Invertebrate Anatomy OnLine
Brine Shrimp ©
Copyright 2001 by
is one of many exercises available from Invertebrate
Anatomy OnLine , an
Internet laboratory manual for courses in Invertebrate Zoology. Additional
exercises can be accessed by clicking on the links to the left. A
glossary and chapters on supplies and laboratory techniques are also available. Terminology
and phylogeny used in these exercises correspond to usage in the Invertebrate
Zoology textbook by Ruppert, Fox, and Barnes (2004). Hyphenated
figure callouts refer to figures in the textbook. Callouts
that are not hyphenated refer to figures embedded in the exercise. The glossary
includes terms from this textbook as well as the laboratory exercises.
Mandibulata, Crustacea sP,
Eucrustacea, Thoracopoda, Phyllopodomorpha, Anostraca C,
Artemiidae F (Fig
by far the largest and most diverse animal taxon, includes chelicerates,
insects, myriapods, and crustaceans as well as many extinct taxa such as
segmented body primitively bears a pair of jointed appendages on each segment. The
epidermis secretes a complex cuticular exoskeleton which must be molted to
permit increase in size. Extant
arthropods exhibit regional specialization in the structure and function of
segments and appendages but the ancestor probably had similar appendages on all
segments. The body is typically divided into a head and trunk, of which the
trunk is often further divided into thorax and abdomen.
gut consists of foregut, midgut, and hindgut and extends the length of the body
from anterior mouth to posterior anus. Foregut
and hindgut are epidermal invaginations, being derived from the embryonic
stomodeum and proctodeum respectively, and are lined by cuticle, as are all
epidermal surfaces of arthropods. The
midgut is endodermal and is responsible for most enzyme secretion, hydrolysis,
coelom is reduced to small spaces associated with the gonads and kidney. The
functional body cavity is a spacious hemocoel divided by a horizontal diaphragm
into a dorsal pericardial sinus and a much larger perivisceral sinus. Sometimes
there is a small ventral perineural sinus surrounding the ventral nerve cord.
hemal system includes a dorsal, contractile, tubular, ostiate heart that pumps
blood to the hemocoel. Excretory
organs vary with taxon and include Malpighian tubules, saccate nephridia, and
organs also vary with taxon and include many types of gills, book lungs, and
nervous system consists of a dorsal, anterior brain of two or three pairs of
ganglia, circumenteric connectives, and a paired ventral nerve cord with
segmental ganglia and segmental peripheral nerves. Various
degrees of condensation and cephalization are found in different taxa.
is derived with centrolecithal eggs and superficial cleavage. There
is frequently a larva although development is direct in many. Juveniles pass
through a series of instars separated by molts until reaching the adult size and
reproductive condition. At
this time molting and growth may cease or continue, depending on taxon.
is the sister taxon of Chelicerata and in contrast has antennae on the first
head segment, mandibles on the third, and maxillae on the fourth. The
brain is a syncerebrum with three pairs of ganglia rather than the two of
chelicerates. The ancestral mandibulate probably had biramous appendages and a
J-shaped gut, posterior-facing mouth, and a ventral food groove. The two highest
level mandibulate taxa are Crustacea and Tracheata.
is the sister taxon of Tracheata and is different in having antennae on the
second head segment resulting in a total of 2 pairs, which is unique. The
original crustacean appendages were biramous but uniramous limbs are common in
derived taxa. The
original tagmata were head but this has been replaced by head, thorax, and
abdomen or cephalothorax and abdomen in many taxa. Excretion is via one,
sometimes two, pairs of saccate nephridia and respiration is accomplished by a
wide variety of gills, sometimes by the body surface. The nauplius is the
earliest hatching stage and the naupliar eye consists of three or four median
includes all Recent crustaceans except the remipedes. The taxon is characterized
by a primary tagmosis consisting of heat, thorax, and abdomen although the
derived condition of cephalothorax and abdomen is more common. Eight is the
maximum number of thoracic segments.
the ancestral thoracopod the thoracic appendages were turgor appendages used for
suspension feeding in conjunction with a ventral food groove. Such appendages
and feeding persist in several Recent taxa but have been modified in many
compound eyes are stalked primitively although derived sessile eyes occur in
is an ancient line of primitive crustaceans found today in inland, relictual,
waters such as snow-melt pools, wet weather ponds, saline or alkaline lakes, or
similar refugia where there are few or no predators.
have no carapace (anostraca = without shell) and are thought to be similar in
many features to the ancestral crustaceans. The body is elongate and shrimp-like
and with little regional specialization of segments or appendages. The
heart is a long dorsal tube extending most of the length of the body and
equipped with segmental ostia. Anostracans
are gonochoric and the gonads are paired, tubular coelomic derivatives. The
excretory organs are saccate nephridia derived from coelomic remnants.
gut is J-shaped with paired digestive ceca. Most
anostracans are suspension feeders and/or bottom scrapers. The
thoracic appendages are broad, flat phyllopods resembling those of the ancestral
exoskeleton is thin, flexible, only weakly sclerotized and is not calcified. The
ladder-like nervous system is typical of primitive arthropods (and annelids) and
exhibits no cephalization of ventral ganglia. Development
includes nauplius and zoea larvae.
are the best examples of primitive crustacean morphology readily available for
study in introductory invertebrate zoology courses. Of
the crustaceans usually available for study in teaching laboratories,
anostracans are the most primitive and provide the closest approximation to the
presumed condition of the ancestral crustaceans.
shrimp, in the genus Artemia,
inhabit inland saline lakes worldwide. They are tolerant of a wide range of
salinities ranging from almost fresh water to saturated. Artemia is
easy to rear from inexpensive, commercially available eggs and is thus easily
studied alive at any time of the year. Artemia
franciscana is the most
common North American species. It
occurs in Great Salt Lake and is the species whose eggs are sold commercially in
North America. This
species produces desiccation-tolerant resting eggs that remain viable in dry
condition for several years. These
eggs can be purchased inexpensively from Carolina Biological, Ward's Natural
Science, or at local pet stores. With
little effort the eggs can be hatched to produce living nauplius, advanced
larvae, and adults for the laboratory. Rearing instructions are provided at the
end of this exercise. Artemia is
restricted to saline lakes and consequently cannot be collected locally in most
is a homogeneous taxon and its members are similar to each other so that any
species can be used for this study. If
possible, living specimens should be used but preserved specimens will also
specimens are suitable for the study of external anatomy but are much inferior
for observation of internal structures. Fairy
shrimps, such asEubranchipus, occur in temporary ponds and ditches, are
morphologically similar to Artemia, and
can be used for this study. Fairy
shrimps are widespread but their occurrence is spotty and unpredictable and they
are not available alive from most supply houses. If
they are available from local wet weather ponds they are recommended for this
prepared wholemount slides of Eubranchipus are
less satisfactory than unmounted preserved specimens.
the dissecting microscope to study an adult brine shrimp (Artemia) or
fairy shrimp (e.g. Eubranchipus). If
the animal is alive, place it in a small square watch glass of
chloroform-saturated water (See Supplies and Recipes Chapter). If
the specimen is preserved, place it in a dish of tapwater. Use
your minuten nadeln to
manipulate the specimen and study it with the dissecting microscope.
anostracan body is divided into three tagma; head, thorax, and abdomen (Fig 1). The
head and its appendages are specialized, as they were in the ancestral
thoracic appendages are similar to each other and are unspecialized phyllopods. The
abdomen lacks appendages.
The head at
the anterior end is composed of five coalesced segments as it is in all
crustaceans (Fig 1). It
bears a pair of stalked, lateral, compound
eyes and a single, median,
unstalked naupliar eye at
the anterior end (Fig 2, 3). The
eyes, although stalked, are not considered to be segmental appendages.
Figure 1. A
female brine shrimp, Artemia
franciscana, viewed from the left. Thoracopod
setae omitted for clarity. Anostraca1La.gif
small chemosensory first
antennae are the appendages
of the first head segment (Fig 1, 2, 3, 19-11A). They
are uniramous, and unjointed.
antennae are larger and are
sexually dimorphic. Those
of adult males are very large and modified to form a clasping organ to hold the
female during copulation (Fig 19-11A). They
are composed of two articles. Female
second antennae are smaller, about the length of the first antennae but much
thicker, and are composed of a single article (Fig 2). Determine
the sex of your specimen.
or upper lip, is a large, median, ventral fold of body wall arising just
posterior to the bases of the second antennae (Figs 1, 2). It
is not paired and is not a segmental appendage. It
extends posteriorly and covers the ventral surface of the head, including the
two, oval, bulging mandibles lie
on either side of the head and are the appendages of the third head segment
(Figs 1, 2, 3). The
mandibles curve medially and touch each other on the midline where the ventral
borders bear teeth.
The mouth is
on the ventral midline between the two mandibles (Fig 2). It
may be necessary to move the labrum aside to see the ventral ends of the
mandibles and the mouth.
first and second maxillae are small and difficult to see. The first
maxilla is larger than the
second and bears a bundle of anteriorly directed setae on its medial edge (Fig
2, 3). The
first maxillae are immediately posterior to the mandibles on the ventral surface
of the head and are used to transfer food from the thoracic appendages to the
tiny, conical second maxillae are vestigial and bear a few setae and the
nephridiopores (Fig 2, 3).
adult excretory organs are the two maxillary
glands, or coxal glands, in the segment of the second maxillae where
they form conspicuous bulges on its dorsolateral surfaces (Fig 3). These
are typical crustacean saccate nephridia. Their
coiled ducts may be visible within the bulges. The
maxillary glands open via the nephridiopores on the second maxillae.
Figure 2. Oblique
view of the right side of the head of a female Artemia.
remainder of the body is the segmented trunk consisting
of an anterior, limb-bearingthorax and
limbless, posterior abdomen. The
thorax consists of 11 independent segments. No
carapace is present and, since none of the thoracic segments is fused with
either the head or with each other, there is no cephalothorax.
thoracic segment bears a ventral pair of leaflike thoracopods (= thoracic
appendages) known asphyllopods (Figs
1, 3). The
11 pairs of phyllopods are similar to each other and exhibit no regional
specialization, differing only in size. The phyllopods are turgor appendages in
which the exoskeleton is thin and flexible and blood pressure is required to
keep the appendages stiff. The
phyllopods are used for swimming, feeding, and respiration.
gross features of the phyllopod are visible with the dissecting microscope but
observation of detail requires preparation of a wet mount and examination with
the compound microscope. At
present you should use the dissecting microscope to study intact phyllopods
while they are attached to the animal (Fig 3). Later
you will remove a phyllopod and make a wetmount. Do
not remove a phyllopod until you have completed the study of the internal
anatomy and then return to the following description for a detailed study. For
now, find only the larger features visible with the dissecting microscope.
are similarities between the parts of the phyllopods and the ancestral biramous
appendage but proposed homologies between these parts are not universally
accepted. The appendages are functionally uniramous although they have parts
that are thought to be homologous to the two rami of a biramous appendage. Each
appendage is flat and leaflike (phyll = leaf) and thus resembles the
phyllopodous portion of the ancestral biramous mixopod. There
is no stenopodous (cylindrical) portion.
Figure 3. Lateral
view of the left side of the head and anterior thorax of a female Artemia.
the large basal protopod attached
along its dorsal edge to the body (Fig 4, 19-11B). Several
processes extend from the lateral and medial borders of the protopod. Any
process from the lateral border of a crustacean limb is an exite and any process
from the medial border is an endite. You can see some of these with the
dissecting microscope but they will be clearer later under the compound
or six endites, some
of which are very small, extend from the medial margin of the protopod (Fig 4,
19-11B). The proximal and distal endites are the largest and easiest to see.
Three much smaller, and harder to see, middle endites lie between the proximal
and distal endites. Note
the array of medially directed setae on the endites.
densely setose proximal endite is often referred to as the gnathobase. Its
setae form a setal comb, or
filter, of finely spaced setae used to filter food particles from the water. The
large distal endite may be homologous to the endopod of
the ancestral biramous appendage.
three large exites on
the lateral margin of the protopod are easily seen with the dissecting
proximal and middle exites do not bear setae. The
middle exite is theepipod, which was once thought to be a gill
although it now appears to be involved in osmoregulation.
distal exite may be homologous to the exopod of
the biramous appendage. It
is the only process attached to the protopod by an articulation. It bears long
plumose natatory setaeused
for swimming. Plumose
setae are feather-like to increase their effectiveness as oars. Later
you can inspect them with high magnification to see the side branches.
Figure 4. The
sixth left phyllopod of Artemia viewed
from its anterior surface. Anostraca4La.gif
anostracans are suspension feeders although a few are carnivores that consume
other species of anostracans. A
longitudinal, midventral food
groove lies between the
gnathobases of the phyllopods (Fig 3, 19-12)). The mouth faces the anterior end
of the groove. Swimming
movements of the phyllopods draw a water current into the groove. The
water is then forced laterally through the setal comb on the gnathobases and
food particles are prevented from leaving the groove. The
food is moved anteriorly in the groove by the setae of the gnathobases. At
the anterior end it is entangled in mucus from the labrum and transferred to the
mouth by the setae of the first maxillae.
a living active brine shrimp in an 8-cm culture dish or square watch glass with
a few milliliters of brine. Add
a drop or two of yeast/Congo red suspension and observe the animal with the
dissecting microscope. Red
food particles quickly accumulate on the setal comb of the phyllopods and in the
food groove. This
colors the food groove red making it easy to visualize. In
a few minutes the red material appears in the anterior gut, rendering it highly
visible as well. Set
the dish aside and return to it later in conjunction with your study of the
digestive system. <
two segments posterior to the thorax are the genital
segments (Fig 1) and bear
the external genitalia. Females
have a conical pouch called the brood
sac (= ovisac) which may
contain eggs (Fig 1, 5). Males
bear a pair of tubular, retractile penes which
can be extended to four times their resting length. The
retracted penes are visible posterior to the last pair of phyllopods in male
The abdomen consists
of the six segments posterior to the genital region and is nearly cylindrical
(Fig 1). The
posterior end of the body is the telson. There
is a caudal furca with
two short rami on the end of the telson. None
of the abdominal segments bears appendages. The anus is
located on the telson at the base of the caudal furca (Fig 5).
specimens should be used for the study of internal anatomy if possible. Many
internal features are not visible in preserved material due to its opacity. Living
specimens should be studied with the dissecting microscope in small dishes of
chloroformed saltwater (for Artemia)
or pondwater (fairy shrimp). This should be supplemented by examination of
wholemounts of small specimens with the compound microscope. Instructions for
preparing the wholemount are provided in the next section.
The gut is
a simple tube extending the length of the animal (Fig 1, 19-11A). Most
of it is easy to see in living specimens, especially if the animal has fed
recently on a yeast/Congo red suspension. The mouth is
located on the ventral midline of the head between the opposing surfaces of the
mandibles (Fig 3). The
short vertical esophagus extends
dorsally from the mouth to open into the stomach above the mouth (Fig 3). The
mouth and esophagus are easiest to see in wholemounts of living specimens viewed
from the side with the compound microscope. The
mouth and esophagus make up the foregut and
arise during ontogeny from the stomodeum, an invagination of surface ectoderm.
Figure 5. Left
side of the posterior thorax and the abdomen of an immature female Artemia. Anostraca5La.gif
The stomach is
an expanded region of the gut in the middle of the head (Figs 3, 6). Two
spherical digestive ceca bulge
from the anterolateral walls of the stomach (Fig 6).
The intestine is
a long tube extending posteriorly from the stomach through the thorax and most
of the abdomen (Fig 1, 5). The
stomach, ceca, and intestine make up the midgut and
are endodermal derivatives. The
midgut is the site of enzyme secretion, digestion (hydrolysis), and absorption. It
is surrounded by the hemocoel and bathed in blood so that uptake of materials
occurs across its thin walls.
intestine joins the short rectum,
or hindgut, in segment 4 of the abdomen (Fig 1, 6). The
hindgut, like the foregut, is ectodermal and is lined by a chitinous
is responsible for formation of fecal pellets and opens to the exterior via the anus between
the caudal furcae. The
anus is equipped with a sphincter.
rectum develops from an ectodermal invagination, the proctodeum. Early
in development the rectum is not yet continuous with the midgut being separated
from it by a partition. Focus
with high power on the junction of the mid- and hindguts and see if there seems
to be a flow of particles from one to the other.
Figure 6. Dorsal,
slightly oblique view of the head of an Artemia protozoea
to the animal left in the yeast/Congo red suspension earlier (or make such a
preparation now). Use
the dissecting microscope to observe the shrimp as it swims in its dish and look
at the gut. By
now the entire length of the digestive tract should be filled with red
particles, making it easy to see. It
takes only about 15 minutes for yeast to move the length of the gut. Remove
the shrimp from the dish and make a wetmount with it for examination with the
compound microscope. The
regions of the gut should be clearly visible with the compound microscope. Save
this slide and refer to it as you study the remaining internal organ systems.
Add tapwater to the slide as necessary so that it does not dry out. <
anostracan hemal system is a good example of the primitive arthropod condition
and it is easy to see how it could develop from the dorsal blood vessel of an
The heart is
a long, median, dorsal tube extending the length of the trunk (Fig 1). It
may be visible in living, and sometimes preserved, material with the dissecting
microscope but it is best studied with the compound microscope using wholemounts
of small living shrimp.
a wetmount of a small (about 5 mm), living, unanesthetized brine
the shrimp on a slide so you will have a side view of the abdomen and then place
a coverslip over the animal. The
coverslip will immobilize the specimen but will not immediately stop the heart. Study
the animal with the compound microscope and find a place where you can see the
abdomen or thorax in side view.
the large, tubular intestine,
which will be opaque if the animal has been feeding. The
transparent, also tubular, heart lies
dorsal to the intestine and is easily seen in good preparations (Fig 1). It
is surrounded by the pericardial sinus, which is a region of the hemocoel and is
not a coelomic space. Anteriorly
the heart becomes the short aorta which empties into the anterior hemocoel.
for the paired openings, the ostia,
in the lateral walls of the heart. Most
trunk segments have a pair. The
easiest one to see, however, is the large, unpaired terminal
ostium at the posterior end
of the heart (Fig 1). Ostia are valved pores in the wall of the heart that admit
blood from the pericardial sinus into the heart lumen.
the beating of the heart. There
is no obvious peristalsis and the entire heart appears to contract
it opens into the hemocoel, whose spaces extend throughout the tissues of the
pressure in the heart lumen closes the ostia so that blood must exit through the
aorta at the anterior end.
for small spherical, or ovoid corpuscles and use them as markers to visualize
the flow of the blood. The
corpuscles tend to move posteriorly in the pericardial sinus outside the heart. Watch
closely and you may see some of them pass through ostia in the walls of the
heart and then reverse direction and move anteriorly once inside the lumen of
the heart. They
move in surges associated with contractions of the heart. Entry
of corpuscles into the heart is easiest to see in the terminal ostium in the
sixth abdominal segment. Sometimes
the blood contains hemoglobin in solution in the plasma. Hemoglobin is most
likely to be present in animals from water with low dissolved oxygen
may be sufficient pigment to impart a pinkish color. <
heart is surrounded by a special compartment of the hemocoel, the pericardial
sinus (Fig 16-7). In
living and preserved specimens this area is relatively free of solid tissues and
is transparent. It
is separated from the rest of the hemocoel, which lies ventral to it, by a
perforated, horizontal septum through which blood flows on its way back to the
of the heart force blood out its open anterior end of the aorta into the body
flows through the hemocoel and over the tissues while making its way
is aided in its flow by movements of the appendages and their muscles. Blood
flows from the body hemocoel into the pericardial sinus through the perforations
in the horizontal partition and then passes posteriorly in the sinus and enters
the ostia of the heart.
Respiratory and Excretory /
gas exchange is accomplished across the permeable surfaces of the phyllopods.
two maxillary glands in
the segment of the second maxilla (Figs 1, 3) are usually referred to as
excretory organs but, in fact, their role is largely osmoregulatory and they
have little to do with the excretion of metabolic wastes. Nitrogen
is lost as ammonia across the phyllopod surfaces.
maxillary gland consists of an enclosed end sac, derived from a coelomic space,
from which a long excretory duct leads to the nephridiopore located on the tiny
second maxilla. The
duct wraps around the end sac and its coils can be seen through the integument
on the side of the head (Fig 1). The
gland is surrounded by hemocoel and bathed with blood. The epithelium of the end
sac is equipped with podocytes and forms an ultrafiltrate of the blood into the
lumen of the end sac. The
ultrafiltrate is modified as it passes down the duct to the exterior. Artemia is
an efficient osmoregulator and is strongly euryhaline, being tolerant of an
impressively wide range of salinities.
keep its blood hyposmotic to environments more saline than about 10 parts per
is something most marine invertebrates cannot do, at least not to the same
extent. Artemia drinks
brine and actively secretes salts from the maxillary glands, epipods, and gut. The
maxillary glands can produce urine four times as salty as the blood. Maintenance
of a hyposmotic blood is facilitated by the impermeability of most of the
exoskeleton, with the exception of the epipods, is impermeable to salts. The
epipods are major sites of active salt secretion. Artemia belongs
to a predominantly freshwater taxon and presumably evolved from freshwater, not
permeability of the epipods in comparison with the impermeability of the rest of
the body surface can be demonstrated with silver nitrate. Remove
some small brine shrimp (about 5 mm is a convenient size) from their dish of
the salt from the outside of the body by placing them in a dish of distilled
water for a minute or two. With
a pipet, transfer one shrimp to a square watch glass of 0.002M silver nitrate. When
the shrimp stops swimming in a few minutes remove it to a microscope slide. Place
it ventral side down on the slide so the phyllopods are splayed to the sides and
add a coverslip. Examine
the specimen with the compound microscope. Silver
nitrate reacts with chloride ions to form an insoluble, opaque silver chloride
precipitate that turns brown in light. Inspect
the shrimp for large dark opaque areas of concentrated silver chloride. Such
areas can form only where a permeable integument allows chlorides from the blood
to come in contact with silver nitrate. Does
the silver chloride seem to be localized or widespread? Pay
particular attention to the phyllopods. Does
any particular area of the phyllopod appear to be especially permeable? If
so, which? Is
this observation consistent with what you already know about the functions of
the parts of the phyllopod? <
nervous system is difficult to study in whole specimens, either living or
consists of a dorsal brain, paired circumenteric connectives, and double,
ventral nerve cord with segmental ganglia. The
brain is a mass of translucent tissue surrounding the naupliar eye in the
dorsal, anterior part of the head. You
may be able to see it in living specimens.
sensory system includes the median naupliar
eye which appears in the
earliest larval instar and persists throughout life. It
consists of three black pigment
cups face laterally and one points ventrally. Two
stalked, lateral compound
eyes, each composed of numerous black ommatidia are also present.
gonads of both sexes are paired tubes located dorsolaterally in the posterior
thorax and anterior abdomen (Fig 1). They
may be faintly visible as transparent, elongate sacs beside the intestine. They
are easiest to see in wetmounts of small, living specimens.
mating the male approaches the dorsal side of the female and holds her with his
enlarged second antennae. The
male twists his body around the female, inserts the penes into the brood pouch,
and deposits sperm. The
partners remain coupled for several hours during which copulation may occur
every few minutes. Eggs
are released unto the brood pouch where they are fertilized and covered with a
kinds of eggs are produced. One
has a thin shell and hatches in the brood pouch. The
other, known as the resting egg, has a heavy shell and can remain viable for
several years out of water and then hatch when immersed in saltwater. This
second type of egg is collected and sold by supply houses and aquarium shops. Both
egg types hatch into nauplius larvae.
now to the postponed study of the thoracic appendages. Use
your nadeln and
fine forceps to remove a phyllopod from your anesthetized specimen and make a
wholemount with it. Select
an appendage from near the middle of the thorax for this purpose. Examine
the slide with the compound microscope. Return
to the description of thoracopods which appeared earlier in this exercise and
find the structures you were unable to see with the dissecting microscope. Be
sure to examine the plumose swimming setae of the endopod and consider how their
structure is adapted for swimming.
a wholemount of a phyllopod if you have not already done so. Use
100 and 400X of the compound microscope to compare the many types of setae characteristic
of the different areas of the phyllopod.
setae of the exopod (distal
exite) are large, strong and plumose, or pinnately branched. Plumose
setae have lateral branches from the central shaft and resemble a feather. These
are swimming setae, or natatory
setae, that function as paddles to increase the surface area of the
appendage and enhance its effectiveness in swimming. The
lateral branches of the seta increase the effective surface area of the seta and
make a little oar of it.
setae of the endopod (distal
endite) are short, fairly stout, and finely serrate. These
are scraping setae used
to scrape algae from hard substrata.
on the three middle endites are
similar but are heavier, generally longer and more coarsely serrate. They
are also scraping setae and these three endites are sometimes called "claws".
setae on the proximal endite are
fine and delicate. They
are very finely plumose although that is not apparent unless the light is
perfectly adjusted. These
are the filter setae of
thesetal comb used
to remove food from the water as it exits the sides of the food groove. Does
the structure of these different kinds of setae seem to be adapted to their
anostracans swim constantly using the phyllopodous thoracic appendages. Waves
of motion pass along the series of phyllopods to draw water and food particles
into the food groove and to create an effective stroke with the swimming setae
so that locomotion and feeding are accomplished simultaneously. Artemia also
feeds by scraping algae from hard surfaces. Anostracans
can make sudden quick moves by flexing the abdomen. The
caudal furca is also equipped with plumose swimming setae.
a dissecting microscope and incident illumination observe a living, active adult
brine shrimp swimming in a dish of water. Note
the orientation of the animal in the water. The
preferred orientation is with the dorsum down butArtemia can
swim right side up also. The
filter-feeding mechanism works best when upside down. Which
appendages are used for locomotion? Do
the antennae seem to be involved in swimming? Does
the animal ever stop swimming? Do
you see any evidence of feeding by scraping the bottom? Watch
for movement resulting from flicking the abdomen. How
does it differ from motion produced by phyllopods? <
with its three eyes, is sensitive to light intensity and exhibits highly
variable responses to light. In
general, Artemia is
positively phototactic at low light intensities and photonegative at medium and
high intensities. The
response varies, however, depending on the physiological condition of the
animal, wavelength, age, salinity, pH, and metabolic condition.
a dish containing larvae in a part of the room where it will receive uneven
the dish for 15 minutes or so and then observe the distribution of shrimp. Do
they seem to be clustered in any particular region of the dish? How
does this relate to the light source? You
may want to design a series of more carefully controlled experiments to
determine the effect of light intensity, life history stage, salinity,
wavelength, or temperature. <
animals exhibit what is called the "dorsal light reaction" in response to the
sun's rays by maintaining their dorsal surface up. A
few animals, such as Artemia,
backswimmers (hemipterous insects), and the fish louse, Argulus (a
crustacean), reverse this response and exhibit a "ventral light reaction" and
keep the ventral surface pointed toward light. Consequently,
brine shrimp (and fairy shrimp) normally swim upside down (ventral side up),
because in nature the light is overhead. A
brine shrimp in a dish on the stage of a dissecting microscope withsubstage illumination,
however, may reverse its orientation and swims with the ventral side down.
a brine shrimp or fairy shrimp in a small culture dish and put it on the stage
of the dissecting microscope with the incident lamp on and the substage lamp
the swimming orientation of the animal. Turn
the incident lamp off and the substage (transmitted) lamp on. Do
this several times noting the response of the animal. <
laboratory culture of Artemia will
provide representatives of the major larval stages of a typical crustacean life
cycle. Artemia requires
about 14 molts to reach its terminal size and achieves sexual maturity in about
12 molts. The
stages between successive molts are instars. Artemia larval
stages are the nauplius, which includes a metanauplius, and the zoea. No
megalops is present. A
nauplius is a crustacean larva that swims with head appendages, whereas a zoea
is an older larva that swims with thoracic appendages. The
megalops, which is older still, swims with abdominal appendages.
In Artemia the
larvae hatch with few segments and gradually increase the number to 19 with
successive molts. The
mitotically active teloblast areas in the telson add new segment buds with each
molt until the characteristic number for the species is achieved. New
limb buds appear on existing segments with each molt. The 19 trunk segments of Artemia are
added in groups, rather than one with each molt.
a fine pipet to select several larvae of each of as many different sizes as
possible from the laboratory culture. Transfer
them to a small (6-cm) culture dish of chloroform-saturated saltwater or add a
few drops of chloroform to the culture dish and wait for the larvae to become
the larvae stop moving, transfer some of the smallest to a glass slide and make
a wetmount with them. Try
to arrange the larvae so some have the venter facing up and others the dorsum. Support
the coverslip with wax feet and press it gently against the larvae but do not
crush or distort them. Refer to the Techniques chapter for instructions on
making wax feet.
the compound microscope find one of the smallest larvae, which should be a nauplius (Fig
7, 19-8). Artemia hatches
as a nauplius with a short body consisting of the first three head segments and
a short trunk but with no external segmentation.
Figure 7. An Artemia nauplius
larva. A. Dorsal, B. Ventral. Anostraca7La.gif
yolk gives the body an orangish color and renders it nearly opaque. Due
to this opacity it is difficult to discern internal structures. (The
interior is much easier to see in an older metanauplius in which the yolk
reserves have been exhausted and the tissues are transparent.) The
nauplius is lecithotrophic, living off its yolk reserves without feeding.
the dark, red or black, median naupliar
eye at the anterior end of
the head. It
consists of three pigment cup ocelli. The
brain may be visible surrounding the eye in older, more transparent embryos. No
carapace is present in adult or larval anostracans.
anteriormost appendages are the small uniramous first
antennae. The second
antennae are the largest
appendages of the nauplius and are the principal swimming organs (Fig 7). They
are equipped with long natatory swimming
setae and are biramous. The
chief swimming setae are on the exopod.
The endopod is
smaller and lacks the row of evenly spaced swimming setae.
third pair of appendages is the uniramous mandibles.
The labrum is
a large, thin, ventral fold of body wall arising between the second antennae and
the mandibles, immediately anterior to the mouth (Fig 7B). It
is not a segmental appendage.
posterior end of the body is the telson (Fig
telson contains the teloblast areas that produce the mesoderm from which new
segments are fashioned. Buds
produced at the anterior edge of the telson differentiate into new segments. The
youngest segments are closest to the telson and the oldest are anterior.
the dissecting microscope observe a few active Artemia larvae
in a small culture dish of salt solution or seawater. Watch
one swim and try to see which appendages it uses. Swimming
with a single pair of appendages is erratic and jerky. Compare this motion with
the smooth swimming of adults. Adult
swimming utilizes 11 pairs of appendages and is much smoother. <
to find a small transparent larva about the size of the opaque, orange nauplii
but with a longer trunk. Such
a larva will be a metanauplius. The
metanauplius is a slightly older larva but, since it swims with its head
appendages, is still considered to be a nauplius.
metanauplius stage includes several molts and instars and lasts for several
ends when the anterior thoracic limbs become functional in the tenth instar. The
metanauplius begins feeding when its yolk reserves are exhausted.
metanauplius has an unsegmented head to
which is attached an elongate thorax. The
anterior end of the thorax is visibly segmented. The
internal organs of these slightly older larvae are easier to see than are those
of the nauplius.
The gut extends
from the head posteriorly to the telson. The
mouth is on the ventral surface of the head, between the bases of the mandibles,
under the labrum but you probably cannot see it. The stomach is
wider than the rest of the gut and is situated dorsal to the mouth to which it
is connected by a vertical esophagus. Two
diverticula, the digestive
ceca, extend anterolaterally from the stomach.
The intestine extends
posteriorly from the stomach. In
the nauplius it is relatively short but it is long, narrow, and getting longer
in the metanauplius. The
posterior end of the gut is hindgut, or rectum, and
it differs from the midgut in appearance. Its
walls are nearly colorless whereas those of the midgut are reddish brown with
the remaining yolk. The
rectum opens to the exterior via the anus at
the end of the telson.
lateral eyes appear in the metanauplius or zoea. They
require several molts to develop their stalks.
zoea stage, consisting of many instars, follows the metanauplius. The
zoeal sequence begins with a long transition period (known as the protozoea) in
which both the head and thoracic appendages are involved in swimming.
of its swimming setae and cessation of swimming movements by the second antennae
signals the beginning of the true zoeal stage. (In Artemia this
period is sometimes referred to as the postlarva. The
larva resembles the adult but is smaller.) During
this period the limb primordia continue developing into functional limbs and the
animal grows to its adult size. The
animal becomes an adult when all appendages are present, complete, and
functional but molting continues every few days through an adult life of several
protozoea or zoea are available, place a dish with a few larvae on the stage of
a dissecting microscope and watch them swim. Compare
the swimming with that of the nauplius. The
swimming motion becomes progressively smoother as more and more pairs of
phyllopods replace the single pair of antennae. <
DP . 1987. Observing
Marine Invertebrates. Stanford
Univ. Press, Stanford. 380p.
Anderson DT. 1967. Larval
development and segment formation in the branchiopod crustaceans Limnadia
stanleyana King (Conchostraca)
and Artemia salina (L.)
J. Zool. 15:47-91.
RM . 1937. A
method for rearing Artemia salina,
in Needham, J. G. et al.,
Culture Methods for Invertebrate Animals. Comstock,
(ed) 1950. Selected
Invertebrate Types. Wiley,
New York. 597p.
RA. 1993. Sex
and the single brine shrimp. Natural
WS . 1958. Practical
Invertebrate Anatomy (2nd ed). MacMillan,
J. 1981. Crustaceans,
in Dales, R. P. (ed) Practical
Invertebrate Zoology. MacMillan,
H. 1924. The
external development of certain phyllopods. Jour.
to the morphology and the taxonomy of the Branchiopoda Anostraca. Zool.
Bidrag. Uppsala 20:101-302.
JW. 1992. Branchiopoda,
pp 25-224 in
Harrison, F. W. & A.
G. Humes (eds.). 1992. Microscopic
Anatomy of Invertebrates vol. 9 Crustacea . Wiley-Liss,
WG. 1957. Studies
on the laboratory culture of Anostraca. Trans.
Am. Micros. Soc. 76:159-173.
Pardi L, Papi F. 1961. Kinetic
and tactic responses, in T.
H. Waterman, The Physiology of Crustacea II. Academic
Press, New York. p
RW . 1989. Fresh-water
Invertebrates of the United States, 3 ed. Wiley,
New York. 628p.
Personne G. et
al. (eds.). 1980. The
Brine Shrimp , Artemia, vols
Press, Wetteren, Belgium.
Ruppert EE, Fox RS,
Barnes RB. 2004.
Invertebrate Zoology, A functional evolutionary approach, 7 th ed.
Brooks Cole Thomson, Belmont CA. 963 pp.
LJ. 1969. Studies
on the larval structure and metamorphosis of Balanus
balanoides (L.). Philos.
Trans. Roy. Soc. London, Ser B 256:237-280.
D. I. 1982. Larval
morphology and diversity, pp. 43-110 in D. E. Bliss (ed), The Biology of
Crustacea vol 2. Academic
Press, New York.
8-cm culture dish or square watch glass
Yeast / Congo red suspension (See Supplies and Recipes
Slides and coverslips
plastic Pasteur pipets
iodine-free salt (e.g. ice cream salt)
5-10 ml 0.002M silver nitrate
Brine shrimp (Artemia) eggs or colony
is convenient to have a self-perpetuating colony in which nauplii, zoea,
postlarvae, and adults are always available. Alternatively,
the necessary life history stages can be produced by adding eggs to saltwater 48
hours, 96 hours, 1 week and three weeks prior to the day of the laboratory in
which they are needed.
Brine Shrimp Cultures
shrimp are easily reared in a salt solution (brine) of almost any salinity. A
suitable concentration can be achieved using 40 g ofnon-iodized salt (e.g.
ice cream salt or non-iodized
table salt) per liter of chlorine-free fresh water. Use pond, spring, distilled,
or aged tapwater. The brine should be in a shallow, non-metallic (i.e. glass)
pan or dish with a large exposed surface area and shallow depth (about 2-3 cm). Do
a lab marker, mark the original level of the brine on the side of the container
and add distilled water as needed replace evaporative losses and maintain this
level. Do not replace evaporative losses with more brine. Do not cover the
container as this restricts the oxygen supply.
have adults and all larvae stages on the day of the laboratory exercise, begin
about 3-4 weeks prior to the day for which the animals are needed. Add
a tiny pinch
of Artemia eggs
to the brine in the shallow dish. Add an additional tiny pinch of eggs every 4
days until about a week before the animals are needed and then add a tiny pinch
of eggs daily. Eggs
will begin hatching about 24-48 hours after being placed in the brine.
second day after the first eggs hatch, a feeding regimen should be instituted
and maintained thereafter for the life of the culture. Prepare
a suspension of one package baker’s yeast in 100 ml of fresh water and store it
covered, in a refrigerator (See Supplies and Recipes chapter). Each
day, use a pipet to add enough of the suspension to render the water in the
cultureslightly cloudy. Do
not overfeed, do not add too many eggs, and do not aerate. Keep
the yeast suspension refrigerated. Do
not add more yeast than the shrimp can remove before the next feeding. The
water should never become opaque rather should be transparent or nearly so.
population density must be low enough that sufficient oxygen can enter the water
by diffusion from the surface. Artemia larvae
are sensitive to increased carbon dioxide concentrations but under the
conditions described can be reared without artificial aeration. Aeration
damages the older, more fragile larvae and adults but does not harm nauplii and
can be reared to maturity by this method and, in fact, continuous cultures can
be maintained with the adults of the first generation producing eggs for
subsequent generations. Nauplii
will be available 24-48 hours after the eggs are added to the water. Other
larval stages will follow in turn. Adults
will appear in about three weeks and will be seen mating (swimming in tandem)
shortly after. Once
adults appear, it is no longer necessary to add eggs. Mature
females produce thin-walled eggs which hatch in the brood chamber and are
released as larvae to perpetuate the colony.
only nauplii or metanauplii are needed, feeding is not necessary and the culture
can be in a deeper container, such as a gallon jar, and aerated with an
these conditions the population density can be vastly increased. Start
such a high density culture (aerated) with 5 ml dry eggs per 4 liters of
saltwater 36-48 hours prior to the time the larvae are needed. A
culture with this density must be aerated vigorously. Aeration does not damage
the nauplii but will kill the more delicate older instars and adults.
eggs are available from biological supply houses, pet shops, and aquarium
Living Eubranchipus are
available Apr 1 to May 15 from Nebraska Scientific, 3832 Leavenworth St., Omaha,
Preserved Eubranchipus are
available from Carolina Biological and Ward's Natural Science.
wholemount slides of small fairy shrimps are available from Carolina Biological
slides of Artemia nauplii
are available from Triarch.
markets, under the name "Living Fossil" culture, vials of soil from the bottom
of temporary ponds that contain eggs of fairy shrimp, tadpole shrimp, and
can be reared from this soil.