the Misaki refer-

ence, listed the habitat of this species as “intertidal, rocks

and gravel”, suggesting a less-restricted habitat.

Saito (2006) also stated that C. propior closely resem-

bles juvenile specimens of C. japonica. According to

Saito (2000), one of the characteristics that separates

the two species is the presence of ‘radiating rows of

granules” on pleurolateral areas of C. propior. In corre-

spondence, he also stated that the morphology of the

larger spicules on the perinotum is an important feature

in separating C. projnor from C. japonica-, these spicules

are “very short, thick and rather blunt at the tip”, while

those of C. japonica are long, slender, and pointed (Saito,

pers. comm.).

SEM photos of the median and posterior areas of the

perinotum of the Jeju specimens (Figures 12, 13) show

spicules that resemble Saito’s diagnosis, as well as the

radiating rows of granules on the valves (Figure 12). Fur-

thermore, SEM photos of C. propior provided by Saito

compare well with those of specimens from Jeju Island

(Saito, pers. comm.). In contrast, Hong et al. (1999), in

their description of Cnjptoplax japonica, mention the

larger perinotum spicules as being slightly curved,

smooth, and sharply pointed at the tip, differing from

the short, thick, blunt, often striated spicules of the spec-

imens in this study. An examination of the valves and

girdle spicules of juvenile and subadult specimens of

Cnjptoplax specimens obtained from the east coast of

the island, revealed a coarser sculpture of fewer radiat-

ing granular ribs and long, slender, pointed spicules

characteristic of C. japonica, thus confirming the pres-

ence of two distinct species on the island.

CONCLUSION

Cnjptoplax propior has both a restricted distribution and

habitat in Japan. It has now been obtained from two

widely-separated localities in Jeju Island, and may have

a wider distribution there. This species and C. japonica

are the northernmost representatives of the genus

Cnjptoplax, with a relatively restricted range, being

reported from the northwestern Pacific, mainly from

Korea and Japan.

ACKNOWLEDGMENTS

We would like to thank Bruno Anseeuw for literature

and correspondence with Hiroshi Saito, who provided

valuable information and illustrations, as well as Jan

Haspeslagh, who supplied the original description and

illustration of C. propior. Enrico Schwabe made avail-

able many references, as well as a translation of

Bergenhayns comments. Thanks also go to Konstantin

Lutaenko, who provided literature, Doug Eernisse, who

reviewed the manuscript, and provided literature and

many valuable suggestions for improvement, and a sec-

ond reviewer who also gave helpful advice. Dr. K.

Hasegawa, editor of The Venus, gave permission to use

the figures of C. propior from the original description.

This study was supported by National Fisheries

Research and Development Institute of Korea (NFRDI

RP-2013-FR-088).

THE NAUTILUS, Vol. 128, No. 4

Page 134

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B.-G. Lee, and K.-S. Choi. 2010. First report on the annual

reproductive cycle of Burehardi’s cockle, Acrosterigma

( = Vasticarclium ) burchardi Dunker 1877 (Bivalvia:

Cardiidae) on a subtidal sand fiat off southern Jeju Island,

Korea. Invertebrate Reproduction and Development

54: 27-34.

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C08.HTM

Min, D. K„ J-S. Lee, D.-B. Koh, and J.-G. Je. 2004. Mollusks in

Korea. Min Molluscan Research Institute, Seoul, 566 pp.

Noseworthy, R. G. and K.-S. Choi. 2010. The Diversity and

Ecology of Mollusks in Seogundo oil Southern Jeju

Island, Republic of Korea. Korean Journal of Malacology

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Noseworthy, R.G., M. R. Mondol, S.-J. Ju, and K.-S. Choi.

2012. The Occurrence of Clithon retropictus (von Martens

in Kobelt, 1879, Gastropoda: Neritidae) in Jeju Island,

Republic of Korea. The Korean Houranl of Malacology 26:

19-31.

Noseworthy, R.G., N.-R. Lim, and K.-S. Choi. 2007. A Cata-

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Florida United Malacologists (FUM) 2015 - First Announcement

I am pleased to announce that the sixth annual meeting of Florida United Malacologists (FUM) will take place

on Saturday, February 28, 2015, at The Florida Museum of Natural History, University of Florida, in Gainesville.

This one-day meeting brings together anyone with an interest in mollusks to discuss a wide variety of topics including

but not limited to biology, paleontology, archaeology, ecology, and conservation of mollusks. This relaxed and informal

venue offers an excellent chance for students, amateurs, naturalists, citizen scientists, and professionals to meet and

share ideas.

If you are interested in giving a presentation please submit a brief abstract of 150 words or less by February 1, 2015.

The gathering is free and box lunches and dinner at a local restaurant (to be arranged) will be available at cost

to participants. Parking is free, as are most museum exhibits. Behind-the-scenes visits to the FLMNH collection at

Dickinson Hall about a mile away can be arranged for the week before or after the meeting.

Please send inquiries and reservations to John Slapcinsky at [email protected]. The deadline for submission of

abstracts is February 1, 2015. The FUM program, abstracts, and times of presentations will be posted on the FLMNH

web site, http://www.flmnh.ufl.edu/fum2015/home/ shortly after the abstract submission deadline.

John Slapcinsky

Florida Museum of Natural History

University of Florida

(352) 273-1829

[email protected]

THE0NAUTILUS

Volume 128

2014

AUTHOR INDEX

Andres, ]. de 22

Araujo, R 22

Amano, K 9, 122

Behrens, D.W. 114

Bertsch, H 101

Bobos, 1 105

Bogan, A. E 22,28

Bragado, M.D 22

Chiesa, I.L 40

Domingues, B 105

Fernandez-Garces, R 1

Ferreira, S 105

Gerber, ] 59

Gonsalves, j.F 105

Harasewych, M.G 18, 55, 91

Harzhauser, M 51

Hinzmann, M 105

Jenkins, R.G 9

Kang, Y.-C 129

Kiel, S 9

Landau, B 31

Lee, H.-j 129

Lopes, A 105

Lopes- Lima, M 105

Machado, J 105

Mandic, 0 51

Neubauer, T.A 51

Nielsen, S.N 31

Noseworthy, R.G 129

Ohara, M 9

Oleinik, A 122

Ornelas, E 114

Pastorino, G 40

Rolan, E 1

Thompson, F.G 97

Valdes, A 114

Watters, G.T. 65

NEW TAXA PROPOSED IN VOLUME 128

GASTROPODA

Attenuiconus marileeae Harasewych, 2014, new species (Conidae) 56

Bartschia ( Agassitula ) peartae Harasewych, 2014, new species (Colubrariidae) 94

Cheilea africana Rolan and Fernandez-Garces, 2014, new species (Hipponicidae) 6

Cheilea arnericana Rolan, Redfern, and Fernandez-Garces, 2014, new species (Hipponicidae) 4

Cheilea atlantica Rolan, Leal, and Fernandez-Garces, 2014, new species (Hipponicidae) 5

Euglandina encladus Thompson, 2014, new species (Spiraxidae) 100

Euglandina hyperion Thompson, 2014, new species (Spiraxidae) 97

Melanopsis fateljensis Neubauer, Mandic, and Harzhauser, 2014, new species (Melanopsidae) 52

Scutellastra arayae Nielsen and Landau, 2014, new species (Patellidae) 35

Scutellastra venezuelana Nielsen and Landau, 2014, new species (Patellidae) 35

U raho rosphaera Amano and Oleinik, 2014, new genus (PBuccinidae) 123

U raho rosphaera kanekoi Amano and Oleinik, 2014, new species (PBuccinidae) 125

BIVALVIA

Archivesica sakoi Amano, Jenkins, Ohara, and Kiel, 2014, new species (Vesicomyidae)

10

REVIEWERS FOR VOLUME 128

Absalao, Ricardo S.

Anistratenko, Vitalij

Bertsch, Hans

Bieler, Rudiger

Coan, Eugene V.

Cummings, Levan S.

Dixon, David

Eernisse, Douglas J

Gosliner, Terrence M

Goldberg, Richard

Hayes, Kenneth A.

Jimenez-Tenorio, Manuel

Kantor, Yuri I.

Kohn, Alan J.

Krylova, Elena M.

Landau, Bernard

Lima, Silvio F. B.

Lindberg, David R.

Lydeard, Charles

McDonnell, Rory

Faustian, Megan

Pearce, Timothy A,

Qiu, Jian-Wen

Sasaki, Takenori

Squires, R.A.

Thompson, F.G.

Tucker, John K.

Valentich-Scott, Paul

Vermeij, Geraat J.

Watters, G. Thomas

Sponsored in part by the State of

Florida, Department of State,

Division of Cultural Affairs and the

Florida Council on Arts and Culture

CULTURE

BUILDS

FLORIDA

FLORIDA DEPARTMENT of STATE

DIVISION ofCULTURAL AFFAIRS

INSTRUCTIONS TO AUTHORS

THE NAUTILUS publishes articles on all aspects of the

biology, paleontology, and systematics of mollusks.

Manuscripts describing original, unpublished research

and review articles will be considered. Brief articles, not

exceeding 1000 words, will be published as notes and do

not require an abstract. Notices of interest to the mala-

cological community will appear in a notices section.

Manuscripts: Each original manuscript and accompa-

nying illustrations should be submitted to the editor pref-

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Text must conform to the dimensions of 8V2 x 11-inch

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