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LIFE HISTORY, MORPHOLOGY, AND SALINITY TOLERANCE OF THE AMPHARETID POLYCHAETE AMPHICTEIS FLORIDUS, HARTMAN 1951
ROBERT ANTHONY ZOTTOLI
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Recommended Citation ZOTTOLI, ROBERT ANTHONY, "LIFE HISTORY, MORPHOLOGY, AND SALINITY TOLERANCE OF THE AMPHARETID POLYCHAETE AMPHICTEIS FLORIDUS, HARTMAN 1951" (1966). Doctoral Dissertations. 841. https://scholars.unh.edu/dissertation/841
This Dissertation is brought to you for free and open access by the Student Scholarship at University of New Hampshire Scholars' Repository. It has been accepted for inclusion in Doctoral Dissertations by an authorized administrator of University of New Hampshire Scholars' Repository. For more information, please contact [email protected]. This dissertation has been microfilmed exactly as received 67-170 ZOTTOLI, Robert Anthony, 1939- LIFE HISTORY, MORPHOLOGY, AND SALINITY TOLERANCE OF THE AMPHARETID POLYCHAETE AMPHICTEIS FLORIDUS HARTMAN 1951. University of New Hampshire, Ph.D., 1966 Zoology
University Microfilms, Inc., Ann Arbor, Michigan LIFE HISTORY, MORPHOLOGY, AND
SALINITY TOLERANCE OF THE AMPHARETID POLYCHAETE
AMPHICTEIS FLORIDUS HARTMAN 1951
BY
ROBERT ANTHONY ZOTTOLI
B.A., Bowdoin College, 1960
M.S., University of New Hampshire, 1963
A THESIS
Submitted to the University of New Hampshire
In partial fulfillment of
The requirements for the Degree of
Doctor of Philosophy
Graduate School
Department of Zoology
June, 1966 This thesis has been examined and approved.
April 15, 1966
Date ACKNOWLEDGEMENTS
I wish to express my appreciation and gratitude to
Dr. Emery F. Swan for his guidance in the preparation of the dissertation and to the other members of the doctoral committee, Dr. George M. Moore, Dr. Arthur C. Borror, Dr.
Albion R. Hodgdon, and Dr. Arthur E. Teeri, for giving generously of their time. Dr. Arthur C. Borror deserves additional thanks for help in editing the dissertation.
Dr. Marian H. Pettibone formerly associated with the
University of New Hampshire and now Associate Curator,
Division of Worms, at the Smithsonian Institution, and Dr.
Olga Hartman of the Allan Hancock Foundation deserve special thanks for their interest and advice.
Summer fellowships received from the University of New
Hampshire in 1963 and from the George F. Dwinnel Memorial
Fellowship Fund of the New Hampshire Cancer Society, Inc., in 1964 enabled me to devote full time to this research throughout those two summers.
Finally I wish to express my appreciation to my wife,
Margaret, without whom this work would not have been possible. TABLE OF CONTENTS
LIST OF FIGURES ...... iii
LIST OF T A B L E S ...... viii
I. INTRODUCTION...... 1
II. THE FAMILY AMPHARETIDAE AND ITS GENERA...... 5
III. MATERIALS AND METHODS ...... 24
IV. GENERAL MORPHOLOGY...... 2 8
V. THE CIRCULATORY SYSTEM ...... 40
VI. THE DIGESTIVE SYSTEM ...... 72
VII. REPRODUCTION AND LARVAL DEVELOPMENT ...... 110
VIII. SALINITY TOLERANCE...... 146
IX. GENERAL DISCUSSION...... 148
LITERATURE C I T E D ...... 151
APPENDIX Formulary of Procedures and Stains . . .160
ii LIST OF FIGURES*
Number Page
1. Trilobed prostomium ...... 16
2 . Dorsal surface showing the arrangement of branchiae...... 16
3. Near-sagittal section of the anterior end with tentacles everted ...... 18
4. Near-sagittal section of the anterior end with tentacles retracted ...... 20
5. Limbate capillary seta of Amphicteis gunneri. 22
6 . Paleal seta of Amphicteis g u n n e r i ...... 22
7. Smooth capillary seta of Neosabellides o c e a n i c a ...... 22
8 . Dorsal hook of Melinna palmata ...... 22
9. Thoracic uncinus of Ampharete grubei. Lateral v i e w ...... 22
10. Thoracic uncinus of Amphicteis procera. Dorsal v i e w ...... 22
11. Abdominal uncinus of Amphicteis procera. Dorsal view ...... 22
12. Dorsal view of the anterior portion of Melinna cristata ...... 23
13. Dorsal hook of Melinna cristata ...... 23
* Unless indicated otherwise in the legend, the dorsal surface of line drawings will be toward the right and the anterior portion toward the top. Except where indicated otherwise all figures are of Amphicteis floridus iii 14. Uncinus of the same s p e c i e s ...... 23
15. Lateral view in which branchiae of the left side have been r e m o v e d ...... 36
16. Uncini of the right twelfth thoracic setiger. 37
17. Uncini of the right sixth abdominal segment . 37
18. Anterior right nephridium dissected out . . . 38
19. Right notopodium of a 18— setiger larval stage showing the muscles which operate the setal s a c ...... 38
20. Cross section at the junction between the fourth and fifth segments ...... 39
21. Diagrammatic dorsal view of the dorsal vessel 58
22. Cross section of the ventral ciliated groove at the mid-portion of the stomach ...... 60
23. Lateral view through the transparent body wall showing blood vessels in the anterior thoracic region...... 62
24. Lateral view through the transparent body wall showing blood vessels in the posterior thoracic and anterior abdominal regions...... 64
25. Diagrammatic ventral view of the anterior portion of the blood sinus...... 65
26. Diagrammatic dorsal view of the oesophageal surface showing blood vessels arising from the annular vessel...... 67
27. Diagrammatic ventral view of the oesophagus showing blood vessels arising from the annular vessel...... 68
28. Diagrammatic dorsal view of the anterior portion of the ventral vessel ...... 70
iv 29. A single dorso-pedal vessel from the anterior thoracic region of Amphicteis gunneri ...... 71
30. Fused dorso-pedal vessels from the posterior thoracic region of Amphicteis gunneri .... 71
31. Dorso-pedal vessel of Melinna palmata .... 71
32. Cross section of a tentacle...... 96
33. Cell types of the upper l i p ...... 97
34. Cell type of the oe s o p h a g u s ...... 97
35. Cell types of the stomach ...... 97
36. Near-sagittal section of the buccal mass. . . . 98
37. Near-sagittal section of the gastric and lateral invaginations ...... 98
38. Near-sagittal section of the lower lip and buccal m a s s ...... 100
39. Cross section through the anterior thoracic r e g i o n ...... 100
40. Near-sagittal section of the junction between the stomach and oesophagus ...... 101
41. Near-sagittal section of the dorsal vessel and the dorsal portion of the body wall ...... 103
42. Cross section the dorsal mid-intestine . . . .105
43. Stomach, intestine, and rectum. The dorsal portion of the stomach is cut away to show lateral and gastric invaginations...... 107
44. Cross section of the mid-abdominal region . . .107
45. Cross section of the posterior portion of the stomach ...... 109
v 46. Unfertilized e g g ...... 133
47. Two-cell stage ...... 133
48. Coeloblastula. Dorsal view ...... 135
49. Early trochophore. Dorsal view ...... 135
50. Early trochophore. Ventral v i e w ...... 135
51. Late trochophore. Dorsal v i e w ...... 137
52. Late trochophore. Lateral view ...... 137
53. One-setiger stage. Dorsal v i e w ...... 139
54. Two-setiger stage. Dorsal view ...... 139
55. Three-setiger stage. Dorsal v i e w ...... 140
56. The first two notopods and accompanying setae of a three-setiger stage. Dorsal view . . . .140
57. Larval uncinus. Lateral view ...... 140
58. Larval uncinus. Dorsal view ...... 140
59. Four-setiger stage. Dorsal v i e w ...... 141
60. Five-setiger stage. Dorsal view ...... 141
61. Seven-setiger stage. Lateral view ...... 142
62. Anterior end of the above specimen showing the single ciliated tentacle .... .142
63. Eight-setiger stage. Dorsal view ...... 143
64. Ten-setiger stage. Dorsal v i e w ...... 143
65. Twelve-setiger stage compressed slightly. Ventral v i e w ...... 144
6 6 . Four teen-setiger stage. Ventral view .... .144
vi 67. Sixteen-setiger stage. Lateral view .... .145
6 8 . Anterior portion of the above specimen . . . .145
vii LIST OF TABLES
Number Page
1. The number of adult uncini per uncinigerous pinnule in one sexually mature female 7 mm long and 1 mm wide ...... 33
2. Distribution of nephridial cell types and measurements ...... 34
3. Distribution and measurements of cell types of the buccal c a v i t y ...... 92
4. Distribution and measurements of cell types of the oesophagus and stomach ...... 93
5. Distribution and measurements of cell types of the intestine and r e c t u m ...... 94
6 . The number of smooth capillary setae per notopodium and uncini per uncinigerous pinnule on setigerous and post-setigerous segments of the 9-18 setiger s t a g e s ...... 128
7. Measurements and numbers of larval stages (Un-r fertilized egg to the trochophore) found in n a t u r e ...... 129
8. Measurements and numbers of larval stages (Late trochophore to the three-setiger stage) found in n a t u r e ...... 130
9. Measurements and numbers of larval stages (Three- setiger to the nine-setiger stage)found in nature 131
10. Measurements and numbers of larval stages (Ten- setiger to the eighteen-setiger stage) found in n a t u r e ...... 132
11. Experimental salinity tolerance 147 1
SECTION I
INTRODUCTION
Proposed groupings of polychaete families generally have met with only brief acceptance, since existing families of polychaetes are very distinct (Dales, 1962)
1963). Benham (1894, 1896) divided the polychaetes into the branches Phanerocephala and Cryptocephala based on head structure. Hatschek (1893), Hempelmann (1931), and
Friedrich (1938) divided the polychaetes into the orders
Errantia and Sedentaria based on living habits. The order
Sedentaria includes all families Benham included in the
Cryptocephala plus several families of Benham's
Phanerocephala. Uschakov (1955) raised Hatschek's orders to sub-classes and divided each group into several orders on the basis of morphology. Dales (1962) grouped polychaete families on the basis of proboscis structure. This grouping appears to merit serious consideration. Gaining of additional information on the proboscis and other characteristics to support or reject Dales' (1962) ideas would seem desirable.
While exploring intertidal mud of several estuarine creeks in the Dover, Durham area of southern New 2
Hampshire, I noted a species of ampharetid polychaete.
Members of the family Ampharetidae are generally small, inconspicuous, and little studied. Hence attempts to identify this worm almost immediately revealed problems.
I decided to study the ecology, life history, and morphology of this animal to further clarify relationships among the polychaete families Ampharetidae, Terebellidae, and Pectinariidae (Amphictenidae).
These three families are members of the sub-order
Terebelliformia of Benham (1894, 1896), the sub-order
Terebellomorpha of Hatschek (1893), the order Terebelli- morpha of Uschakov (1955), and the group Terebellida of
Dales (1962). Benham (1894, 1896) also included the
Cirratulidae in his sub-order Terebelliformia.
Uschakov (1955) also included the Trichobranchiidae in his order Terebellimorpha.
An ampharetid polychaete named Amphicteis gunneri floridus by Hartman (1951) is recorded from Englewood and
Ocklockonee, Florida. A similar form named Hypaniola grayi by Pettibone (1953) is reported from a salt pond on
Martha's Vineyard, Massachusetts. The worms from the intertidal muds of the several estuarine 3 creeks* in the Dover, Durham area of southern New
Hampshire also appeared similar to both.
The animals collected in New Hampshire dwell in tubes about five times the length of the worm’s body. The upper third of this tube projects above the surface of the mud.
The tube consists of mud, sand, and occasional bits of animal and plant debris, cemented together by a mucus-like substance probably secreted by the ventral shields of the worm.
The worm feeds by extending its anterior end from the upper portion of the tube and spreading its tentacles over the mud surface (see section VI for details). Gut contents of 24 sexually mature worms (10 males and 14 females) included Foraminifera, diatoms, and green algae, as well as particles of sand and multicellular animal and plant material.
Other than the works of Wiren (1885) on Amphicteis gunneri and Melinna cristata, Meyer (1887) on Melinna
* Approximate N. Latitude and W. Longitude W N Bunker's Creek 70° 53’ 43° 08* Crommet Creek 70° 53* 43° 06' Fresh Creek 70° 50' 43° 11' Johnson's Creek 70° 54' 43° 08' 4
palmata, and Fauvel (1897) on Ampharete grubei. Amphicteis gunneri, Melinna cristata, and Samytha adspersa, studies on the circulatory systems of ampharetids are relatively few.
The anatomy of digestive systems of ampharetids have been studied by Wiren (1885) in Amphicteis gunneri and
Melinna cristata, by Fauvel (1897) in Ampharete grubei,
Samytha adspersa, and Melinna palmata; and by Djakonov
(1913) in Amphicteis gunneri. Newell and Baxter (1936) described cell types in portions of the digestive tract of
Melinna palmata. Dragoli (1961) investigated the feeding habits of the latter species.
Okuda's (1947) study of Schistocomus sovjeticus and
Nyholm’s (1950) study of Melinna cristata are the only published approaches to complete life histories of members of the family Ampharetidae. Fauvel (1897) illustrated one non-pelagic larval stage of Ampharete grubei. Ostrooumouv (1899) briefly described several larval stages of Hypania invalida. Wesenberg-Lund (1934) recorded the presence of larval stages of the viviparous Alkmaria romijni, and Thorson (1946) illustrated larvae of the same species. 5
SECTION II
THE FAMILY AMPHARETIDAE AND ITS GENERA
Taxonomy
The genera Amphicteis with three species and
Sabellides with five species, previously considered members of the Terebellidae, were transferred to the newly created Ampharetidae (Malmgren, 1866) because they possess
tentacles that are retractable into the buccal cavity.
Malmgren (1866) and Meyer (1912) classified genera
in the family on the basis of the number of thoracic
setigerous segments, presence or absence of paleae, condition of the tentacles (smooth or ciliated), and the
number of branchiae. Hessle (1917) criticised the
overemphasis on tnese characters, and attempted to construct
a more precise key using such characters„as presence or
absence of ventral acicular setae on the anterior segments,
condition of the branchiae, presence or absence of glandular
bands on the prostoraium, position of the notopodia, and
characteristics of the nephridia and nephridiopores.
Chamberlin (1919) divided the family into three sub-families,
the Ampharetinae, Samythinae, and Melinninae,on the
basis of presence or absence of paleae and dorsal hooks.
Presence or absence of paleae is a questionable character because of the variation in setae of the branchiferous segments III to VI and the difficulty of distinguishing between small capillary setae and paleae of segment four. Likewise arrangement of nephridia and other internal structures such as a gastric invagination and anterior lobes are variable within genera, of questionable significance, and difficult to observe because of the time-consuming dissection required (Day, 1964).
Day (1964) separated genera in the family Ampharetidae on the basis of the number of pairs of branchiae, and the number of uncinigerous thoracic segments. Day's (1964) tentative key, in which 16 of the approximate 49 genera are reduced to synonomy, is essential for checking diagnostic characteristics and similarities among genera.
Though Day's (1964) work is valuable, a thorough study of the generic characteristics within the family is still needed.
Description
Members of the family Ampharetidae are small
(usually 5-80 mm in length), with the body divided into thorax and abdomen. The thorax is widest anteriorly and characteristically bears notopodia possessing capillary setae and neuropodia (uncinigerous pinnules) bearing 7
pectiniform uncini. The abdomen generally lacks notopodia possessing capillary setae but has neuropodia bearing pectiniform uncini. However capillary setae have been reported to occur to the caudal end of the body in the genus
Otanes (Kinberg, 1867).
The surfaces are relatively smooth, except for thick
glandular shields (ventral shields), generally one per
segment, on the ventral surface of the thorax.
Segment number ranges generally from 20 to 40 with
as many as 70 in the genus Melinna.
The prostomium is generally trilobed (Fig. 1, p. 16)
with eyespots visible through its more or less transparent
epithelium. Generally only one pair of eyespots is present,
but in some species of Amphicteis numerous eyespots have
been reported (Fauvel, 1927).
Tentacles lie between the cephalic lobe and upper lip
in the retractile anterior portion of the buccal cavity.
When it is extended, the tentacles are everted (Fig. 3,
p. 18). When it is folded inward, they lie posteriorly
directed in the oesophageal cavity (Fig. 4, p. 20). In
the genera Auchenoplax, Glyphanostomum, Phyllocomus,and
Schistocomus, tentacles are reported as being absent.
However, they may have been overlooked. The filiform
tentacles may be ciliated (Sabellides), smooth (Amage), or may possess ciliated pinnules (Ampharete).
Branchiae occur in two to four pairs, are either filiform or subulate (wide at the base and tapering towards the tip), ciliated, and may be either smooth or pinnately branched (Fig. 2, p.16). In a specimen with four pairs of branchiae, they occur on the dorsal surface of the paleal or corresponding segment and on the first, second, and third thoracic setigerous segments (Fig. 2, p. 16).
Generally setae are, paleae, dorsal spines or hooks, smooth capillary setae, limbate capillary setae, and pectiniform uncini (Figs. 5-11, p. 22).
Paleae are the setae of the segment immediately following the buccal segment and, when present, are borne generally on small ridges which represent rudimentary notopodia (Fig. 2, p. 16).
In members of the sub-family Melinnae. there is a pair of dorsal hooks or spines which in Melinna cristata are located a short distance behind the posterior pair of branchiae (Figs. 12, 13, p. 23). Also in this group there are generally rows of anterior thoracic, fine, ventral, smooth capillary setae (Ac.S.) which in Melinna cristata are present on segments 2, 3, 5, and 6 (Fig. 12, p. 23).
Ampharete grubei possesses uncini with 9-10 teeth in two parallel rows. Sabellides octocirrata has uncini with 9
3-4 teeth in a single vertical row in the thoracic region and uncini with 8 teeth arranged in 2 or 3 vertical rows in the abdominal region. Other species, including Amphicteis floridus, have uncini with teeth in a single vertical row
(Figs. 16, 17, p. 37).
Cylindrical notopodia, often with a small distal cirrus, are present on most of the post-paleal thoracic segments, and are rarely represented on the abdomen. Several species, including A^_ f loridus, possess rudimentary notopodia on the first few abdominal segments.
Neuropodia (uncinigerous pinnules), sometimes with cirri, occur on each segment from the 4th (Moyanus) or the
6 th or 7th segment posteriorly. Malmgren (1886) and Fauvel
(1927) recognized two segments in front of the paleal, making it the third while Nilsson (1912) and Hessle (1917) recognized one bi-annular buccal segment in front of the paleal, making it the second. The latter numbering system will be used throughout this work.
Two or more cirri sometimes occur on the pygidium.
The gastric invagination is formed by the inpocketing of the antero-ventral stomach floor (Figs. 37, 43, pp. 98
107 ). Externally directed lateral lobes may occur anteriorly on the stomach (See section on the digestive system). 10
Two to five pairs of nephridia have been reported in the anterior thoracic region of all ampharetids thoroughly investigated. The first pair in most cases occurs posterior to a diaphragm located either between the third and fourth or the fourth and fifth segments. The diaphragm divides the thoracic cavity into an anterior and a posterior region.
Each nephridium opens to the outside through nephridiopores generally located on small papillae located at the base of a notopodium or neuropodium.
Ampharetids live in tubes covered by mud, sand, and animal and plant debris. Most ampharetids live in deep water. Hypania invalida and Hypaniola kowalewskii, however, occur in shallow areas of the Caspian Sea.
Amphicteis floridus occurs in intertidal mud in several regions on the Eastern Coast of North America. Ampharete grubei occurs in the Tamar Estuary in England with a population density of 4033 individuals per square meter
(Spooner and Moore, 1940).
Taxonomy of A. floridus
In 1953 Pettibone described Hypaniola grayi from
Martha's Vineyard. It is conspecific with the worm here studied (Pettibone, in litt.) and possibly identical with
Amphicteis gunneri floridus Hartman, 1951. Thus several questions may be raised. 11
1. Are Hypaniola grayi and Amphicteis gunneri
floridus the same?
2. To what genus should this species be assigned?
3. Is this worm specifically different from
AmphictoiB gunneri?
Paratypes of Amphicteis gunneri floridus and of
Hypaniola grayi, and several specimens from the populations used in this study all appear to me to be externally identical. Internally each possesses a gastric invagina tion.
The genus Hypaniola was erected by Annekova (1927) to separate Amphicteis kowalewskii Grimm 1877 from other species of Amphicteis. As originally defined it included those ampharetids with trilobed tentacular membrane without glandular crests or ridges; one pair of eyespots; paleae shorter than the capillary setae; three or four pairs of branchiae, the first three of each side fused at their bases; 17 thoracic notopodia with setae beginning on the third segment; notopodia without cirri; rudimentary notopodia on the anterior abdominal segments; the tooth bearing portions of the uncinate setae not clearly attached to the basal part; thoracic uncini with a single vertical row of teeth; abdominal uncini with three vertical rows of teeth; no anal papillae or cirri; 12
stomach without side lobes or a gastric invagination; nephridia (3 pairs) with the long elbows, those of the
fifth segment being the longest; the outer openings of the nephridia not shifted dorsally; ventral giant cells
(large cells which lie between the ventral thoracic shields
and the ventral nerve cord) lacking.
Annekova (1927) considered the form of the tentacular
membrane (prostomium), the condition of the paleae and the
number of thoracic setigerous segments as the distinguishing generic characteristics.
The genus Hypaniola was emended by Pettibone (1953)
to include H_^ grayi. The concept of the genus was emended
in the following manner: glandular prostomium crests
variable depending on the amount of folding; oral
tentacles retractile and smooth, dorsal hooks posterior to
the branchiae (such as are present in the genus Melinna)
lacking; thoracic uncinigerous pinnules beginning on
segment 6 (thoracic setiger 4); thoracic uncini with 1 or
3 vertical rows of teeth; abdominal uncinigerous pinnules
may have cirri; three pairs of nephridia located
respectively in segments 4-6 (setigerous segments 2-4).
Pettibone (1953) did not mention the gastric
invagination. Presumably no deviation from the original
description of the genus in this regard was observed and thus a gastric invagination would appear to be lacking.
Day (1964) placed the genera Hypaniola and
Amphisamytha in synonomy with Lysippides. Day (1964) characterised Lysippides as follows:
'•Prostomium without glandular ridges. Buccal tentacles smooth with a groove along one side. Four pairs of gills. Segments III-VI without neurosetae. Notosetae present on segments V and VI and usually III and IV as well. Fourteen uncigerous thoracic segments. Notopodial cirri absent. Thoracic uncini with 1-3 series of teeth. Between 15 and 20 abdominal segments sometimes with rudimentary notopodia as well as uncigerous pinnules."
Day (1964) has retained the genus Amphicteis and characterizes it as follows:
"Amphicteis Grube 1851 synonyms. Crossostoma Gosse 1855,? Rytocephalus Quatrefages 1865, and Paramphicteis Caullery 1944.
Prostomium with a pair of glandular ridges. Buccal tentacles smooth with a groove along one side. Four pairs of gills. Segments III-VI without neurosetae. Segment III often with paleae; segments IV-VI with notopodal capillaries. Fourteen uncigerous thoracic segments. Notopodial cirri present. Thoracic uncini with a single vertical series of teeth. Thirteen to 19 abdominal segments sometimes with rudimentary notopodia as well as uncigerous pinnules.
Type species Amphitrite gunneri Sars 1835."
Specimens investigated here differ from the genus
Hypaniola emended by Pettibone (T953) in possession of notopodial cirri, possession of only two pairs of nephridia located in segments 6 and 7 and possession of 14 a gastric invagination. A_^ floridus differs from the genus Lysippides as characterized by Day (1964) by the possession of glandular ridges on the prostomium, and by the possession of notopodial cirri. Internal characteristics were not used by Day (1964).
Specimens investigated in this study agree with the description of the genus Amphicteis as characterized by Day (1964). They differ from Amphicteis gunneri in possession of two pairs of nephridia; (four to five pairs are present in A. gunneri); by the lack of anal cirri; (one pair is present on A. gunneri); by the possession of a greater number of abdominal segments with uncinigerous pinnules (There are 22-28 in the species here studied and 15 in A^_ gunneri) ; possibly by the nature of the eyespots (the majority of the specimens used in connection with this study possess one pair while A_^ gunneri is described as having paired patches of eyespots).
These differences justify designating the present species as A^_ floridus Hartman, 1951.
The synonomy of the animal here studied is as follows: Amphicteis gunneri floridus Hartman, 1951;
Hypaniola grayi Pettibone, 1953; Amphicteis floridus
Hartman, 1951. 15
Ba. B . Basal Part —1 i A 03 n • i Branchiae 1-4
Buc. S . Buccal Segment
By. S. Eyespot
Lat. L. Lateral Lobe
Med. L. Median Lobe
Med. P. Median Part
Pa. Palea
Pa. S. Paleal Segment
Pro. Prostomium 16
Med. L
,M ed, P.
La 1.1
Fig. 1. Trilobed prostoraium. X10
Pro.
Pa.S.
Br. 1. Br.2,
Br.4.
Br.3
Fig. 2. Dorsal surface showing the arrangement of branchiae. X25 Fig. 3. Near-sagittal section of the anterior end with tentacles everted, dorsal surface toward the right. X100
B. M. Buccal Mass
Br • Branchia (1st pair)
B. S. W. Blood Sinus Wall
C. G. Cerebral Ganglion
C. L. Cephalic Lobe
D. V. Dorsal Vessel
I . L. Inner Lip
L. L. Lower Lip
Oe • Oesophagus
S. E. Stomach Epithelium
T. Tentacle
U. L. Upper Lip
V. N. C. Ventral Nerve Cord 18
U.L
C.L
C.G.
.B.M
V.N
D.V.
Oe
S.E,
B.S.W.
,______0.4 mm
Fig. 3. 19
Fig. 4. Near-sagittal section of the anterior end with tentacles retracted, dorsal surface toward the right. X100
Br. Branchia (2nd pair)
B. M. Buccal Mass
C. G. Cerebral Ganglion
C. L. Cephalic Lobe
Cil. Cilium
D. V. Dorsal Vessel
H. B. Heart Body
I. L. Inner Lip
L. L. Lower Lip
Oe. Oesophagus
T. Tentacle
T. M. Tentacular Membrane
U. L. Upper Lip 20
C.G,
L.L. UL
T.M
.B.M
O e
D.V.
0.4 mm
Fig. 4. 21
Fig. 5. Limbate capillary seta of Amphicteis gunneri (after Fauvel, 1927).
Fig. 6 . Paleal seta of Amphicteis gunneri (after Fauvel, 1927).
Fig. 7. Smooth capillary seta of Neosabellides oceanica (after Fauvel, 1927).
Fig. 8 . Dorsal hook of Melinna palmata (after Fauvel, 1927).
Fig. 9. Thoracic uncinus of Ampharete grubei. Lateral view (after Fauvel, 1927).
Fig. 10. Thoracic uncinus of Amphicteis procera . Dorsal view (after Ehlers, 1887).
Fig. 11. Abdominal uncinus of Amphicteis procera . Dorsal view (after Ehlers, 1887). 22 23
Fig. 12. Dorsal view of the anterior portion of Melinna cristata (after Fauvel, 1927, modified).
Ac. S. Acicular Setae
D. H. Dorsal Hook
Fig. 13 Fig. 14
Fig. 13. Dorsal hook of Melinna cristata (after Fauvel, 1927).
Fig. 14. Uncinus of the same species (after Malmgren, 1866). 24
SECTION III
MATERIALS AND METHODS
Living worms were examined for external and internal characters with a Bausch and. Comb Stereozoom (Model AVB-73)
dissecting microscope.
The gross anatomy of the digestive system was
observed in dissection of living (50) and preserved (30)
specimens, and further details were elucidated with study of serial sections. Twenty specimens were prepared for
sectioning in the following manner: 1. Relaxation in a
mixture of one part Chloretone saturated in distilled water
and two parts sea water having a salinity of 32°/00
2. Fixation in Bouin's Duboscq for 24 hours (Appendix)
3. Dehydration with a graded series of alchohols
(Appendix) 4. Imbedding in "Paraplast", a prepared
tissue imbedding medium manufactured by Biological
Research Inc. (subsidiary of Brunswick Corp.)
5. Sectioning with an American Optical Co. "800"
rotary microtome at from 5-12 ju. 6 . Affixing the
sections to slides with a mixture of albumen and distilled
water (Appendix). 7. Staining with alcian blue, aldehyde
fuchsin counterstained with Halmi's mixture, azure-A, 25 diluted Foot's, haematoxylin-eosin, modified iron haematoxylin-orange G, periodic acid-schiff (sections treated with amylase), safranin 0, and Van Gieson's
(Appendix)
Line drawings were made with the aid of an American
Optical Co. microprojector and a series of Rapidograph pens
(0 0 ,1 ,2 ).
Measurements of cell types were made with the aid of an ocular micrometer, on specimens which were approximately
7 mm long and 0.9 ram wide. The range of variation in measurement is indicated.
Photomicrographs were made using an Exacta XV camera with a waist level view finder, and a Science and Mechanics
supersensitive Model 250 P light meter.
The structure and arrangement of blood vessels was
determined by observing live animals (70) as well as cross
and longitudinal sections of adults and larvae which were
prepared for sectioning in a manner similar to that used
for a study of the digestive system. Sections cut at from
7-12 ji were stained with aldehyde fuchsin counter stained
with Halmi's mixture, diluted Foot's, Gomori's trichrome,
haematoxylin-eosin, and Van Gieson's iron hematoxylin
(Appendix).
In experiments to determine how tentacles move, ten 26 live worms were dissected with a razor blade on the ventral surface so that the anterior coelomic cavity was opened.
The cut surfaces were pinned to the side so they could not rejoin.
Larvae were removed from the parent tube by opening
the tube lengthwise with a pair of iridectomy scissors.
Drawings were made from living specimens with the aid of a camera lucida. Larvae were prepared for sectioning in a similar manner to that used for a study of the digestive system. Whole mounts were prepared according to Herman's
(1964) method (Appendix).
The induction of spawning was attempted by subjecting sexually mature worms (3 males and 3 females for each
test) to waters of varying salinity (0.5, 10, 30, 40, 50,
70o/oo) for 30 minutes each and in a separate experiment
to waters (all of 30 o/oo salinity)at 0, 20, and 30°C.
In attempting artificial fertilization, eggs were
removed from ripe females by opening the body cavity.
These were then washed in fresh sea water (30 o/oo),
and a drop of concentrated sperm suspension was added to
a Syracuse watch glass containing about 20 eggs in 10 ml of sea water (30 o/oo).
Early cleavage stages were not plentiful. Late cleavage stages were taken from the tubes of female worms 2 7
and placed in sterile plastic, Petri dishes containing sea water (27 o/oo) and allowed to develop. A small amount of surface mud from the area in which the worms were collected was placed in each Petri dish to provide food for the developing larvae. The temperature was maintained at between' 20° and 30° C.
Salinity was measured in the field with a Gemware hydrometer (Gemware salinity testing set; G.M. Manufactur ing Co.) and in the laboratory by the Mohr method for titrating chloride. Sea water collected in Great Bay,
New Hampshire was allowed to evaporate to a salinity of
70 o/oo. This in turn was diluted with ion free distilled water to 0.5, 1, 2, 5, 10, 20, 30, 35, 40, 45, 50, 60,
70 o/oo. For testing their salinity tolerance ten worms were removed from their tubes and placed in each of
thirteen covered (9") finger bowls filled with 1" of sea
water of known salinity. They were examined periodically for eight days. Each experiment terminated when all the worms no longer could be observed pumping blood. Sudden exposure
to extreme salinities did not cause cessation of pumping of blood in any worms. The salinities had not changed
significantly at the termination of each experiment. 28
SECTION IV
GENERAL MORPHOLOGY
External Anatomy
The animal is fusiform, wide anteriorly, tapering gradually towards the posterior end. The dorsal body wall is transparent; the ventral surface is opaque because of thick ventral longitudinal muscle bundles. Distinct ventral annulations indicate segmental boundaries; the dorsal surface is indistinctly annulated. Sexually immature individuals have an orange-green tinge with white flecks on the surface while egg bearing females are green and ripe males white corresponding with the color of the gametes in the coelom. A thin cuticle imparts an irridescence to the animal. Through the transparent wall one can see many of the blood vessels.
The prostomium is trilobed (Fig. 1, p. 16). The median lobe has a basal and a median part and is elevated.
Laterally it bears the eyes and nuchal organs; the median part is shovel shaped and beans raised glandular crests on its lateral edges. Usually a pair of pigment spots occurs on the ventro-anterior edge of the median part.
Two lateral lobes surround the median lobe.
The buccal segment is bi-annular (two-ringed), is 29 complete dorsally, and projects forward on the ventral surface beneath the prostoroium (Fig. 15, p. 36). It can be discerned in larval development as early as the late trochophore stage; later it fuses with the peristomium (Fig. 51, p. 137).
Ventrally ciliated digitiform oral tentacles (8-15) are present and can be seen projecting anteriorly from the mouth when the anterior buccal cavity is everted.
The paleal or second segment possesses a pair of rudimentary notopodia, each with a fan shaped bundle of five to ten paleal setae (Fig. 15, p. 36). Each paleal seta is the same length as the notopodial seta, and tapers gradually to a capillary tip. The first pair of branchiae is borne on the dorsal surface of this segment (Fig. 15, p. 36).
Each of the first three thoracic setigers (segments
3-5) is narrow and bears a pair of notopodia with bundles of five-seven slender capillary setae and with a small distal cirrus (Fig. 15, p. 36). A study of developmental stages revealed that the next three pairs of branchiae occur on the third, fourth, and fifth segments. All branchiae are long and tapering (Fig. 15, p. 36).
Segments 6-19 (thoracic setigers 4-17) each bear a pair of notopodia with a distal posterior cirrus and a 3 0 bundle of five to seven slender setae which taper to slender capillary tips, a pair of uncinigerous pinnules
(neuropodia) each possessing twenty-two to twenty-seven pectiniform uncini (Table 1, p. 33) (Fig. 16, p. 37).
Each uncinus bears five teeth in a single row above a rounded basal part (Figs. 16, 17, p. 37). Each un cinigerous pinnule bears a cirrus which projects posteriorly from its upper distal edge (Fig. 15, p. 36).
Segments 20-40 (-45) are abdominal segments, each with a pair of uncinigerous pinnules. The first 2-6 abdominal segments bear achaetous rudimentary notopodia.
Each uncinigerous pinnule bears a single vertical row of uncini and possesses at its dorsal posterior edge a cirrus
(Table 1, p. 33, Fig. 15, p. 36). The uncini are pectinate and possess five teeth in a single row above a rounded basal part (Figs. 16, 17, p. 37).
Between the last setigerous abdominal segment and the pygidium there may be one or two achaetous segments.
The pygidium is short, rounded, slightly lobed and generally has two clearly visible pigment spots, one on either side of the anus. Anal cirri and papillae are absent.
Internal Anatomy
There are two pairs of nephridia; the first 31
originates in the posterior part of the third setiger.
The second (visibly shorter than the first but with a
larger diameter) originates in the posterior part of the fourth setiger.
Each member of the first pair has a ciliated nephrostome extending dorsally from the edge of the ventral nerve cord to the upper edge of the ventral longitudinal muscle bundle (Table 2, p. 34, Figs. 18, 20, pp. 38, 3 9 )
leading to a nephridial elbow. This consists of a U-shaped
tube extending to about the anterior portion of the
tenth thoracic setiger and connects the nephrostome to the bladder (Fig. 18, p. 38). The short thin-walled bladder
opens dorsally to the outside through a nephridiopore on
a small (-10 p in height) papilla posterior to the notopod
of the third setiger (Table 2, p. 34, Fig. 18, p. 38).
Each member of the second pair of nephridia is
similar to the first except that the bladders open on a
small papilla posterior to the notopodia of the fourth
setiger, and the elbows end in the posterior portion of
the seventh setiger.
Each pair of nephridia is supplied and drained by a
pair of lateral branches of the ventral blood vessel
(See section on circulation). 32
The musculature comprises circular, longitudinal and oblique fibers, arranged as in Ampharete grubei (Fauvel
1897).
The oblique muscle bands divide the body into a median
intestinal and two lateral nephridial chambers. Pro
tractor and retractor muscles operate the notopodial
setae (Fig. 19, p. 38)*
A diaphragm or septum between setiger 1 and setiger 2
containing muscle fibers divides the coelomic cavity into
an anterior and a posterior thoracic chamber.
The structure of the epidermis of Amphicteis
floridus appears to differ little from that described
for Ampharete grubei, Amphicteis gunneri, Melinna palmata, and Samytha adspersa by Fauvel (1897). 33
Tablo 1. THE NUMBER 0? ADULT UNCINI PER UNCINIGEROUS PINNULE IN ONE SEXUALLY MATURE PEMALE LONG AND lna VIDE
Thoraolo S#tlgor Numbar Ku&bor of Unolnl Abdominal Sagmant Nuabar Nuabar of Unolnl
4 24 1 19
5 25 2 19
6 27 3 19 7 27 4 19
8 28 5 19 9 24 6 19
10 28 7 18
11 28 8 18
12 28 9 18
13 26 10 17
14 26 11 16
15 26 12 16
18 25 13 16
17 22 14 16
15 16
16 16
17 14
18 11
19 11
20 11
21 11
22 11
23 9 24 8 Table 2. DISTRIBUTION OF KEPHRIDIAL CELL TYPES AND MEASUREMENTS fy)
Elbov(vlde portion) Elbov(narrov portion)
FIRST PAIR Nephrostome Bladder Anterior Middle Posterior Anterior Middle Posterior
Ciliated Cells 6-10x6-6*
Ciliated Cell Nuclei 8x8
Ncn-Clllated Cells 12x9 30x30 25x15-20 25x15-20 25x17 25x17 25x18
Non-Cillated Cell Nuclei 5*5 8x8 8x8 8x8 8x8 8x8 8x8
Diameter of Nophridlal Regions 100 75 70 65 60 50 SECOND PAIR
Ciliated Cells 10-12x5-?
Ciliated Cell Nuclei 5x5
Non-Cillated Cells 15x10 25x15-20 25x15-20 25x15-20 25x15-20 25x15-20 25x15
Non-Ciliated Cell Nuclei 5*5 8x8 8x8 8x8 8x8 8x8 8x8
Dicmeter of Nophridlal Regions 120 90 JO JO 85 85 JO
* Height is indicated first and vldth second 35
Fig. 15. Lateral view in which the branchiae of the left side have been removed. The dorsal surface is toward the right
Br. Branchia
Buc. S . Buccal Segment
Cap. S. Capillary Seta
No to. Notopodium
Noto. C. Notopodial Cirrus
Pa. Palea
Pa. S. Paleal Segment
Pro. Prostomium
Py. Pygidium
Un. P. Uncinigerous Pinnule
Un. P. C. Uncinigerous Pinnule Cirrus
Un. Uncinus
1 Th. S. First Thoracic Segment
4 Th. S. Fourth Thoracic Segment 36
Br.
Pro.
Buc. S.
ftv-' * - jji Po. s. '•■ < i ? j\ '• 1. Th.S,
4. Th.S
U n. P.
Noto.C
Cop^S.
Un
,Un. P. C
. 0.6mm. F i g . 15. 37
Fig. 16. Uncini of the right, twelfth thoracic setiger. The dorsal surface is toward the right, anterior toward the top. X200
Fig. 17. Uncini of the right sixth abdominal segment. Orientation is the same as above. X200 i Nephridipore
Elbow
Nephrostome
Fig 18. Anterior right nephridium dissected out. X50
Fig 19. Right notopodium of a 18-setiger larval stage showing the muscles which operate the setal sac. X900 39
Nephrost. V.N.C.
Fig. 20 Cross section at the junction between the fourth and fifth segments. The dorsal surface is toward the top, and the right side toward the right. X75
D.L.M.B. Dorsal Longitudinal Muscle Bundle
Br. Branchia (4th Pair)
H.B. Hear t Body
Nephrost. Nephrostome
Oe. Oesophagus
V.L.M.B. Ventral Longitudinal Muscle Bundle
V.N.C. Ventral Nerve Cord
v.v. Ventral Vessel 40
SECTION V
THE CIRCULATORY SYSTEM OF AMPHICTEIS FLORIDUS
A large blood sinus surrounds the stomach and intestine.
Extending anteriorly from its dorsal part is the "heart" or dorsal vessel from which arise laterally four pairs of afferent branchial arteries (Fig. 21, p. 58). Within the dorsal vessel and the anterior part of the sinus is a mass of tissue, the "heart body" (Fig. 21, p. 58). The afferent branchial vessels extend to the tips of the branchiae. The blood descends in four pairs of efferent branchial vessels to the ventral vessel, which extends above the ventral nerve cord to the posterior end of the body before joining the blood sinus.
The extensive network of lateral blood vessels includes branches of the peri-intestinal sinus (dorso-pedal vessels) extending to the thoracic notopodia and abdominal neuropodia (Figs. 23, 24, pp. 62, 64); branches of the ventral vessel draining ventral longitudinal muscle bundles (ventral clypeal vessels) and neuropodia and notopodia (ventro-pedal vessels) (Figs. 23, 24, pp. 62, 64); lateral vessels (neuropodial and notopodial) which run the length of the body eventually joining the blood sinus in the pygidium (Figs. 23, 25, pp. 62, 65) 41
and additional vessels connecting notopodia and neuro podia.
Blood flows from posterior to anterior in the blood
sinus and the opposite direction in the ventral vessel.
Blood Sinus
The blood sinus extends from a ring around the
rectum in the pygidium to the junction between the stomach
and oesophagus. At this latter point the anterior face
of the blood sinus is swollen, forming what has been
called an annular or ring vessel, and is continuous
dorsally with the dorsal vessel or ’’heart" (Fig. 21,
p. 58).
The blood sinus is continuous internally with the
gastric invagination (See section on digestive system).
The wall of the blood sinus is tightly pressed mid-ventrally against the base of the mid-ventral groove
which extends posteriorly from slightly behind the
entrance of the gastric invagination (Fig. 22, p. 60).
A pair of unbranched dorso-pedal or transverse
vessels arise from the ventro-lateral side of the blood
sinus in each thoracic segment posterior to setiger 2 ,
and from the dorso-lateral side of the blood sinus in
each abdominal segment (Figs. 23, 24, pp. 62, 64).
The dorsal vessel begins where it leaves the 42
annular vessel and extends into the anterior portion of
the paleal segment (Fig. 23, p. 62).
It bears four pairs of lateral branches, the afferent branchials (Fig. 21, p. 58). The fourth pair of afferent branchial vessels in the third setiger loop posteriorly before continuing dorso-anteriorly into the
fourth pair of b unchiae (Fig. 21, p. 58). The third,
second, and first pairs of afferent branchial vessels
continue dorso-anteriorly into the branchiae on the
first two setigers and the paleal segment. Each afferent branchial vessel ascends to the tip of a branchia, loops
and descends as an efferent branchial vessel (Fig. 23, p. 62).
Blood flows from posterior to anterior in the blood
sinus and the dorsal vessel. From the dorsal vessel
blood passes to the afferent branchial vessels and to the branchiae. From here blood enters the efferent branchial
vessels and flows postero-ventrally into the ventral
vessel.
Heart Body
A greenish black homogeneous mass of tissue, the heart body (Figs. 21, 25, pp. 58, 65) lies unattached within
most of the dorsal vessel (Fig. 21, p. 58). It bifurcates 4 3
posteriorly, encircling the digestive tract within the annular vessel, and reunites mid-ventrally (Fig. 25,
P- 65).
The heart body has no central cavity nor is its tissue continuous with the gastric epithelium, as was indicated by Fauvel (1897) for the ampharetids he studied.
Djakonov (1913) found a similar situation in Amphicteis gunneri. A study of sections of larvae shows that the heart body is probably formed from peritoneal cells.
From observations on living specimens, I believe the heart body acts as a solid support against which the wall of the dorsal vessel contracts, when forcing blood into the branchiae. Also it seems to be forced against the openings of the afferent branchial vessels after con traction of the dorsal vessel, acting as a valve prevent ing the backward flow of blood into the dorsal vessel.
The tissue of the heart body may have a haematopoietic function as suggested for the ampharetid Melinna sp. by
Kennedy and Dales (1958).
Oesophageal Vessels
Numerous (11-15) oesophageal vessels, all ventral to
the dorsal vessel, arise from the anterior face of the
annular vessel. In worms with eleven such vessels, 44
vessels 1-5 arise from the dorsal portion of the annular vessel while the rest arise ventrally. Vessels 1-8 and
11 unite on the mid-dorsal surface of the oesophagus
(Fig. 26, p. 67). Vessels 9 and 10 continue along the ventral surface of the oesophagus for a short distance and
end (Fig. 27, p. 6 8 ). These vessels and their branches
make the oesophagus appear to be surrounded by a blood
sinus.
The single mid-dorsal vessel gives rise anteriorly to
six pairs of lateral branches (A-F) (Fig. 26, p. 67).
The first two encircle the oesophagus slightly anterior to
the origin of the third pair of afferent branchial vessels
(Fig. 26, p. 67). The third and fourth pairs arise at
about the level of the origin of the second pair of
afferent branchial vessels, and run laterally over the
surface of the oesophagus joining the fifth pair
(semi-circular vessel) at about the end of the dorsal
vessel (Fig. 26, p.67).
The sixth pair of vessels pass around either side of
the brain forming a cerebral sinus (Fig. 26, p. 67).
The mid-dorsal oesophageal vessel continues
anteriorly into the upper lip.
The oesophageal vessels supply the oesophagus, buccal
cavity, the brain, the bases of the tentacles, and the 45 upper lip. The number of branches of the anterior portion of the mid-dorsal oesophageal vessel has not been determined.
Blood is forced into the oesophageal vessels by contractions of the stomach sinus and from here via the mid-dorsal oesophageal vessel to the buccal cavity, brain, and base of the tentacles. The pathway of blood from
these areas to the ventral vessel has not been determined.
Dorso-Pedal Vessels
A pair of unbranched dorso-pedal vessels arise generally from the ventro-lateral side of the blood sinus in each segment of the thorax posterior to the second
setiger, and from the dorso-lateral side of the blood
sinus in the abdomen (Figs. 23, 24, pp. 62, 64). The dorso-pedal vessels of the 3rd and 4th setiger arise from either side of the ventral opening into the gastric
invagination (Fig. 23, p. 62). In the thorax and in
abdominal segments with rudimentary notopodia, the dorso-pedal vessels anastomose with the notopodial vessel and continue into each notopodium as a short branch (Fig. 23, p. 62). In the other abdominal segments, the dorso-pedal vessels end directly in the neuropodia (Fig. 24, p. 64). 46
Blood flows from the blood sinus via the dorso-pedal vessels to the thoracic notopodia and abdominal neuro podia. I believe that blood passes from here through the ventro-pedal and ventral clypeal vessels to the ventral vessel.
Ventral and Connecting Vessels
A vessel from the center of the inner lip (inner lip vessel) and a vessel from the center of the lower lip
(lower lip vessel) merge mid-ventrally to form the ventral vessel (Fig. 28, p. 70). Each of the vessels mentioned above branches several times supplying and draining the lips in the ventral region.
I believe that blood is supplied to and drained from the inner and lower lips by alternate filling and emptying of the anterior branches of the ventral vessel. This has been reported in the peripheral vessels of certain serpulids (Hanson, 1950) and in the blind capillaries pro jecting into the coelom of Ophelia denticulata (Gilmore,
1965) and Ophelia radiata (Claparede, 1870). This kind of circulation is well known in phoronid tentacles.
In the anterior part of the first setiger, a single circum-oesophageal vessel joins the dorsal surface of the ventral vessel (Fig. 28, p. 70). Just before reaching the surface of the oesophagus, it bifurcates, each branch 4 7 passing half way around the oesophagus. It is believed that the circum-oesophageal vessel joins the oesophageal vessels and drains! blood from this region.
The ventral vessel appears to arise from the fusion of a pair of lateral trunks in the posterior part of the first setiger. Bach of these trunks in turn results from
the confluence of three tributary vessels: the first efferent branchial vessel, a clypeal vessel, and, a neural vessel (.Fig, 28, p. 70).
The first efferent branchial vessels descend from the first pair of branchiae and join the ventral vessel
(Figs. 23, 28, pp. 62, 70). The clypeal vessels, which
always pass between the ventral circular and ventral
longitudinal muscle bundles, end blindly. The neural vessels pass under the ventral nerve cord and join the
single sub-neural vessel on either side (Fig. 28,
p. 70).
Two pairs of similarly branched trunks occur in the
second setiger and receive blood from the second and third
efferent branchial vessels as well as from the clypeal
and neural vessels (Figs. 23, 28, pp. 62, 70).
The third setiger contains a pair of trunks joining
the ventral vessel which result from the confluence of
the fourth efferent branchial vessel with vessels com- i I i 48 parable to the others previously described (Figs. 23, 28, pp. 62, 70). Extensions of the fourth efferent branchial vessels continue laterally the whole length of the body slightly below the level of the neuropodia. (Figs. 23,
28, pp. 62, 70). Because of their location and apparent function, they are here called neuropodial vessels. Each connects with the end of the blood sinus in the pygidium.
All of the remaining clypeal and ventro-pedal vessels connect with the neuropodial vessels (Fig. 28, p. 70).
Each neuropodium is drained by a branch from one of the neuropodial vessels (Fig. 28, p. 70).
Nephridial Vessels
Two pairs of lateral nephridial vessels join the ventral vessel in the third setiger and both supply and drain their respective nephridia (Fig. 28, p. 7q) . Each vessel passes over the posterior surface of the nephrostome and travels posteriorly between the upper and lower por tions of the elbow, giving off numerous blind branches.
I believe that direction of blood flow is similar to that discussed for the anterior branches of the ventral vessel.
Ventro-Pedal Vessel
From setiger 3 posteriorly, segmentally arranged pairs of ventro-pedal trunks join the ventral vessel at the level of the neuropodia. Each trunk results from the confluence of a ventro-pedal vessel, a clypeal vessel and a neural vessel (Fig. 28, p. 70). The ventro-pedal vessels pass from the notopodia or neuropodia, anastomose with the neuropodial vessel, and pass over the inner face of the ventral muscle bundles before joining the ventro-pedal trunks. Each member of the first pair of ventro-pedal vessels has a branch which extends to the appropriate notopodium of the third setiger (Fig. 28, p. 70). The neural vessels meet under the ventral nerve cord and join on either side the single sub-neural vessel. The clypeal vessels posterior to the third setiger anastomose with the neuropodial vessel.
The sub-neural and neural vessels extend into the twelfth setiger where the sub-neural vessel ends blindly.
The nerve cord posterior to this is contiguous with the body wal1.
In the fourth setiger, the first with neuropodia, four pairs of clypeal trunks connect laterally with the ventral vessel (Fig. 28, p. 70). In the first twelve setigers each trunk results from a confluence of a clypeal and a neural vessel. An additional pair of clypeal vessels joins the ventro-pedal vessels of the fourth setiger. The positions and destinations of the various vessels are 50 similar to those of the third setiger.
From the thirteenth setiger to the mid-portion of the abdomen unbranched clypeal vessels connect with the ventral vessel.
The ventrally located clypeal vessels are present from the first setiger to the tenth abdominal segment. There are usually five vessels per thoracic segment, (range=3-5) and three to four per abdominal segment.
A pair of vessels (ventro-neuropodial connectives) connects the ventral vessel in the 14th thoracic setiger with the neuropodial vessel in the third abdominal segment. These vessels fuse with the ventro-pedal vessels of segments between the 14th thoracic setiger and the third abdominal segment.
Connecting Vessels
In thoracic setigers 4-17, segmentally arranged connecting vessels connect the dorsal surface of the ventral vessel at about the level of the ventro-pedal vessels with the ventral surface of the blood sinus just to the left of the base of the ventral stomach groove
(Fig. 28, p. 70). Direction of blood flow was not determined. 51
Neuropodial Vessels
Neuropodial vessels, lateral extensions of the fourth efferent branchial vessels, join the blood sinus in
the pygidium. Each vessel lies slightly below the neuro podia, and at the junction of it and the ventropedal vessels, a short branch passes to each neuropodium
(Figs. 23, 24, pp. 62, 64). All but the first three clypeal vessels and all of the ventropedal vessels connect with these neuropodial vessels.
Notopodial Vessels
In segments 5-24 paired lateral notopodial vessels pass slightly above the notopodia (Figs. 23, 24, pp. 62,
64). Short segmental vessels pass from these vessels to the
thoracic notopodia, and to the rudimentary notopodia of the
first three abdominal segments (Figs. 23, 24, pp. 62,
64).
Inter-ramal Vessels
Posterior from the third setiger, an inter-ramal vessel connects the notopodium of each segment with the neuropodiura of the next posterior segment (Figs. 23, 24,
PP. 62, 64). Inter-ramal vessels also connect the
rudimentary abdominal notopodia with the neuropodia of
the next posterior segment. 52
Intra-ramal Vessel
Posterior from the fourth setiger, a pair of intra-ramal vessels connects the notopodia and neuropodia of the same segment (Figs. 23, 24, pp. 62, 64 )• The ends of these vessels connect with the ends of the inter-ramal vessels. Intra-ramal vessels also connect the rudimentary abdominal notopodia directly with the neuropodia.
General Description of Blood Flow
Patterns of blood flow were determined by observing living worms. Possible direction of flow in vessels too small for visual determination is also suggested.
Contraction of circular muscle fibers in the outer wall of the blood sinus forces blood anteriorly into the visible dorso-pedal vessels, and probably into all of them. The blood then passes to the abdominal neuropodia or thoracic notopodia. In the abdomen it is believed that the direction of flow is from the neuropodia via the neuropodial vessel to the blood sinus in the pygidium or from the neuropodia via the ventro-pedal and clypeal vessels to the ventral vessel and from here posteriorly to the blood sinus in the pygidium. Blood flows ventrally in the ventro-pedal vessels and posteriorly in the ventral vessel.
In the thorax it is believed that blood passes from 53
the dorso-pedal to the notopodial vessel and from here it either flows ventrally via the notopodial vessel to the ventral vessel or passes into the notopodia and then via the inter or intra-ramal vessels to the neuropodia and then to the neuropodial vessels. These possibly empty into the ventral vessel. Blood may pass from the blood sinus directly into the ventral vessel through connecting vessels.
The net effect postulated is that blood passes from the blood sinus laterally to the neuropodia and notopodia
through dorso-pedal vessels, and then to the ventral vessel where it enters the posterior part of the blood sinus to be pumped forward again.
Blood enters the dorsal vessel from the stomach sinus. It is pumped into the afferent branchial vessels and returns to the ventral vessel via the efferent branchial vessels.
Comparative Anatomy
Ampharete grubei and Amphicteis floridus have four pairs of afferent branchial vessels arising from the anterior portion of the dorsal vessel (Fauvel, 1897,
Plate XVI, Figs. 12, 15). Amphicteis gunneri and Samytha adspersa have three pairs of afferent branchial vessels, 54
the anteriormost of which bifurcates, one branch going to
the first and the other to the second branchia (Wiren,
1885, Plate I, Fig. 3). Melinna cristata and NU_ palmata have one pair of afferent branchial vessels, each of which branches into four vessels which go to the four pairs of branchiae (Wiren, 1885, Plate I, Fig. 13).
In Ampharete grubei, Amphicteis floridus, M . cristata,
M. palmata, and adspersa, the heart body is attached
to the dorsal surface of the stomach at the junction between the stomach and the oesophagus (Fauvel, 1897,
Plate XVI, Fig. 15). The blood sinus, and heart body of
the above species are similar. Fauvel (1897) attributed
a valvular and excretory function to the heart body of
ampharetids.
The dorso-pedal vessels of Ampharete grubei arise
ventrally from the blood sinus on either side of the
ventral ciliated groove (Fauvel, 1897). The first nine
arise from the sinus each as a single tube while those of
the fifteenth through seventeenth segments each arise as
two vessels which join after leaving the sinus (Fauvel,
1897, Plate XVI, Fig. 11). These vessels are otherwise
similar to those of Amphicteis floridus. The first five
dorso-pedal vessels of Amphicteis gunneri arise from the
ventral side of the blood sinus as a simple tube. The 55 remaining thoracic dorso-pedal vessels arise as a series of simple tubes joining one vessel giving the appearance of a ramifying arc with numerous pectinate ramifications
(Wiren, 1885, Plate I, Fig. 4) (Fig. 30, p. 71). In the abdominal region the dorso-pedal vessels of each side join above the blood sinus and form a semi-circular arc with numerous pectinate ramifications united by a membrane
(Wiren, 1885, Plate I, Fig. 4 ) (Fig. 30, p. 71). The first two thoracic and all abdominal dorso-pedal vessels of
Melinna cristata and M_^ palmata arise unbranched from the blood sinus while the remaining thoracic vessels join the blood sinus by a series of short branches (Wiren, 1885,
Plate I, Fig. 14) (Fig. 31, p. 71).The first five dorso-pedal vessels of Sarnytha adspersa arise ventrally from the blood sinus as simple tubes while the remaining vessels
arise by a number of tubes which embrace the dorsal portion of the blood sinus and which join in a single vessel
(Fauvel, 1897). Amphicteis floridus is the only
a-pharetid so far studied in which all of the dorso-pedal vessels appear to be unbranched.
A number of vessels arising from the annular vessel of Ampharete grubei, similar to the oesophageal vessels of
Amphicteis floridus, and illustrated but not named by Fauvel
(1897), branch over the oesophageal surface. Fauvel (1897) 56 reported that some of these oesophageal vessels supply the pharynx, brain, and the base of the tentacles. A network of oesophageal vessels has been described in
Amphicteis gunneri (Wiren, 1885, Plate I, Fig. 1).
The ventral vessel in all amph&retids which have been sufficiently studied is similar to that of Amphicteis floridus.
Fauvel1s (1897) "vaisseau anastomotique", in
Ampharete grubei, Amphicteis floridus, A. gunneri, and
Samytha adspersa, is here called the neuropodial vessel to give a better idea as to its location.
Fauvel1s (1897) "anastomose", in Ampharete grubei, and Amphicteis floridus, is here called the notopodial vessel.
The "anastomose dorso-ventrale" probably comparable to our inter-ramal vessel, connecting the notopodia to the neuropodia is mentioned by Fauvel (1897). 57
Fig. 21 Diagrammatic dorsal view of the dorsal vessel. X80.
Aff. Br. V. (1-4) Afferent Branchial Vessel
A. V. Annular Vessel
B. S. W. Blood Sinus Wall
D. V. Dorsal Vessel
H. B. Heart Body 58
Aff. Br
Aff. Br V.
Aff. Br V.
Br.V.
A.y.
B. S
0.5 mm 59
Fig. 22 Cross section of the ventral ciliated groove at the mid-portion of the stomach. The right side is toward the bottom of the page. X900.
B. G. Basal Granule
B. S. Blood Sinus
B. S. W. Blood Sinus Wall
Cil. Cilium
Cil. C. Ciliated Columnar Cell
C. R. Ciliary Rootlet
M. C.2. Mucous Cell(type 2)
N-Cil. C. Non-Ciliated Columnar Cell
V. G. Ventral Groove 60
*vv.<
B. S.W.
Ci i.e.
M.C.2
Fig. 22 50-u. 61
Fig. 23 Lateral view through the transparent body wall showing blood vessels in the anterior thoracic region. The branchiae of the right side have been removed. The dorsal surface is toward the right. X75
Cly. V. Clypeal Vessel
D-P. V. Oorso-Pedal Vessel
1-4 Eff. Br. V. 1-4 Efferent Branchial Vessel
Intra-R. V Intra-Ramal Vessel
Inter-R. V Inter-Ramal Vessel
Neur Neuropodium
Neur. V . Neuropodial Vessel
Noto. Notopodium
Noto. V Notopodial Vessel
V. V Ventral Vessel
V-P. V. Ventro-Pedal Vessel 62
Cly. v. v.v.
V-P.V.
I nter-R. V N e u r. V. Intra-R» V.
D-P. V.
N o to . V.
0.4 m m Fig. 23 63
Fig. 24 Lateral view through the transparent body wall showing blood vessels in the posterior thoracic and anterior abdominal regions. The dorsal surface is toward the right. X65.
Cly. V. Clypeal Vessel
D-P. V. Dorso-Pedal Vessel
Inter-R. V. Inter-Ramal Vessel
Intra-R. V. Intra-Ramal Vessel
Neur. Neuropodium
Neur. V. Neuropodial Vessel
Noto. V. Notopodial Vessel
V. V. Ventral Vessel
V-P. V. Ventro-Pedal Vessel 64
D-R V.
N o to . V.
lntro*R. 'V.
Inter-R.
Cly. V.
V -P . V. V.V.
DU V.
Neur. Neur. V. Oe.
H.B.
A.V.
B.S.W.
,_____ 0.5mm
Fig. 25 Diagrammatic ventral view of the anterior portion of the blood sinus. X30.
A. V. Annular Vessel
B. S. w. Blood Sinus Wall
H. B. Heart Body
Oe • Oesophagus 66
Fig. 26 Diagrammatic dorsal view of the oesophageal surface showing blood vessels arising from the annular vessel. Anterior is toward the top of the page. X80.
A-F. Branches of the mid-dorsal vessel
.A. V. Annular Vessel
M-D. V. Mid-dorsal Vessel
Oe. Oesophagus
S. V. Semi-circular Vessel
1-5. Oesophageal vessels arising from the annular vessel Fig. 26 0.5 mm A. V.
0.5 m m c
Fig. 27 Diagrammatic ventral view of the oesophagus showing blood vessels arising from the annular vessel. Anterior is toward the top of the page. X80
A. V. Annular Vessel
6-11 Oesophageal vessels aris ing from the annular vessel 69
Fig. 28 Diagrammatic dorsal view of the anterior portion of the ventral vessel. The anterior is toward the top of the page. X65.
C-0. V. Circum-Oesophageal Vessel
Conn. V. Connecting Vessel
Cly. T. Clypeal Trunk
Cly. V. Clypeal Vessel
1-4 Eff. Br. V. Efferent Branchial Vessel
I. L. V. Inner Lip Vessel
L. L. V. Lower Lip Vessel -
L. T. Lateral Trunk
N. V. Neural Vessel
Nep. V. Nephridial Vessel
Neur. V. Neuropodial Vessel
S-N. V. Sub-Neural Vessel
V. V. Ventral Vessel
V-P. T. Ventro-Pedal Trunk
V-P. V. Ventro-Pedal Vessel 70
L.L.V. Segmental Bounda r ies .C-O. V. ,C ly.V. Fi rs f Setiger .1. Eff. Br. V. L.T.
N.V. 2.E f f. Br. V.
Secor. S-N. V.
3. Eff.Br. V.
V-P. V.
N e p. V. Third Se t ig e r
N ep. V.
Conn. V,
V-P.T,
,V-P. V.
F o u r th C ly.T. Setiger V. V.
Conn. V.
Cly. V.
0.4 mm Fig. 28 Fig. 29 A single dorso-pedal vessel from the anterior thoracic region of Amphicteis gunneri (after Wiren, 1885).
Fig. 30 Fused dorso-pedal vessels from the posterior thoracic region of Amphicteis gunneri (after Wiren, 1885).
Fig. 31 Dorso-pedal vessel of Melinna palmata (after Wiren, 1885). 72
SECTION VI
THE DIGESTIVE SYSTEM
The digestive tract of Amphicteis floridus consists of buccal cavity, oesophagus, stomach, intestine, and rectum.
Buccal Cavity
The buccal cavity, (Table 3, p. 92) in specimens with retracted tentacles, extends from the ventral surface of the cephalic lobe and dorsal surface of the lower lip to about the end of the buccal segment (Figs. 3, 4, pp. 1 8 f
20). The epithelia of the tentacular fold, tentacles, upper lip, lower lip, buccal mass, and inne- lip constitute contiguous elaborations of the epithelium of the buccal
cavity. The epithelial lining is continuous posteriorly with the oesophageal epithelium, which in turn joins the
enlarged stomach.
The epithelium of the cephalic lobe is continuous
anteriorly with a semi-circular layer of columnar non-ciliated cells, the tentacular fold(Table 3, p. 92; Figs.
3, 4, pp. 18, 20). The tentacles (10-20) arise along the
lateral edge of this layer. The outer surfaces of these cells are covered by a cuticle approximately 2.5 .p thick. 73
Each tentacle is ciliated on its ventral surface and can elongate to body length.
I believe that in Amphicteis floridus elongation
and contraction of the tentacles is largely brought about by the circular and longitudinal muscles acting in opposition to each other against the tentacular coelomic fluid. Dales (1955) indicated that in the terebellid
Amphitrite johnsoni:
Most of the extension of the tentacles of Amphitrite johnsoni is due to ciliary creeping, by rolling over and opening out the ciliary groove in order to represent a flat ciliated surface to the substratum, the tentacle can crawl along rather like a planarian.
It appears probable that extension of the tentacles of
ampharetids is effected to some extent by this method of
ciliary creeping.
Meyer (1887) reported that tentacle extension in
terebellids was mainly due to thoracic coelomic fluid
pressure created by muscular contraction of four pouches
located on the first diaphragm. Dales (1955) found that
tentacle extension in the terebellid Terebella lapidaria
was not mainly due to thoracic coelomic fluid pressure,
since specimens with the anterior coelomic cavities opened,
could extend their tentacles normally. Similar operations
carried out in this study gave similar results. 74
When the buccal cavity is extended, the ciliated semi circular upper lip is directed anteriorly;in retraction, it
is within the oesophageal cavity( figs. 3, 4, pp. 18, 20).
The semi-circular lower lip is continuous posteriorly
with the buccal mass and externally with the integument of
the ventral surface of the body (Figs. 3, 4, pp. 18, 20).
The buccal mass consists of two distinct lobes, one
behind the other (Figs. 36, 38, pp. 98,100). Each lobe has
internal branching muscle fibers originating from diagonal
muscle sheets (Fig. 39, p. 100), and joining the basement
membrane of the buccal mass epithelium. Muscle fibers
entering the buccal mass from the diagonal muscle sheets
branch into the lower and inner lips.
A striated cuticle persists on the surface of the
lower lip and buccal mass, but appears to be replaced in the
rest of the digestive tract by a non-striated hardened
layer of mucous.
The buccal mass presses against the anterior
oesophageal wall when a ball of food cemented together with
mucus is taken into the mouth and oesophagus. It may
assist in swallowing by forcing food posteriorly through
the oesophagus.
The buccal mass is designated as the internal lip by
Fauvel (1897). It was decided to change the name to one 75 of more functional significance.
The buccal mass is continuous posteriorly with a small rounded inner lip.
Oesophagus
The epithelium of the buccal cavity merges im perceptibly with that of the oesophagus (Fig. 3, 40, pp.
18, 101). The oesophagus (Table 4, p. 9 3 ) is thin-walled anteriorly, and thicker near the stomach (Fig. 40, p. 101).
Peritoneal cells through which circular muscle fibers seem to pass cover the outer walls of the buccal cavity, oesophagus, dorsal vessel, and blood sinus. Maceration with Bela Haller's fluid (Appendix) indicates that the peritoneum may be syncytial. The basement membrane of the peritoneal cells rests against the basement membrane of the buccal and oesophageal epithelial cells.
The oesophagus pushes into the stomach forming a circular fold of tissue which may act as a valve in prevent ing the backward movement of food (Fig. 40, p. 101).
Stomach
Three invaginations, one central and two lateral, occur antero-ventrally in the stomach (Figs. 37, 43, pp. 98, 107). The two lateral invaginations are short; their cells resemble those of the anterior wall of the 76
stomach (Table 4, p. 93). The central invagination .called
'•inner blindsack" by Hessle (1917) and "internal diverti culum" by Day (1964), and hereafter termed the gastric
invagination.extends posteriorly in the stomach about
2/3 to 3/4 of the total length of the latter (Fig. 43, p.
107 ). The blood sinus is continuous with the inner cavity of each invagination.
The epithelium is smooth except posteriorly, where
it has numerous inwardly directed longitudinal folds
(see histology section, p. 78). A ventral ciliated groove
extends posteriorly from the base of the gastric invagina
tion (Fig. 23, p. 62).
The extreme posterior portion of the stomach is
constricted and gradually joins the intestine at about
the level of the 17th thoracic setiger (Fig. 43, p. 107).
Two bands of mucous cells in the ventral groove
opposite the heavy concentration of mucous cells on the
ventral surface of the gastric invagination provide a
possible means of effectively mixing mucus with the balls of
food passed into the stomach.
The stomach, intestine, and rectum are supported by a
continuous dorsal longitudinal mesentery consisting of
two layers of peritoneal cells (Fig. 44, p. 107). Muscle
fibers are found inside each layer of peritoneal cells 7 7 just above the basement membrane (Fig. 41, p. 103). The mesentery attaches the blood sinus to the body wall mid-dorsally between the two dorsal longitudinal muscle bundles (Fig. 44, p. 107).
There are numerous connections between the basement membrane of the epithelium of the digestive tract and the basement membrane of the peritoneal cells of the blood
sinus wall (Fig. 43, p. 107).
A peritrophic membrane such as is described in the
hind stomach of Melinna palmata by Newell and Baxter (1936)
is lacking in floridus. Its function is presumably protection of cells of the digestive tract wall from
abrasion.
Intestine
The intestine (Table 5, p. 94 ) loops left
antero-laterally, then twists;postero-dorsally and
continues straight posteriorly (Fig. 43, p. 107).
Bordering the ventral groove the intestinal
epithelium is longitudinally folded. The intestine is
continuous with a thin smooth-walled rectum at the 15th
abdominal segment (Fig. 43, p. 107).
The ventral groove ends posteriorly in the intestine.
Ventrally the intestine and rectum are supported by strap like mid-ventral mesenteries found at the boundaries of each abdominal segment beginning with the third. Each encircles the ventral vessel and attaches the surface of the ventral nerve cord to the mid-ventral surface of the intestine and rectum (Fig. 44, p. 107). Thin, lateral (transverse) mesenteries at the boundaries of each abdominal segment posterior to.the third connect the lateral walls of the intestine and rectum to the lateral body wall between dorsal and ventral longitudinal muscle bundles (Fig. 44, p. 107). All mesenteries are structurally similar to the dorsal longitudinal mesentery of the stomach.
Histology
The cells of the tentacular fold are nonciliated and columnar. Nuclei of all digestive tract cells contain a central nucleolus which stains red with Gomori's tri chrome method and hematoxylin-eosin sequences (Appendix).
The epidermis of each tentacle includes dorsal non-ciliated, ventral ciliated, and mucous cells (Fig. 32,
P. 96 )•
Cilia of ciliated cells arising from basal granules
(Fig. 33, p. 97) are best shown using a modified iron haematoxylin--orange G technique (Spencer and Monroe, 79
1961). The cilia pass through a cuticle 2.5 p thick.
Each basal granule connects to a rootlet; collectively all rootlets pass toward one side of the nucleus (Fig. 34 p. 97).
The basal area of all cells is lined by a membrane
(basement membrane) which stains purple with aldehyde fuchsin and red with PAS after treating sections in amylase
(Appendix).
The dorsal non-ciliated cells are similar to ventral
ciliated cells, but lack a ciliary apparatus.
Mucous cells, type 1*, appear blue green with alcian blue so probably secrete the copious mucus used in entrapping
food (Fig. 33, p. 97).
Layers of the tentacles from the outside inward are
the epidermis, epidermal basement membrane, peritoneal
basement membrane, peritoneal cells, circular muscle
fibers inside peritoneal cells, and longitudinal muscle
fibers originating from longitudinal muscle bundles (Fig.
32, p. 96). Each bundle sends branches into the tentacles
of its particular side. Each muscle fiber lies within a
frilled cytoplasmic membrane with a nucleus raised above
the surface similar to the parapodial muscles of some of
the Serpulimorpha (Hanson, 1948) and the muscle fibers
* See footnote in Table 3, page 92 for staining reactions from the heart of Lumbricus terrestris (Goodrich, 1942).
Transverse muscle fibers cross the tentacular cavity at regular intervals from the dorsal to ventral surfaces and are continuous with the circular muscles. Each of the transverse muscle fibers appears to be contained within a peritoneal cell.
The tentacular surface is covered by a densely stain ing layer of mucus.
The upper lip is covered dorsally by columnar
non-ciliated and ventrally by columnar ciliated and type1 mucous cells (Fig. 33, p.97). The ciliary apparatus of the ciliated columnar cells is similar to that of the tentacular cells.
The semi-circular lower lip is similar to the upper lip but lacks ciliated cells.
The buccal mass is covered by non-ciliated columnar cells (Fig. 36, p. 98).
The inner lip is covered by ciliated columnar cells.
The posterior buccal cavity and oesophagus are covered mostly with ciliated columnar cells (Table 2, p.
34 , Fig. 34, p. 97 ) and a few mucous cells (Type 1).
The dense layer covering the oesophageal mucosa stains blue-green with alcian blue, purple with aldehyde 81 fuchsin (counterstained in Halmi's mixture), red with periodic acid-schiff, and purple with azure A, and hence is probably mucus.
Numerous granules of undertermined function occur in ciliated columnar cells of the upper lip, inner lip, and oesophagus.
The extreme anterior portion of the stomach is covered by non-ciliated columnar cells with numerous projecting microvilli 2 ji thick. (Fig. 35, p. 9 7 ).
The surface layer of the gastric invagination facing the lumen of the stomach consists of non-ciliated columnar cells with microvilli, mucous cells of type 2*, and gland cells. Flask shaped mucous cells, generally restricted to the ventral surface, taper to fine points at their free edges and open into the stomach lumen by a pore1 p. in diameter (Fig. 35, p. 9 7 ). The gland cells are sparsely and evenly distributed, and are filled with granules about1 y. in diameter.
The ventra- groove consists of ciliated columnar cells
(Fig. 22, p. 6 0 ). The cilia in the right side of the groove are shorter than those of the left. A band of five to nine mucous cells mixed with non-ciliated columnar cells (Fig.
2 2 , p. 60) lies to either side of the cells forming the ventral groove in the stomach.
* See footnote in Table 4, page 93 for staining reactions 82
The anterior portion of the stomach, except in the ventral groove, is lined by non-ciliated columnar cells with microvilli, raucous cells (type 2), and gland cells (Fig. 35, p. 97).
The intestine is lined mainly with basally narrowed ciliated columnar cells (Fig. 43, p. 107). At the bottom of the longitudinal intestinal folds these cells are shorter
(Fig. 42, p. 105). Cilia, less numerous here than anterior
ly, arise from basal granules and pass through a dense surface mucous layer; ciliary rootlets are absent (Fig.
42, p. 105).
Mucous cells (type l) are sparsely distributed through the posterior intestine.
The rectum is lined by cuboidal, ciliated cells similar to those of the intestine.
Com parative Anatomy
A. en eral
Wiren (1885), Fauvel (1897) and Djakonov (1913) con sidered the digestive tract of ampharetids to include the following regions: pharynx, oesophagus, stomach, and i n t e s t i n e .
B. Pharynx 83
The pharynx as defined by the above workers included the various lips as well as the tentacles, all of which are elaborations of its wall. The naming of parts of the anterior portion of the digestive tract used by Dales (1955) for the terebellid, Amphitrite johnsoni. is accepted in the present study. The pharynx of the above workers thus becomes the buccal cavity, and the epithelium of the buccal cavity is considered as merging directly with the oesophagus. A pharyngeal region could not be distinguished in Amphicteis floridus nor by Dales (1955) in Amphitrite jo h n s o n i.
All ampharetids which have been investigated possess a superior or upper lip, an inferior or ventral lip, and a buccal mass. Amphicteis floridus possesses in addition an internal lip, figured but unnamed for Ampharete grubei by
Fauvel (1897).
Species belonging to the genera Ampharete and
Amphicteis possess a buccal mass similar to that of
Amphicteis floridus (Fauvel, 1897). Dales (1955) described in the terebellid Amphitrite johnsoni an upper lip, and two inner lips designated respectively as C. and D. The upper lip, because of its position and the fact that the tentacles are borne on a fold of tissue above it, appears homologous to that of the ampharetids. The two outer lips 84
are similar in position and general appearance to the one lower lip of ampharetids. The inner lips C. and D. are similar in structure and position to the buccal mass of ampharetids.
Sutton (1957) in her description of the terebellid
Terebella lapidaria described an inner lip region and a ventral sac which corresponds to the inner lips C. and D. of Dales (1955). Sutton (1957) in addition described a lip above the ventral sac which she calls the buccal process of the oesophagus. This structure seems to correspond to the inner lip of Amphicteis floridus.
In all ampharetids which have been investigated, the anterior portion of the buccal cavity is evertible
(Wiren, 1885, Fauvel, 1897).
The part of the body wall to which the tentacles of the pectinariids and terebellids are attached is not retractile and hence their tentacles cannot be withdrawn into the oesophageal cavity.
C. Tentacles
The tentacles of ampharetids are inserted on a fold of tissue located between the cephalic lobe and the base of the superior or upper lip (Wiren, 1885, Fauvel, 1897,
Djakonov, 1913). 85
Ampharete grubei possesses 60-80 hollow tentacles whose inner cavities are confluent with the coelom
(Fauvel, 1897). Each tentacle possesses ventro-laterally located, strongly ciliated pinnules (Plate XX, Figs. 76,
77, 73, 79, Fauvol, 1897).
Amphicteis gunneri possesses 20 hollow tentacles
(40 reported by Djakonov, 1913) which lack pinnules, and in which the glandular elements are less localized (Fauvel,
1897) . The tentacles of A_^ gunneri are ventrally ciliated and possess circular and longitudinal muscles under the epithelium (Djakonov, 1913) as in Amphicteis floridus.
The mucous cells of the tentacular epithelium of A^ gunneri are small and arranged without order on the edges of the ventral, ciliated, epithelium (Fauvel, 1897).
Melinna palmata has large, smooth tentacles in which mucous cells are distributed uniformly on the ventral, ciliated, epithelium (Fauvel, 1897).
Samytha adspersa possesses tentacles which are similar to those of Amphicteis, but in which the mucous cells are arranged in groups along the edges of the denticulated, ciliated, ventral epithelium as in
Ampharete (Fauvel, 1897).
Dales (1955) described in detail the tentacular structure of the terebellid, Amphitrite johnsoni. Each hollow tentacle ciliated only on its ventral surface is in communication with an anterior coelom and possesses longitudinal, transverse, and oblique muscle fibers under the epithelial lc.yer (Fig. 4, C. and D. Dales, 1955).
Transverse muscle fibers which traverse the hollow cavity are also found (Dales, 1955). Such tentacles bear a strong resemblance to those of ampharetids.
D. Oesophagus
The thin-walled oesophagus of Ampharete grubei communicates, as in Amphicteis floridus, with the stomach via a narrow circular opening bordered by a thick padded epithelium which forms a kind of valve (Fauvel, 1897).
The oesophageal epithelia of Amoharete grubei,
Amphicteis gunneri, Sarnytha adspersa, Melinna cristata
(Fauvel, 1897), Melinna palmata (Newell and Baxter, 1936),
Amphitrite johnsoni (Dales/ 1955), Clymenella torquata
(Ullman and Bookhout, 1949), and Lagis koreni (Brasil,
• 0 0 4 ) are similar to that of Amphicteis flor idus.
Djakonov (1913) reported mucous cells in the oesophagus of Amphicteis gunneri; this is the only previous report of mucous cells in the oesophagus of any ampharetid.
The folded oesophageal epithelium of the terebellid
Amphitrite johnsoni consists of ciliated columnar 87 and mucous cells (Dales, 1955).
E. Stomach
The stomach epithelium of Ampharete grubei consists of high columnar non-ciliated cells and gland cells, both of which seem similar to those of Amphicteis floridus although microvilli were not reported from the former
specie-. A ventral ciliated groove in Ampharete grubei
Sarnytha adspersa (Fauvel, 1897) begins, as in
Amphicteis floridus, in the anterior portion of the stomach.
Fauvel (1897) reported that only one side of this groove is
ciliated (plate XX, Fig. 85, Fauvel, 1897), while in A.
floridus both sides possess cilia, those of the left side being stouter and longer. The stomach in most ampharetids
extends to the posterior end of the thoracic cavity,
narrows, and joins the intestine through a loop similar to
that described for the species here studied. The stomach
is supported dorsally by muscular straps which join the
body wall to the mid-dorsal portion of the blood sinus
(Fauvel, 1897) instead of a continuous dorsal longitudinal mesentary as in Amphicteis floridus. Ventral supports are
lacking in both Ampharete grubei (Fauvel, 1897) and
Amphicteis floridus.
Amphicteis gunneri (Wiren, 1885, Fauvel, 1897, 88
Djakonov, 1913) Samytha adspersa (Fauvel, 1897), and
A. floridus possess a gastric invagination which extends posteriorly inside the stomach cavity. Djakonov (1913) reported that this structure was folded inward on its ventral surface, and that mucous cells similar in position to those of A^_ flor idus were borne on its ventral surface
(Figs. 1, 16, 18. Djakonov, 1913). The stomach epithelium of Amphicteis gunneri consists mainly of ciliated columnar cells (Wiren, 1885, Fauvel, 1897, Djakonov, 1913, Figs.
17, 19). Such cells are lacking in the stomach wall of the species here discussed. Circular muscle fibers, similar to those of Amphicteis floridus, are present in the blood sinus wall (Wiren, 1885, Fauvel, 1897,
Djakonov, 1913). Longitudinal muscle fibers, absent in
Amphicteis floridus are found in the wall of the blood sinus at the base of the ventral groove in Ampharete grubei, Amphicteis gunneri, and Melinna cristata
(Fauvel, 1897, Wiren, 1885).
In Melinna cristata the stomach is constricted in each segment giving it a moniliform appearance (Wiren,
1885, Fauvel, 1897). It is lined with a feebly ciliated epithelium except in the ventral groove where the cilia are most strongly developed (Fauvel, 1897).
The epithelium of the stomach of Melinna palmata as 89 described by Newell and Baxter (1936) is divided into anterior, mid, and posterior regions. The anterior region, lacking in A^_ floridus, consists of ciliated columnar cells about 90 ju high, 5 to 1C wide, and with basally located nuclei (Newell and Baxter, 1936). The
cells of the mid portion of the stomach lack basal granules,
distal granules and cilia; but possess non-motile rods at
their free edges (Plate 6 , Fig. 3, Newell and Baxter, 1936).
Perhaps these non-motile rods are the microvilli here
reported. Newell and Baxter (1936) reported a peritrophic membrane above the inside of the stomach epithelium of M. palmata. Such a membrane is lacking in Amphicteis floridus.
The stomach epithelium of the terebellid Amphitrite
johnsoni is divided into fore and hind regions (Dales,
1955). The fore stomach is thrown into longitudinal folds,
and both portions are lined with non-ciliated columnar cells
with a brush border and secretory cells (Fig. 8 E, Dales,
1955). Only the former is found in Amphicteis floridus.
The contents in this region are surrounded by a chitinous
peritrophic membrane (Dales, 1955). Perhaps this portion
of the stomach is similar to that described by Newell and
Baxter (1936) for Melinna palmata.
Brasil (1904) did not recognize a stomach region in 90
the pectinariid Lagis koreni. He divided the gut into an oesophagus and three intestinal regions. Possibly the first two intestinal regions correspond to the stomach of ampharetids and terebellids. A ventral typhlosole (a longitudinal infolding of the intestinal epithelium) is present in the first two regions of the intestine (Brasil,
1904). It is significantly different from the gastric invagination possessed by some of the ampharetids. The ciliated columnar cells in all regions of the intestine are similar to the oesophageal cells of Amphicteis floridus.
A ventral ciliated groove similar to that of Amphicteis floridus is initiated in the posterior part of the second intestinal region (Brasil, 1904).
The cells of the stomach epithelium in the maldanid
Clymenella torquata and the oweniid Owenia fusiformis bear no resemblance to those of Amphicteis floridus
(Ullman and Bookhout, 1949; Dales, 1957).
F. Intestine and Rectum:
Ampharete grubei, Amphicteis gunneri, and Melinna palmata possess a ciliated intestinal epithelium similar to that of Amphicteis floridus (Wiren, 1885, Fauvel, 1897,
Djakonov, ^.913). As the posterior portion of the intestine of Ampharete grubei is approached, the height of the cells diminishes to the point of their being cuboidal. The
intestine terminates in a thin star-shaped cavity which is probably homologous with the rectum of the species here
studied. Yellow refringent granules of the type found only
in the upper lip and oesophagus of Amphicteis floridus are
reported by Pauvel (1897) in cells of the intestinal
epithelium of Ampharete grubei. The intestine is
supported by dorsal, ventral, and transverse mesenteries
(Fauvel, 1897). The dorsal mesentery differs from that of
Amphicteis floridus in that it consists of series of muscular straps (Fauvel, 1897). The ventral and
transverse mesenteries are similar to these of Amphicteis
floridus.
The fore-intestine of Amphitrite johnsoni, the region
in which the ventral ciliated groove begins, consists of
columnar cells with a brush border, secretory cells with
heavy staining grannules, and mucous cqlls (Fig. 8, G. H.
Dales, 1955). This region appears similar to the stomach,
and the hind intestine similar to the intestine of
Amphicteis floridus.
The posterior portion of the intestine of Lagis koreni
and the intestine of Clymenella torquata are very different
from comparable regions in the ampharetids. T a b l e 3. DISTRIBUTION AND KEAEUnEKlNTS^i) 0? CELL TYPES 0? 1" BUCCAL CAVITY
Tentacular Fold Tentacles Upper 1-ip Lover Lip Buccel l-'.ass Inner Lip Posterior Epithelium
Non-Ciliated Cells 10-20x7-5* 15-20x>j-6 lo-?ox7-s 15-25x5-8 10-20x5-8 Non-Clllated Cell Nuclei 5*7
Ciliated Cells 15-20x>4-6 20-30x6-8 20-30x5-7 15-20x7-10 Ciliated Cell Nuclei >4x7 3*5 5*7 5*7
Kucous Cells(type l)** 15-20x6-8 20-30x>+ 15-20x7-10 Kucous Cell Kuelei 3*5 3*5 5*7
Kucous Covering f - V
Cutlculor Covering - }
(cross) * Height Is indicated first end width second. Measurements mado on complete serial sections of ten worms Jtrji long and 1cm vide.
** Kucous(type l) cells stain bluc-grocn with nlclan blue, purple with aldehyde fuchsin cour.torstalnod with Kilci's mixture, red with poriodic-acid schlff on sections treated with saliva. Tests with adjacent longitudinal sections of a 7 O K> T c b l o *4. D IST R IB U T IO N AND KABUR£K>:.NT S ( j i ) OF CELL TYPES OP THE OESOPHAGUS AND STOMACH Anterior end Posterior Ju: ' ion vith Anterior Stomach Gastric Kid-Stonoch Post-Stonach Ventral Ocsoph;. ;■■■: 1 Fpltheliub Stomach Invagination Croovc Ciliated Colls 25-35^-7* 30->40x5 15->40x5-7 Ciliated Cell Nuclei *4X7 >4x7 5x7 Columnar C c II b vith Klcrcvilll 15-20x5-6 20-35x6-10 15-20x5-8 30-1)0x6-10 Columnar Coll Kuolel 5x7 5x7 5X7 5x7 Kucous Colls(type 1) 25-35*5 Kucous Coll Nuclei *4x5 Kucous Colls(typa 2)** 20-35*5 20-35*5 3 ° ~ ,,0* 5 Kuoous Coll Nuclei 5*7 5*7 5*7 Gland Cells(typo 1) 30-35x5 20-35xl5-2p 30-'l0xl5-25 Gland Coll Ku ' 1 5*7 5*7 5*7 Kucous Covoring f 4 4 ♦ ^ ♦ (cross) * Height is indicated first end width second. Measurements vsro cede on oompleto sorlal coctlcns of 10 vorirs 7Ka long and lien vide . ** Kucous(typs 2) colls stain r.etachromatloally vith azure A and Safronin 0. vO U) Table 5. DISTRIBUTION AND KEASUREKEKTSQi)OF CELL TYPES C? IT : INTESTINE AND RECTUM Anterior Inteatin3 Kld-Intestlne Posterior Intestine Ventral Croovo Anterior and Posterior Roctum Ciliated Cell* 10-30x6-5* 10-35x5-8 10-20x5-8 10-35x5-8 7-10x5-8 Ciliated Cell Kuclol 5x7 5x7 3x5 5x7 4x5 Kucous Collc(type l) 10-35x5-8 10-20x5-8 Kucous Cell Nuclei 5*7 3*5 Kucous Covering f f t f f * Height is indicated first and vldth socond. Keasurccents vore nade on cocploto serial cross sections of ten vo r s . s 7 lr--o long and lea vide . vO 95 Fig. 32 Cross section of a tentacle. The ventral surface is toward the right. X1100. B. G. Basal Granule B. M. Basement Membrane C. M. C irc u la r Muscle C. R. Ciliary Rootlets C il. C ilium Cu. C u tic le L. M. Longitudinal Muscle P e r . Peritoneum 96 Fig. 32 97 C i I i u m Ba sa I G ranuI Mucous Cell ] Basement Membfone Fig. 33 Cell types of the upper lip. X1200. j Fig. 34 Cell type of the oesophagus. X1200. M icrovil lu Non Ciliated Cell Mucous Cell 2 Gland Cell Fig. 35 Cell types of the stomach. X1200. 9 8 Fig. 36 Near-sagittal section of the buccal nass. In both figures the anterior is toward the right and the ventral surface toward the bottom. X350 feral Invagination Fig. 37 Near-sagittal section of the gastric and lateral invaginations. X350 99 Fig. 38 Near-sagittal section of the lower lip and buccal mass. Anterior is toward the right, and ventral toward the bottom. Cellular details omitted. X2 0 0 . Fig. 39 Cross sec .ion through the anterior thoracic region. Cellular details omitted. X70. Buc. M. Buccal Mass C. M. Circular Muscle D. M. Diagonal Muscle L. L. Lower Lip L. M. L o n g itu d in al Muscle T. T en tacle 100 3uC. M . IX. 0.1mm F ig . 38 l . M , D. M Bue.M 0.3mm Fig. 39 1 0 1 Fig. 40 Near-sagittal section of the junction between the stomach and oesophagus. Anterior is toward the bottom and dorsal toward the right. XI00 H.B. Heart Body St. Stomach Oe. Oesophagus 1 102 Fig. 41 Near-sagittal section of the dorsal vessel and the dorsal portion of the body wall. Dorsal surface is toward the top. Cellular details omitted. X1600 B. M. Basement Membrane B. S. Blood Sinus C. C. Coelomic Cavity C. M. Circular Muscle Cu • C u tic le D. Ep. Dorsal Epithelium D. V. Dorsal Vessel H. B. H eart Body L. M. Longitudinal Muscle Pe a>• • Peritoneum 103 '.■■l: ■ \w.«i ■ ■ ' * ■ & ? . ’• > J-*. O'- i* 2 5>u Fig. 41 104 Fig. 42 Cross section of the dorsal .d-intestine. Dorsal is toward the bottom. X2000 B. G. Basal Granule B. M. Basement Membrane B.S. W. Blood Sinus Wall C. M. C irc u la r Muscle Cil. Cilium M. C. 1 Mucous Cell Type 1 105 C/I. M.C.l S __B. M. - B. M. — C. M. L B.S.W. Fig. 42. 106 Fig. 43 Stomach, intestine, and rectum. The dorsal portion of the stomach is cut away to show lateral and gastric invaginations. X25. Fig. 44 Cross section of the mid-abdominal region. Cellular details omitted. X100. An. Anus B. S. Blood Sinus B. S. W. Blood Sinus Wall C. M. C irc u la r Muscle D. Mes. Dorsal Mesentary G. I. Gastric Invagination I n t . I n te s tin e L. I. Lateral Invagination L. M. Longitudinal Muscle R ec. Rectum S t . Stomach Tr. M. Transverse Mesentary V. Mes. Ventral Mesentary 107 A n Re c I n r 0.8mm Fig. 43 B.S. Tr.M. B.S.W. V. M es 0.2mm . Fig. 44 Cross section of the posterior portion of the stomach. Dorsal is toward the left. Cellular details omitted. X120 B. S. W. Blood Sinus Wall S. E. Stomach Epithelium V.G. Ventral Groove 0.25mm. 110 SECTION VII REPRODUCTION AND LARVAL DEVELOPMENT Cleavage stages and larvae occur in female tubes from iare May to early September. The eggs develop in the central portion (anteoro-posterior ly ) of the female parent's tube and are not contained within any sac-like membrane. Eggs probably pass individually through the nephridia and nephridiopores into the tube where they are fertilized by sperm brought in by ciliary currents from the surrounding water, since both cleavage stages and early larvae are found only in the tubes of female worms with eggs in their coeloms. It appears likely that it is through the second pair of nephriaia that the eggs pass to the exterior, because they are shorter and wider. Egg re le a s e was not observed, although a tte m p ts were made to obtain shedding of gametes by changing the temperature and salinity of the water in which presumably ripe specimens were immersed. Okuda (1947) was also unable to observe shedding of gametes from Schistocomus sovjecticus. Nyholm (1950) observed shedding of eggs from the anterior end of the tube of Melinna cristata. He suggested that the eggs leave through the nephridia. I l l The eggs of Ampharete grubei and Amphicteis gunneri are shed singly into the sea through the pair of nephridia opening on the fourth setigerous segment (Fauvel, 1897). Larval development in Alkmaria romijni, Ampharete grubel, an. ’. Amohicteis f lor idus appears to be non-pelagic. Thorson (1946) said of A^ grubei, The wide distribution of this species in high-Arctic seas where, as is well known, the pelagic development is suppressed in nearly all species, seems to support the assumption that the larval development is non- p e la g ic . Larval stages beyond the three-setiger stage are non-pelagic and crawl on the mud surface. Hence it is assumed that they crawl out the anterior opening of the tube between the two- and three-setiger stage. Nyholm (1950) reported that the larvae of Melinna cristata normally have three setigers when they become truly benthonic. Although 50 ripe specimens were kept in circulating sea water two months, none of these spawned either naturally or under inducement. Twenty attempts at artificial fertilization of eggs of females with larvae in their tubes were unsuccessful, perhaps because the eggs were not naturally shed or that the gametes were unripe. Measurements of the larval stages are given in 112 Tables 7-10 (pp. 129-132 ). The unfertilized egg is greenish, opaque, irregularly elliptical, with a large germinal vesicle (visible nucleus) (Fig. 46, p. 133). The egg is enclosed within two thin membranes, both approximately 2 ju thick, and contiguous with the egg surface. After fertilization the outer membrane only is raised from the egg surface. The chemical nature of the membranes was not determined. An estimated timed Table of Development has been worked out for larval stages from the late cleavage to the eighteen-setiger stage, and the times are given in parentheses below the section headings. Early Embryology (Figs. 47-50, pp.133-135 )(Blastula--! hour, Gastruia--3 hours, Early Trochophore--5 hours, Trochophore--8 hours) At fertilization the outer membrane rises from the egg and the germinal vesicle disappears. The two-cell stage has unequal blastomeres (Fig. 47, p. 133 ). The four-cell stage has three similar nearly spherical cells and one larger cell. The eight-cell stage has seven similar cells and one larger cell. Cleavage culminates in a coeloblastula (Fig. 48, p. 135 ) with an apical tuft of long and short cilia and a broad prototroch of moderately long cilia. Gastrulation was not observed. The stereogastrula lengthens between the prostomium and pygidium; a pygidial 113 telotroch forms, resulting in the early trochophore (Figs. 49, 50, p; 135). A prototroch (band of cilia) completely surrounds the anterior portion of the larva (Figs. 49, 50, p. 135 ). The apical tuft persists, and large endodermal cells, presumably forerunners of the digestive tube, fill the interior. The stomodeura is a slight raid-ventral depression posterior to the prototroch (Fig. 50, p. 135 ). No proctodeum, eyes, or other pigment spots are evident at this stage. In the late trochophore, dorsal red eyespots occur anterior to the prototroch (Fig. 51, p. 137). Other red pigment spots of undetermined nature occur, on the anterior and posterior borders of the prototroch, and lateral to the presumptive proctodeum at the tip of the pygidium (Fig. 51, p. 137). Still later, as the trochophore elongates, the prototroch and telotroch are narrower, with fewer cilia, possibly due to resorption. Pigment spots are more evident and clefts in the seemingly solid mass of endodermal cells indicate resorption of yolk. Still later, only the stiff sensory cilia remain apically. A new ventral band of cilia extends from the anterior border of the prototroch to the anterior edge of the prostoraium. This stage possesses a prototroch with stiff sensory cilia and eyespots, an obscure peristomium, a 114 bi-annular achaetous segment, and a pygidium with a telotroch (Figs. 51-53, pp. 137, 139). One S e tig e r S tage (Twenty hours) The apical cilia have disappeared and the prototroch is narrower. Two new segments have been added between the bi-annular achaetous segment and the pygidium. The first of these bears the first pair of notopodia, each equipped with a single spatulate seta (Fig. 53, 56, pp. 139,140). The second is achaetous, but will later bear the second pair of notopodia. The first new segment corresponds withthe paleal segment of the adult, while the second corresponds with the first post-paleal setiger of the adult. Two new dorso-ventrally complete bands of cilia (metatrochs) occur, one in the center of the groove separating the halves of the bi-annular achaetous segment, and the other in the center of the paleal segment. In this paper the larval stages are named on the basis of the number of segments (setigers) equipped with notopodia possessing setae. In the adult, the term setiger is used to denote a segment possessing setae of the capillary or uncinate type. The paleal segment of the larval stages is the first setiger. In addition, when 115 describing the segmentation of larval stages, the setigerous segments which follow the paleal may also be referred to as post-paleal setigers. Two-Setiger Stage (1 day 6 hours)(Fig. 54, p. 139) The prototroch is further narrowed and the first post-paleal setiger has a metatroch completely around the middle of the segment. The first post-paleal setiger also bears a pair of notopodia each of which has a single spatulate seta. Three-Setiger Stage (1 day, 18 hours--2 days)(Fig. 55, p. 140) A new segment is formed between the pygidium and the first post-paleal setiger, which develops in succession a pair of spatulate setae. When this second post-paleal segment is complete, a single smooth capillary seta is formed in each of the four notopodia of the paleal and first post-paleal setigers. A metatroch develops and completely encircles the second post-paleal setiger. The prototroch is further narrowed and the first pair of larval uncini appear on the first post-paleal setiger on a pair of rudimentary neuropodia. The larval uncini are pectinate 116 and bear thirteen teeth above the basal part (Figs. 57, 58, P* 140 )• The stomodeum and proctodeum are continuous with the now clear lumen of the digestive tube and possess cilia on their inner surfaces. The setae of the second post-paleal setiger are decidedly smaller than those of the paleal and first post-paleal setigers. Later two segments can be discerned between the pygidium and the second post-paleal setiger. The first of these bears a pair of notopodia which lack setae, and also a pair of rudimentary neuropodia each with a single pectinate larval uncinus. The second of these segments lacks notopodia, but bears a pair of neuropodia, raised a distance from the body wall, each of which is equipped with a single pectinate larval uncinus. Four-Setiger Stage (2 days 16 hours--5 days 16 hours)(Fig. 59, p. 141) Each member of the pair of notopodia of the segment immediately following the second post-paleal setiger, has a single smooth capillary seta. This segment becomes the third post-paleal or fourth setiger. Setae formed in the notopodia of the third post-paleal and subsequent setigers are of the adult smooth capillary type, and no new 117 spatulate setae are formed. A metatroch completely encircles the third post-paleal setiger. An oesophagus, stomach, and intestine can be discerned. The intestine, just after leaving the stomach, curves first anteriorly, then dorsally, continuing in a straight line posteriorly to the anus, which perforates the center of the pygidium. The red pygidial pigment spots remain as a pair of compact red massestone on either side of the pygidium. The prototroch has been reduced to a single complete ring of cilia, and the telotroch has been obliterated except for a few stiff sensory cilia present at the tip of the pygidium. Five-Setiger Stage (6 days 5 hours--8 days 5 hours)(Fig. 60, p. 141) The third post-paleal setiger has an additional smooth capillary seta per notopodium,and the segment which follows it has acquired a pair of notopodia each with a single smooth capillary seta. A segment can be discerned following the fourth post-paleal setiger, bearing a pair of neuropodia each with a single pectinate larval uncinus. The prototroch remains and a metatroch develops which completely encircles the fourth post-paleal setiger. Later an additional smooth capillary seta is added to each notopodium of the fourth post-paleal setiger, and 118 four segments, three added since the above five-setiger stage are formed between the pygidium and the fourth post-paleal setiger. Each segment which is subsequently added first develops a pair of neuropodia, each member of the pair with a single pectinate larval uncinus, and then (up to the eighteenth post-paleal setiger) a pair of noto podia, each with a smooth capillary seta. More uncini and smooth capillary setae are subsequently added. The heart body is visible as a mass of tissue located dorsally above the junction between the oesophagus and stomach. Six-Setiger Stage (9 days 6 hours--10 days 13 hours) Four segments, one added since the above stage, are present between the pygidium and the fifth post-paleal setiger. The metatrochs circling the paleal and first through fourth post-paleal setigers disappear, . However the metatroch around the middle of the bi-annular achaetous segment remains. The prototroch is completely lost when the larva passes from the five to the six-setiger stage. A single tentacle, ciliated only on its ventral surface, projects anteriorly from the tentacular fold (Fig. 62, p. 142). 119 Seven-Setiger Stage (11 days 13 hours--13 days 14 hours)(Figs. 61, 62, p. 142) There are three segments, none added since the previous stage, between the pygidium and the sixth post-paleal s e t i g e r . Eight-Setiger Stage ;14 days 11 hours--16 days 10 hours)(Fig. 63, p. 143) There are five segments, three added since the previous stage, between the pygidium and the seventh post-paleal setiger. A pair of evaginations of the body wall are present on the dorsal surface of the larva slightly above the notopodia of the paleal setiger; these develop into the first pair of branchiae. Nine-Setiger Stage (17 days 10 hours--18 days 10 hours) There are six segments, two added since the previous stage, between the pygidium and the eighth post-paleal setiger. This stage is otherwise similar to the eight setiger stage. The number of capillary setae per notopodium and the number of uncini per neuropodium in the nine- through the eighteen-setiger stages are included in Table 6, page 128. 120 Ten-Setiger Stage (20 days 12 hours)(Fig. 64, p. 143) There are seven segments, two added since the previous stage, between the pygidium and the ninth post-paleal setiger. This stage is similar to the eight- and nine-setiger stages except that there is a general increase in the number of adult smooth capillary setae. Eleven-Setiger Stage (21 days 14 hours--22 days 15 hours) There are seven segments, one added since the previous stage, between the pygidium and the tenth post-paleal setiger. The metatroch circling the bi-annular achaetous segment disappears. The number of tentacles has increased to three. One long pair of branchiae is present on the dorsal surface of the paleal setiger. The number of larval uncini has increased since the ten-setiger stage (Table o, page 128). Tv/elve-Setiger Stage (24 days 13 hours)(Fig. 65, p. 144) There are seven segments, one added since the previous stage, between the pygidium and the eleventh post-paleal setiger. The number of capillary setae as well as the number of uncini has increased (Table 6, p. 128). 121 Thir teen-Setiger Stage (26 days 12 hours--28 days 11 hours) There are six segments, none new, between the pygidium and the twelfth post-paleal setiger. No change was noted in the numbers of setae per notopodium or neuropodium with the exception of the one pair of smooth capillary setae added to the twelfth post-paleal setiger. Four teen-Setiger Stage (30 days 15 hours) There are seven segments, two added since the previous stage between the pygidium and the thirteenth post-paleal setiger. There are two pairs of branchiae located dorsally on the paleal and the first post-paleal setigers on either side of the mid-line just behind the notopodia. There is a general increase in the numbers of setae per notopodium and the neuropodium (Table 6, p. 128). Adult uncini are found for the first time (Table 6, p. 128). The larval uncini on the rudimentary neuropodia of the first and second post-paleal setigers have disappeared. Fifteen-Setiger Stage (33 days 9 hours) There are six segments, none new, between the pygidium and the fourteenth post-paleal setiger. The number of setae 122 has increased (Table 6, p. 128). Sixteen-Setiger Stage (Figs. 67, 68, p. 145) There are seven segments, two new, located between tha pygidium and tha fifteenth post-paleal satiger. The number of setae is about the same as the previous stage (Table 6, p. 128). Seventeen-Setiger Stage (35 days 14 hours) Six segments, none new, are found between the pygidium and the sixteenth post-paleal setiger. The number of setae is about the same as the two previous stages (Table 6). There is in addition to the two well developed pairs of branchiae, a third pair which is located on the dorsal surface of the second post-paleal setiger just above the notopodia. Eighteen-Setiger Stage (36 days 14 hours) Twelve segments, six new since the previous stage, are found between the pygidium and the seventeenth post-paleal setiger. The number of setae has increased (Table 6, p. 128). Neuropodial cirri are present on the upper portion 123 of all but the last six abdominal neuropodia. No cirri are present on the notopodia. The changes which occur between the above mentioned stage and the adult worm are: 1) an increase in the number of abdominal segments to about 23, 2) an increase in the number of paleal setae to about 8, 3) an increase in the number of adult uncini per neuropodium, 4) the loss of all larval uncini, 5) the addition of a fourth pair of branchiae on the dorsal surface of the third post-paleal setiger just above the notopodia, 6) the addition of a small cirrus on each notopodium, 7) the development of a gastric invagination and 8) the development of up to twenty oral tentacles. Larval Ecology Larvae began tube building just after leaving the female parent tube. Several 3-setiger larvae placed in a Petri dish containing a small amount of surface mud had formed a typical mucous tube within a few minutes. The ventral shields are the only apparent source of mucus used in forming the tube. Nyholm (1950) reported that the first stage in tube formation of Melinna eristata larvae, is a mucous secretion from just behind the prototroch. Larvae first contain microscopic plant and animal 124 material in the digestive tract. Until the formation of tentacles, they feed by forcing material from the mud surface into the digestive tract by cilia on the upper lip and by muscular action of the buccal mass. Com parative Anatomy A. G eneral Embryos of Alkmaria romijni were found in the coelom of adult females and within female tubes. Thorson (1946) reported that A_^ romijni was a protandric hermaphrodite, and rhat in ripe females there were sometimes twenty to thirty larvae attached around the mouth of the female tube. Thus we have evidence that this species has a non-pelagic form of development. B. Spawning Natural spawning has been observed in Ampharete grubei and Amphicteis gunneri by Fauvel (1897) and by Nyholm (1950) in Melinna cristata. C. Artificial Fertilization Fauvel (1897) was unsuccessful in obtaining artificial fertilization of the eggs of A_^ grubei . as was Nyholm (1950) for ^4elinna cristata. Okuda (1947), however, artificially fertilized the eggs of sovjeticus, and raised the subsequent larvae to the three-setiger stage. 125 D.Unfertilized Egg, Early Cleavage Stages, and Trochophore The unfertilized egg, early cleavage, trochophore, and one-through three-setiger stages of Schistocomus spy/jeticus (Okuda, 1947) and Molinna cristata (Nyholm, 1950) correspond in general appearance to those of Amphicteis floridus. The two-cell stage of sovjeticus is formed two hours after fertilization, the four-cell stage 30 minutes later and the trochophore stage less than 18 hours later. The trochophore of sovjeticus lengthens in a manner similar to the species here studied. The only observed difference between the trochophores of these two species appears to be that the prototroch of sovjeticus has a slight gap mid-ventrally while that of A. floridus is complete. The late trochophore of Amphicteis floridus consists of a prostomium, a peristomium, a bi-annular achaetous segment, and a pygidium (Fig. 51, p. 137). The peristomium, indistinguishably separable from the prostomium until the prototroch is resorbed, is fused as in Scoloplos armiqer to the first definable trunk segment (Anderson, 1959). Dales (1950), as a result of his work on Nereis 126 diversicolor, gives the following definitions of the prostomium and peristomium: 1) prostomium: that region which lies anterior to the mouth and contains the brain, bears the eyes and the various appendages, and shows no obvious signs of a segmental nature. Its composition is disputed, but whatever the structure of the prostomium may be, it is comparable unit throughout the polychaetes, 2) peristomium: a variable structure which should always be defined for individual species. It often includes more than one segment, but typically is the first segment of the body surrounding the mouth. The prostomium and peristomium of the ariciid Scoloplos armiger are pre-segmental; the mouth region, in giving rise to the peristomium, may become fused with the first definable trunk segment (Anderson, 1959). E. Tw o-Setiger S tage The two-setiger stage of S^_ sovjeticus possesses a short band of fine cilia (Neurotroch) extending ventrally from the prototroch to the pygidium (Okuda, 1947). Such a band is lacking in Amphicteis floridus. No ciliary bands other than the prototroch and telotroch appear on M. cristata larvae (Nyholm, 1950). S. sovjeticus larvae which reached a length of 260 p, 70 hours after fertilization, included besides 127 the prostomium, peristomium, and pygidium, three middle segments (Okuda, 1947) the first was achaetous with no indication as to whether or not it possessed a metatroch. The second bore a metatroch and the first pair of notopodia. The third possessed the second pair of notopodia and a pair of neuropodia with each neuropodium bearing a single pectinate uncinus with three teeth above a basal part. No mention is made by Okuda (1947) of larval uncini like those found in A_^ floridus. F. Three- Through Five-Setiger Stages The three-setiger stage of S_^ sov jeticus is formed when the larva is four days old and 280 p in length (Okuda, 1947). Spatulate and capillary setae can be seen in a photograph of a three-setiger stage of M. cristata (Nyholm, 1950). Okuda (1947) working with S_^ sov jeticus and Nyholm (1950) working with NL_ cristata were unable to rear larvae past the three-setiger stage, but Ostrooumov (1899) briefly described the four and five-setiger stages of Hypania invalida similar to those of Amphicteis floridus. Tables 6. THE: NUMBER OF SMOOTH CAPIM ARE SET/." PER NOTOPOOIIT! AMO UMGi::i PER Ull:WIHEROUG PINNULE OH SEEIGt'RG'j:. A.NO POSE -SETIOERCUo SEGMENTS CF THE J-l8 SET I PER STAGFS Number of Smooth Capillary Setae Number of Unolni Adult Larval Adult Setlgor Stege 9 10 11 12 13 14 15 16 17 18 9 10 ll 12 13 14 15 16 17 13 9 10 11 12 13 14 15 16 17 18 Setiger Nur.oor 1 1 1 1 1 1 1 1 1 1 3 0 0 0 0 0 0 0 0 0 0 0 0 ’ 0 0 0 0 0 0 0 0 2 1 1 1 1 1 1 2 1 2 4 1 1 1 1 l 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 3 1 2 1 2 2 2 2 2 2 4 1 1 1 1 l 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 4 2 3 3 4 4 4 3 4 4 5 1 1 1 1 l 1 1 1 0 0 0 0 0 0 0 0 0 0 0 0 5 2 3 3 3 3 3 4 4 4 5 1 2 2 2 2 3 3 3 3 2 0 0 0 0 0 0 0 1 1 2 6 2 3 3 3 ✓ 3 4 4 4 5 1 1 2 2 2 2 2 3 3 2 0 0 0 0 0 1 1 1 1 2 7 2 2 2 3 3 3 4 4 4 5 1 1 2 2 2 2 2 2 3 3 0 0 0 0 0 1 1 1 1 2 6 2 2 2 2 2 3 4 4 4 5 1 1 1 2 2 2 2 2 2 2 0 0 0 0 0 1 1 1 1 2 9 1 2 2 2 2 2 3 aJ 3 4 1 1 1 2 2 2 2 2 2 2 0 0 0 0 0 1 1 1 1 2 10 2 2 2 2 2 3 3 3 4 1 1 2 2 2 2 2 2 2 0 0 0 0 0 1 1 1 2 A ll 1 2 2 2 3 3 3 4 1 1 1 1 1 2 2 2 0 0 0 0 o- 0 0 12 1 2 2 3 3 3 4 1 1 1 1 2 2 2 0 0 0 0 0 0 2 13 1 1 2 3 3 4 1 1 1 1 2 2 0 0 0 0 0 2 14 1 1 2 3 3 1 1 1 2 2 0 0 0 0 2 15 1 2 2 3 1 1 2 2 0 0 0 2 16 1 2 3 1 1 2 0 0 2 17 1 3 1 2 0 1 13 2 2 • 1 ”?ost-Setl£crGUs" Table 7. MEASUREMENTS^) and n o sers 0¥ LARVAL stages( unfertilized egg to the trochophore )found in nature Unfertilized Egg Fertilized Egg Cooloblastula Early Trochophoro Trochophoro Late Trochophore 1 Length Kean 170x155* 155xl6oe 153 162 180 198 Observed Range 175x150 tc 145-165 150-155 160-165 170-190 193-205 160x14-3 Prototroch Vldth Kean 144 126 126 126 Observed Range 142-145 120-128 123-130 124-129 Prototroch Breadth Keen 72 72 No Measurements for both Ob30ivod Range 70-74 70-74 Tclotrcch Vldth koan 72 81 77 Observed Rango 71-74 77-85 75-85 Telotrcci. Breadth Kean ~ 35 No Measurements for both Observed Range 30-33 Number of Individuals 10 eggs from 4 eggs from 5 from one 20 frcn one 20 frca or.s 10 froh or.e each of 20 tubes one tube tube tube tuts tube * Diameter 9 2 1 Table 8. KEASUREKEH KUK3ERS OF LARVAL STAGES( LATE TROCHOPHORE TO THE THREE-SETIGER STAGE )F0UHD IN NATURE Late Trcohophore 2 Late Trochophore 3 1-Sotlgor 2-Sctlger 3-Setlgcr( early) 3-Set lger(nld) Length Kean 22* 252 270 2 70 315 432 Observed Renge 220-235 21*5-255 265-275 265-275 290-320 400-450 Prctotroch Vldth Kean 108 113 81 72 72 72 Observed Range 103-110 110-115 75-86 68-76 70-75 Prototroch Breadth Keen Ho Kcasurcnonte Ho Koasurcnonts 4o 35 25 Ho Keasurenents Observed Rango 35-^5 30-40 20-30 Telotrcch Vldih Keen 67 63 60 63 63 No Keasurenents Obcorved Range 63-69 60 55-65 60-67 60-66 Nucbor of Individuals 4 fron one tube 5 fron tvo tube# 20 fron 80 fron 4o 5 four tubes fivo tubes T a b l e 9. KEASUREKENT3 (ji) D ITO'.BFHS C? LARVAL STAGES( THREE-SETIGER TO THE 1JIHE-SETICER STAGE )EO’Jl!D D1 NATURE 3-Setl£or(lato) •t-Setlger 5-Setl£cr(cerly) 5-Sctlccr(late) 6-Sotlger 7-Setlger 8-Setiger 9-s< Kean *♦*+1 1*68 **S5 £>10 657 632 536 S50 Observed Rcn£9 *430-450 *460-475 •*85-515 630-655 650-6CO Knxlrcuni Vldth Keen 72 76 SO 100 SO 117 135 150 Observed Rangs 70-75 73-So 85-95 90-110 85-100 Nur.bcr of Individual* 6 10 8 >1 3 1 1 1 131 T able 1 0 . KEA.SUREKEMTS(ji)A!JD NUMBERS CP LARVAL STAGES (TEN-SF.TIGER TO Ti'.E EIGHTEEN-SETIGER STAGE) FOUKD IN MATURE 10-Setigor 11-Setigcr 12-Setiger 13-Setlgor l4-SotIger 15-Sotlger l6-Setlgcr 17-Setiger l8-Sctiger Length Kean 95° 962 1220 1260 I30O 1405 1600 1900 2370 Observed Range - 955-970 1200-1240 I25O-I265 I25O-I350 1350-1420 230O-25OO Kaxltrua Vldth Kean 160 170 165 185 180 210 220 260 2f0 Observed Range IC5-I75 I8O-I9O I8O-I9O 175-190 200-220 255-270 Kucher of Individuals 1 5 2 4 6 3 1 1 •> 133 ler Membrone Nucleol us Inner Kembrane Germ i no I Vesicle 9 0 u Fig. 46 Unfertilized egg 9 Ou - 1 Fig. 47 Two-cell stage 134 Fig. 48 Coeloblastula. Dorsal view. Fig. 49 Early trochophore. Dorsal view. Fig. 50 Early trochophore. Ventral view. A. T. A pical T uft D. T. Digestive tract Pro. Prostomium Pt. Prototroch Py. Pygidium S t. Stomodeura (Developing) Tel. Telotroch 1 135 A.T. P ro Fig. 48 6 5m F ig . 49 St. Fig. 50 -6V , 136 Fig. 51 Late trochophore. Dorsal view. Fig. 52 Late trochophore. Lateral view. The dorsal surface is toward the left. A. Pro. P. S. Anterior Prostomial Pigment spot Ey. S. Eyespot P. Pt. P. S. Posterior Prototroch Pigment Spot Py. P. S. Pygidial Pigment Spot V. Pro. Cil. Ventral Prostomial Cilia 137 Fig. 52 70K f 138 Fig. 53 One-Setiger stage. Dorsal view. X400. Fig. 54 Two-Setiger stage. Dorsal view. X280. B . A . S . Bi-annular Achaetous Segment Met. Metatroch No to. Notopodium Perist. Peristomium s . c . s . Smooth Capillary Seta s. s. c. Stiff Sensory Cilia Sp. s . Spatulate Seta 139 • s. S.C. /Si • . • >/ / • • ■ ' ' - p MiMW'1 >i /&x 7V<*. . 50m Fig. 53 m m w m - ::.\V:VV-W • s.c.s. 80u Fig. 54 I n i 2 140 Fig. 55 Three-Setiger stage. Dorsal view. Fig. 56 The first two notopods and accompanying setae of a three-setiger stage. Dorsal view. Fig 57 Larval uncinus. Fig. 58 Larval uncinus, Lateral view. Dorsal view. 141 Fig. 59 Four-setiger stage compressed slightly Dorsal view. X200 Fig. 60 Five-setiger stage. Dorsal view. X165 Fig. 61 Seven-setiger stage. Lateral view. X140 Fig.682 Anterior end of the above specimen showing the single ciliated tentacle. X500 Fig. 63 Eight-setiger stage. Dorsal view. X100 Fig. 64 Ten-setiger stage. Dorsal view. X95 144 Fig. 65 Twelve-setiger stage compressed slightly. Ventral view. X85 Fig. 66 Fourteen-setiger stage. Ventral view. X70 145 Fig. 68 Anterior portion of the above specimen. X200 146 SECTION V III SALINITY TOLERANCE Salinity at the local collection sites of A_^f lo r id u s normally varies from approximately 27 o/oo at high tide down to approximately 10 o/oo at low tide largely as the result of variation in fresh water runoff. Some worms, however, occur in high intertidal pools, where salinity reaches 40 o/oo at low tide because of evaporation, though mixed with water of lower salinity at high tide. Animals exposed to salinities exceeding 35 o/oo or less than 0.5 o/oo in the laboratory survived only short periods (Table 11, p. 147). For te s tin g th e ir s a l i n i t y to le ra n c e , ten worms were removed from their tubes and placed in each of thirteen covered (9" ) finger bowls filled with 1" of sea water of known salinity. They were examined periodically for eight days. Each experim ent term in ated when a l l the worms no longer could be observed pumping blood. Sudden exposure to extreme salinities did not cause cessation of pumping of blood in any worms. The salinities had not changed significantly at the termination of each experiment. 147 TABLE 11 EXPERIMENTAL SALINITY TOLERANCE Survival S a l i n i t y in o/oo Number o f Days Number o f Hours 0 0 0 0. 5 8 0 1 8 0 2 8 0 5 8 0 10 8 0 20 8 0 30 8 0 35 8 0 40 1 0 45 1 0 50 0 2 60 0 2 70 o . — 0 148 SECTION IX GENERAL DISCUSSION In the past external characters have been used for the most part in separating genera and species in the Ampharetidae. Internal characters such as the number of pairs of nephridia and the presence or absence of a gastric invagination, though they were used by Hessle (1917), were not considered useful by Day (1964). Such characteristics appear to me to be of increasing taxonomic significance. The question arises as to whether such internal characters are valid criteria for full generic or specific separation. The value of such characters can be determined only after they have been established for each member of the group. As more species are thoroughly studied,a considerable body of knowledge on internal anatomy will be developed within the family, and it is believed that the usefulness of such characters will be demonstrated. A study of the various organ systems is also important in determining phylogenetic relationships within the family and closely related families. The upper lip, lower lip, buccal mass, and tentacles of ampharetids appear to have structurally similar counter parts in the terebellids, Amphitrite johnsoni (Dales, 1955) and Terebella lapidaria (Sutton, 1957). Also the oesophageal epithelia of the terebellid Amphitrite johnsoni (Dales, 1955),the maldanid Clymenella torquata (Ullman and Bookhout, 1949), and the pectinariid Lagis koreni (Brasil, 1904), appear similar to that of ampharetids. On the basis of anatomy, the morphology of the digestive and circulatory systems Amphicteis floridus shows resemblances to genera within the family Ampharetidae. Notopodial cirri and a gastric invagination are shared by Amphicteis floridus with the genera Amage and Amphicteis (a few species). External and internal characters have not been adequately investigated in all genera for detailed comparisons to be made. The genera Ampharete and Amphicteis have tentacles, a buccal mass, a network of oesophageal vessels, and a digestive tract similar to those of Amphicteis floridus. Amphicteis gunneri and A^ floridus are the only ampharetids reported with mucous cells in the oesophagus; both possess a gastric invagination and a similar ciliated intestinal epithelium. The fact that Amphicteis flor idus can tolerate waters 150 of such a wide range of salinity suggests that it might be capable of colonizing a wide array of habitats. Further work on this problem would be of interest, especially a study of the salinity tolerances of developmental stages. 151 BIBLIOGRAPHY A nderson,D. T. , 1959. The embryology o f the p o ly c h ae te Scoloplos armiger. Quart. J. Micr. Sc., 100:89-166. Annekova, N., 1927. Ueber die pontokaspischen Polychaeten. I. Die Gattungen Hypania Ostrooumov und H y paniola, n. gen. Ann. Mus. Zool. Acad. S ci. U. R. S. S. 28:48-62. Benham, W. B., 1894. 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U. K. 29:321-360. Dales, R. P., 1955. Feeding and digestion in terebellid polychaetes. J. Mar. Biol. Ass. U. K. 34: 55-79. Dales, R. P., 1957. The feeding mechanism and structure of the gut of Owenia fusiformis Delle Chiaje. J. Mar. Biol. Ass. U. K. 36:81-89. Dales, R. P., 1962. The polychaete stomodeum and the interrelationships of the families of Poly chaeta: Proc. Zool. Soc. London. 139:389-428. 153 Dales, R. P., 1963. Annelids. Hutchinson and Co. London 2 0 0 p. Day, J. H., 1964. A review of the Family Ampharetidae (P o ly ch a e ta). Ann. S. Afr. Mus. ^*8: 97-120. Djakonov, A. M., 1913. Anatomisch-histologische Untersuchungen des Darmes von Amphicteis gunneri. Soc. Nat. Zool. Phys., St. Petersburg, Trav. 42 (Livr. 4)(2): 297-346. Dragoli, A. L., 1961.{peculiar feeding habits in the Black Sea polychaete Melinna palmata Grube.^ Doklady Akademii Nauk SSSR. 133: 970-973. (Translated in Biol. Sci. 138: 534-536). Ehlers, E., 1887. Report on the annelids of the dredging expedition of the U. S. coast survey steamer B lak e. Mus. Comp. Zool. H arvard, Mem. _15: 1-335. Fauvel, P., 1897. Recherches sur les Ampharetiens, Annelides polychaetes sedentaires. Morphologie, Anatomie, Histologie, Physiologie. Bull. Sci. Fr. Belg. 30: 277-488. 154 Fauvel, P., 1927. Polychetes sedentaires. Faune de France. 16: 1-494. Foot, N. 1945. Pathology in surgery. Lippincott, Philadelphia xi+511 p. Friedrich, H. 1938. Polychaeta. Die Tierwelt der Nord-und Ostsee. Gegriindet von G. Grimpe und E. Wagler, Leipzig. 32 (6b):1-201. Gilmore, C. R., 1965. Personal Communication. Goodrich, E. S., 1942. A new method of dissociating cells. Quart. J. Micr. Sci. 8_3: 245. Gosse, P. H. , 1855. On new or little known marine animals Ann. Mag. Nat. H i s t ., ser 2.1_6: 31-35, 308-312. Grimm, 0. A., 1877, Das Kaspische Meer und seine Fauna. Berichte der Aralo-kaspichen Expedition, 1876-1877 (In Russian). 1-168 and 1-105. Grube, A., 1851. Die Familien der Anneliden. Arch. Naturg. Berlin. 16.1: 249-364. Gurr, E., 1962 Staining practical and theoretical. The W illiam s and W ilkins Co. B altim o re, x +631 p. 155 Guthrie, M. and J. Anderson, 1959, Laboratory directions in general Zoology. John Wiley & Sons. New York. 233 p. Hanson, J. , 19*1-8. An unusual type of muscle fibre. Quart. J. Micr. Sci. jS9: 139. Hartman, O., 1951. The littoral marine annelids of the Gulf of Mexico. Publ. Inst. Mar. Sci., Univ. of Texas. 2J: 7-124. Hatschek, B., 1893. System der Anneliden, ein vorlaufiger Bericht. Lotos, Prag. _13: 123-126. Hemplemann, F., 1931. Erste und zweiter klasse der Vermes Polymera (Annelida). Archiannelida und Polychaeta. Kukenthal und Krumbach, Handbuch der Z oologie, 2_, T heil 2, L ief 12 u. 13: 1-212. Herman, C. O., 1964. The reproductive and developmental biology of the ophelid polychaete, Armandia brevis 'Moore'. Unpublished Master's Thesis University of Washington, Seattle, 131p. Hessle, C., 1917. Zur Kenntnis der terebellomorphen Polychaeten. Zool. Bidr. Uppsula. 5: 39-258. 156 Kennedy,G. Y. , and R. P. Dales., 1953. The Function of the Heart-Body in Polychaetes. J. Mar. Biol. Ass. U. K. 37: 15-31 K in b erg , J ., 1867. Annulata nova. Oefv. Vet. Akad. Stockholm, Forh. 23: 337-357. Malmgren, A. J ., 1866. Nordiska Hafs-Annulater. Oefv. K. Vetensk. Akad. Stockholm, Forh. _22: 355-410. Meyer, A. , 1912. Die Amphicteniden, Ampharetiden und Terebelliden der Nord-und Ostsee. Inaug. Dissertation. Kongl. Christian-Albrechts Univ., Kiel, 1-68. Meyer, E. , 1887. Studien uber den Korperbau der Anneliden. Zool. Sta. Neapel, Mitt. 7i 592-741. Newel1, I. E. and E. W. Baxter., 1936. On the nature of the free cell border of certain mid-gut epithelia. Quart. J. Micr. Sci. 79: 123-150. N ils s o n , D., 1912. Beitrage zur Kenntnis des Nerven- systems der Polychaeten. Zool. Bidr. Uppsala 1: 85-161. 157 Nyholra, K. 1950. Cantributions to the Life-History of the Ampharetid, Melinna cristata.Zool. Bidr. Uppsala. 29:79-91. Okuda, S., 1947. On an ampharetid worm Schistocomus sovjeticus Annekova, with some notes on its larval development. Jour. Faculty. Sci. Hokkaido. Imp. Univ., ser. vi, 9^: 321-329. Ostrooumouv, A., 1899. Note sur la larve de la Hypania invalida Grube. Annuaire du Musee. Zool. de L1 Ac. Imp. des Sc. de S t. P ete rsb o u rg . 4:350. Pettibone, M. H., 1953. A new species of polychaete worm of the family Ampharetidae from Massachusetts. Jour. Wash. Acad. Sci. 43 (11): 384-386. Quatrefages, A., 1865. Histoire naturelle des Anneles marina et d'eau douce. Annelides et Gephyriens. Paris, Libr. Encycl. de Roret. 1_: 1-588. Spencer, F. M., and L. S. Monroe., 1961. The color atlas of intestinal parasites. Charles C. Thomas, Springfield, 111. 150 p. Spooner, G. M. and H. B. Moore., 1940. The ecology of the Tamar estuary. VI. An account of the macrofauna of 158 the intertidal muds. J. Mar. Biol. Assoc. U. K. 24: 283-330. S u tto n , M F ., 1957. The feeding mechanism, functional morphology, and histology of the alimentary canal of Terebella lapidaria L. (Polychaeta). Proc. Zool. Soc. Lond. 129: 487-523. T horson, i. , 1946. Reproduction and larval development of Danish marine bottom invertebrates. Medd. Koma. Denmarks Fisk.-HavunderS'/gelser . Ser . : Plankton, 4 (1): 1-523. Ullman, A and C. G. Bookhout., 1949. The histology of the digestive tract of Clymenella torquata (Leidy). Jour. Morph. Phila. 84_ (1): 31-56. U schakov, P. V., 1955. £The polychaete worms of the far eastern seas of the U. S. S. R.J Tabl. anal. Faune.U. R. S. S. 56: 1-445. (In Russian). Wesenberg Lund, E., 1934. A viviparous brackish-water ampharetid Alkmaria roni jni Horst from Ringk^bing Fjord. Vidensk. Medd. naturh. Foren. Kjpb. 98: 215-222. 159 Wiren, A., 1885. Om zirkulations och digestions-organen hos Annelider af familjerna Ampharetidae, Terebellidae, och Amphictenidae. K. Svensk. Vetensk. Akad. Handl. 21: 1-58. 160 APPENDIX FORMULARY OF STAINS AND PROCEDURES Bouin1s Duboscq Fixative (Gurr, 1962) P ro c e d u re : . Fix 12-34 hours Wash in 70% alcohol* for 24 hours Store if desired in 70% alcohol Bela Haller's Fluid (Guthrie and Anderson, 1959) Recipe: 50 ml. distilled water 25 ml. glacial acetic acid 25 ml. g ly c e rin P ro ced u re: Add a drop of the above mixture to a piece of t i s s u e . Remove f l u i d a f t e r 30-60 seconds. Add a drop of 0.5% aqueous methyl violet. After 2 minutes remove the stain and add a drop of sea w a te r. Put on a coverslip and tap gently. Alcian Blue (Gurr, 1962) R ecipe: 1% aqueous Procedure (Modified from Gurr, 1962) Fix in Bouin's a Duboscq and embed in Paraplast. Fix sections to slides and carry through to * Alcohol refers exclusively to ethyl alcohol 161 distilled water as usual.** Stain 15-45 minutes Dehydrate through the usual series of alcohols** Mount in Piccolite WW 85 Aldehyde Fuchsin counterstained with Halmi's mixture (Gurr, 1962) Recipe (Halmi's mixture): Light green SF yellowish 0 .2 gm. Orange G 1 .0 gm. Chroraotrope 2R 0 .5 gm. Phosphotunsic acid 0 .5 gm. Glacial acetic acid 1 .0 m l. Distilled water 1 0 0 .0 ml. Procedure (Modified from Gurr, 1962) Fix in Bouin1s Duboscq and imbed in Paraplast. Fix sections to slides and carry through to distilled water as usual. Oxidize in Gomori's fluid (0.15 gm. KMn04 in 50 ml. of 0.2% H2s04) for 1 minute. Rinse in 2.5% sodium bisulfite for a few seconds. Rinse in distilled water. Place in 30, 50, and 70% alcohol, 1 minute each. Stain in aldehyde fuchsin 2-10 minutes. Wipe the back of the slide and rinse quickly in 95% alcohol. Place in 95% alcohol, 2-5 minutes, until no more stain comes out. Place in 70, and 80% alcohol, 1 minute each. Counterstain in Halmi1s mixture 20-30 seconds. Wipe back of slide and differentiate in acid alcohol (95% alcohol/0.2% acetic acid) for 2-3 m in u te s. Rinse in 95% alcohol. Place in 100% alcohol, two changes, 2 minutes each and into xylene. Mount in Piccolite WW85. **Usual r.e'...ods of attaching sections to slides, carrying slides to water, and dehydrating are included at the end of this appendix. 162 Azure A (Gurr, 1962) R ecipe: 0.1% in 30% alcohol. Procedure: Fix and carry through to water as above. Stain for 30 minutes. Rinse in 70% alcohol. Dehydrate in 100% or acetone. Clear in xylene. Diluted Foots (One part Foot's (1945) "single differential stain" and three parts 95% alcohol) P ro ce d u re : Fix and carry through to water as with aldehyde f u c h s in . Stain in Ehrlich's acid haematoxylin for 90 sec o n d s. Dehydrate through the usual series ox alcohols to 85% alcohol. Stain in diluted Foot's 90 seconds. Dehydrate in 95% alcohol, two changes, two minutes each; in 100% alcohol, two changes, three minutes each; and xylene, two changes, five minutes each. Mount in Piccolite WW 85. Gomori's Trichrome (Modified from Gurr, 1962) R ecipe: Chromotrope 2R 0 ., 6 gm. Fast green FCF 0 .,3 gm. Phosphotungsic acid 0 .,2 gm. Glacial acetic acid 1 . 0 m l. Distilled water 1 0 0 ..0 m l. Procedure: Fix and carry through to water as with aldehyde f u c h s in . 163 Stain in Ehrlich's acid haematoxylin 2 minutes. Wash in tap water 5 minutes. Stain in trichrome 5-10 minutes. Rinse in distilled water. Rinse in acid water (0.2% glacial acetic water or tv/o drops concentrated HC1 in 50 ml. of distilled water) 10 seconds. Rinse in 70% acid alcohol (0.2% glacial acetic acid) 10 seconds. Rinse in 95% acid alcohol 10 seconds. Dehydrate in 95% alcohol 10 seconds, 100% alcohol 3 minutes, and xylene 5 minutes. Mount in Piccolite WW 85. Haematoxylin (Ehrlich's acid)--Eosin (Gurr, 1962) Modified Iron Haematoxylin-Orange G (Spencer and Monroe, 1961) R e c ip e : Solution 1: Haematoxylin 10 gms. in 1000 ml. of absolute alcohol. Allow to remain in the light for one week. Solution 2: Ferrous ammonium sulfate 10 gm. F e rr ic ammonium s u l f a t e 10 gm. Concentrated HC1 10 ml. Distilled water 1000 ml. This solution will keep for approximately six months. For the working solution mix 15 ml. of each solution, It will last for about seven days. Procedure: Fix and carry through to 95% alcohol as with aldehyde fuchsin. 95% alcohol 5 minutes. 70% alcohol 5 minutes. Tap w ater 10 m inutes. Stain in working solution 4-5 minutes. Tap water 10 minutes. Stain in a 0.5% aqueous solution of orange G 3 m inutes. D ehydrate in 70%, 95%, and 100% fo r 5 minutes each. Xylene 10 minutes. Mount in Piccolite WW 85. P e rio d ic A c id -S c h iff R eaction (PAS) (Gurr, 1962) P ro c e d u re : Fix and carry through to water as with aldehyde fuchsin. Place in saliva, strained through cheese clo at 37°C for one hour. Running tap water 5 minutes. Hydrolize 7-8 minutes in solution A. S o lu tio n C 15 m inutes. Running tap water 5 minutes. Solution D, three changes, 2 minutes each. Running tap water 20-30 minutes. Dehydrate, clear, and mount as usual. Safranin 0 (Gurr, 1962) R ecipe: 1:50,000 aqueous solution. P ro c e d u re : Fix and carry through to water as with aldehyde fuchsin. Stain 15 minutes. D istilled water 1 minute. Dehydrate in 70% and 95% alcohol 10 seconds each, and in 100% alcohol 3 minutes. Xylene, two changes, 3 minutes each. Mount in Piccolite WW 85. Van Gieson's Stain (with haematoxylin) (Gurr, 1962) R ecip e: Acid fuchsin 0.1 gm. Picric acid, saturated, aqueous 100.0 ml. 165 P rocedure: Fix and carry through to water as with aldehyde f u c h s in . Stain in Ehrlich's acid haematoxylin 90 seconds. Tap w ater 5 m inutes. Stain in the above mixture 3-5 minutes. Rinse in tap water. Dehydrate in 95% alcohol 3 minutes, 100% alcohol 5 minutes, and two changes of xylene 3 minutes each. Mount in Piccolite WW 85. Fixing Sections to Slides: Individual sections or short ribbons are floated on a few drops of a solution which consists of two drops of album in mixed w ith 10 ml. o f d i s t i l l e d w a te r. The slide containing the solution and sections is then placed on a warming table at about 40° C. When the sections have properly flattened, the solution is drained from the slide and the sections blotted with a piece of filter paper soaked in distilled water. The slides containing the sections are then placed on a warming table to dry. Procedure for Carrying Sections to Water: Place slides containing sections in two changes of xylene, 10 min. each. Place in two changes of absolute alcohol, 3 min. each, and one change o f 95%, 85%, 75%, 50%, and 35% alcohol 5 minutes each. Procedure for Dehydrating Sections: The reverse process of the procedure for carrying sections to water. Method of Hermans (1964) for staining and mounting larvae of the ophelid polychaete Armandia brevis 'Moore' : The Bouin's was washed out in 70% alcohol. When the yellow color disappeared, a drop or two of 1% chlorazol black E in 70% alcohol was added to less than one ml. of 70% alcohol containing the specimens. The staining takes place rapidly. The tissue is then washed in 85% alcohol 166 and cleared by transferring it from 85% alcohol to clove oil or terpineol. Specimens were mounted in Piccolite WW 85. Specimens could be de-stained by adding a drop of pyridine to one ml. of 85% alcohol containing the specim ens.