larval cultures

Marine Biology - San Diego City College

Larval Cultures Adult animals supplied by:

Notes
Observations & descriptions of developmental stages

Development stage

Adult sea urchins (Strongylocentrotus purpuratus) and sand dollars (Dendraster excentricus) were spawned on Monday March 6. Newly fertilized egg of S. purpuratus. Approximately 30 seconds after insemination, a fertilizaiton envelope forms around the egg, starting at the point of sperm entry. This is the slow block to polyspermy.
S. purpuratus (urchin)

March 8 (d.2): 48 hrs. old. Both cultures look healthy. Temp. is controlled at 13 deg. C by water jacket in aquarium. First time culturing with artificial seawater (35 o/oo). Mesenchyme blastula stage. Hyaline layer is visible, which the embryo will soon hatch out of. It will begin to spin via ciliary beating and then break free of the membrane. The mesenchymal cells visible within the blastocoel have begun to cluster and will soon lay down the matrix of the calcium carbonate spicules. These spicules are analogous to those we saw in the Porifera.
S. purpuratus (urchin)

March 9 (d.3) : 3 days old. Should be ready to feed in one more day. Prism stage. Spicules (Sp)have begun to form. Mesechymal cells are clearly visible along the growing spicules. Mouth is not complete. The larva can now swim quite fast and would be swimming from right to left.
S. purpuratus (urchin)

March 10: (d.4)
Dendraster culture looks great (4-arm pluteus stage). S. pupuratus culture not forming larval arms as expected. Both cultures are using the same water, however, D. excentricus is mixed with aeration and S. purpuratus is being stirred mechanically. Feeding has begun with phytoplankton (Dunaliella and Isochrysis). Pluteus stage larva. Spicules are clearly elongating and the first pair of larval arms are starting to extend. The digestive tract is now complete: mouth (M), esophagus (Es), stomach (St), intestine (I) and anus (A).
S. purpuratus (urchin)

March 22 (d.16) - Some S. purpuratus have survived and look normal. The majority of this culture had been abnormal but these have died off. S. purpuratus.
Larvae at 8-arm stage. All larval arms are formed. Soon rudiment formation will begin if these cultures continue to develop.

March 22 (d.16) S. puupuratus.
Close up of mouth and bands of cilia drawing the feeding current.

March 26 (d.20) - Numerous D. excentricus seen with rudiment present. Rudiment formation is the beginnings of the juvenile forming within the larva. Tube feet (Tf) are clearly present

March 26 (d.20) - Several S. purpuratus showing signs of rudiment formation. Most of them are on the bottom of culture. Beginnings of Rudiment (R) formation. Pentamerous symmetry is clearly visible here.

March 28 (d.22) - Several S. purpuratus near metamorphosis. No more swimming larvae. They are no longer buoyant. Time to settle! S. purpuratus. Rudiment (R) formation is nearing completion with the juvenile forming within the larva. Tube feet (Tf) are clearly present. Larval arms are still present but will soon be dropped off or resorbed

April 7 (d.31) - Metamorphosis!!
Approxiamately 50% of each culture has undergone metamorphosis. Remainder of survivors are inbetween (still have larval arms). D. excentricus. Metamorphosis - The individuals that have undergone metamorphosis are now juveniles. The larval body has been discarded (possibly resorbed) and now the echinoid shows its pentamerous symmetry (as opposed to the bilateral symmetry observed in the larvae).

April 7 (d.31)
Juveniles are now 'settled' (no longer planktonic) and are grazing on the sheets of algae on the bottom of the culture. S. purpuratus.
Tube feet (Tf) and spines (S) are clearly present. Test size is approx. 1 mm. Lots of growth when compared to the 80 um egg.

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Page by Jay Vavra

Notes	Observations & descriptions of developmental stages	Development stage
Adult sea urchins (Strongylocentrotus purpuratus) and sand dollars (Dendraster excentricus) were spawned on Monday March 6.	Newly fertilized egg of S. purpuratus. Approximately 30 seconds after insemination, a fertilizaiton envelope forms around the egg, starting at the point of sperm entry. This is the slow block to polyspermy.	S. purpuratus (urchin)
March 8 (d.2): 48 hrs. old. Both cultures look healthy. Temp. is controlled at 13 deg. C by water jacket in aquarium. First time culturing with artificial seawater (35 o/oo).	Mesenchyme blastula stage. Hyaline layer is visible, which the embryo will soon hatch out of. It will begin to spin via ciliary beating and then break free of the membrane. The mesenchymal cells visible within the blastocoel have begun to cluster and will soon lay down the matrix of the calcium carbonate spicules. These spicules are analogous to those we saw in the Porifera.	S. purpuratus (urchin)
March 9 (d.3) : 3 days old. Should be ready to feed in one more day.	Prism stage. Spicules (Sp)have begun to form. Mesechymal cells are clearly visible along the growing spicules. Mouth is not complete. The larva can now swim quite fast and would be swimming from right to left.	S. purpuratus (urchin)
March 10: (d.4) Dendraster culture looks great (4-arm pluteus stage). S. pupuratus culture not forming larval arms as expected. Both cultures are using the same water, however, D. excentricus is mixed with aeration and S. purpuratus is being stirred mechanically. Feeding has begun with phytoplankton (Dunaliella and Isochrysis).	Pluteus stage larva. Spicules are clearly elongating and the first pair of larval arms are starting to extend. The digestive tract is now complete: mouth (M), esophagus (Es), stomach (St), intestine (I) and anus (A).	S. purpuratus (urchin)
March 22 (d.16) - Some S. purpuratus have survived and look normal. The majority of this culture had been abnormal but these have died off.	S. purpuratus. Larvae at 8-arm stage. All larval arms are formed. Soon rudiment formation will begin if these cultures continue to develop.
March 22 (d.16)	S. puupuratus. Close up of mouth and bands of cilia drawing the feeding current.
March 26 (d.20) - Numerous D. excentricus seen with rudiment present.	Rudiment formation is the beginnings of the juvenile forming within the larva. Tube feet (Tf) are clearly present
March 26 (d.20) - Several S. purpuratus showing signs of rudiment formation. Most of them are on the bottom of culture.	Beginnings of Rudiment (R) formation. Pentamerous symmetry is clearly visible here.
March 28 (d.22) - Several S. purpuratus near metamorphosis. No more swimming larvae. They are no longer buoyant. Time to settle!	S. purpuratus. Rudiment (R) formation is nearing completion with the juvenile forming within the larva. Tube feet (Tf) are clearly present. Larval arms are still present but will soon be dropped off or resorbed
April 7 (d.31) - Metamorphosis!! Approxiamately 50% of each culture has undergone metamorphosis. Remainder of survivors are inbetween (still have larval arms).	D. excentricus. Metamorphosis - The individuals that have undergone metamorphosis are now juveniles. The larval body has been discarded (possibly resorbed) and now the echinoid shows its pentamerous symmetry (as opposed to the bilateral symmetry observed in the larvae).
April 7 (d.31) Juveniles are now 'settled' (no longer planktonic) and are grazing on the sheets of algae on the bottom of the culture.	S. purpuratus. Tube feet (Tf) and spines (S) are clearly present. Test size is approx. 1 mm. Lots of growth when compared to the 80 um egg.