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Tanabe, K., Shigeta, Y., Sasaki, T. & Hirano, H. (eds.) 2010. Cephalopods - Present and Past
Tokai University Press, Tokyo, p. 77-84.
Introduction
Spirula, with its well-developed internal chambered
shell, is one of the most unusual Recent cephalopods.
Empty shells of Spirula can be found in great numbers
on oceanic beaches in the Atlantic, Indian and West
Pacific Oceans (Bruun 1943, Clarke 1966). As with
many other epi- and mesopelagic circumtropical spe-
cies, Spirula’s distribution is patchy, apparently there is
no gene ow around South Africa between the Atlantic
and Indo-Pacific parts of its range (Nesis, 1998). This
observation challenges the classical taxonomic opinion
that only one biological Spirula species exists. Cryptic
species could be relatively easily detected by molecular
analysis but, unfortunately, well-preserved material of
whole Spirula is scarce, and most collections of Spirula
were preserved in formol; and so it is quite rare to nd
ethanol xed tissue on which DNA molecular analyses
can be conducted. A preliminary study of four Spirula
specimens caught near Fuerteventura (Canary Islands,
Spain) and sequences (from the gene bank) of one Spir-
ula from New Caledonia were examined for indications
about the diversity of the geographically fragmented
populations (Warnke, 2007). Fragments of mitochon-
drial genes, 16S rRNA (16S) and cytochrome oxidase
subunit III (COIII) were analysed. The 16S sequences
proved to be nearly identical (99.8–100%) whereas the
nucleotide sequences of COIII revealed moderate di-
vergence (0.7–1.3%) between the sequences of Spirula
from Fuerteventura and New Caledonia. The translated
COIII amino acid sequences displayed substantial di-
vergence (3.9–4.6%), but this result was supported by
single sequence from a single Spirula from New Cale-
donia, so the marked sequence divergence may be an ar-
tefact. In short, the analysis provided no clear answer to
whether there are multiple Spirula species. The present
study proposes further evidence on this issue based on
shell shape and growth. These morphometric analyses
are computed using intrapopulational (i.e. geographi-
cal), interpopulational and ontogenetic investigations.
Taxonomic framework
Since Linnaeus (1758) first defined the species he
termed Nautilus spirula, taxonomic history has re-
corded a rise in the number of reported Spirula species
(see Young and Sweeney, 2007 for a review) before the
eventual return to just the one Spirula spirula (Linnaeus,
1758). The genus Spirula was defined by Lamarck
(1799). It is now generally considered that this genus is
monospecific. Some have suggested the occurrence of
geographical subspecies (Bruun 1943), one in the Atlan-
tic and the other in the Indo-West Pacic (Nesis, 1998),
but evidence for this is still lacking.
Just how many species of Spirula are there?
A morphometric approach
PASCAL NEIGE*1 AND KERSTIN WARNKE2
1Laboratoire Biogeosciences, Université de Bourgogne, CNRS, 6 Boulevard Gabriel, F-21000 Dijon, France
(*Corresponding author; e-mail:Pascal.Neige@u-bourgogne.fr)
2Freie Universität Berlin, FR Paläontologie, Malteserstr. 74-100, Haus D, 12249 Berlin, Germany
(e-mail: warnke@zedat.fu-berlin.de)
Received May 7, 2008; Revised manuscript accepted November 7, 2008
Abstract. We compare 110 Spirula shells from ve geographical areas. Morphometry provides a criterion for
determining when shell growth ends (decrease in whorl height). Characteristics of adult shells apparently
vary with geographical origin: specimens from Madagascar, New Zealand and Brazil are larger than those
from North-West Africa and Australia. These ndings challenge the monospecic status of the genus Spirula
but fall short of proving the occurrence of more than one species. Supplementary molecular investigations
are called for.
Key words: Spirula, morphometry, ontogeny
Pascal Neige and Kerstin Warnke78
Figure 1. Geographical distribution of Spirula spirula after Okutani (1995) (dark grey) and Reid (2005) (light grey), and location
of sampling areas (each sampling area is indicated on the map by its code and the number of specimens measured). Question marks indicate
imprecise locations. Stars indicate specimens reported by Haimovici et al. (2007). Five main geographical units are dened: NWAF (North-
West Africa), BRA (Brazil), MADA (Madagascar), AUS (West Australia) and NZEL (New Zealand). See Table 1 for further abbreviations.
Main geographical
areas Label n
NWAF West Morocco (coll. Warnke) Mo 7
Fuerteventura (coll. Warnke) Fu 14
Playa del Ingles (coll. Warnke) Ca 6
BRA Caponga Beach (coll. J. Geraldo de Aratanha) BRA-F 3
Canoa Quebrada (coll. M.A. Reis Jr.) BRA-A 10
MADA Port Ostafrika (coll. Museum für Naturkunde, Berlin) PoO 8
Madagascar NW (coll. S. Kiel) MaNW2 10
Manakara (coll. J. Hartmann) MKSE 22
Madagascar SE (coll. J. Hartmann) MaSE 9
AUS South West Australia (coll. K. Bandel) KB 3
NZEL Ninety Mile Beach (coll. F. Riedel) NMB 9
New Zealand (coll. F. Riedel) NZ 9
Table 1. Location and number of specimens used in this study (see also Fig. 1). coll. = collection. n = number of measured speci-
mens for a given locality.
How many species of Spirula? 79
Material and methods
A set of 110 shells was measured in this study. All
were collected from beaches. The specimens measured
were the best preserved ones from a larger sample. They
come from various areas covering a broad span of the
known geographical distribution of Spirula. Five main
geographical areas are dened here based on sampling
localities (see Figure 1 and Table 1): North-West Africa
(NWAF), with specimens from the Canary Islands and
from the coast of Morocco; Brazil (BRA), all specimens
from two localities close to Fortaleza; the Madagascar
area (MADA), with specimens from the east and west
coasts of Madagascar and from Mozambique; South-
West Australia (AUS), with specimens collected not
far from Perth; and the North Island of New Zealand
(NZEL). Because this analysis is based on shells col-
lected from beaches, we cannot exclude the effect of
post-mortem drift. This may be the case particularly
for BRA shells, located outside the known geographi-
cal distribution of Spirula. The results will thus be dis-
cussed (see “Discussion”) on the assumption that ana-
lyzed samples might represent nearby populations.
A new measurement protocol was defined for de-
ciphering shell growth as accurately as possible. The
anterior boundaries of ventral and dorsal sutures (sensu
Naef, 1923) were used as landmarks (Figure 2A). Their
coordinates being known throughout growth (i.e., mea-
sured for each septa), basic trigonometric equations
were used to calculate several parameters with which to
describe the shell (Figure 2B).
A particularly complicated issue is to dene the cen-
tre of the supposed logarithmic spiral of Spirula shell
growth. Several workers have addressed this issue (e.g.,
Landman 1987, Neige 1997 for ammonites). Schindel
(1990) attempted to locate such a starting point for
growth (the apex) when studying gastropod ontogeny.
He concluded that radius measurements included an
oscillating error term arising from mislocation of the
Figure 2. Shell measurements. A. Location of P1 (0;0) and details (left) showing the locations of the successive landmarks. B. Cal-
culated parameters (from landmark coordinates, see main text) used to study shell growth: r is radius, H is whorl height, and α is the angle
between two successive septa.
Pascal Neige and Kerstin Warnke80
Figure 4. Comparative shell growth for the ve main geographical areas (see Table 1 for abbreviations).
Figure 3. Example of individual growth (case of specimen NZ-02). A. radius versus whorl height. This pattern shows
a decrease in whorl height at the end of growth (see main text). B. radius versus α.
How many species of Spirula? 81
apex (see also Neige 1997 for similar considerations on
ammonites). Certain mathematical calculations might
help in defining a theoretical initial growth point of
the spiral but would necessarily assume a perfect loga-
rithmic growth throughout ontogeny. Such mathemati-
cal extrapolation might mask ontogenetic changes in
growth of the shell. Because the present study explores
ontogeny, we prefer to locate the initial growth point at
an anatomical landmark: the dorsal attachment of septa
number 1. By convention, P1 coordinates (dorsal attach-
ment of septa 1, Figure 2A) are x = 0 and y = 0. This
excludes any articial correction of growth parameters,
but produces more prominent articial oscillations dur-
ing early growth stages.
The anterior boundaries of ventral and dorsal sutures
were located using a microscope with transmitted light.
Measurements were made with an optic measuroscope
(Nikon Measuring Microscope MM-60; precision of 1
μm). Measurement error was estimated by measuring
one specimen nine times with complete repositioning
and measurement. Error was very low compared with
differences between specimens from a given locality.
A Kruskal-Wallis test was used to test for differences
in the adult-stage parameters: radius, whorl height,
number of septa and rotational angle. The Kruskal-Wal-
lis test can be applied as a single factor non-parametric
ANOVA case. It is useful for situations where the usual
ANOVA normality assumptions may not apply, which is
the case here because of small sample sizes for Austra-
lia (n = 3) and Brazil (n = 7). This test indicates whether
observed differences between means of a given param-
eter and for several populations are statistically signi-
cantly different. H0 is that observed differences are due
to a sampling effect. H1 is that at least two populations
have statistically different means.
Results
Two complementary analyses were conducted: one
focusing on individual growth; the other comparing the
various growth parameters for the ve main geographi-
cal areas dened here.
Growth
The growth parameters of each of the 110 specimens
were investigated. Because the anterior boundaries of
suture attachments were measured for all the septa of
each specimen, growth could be described very pre-
cisely, particularly in terms of whorl height versus shell
radius.
One pattern is largely dominant (68 specimens; i.e.,
64.5% of the complete sample). This pattern involves
a characteristic decrease in whorl height at the end of
growth (Figure 3A). Only in three specimens (i.e., 2.7%
of the complete sample) is this decrease followed by a
fresh increase in shell height, and even then it is a tiny
increase limited to only one or two septa. 39 specimens
display no decrease in whorl height during growth.
These are interpreted as juvenile or broken shells. The
decrease in whorl height may be considered as an indi-
cation of the imminent end of growth in Spirula. It is
considered here as a homologous feature, indicative of
a homologous stage of growth. For convenience, this
stage (onset of the end of growth) is termed the ‘adult
stage’ in what follows.
Angle variations between two successive septa are
similar between specimens. Specimen NZ-02 (Figure
3B) is used as an example. A rst phase displays high
values (nearly 55°). These large initial angles can be
explained by the bulbous shape of the first chamber.
Thereafter the angle decreases in the course of growth
to about 20°. A further small decrease in septal angle
occurs at the very end of growth.
Population approach
A population analysis is made on adult shells only
(i.e., those meeting the criterion for the end of growth
defined above) to explore geographical differences in
shell characteristics. The five main geographical areas
are used for that purpose (see Figure 1).
Shells from the same areas display similar patterns
of growth in terms of whorl height increase (Figure 4).
Moreover, the growth of specimens is conservative at
the complete sample scale. The main difference between
specimens from the ve main geographical areas occurs
at the end of growth. On average (see Figure 4), shells
from Madagascar attain the greatest whorl heights at
the end of growth. Shells from New Zealand and from
Brazil display intermediate whorl heights; those from
Area Mean Min Max n
AUS 27 25 29 3
BRA 28.7 25 31 7
MADA 30.5 27 34 28
NWAF 27 24 30 17
NZEL 29 26 34 13
Table 2. Mean, minimum and maximum of septa at adult stage (see
main text for end of growth criterion) for the ve main geographical
areas (NWAF (North-West Africa), BRA (Brazil), MADA (Madagas-
car), AUS (West Australia) and NZEL (New Zealand). n = number of
adult shells.
Pascal Neige and Kerstin Warnke82
North-West Africa and Australia the smallest whorl
heights. These results may be seen differently on graphs
and statistically tested. For that purpose, we calculated
the mean whorl height at the adult stage as defined
above, associated, at this adult stage, with the mean ra-
dius, the mean number of septa and the mean rotational
angle for each of the ve main geographical areas (i.e.,
using individual measurements of radius, number of
septa and rotational angle associated with the maximum
whorl height).
Mean values and associated standard deviations are
presented in Figure 5. This conrms previous observa-
tions about differences in whorl height at the adult stage
between geographical populations (Figure 5A). The pat-
tern for the number of septa is very similar (Figure 5B).
On average, at the adult stage, shells from Madagascar
have 30.5 septa (Table 2). By contrast, at the adult
stage, shells from Australia and North-West Africa have
only 27 septa. The pattern for rotational angle at the
adult stage is less differentiated between geographical
Figure 5. Bivariate plots showing differences in measurements for adult shells only. Means and standard deviations are given
for shells from the ve geographical areas (NWAF (North-West Africa), BRA (Brazil), MADA (Madagascar), AUS (West Australia) and
NZEL (New Zealand).
How many species of Spirula? 83
areas (Figure 5C). Table 3 shows the Kruskal-Wallis test
results for these observations. All comparisons reveal
statistical differences among populations, although the
difference for rotational angle is less marked (p = 0.023).
Discussion
This study uses a morphometric exploration of shell
growth to propose pointers for investigating species
structuring within Spirula. This should be seen as a rst
step towards a comparative analysis using both morpho-
metric and molecular approaches. However, a number
of ndings are worth discussing:
1. We propose a criterion for detecting the end of
shell growth based on the recognition of a decrease in
whorl height. This ontogenetic pattern is common in the
populations under study (64.5%). It would be interesting
to compare this criterion with others based on the soft
parts of the animal. This criterion has made it possible
to compare shells from different geographical popula-
tions at an equivalent (adult) growth stage.
2. The comparisons revealed differences in measure-
ments between populations at the adult stage. We sta-
tistically tested differences in values of radius, whorl
height, number of septa and rotational angle. Differenc-
es between populations may be relatively large: for ex-
ample the mean number of septa at the adult stage is 27
for North-West African specimens versus 30.5 for shells
from Madagascar. Similarly, whorl height is nearly 5.78
mm for shells from Madagascar versus nearly 4.86 mm
for shells from North-West Africa, a difference of nearly
20%. Schematically, two separate clusters can be recog-
nized: specimens from Madagascar, New Zealand and
Brazil, with larger dimensions at the adult stage, and
specimens from North-West Africa and Australia. It is
difcult to explain this clustering in terms of geography.
It should be remembered that the analyzed popula-
tions contain shells that might have been subjected
to post-mortem drift. Post-mortem drift is a common
phenomenon within recent shelled cephalopods and
has lead to misinterpretation of geographical ranges of
some of them. For example, cuttleshes have been er-
roneously supposed to occur in the Western Atlantic
Ocean because of cuttlebone post-mortem drift (see
Voss, 1974). Analyzes of shell preservation and epizoan
occurrences on Spirula shells collected from the beach
by Dauphin (1979a, b) were inconclusive regarding the
exact geographical origin of the living population, and
thus the possibility of intense geographical drift. As a
result, she favoured a distant origin because of the lack
of any living specimen caught in the area. Similarly,
shells used here for BRA locality are not included in the
known geographical distribution of Spirula spirula (see
Figure 1). However, Haimovici et al. (2007) recently
reported S. spirula along Brazilian coasts (Bahia state)
southward of the beaches studied here. Only few com-
plete specimens have been found (e.g., not only shells
but nor a living population). Thus, the Brazilian shells
used in the present study may have been drifted, but for
a short distance. In addition, the relatively homogenous
distributions of morphological features within popula-
tions compared to statistical differences of morphologi-
cal features among populations suggest that shell popu-
lations reect natural living populations. More work is
needed to clearly determine whether Spirula popula-
tions live along the Brazilian coasts and thus to identify
the exact extent of geographical drift.
The results obtained here clearly challenge the tradi-
tional monospecic status of Spirula. However, they are
only preliminary findings and do not demonstrate the
existence of two or more species. A molecular analy-
sis is now needed. It would be interesting also to use
a similar morphometric approach but based on sexed
specimens to control for any sexual dimorphism. An
alternative hypothesis to the occurrence of two or more
species to account for the observed morphometric dif-
ferences is ecophenotypy. This could also be tested by a
molecular approach. In practice, this will prove difcult
because complete Spirula specimens are seldom caught.
Although limited in geographical scope compared to the
present study, an analogous analysis based on Nautilus
may help interpreting the present results (Tanabe et al.
1990). In spite of their non-overlapping distributions
(live-caught specimens were coming from the Philip-
H p-values
Radius at adult stage, Geographical areas 43.6 <0.0001 ***
Whorl height at adult stage, Geographical areas 49.7 <0.0001 ***
Number of septa at adult stage, Geographical areas 25.9 <0.0001 ***
Rotational angle at adult stage, Geographical areas 11.3 0.023 *
Table 3. Non parametric Kruskal-Wallis test for various data sets (see main text). H is the Kruskal-Wallis test statistic. Results are
given for ex-aequo correction. *: test signicant at 95% condence level; ***: test signicant at 99.9% condence level.
Pascal Neige and Kerstin Warnke84
pines, Fiji and Palau), Nautilus from the three popula-
tions shared quite similar overall shell morphology but
could be distinguished by adult features such as the
dimensions of the shell and total number of septa. Using
such results and some genetic data (from Woodruff et al.
1987), the authors concluded that the three populations
belong to the same wide-ranging species. Based on ge-
netic and morphological exploration of Nautilus coming
from different geographic areas, Wray et al. (1995) con-
cluded similarly that some of the morphological features
used to dene Nautilus species may represent variation
within one widespread species. Applied to the present
analysis, this would mean that observed morphological
differences between populations could represent varia-
tion within a single species. However, it is worth noting
that the geographic variation described here for Spirula
is much more extensive than that for Nautilus. Finally,
new genetic and morphological data are needed before
to determine Spirula species number.
Acknowledgments
We thank M. Aeppler, J. Geraldo de Aratanha, K.
Bandel, J. Hartmann, S. Kiel, F. Riedel, M.A. Reis
Jr, Ignacio Santana and the Museum für Naturkunde
(Berlin) for kindly providing Spirula shells. Thanks to
Emmanuelle Pucéat for her help in computing trigono-
metric equations to transform landmark coordinates into
distances and angle measurements. We are grateful to
participants at the 7th International Symposium, Cepha-
lopods – Present and Past for their discussions during
the meeting. We thank Kazushige Tanabe for his helpful
comments, and Sylvain Gerber, Melanie J. Hopkins and
Michael Labarbera for checking our English. This work
was supported by the DFG WA 1454/2-1 grant from the
Deutsche Forschungsgemeinschaft (DFG) to Kerstin
Warnke. This paper is a contribution to the FED (Forme,
Evolution, Diversité) team of UMR CNRS 5561 Bio-
géosciences, and to GDR MEF (Morphométrie et Evo-
lution des Formes).
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