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The Biological Characteristics, Life Cycle, and System Design for the Flamboyant and Paintpot Cuttlefish, Metasepia sp., Cultured Through Multiple Generations

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  • Monterey Bay Aquarium, CA, United States

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The Biological Characteristics, Life Cycle, and System Design for the Flamboyant
and Paintpot Cuttlefish, Metasepia sp., Cultured Through Multiple Generations
Bret Grasse, Aquarist II, bgrasse@mbayaq.org
Monterey Bay Aquarium, 886 Cannery Row,
Monterey, CA 93940
BACKGROUND
Metasepia is a genus of small cuttlefish found in the Pacific Ocean. They are characterized
by several definitive traits. The cuttlebones of this genus are small and diamond or
rhomboidal shaped. Cuttlebones are also typically much shorter than the mantle and
located in the anterior half to two-thirds of the mantle. It is believed that the small
cuttlebone size results in negative buoyancy, allowing a unique method of locomotion in
which they spend most of their time walking on the sea floor. The mantle itself is also
unique, as the dorsal anterior edge lacks the traditional tongue-like projections of most
Sepiidae genera. (Jereb and Roper, 2005)
There are two species in the genus Metasepia. Metasepia pfefferi, the flamboyant
cuttlefish, is found in the Indo-pacific waters around Australia, New Guinea, the Philippines
and Indonesia. Metasepia tullbergi, the paintpot cuttlefish, is found from Hong Kong to
southern Japan. The two species can be quite challenging to differentiate based on
morphology, making genetic analysis an important tool in species identification. Both
species of Metasepia are neritic demersal species generally found in sand and mud
substrates at shallow depths from 3-100m. Metasepia breeding is seasonal with mature
individuals migrating into shallow water to spawn in the spring, and juveniles migrating to
deeper depths in the late summer. (Reid et al., 2005) Metasepia lay individual eggs in
crevices, on coral rubble, or under overhangs. Adults will live an average of 6 and 8
months, and throughout their life span they feed on various crustaceans and fish.
Commercially, Metasepia have been collected for the public and private aquarium trade.
Unfortunately, success in captivity has been limited until recently. Ultimately, shipping has
made it quite challenging to get healthy individuals from source to destination. These
animals traditionally have shipped very poorly, which generally results in significant
mortality occurring in transit. Furthermore, in the past, successful rearing of newborn
hatchlings has been highly problematic.
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BIOLOGY
For both species of Metasepia, gender can be determined starting at 90 days. At this point
in time, there are some behavioral and physical distinctions indicating gender. The males’
bodies will typically elongate and turn almost completely white in color during courtship.
Males will retain some pink color at their arm tips and a yellow/brown coloration on their
posterior mantle. During typical courtship display, males will approach females face to
face and bob their heads back and forth on a horizontal axis (Image 1).
Another predictable method for determining gender is the size differential of males and
females at maturity. At around 90 days of age, the growth rate of each individual changes
depending on its sex. This results in a drastic sexual dimorphism in adults whereby
females are larger than males. It has been noted that with some cephalopods it is
necessary to create two growth curves: one exponential for the immature individuals and
one logarithmic for the adults, creating a sigmoid pattern (Domingues et al. 2006). This is
the case for Metasepia, and the following curves demonstrate the increase of dorsal mantle
length and mass (Figs 1, 2) with respect to age. The immature individuals follow a typical
exponential growth pattern, which slows upon maturity for males, but remains exponential.
Females, however, transition to logarithmic growth upon maturity. The curves fit to these
data are significant, with most R^2 values above .8. The rate of growth of a mature female
is greatest upon maturation, and decreases with age. And so females grow faster and
reach larger sizes, slowing down as they approach appropriate reproductive size.
Figure 1
Mantle Length (cm) growth curves in
developing Metasepia at MBA
Image 1
Male depicted on left displaying courtship
behavior to female on right. Females have
occasionally been observed demonstrating
this behavior in territory disputes but it is
uncommon.
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Additionally, the ratio of mass to mantle length is much higher in females, demonstrating
that the females invest more energy into increasing their body mass (i.e. gonad
development) than they do to their length (Fig. 3)
Reproductively mature females began laying eggs at four months of age. The youngest
female to record a clutch was 108 days old. The oldest female to record a clutch was 213
days old. The median age at which females deposit egg clutches is 151 days. Female
Metasepia can lay eggs through senescence. On average there were seven clutches per
female through their lifespan. Females typically laid larger clutch sizes as they aged
through their reproductive period. Figure 4 depicts the reproductive trend for one of our
mature females. This trend is typical for most of our female broodstock. For this particular
female, the first two clutches were small and non-viable and the clutch size increased
exponentially as she aged with a minimum of 11 eggs and a maximum of 106 eggs.
Figure 2
Mass (g) growth curves in developing
Metasepia at MBA
Figure 3
Mass differentials for each gender post
maturity at MBA
Figure 4
Clutch size in females during laying period
and embryo viability trends
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In captivity, Metasepia will lay their eggs in live rock crevices, empty coconut shells, or in
any other well protected artificial décor arrangement. Eggs are opaque and ovular when
they are first laid (~1cm in diameter). As cell division proceeds and the embryo develops,
the eggs become transparent and increase in size. Gestation period was a minimum of 18
days to a maximum of 32 days. Average gestation was between 23-29 days. After
hatching, juvenile growth was exponential until reaching sexual maturity at around 90 days
(Fig. 5; Fig. 6)
HUSBANDRY
System Design
The most commonly used holding at MBA for our sub-adult/adult system is a 230 L
fiberglass tank with dimensions 60cm X 90cm X 50cm. Our holding tanks have three
opaque sides that aid in prey detection and one clear acrylic window for observations.
The Monterey Bay Aquarium utilizes a semi-closed, flow through system which greatly
assists with maintaining optimal water quality. Protein skimmers or other mechanical
filtration can be used depending on system specifics and the bio-load of the Metasepia
culture. These extra filtration methods will assist during inking events, a relatively
uncommon event in our experience with Metasepia cultures when compared to other
sepioid species.
Figure 5
Juvenile exponential growth rate trend in mantle length (cm)
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Like many cephalopods, the skin of Metasepia possesses innumerous microscopic
epidermal projections, a microvillus epidermis, which drastically increases the total surface
area of the skin. This increases susceptibility to elevated ammonia and nitrogenous
compounds in the water column. Due to this sensitivity, water quality needs to be carefully
monitored and managed.
A water temperature of 77° F (25° C) has resulted in consistent and reproducible culture
results, however, this temperature may be raised or lowered by (+/- 2° F; 1° C) as needed
depending upon institutional culture goals. Higher temperatures result in more rapid
growth, but a shorter lifespan. Conversely, lower temperatures provide for a longer
lifespan, but a reduction in growth rate (Domingues et. al. 2002).
Substrate is important to incorporate on the holding tank’s bottom so individuals don’t rub
their sensitive ventral mantle on the tank’s fiberglass surface. The substrate also helps
Metasepia with their crawling locomotion behavior. Crushed coral or sand works well. For
sub-adults and adults, it is important to add vertical relief in their holding. Live rock, faux
sea grass, or empty coconut shells work well. The additional décor helps maturing
Metasepia by providing environmental complexity and visual security in the form of refugia
and/or opportunities for escape from conspecifics.
Air diffusers should not be utilized because the tiny bubbles they create can occasionally
get trapped inside the mantle cavity. Trapped air will negatively impact buoyancy control
and increase stress. Traditional routine maintenance used in aquaria tends to increase
stress levels in Metasepia and may hamper development of this culture. Siphoning/gravel
washing should not be performed more than once daily. Similarly, window maintenance
should only be conducted as needed. This maintenance will only add stress to the captive
population and hinder culture progression. Using live food and low light levels to decrease
the necessity of stressful “routine maintenance” is recommended for Metasepia.
Feeding
Metasepia sp. larger than 2cm mantle length (~2 months old) should be on a diet of glass
shrimp, Palaemonetes sp., or other similar live feeder shrimp (see the “Hatchling Care”
section for feeding instructions for younger individuals). These Palaemonetes are a small
(1-3cm) brackish water species of shrimp. These can be obtained from a number of
venders and retailers, MBA orders through Northeast Brine Shrimp in Oak Hill, FL.
Regardless of life stage, Metasepia prefer shrimp to crabs and fish. The mono-specific
shrimp diet resulted in no observable untoward effects on growth or reproductive output of
Metasepia. Glass shrimp should be size-selected based on the size of the cuttlefish cohort
being fed. Prey items should be smaller than 3/4 the length of the mantle. Larger prey
items may damage Metasepia’s delicate feeding tentacles. Prey items of inadequate/small
size will not provide an adequate nutrition for healthy growth and reproduction.
Feeding sessions should be carried out three times daily. The Metasepia should be fed to
satiation at each feeding session. Most individuals will accept 2-4 dietary items during
each session (depending on the size of the food offered). Attempts to entice Metasepia to
consume frozen food have been moderately successful at best. The use of a clear feeding
stick to offer thawed, frozen food (shrimp, prawn, fish, etc) has been associated with the
most successful non-live feedings. When one considers the time investment in offering
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frozen diets versus the minimal expense of live glass shrimp, little to no advantage to
frozen diets has been demonstrated.
Excessive stress-related behavioral displays, such as uncontrolled buoyancy or
intense/abnormal color flashing, may be an early indication of nutritional deficiency. Live
food should be added immediately. If there is obvious prey present and the stress-related
behavior persists, do not add more food and look for other sources of stress (i.e.
aggression, water quality, etc).
Reproductive considerations
When selecting broodstock, the male to female ratio in each holding tank warrants
consideration. If there are too many males, excessive competition for available females will
occur and stress-related behaviors will likely ensue. Signs of stress-related behavior
include uncontrolled buoyancy, excessive color flashing, inking, and jettisoning into the
holding tank walls. Additionally, males may flush out a competing male's spermatophores
with their siphon (Hanlon et. al. 1999), thus hindering reproductive efficiency. If there is a
heavy female to male ratio, reproductive females will be forced to compete for food
resources and laying grounds. This concern is dependent upon the size of the enclosure.
A male to female ratio of 1:2 seems to be a reproductively efficient pairing for typical 75-
200 liter holding spaces. Maintaining a small reproductive cohort also facilitates more
accurate tracking of genetic lineage in multi-generational cultures.
Sustainable cultures benefit from fresh genetic input every second or third generation. In
Sepia officinalis and Sepia pharaonis, inbreeding directly affects morphological
appearance/condition and causes a reduction in size through progressing generations (Lee
et. al. 1994). This does not appear to be the case with Metasepia. The Monterey Bay
Aquarium has reared five generations of Metasepia pfefferi and three generations of
Metasepia tullbergi with limited morphological variance through each generation.
Decreasing female fecundity, mating compatibility, and overall reproductive success were
the primary negative outcomes observed in multiple generations of Metasepia cultures.
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Egg care
There are advantages and disadvantages for either natural maternal or artificial incubation
of cuttlefish eggs. Maternal incubation provides consistent and reliable incubation for all
eggs in the clutch. There is also less work for the attending biologist. Maternal incubation
is a consistently reliable option if the goal is to simply have reproductive success. In order
to maximize the reproductive efficiency of broodstock, artificial incubation should be
considered. One significant negative aspect of maternal incubation is the fact that the
female must allocate a large portion of her energy towards egg incubation rather than
maximizing her reproductive productivity. If the eggs are removed for artificial incubation,
the laying female quickly abandons brooding behavior and returns to mating and
subsequent oviposition. Eggs can be removed with medical forceps. Artificial incubation
permits biologists to record gestation period(s), egg hatchability statistics, and note exact
hatch date of progeny. Artificial incubation also allows for an extensive knowledge of the
development and progress of nearly every embryo produced. This greatly reduces the
chance of an unmonitored hatching where the sensitive progeny may be in danger of
cannibalism, starvation, or being flushed to discharge. Harvesting eggs for artificial
incubation also allows alternate males to be introduced to broodstock females, genetically
diversifying the female’s reproductive output.
If eggs are artificially incubated, they should be agitated vigorously for the first two or three
weeks. This initial vigorous agitation/circulation is critical, as it keeps the embryos
oxygenated and prevents the settling of debris and bio-fouling organisms (hydroids,
bacteria, and fungal recruitments). If the agitation is too intense, damage can occur to the
egg tunic. It is important to find the correct balance.
This is one example of a vigorous incubation model that was successful at the Monterey
Bay Aquarium (Image 2).
Image 2
Embryo incubation device used at MBA
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This design was constructed using two 12 fl oz empty plastic bottles as
the incubator shell. A screen mesh partition was adhered to the inside of
one of the bottles using non-reactive bonding compound (i.e. Dow
Corning silicone 999). The two bottles are then combined using the
same bonding compound. The upward facing cap should be removed to
allow a discharge point for air bubbles and exiting sea water. The bottom
facing cap face is removed and replaced with screen mesh, and adhered
to the cap rim for sea water supply. The exiting air bubbles from the air
stone drive fresh sea water in through the bottom and incubate the
embryos. The diameter of the supply/discharge cap openings dictates
flow mechanics within the incubator.
Embryonic development can be observed with Metasepia eggs as the tunic/albumen is
transparent. Days before the yolk is completely absorbed by the developing embryo, the
egg should be transferred to a more passive incubation system. Cephalopod eggs
incubated at the upper end of their species-specific temperature requirements develop
more quickly than at lower temperatures (Domingues et. al. 2002). Therefore temperature
will have direct influence on how long the embryos should be in each style of incubator.
For our culture purposes we found this passive incubation model to be effective (Image 3).
This design utilizes ¾ inch PVC as the primary structure. Sea water supply ports are drilled
to guide fresh sea water over the plastic mesh basket. Sea water supply ports should be
drilled so that there is equal flow over the entire mesh basket. As the hatchlings emerge,
they can drift safely out of the flow and come to rest on the culture tank bottom.
Image 3
Passive incubation device used at MBA
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If excessive bio-fouling occurs on the egg tunic, the permeability of the embryo can be
reduced. This can cause asphyxiation in developing embryos and eventually lead to
embryonic mortality. Similarly, bio-fouling organisms can degrade and weaken the tunic,
leading to pre-mature birth. To remedy this, an immersion bath of 1ml of iodine/Betadine to
1 L of seawater for 10 minute treatments every other day as needed can be applied (Lee
et. al. 1994). This method has resulted in no harmful effects on embryos and has been
successful in limiting the biological growth on the eggs’ external tunic.
Hatchling Care
As with many aquatic invertebrates, the hatchling stage is when Metasepia is the most
sensitive. Positioning the holding system away from high traffic areas or areas with
excessive noise/vibrations is recommended.
Metasepia hatchlings should be kept in smaller holding
tanks ranging from 15 to 200 liters depending on the
hatchling population density. Tanks should be more
horizontal than vertical, since hatchlings spend the
majority of their time on or near the bottom. If the tank is
too vertical, live food items may go un-noticed near the
surface. Tank walls should not be transparent to assist
in the visual detection of prey, as tiny prey items are
difficult for hatchlings to locate with transparent walls. This may be achieved by adhering
an opaque, non-permeable, non-reactive material inside of the holding walls (i.e. kydex).
Seams where this material meets are also sealed with silicone to avoid hatchlings getting
wedged in a corner. Tank discharge areas need to be screened (<3mm) such that
hatchlings are not inadvertently swept out.
Smaller substrate (<1mm) is preferred to aid in locating hatchlings, facilitating population
counts, and making behavioral observations. Smaller substrate also allows hatchlings to
locate food with greater efficiency. Large decor is not necessary for the first two months of
Metasepia sp. life span as it obstructs visual observations of the cohort and hinders prey
detection by the hatchlings.
Flow rates should be low (< 2L/min) during this sensitive developmental period to decrease
stress. Low flow will also allow dietary items to remain in the tank for longer periods of
time, thus extending feeding opportunities.
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Captive light cycles should mimic natural light cycles with 12 hours of daylight and 12 hours
of darkness. Intense light is not recommended as it tends to increase bio-fouling.
Moderate florescent or LED lighting is preferred. Blue-tinted “moonlight” during the night
period is necessary as it decreases captive stress and allows for crepuscular feeding.
Hatchling stocking density should never be greater than one individual for every 7.5 cm².
Hatchlings exhibit uncontrolled buoyancy and stressed behavior if densities are too high.
There is also greater competition for food resources and habitat.
For the first two months of life, captive Metasepia sp. should be fed exclusively live mysid
shrimp, Americamysis bahia. Adult Artemia salina are too nutrient deficient to nourish a
growing cuttlefish. A culture utilizing Americamysis bahia nourished with a healthy Artemia
nauplii (small Artemia hatchlings) diet has proven successful. Mysids should be offered
three times a day at a rate of 3-5 mysids per individual. Excessive prey in the holding tank
may be stressful for the hatchlings and may suppress their natural predatory behavior.
Before each feed, be certain to search for any remaining mysids from the previous feed to
prevent overfeeding. If there are still two or more mysids per hatchling, do not add more.
Offering the correct mysid size is paramount to hatchling survivorship. Metasepia less than
three weeks of age should not be fed adult mysid shrimp. In the initial weeks of
Metasepia’s life span, they are too weak to capture adult mysids, therefore younger
shrimp (1-8 days of age) is recommended. Although Metasepia are diurnal, some feeding
occurs at night. It is important to leave two or three extra mysids per hatchling after the
final feed of the day.
Tank maintenance, such as siphoning or gravel washing should only be conducted once a
week. The live feeding regime and the small size of the Metasepia help preserve adequate
water quality for a longer time period. Additionally, gravel washing is an added stressor
that is important to avoid during early development.
SHIPPING CONSIDERATIONS
Hatchlings should be at least four weeks old (~1.5 cm ML) before shipping. Use 1-10 liters
of sea water per individual, depending on the size of the Metasepia. The shipping water
needs to be highly oxygenated prior to shipping (150%-200% dissolved oxygen). The
preferred shipping method does not utilize an air/oxygen air space sealed inside the
shipping bag. Removing any empty air space greatly reduces agitation inside the shipping
bad and decreases stress during transit. Empty air space can also lead to air becoming
entrapped in the cuttlefish's mantle cavity and subsequent buoyancy
issues and stress. Shipping with no empty air space decreases shipping
stress but reduces available oxygen and has other water quality
implications. This concern may be mitigated by shipping Metasepia with
an adequate volume of highly oxygenated sea water. Metasepia should
be packed in individual bags. Smaller individuals are not only less
expensive to ship, but are also associated with a higher survival rate.
Temperature considerations are necessary depending on the geographic location the
cuttlefish will be shipped to. Heat/cooling packs will need to be implemented accordingly.
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Shipping animals on the cold side of their biological requirements will lower the metabolic
rate of the animals and hold more oxygen.
Metasepia less than six weeks of age should not be fasted prior to shipping. Fasting
hatchlings at this age is dangerous due to the high metabolic requirements at this early
developmental stage (Foresthye et. al. 1994). The impact of malnutrition over a 24 hour
shipping period is actually of greater concern than deteriorating water quality.
Consistent and efficient shipping methods are important in facilitating successful Metasepia
culture collaboration between institutions. This collaboration will prove to support genetic
diversification between various cohorts of the species The ability to maintain and share
diverse genetic populations not only enhances the sustainability of the species in zoos and
aquariums, but also significantly reduces potential collection pressures of wild populations.
Conclusion
The flamboyant and paintpot cuttlefish (Metasepia) are highly charismatic marine
invertebrates with significant potential for public display. Until recently, however, the
transport and captive culture of these species have proven to be significantly challenging
and largely unsuccessful. Advancements in Metasepia propagation and transport
techniques at the Monterey Bay Aquarium have proven that these species can be
successfully maintained in captivity over several generations. Cultured offspring have
subsequently been provided to several other AZA institutions. MBA developed husbandry
techniques have allowed some of those institutions to begin successful propagation efforts
of their own.
These techniques will serve to relieve wild collection pressure on what has historically been
a challenging genus to handle and bring to public display. Ultimately, we hope that what
we have learned about Metasepia biology and captive life cycle will help to inform future
understanding and conservation of these species in the wild.
Acknowledgments
I would like to acknowledge the Monterey Bay Aquarium staff, volunteers and interns for
their diligent, detail oriented work with throughout this culture. In particular, Ellen Umeda
(now at SLP Toronto) was a major contributor to this project during her internship at MBA. I
would also like to thank Alicia Bitando for assisting with data analysis and graph creation.
Thanks to Marcus Zevalkink, Paul Clarkson, and Dr. Mike Murray for assisting with the
writing and editing process. Thanks to Randy Wilder for the beautiful photographs. A final
thanks to the MBA husbandry management team for their continued support on this project.
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... All water parameters were kept within optimal ranges as outlined in previous reports of cephalopod culture (Hanlon and Forsythe, 1985;Grasse, 2014). Temperature, salinity, pH, and dissolved oxygen levels were constantly monitored via automated systems (Apex Fusion Neptune System) and an additional water chemistry panel was conducted weekly. ...
... These findings emphasize the need to periodically supplement the population with new genetics via wild-collected octopuses. Decreased fertility in later generations of laboratory culture has been demonstrated in several cephalopod species including the common cuttlefish, S. officinalis (Forsythe et al., 1994), the flamboyant cuttlefish, Metasepia pfefferi (Grasse, 2014), and the bigfin reef squid, Sepioteuthis lessoniana (Walsh et al., 2002). More research on inbreeding depression and fecundity in O. chierchiae is required to help identify genetic bottlenecks and modify protocols to optimize breeding efficiency. ...
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... With regard to camouflage, Thomas and MacDonald (2016) show only one image of camouflage (their Fig. 4C) and reported that it is seldom seen in aquaria; moreover, camouflaged body patterns are not often observed in most public aquaria (see also Grasse, 2014). By contrast, in nature this species is camouflaged nearly all of the time, due in part (or whole) to the very different sensory input in nature versus laboratory environments. ...
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... The gladius extends exactly from the anterior to the posterior part of the mantle in hatchlings of nearly all Decabrachia, so the ML is effectively the gladius length. There are some rare exceptions that were taken into consideration: in the genus Idiosepius a delicate membranous gladius covers only about two-thirds of the ML (Hylleberg & Nateewathana, 1991), in the sepiolids Rossia and Semirossia the gladius extends for some 70-75% of the ML (Bizikov, 2008), and the cuttlefish genus Metasepia have a sepion of half to two-thirds the ML (Grasse, 2014;B. Grasse, personal communication). ...
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Family Sepiidae Cephalopods of the World. An Annotated and Illustrated Catalogue of Cephalopod Species Known to Date
  • A Reid
  • P Jereb
  • C F E Roper
Reid, A., Jereb, P. and Roper, C.F.E. 2005. Family Sepiidae. In: P. Jereb and C.F.E Roper (eds), Cephalopods of the World. An Annotated and Illustrated Catalogue of Cephalopod Species Known to Date. Volume 1. Chambered Nautiluses and Sepioids (Nautilidae, Sepiidae, Sepiolidae, Sepiadariidae, Idiosepiidae and Spirulidae), pp. 54-152. FAO, Rome.
  • M D Norman
Norman, M.D. 2003. Cephalopods A World Guide. ConchBooks, Hackenheim, Germany.