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O’Shea et al.—First records of Nototodarus gouldi egg masses 161
from 1.0 to 2.0 m diameter were reported by divers
off the Poor Knights Islands (Fig. 1); a further sight-
ing was reported from Deep Water Cove, Bay of
Islands, in November of 1997 (Table 1).
In photographs, these structures bore striking re-
semblance to egg masses from laboratory-spawned
ommastrephid squid genera Todarodes and Illex
(O’Dor & Balch 1985, Bower & Sakurai 1996).
Accordingly, the gelatinous spheres were tentatively
identified as the egg masses of an unknown species
of squid.
This paper reports the most recently observed
sphere, a 1.5 m diameter mass encountered in the
entrance to Rikoriko cave, Poor Knights Islands on
26 March 2003, for which excellent photographs,
film, and tissue samples are available. The egg mass
sample contained embryonic squid, but the embryos
were too undeveloped to attribute to any recognised
taxon based on morphology (mantle and funnel-lock-
ing cartilages, tentacles and arm suckers were still
undeveloped). The egg mass disintegrated 2 days
later, resulting in the loss of all embryos. As a pre-
cautionary measure, in the event the embryos could
not be hatched, five had earlier been preserved in
90% ethanol.
Egg masses have been described or their type is
known for the following squid in New Zealand
waters: two Sepioteuthis and three Sepioloidea
species, both of which deposit and attach eggs in
gelatinous clusters to the seabed; seven locally
distributed enoploteuthid squid (Enoploteuthidae)
and one Brachioteuthis (Brachioteuthidae) species,
all of which release eggs individually into the plank-
ton (Young & Harman 1985; Young et al. 1985); and
a commercially important squid, Thysanoteuthis
rhombus (Thysanoteuthidae), for which the egg mass
is described as a cylindrical structure of 0.6–0.7 m
length and 0.15–0.2 m diameter (Sanzo 1929).
The egg masses of the other 76 (approximate)
species of squid represented in museum collections
from New Zealand waters were unknown.
In the present study we have used mitochondrial
DNA (mtDNA) sequences to identify the first-
known egg mass of the commercially important
STEVE O’SHEA
KATHRIN S. BOLSTAD
Earth and Oceanic Sciences Research Institute
Auckland University of Technology
Private Bag 92 006
Auckland 1020, New Zealand
PETER A. RITCHIE
Allan Wilson Centre for Molecular Ecology
and Evolution
Massey University
Private Bag 102 904 North Shore
Auckland, New Zealand
email: steve.oshea@aut.ac.nz
Abstract The egg mass and embryos of the
ommastrephid squid Nototodarus gouldi McCoy,
1888 are reported for the first time, their identity
confirmed by mitochondrial 16S rDNA sequence
determination. The egg mass is a free-floating ge-
latinous sphere of at least 1.5 m diameter and con-
tains an estimated several thousand randomly
distributed eggs; similar egg masses recorded from
north-eastern New Zealand waters of 1.0–2.0 m di-
ameter are reported. Observed fluctuations in
populations for this and other squid species may be
a partial result of trawl damage to the egg masses.
Keywords Nototodarus; Cephalopoda; Ommas-
trephidae; egg mass; trawling
INTRODUCTION
In recent years, nine reports of large gelatinous
spheres sighted off north-eastern New Zealand have
been brought to our attention. Between November
2001 and March 2003, eight of these spheres ranging
Z03030; Online publication date 12 May 2004
Received 24 July 2003; accepted 3 November 2003
New Zealand Journal of Zoology, 2004, Vol. 31: 161–166
0301–4223/04/3102–0161 © The Royal Society of New Zealand 2004
First records of egg masses
of
Nototodarus gouldi
McCoy, 1888
(Mollusca: Cephalopoda: Ommastrephidae), with comments on
egg-mass susceptibility to damage by fisheries trawl
162 New Zealand Journal of Zoology, 2004, Vol. 31
Fig. 1 Location map (Poor
Knights Islands, Bay of Islands,
north-eastern New Zealand),
depicting sites where egg masses
have been observed.
Table 1 North-eastern New Zealand records of gelatinous squid egg spheres.
Date Location Depth Diameter
26 Mar 2003 Rikoriko Cave 22 m; 20°C 1.5 m1
17 Mar 2003 Middle Arch <30 m Not cited2
16 Dec 2002 Trevor’s Rocks <30 m ~1 m3
9 Apr 2002 Anne’s Reef 14 m ~1 m2
23 Mar 2002 Butterfish Bay 30 m ~2 m4
23 Mar 2002 Landing Bay Pinnacle Not cited Not cited4
5 Feb 2002 Eastern side of Ngaio Rock 10 m 1.5–2.0 m5
Nov 2001 Middle Arch 15 m ~1.3 m6
Dec 1997 Deep Water Cove, Bay of Islands 12 m 1.0 m7
Observations courtesy of: Pete Mesley1; Wade Doak2; Haydon Spencer3; Ross & Diane Armstrong4;
Tony & Jenny Enderby5; Dave Abbot6; Toby Bernard7.
ommastrephid squid genus Nototodarus, and
describe the first structure of its type from New Zea-
land waters. Furthermore, in light of the tenuous
nature of the egg mass, its size and bathymetric dis-
tribution, we take the opportunity to express concern
regarding the inadvertent damage to or destruction
of these spheres by trawlers passing through spawn-
ing aggregations of squid.
METHODS
In 1981, the ocean surrounding the Poor Knights
Islands, off the East Coast of Northland, was estab-
lished as New Zealand’s second marine reserve. This
reserve extends for 800 m offshore around these is-
lands; the taking or disturbing of any marine life
within the reserve is prohibited. In anticipation of
summer egg-mass sightings in this region, a special
permit issued under the Marine Reserves Act by the
New Zealand Department of Conservation, after
consultation with the Ngati Wai Trust Board, was
granted. This allowed divers to collect a sample of
~50 eggs from an egg mass from within the reserve,
to enable species identification (through both mor-
phological and DNA analyses). On 26 March 2003
an egg mass of 1.5 m diameter was sighted, and a
sample of ~50 eggs was removed from it. Within
10 h of collection the egg and gel matrix was trans-
ported ashore in a container of water, returned to
Auckland (AUT) and placed within a fine-meshed
bag suspended within a 200-litre hexagonal tank, in
which the seawater was maintained at a constant
temperature of 20°C. The status of the gel and
O’Shea et al.—First records of Nototodarus gouldi egg masses 163
Fig. 2 Egg mass, 2 m diameter, 30 m depth, Butterfish
Bay, Poor Knights Islands (photograph courtesy Ross
Armstrong).
Fig. 3 Close up, egg mass, 2 m diameter, 30 m depth,
Butterfish Bay, Poor Knights Islands (photograph cour-
tesy Ross Armstrong).
Fig. 4 Egg mass, 1.5–2.0 m diameter, 10 m depth, east-
ern side of Ngaio Rocks, Poor Knights Islands (photo-
graph courtesy Tony Enderby).
Fig. 5 Egg mass, 1.5 m diameter, 22 m depth, entrance
to Rikoriko Cave, Poor Knights Islands (photograph
courtesy Pete Mesley).
Fig. 6 Embryos removed from 1.5 m diameter egg
mass, 22 m depth, entrance to Rikoriko Cave, Poor Knights
Islands (photograph K. Bolstad).
2
4
3
5
6
164 New Zealand Journal of Zoology, 2004, Vol. 31
embryonic tissues was monitored daily. Five em-
bryos were removed from the gel matrix and pre-
served in 90% ethanol solution for DNA analysis.
As GenBank mitochondrial 16S rDNA sequences
were not available for N. gouldi, voucher tissues
were sourced for comparative purposes. Only male
N. gouldi, identified on grounds of hectocotylus de-
tail (sensu Smith et al. 1987), were used for genetic
comparison, as female N. gouldi and N. sloanii can-
not be reliably differentiated (they are sympatric in
the area from which fresh Nototodarus tissues were
sourced, the Wairarapa Coast, south-eastern North
Island, New Zealand; Smith et al. 1987).
DNA was subsequently purified from four of the
embryos and one adult male N. gouldi using stand-
ard procedures of phenol-chloroform extraction and
ethanol precipitation (Sambrook et al. 1989). Mock
extractions were performed at the same time to con-
trol for cross-contamination of samples. DNA yields
from each sample were quantified using a Hoefer
DyNA Quant 200 fluorometer. The mitochondrial
16S rDNA gene was amplified with the primers L-
16SAr (5¢-CGCCTGTTTATCAAAAACAT-3¢) and
H-16SBr (5¢-CCGGTCTGAACTCTGAT CAT-3¢)
(Piertney et al. 2003). For polymerase chain reaction
(PCR) amplifications we used 25 ml volumes using
200 ng of DNA and 10 mM Tris pH 8.0, 50 mM KCl,
1.5 mM MgCl2, 0.4 mm of each primer, 200 mm of
each dNTP and one unit of Taq DNA Polymerase
(Roche). Thermal cycling was carried out on a
Hybaid Omn-E at 94°C 10 s, 60°C 10 s, and 7°C 45
s for 30 cycles. The resulting PCR products were
purified using the High Pure PCR Product Purifica-
tion kit (Roche), sequenced using the PRISM®
BigDye™ Terminator version 3.1 Cycle Sequencing
kit (Applied Biosystems), and analysed on a 3730
automated sequencer (Applied Biosystems). DNA
sequences were deposited in GenBank under the
accession number AY380810.
The four sequences from the egg mass were
aligned to known sequences retrieved from GenBank
using Clustal w (Thompson et al. 1994) and the
number of pairwise differences between all se-
quences were estimated using PAUP*4.0b10
(Swofford 2002).
RESULTS
DNA was successfully extracted and consistently
amplified from four embryos. The eggs measured
approximately 2 mm in diameter and yielded 5.8–
6.3 mg of total genomic DNA. The replicate egg
samples all produced identical sequences. The result-
ing DNA sequences from the four embryos and the
known N. gouldi sample were aligned to 60 squid
sequences that were retrieved from the database
GenBank. Using the genetic distance analysis the
egg samples were unambiguously assigned to the
species N. gouldi. The number of pairwise sequence
differences between the egg samples and the five
most closely related species were N. gouldi = 0, N.
hawaiiensis = 16, Todarodes filippovae = 18, Illex
argentinus = 19, Sepioteuthis australis = 25, and
Sthenoteuthis oualaniensis = 31.
DISCUSSION
Two species of Nototodarus live in New Zealand
waters: N. gouldi and N. sloanii. Although tissues
of N. sloanii could not be procured in time for in-
clusion in this analysis, because it is the
southernmost of the two species found in New Zea-
land waters it is not possible that N. sloanii released
the large egg spheres herein reported from north-
eastern New Zealand. It is excluded from compari-
son on biogeographic grounds.
Mattlin et al. (1985) suggest that Nototodarus
spawns in the upper 100 m of the water column
(Uozumi et al. 1995), a theory herein supported, in
part, by the identification of N. gouldi egg masses
in the upper 10–30 m.
McGrath & Jackson (2002) estimated mature fe-
male N. gouldi oviduct-egg counts in the range
2176–82 395 and concluded that this squid releases
eggs in series of small clutches, rather than in one
terminal spawning event. Although the density of
eggs in the egg mass here reported could not be ac-
curately determined, we estimated that it contained
at least several thousand eggs. This low estimate
lends support to McGrath & Jackson’s suggestion
that N. gouldi does in fact spawn more than once.
However, for N. gouldi to spawn several times it
must also mate several times. Balch et al. (1985)
reported relative fertilisation rates for eggs spawned
by a captive Illex illecebrosus (Ommastrephidae), as
15% for the first of two masses, to <1% for the sec-
ond. The reduced success for the second mass was
attributed to sperm depletion, because most
spermatophores were discharged to fertilise the first
mass. Although Bower & Sakurai (1996) report
laboratory-spawned egg masses of Todarodes
pacificus with fertilisation rates of 90–95%, these
females died immediately after spawning, with
93 000–110 000 eggs remaining in their oviducts.
O’Shea et al.—First records of Nototodarus gouldi egg masses 165
Given the known range of intraspecific variation
in egg mass diameter described for ommastrephid
squid (Illex illecebrosus 0.4–1.0 m (Balch et al.
1985), Todarodes pacificus 0.4–0.8 m (Bower &
Sakurai 1996)), it is possible that the nine spheres
reported from north-eastern New Zealand waters all
belonged to N. gouldi. If so the species must spawn
off north-eastern New Zealand from late November
to early April. Since the egg masses of the 75 other
species of squid (excluding N. gouldi and the afore-
mentioned taxa) are unknown, it is not clear whether
or not these egg masses are truly conspecific.
Uozumi & Förch (1995) determined that most
larval N. gouldi in central and southern New Zea-
land waters were hatched during the months of April
and June. Artificially fertilised Illex argentinus eggs
hatched in 5–16 days (Sakai et al. 1998), and those
of I. illecebrosus hatched 6–16 days post-spawning
(Durward et al. 1980; O’Dor et al. 1982). Tempera-
ture determined the rate of embryonic development
in both species. Eggs of the confamilial Todarodes
pacificus hatched in 4–6 days (Bower & Sakurai
1996), and 7 days (Watanabe et al. 1996). If embryos
of N. gouldi develop at comparable rates, then the
egg-mass life span is short and paralarval squid
should start appearing in the water column in De-
cember, extending through until mid April. These
estimated hatching dates are considerably earlier
than those proposed for this species from central and
southern New Zealand waters (Uozumi & Förch
1995), indicating that N. gouldi has a separate sum-
mer-spawning population in waters off and north of
the Poor Knights Islands.
CONSERVATION
Development of eggs isolated from the egg mass gel
in Illex illecebrosus has been described as abnormal,
compared with those developing within intact egg
masses (Balch et al. 1985). Collision with a trawl,
or retention of the egg mass in the net itself, may
destroy the structural integrity of the mass and
viability of developing embryos and/or expose
thousands of fragmented embryos to predation by
fish and planktonic organisms. After the outer
surface of the N. gouldi egg mass reported here was
cut for sample collection, video footage revealed
near-instantaneous predation on the inner gel and egg
matrix by reef fish. Moreover, crustaceans,
protozoans, and bacteria rapidly infested the egg
mass once its outer layer was damaged (Bower &
Sakurai 1996).
Since Nototodarus species are locally caught by
both jig and trawl (Uozumi & Förch 1995), it is
likely that any trawl fishery targeting spawning
aggregations will inadvertently damage or destroy
any egg masses present, killing the developing
embryos. This impact on the earliest stages of the
life cycle could be a contributing factor to fluctu-
ating squid populations, both in New Zealand
waters and on an international scale. Serious
consideration should be given to introducing
regulations that would limit fishing activity amid
spawning squid aggregations solely to jigging
techniques.
ACKNOWLEDGMENTS
We thank Keith Hawkins (Department of Conservation),
Wade Doak, Glenn Edney, Tony and Jenny Enderby,
Phil Endle, Peter Mesley, Ross Armstrong, and many
other divers who contributed their time and resources to
look for egg masses and respond to queries on Wade
Doak’s website, www.wadedoak.com. We are
particularly grateful to Glenn Edney and Pete Mesley
for the retention of a sample of one of these egg masses,
without which this contribution would not have been
possible. We are also grateful to Martin Cryer from the
National Institute of Water & Atmospheric Research
Ltd (NIWA), Auckland, New Zealand, for procuring
fresh voucher specimens of Nototodarus gouldi; and
Bruce Marshall, Museum of New Zealand Te Papa
Tongarewa, Carolyn King, University of Waikato, and
one anonymous reviewer for their valued critique of the
manuscript. This research was supported by the Auckland
University of Technology, Massey University, and the
Discovery Channel (USA).
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