STUDIES ON THE LIFE HISTORY OF
MESOSTEPHANUS KENTUCKIENSIS (CABLE, 1935)
COMB. NOV. (TREMATODA: CYATHOCOTYLIDAE)
’ DISSERTATION
Presented in Partial Fulfillment of the Requirements for the Degree Doctor of Philosophy in the Graduate School of The Ohio State University
By '
DONAL GENE MYER, B, Sc., M. Sc.
The Ohio State University . 1958
Approved by
Adviser1 MAM » tepartntent of Zoology and Entomology ACKNOWLEDGMENTS* e e
• * This study was made possible through the assistance of many of the faculty members, technical personnel, and graduate students in
.the Department of Zoology of The Ohio State University. The author wishes to thank Mr. Stanley S. Sroufe for his assistance in collect ing material., and Mr. Russell Williams for'his assistance .in preparing photomicrographs, Mr, George Phinney and Mr. David H,
Stansbery kindly helped collect and identify fish for the author,'
'Appreciation is expressed to Dr, Allen McIntosh of the Animal Dis ease and Parasite Research Division of the Agricultural Research
Service for the loan of slides from the United States National
Museum Helminthological Collection,
The author .wishes especially to acknowledge the many helpful • suggestion's and constructive criticisms given by his adviser,
Dr. Joseph N, Miller of the Department of Zoology and Entomology. SABLE OF CONTENTS t * * * fc INTRODUCTION...... 1
HISTORICAL REVIEW...... -...... 3
MATERIALS AND METHODS...... *...... 17
OBSERVATIONS AND RESULTS ......
Life Cycle of Mesostephanus kentuckiensis .
The First Intermediate Host.®...... 21
The Second Intermediate Hosto®.#..••••••••o o 23
The Definitive Host...,«...®...... o0.oo.•••••••••••.••• 2$
Comparison and Discussion® ••••••••••«« 28
Descriptions of Stages in the Life History "of.
Mesostephanus kentuckiensis
The Egg...... 3U
The Sporocysts...... 36
The Cercaria...... 3?
Redescription of the cercaria of Mesostephanus
kentuckiensis...... ho • » » Development of the excretory system of tha * * ' . * * * * # cercaria......
<6 Comparison ...... 1x1 « Activity of the cercaria.••«...... $0
Penetration of the°second intermediate host.,,..., j?l e The Metacercaria...... $3 0 o o o 0 'The Adult...... : 55 * e o , * Description 9 * kentuckiensis...... 55 * * * Comparison...,,.,*...... 57 Longevity of arlult worms ...... 58 DISCUSSIOK •'General Aspects of the Life Cycle...... _ 59 General Taxonomic Considerations...... 66 Some Taxonomic Relationships Within the Subfamily . Prohemistominae ...... 69 CLASSIFICATION...... '...... 76 ' SUMMARY...... 78 . LITERATURE CITED...... 81 PLATES...... 88 AUTOBIOGRAPHY...... 100 IV o O 0 o e 9 LIST OF TABLES • Table Page 1 Collection of G’oniobasis livescens from central Ohio 22 2 Fish from the Olentangy River examined for natural infections of metace-rcariae of Mesostephanus kentuckiensis...... 2h .3 Results of feeding experiments using one-day-old chicks as definitive hosts...... '...... 26 h Second intermediate hosts of species in the subfamily Prohemist.ominae ...... 30 and 31 v o TEXT ILLUSTRATIONS 0 0 Figure Page 1 The natural emergence of cercariae of Mesostephanus kentuckiensis from a single-snail during eight hour intervals starting at 8:00 A. M ...... 52 2 Location of the ventral sucker from the anterior end in relation to the length of the b o d y in Mesostephanus kentuckiensis from cercaria through the adult...... 75 o vi o APPENDIX ILLUSTRATIONS PLATE I* ...... !...... 89 Fig. 1 The egg -of M. kentuckiensis. Fig. 2 Daughter sporocyst of M. kentuckiensis containing • • germ balls and cercariae in various stages of development Fig. 3 Developing cercaria with three pairs of flame cells ' Fig. U Developing cercaria with six pairs of flame cells in body, two pairs in the tail* ' * Fig. 5> The cercaria of M. kentuckiensis. Dorsal view Fig. 6 Variations which occur in the canals anterior to the ascending trunks Fig. 7 View of posterior surface of furca showing-fureal flagellets ' PLATE II...... ;...... *....’...... 91 Fig. 1 The metacercaria of M. kentuckiensis. Ventral • • • view Fig. 2 Immature worm after four days in the chick. Ventral view o o Fig. 3 Adult of M. kentuckiensis. Dorsal view PLATE III...... 93 Fig.°l Longitudinal section through hindbody of adult 0 of M. kentuckiensis showing uterus and vaginal sphincter. X 3^0 0 o ° vii PISTE IV...... 95 • * o > Fig. 1 Cross section through the pharyngeal region of „ the adult of M. kentuckiensis. The cavity of the pharynx is flattened laterally. X 150 Fig. 2 Cross section through the acetabular region of the adult of M. kentuckiensis. X 15>0 PLATE V..:...... 97 Fig. 1 Cross section'through the region of the tribocy- • tic organ of.the adult of M. kentuckiensis • • showing the glandular tribocytic organ, .some vitellaria, and the intestinal-caeca. X 150 Fig. 2 Cross section through the region of the anterior . testis of the adult 'of M. kentuckiensis showing the ascending and descending portions of the uterus ventral to the testis. X 150 PLATE VI...... •...... 99 Fig. 1 Cross section through the region of the anterior • * testis and the ovary of the adult of M. . kentuckiensis." In addition, ventral to the testis is the anterior portion of the cirrus sac, and 0 ventral to the ovary is the ascending and descend ing limbs of the uterus. The ascending limb contains an egg. X 150 vxii O 0 0 Fig. Z Cross section through region of the posterior testis of the adult of M. kentuckiensis. Ventral to the testis are the muscular cirrus sac and the descending limb- of the uterus-. X 15>Q • INTRODUCTION* • . _ ... 'As a result of this investigation the life history of a cyathocotylid trematode, Mesostephanus kentuckiensis ('Cable,■ 1935) comb, nov, has been elucidated. This trematode, until now, has been known only from a larval stage, the cercaria, described by Cable. (1938) from the snail, Goniobasis semicarinata, in Kentucky* This study presented itself.while the writer was working on par.thenital stages of trematodes, Cercaria kentuckiensis was found to be common in one of the pr.osobranch snails, Goniobasis liveseens, of central Ohio, .Because of'the abundance of this snail and the cercariae, definite prerequisites for- life history work, this inves*? tigation was undertaken. Taxonomy in this family, as' in others of the trematodes, is ‘ based largely on the morphology of adults. Parasitism in general is associated with reduction and modifications.of many,structures - which, if they were present, might otherwise.be used to establish • relationships. For this' reason additional information obtained con* cerning the life history and larval stages of trematodes is invalu able in establishing relationships between genera and families. The relationship of the family'Cyathocotylidae to other families of the suborder Strigeata is not known despite‘the placing, of-this * , >*; family by La Rue (19!?7) and others along with various other families * • in the superfamily* Strigeoxdea. This is the first complete life history of any species of the genus Mesostephanus, and is therefore of special interest in regard to the systematies of the subfamily and family to which it belongs* '• 'V r», ^ Many of the published life histories of Cyathocotylidae are, indeed, A •incomplete, especially where descriptions of cercariae are con-' C*' 0> ?l 0 cernedo ' The relationships between genera within the' family are poorly understood, being complicated by incorrect.descriptions of the genera and species*. _• HISTORICAL REVTEW® • » • * i ■ Cercaria ‘kentuckiensis was first Reported by- Cable (1935), and ® * ■was later described % him (1938) from cercariae obtained from . '• Goniobasis semicarinata, a species of prosobranch snail which he ' collected from. Scaffold Cane Creek, Madison County, Kentucky* . It was again reported by Cable (1937) and by Anderson. (I9hh), in Goniobasis sp# and Goniobasis depygis respectively, from McCormicks Creek, Indiana® Banks (19h9) reported' finding the cercaria in • Goniobasis liveseens from Big Walnut Creek in central Ohio® It was again reported by Vernberg (1952) from Goniobasis liveseens collected from Clifty Creek in southern' Indiana® •; The first cyathoeotylid cercaria described was Cercaria vivax * Sonsino, 1892 from Cleopatra bulimoides in Egypt® Detailed descrip- ■' tion.s of this cercaria and the sporocysts were given by Looss (I896)® At present there have appeared in the literature descriptions of . approximately thirty-eight additional species of cyathoeotylid , .. cercariae® Many of these descriptions are inadequate®. f*i y. Various authors have reviewed-the literature, and have classi- . fied the known cyathoeotylid cercariae into various groups. Among the-more complete reviews are those'of Sewell *(1922), Szidat' (19-33), ^Cable (1938), .Goodman (1951), and Dubois (195l)« »The cercariae of trematodes .of the family Cyathocotylidae ® ... « ' ■? Poche, 1925, form a rather homogenous group !of furocercous cercariae ® <•) easily characterized by their distinctive excretory and prominent 0 © © 0 o * • . It & <*. * * • . digestive systems (Plate I, Pig., $ )• The excretory system,, consists • of a bladder with four ascending canals, two lateral* and two « •median#, ' * The median 9 * pair unite a short distance® anteriorly, forming a single median canal, which in turn bifurcates just posterior to « ,0 * . .* . . , . the esophagus where each branch connects with a lateral ascending * - * o 0 . ® ® e, canal. " Another canal emerging caudally from the bladder passes down , ^ ° _ o 0 0 * - ® ° the tail-stem and divides into two,branches which then traverse the 0 * * ° and Atkinson (!9!5>)o » The two new cercariae described by Sewell w§re ”, Co indicae XV from Melanoides tuberculatus and M* llneatus, and C, © o, ® indicae LVIII from Paludomus baeulus.■ Within the sporocysts of one •0 0 * of the Indian cercariae, £o indicae IV, Sewell reported the presence © ® ® of miracidia, a unique observation until recently (vide infra)# The following features are characteristic of the cercariae of-the Vivax , subgroups ' , ! * * s j * * * . • ‘ _ s * f (1) The acetabulum is extremely-small aiid rudimentary. (2) The fureal rami are provided with fin-folds that •extent the whole length of the margins, and are continous around the distal end, (3) The excretory system is very highly developed, and there are twelve pairs of flame-cells in the body # * and three pairs in the tails so that the excretory formula is 2 X 12 (X 3) X 1 = 2b (+6) flame ceUs. 0 0 ° s C. vivax.was again reported by Sonsino in 189 b from Tunisia, in two species of snails, Cleopatra bulimoides and Melanopsis praemorsa. e © © ® © © © © © * 5 * . * ^ '* •• > ... -s; This cercaria was studied by Langeron (I92hj, who concluded that it' was distinct' from C# vivax Sonsino, 1892, which was'reported o from Egypt;’ Since Langeron*s report, the life histories of both of co ' 0 a a ° © * o a ° these cercariae have been demonstrated; ®° ° g 0 * # E *> ’ • • 0 “ 0 s Azlm (1933) showed that C« vivax Sonsino, 1892 encysted1 as a. . • so®"* "" * 0 ® j ! © ’•When the metacercarias were fed to dogs, mature adult worms were ® * © 0 © ® • * 0 „ fi‘ ® « s ® o obtained#* These were identical to * Prohemlstomum splnulosum Odhner, © « ® ® * s ,1913 from the. common Egyptian kite, Mllvus migrans aegyptius; This * ® ' 0 0 0 . 8 © r ® 0 ® . trematode, then, is Prohemistomum vivax (Sonsino, 1892) Azim, 1933#s Through the-work of Joyeaux and Baer (193^, 19^1) it was * 0 ; 8 % s * ® ® .determined 'that C 0 vivax Sonsino, 189U encysts naturally in the ’"** " .... ® 0 0 e ®’ . frog Rana e'sculenta ridibunda and the fish Astatotilapia ® . • ' 0 # '' e desfontainesi, becoming mature in experimental infection’s of the ® " 0 s © ro 0 © 0 it) . snake.Tropidonotus natrix persa# These authors considered the . cercaria described by Callot (1938) from, Melanopsis pseudoferussaci * a © o by Tubangui (1928) from the snail Ampullaria lagunaensis in the o © * Phillippine-Islands was also assigned to this group* From two species of snails in Germany, Faludina vivipara and P* fasciata, ■ Peterson (1932) described briefly a cercaria which he called Furcocercaria No. U, but neither mentioned nor illustrated flame cells in the cercaria* Lutz (1933) described two cyathoeotylid, • „ ° ° ,.i 0- ' (., ® cercariae from Brazil, which he stated were similar to"Cercaria 9 ®9 •, ' ’ 0 . * ® ® o ’ vivax* These he designated as•Dieranocerearia utriculata from " ® © ® f the marine snail Cerithium stratum and Die ran o c e r e aria conchleula * * from a clam (Unionidae). Later, Lutz (1935) expressed the opinion that C* utriculata was the larva of one of'two species of ° e0e » ® ® “ * » * °s , Mesostephanus found in sea birds in Brazil* * Another cercaria, the, ® ”» "Cercaria vivax1* of Wesenberg-Lung (193U), from Bithynia ? 0 9 * ® ° , 0 * tentaculata in Denmark, "probably belonging to the Vivax group is » most likely a different- species from that reported by’Sonsino* Weseriberg-Lund also reported two other cyathoeotylid cercariae,) e ’ ° o , ® ® ® © 0 Cercaria No. h Peterson and Cercaria sp*, both from the snail e ® ® o s Bithynia tentaculata.0 In the former, he states that no flame cells ...... ■ ■ — * — «■ ■ tim © * * f,) ® were visible^ and in the latter, only one-pair in the tail-stern* - ® .* © ^ 0 ® ' * Cable (1938). included the subgroups’Leptoderma and Vivipara is _ in the Vivax subgroup as emended by himself at that time, thus including in one subgroup all cyathoeotylid cercariae with six • ® ♦- ■#: flame cells' (three pairs) in the tail-stem* The Leptoderma sub- * ® • group was established by Faust to contain C. leptoderma Faust, 1922 © * ® 01 found in Viviparus quadratus in China. This subgroup was characterized by being brevifurcate instead of longi^urcate as were © 0 • * ‘ - T 7 „ 0 rt* © ‘,J all the others then described* The Vivipara subgroup was erected “ „ ** . 0 "* ° 0 O o« " by Szidat (1933). to contain the cercaria of Linstowiella viviparae (Linstow, 1877).from the snail Vivipara vivipara in Prussia* This ® « snail served also as the second intermediate.host, the'definitive «„a * <«, <*• « ® jj. * host being the tern, Sterna paradisea, .The new .subgroup was’charac terized by the'absence of fin-folds on the furcae and by an excre- r " * » v ," 8 * * * - tory pattern of nine flame cells in' the body and six in the tail* »0 Szidat later (1936) stated that he considered the inadequately •described cercaria, Furcocercaria’-No®* U Peterson, 1932, from the * * '«* ,* * © ° m m t © it * * * ' ■* *' ® same snail host, to be identical to that of Linstowiella viviparae, * .In the same paper Szidat (1933) described two other new $) »• ► * ® ® f 1 • ,* « t * c ffi Si species, C, curonensis and C0 balthica from Bithynia tentaculata, . « which he assigned to the Tetis. subgroup of Sewell (1922)® This sub group, until then, contained only C» indica XXXIII Sewell, 1922 from Paludomus baeulus in India® » It was characterized by furcae * .without fin-folds, no acetabulum, and a flame Cell pattern of five pairs in the body and-two pairs in the tail® The life cycle ' • «, © * * © * ® ® t, ^ * * ® * ® * of Cyathocotyloides curonensis (Szidat, 1933) (i.e®, Cercaria curonensis) was determined’experimentally by Szidat (1936)® In his report he showed 'that leucasplus delineatus served as the. ® © * second, intermediate host while the domestic duck, Anus platyrhyncha, ® © ‘ served as the definitive host® He also proposed the subgroup Tauiana to contain Cercaria tauiana Faust, 1930 from Bithynia ® a j . ® fushsiana in China® This is a cercaria in which there are six pairs 1 m ;» © of flame cells in the body and none in the tail. Faust had not • © e o ® «• ® assigned this cercaria to any group, refering to0it only as a © © O © strigeid or holostomate fluke* o ® „ s ® p ® ' 0 • • • «• © B * ® © m ® ^ > ® “Dubois (1938) considered cercariae of the subgroups ..‘Vivax • 0 ’» s® ® and Vivipara to be larvae of trematodes of the" subfamily1 ‘f . % s a " . 0 <9 © ® o » ® Prohemistominaej‘while those of, the subgroup Tetis were'larvae of members of the subfamily CyathoOotylin'ae* These, considerations • * , * * *' * ■ • *' • were based upon the knowledge of one or more life histories known » , * *■ i n ‘these groups* Because of the inclusion of Cyathocotyle gravieri * » * * • Mathias, .193!? in the later subfamily, he concluded'that the inade- • quately 'described cercaria of this fluke must belong to the Tetis % * ■* 'i s * * * ; group* ' This fluke was-obtained by Mathias (1933) from a domestic duck which had been fed fish which'in turn had been exposed to *’ , fi. o ’ * • ’ 0 cercariae from the.snail Bithynia tentaculata in France*• Con- cerning the ‘two* remaining‘ monotypic ' subgroups, * Leptoderma and . * *> * .■(, * <« ^ , » Tauiana, Dubois stated that nothing of, their systematic position , ' * **■ * . 0 ® ® 0 a ^ « ® t * could be determined* 0 ® ® ® «* Porter (1938) reported another species, C* theodoxa from the * 4' 0 ® L ® ® * * ■ * * g, snail Neritina natalensis in South Africa, as a member of the’ *. ‘ ' ® „« Tetis subgroup* , ^ s a • ® s *" * r" r ^ 0 • Komiya (19,39) reported the life history of the cyathoeotylid, •i> ’ ® Paracoenogonimus ovatus (Katsurada, I9lh)j however, he did not <<■ •*; r$> assign its cercaria to any group*, The natural first intermediate * ** ® ® © ©' - © © ® host in Germany was Vivipara viviparus*, The natural second inter mediate hosts we^e various species of fresh-water fish* Experi«f e mentally infected mice seyved as the definitive host*0 ® 0 © 0 © 0 0 The cercaria and life©history of Cyathocotyle oriental!s © • Faust, 1922 was reported by Yamaguti (19h0). He found the gastropod Bulimug.■-> striatulus japonicus to be the natural first intermediate host, a species of fresh-water fish, Pseudorasbora parva as an experimental second intermediate host, and Milvus mlgrans lineat us as the experimental definitive host in' Japana He did not compare the cercaria to other related ,ohes0 .. .Johnston and Angel.(19h0) reported -the first cyathoeotylid * » cercaria from Australiao 'This was Cercaria tatei from the snail , Plotiopsls tatei. ' They pointed put the relationships of this. • cercaria to members of the subgroups Vivax and Vivipara, and' mentioned that the cercaria penetrates and encysts-in the fish Gambusia affinis#. * * „ " b 4 * • • • 0 Tang (I9hl) reported from China the life history of'. Prosostephanus industrius (Tubangui, 1922)' Lutz,- 1935. This ,trema»‘ tode, which was obtained'as natural infections of many domestic . * mammals of the area, was found to infect the snails Parafq s sarulus * eximius and P. striatulus as first intermediate hosts while many * ® ® it. ® species of fresh-water fish served, as second intermediate hosts.s Tang also gave the first description of a miracidium (obtained <9 * from eggs) of a fluke of this family# He assigned the.cercaria to : 0 a 0 . the Vivax subgroup. 8 % j s ® ea . * • . , s s Anderson (19hh) described,Cercaria szldati.from Campeloma rufum ® 0 0 * @ ® in Indiana. This was the second cyathoeotylid cercaria to be a s 0 © q ® 0 0 , ® ® ® eS ^9 reported in ;North America., The life history of this worm,) e a ® ® ® 0 Paracoenogonimus szidati (Anderson, 19UU) Dubois, 1951, was reported ® • ® ® by Anderson^and Cable® (1950 )• ®They found that the .fish Notropis cornutus harbored the metacercaria. *They obtained adults® & by * ' © • • , • 10 • *o <•> experimentally feeding metaeercaria to chicks. Because the cercaria G ® 0 0 0 G ° possessed characteristics of both the Vivax and Tetis subgroups, ;• - & O : ® e ®0 Anderson proposed combining the two into one subgroup, the VivaX ‘® « «, 0 Q W 0° °« * o Q subgroup. * * B, 55 ® s J . ' „ * ' . . 9 * .. * ° •C. notopalae was described by Johnston and Beckwith (I9h?) 0 ■ from the mollusc, Notopala hanleyi, in Australia. This snail also • serves as host for'the metacercaria. They pointed out the relation ships of this cercaria to those of the Vivipara'subgroup. & « ' * '.« * f * ® * « -'An unnamed cercaria from-the marine - snail Cerithidea * .Californica was described by Maxon and Pequegnat (1?U9) from'the ’ •. * coast of California. This cercaria has characters of both .the - * 'Tetis and Vivax subgroups. Maxon and Pequegnat concurred with Anderson's suggestion that- the groups be combined.* » *' . Goodman (19^1) described C* yankapinensis from Viviparus ’ intertextus- in Tennessee,,-the first,cercaria of the Tetis sub group in North America. Another'North Americari cercaria, .that of . Prohemlstomum chandleri, was reported by Vernberg (19$2)0 She* '* ® 9 a ■* described it as the Vivax type.0 The fish Huro salmoides and ® ® ® Micropterus dolomieui served as second intermediate'hosts, both © ® © naturally and 'experimentally. She obtained immature flukes . 0 • 0 3 ® . ® S) ® ©0 0 * • experimentally in Natrix sipedon. a ° 0 J”» ®* . . *- " * 1 0 © ® » * * * ® ® 0 * - Pubois (19!?1) redefined the Vivax subgroup and proposed a /i, ® new subgroupf Novena., The Vivax subgroup was emended to include 0 ® * cyathoeotylid cercariae with the excretory formula of • • • ® • © * » 2 [(3+3+3) + (3+3+(3))l " 36, i.e., those in-the previous sub- O groups Vivax, Vivipara, and Leptoderma. He further subdivided this 0 © e ® ® • e © a o 8 • • •• 8 . 0 ; into five groups, (a) through (e), based upon the extent of develop ment of the fin-folds,; length of the furcae, and presence or absence of a ventral sucker.. The new subgroup Novena which was established to contain the cercaria of Cyathocotyle orientalis and that of Cyathocotyle prussica (synonym C0 graviri) was characterized by .cercariae with an excretory formula 2. [(3+3) + ,(3+(3))J ® 2U, long furcae without fin-folds, and no ventral sucker0 The sub groups Tetis and Tauiana were retained. .According to Dubois, all members of the -supersubfamily Prohemistomatidi have cercariae of the Vivax type, and those of’the subfamily Cyathocotylinae have cer cariae of the Novena or Tetis types0 ‘Nothing was said concerning. • • the Tauiana type® . ^ „ * ,* , •• *’ From the Belgian Congo, Fain (1953) reported two new species occurring in the snail- 14ela.noides tuberculata. ’ .Both of these, C« kasenyi ahd’C* schoutederii, while appearing to be of the Tetis subgroup, were referred to only as pharyngeal longifurcate'mono- * * * t ® , * * • „ • •» • • • • 9 • *, • stomes. , * s * ® 9 9 »s . » » . . . - * * 9 ® ® * * ® © “ „ * * « # "J & ® -« * the snail Melanoides tub’ereulatus in India. This-cercaria* un assigned by the author, to any subgroup, is produced in sporocysts* .. some of which produce miracidiai* Cercaria multiplidata thus ® © ® ® * * * ® ® o © furnishes the second reference to sporocysts producing miracidia# ■* * ® 0 ® ® 9 * ® From marine snails in the Caribbean, Cable -(3.956 5 reported a ' % ® ! ° © 0 ° # twoa new cyathoeotylid cercariaeof the Vivax subgroup,.. These were o ‘ 0‘ * • • • • * & • C . caribbea S* and carlbbea LI, both found in Cerithium alglcola, 0 © ® ° - & and the latter also recovered0in Cerithium literatunU * The genus* Mesostephanus was .erected 'by Lutz (1935) for the reception’of certain species of cyathocotylids formerly assigned to the genus Prohemistomum© These were M© fajardensis ’(Price, 193k); * ■- ’ ,s>" S> I’’ . ® ' ■ ' '• M. appendiculatoides (Price, 193k); M* appendieulatus (Ciurea,. 1916) r m © ?1 * M. infecundus Lutz, 1935; and M© odhneri- (Trayassos,‘l93k)® He., : 7 • "* *« *. • o *. * characterized the genus as follows: ® e 'e p CO <0 ■>, * * ■* e Mesostephanuso Vorderkorper einfach, zungenformig Seiten- ' rander meistens nicht eingeschlagen© Dptterstoche kranzformig, • , mit dem Vorderende das Acetabulum kaum-erreichend0 (Simple * linguiform anterior body® Lateral-margins not mostly folded' _“ downwards, Vitellaria wreath-shaped and hardly reaching the ventral sucker anteriorly©) * , ' • • * * s'* Szidat (1936) accepted the genus, presented other character-' ‘ istics of it,’ and designated Mo fajardensis (M», jprolificus of Lutz) as the type© Both. Szidat (1936) and, later Dubois (1938# ♦ « 4 5 * . ‘ \ . 19UU, 1951) list Mo infecundus Lutz, 1935 as a-species o f ’ - Mesostephanus, but this species was neither-described nor illus- ® * «. * ® « * * * i' ...-trated by Lutz, who referred to. it only in making comparisons ** • - » « „ * • . with M© prolificus Lutz, 1935o Dubois does not include it in a later work (1953)® „ „ • * , ° 9 •, “ *. .* . . • • • •• \ * * • . Other species assigned to the genus Mesostephanus are M® milvi, by Yamaguti (1939); M© fregatus and M© haliasturus by . * D * ® .,m m » * r . • © ® ® © 9 Tubangui and Masilungan (I9kl); and M, burmanicus by, Chatterji ® ’ ® ® ® * * ’ (19I1O)© The latter species was transferred to the genus Gogatea © , © ® - - 111 ® © q © by Mehra (I9k7) who* also assigned to the genus Mesostephanus a. new .species, M© indicum© Chandler* (1950) "described a new -species, * # M, longisaccus from a dog: In the tJnited States# The latest species a „ ® (f . 3 * assigned to this genus is A tiicrgbursa, described by Caballero, Grocott, and Zerecero (1953)• Dubois (1951) reviewed and redefined the genus Mesostephanus, • removing several species previously assigned.'to it* He transferred M. odhneri (Travasso, l92li) Lutz, 1935 back to Prohemistomum odhneri where it had been placed originally by Travasso* The genus Mesostephanoides was.established by .Dubois (1951) to contain the • . worm originally described as Mesostephanus burmanicus Chatterji, '• 19U0, which Mehra had later assigned to the genus Gogatea* Five . other species that had been-assigned to the genus Mesostephanus were transferred by Dubois.-to the genus Prohemistomum,' two of the * • species being declared.synonyms .of -two other species* Mesostephanus milvi Yamaguti, 1939-became Po milvi (Yamaguti, 1939)9 'with M* ‘indicus Vidvarthi, 19 W nee'Mehra, 19h7 a synonym* -Mo fregatus Tubangui and Masilungan, 19^1'became P* fregatus (Tubangui and ' Masilungan, l9til)* M* 'indieum Mehra, 2$h7 became P* indlcum , (Mehra, 19U7) with M® lutzi Vidyarthi, 19^8 a* synonym* * ,* Up to'the present, the-genus Mesostephanus has contained* * seven species," the ' six listed by Dubois8 §(1953) ^ \ in the supplement 0 ® . " • * r. 5 ® • « e to his monograph and the one described by Caballero, Grocott, and Zerecero ’(1953)o These species have been reported from birds of the orders Falconiformes and Pelecaniformes,- and from mammals of"the " * * „ ® © . © order Carnivora* The species of the genus Mesostephanus apparently 0 ’ ,« are rather widespread,* being reported from Europe, North America, 0 ... >:> Central America, South America, the Phillippine Islands, and Puerto © * * # ft °Rico0 The fecognized species of Mesostephanus are* o O 00 o o o 0 o . . 0 0 * iu M.' fa’jardensis (Price, 193k) Lutz, 1935 (Type) _ ‘ Synonyms? Prohemistomum fajardensis Price, 193k ■. Mesostephanus prolificus Lutz, 1935 Hosts; Sula leucogastra (Type), Sula sula Geographic distribution? Puerto Rico, Brazil References; .Price (193k), Lutz (1935), Dubois (1938) M. appendiculatoides (Price, 193U) Lutz, 1935 Synonym: Prohemistomum appendiculatoides Price, 193U Hosts: Pelecanus occidentalis (Type) •’ P. o. californicus Geographic distribution: Dominican Republic, Panama • • (Canal Zone) , . References: Price (193h)> Caballero, Grocott, and Zerecero (1953) . * M® appendlculatus (Ciurea, 1916) Lutz, 1935 " „V* * Synonym: Prohemistomum appendlculatus Ciurea, 1916 Hosts: Canls familiaris and Felis tiomestica , experimentally (Types] "1" " " y . Geographic distribution: Rumania, U0 S ®‘, Ukraine • © References: Cuirea (1916), Price (1928), Prendel (193^)# Dubois.(1938) ® ® ® ® 0 a * g 0 . ’• M. cubaensis Alegret, I9hl »G e s 0 V »*” a8’‘ S ° 8 0 a s ^ o s t s Phalocrocoraxauritus floridanus (Type) \ e J ' e ~ ~ ^ ° “* | Host: Canis familiaris (Type) Geographic distribution; U. S, (Texas) References: Chandler (1900), Anderson and Cable (1950), Dubois (1951) • M. microbursa Caballero, Grocott,. and Zerecero, 1953 Host; • Pelecanus•Occidentalis californicus (Type) Geographic distribution; Panama, Mexico . References; Caballero, Grocott, and Zerecero (1953) •Luts (1935). erected the' subfamily Prohemistominae for all, the species then belonging-to the genus Prohemistomum, which he split , into four genera,' .Prohemistomum and three newly' erected genera, Mesostephanus, Prosostephanus, and'GogateaB . He' erected this sub« . family on the basis of an elongate forebody as opposed to those forms with a short forebody0 The- latter forms he assigned to the subfamily Cyathocotylinae Muhling, 1898® Both subfamilies were . assigned to the family Cyathoc0tylidae Poche, 1925 of,the suborder Strigeata La Rue, 19260 Although some genera have been split off "from the subfamily Prohemistominae by various authors to form other © $ ® • subfamilies, the genus Mesostepha'nus has remained associated with • « * • ■■ 0 ® ^ ® ® 0 0 8 0 0®O the subfamily Prohemistominae <■>- ® 0® ® e ^ “ ® s s 0 0 „ Ten life histories*of members of the family Cyathocotylida@ 0 'e . ® e are known# Three of these are of species® ® of the subfamily ® 0 ® • . • • • 9 ' I Cyathocotylinae; Cyathocotyle prussica, £, oriental!s, &n<| 0 I * ' ® ® ® c 0 © ®» Holostephanus curonensis® The 'life history of Prosostephanus a * J . 0 industrius is the only one known in the subfamily prosostephaninae §, ® 5 .!> . • * 16,. , * The remaining six life histories are. of species in the subfamily Prohemistominae: Prohemistomum vivax, P. chandleri, Linstowiella viviparae, Paracoenogonimus ovatus, P. szidati, and Szidatia joyeuxi. Only two of these, Prohemistomum chandleri and Paracoenogonimus szidati, are found in the United.States. In both of these the adults have'been recovered in experimental infection only. No life histories are known for species of the genus Mesostephanus, although Lutz C1935) expressed the opinion that his Cercaria utriculata might be the larva of one of two species of Mesostephanus found in' sea birds in Brazil*. 0 0 MATERIALS AND METHODS Goniobasis snails, collected in the rivers and streams by hand or dip-net, were brought into the laboratory and placed individ ually in small glass jars .three-fourths filled with tap water. Cercariae shed 'from these snails were studied both living and fixed. Living material was examined unstained or stained with neutral red, nile blue sulfate, or Bismark brown*' Observations upon the excre- tory system were made when the cercariae were flattened considerably as a result, ’of evaporation of the water from under the cover slip* Mounting fluids for living cercariae were tap water, various physio logical salt solutions, or polyvinyl alcohol* Fixed cercariae were . • prepared by adding water containing the cercariae to an equal volume of boiling ten percent formalin* For measurements, these cercariae'were given no other treatment, being measured in. the re sulting five percent formalin* . • Sporocysts were obtained for observation and measurement by crushing the snail.shell and extracting the body of the snail* Living sporocysts were dissected from'the liver with dissecting needles and examined, using the same methods as for cercariae# In addition, whole snails were fixed, sectioned, stained, and mounted for study of details of the larval stages# Guppies, used as experimental second intermediate hosts, were purchased from a local, tropical fish hatchery® Young tadpoles were '* " .i, * '* *' obtained from a pond which had been found free of snails of the genus Goniobasis. * It also had been determined that none of the tadpoles from this pond were previously.infected with the meta- cercaria of the trematode* Adult Rana pipiens were purchased from a supplier in Vermont. Both fish and tadpoles were maintained in aquaria in .the laboratory; the former were fed crushed Purina Dog Ghow Checkers, the latter dried egg yolk. ' • . Fish examined as natural intermediate hosts were collected •by using either a six foot-seine with quarter inch mesh or a twenty foot seine with'half inch mesh. Small fish (under an inch and a half) were examined directly for cysts under a dissecting binocular microscope using transmitted light. larger fish were sliced into thin strips which then were examined in a similar manner. Cysts were' • rempved with dissecting needles for observations.and .measurements. Metacercariae. were released by breaking the cysts with number 0 insect pins, after which they were examined or fixed in the same ' manner as the cercariaeo ’ *' • ’ * •' • *' * * * ■ Water-snakes, chicks, mice, and raccoons were.used as experi mental definitive hosts© The chicks Were obtained from the Poultry Department of The Ohio State University©' They were always one- day-old chicks, obtained, directly, from the incubator® They were fed commercial chick starter ‘feed after being infected® The laboratory mice were fed Purina Dog Chow Checkers and kept in cages in the animal room of the basement of the Botany and Zoology Greenhouse. The, raccoons were iwosof a litter of five"obtained before weaning, * (!) ^ These were weaned to Purina Dog Chow Checkers and raw hamburger, ii, 19 • • and, therefore, were not infected with the trematode at the outset of the investigation. The snakes which were used were captured in the vicinity of Columbus. They were found to be uninfected after examination of their feces for. the eggs of the trematode. Various methods were employed in infecting the animals. Fish and tadpoles were exposed to cercariae by placing them for a period of twenty-four hours in an aquarium together with snails that were shedding cercariae. Penetration of the cercariae was observed by restricting these experimental animals to a .small area in a dish of . water. This was observed under a dissecting binocular microscope. • The snakes, .mice, and chicks were fed by holding open their jaws and forcing pieces of fish or tadpoles down the esophagus. The mice and chicks responded by swallowing such food, but in the case of the snakes, it was necessary to-work the food particle down the esophagus, to the stomach by massaging on the outside0 The raccoons were ex perimentally fed chopped pieces of infected intermediate host in raw. hamburger. • ' • ■ • Eggs of the adult cyathocotylid were obt'aine'd from the feces' •• of infected chicks kept in a box .with a wire-mesh bottom* Shallow ' pans of water, into which the feces fell-, were, placed below the wire floor. ' The feces then were suspended in tap water and allowed • to settle repeatedly until the supernatant fluid was clear0 The residue containing the eggs was examined under a dissecting ® . * binocular microscope, and the eggs were removed to*clear tap water * i ® _ , •• ■« ® ® :gi using a small bore pipette. In an attempt@to obtain miracidia, some © O eggs were incubated" in 0.5 percent formalin, others in tap water. Some eggs in each solution were aerated for an hour daily; others, not at all. To recover adult worms, the intestine of the experimentally infected animal was removed to a Petri-dish containing Ringer's, solution. It then was opened with scissors and examined under a dissecting binocular microscope. The worms were removed to a dish of clean Ringer’s solution and then fixed. Some were mounted as stained whole mounts; others, as stained serial sections. All stages obtained were fixed in either ten percent formalin, • L'avdowsky’.s formula of AFA, or Carnoy's fixing reagent. When for malin was’ used, it was heated to a temperature between 55>° and 70° C, before pipetting in the living material, 'All sections were’ cut at either six or ten microns and stained with either Semichon's stain, Delafield's hematoxylin, or Harris’s hematoxylin. The ■latter two were counterstained with eosin. Whole mounts were • stained with Semichon’s stain or Harris’s hematoxylin, Measurements were made either with an ocular micrometer scale or with the aid of a camera lucida. Drawings were made both free • hand and with the camera lucida® . . . . . OBSERVATIONS AND RESULTS . • * • ♦ • Life Cycle of Mesostephanus kentuckiensis • « The First Intermediate Host* After isolation of various. . • prosobranch snails in individual jars and later examination of the water in the jars for cercariae, it was determined that the snail Goniobasis livescens is probably the only natural first interme diate host in central Ohio for the cercaria of Mesostephanus kentuckiensiso An examination of 1880 individuals of this species of snail revealed that sixty-two (3»l8 percent) were infected with this cercaria* These snails were collected over a period of one. year* They represent collections in all months of the year except October, November and March (Table l)o / A few scattered collections of Goniobasis livescens from locations ‘ other-.than central Ohio were made0 Forty-seven snails were collected from the shore of Lost Ballast Island in Lake Erie during September; seventy were obtained' from Clear Fork Creek at Mohigan-State Park in Ohio during October; and three hundred and twenty-seven were collected from the White River near Munsie, Indiana in March* None of these snails were infected with this cercaria* In view of the -low percent of infection found in the central Ohio snails-,_ it is possible that not enough snails were collected from Lake Erie and from Clear Fork Creek .to preclude the * • possible occurrence of the cercaria of M. kentuckiensis in these * o areas• * • • . * TABLE 1 Collections of Goniobasis livescens from central Ohio Number. Number Percent Date ‘ Location- Examined Infected Infected 7/29 Olentangy•River - Lane Ave# 5h 2 • 3.7£ 8/11 Olentangy River -.Lane Ave# 59 2 3. h 8/13 • Big Walnut Creek - Rt, 33 58 0 0.0 7? CO Big Walnut Creek - Rt. 33: •' 202 • 1 ' / 0.5' 8/20 Olentangy River - Lane Ave# . 9' 1- .11.1. ■ 8/22> Rocky Fork Creek ' . - 159 7 M.S 8/25 Big Darby Creek - Rt# lOlj . 87 . 8 9.2 8/25 . Big Darby Creek - Darbydale . ' 70 0 . 0.0 ' 9/19 Olentangy River - Lane Ave#- 8U ’ .3 3.6 12/31 Big Darby Creek - Circleville' Waterworks . 35 2 ' 5.7 1/19 Olentangy River.- Fifth Ave• . "• • 71 •1 • l.U 2/23 Olentangy River => Fifth Ave# bk ' 1 . 2.3 fc/3 • Big Darby Creek - Rt0 lOlj ' 362 • ' 8 ' ’ ' 3.1 5/25 Olentangy River «=* Fifth Ave# • 132 8 1.9 S/25 Olentangy River « Lane' Ave# 22 h ’ 18.2 Olentangy River » Lane Ave® • 28 ' ‘ U.1 5/31 • ' 1 . ’ 6/7 ' • Big Darby Creek «•- Rt # 10l* ' ' , _ 1'iU • . • 8 7.0 7/3 •> Olentangy River - Lane'Ave® * 2 20.0 » ' 10 ** • The* Second .Intermediate Host. In the known life cycles of • • * 9 cyathocotylids, the second intermediate hosts have been* found to be either fish, amphibia,’or snails. Cable (1938) reported that Cercaria kentuckiensis encysted in several species of cyprinid fishes. Vernberg (19^2) found that while the cercariae penetrate several species of fish, they encyst only in a single species of darter, the name of which was not given. Common'guppies, Lebistes reticulatus, tadpoles of Raha clamitans and R. catesbiana, and adult specimens of R. pipiens were exposed to the cercaria of M. kentuckiensis in order to determine whether any of these animals would serve as experimental hosts. This controlled feeding of .ex perimental animals also made it possible to determine the time necessary for the metacercaria to. become infective; It was found, that cercariae readily penetrated and encysted as•metacercariae in all of the. animals exposed to them0 Metacercariae were found in the muscles of the fish and frog and in the muscles as well as in the fibrogelatinous tissue'of the tadpoles© ... * * Examination of fish and tadpoles from areas in which ahigh incidence of infected snail.s occurred revealed that manyspecies of ; fish and amphibians were infected naturally with the metacercariae of M. kentuckiensis & The cysts of t'his fluke are easily distin guished from cysts of other strigeids and from gasterostome larvae^ which also were common in the’fish examined, by their size, * subspherical shape, and lack of pigment cells surrounding the * cyst®, The results of a suryey of fish obtained from the e ,, P <*> % Olentangy River * * • Fish from the Olentangy River examined for natural infections of • '' metacercariae of Mesostephanus kentuckiensis , . Number Number Host Examined Infected Family Ictaluridae Ictalurus natalis - Yellow Bullhead . 1 • 1 . Family Cyprinidae Campostoma anomalum - Stoneroller Minnow ■ 3 3 Cyprinus carpio - Carp ' . 1 ‘ 1 Notemigonus crysoleucos Golden Shiner • 1 '■ o Notropis cornutus - Common Shiner ■ 2 ■ Notropis spilopterus - Spotfin Shiner. . . 2 ■■ 0 ■Pimephales notatus » Bluntnose Minnow 1 ' 1 Semotilus atromaculatus - Creek. Chub '• 1 ■ 1 Family Clupeidae • • Dorosoma cepedianum - Eastern Gizzardshad h ■' 0 • Family Centrarchid.ae • .Ambloplites rupestris » Northern Rockbasg 25 ■ ’/ - Lepomis macrochi rug «=• Northern Bluegill Sun- fish 12 . k „ . ’ Micropterus dolomieui » Smallmouth Blackbass 6 h 9 Pomoxis annularis - White Crappie ' 3 1 * ♦. • Family Percidae . Etheostoma caeruleum •- Rainbow Darter . 2 • 0 Etheostoma zonale - Banded Darter * * « 2 9 0 Percina caprodes - Logperch Darter . 1 0 “A single specimen of Micropterus dolomieui, the Smallmouth Blackbass, • • • < from Big Walnut 'Creek, located on U. Si Route 33 south of Columbus, Ohio, was found to be,infected with the metacercaria-of this trema— * tode. Tadpoles of .Rana clamitans and R. catesbiana from the latter stream were also found to be naturally infected. A tadpole of R. clamitans- from the Olentangy River.was heavily infected, : •. The Definitive Host. The cercaria of M. kentuckiensis most closely resembles the ' cercariae of Szidatia joyeuxi and Prohernis- tomum chandleri. The adult of S. joyeuxi occurs naturally in water snakes in Tunisia. Vemberg (19^2)'recovered immature specimens of P. chandleri three days after feeding xnetacercariae to a Natrix sipedon. Because of the above.relationships, water snakes collected' from along the banks of the Olentangy River were examined for cyathocotylids. Three Natrix sipedon and'two N. septemvittata’which' • were examined were found to be uninfected, • • A specimen' of Natrix sipedon which had been fed two small naturally infected Ambloplites rupestris was found .to be negative for flukes upon post mortum examination six days later0 Anderson and Cable (19^0) found that chicks which.had never received food could be infected with the eyathocotylid, Linstowiella szidati, but they •. were unable to infect chicks which had been previously fed. After the author determined that chicks would harbor Meso— ' stephanus kentuckiensis when fed naturally infected Ambloplites • * * rupestris, a number of other feedings were’undertaken in which natu- « 9 rally and experimentally infected fish and tadpoles were fed to chicks. © ® * © * The results of these feedings to the ehicks are tabulated8 in Table 3‘* o 0 O O© » « It 26 . •. * « • . ' TABLE 3 / ‘ Results of feeding experiments using one-day-old chicks as definitive hosts • ■ . • • -■ Age of Interval Host of Metacercaria after Feeding Worms Recovered Metacercaria (Days) (Days) -. ■ Immature Adult •> Ambloplites rupestris 9 ■ 2 ’ . 1 0 n in V• .h 6 . 0 i* it- ' f• ■h 0 0 U? • li . •>• • . . 5 - ■ 0 0 * * * tit nt • • • . • 10 O- ' 0 . tr in •> 9 10 - 0 0 Micropterus dolomieui • • ? * .» 9 0 0 11 nr• •? • 10 . 0 • 1 .Mi • - 10. 0 0 ' Lebistes reticulatus ' 1 2 . • • 6 0 0 .n * ll -. 12 • • -16 ' 0 0 ii .li 12 • -• ”• . 16 0 0 ■ II: ' ’ . lt - • • 2 2 . ■ . 6 . 0 0 Ill .11 •. 22 ■ '6 •. 0 ■ • • 0 III .- lit 26 - 8-| ••• 0 0 II . • li ." •28 h 1 . 0 II II . 29 • ‘ u . . . -3 : 0 ’•II’’-' iii . ’ • . - 29 / • - '5 ■5 0 II 11 ' 29 . 5 . . ‘ 3 ' -o ' II ' ii- - ■29 9 0 0 II 11 : • 2 9 - . :'10 • ‘ : 0 13 II .11 . ' • V 36 . ' 9 ' ' -• • •• 0 . 0 111 ii - - -.36 10 • • • 0 . 0 • III . . ir - 36 ■ • 2h 0 • 0# II ‘ • in 39 ; 12 ■ ■ • - 0 - 3 in • 39 . , .13 • ' ' O' 0-x-: Tadpole • of R • clamitans •• , h6 ’ / . 3 2 . 0 11 in. in -* ' I46 , » - 9 0 0 . . 11 11 11 ir " h6 - 10 0 2 11 • 111 • in if - • h6 ■ ‘ * ' * 17 '• 0 . 1 c * t & -*Eggs appeared ih the feces from? days, until 17 days after feeding the metacercariae« ,> ■M-B-Eggs appeared in the feces from ^ days tintil 1h days after feeding the metacercariae® m © © @ O © O G o o 0 9 * . • ’ - . 2? * Of a total of thirty chicks fed metacercariae seven .were in fected with, immature and seven others with adult cyathocotylids* By examining the chicks for flukes at varying periods of time after feeding them metacercariae, it was determined that this fluke matures and commences egg laying in from six to eight days* Fecal examina- • tion for. ova confirmed this observation, ova being found in the feces of three of the chicks on the seventh day after feeding them meta cercariae* . Feedings of experimentally infected guppies were conducted to . determine the age. at which the metacercariae in the guppy became infective* The evidence presented’here would indicate that meta cercariae are infective between the ages of twenty-six and twenty- eight days after becoming established in the guppy0 However, in view of the fact that only a single feeding was made with metacer caria that were twenty-six days old, the age of infectivity may be between twenty-two and twenty-eight days* ’ • • v After determining the approximate age that metacercariae . .become infective, additional feedings of infected guppies to rep- • tiles- and mammals were undertaken* A Matrix septemvittata was fed a • guppy with metacercariae that-were forty days old, and a N. sipedon was’fed a .tadpole with metacercariae that were eighty-five days old, » ,f Examination of both snakes after sixteen days revealed-that neither was infected, .Two laboratory mice were fed guppies containing metacercariae that were forty-six, days old. Both mice were uninfected when examined twelve days after feeding. Two rapeoons .were each fed an infected tadpole of R. clamitans. One of the tadpoles was # naturally infected, tjie other-had been experimentally infected ninety-seven days previously* Examination of the raccoons eight days after feeding revealed no cyathocotylid worms. Comparison and Discussion* All hut one of the cyathocotylid cercariae are. harbored by snails of the subclass -Prosobranchia, .. All the marine cyathocotylid cercariae'are found in snails of the family Cerithiidae0 The freshwater cyathocotylid cercariae infect • snails of six'or seven' other families. . The natural first intermediate hosts of the cercaria of ■ Mesostephanus kentuckiensis are. Goniobasis semicarinata, G. 'depygis, and G. livescens of the family Pleuroceridae0 No other cyathocotylid cercariae have been' described from snails of the genus Goniobasis. However, one other cyathocotylid cercaria has been described from a '• snail belonging -to the family Pleuroceridae. ' That one is the cercaria of Prohemistomum chandleri from the snail Pleurocera acuta. ■ • . Dicranocercaria conchicula described by Lutz (1933) as occurring • ’ in a unionid clam is the only cyathocotylid cercaria not found in a prosobranch. It is quite possible that-this poorly described cercaria is not a cyathocotylid, Dubois (1901) stated that the .cercaria of Cyathocotyle orientalis was harbored- at the expense of the pulmonate gastropod Bulimus striatulus japonicus, constituting a notable and unique exception among, the cyathocotylid larvae. According to*Baker (1928) and Fain (1953) snails of the genus * • Bulimus Scqpoli (syn. Bithynia Leach) belong to the family . Amnicolidae.s ’This family is one included in the subclass Prosobranchia & 0 * It is possible that®Dubois confused this genus with’another* © @ There have been five other species of cyathocotylid cercariae reported as occurring in more than one species of snail# Four of these have been reported in two species of snails of a single genus# The cercaria of Szidatia joyeuxi has been reported from six diff- ■ erent species of Melanopsis, . The cercaria of Mesostephanus kentuckiensis will penetrate, ; .encyst, and develop in many-species of fish and amphibians#. Only •one other cyathocotylid is known in which-the metacercariae occur in both fish and amphibians. The metacercaria of Szidatia joyeuxj- has been found in two different amphibians and five different fish# - In addition two species'of adult amphibia'were experimentally in- .fected by Joyeux and Baer (19Ul)o The second intermediate hosts of species in the'subfamily Prohemistominae whose life histories are known are listed in Table iio • ■ • The metacercaria of Mesostephanus kentuckiensis becomes infec tive for chicks after twenty-two to twenty-eight days encystment in the experimental second, intermediate host, Lebistes reticulatus# This interval agrees with the time required for maturation of the metacer- •cariae of those other cyathocotylids for which the interval for devel opment known# Azim (1933) stated that metamorphosis of the metacer- ' caria of Prohemistomum vivax continued for three weeks, but he did not r —1 " " - T • mention the age of the metacercariae which he had used in his experi- 9 ments. Komiya (1939) found that metacercaria of Paracoenogonimus , o ovatus that were twenty-eight days, thirty-five0 days, and one year old O 0 were infective,O while those A twenty-one days old were not. TABLE U Second intermediate hosts of species in the subfamily Prohemistominae © • Natural (N) &e * Species Second Intermediate Host Experimental (E) Author Linstowiellia viviparae . Vivipara vivipara N Szidat (1933) Paracoenogonimus ovatus Leuciscus runtilus N Katsurada (191B) o Blicca bjoerkna H ■ Katsurada (191ii) . o • Abramis brana . N . Katsurada (I93ii) Komiya • (1939) . Alburnus lucidus • ■ N - Katsurada (I91li) Komiya o ©0 * - (1939) Barbus fluviatilis . Katsurada (I9U 4.) o . Lucioperca sandra . • N • Katsurada'(19lB) • Rotfedern • ' . ‘ ‘H • Katsurada (191U) Komiya e (1939) ■ Paracoenogonimus szidati Notropis. cornutus N &'E Anderson & Cable (1950) Blue gill • E Anderson & Cable (1950) Prohemistomum vivax Gambusia affinis • E Azim (1933) • Tilopia nilotica E ® ® ’ Azim (1933) Prohemistomum chandleri Hur-o salmoides N & E . * Verriberg (1952) (2) ’ ■l""' ' "" ' ' ’ " " ' Micropterus dolomieui-• • N & E Vernberg (1952) e (Continued on Page 31) TABLE h (CONTINUED) , . Second intermediate hosts of species in the subfamily Prohemistominae ' Natural (-N) Species ' . • Second Intermediate Host ■' Experimental (E) Author Szidatia ^joyeuxi Bufo mauritanicus N Callot-(1936) Hyla arborea “ • E. • Joyeux & Baer (19Ul) Rana. esculenta N • Joyeux (1923) • N Callot (1936) 9 . Dollfus (1951) • Anguilla anguilla .to .Balozet (1953) Astatolilapia,desfontainesi Joyeux (1923) * 9 N ■ Langeron (192U) e Barbus antinorii “ ' : to 5 Callot (1936) * Gambusia affinis * , n ■ Callot (1936) ■ N ■ Balozet (1953) - Hemichronis bimaeulatus ' • • N Callot (1936) Mesostephanus kentuckiensis Rana catesbiana tadpole ■ ■ to E ' Present 9 • Rana clamitans tadpole to & E Present m Rana pipiens adult E Present ■ Lebistes reticulatus • E - Present * ' Cyprinid fishes ? . Cable (1938) » A darter ' E Anderson & Cable (1950) * Other fish (see TABLE 3) - N ■ Present •• 32 n According to Anderson and Cable (1950)j the metacercariae of Paracoenogonimus saidatl were fully developed at,three weeks* and were infective between twenty and twenty-five days as determined by feeding# The metacercariae'of Prohemistomum chandleri were presumed to be infective after three weeks by Vernberg (1952) since the primordia of adult structures ceased further development at '* that time#*- * ’• " * ' While ‘the natural definitive host or hosts of Mesostephanus kentuckiensis have not been disclosed, it was found that chicks will harbor the adult worms* Failure of the- snakes and raccoons to be- . ■ . . come infected would seem &© preclude the possibility of their being , natural definitive hosts* * There is the possibility that-the metacercariae fed to one of the snakes.and one of the raccoons were too old to.be infective# , . However, it is the author's-opinion that.this is .not the reason that , these animals did not acquire the infection* Komiya (1939) found,that -' the metacercaria of Paracoenogonimus ovatus were still infective after * one year® ■ The present author' examined metacercaria of Mesostephanus . kentuckiensis which had been'encysted for ninety days and found them « * to be similar.in all respects to-metacercaria thirty days old# Un fortunately, metacercaria older than 16 days were not fed to chicks# Dubois (1938, 1911, 1951,. 1953) is of the opinion that birds of the order Pelecaniformes are natural hosts of the adults of the genus; * Mes ostephanus # His reasoning is based upon the fact that four of the * • . • o six previously»known^species of Mesostephanus occur in Pelecaniforrftes, 0 * » * * 0 The other two are known only from dogs* Dubois (1951) refers to them as secondary parasites or erratics in the dog.., In experimentally fed chicks, ova of M. kentuckiensis first ' appeared after seven days# T.he time-between the feeding of the metacercariae and the. production of eggs by the adult worms seems to vary considerably among the different species of cyathocotylids# For Cyathocotyle orlentalis in ducks, Yamaguti (19h0) found ova after eighty hours# In a closely related species, £#, gravierl, Mathias (1935) reported .eggs in feces of ducks after three and a half days# These worms belong to the subfamily Cyathocotylinae#■ Three days after feeding metacercariae of Paracoenogonimus szidatl to chicks, Anderson and Cable (1950) recovered eggs in the feces of the chicks# • Katsurada (191U) recovered a specimen of Paracoenogonimus ovatus which contained a single egg four days, ■after feeding fish infected with the metacercariae to a mouse# Feeding metacercaria of the same species to mice, Komiya (1939)' found eggs in the feces after five to seven days0 Azim (1933) re ported ova of Prohemistomum vivax in the feces of mice seven days after feeding them the metacercariae0 Joyeux and Baer (193^, l9hl) recovered young immature worms of Szidatia joyeuxi, none of which contained eggs, nine and eleven days following the feeding of the metacercariae to the snake Tropidonatus viperinus♦ The latter instance is set apart from the others since the definitive host is o poikilothermic. * ° ® Descriptions of Stages in the Life History of Mesostephanus kentuckiensis (Cable, 1935) comb, nov. The Egg, The operculated eggs are oval, the opercular end being slightly more pointed (Plate I,. Fig, 1). The, color is light yellow. Twenty eggs recovered from feces of infected chicks had an average measurement of 102,5 by 67.3 microns, with a range of 9U.8 - 113.5 mi- crons by 61,8 - 7h,6 microns. The operculum averaged 20,8 by U,1 microns. Many of the eggs show a thickening of the egg shell ' opposite the opercular end. Eggs recovered from freshly deposited feces of chicks were filled with vitelline cells,, and each .contained an unsegmented ovum, usually located at the opercular end. These eggs cannot be distinguished from those of most of the other species of Mesostephanus nor from eggs of many other species of strigeate worms. Eggs were incubated in a variety of ways, none of which re sulted in the hatching of miracidia. Although eggs were observed daily for three weeks, it was apparent after the. first week that none were developing further. Most of the eggs showed rapid degenera tion of the yolk cells, In other .eggs the ovum never completed the first cleavage while a few others developed for a couple of days, after which the egg contents became vacuolated and no further develop- « jnent was observed, o >#\ <*• & 9 The eggs of two species of cyathocotylids have been success fully incubated with the resulting production of miracidia, Tang (l9Ul) incubated eggs of Prosostephanus industrius in water and recovered miracidia eighteen days later.' The miraeidium had a pair of pigmented eye spots, two* pairs of flame cells, and a body covered with four rows of ciliated epidermal plates* The plate formula from anterior to posterior was 6-8-1;-3* Anderson and Cable (1950) were successful in hatching a few miracidia of Paracoeno- gonimus szidati* They found that two pigmented eye spots appeared on the fifth day, the flame cells at eight to eleven days, and hatching occurred in eleven to’eighteen days* • The miracidia of two other cyathocotylids have'been described,, both occurring in sporocysts*. Sewell .(1922) first described this .’ phenomenon in sporocysts of Cercaria indica. XV, He found’miracidia .• in both mother and daughter sporocysts*. Premvati (1955) published the second account of miracidia produced in sporocysts* She.re- . ported three generations of sporocysts for C* multiplicata, of which the first generation or mother sporocyst produced only mira- cidia. Her results indicate that the second generation spor.ocysts, which came from the miracidia, produce both third generation ' sporocysts and miracidia, while the third generation sporocysts produce only cercariae* While the cercariae described by Sewell and by Premvati were quite different, the miracidia were very similar. Especially noticable is the fact that-the miracidia of both lack eye-spots. The miracidia.are otherwise similar to those of other cyathocotylids, all of.which have ciliated epidermal plates and two pairs of flame ceils. The Sporocysts» . Over half of the sixty-two snails that were shedding cercaria of M. kentuckiensis were crushed and examined for mother sporocysts# In none of these were mother sporocysts found# A possible explanation for this situation is that the mother sporo- cyst may/disintegrate or degenerate after releasing daughter sporo- cysts, leaving ho trace of itself by the-time cercariae are being produced. With this'in mind, sixty-three snails’collected after ’ • April 195>8 which were not shedding cercariae were crushed and . examined for sporocysts#■ In none of these were any parthenital stages of cyathocotylids found# Infected snails collected during the winter months did not shed .cercaria until as long as fourteen days after they were brought into, the laboratory# During the'warmer months of the year the cercaria are shed the first or second day# ‘ • • It is possible that mother sporocysts would have been recovered by’ examination of crushed snails immediately after their collection . .. during cold weather# ' There have been only four reports of mother sporocysts of . cyathocotylids being found# Looss (I896) described finding mother' sporocysts of C. vivax Sonsino, 18920 These were always found singly in the branchial chamber0 They were approximately mm# long by 0,1 mm# wide, ,and they had a subterminal birth pore# The mother sporocysts of C, indica XV were approximately 9*6‘mm. long, and were provided with a terminal birth pore# They were found in the tissues * f r * * in I92J4. found two "generations of sporocysts for Szidatia joyeuxl. The final report-of mother sporocysts is that of Premvati (1955) mentioned above* Thes'e mother sporocysts produce miracidia which are also similar to those of £* indica XV in that they also have a ter- •minal birth pore*- They are 2,62 to 5.55 mm, in length by 0,27 mm, in width, and they were' found in the mantle cavity of the snail* . Elongate, hollo;-; daughter sporocysts of Mesostephanus kentuckiensis were recovered from the. digestive gland and also the. tissues of the’ upper branchial cavity of infected snails* Twenty- one were dissected from one-snail. Ten living second generation sporocysts averaged 3o70 mm* long and-0,176 mm, wide (Plate I, Fig* 2) They are opaque and very motile when extracted from the snail tissues. The anterior end is provided with a solid tapered snout which is very, active. The cells packing the snout stain vividly with neutral red. The - subterminal birth pore in living individuals is located fifty-six to seventy-two microns from the anterior end. The posterior end is broadly rounded. Small hairs or flagellets, arising from papillae, are found scattered over all the spOrocyst , being most numerous at the anterior end. The flagellets are 3.U to U.8 microns .long. Occasionally, annulationis in the form of trans verse bands are present, usually starting posterior to the, birth pore.- - These a!re produced apparently as the result of contraction • « • ® of circular muscle bands in the sporocyst walls. The wall surrounding the cavity of th*e sporocysts is about 6,8 microns thick in living sporocysts* but only Iu8 micrpns in o ' ^ fixed sporocysts# The wall of the Sporocyst observed in optical section is composed of three layers#- The outermost layer is the cuticle which is about .0,68 to 1.2 microns ‘thick# The layer beneath the cuticle is about 3»U microns thick and appears to be made up" of longitudinal and circular fibers,.which possibly are muscle fiberso The.innermost layer is made up of parenchymal cells which form a very loose tissue layer# The cells forming the solid snout are similar to these lining the wall of the cavity# There are many refractile yellowish granules, which are approximately one micron in diameter, in the wall of the sporocyst, especially in those sporocysts removed from snails that-had been • kept in the laboratory for more than a few weeks. Flame cells are abundant and very large, being 13.6 microns in length. In young sporocysts, the flame cells appeared to be restricted .to-certain areas of the body.' These areas were easily located by the presence o.f rather hyaline, irregularly shaped bodies about fourteen to fifteen microns long and three to four microns wide which have the appearance of slivers of wood. In larger sporocysts with refractile ' * spherical granules, the sliver-shaped bodies are no longer present, and the flame cells are more abundant and have no detectable pattern of distribution. The body cavity of the second generation spor.ocyst is filled with germinal masses and cercariae in various stages of development. In sporocysts obtained from snails crushed within a few days after ® • O bdlng® collected, there ° are ° usually forty-five to fifty mature © cercariae which actively move about within the-sporocyst. ■*„ The brevity of or. lack of descriptions of the sporocysts of <>> , ‘other cyathocotylids makes comparison of those of M. kentuckiensis and other species impossible# Cyathocotylids with sporocysts that have a subterminal birth, pore are Prohemistomum vivax (Sonsino, 1892) as described by Looss (I896), Paracoenogonimus szidati as described by Anderson and Cable (1950), Prohemistomum' chandleri' described by Vernberg (1952), C„ kasenyi and.C. schoutedeni described by Fain (1953), and C„ caribbea LI described by Cable (1956)o A terminal birth pore has been reported for sporocysts of £. indica XV by Sewell (1922) and sporocysts of C, multiplicata by Premvati (1955)® A few•sporocysts have been described which have transverse muscle bands. ■ • ■ . Cercaria tatei and the cercaria of Prohemistomum chandleri are most like the’ cercaria of Mesost'ephanus kentuckiensis. The former was described by Johnston and Angel (19h0) as being produced by sporocysts with muscle bands, while the sporocysts of P. chandleri are described by Vernberg (1952) as lacking ’the bands. While many . fresh sporocysts of M. kentuckiensis exhibited these transverse bands, many others did not. The bands.are not apparent'in fixed material or in living material under pressure of a cover slip. For these reasons the bands may be of no importance in the identi- fication of "species. Cable (1938) and Vernberg (1952) did not report, them for C. kentuckiensis. The Cercaria. Measurements of cercariae are from eighteen G specimens killed in hot 10 percent formalin. The first figures are fr • ' * 4- * ■ ho the range in size, the figure in parenthesis is the average size. Redescription of the cercaria of Mesostephanus kentuckiensis (Plate I, Fig. 5)*- The cercariae are longifurcous, pharyngeate distomes with characteristics of the .Vivax subgroup of Sewell, 1922 '' as emended by Dubois (195l)« The subgroup is characterized by the formula of the excretory system: 2 [(3t3+3) ♦ (3+3+(3))] *36® . The cercaria of M. kentuckiensis belongs to the subsubgroup (e) of Dubois (1951), characterized'by long'furcae with fin-folds upon their distal half; ventral sucker present; and eight gland cells • in 'the oral sucker.' . . • _ . •The body is flattened dorso-ventrally, concave ventrally, and' . is oval to pyriform in shape; length 0.2h0 - 0.289 mm, (0.262 mm.), vridth 0.117 - 0.15U mm. (0.138 mm.)0 The body is spinose, except for the ventral surface in the region where the future tribocytic organ will form behind- the ventral sucker. Spination is heaviest at ■ the anterior end,- where the spines are thorn-shaped, and at the posterior end, where they are long and narrow. There is a brownish tinge to the lateral' and posterior portions of the body in many of •the cercaria observed. Fifteen flagellets which are about four microns long are visible along the lateral margins of each side. The tail is inserted dorsally; tail-stem length 0.323 - O.lUl-O mm. (0.386 mm.), width 0.05U - O.O69 mm. (0.060 mm.). The furcae are flattened laterally; measuring 0.258 - 0.316 mm. (0.283 mm.) in length, They are provided with dorsal and ventral fin-folds, which are continuous around their tips.. The furcal fin-fold extends about o c o e O A ® 9 . ° 6 • ■ . OO ® thirty microns beyond the tips of the furcae. The ventral fin-folds extend approximately two-thirds of the way up the furcae from the 'tip, while the dorsal fin-fold is slightly shorter* A -ridge of spines seems to be continuous with the furcal fin-folds for a. short ’ distance anteriorly* The surface of all the tail-stem and the'proxi- ’mal.one-fourth of the furcae is finely spinose* ’ • « * * . 1 'The lateral edges of the tail-stem have flagellets, approximately eighteen to twenty-three microns long, arranged in a definite pattern. There are thirty-eight to forty of these hair-like processes on each ' side. The anterior twenty on each.side.are rather far apart and occupy two-thirds of the tail-stem* They alternate dorsally.and ventrally along the margin of the tail-stem* The posterior twenty are arranged in pairs on each side* One of each pair is ventral, • the other, dorsal. Each flagellet arises from a granule in the cuti cle* Other flagellets are located on the-dorsal and ventral surfaces of the tail-stem. The pattern of these has not been determined since they are difficult to observe* There is a single flagellet located on the posterior surface near the -tip of each furca. This flagellet is about half the length of those on the tail-stem, and is located five to ten microns from .the tips of the furcae or thirty-five to forty microns from the tips of the furcal fin-folds. This flagellet is‘visible only when the • Tf, * • furcae are viewed from either a «dorsal or ventral position^ and not 9 ® after the lateral surfaces of the furcae are exposed to the pressure of the cover slip. Other/smaller flagellets, each of which emerges from a small papilla, are located on. the posterior'‘surface of the furcae where the latter* joins the fin-folds (Plate I, Fig..7).These can be observed only when the furcae are flattened by cover slip pressure in order to expose the flat surfaces of the furcae. The; number of these small furcal flagellets seems to’ vary between six-., teen and twenty on the dorsal edge and sixteen to .twenty-two on the ventral edge of each furca, The papillae from which these flagellets arise are about 1,5 microns in diameter, and the flagellets.are about 5*5 microns long. The- papillae are .not evenly distributed along • their course. Some of'them are paired, but there are usually no more than three pairs along an edge, dorsal.or ventralo The' second papillae and their flagellets are smaller than the others, and are . located more centrally on the furcae0 ' . The oral sucker which is very protrusible has a length of 0.061 » 0.068 mm. (0.06limm.) and a width of'Q.Oh? - O.OI48 mm, . * (0.0U7 mm.). There are at least eight gland cells located in the oral sucker. These open at the anterior end of the oral sucker, each through its own duct. In stained sections the contents of the ducts are eosinophilic while the glands are not. The short pre pharynx, nine microns or less in length, is followed by a muscular pharynxj length 0.022 - 0.02U mm. (0.023 mm.), width 0.019 - 0.021 mm.' (0.020 mm.). The esophagus is about twenty microns long. It leads to the prominent intestinal caeca which extend posteriorly to the , sides of the genital primordia. The caeca follow a sinous course posteriorly. The walls of. the caeca often appear crenateds. Within 9 ® the caeca are0found irregularly, shaped bodies which exhibit . birefringence when- viewed with polarized light. These free-floating bodies are plastic, changing their shape readily under stress. .Between the intestinal caeca are two bodies* The most anterior is the ventral sucker;'length 0.209 mm., width 0.220 mm. This structure is located 60/100 - 65/100 (62/100) of the length from the anterior end of the .body* Behind this rudimentary sucker is the second body, an irregularly shaped mass of cells, presumably the genital primor- . dium,. which stains strongly in fixed whole mounts. • . / The basic plan of the excretory system is typical of that of cercariae in the Vivax subgroup.From the excretory bladder arise • four ascending trunks. The two median ones fuse just anterior to-- . • the ventral sucker where they form a single, median trunk which bifur cates just’ posterior to the esophagus into right and left cross commissures;, These commissures connect with the lateral ascending trunks across the intestinal caeca. At the junction, of the lateral trunk and the cross commissure on each side is a canal which extends 'anteriorly. -This may-be unbranched or may be divided into an anterior median and an anterior lateral branch. In the latter case, the median anterior branches may end blindly at the level of the pharynx or unite with each other across the pharynx. (Plate I,' Figo 6 a-d). All'the excretory canals in the cercarial body.contain refractile granules except the paired median ascending canals and the lateral ascending canals posterior to where the collecting tubules enter. The granules * * • • * are approximately lo5 microns in diameter. The main collecting tubule ° o 6 on each side enters the lateral trunk just posterior to where the e anterior canal joins the lateral trunk and cross commissure* It divides shortly into anterior and posterior secondary collecting tubules. Each of these receive the capillaries of three groups of three flame cells. The flame cell formula is, therefore,- 2 £(3*3*3) + (3*3*(3))J 3 of. which the last three on each side are located in the tail-stern. Another excretory trunk emerging caudally-from the bladder ex- ■ tends posteriorly down the tail-stem# It divides into two branches which traverse the furcae and empty by pores on the posterior surface • of the furcae just proximal to their tips,' . The undivided portion of the caudal excretory trunk in.the tail-stem is suspended rather loosely and moves freely in the tail-stem. Surrounding t.he caudal trunk is a matrix of cells with large nuclei to which the flame cells and their capillaries are.attached. The three flame cells on each . .side are connected to each other by their capillaries from which a . '•single collecting tubule extends'anteriorly<> This is attached to the matrix surrounding,the caudal trunk0 'This collecting tubule enters the body and turns laterally at the level of. the Island of Cort, and finally joins the collecting tubuie of the posterior three flame cells • of the cercarial body. Caudal bodies are present, on each side of the • ‘caudal trunk. These are large and contain conspicuous nuclei in either. cercariae within the sporocysts or in freshly-emerged-cercariae. They are less obvious' or even may be completely degenerated in older cer- * ■ '. * * • ■ - cariae, * . . „ . * , >• « . % *' 9 8 Beneath the cuticle in the tail-stem 5are prominent nuclei which x * 8 form four masses, two lateral, % one dorsal and one tentral, each ® of which extend the length of the ,tail-stem. On each side of the caudal excretory canal are bands of muscle composed of prominent . oblique fibers. Examination of sectional material revea3..s the . presence of four muscle bands, two' dorsal and two ventral, attached to the inner surfaces of the lateral walls of the tail-stem. Each muscle band has dorsal and ventral flutes which are obliquely situated, giving the'appearance of muscle fibers extending postermesad •' Besides the gland cells occurring in the oral sucker, there are ill defined glands' in the body lateral.to the pharynx. These probably empty to the outside by ducts which extend through the ' oral sucker. In stained sections at least twenty-two ducts (eleven pairs) have been observed. Eight of these are from gland cells in the oral suckero The remaining fourteen may be those of the glands lateral to the pharynx.- Numerous unicellular, cutaneous glands, each- opening immediately upon the surface of the worm, are scattered over the entire cercarial body. While these may be used in cyst forma tion, it has been observed that at least some of them are still present in metacercariae which have been encysted for ninety days. Vital stains are taken up most strongly by the concretions in the intestinal caeca. . ’ : Cercaria develop in daughter sporocysts as described above. Development of the excretory system of the cercaria. The deve lopment of this system has been presented in detail for.other species o • of cyathocotylid cercariae by Looss (I896), Komiya (1939), and o Anderson and° Cable (19S>0). In every case the definitive number of O flame cells is attained by the addition of new pairs of flame cells « • and their ducts. Since most of the developmental stages observed by the present’author have been described previously, the excretory systems of only two developmental stages, not previously described, are discussed here* ’ In all previous descriptions of developing cercaria, a stage having four pairs of flame cells follows one with two pairs. On two occasions, cercarial embryos of M. kentuckiensis were observed which possessed three pairs of flame cells in' addition to a ciliated patch in the main collecting tubules (Plate I, Fig. 3)* The duct • of each of the third pair becomes associated with that of the anterior flame cell of the original two pairs, A stage with three pairs of flame cells in the tail-stem has been described as-following one with but one pair. On one occasion a stage was observed by the author in which there were two pairs of flame'cells in the tail-stem (Plate I, Fig. U), In the body of this stage there were five pairs of flame cells. The two median ascending tubules in the cercarial body as well as the two caudal tubules in the tail-stem have fused for a part of their length. The Island of Cort has not yet been formed. ° ° * The appearance of other flame cells in the body is apparently • o 0 rather rapid, since larger embryos show the complete complement of o fifteen pairs of flame cells in the body and three pairs ill the tail- o ° stem. °The only .other undescribed feature is the presence of bladder like enlargements of the excretory ducts at the tips of the furcae. O These were first evident in th.% stage with only three0 pairs of flame cells, but were not observed in embryos with the full complement* of flame, cells. . * • • Comparison. The cercaria, as described above,.differs in only a fet* points with the description given of it by Cable (1938); however, additional morphological features have been described which will per- ’ haps permit it to be readily identified and separated from other . cyathocotylid cercaria* • • . • • • The specimens measured by the present author were killed in hot ten percent formalin, while those of Cable were measured alive under slight cover slip pressure. . The single measurement given by Cable falls every time within the range given for- fixed' specimens. Since •fixation invariably shrinks .specimens, it can be concluded that the cercariae of the present author are larger than those of Cable, a fact' substantiated by measuring some cercariae by the method of Cab3= . A comparison 'of measurements of fixed and living cercariae from the same snails reveals that the fixed ones are twenty to twenty-five •• percent smaller* ' . • • Cable did not observe the connections of the flame cells, and he reported only fourteen flame cells in the body instead of fifteen* The full number of fifteen was reported by Vernberg (1952) for this cercaria* Cable reported the tail-stem to be without spines, how ever, Anderson (19UU) and the present author have observed these on the tail-stem of the cercaria of Mesostephanus kentuckiensis* While the present author has reported fifteen flagellets on the margins "of the body, Cable reported only about five* These are viewed vit^ great difficulty, and usually no more than six or eight are seen; how- .. • ever, the largest number seen was fifteen, aji observation confirmed many times. The pores of the excretory tubules in the furcae were not seen by Cable who believed them to be absent. Other additional features described by the present author are limited mainly to the number and distribution of flagellets on the tail-stem and furcae. Their arrangement in rather definite patterns is probably of value in species identification. Other adequately described cyathocotylid cercariae which closely resemble the cercaria of Mesostephanus kentuckiensis are Cercaria tatei Johnston and Angel, 19^0 and the cercariae of Szidatia • joyeuxi and Prohemistomum chandleri. Cercaria tatei is remarkably similar to the cercaria of M. kentuckiensis in size, flame cell number, anterior canals which branch into anterior, lateral, and median branches, and in fhe presence of small refractile granules in the anterior portion of the ascending excretory trunks and the anterior canal .and branches. It differs in not having concretions in the intestinal caeca and in lacking flagellets on the body, tail-stem, and furcae. It is © possible that these structures had been overlooked in Cercaria o tatei. The cercaria of Szidatia joyeuxi is also similar in size to the e o cercaria of M. kentuckiensis. However, it differs in the absence of ° O O ° .anterior canals from the anterior portion of the lateral ascending 0 O O trunk; in the absence of refractile granules in the anterior portions 9 of the ascending trunks; and in the absence of flagellets anywhere°on the cercaria. The flame cell pattern has never been completely / investigated for the cercaria of Szidatia .joyeuxi. -Of four des criptions given of the cercaria, two describe and illustrate concre tions in the intestinal caeca, one describes the intestinal caeca as being empty, while 'the fourth does not discuss them, but does contain an illustration of the cercaria.devoid of intestinal concretions. ' • • ' Vemberg (195>2) describes the cercaria of Prohemistomum chandleri as being almost identical to C . kentuckiensis. She states that the only differences are those in size, P. chandleri being the larger, and in the development of the anterior canal which arises from the anterior end of the lateral ascending trunk on each side. According to her this canal is always undivided in P. chandleri5 while in C. kentuckiensis the canal branches anteriorly into a lateral and a' median canal. The median canal may or may not unite with the one on the opposite side (Plate I, Figs. 6 b, c). The present author has found cercariae from a single snail, in which some of the cercaria had the characteristic unbranched canal of the cercaria of P. chandleri, while others had the characteristic • ■ branched canal of C. kentuckiensis as described by Vernberg (1952)* The second difference between the two .cercariae observed by Vernberg was the difference in size. The cercaria described in the present paper, when measured alive, has approximately the same measurements as those given by Vernberg for the cercaria of P. chandleri» * * ® Vernberg had recovered C. kentuckiensis from Goniobasis llvesceng * collected from Clifty Creek in southern.Indiana; while the’-cercaria of P. chandleri had been recovered from Pleurocera acuta collected in northern Indiana. It has been observed that cercariae obtained from different individual snails will vary in size, even if the snails are from the same stream. It would seem likely that cercariae 'obtained from different species of snails from widely separated streams might vary in size as much as has been described for these two cercariae recovered by Vernberg. A third difference between these cercariae observed by Vernberg was in the degree of deve lopment of the caudal bodies. , Their prominence is believed by the present author to depend upon the age of the cercaria. Activity of the cercaria. • The cercaria.of M. kentuckiensis responds to shadowing by rapidly swimming tail-first in a series of jerks. After reaching the surface, it settles to the bottom with the body directed down, the furcae spread, and the.tail-stem flexed'either to-the right or left. ' The direction.of tail flecture . is apparently-due to chance, being observed as many times.to the ‘ right as left,' / . ' ■Sixteen cercariae were placed in two inches of water in-a crystalization dish six inches in diameter. The.dish with the cercariae was placed in a-room darkened except for a single small., light located about'six feet to one side of the dish. When exam-'" . _ ined at fifteen minute intervals, an average of twelve of*the cer- .}. ■* * * !> * * « cariae were in the two inches opposite the/light source, thus „ ,, » • * * ^ . » „ • * exhibiting a definite photonegative "response, * ; ® ■ » 5) ® ® • The number of .cercariae® shed by a * single snail ® was observed © « ® © © © • © at eight h o w intervals over a period of eighty hours® The results are presented in Text Figure '1. Examination of the data shows that the most unproductive eight hour period is that between 8:00 A. M. and U:00 P. M., the period'of continuous light. The period during which the most cercariae were'shed was the U:00 P. M. to 12:00 midnight period.. The greater number during this period is to be expected since it follows a period during which, presumably,' . cercariae are still being, produced; ' • Penetration of the second intermediate host. . Penetration of- cercariae into the guppy was observed.' After making contact with the surface of the fish, the .cercarial body and tail are orientated at an angle of approximately 3 b - ^0 ° to the long axis of the' fish. The cercariai body now penetrates very rapidly, this- occurring in approximately UO - 60 seconds. Apparently the tail does not aid ■ directly in penetration since' it remains motionless while the cercarial body is making entrance into'the fish. While the tail- becomes detached after the body has completed its penetration, it may adher to the surface of the fish for as long as 2 0 - 3 0 minutes during which time it periodically flails, " . •• . ' ; Penetration of the cercaria into tadpoles -also occurs in a similar manner; however, shortly after entrance a pustule appears on the skin of the tadpole at the site of penetration. "Such fluid #■ ’ * filled pustules* were observed to contain red bloqd cells. © NUMBER OF CERCARIA TEXT FIG. 1. The natural emergence of cercariae of Mesostephanus kentuckiensis from a single a from kentuckiensis of Mesostephanus ofcercariae emergence natural The 1. FIG.TEXT 0 0 4 snail during eight hour intervals starting at 8:00 M. atstarting A. intervals hour eight during snail j2:oo 00 .0 4 PM i IE F A ' • ' DAY OF TIME 12.00 8100 AM 00 0 :0 4 PM 8:00 AM 4:00 PM P O The Metacercaria (Plate II, Fig. l). Dissection of a guppy three hours after exposure to cercaria showed that the metacercaria .were encysted. The worm was still active within the cyst which ‘measured 0.1^5 nun. by O.IJ4O mm. The cyst at this time consisted of a single thin wall of cercarial origin. After a few days, cysts dissected from the muscles and other tissues of the fish have two walls, the new outer wall being produced by the host, apparently ’ as a response to the presence 6f a foreign body. The outer cyst • is yellow in color, lacks melanophores, and has a very loose con struction. It is easily removed from around'the inner cyst. The hyaline inner cyst of worm origin, possibly formed by the cutaneous glands, is smooth and tough. It can be removed only with great difficulty, and the worm is frequently damaged when this is done* Within the cyst, the most obvious structures of the metacercaria are refractive concretions or granules which fill the excretory system. These make the bladder and the enlarged tubules appear • black, and they obscure other structures in the worm which are not already obscured by the vacuolated tissues of the -body* The following measurements are of the inner cysts of living encysted metacercaria. These were made while the cysts were ® 0 floating free in saline solution with no coverslip over them* At eight days the inner cysts measured 0 . 2 1 3 “ 0 ,2 6 ? mm* by 0 . 2 0 6 «■ S' ® ' ® @ 0.220 mm. (0.2^0 X 0.211* mm.). At sixteen days the cysts measure * ® © " © ® 0.2oU.- 0.311 mm.®by 0 . 2 1 8 - 0^281* mm. (0.283 % 0-23& Cysts e G 0 , after ninety days are found to vary considerably In size, possibly depending upon the tissue site in -which they formed.® $tost are only slightly larger than'the sixteen day old cysts; however, the outer cyst wall of connective tissue is considerably thicker in the older ' metacercariae. Observation of living metacercariae reveals that the flame cell number has not increased, it still being fifteen pairs in meta-' cercaria ninety days old. Following are measurements from twelve metacercariae ninety days old, released from their cysts, fixed in hot ten percent formalin, stained, and mounted. Length of the body 0.190 - 0.3^1 m. (0.270 mm.), width 0,165 - 0.219 mm. (0.180 mm.); oral sucker, length 0.030 - 0.037 mm. (0 ,03)4 mm.), width 0,032 -. 0.0l;5 mm. (0 .03L mm.)j pharynx, length 0.02L - 0.03U mm. (0.030 mm.), width 0.021 - 0.029 mm. (O.O2I4 mm.); ventral sucker, length 0.021 - 0.026 mm. (0.023 mm.), width 0.021 - 0.026 mm. (0.02h mm.); tribocytic organ, length O.0I4O 0.058 mm. (0.0ii7 mm.), width 0.035 - O.OI48 ram. (0.01*3 mm.); genital primordium, length 0.027 - 0,0i*2 mm. (0.03L mm.), width 0.019 - 0.027 mm.. (0.021* mm.). The ventral sucker 'is located 1*3/100 - 57/100 (5l/l0Q) the length from the .. anterior end. These measurements are ten to thirty-four percent smaller than measurements of living metacercaria under light cover slip pressure." * • • •, .Only the metacercaria of Szidatia j oyeuxi has been suffiqiently described to make a comparison with the present form. It is about twice the size of the metacercaria of M. kentuckiensis. The most conspicuous difference between them is in the relative sizes of oral sucker and®pharynx. In S. joyeuxi the oral sucker is fifty to © 0 ® one-hundred percent larger than the pharynx, while in M. kentuckiensis the oral sucker is never more than twenty percent larger than the pharynx. The Adult (Plate II, Fig. 3), Measurements in the following description are from five adult specimens obtained from the intes- time of a chick which had been fed metacercaria ten days previously The metacercariae were obtained from a specimen of Lebistes reticulatus which had been.experimentally infected with cercaria twenty-nine days previous to being fed to the chick. The presence of trematode eggs in the feces of the chick on the seventh day indicated that the five worms which were measured had been egg-' laying adults fcpr about three days. Other adult specimens from the same host were sectioned for other details (Plates III, IV, V, and VI). ’ Description of the adult of Mesostephanus kentuckiensis (Cable 1935) comb, nov. Specific diagnosis; with the characters of the genus Mesostephanus. Body pyriform, spinose to level of ovary, divided into two regions; the anterior region yellow - orange in life, flat and wide; the posterior region shorter, narrow, cylin drical, and truncated. Total length 1.078 - 1.5U5 mm. (1.28l mm.); maximum width at level of ventral sucker 0,350 - 0.522 mm. (0.U00 mm.). Oral sucker subterminal, measuring 0.058 «• 0,0?0 mm. in length, 0.061 - Q.06? mm. in width (0.0614.X 0.06U mm.)% No fire— pharynx. Pharynx'muscular; length 0.070 - 0.086 mm.,, width 0.068 - 0.089 mm. (0.079 X 0.079 mm.); with cavity flattened laterally 9 (Plate IV, Fig.-l). Esophagus frequently contracted, measuring D.Oii.0 - 0.088 mm. (0.065> mm.) in length. IntdStinal caeca sinuous * * # 0 'to level of ventral sucker, posterior to which, they converge slightly before extending to posterior border of posterior testis. Ventral sucker feebly developed, (Plate IV, Fig, 2), located anterior to tribocytic organ and vitellaria, 0,028 -> 0.0UU mm, X 0,032 - 0.0U7 mm. (0.033 X O.Okl iron.),•located 23.5/100 - 3k.3/100 in the length of the body (aver. 30.2/100), Tribocytic organ well developed with a median slit, glandular (Plate V, Fig, 1); 0.273 - 0«3?6 mm, X 0.17k - 0.2k8 mm. (0.323 X 0.207 mm.), ’ . Testes tandem, oblong or subspherical, in posterior half of body, median or slightly to the rightj linear extent 0.277 - 0.363 mm (0.328 mm.)j anterior testis slightly larger, measuring O.lkk ' 0.18k mm. X, O.I36 - 0.176 mm. (0.163 X 0.137 mm.); posterior testis 0.118 - 0.18k mm. X 0.102 0.160 mm. (0.lk7 X 0.127 mm.). Cirrus • sac large, cylindrical, anterior end extending to. posterior level ' • of vitellaria (Plate VI,- Fig. 1), .6.360 - 0,363 mm. X 0.083 - 0.10k mm. (0.k53 X 0.093 mm.).- Convoluted seminal vesicle occupies anterior one-fourth of cirrus sac, glandular pars prostatica the middle one-half or more (Plate VI, Fig.- 2), the cylindrical cirrus the last one-fourth or less. Genital atrium small, genital pore terminal. Ovary between and to right (.or left in some specimens) ' of testes measures 0,036 - 0.10k mm. X 0.080 • O.096 mm, (0.080 - 0.085 mm.). Mehlis* gland diffuse, medial and posterior to ovary. No Laurer’s canal observed.' Uterus with ascending limb and anterior portion of descending limb occupying area between, vitellaria, (Plate V, .Fig. 2 and Plate VI, Fig. l), posterior portion of ■* descending limb parallel or ventral to cirrus sac, (Plate 6, Fig. 2), terminated by vaginal sphincter before entering genital artium. Vaginal' sphincter 0.056 mm. in diameter, O.Olh - 0.019 mm. in thickness (Plate III, Fig. 1). Eggs large, few (one to four), . • operculate, yellow, measure 0.095- O'.llh mm. X 0.063 - 0.075 mm. . (0.103 X 0.067 mm.). Vitellaria composed of large vitelline follicles extending from anterior, border of tribocytic organ to anterior border of posterior testis, ventral and lateral to the genital apparatus, follicles disposed in two rows whose anterior portions are confined to the anterior half of the tribocytic organ* Linear extent of • follicles-0,311 - 0.501 mm. (O.I4.2I4■ mm.), - . . . . Comparison* M. kentuckiensis can easily be distinguished from other species, of Mesostephanus by the large pharynx, which’ is always . longer and wider than the oral sucker in M. kentuckiensis. It is- ' also separated clearly by the location of the ventral sucker, •23/100 3U/100- of the length from the anterior end of the body.’. In other species it is never less'than 37/100. , M. kentuckiensis is longer in comparison to the width than any other species except M. cubaensis as described by Dubois (1951). In life, the forebody ■' of M. kentuckiensis is a bright yellow-orange, a condition not •• reported for other- species of the genus. • ’ ..- ■ ’ • Chandler and Rausch (19U7) reported the orange color as occurring in the vitellaria of Neogogatea bubonis. The type slide- of this worm was kindly loaned to the author by Dr. Allen McIntosh from the U. S. N. M. helminthological collection, along with the • « • ® • type slides of Neogogatea pandionis, and Mesostephanus longisaccus. All three of these” species had been originally described as lacking ,a ventral sucker. Dubois (19!?1) had reported finding a ventral sucker thirty-seven microns in diameter on specimens of M. longjsaccus loaned to him by Dr. Chandler. This observation was confirmed by the present author. In addition a ventral sucker has been located on both the species of Neogogatea by the present author Longevity of adult worms. • Data obtained from three experi mentally infected chicks would indicate that the life span of the adult of M. kentuckiensis is between fourteen and seventeen days • in this host. The feces of the three chicks were examined daily • for trematode eggs. In the feces of one, eggs were not found fourteen days after feeding the metacercaria. In the feces’ of a second chick, no eggs were found after seventeen days. .'The- \ • • intestine of a third chick was examined seventeen days after feed ing, ’at- which time an abrupt decline in the number of eggs in. the' feces was observed. A single specimen of M. kentuckiehsis was recovered in the lower third of the small intestine. The usiml ■ • site'of infection in.chicks with both immature and adult specimens is the first'one-third of the small intestine. The above relation ships suggest that the fluke recovered in the lower third of the small intestine was a senile individual in the process of leaving DXSCUSSICM ‘ ; Geheral Aspects of the Life Cycle . » S' -i, ,f' » ■ ° ’ Enough life histories in the family*Cyathocotylidae are ..known that many generalizations may be made concerning them# The first • intermediate host is a prosobranch snail, marine or fresh-water* Earlier, in .this paper two possible exceptions were discussed; one was dismissed as an error and the other, considered' doubtful. Snails implicated as hosts can be assigned to only seven or eight families , .within,the subclass Prosobranchia, The five' marine cercariae infect a single family of snails, the family Cerithiidae,-- ■Although the mother sporocyst has been reported for only four of. the, twenty-six species, it is probably' a stage which all cyatho- cotylids have .in common. The mother sporocysts, in most cases where. this stage has been observed, give rise to .daughter sporocysts which in .turn produce cercariae, ' Two exceptions have been previously mentioned; those in which the first and second generation sporocysts produced miracidia. In discussing the occurence of miracidia in * * * ' • 'mother and daughter sporocysts of C. indica XV, Sewell (1922) points .out that the production of miracidia by sporocysts lends support to the view presented by Looss in 1892, that miracidia, redia, and * c* , cercaria are homologous with each other, *In all these stages the. * , • ' ' • * body structure is fundamentally the same, Anderson and Cable (195>0) * * * ' . ® « point out that the interpretation of Sewell’s findings from the * phylogenetic standpoint is difficult. They further.state that until this phenomenon, is confirmed in other species, and the manner in which the miracidia develop is understood, the interpretation of Sewell*s observation as either the retention of a primative phase*or a coenogenetic intercalation in the life cycle would be mere con- jecture* The second description'of miracidia being produced by * 'sporocysts, Premvati (195>£), adds some to our interpretation of this .phenomenon. She'described three generations of sporocysts of ' Cercaria- multiplicata. The first generation sporocysts produce only miracidia. The miracidia leave the first generation sporocyst by a terminal birth pore and devlop into second generation sporo cysts which in turn produce miracidia. She pointed out that.the ■' miracidia in the second generation sporocyst of Cercaria multiplicata seem to elongate and develop into a third generation sporocyst before .leaving.the second generation sporocysts. Cercaria only are* produced in the third generation sporocysts. While Sewell described only two generation of sporocysts for C. indica XV, it is possible .that he observed three generations.' He states that he observed mother sporocysts with only miracidia within them, while on one. occasion he observed mother sporocysts with daughter sporocysts which in turn contained miracidia. Later' in his discussion he states that he never found'miracidia'and cercaria. developing together within the same sporocyst, thus implying three generations of sporocysts'0 .Both Sewell and Premvati described miracidia as developing within the sporocysts from germ balls, even having the layer of germinal* j. ■% • 9 epithelium of the germ ball surrounding them for a time. More * ® © important is the fact that both describe the miracidia as lacking 61 ' • ' pigmented eye-spots and as being produced in both first and second ; generation sporocysts which had terminal birth pores*' Also the cercaria described by Premvati, C. multiplicata, is different from any previously described cercaria in possessing six pairs of flame cells in the body in combination with two pairs in the tail-stem. Five pairs in the body and two pairs in the tail- •' stem is characteristic of the Tetis subgroup. Sewell's C. ihdica XV belongs to the Vivax subgroup which has.fifteen pairs in the body' and three pairs in the tail-stem. ' ...• ' ; ' • These two cyathocotylid cercaria, which probably are not closely . related, are produced in third generation’ sporocysts which in turn are produce miracidia. All of the sporocysts of these cercariae are provided with terminal birth pores0 All other cyathocotylid sporocysts have sub-terminal birth pores„ In both cases the miracidia are typical except for the absence of eye-spots which are' present in all miracidia "that develop in the eggs of adult strigeoid worms, .These facts are interpreted by the present author to be an indication that the production of miracidia in sporocysts is a . primative characteristic. .That these three features could develop and become established independently in the life histories of two' cercariae which are so different, is highly improbable. -The causes leading to the retention of these primative characters, if they are • • . such, remain obscure. * • 0 • * • In contrast to the parthenital stages which, are restricted to ° * 9 snails belonging to a single subclass, the metacercaria of the 0 o • 0 cyathocotylids are found to encyst in a great variety of cold-blooded © © 0 0 © © aquatic vertebrates, fish and amphibia. Again, there have been two exceptions reported. In both instances the first intermediate host, the snail, acts also as the second intermediate host. Encystment in the second intermediate host occurs shortly after penetration by the cercaria. An exception, reported by Balozet (19^3is the metacercaria of Prohemistomulum expeditum from the kidneys of Rana ridibunda* This metacercaria remains unencysted. . In all but a few cases, only one or two species of fish.or amphibian are reported as being the intermediate hosts of such Speeies of cyathocotylid. The number of intermediate hosts known to harbor the metacercaria of Szidatia joyeuxi is a notable excep-* tion. .The fact that many species of fish and amphibians can act as intermediate hosts of Mesostephanus kentuckiensis and Szidatia . joyeuxi. indicates a low degree of host specificity of the larvae of. these worms. This is probably true also of other cyathocotylids* It is possible that it has not been .apparent because other hosts have not been checked for the presence of the metacercariae*' In the present work, fourteen species of fish from the Olentarigy River were checked for the presence of metacercaria (Table 2), Some individuals of eight different species were found to harbor the metacercaria of M. kentuckiensis, although in many there was found but a single cyst. Concerning the other six species of fish, so . * few were examined that it is safe to say only that the individuals examined were negative.- Another factor determining the presence * 4- c or absence of metacercaria would be the availability of the host to® m the cercaria. Fish*,which inhabit fast moving water*1 may not be subject to attack by cercaria.* There are at least eighteen other species of fish reported to be common in the' Olentangy River which were not examined* A search for metacercariae in these and also in other species previously found to be uninfected, would undoubtly reveal many additional second intermediate hosts of M. kentuckiensis* Adult cyathocotylids have been reported from a variety of vertebrates; however, out of over fifty species of these worms re- covered in natural infections, all but thirteen are found in birds*. Of the thirteen species not found in birds, six are found in mammals, ' six in reptiles, and one in a fish, Dubois (1953) considers species of the genus Mesostephanus to be natural parasites of birds of the order Pelecaniformes, He indicates two species from dogs, M, appendiculatus and M. longisaccus, as secondary or erratic parasites in that host. The present author agrees with Dubois that dogs are • probably not the natural hosts for these parasites. An. examination ■ of the remaining species reported from mammals suggest that mammals as a group are not natural hosts of cyathocotylids. Besides the two reported from dogs, there are four other species of cyathoco tylids reported from mammals. Two of these have been reported only once. These are Muhlingina lutrai Mehra, 1950 from Lutra lutra in India, and Eaboisia pgtrvovoipara (Faust and Tang, 1938) In Meles leptorhynchus in China, Prosostephanus pagumae (Faust and Tang, 1938)e was e found in Paguma V larvata1...... and Mustela...... sp. in China, The fourth species, Prosostephanus industrius, has been reported in o 0 Felis domestica, Vulpes vulpes, Nycterentes procyonoides, Herpestes urVfe., Felis viverrirjg, and Canis familiar is. All of these were ® reported in- China; however# it should be noted .that all but the last two hosts were zoo animals# which had died as a result of being fed ' • carp infected with the metacercaria# Tang (19U1) reported examining ■ one hundred and twenty cats and twelve dogs from the same region in China without' finding this worm# Of'all the. above mammals harboring these four species of worms, only one, Lutra lutra, could be considered as chiefly piscivorous# The remaining are carnivorous or omnivorous# • In addition they are not particularly associated with water; a • requirement for the continuance of the life cyle of cyathocotylids# Among the seven species found in reptiles and a fish, only one, • Szidatia joyeuxx, has been recovered more than once from its cold blooded host. Of the remaining six species two were reported once from crocodiles,' three species once from snakes, and one species once • from a catfish. These seven species of trematodes do not form a" ; • homogeneous taxonomic group; the two found in crocodiles belong to •' the genus Cyathocotyle, the subfamily Cyathocotylinae, the remaining five speci'es from snakes and a'fish belong to four different genera in the subfamily Prohemistominae, Since all of'these cold-blooded . hosts eat fish or amphibians at times, they are understandably vulnerable to infection with cyathocotylids which show low host. '• specificity# • ’ • • ' ' Returning to the proposal of Dubois, that the natural hosts of species in the genus Mesostephanus are birds of the order Pelecani- formes, we find that all but one were reported from this group of « birds. The one exception is Mesostephanus haliasturus, described by © Tubangui and Masilungan (19Ul) from Haliastifr indus of the order 8 ' — — Falconiformes, Dubois (1951) 'has observed this uncomformity .and he points out that Tubangui and Masilungan describe in the same . paper another species of cyathocotylid, Prohemistomum fregatae, ' , . .from Fregata ariel. Dubois further shows that all other known species of Prohemistomum are parasites of birds of the order Falconiformes, Y and he suggests that perhaps Tubangui and Masilun'gan might have accidentally exchanged the collection labels'of the two species. Even accepting the proposal that the species of the genus' Mesostephanus .are natural parasites of birds of the order Pelecarii- • formes, it is not too hard to imagine that one-might occasionally appear in a bird of another order, especially in view of the apparent . lack of host specificity as shown by the appearance•of two species in dogs. ' The author has examined two specimens of a cyathocotylid obtained by L. F. Conti of’San Diego, California from Fregata sp,- Conti haa deposited these worms' in the U. S. N. M. Helmintholo gical Collection (No. hhl30), having'identified them as Mesostephanus 'infecundus. As pointed out earlier this species name was proposed by Lutz (193$) for a parasite which he failed to describe or illustrate •and which he had obtained from Fregata aquila. Examination by the ‘ present author of the worms recovered -by Conti indicates that they are‘probably an undescribed species of Prohemistomum. -This finding indicates that the host specificity of these two genera of worms is probably not so strict as Dubois has proposed, . Restricting species of the genus Mesostephanus to birds of the *s i order Pelecaniformes would implicate either the White Pelican,. * 9 * 9 ■* 9 & ® Pelecanus erythrorhynch’os or the Double-crdsted Cormorant, Phalacrocorax auritus as the possible natural definitive host of M. kentuckiensis.* Borror (1950) lists the former as a .rare transient, in Ohio, the latter as a rare to common spring and fall transient. Phalocrocorax auritus appears the most available possibility as the host. This species has been'reported as the host of a species of Mesostephanus, M. cubaesis. There are other piscivorous birds in Ohio which are more common and which may act as hosts of M. kentuckiensis. . General Taxonomic Considerations According to Dubois (1951) there"is general" agreement among investigators working with cyathocotylids that these trematodes form a homogenous grouping upon the basis of the anatomical uniformity of the excretory system of the cercariae. He points out that the diversity of the adult forms contrasts with the uniformity of the cercaria. Anderson and Cable (1950) believe that the Cyathocotylidae are more primative than are the Strigeidae, basing this belief in part upon the fact the cercaria of cyathocotylids develop' in prosobranch snails which are more primative than pulmonate snails, the principle snail hosts of other Strigeata, • In the hope of finding a clue to'the origin of the cyathoc'o- tylids based-upon their unusual specificity for prosobranchs as . first intermediate hosts, descriptions of over three hundred e strigeate cercaria other than cyathocotylids were examined. Out of these, only fifteen are found in prosobranch snails. These fifteen 0 © 0 © ® © occur in four natural groups of strigeate cercaridte, of which the relationships to adults are known for only two of the groups, the schistosomes and the strigeids. Three of the cercaria are definitely of the family Schisto- Somatidae, superfamily Schistosomatoidea. The cercariae are similar to those of cyathocotylids in having the caudal excretory pores opening at the tips of the furcae, and, among some' species, in having ; dorsal and ventral fin-folds on the furcae. It might also be pointed out that the miracidia' of members of the family Schistomatidae do not have pigmented eye-spots, a characteristic of the miracidia produced by mother and daughter sporocysts of the two cyathocotylids Cercaria indica XV and Cercaria multiplicata. There are many other differences between the schistosomes and cyathocotylids. The main differences are that the schistosomes have a two-host life cycle instead of one involving three hosts, they are apharyngeatej and their' cercaria are brevifurcate and develop in sporocysts that do not have.birth- pores. • . . The cercariae belonging to the other recognized group are of one of the other families in the superfamily Strigeoidea to which La Rue (1957) and others assign the family Cyathocotylidae._ The .cyathocotylids resemble most of the other'Strigeoidea in having pharyngeal, longifurcate. cercaria which develop in elongate sporo cysts that have birthpores. They are also similar in having mira- • * cidia (from eggs) with a pair of pigmented eye-spots and in having « ®a life cycle involving three hosts. They differ mainly in that the IE) ® ® © ® **> other Strigeoidea have the furcal excretory ducts opening upon the surface somewhere between the'tail-stern and the fureal tip instead of at the tip. Cercaria tetradena Johnston and Beckwith, 19U5, has characteristics of both the cyathocotlylids and the other Strigeoidea It is a typical cercaria of the subsuperfamily Strigeines (families Strigeidae and Diplostomatidae), except that the excretory ducts open at the furcal tips and the cercaria develops in a prosobranch snail. Stunkard (19!#)* in-discussing early developmental and - 'larval stages as evidence of systematic relations among animals, presents a .principle which may be of value in interpreting the above relationship's between the larval stages of cyathocotylids and the larval stages of other strigeates. He states that larvae of related groups show greater resemblance than do the adults, and the resem blances are greater, the .earlier the developmental stage. . • If the supposition is accepted that the production of mira cidia in sporocysts is a primative character, then we could possibly interpret the resemblance between those miracidia produced in sporo cysts of cyathocotylids and those produced in eggs of schistosomes as being due to common origin or ancestry. The excretory openings at the furcal tips may be additional evidence of relationships. Stunkard {!$$$) also points out that the free-living existence of young individuals has resulted in development of adaptive, coeno- genetic features, which have no phylogenetic significance. The presence or absence of furcal fin-folds would seem to be such an * * • adaptive feature. In addition fin-folds are not restricted to only • . * some species of cyathocotylid and schistosome cercariae. ® • • . 69 The comparison of cercaria and other larval stages is hampered con siderably by inadequate and, quite often, inaccurate observations of these larvae stages. ‘ ' •••'•• Some Taxonomic- Relationships Within the Subfamily Proheroistomtinae The classification of cyathocotylid cercariae has been reviewed earlier. .The most comprehensive attempt towards this end is that of Dubois (1931). The groups are characterized by Dubois as followst (l) Subgroup Vivax Sewell, 1922 (emend.;Dubois, 1951). Formula of excretory system: 2 £(3+3+3) ♦ (3+3+(3))Ja 36. (a) Furcae long with fin-folds all their length; ventral ' . - . sucker present; 6? or 8 glandular cells included in " ■ the 'oral sucker. • C. of Prohemistomum vivax (Sonsino, 1892) ? C. indica XV Sewell, 1922 1 ,C. dorosocauda Tubangui, 1928 . ? C. .utriculata Lutz, 1933 Cercaria R McCoy, 1928 (b) Furcae long with fin-folds upon their distal half; ventral sucker present; eight gland cells in the oral sucker. C. tatei Johnston and Angel, 19UQ ? C. kentuckiensis Cable, 1935 ? C. of Szidatia joyeuxi (Hughes, 1928) (c) Furcae long with fin-fold only at extremity; ventral sucker present. C . of Pr.osostephanus- industrius (Tubangui, 1922) (d) Furcae long without firi-folds, no ventral sucker. C. of Paracoenogonimus ovatus Katsurada, 19lU C. of Paracoenogonimus szidati (Anderson, 19UU “ nov. comb. C. notoplae Johnston and Peckwith, 19U3 ?. C. of Linstowiella vivparae (v. Linston, 1877) * * * (e) Furcae short without fin-folds; po ventral sucker. C* leptoderma Faust, 1922 * * m 9 0 m * o p * : • o ° 0 ° o • s ° * 0 0 o °e * . ° o ® o o * % » * ® o • « 70 • ‘ (2) Subgroup Novena. Formula of excretory system: 2 [(3+3) + (3+(3))] = 2k • C. of Gyathocotyle orientalis Faust, 1921 C. of Cyathocotyle gravleri Mathias, 1935 (3) Subgroup Tetis Sewell, 1922 (emended) (Dubois 1951) Formula of the excretory system: ' 2 [5 ♦'(2)] = llu Furcae long without fin-foldsj no ventral sucker. C. of Holostephanus curonensis Szidat, 1933 C. balthica Szidat, 1933 C. indica XXXIII Sewell, 1922 C. theodoxa Porter, 1938 (U) Subgroup Tauiana Szidat, 1933 Formula of the excretory system: 2[2+2+2+(0)] =12. Furcae long without fin-folds, no ventral sucker. C. tauiana Faust, 1930 In the scheme above Dubois has united all the cercariae of. the subfamilies Prohemistomatinae and Prosostephaninae in the Vivax sub group while the cercariae in the subgroups Novena and Tetis are of the subfamily Cyathocotylinae. • Since the presentation of this classification of cercaria, there have appeared descriptions of seven additional cercariae. Two cercariae, described by Cable (1956)' C. caribbea L and LI can be • assigned to the Vivax (b) grouping. The remaining five cannot be assigned to any existing grouping since they possess new features or Combinations of features previously unknown. C. yankapinensisj, described by Goodman (1951), has the flame cell pattern of the Tetis - subgroup to which he assigned the^ cercaria, but an examination of his * ■* illustration shows the presence of a ventral tucker. By adhering to * the system of, Dubois, the Tetis subgroup should be emended aid sub- 9 » ® ® O divided into groups (a) and (b). @ © O O o © o # o' o ©■ © ' • 71 ' Subgroup Tetis Sewell, 1922 (emend.)# . • ' Formula of the excretory system: 2 [5 + (2)] 3 111. ' (a) .Furcaje long without fin-folds; no ventral sucker, C. of Holostephanus curonensis Szidat, 1933 . • • C. balthica Szidat, 1933 ' • C . indica XXXIII‘Sewell, 1922 . C. theodoxa Porter, 1938 ' • • . (b)’ Furcae long without .fin-folds; 'ventral sucker present, .. C‘. yankapinensis Goodman, 1951 The four other recently described cercaria, the unnamed cercaria of Haxon and Pequegnat, C. kasenyi, C, schontedeni, and C. multiplicata, form a homogenous group with regard to the excretory system,.'Six" ‘• pairs of flame cells are reported in the body, two pairs in the tails,' There is some difference from the other three in the connections of ■ the flame cells in C, multiplicata which was described by Premvati. ' / She has shown the pattern to be 2lj3+3+(2)J 3 16 in C, multiplicata, while in the other three the pattern is 2 [(2+2) ♦ (2+(2))] 3 16. In addition, the furcal excretory pores are located just posterior to the furcal junction in C, multiplicata.- This may. not be as presented by Premvati since these ducts are sometimes extremely difficult to ■ observe to their ends. For this new subgrpup the name Kasenyi is proposed by the present author. Upon the basis of the development of the fin-folds on these cercariae, the group probably should be sub-, divided into three divisions. However, until more species are des cribed this move is unwarranted. Subgroup Kasenyi , r Formula of the excretory system: 2 [6 +(2)] =16, No ventral sucker; fin-fold absent, partial, or complete,* * # C. of Maxon and Pequegnat, 19^9 # £. kasenyi Fain, 1953 , * * * G°. schoutedeni Fain, 1953 » ^ m— _ - - A ' , , ® . C. multiplicatanPremvati, 1955 0 - - » « O 0 o ® Q 0 • Cercaria. kentuckiensis has been placed by Dubois in the (b) ; * • group of the subgroup Viva.x# indicating a dose relationship with the cercaria of Szidatia joyeuxi and C. tatei» The similarities and differences between these cercaria have been discussed following the description of the cercaria. A close relationship between these cercaria would now indicate also a close relationship between the adults of the genera Mesostephanus and Szidatia. . Dubois (1938, ’1988) placed these two genera in two different subfamilies, Prohemistomatinae and Szidatinae, mainly upon the basis of host specificity. The flukes in the latter subfamily are all parasites of snakes. Later Dubois (1951) transferred a 11 the members of.the .latter' subfamily to the family-Prohemistomatinae.. His principle ’ reason for this transfer was the discovery in a snake of a species of fluke which possessed’characters intermediate between Hesostephanus and Szidatia. This worm, Mesostephanbides burmanicus (Chatterji, . 1980) Dubois, 1951* was originally placed in the genus Mesostephanus. by Chatterji, than later transferred to the genus Gogatea by Mehra (1987) until Dubois (1951) erected a new genus for it. Dubois thus recognized a closer relationship between the genera Mesostephanus and Szidatia-than he had previously, partly because of the interme- • diate form Mesostephanoides burmanicus and also because the 'cercaria Szidatia joyeuxi, which is in the Vivax (b) -grouping, is intermedi ate in structure between cercariae in the groups (a) and (c), which • • • are cercariae of'flukes in the subfamily Prohemistomatinae. o e © © There is a great monphological similarity between the genera s Prohemidtomum and Mesostephanus, The great .number of .speqies assign ed incorrectly to one or the other of these two genera testifies to,* ' their similarity. They were not clearly separated until some distinct characters of each genus’were recognized by Dubois (1951), who pointed* out such differences as the length in relation to the width of the • » “ ® body; the location of the acetabulum in the bodyj the length of the \ body in relation to the diameter of the vitelline crownj and the /nature of the termination of the uterus* In Prohemistomum the‘uterus ends in a metraterm while -in Mesostephanus it ends with a vaginal ° * sphincter.- However, with regard to the- location of the acetabulum •- •' •in.the body, M* kentuckiensis shows a resemblance to the genus Prohemistomum. The acetabulum is located between 23-3^/10^ °f the •body; while‘in all other species of Mesostephanus it is between h2-£8/lOO. In species of the genus Prohemistomum the ventral'sucker is between 2£-ltO/lOO. -Dubois (1953) states that the ovary is generally to the left of the median plane in Prohemistomum, except for P.•vivax, and to the right of the median line .in Mesostephanus, except for M. ■ .appendiculatus. Examination of'the adults of Mesostephanus kentuck iensis by the present author reveals that the ovary is on the right side of the median. plane as often as-on‘the left side'of the mediae • r-i * $ « * plane* . *. ^ , There have been but two described species in the genus Szidatia^ b . » joyeuxi and S. nemethi, both occurring i n ’snakes in North Africa# * * t ■ — - O m * .« * These species are similar in many respects to species of the genus • O Mesostephanus, but they arse equally similar to species of the gemfs e _ 8 ® • o Prohemistomum, to which they are not quite so eclosely related on the basis of cercarial characters. They resemble species in the genus * ■**'’'.» v * * Prohemistoritum in the location of the ventral sucker* The present 3, * * * * * -■*' author has pointed cut earlier that the cercaria of Prohemistomum’ < ■ n ■ ■ chandler!, as described by Vernberg (1952), was .similar to the cercaria of Mesostephanus kentuckiensis* Dubois (1953) doubts the generic " attribution of Prohemistomum chandler! upon the basis that vitelline glands had not yet developed and that the ventral sucker, as illus-.’ trated, is 53/100 of the length from the anterior end of the body,-•* . the latter'being a character of the genus Mesostephanus. If'the • worm described by Vernberg is identical to M*'kentuckiensis re- covered by the present author, the ventral sucker should hot be . . located as in typical Mesostephanus, since M». kentuckiensis is not .typical in this character. Text Fig* 2 illustrates the effect.of- \ •differential growth rates of the anterior and posterior regions of ** the body upon the position of the acetabulum in the body of M« kentuckiensis from cercaria’through the adult* Examination ©£ the,:" figure shows that in an- individual of M*. kentuckiensis recovered • # ■ * s n three days after feeding, the acetabulum is an average of Ia9/1Q0 * * * * * 0 of the length from the anterior end of the worm* While this does* 0 ,e •*’ . ’ * • ® J * . ® not definitely establish Prohemistomum chandler! as identical to Mesostephanus kentuckiensis, it does indicate that the specimens 4 • referred to by Vernberg belong to the genus Mesostephanus* The* 9 slides of P. chandleri have been requested from Dr. Vernberg*who — ...... — * ® • a & •> ■« ha;s replied, that she does not have them in hand at present.,8, METACERCARIA IMMATURES ADULTS 80 70 60 50 4 0 30 20 0 4 6 12 16 20 24 • 28 32 36 4 0 TIME IN DAYS STARTING WITH MATURE CERCARIA % . TEXT FIG. 2. ■ Location of the ventral sucker from the anterior end in relation to-the • * * s * length of the body in Mesostephanus kentuckiensis from cercaria through the- adult • ' , ' ,, ■ CLASSIFICATION. , • •' ; ^ f-’ O o ° 03 * "the great number of trematode life histories which have, become known in the last thirty years has contributed greatly to an under- fr o ' , , '» standing of the relationships between the higher taxonomic groups# The following classification is from La Rue (1957) and' Dubois (1953) o 0O Phylum Platyhelminthes ° class Trematoda Subclass Digenea. “ ! . Superorder Anepitheliocystidia La Rue, 1957.- • 4 Order Strigeatoidea La Rue, 1926 • Suborder Strigeata La Rue, 1926 Superfamily Strigeoidea Railliet, 1919 ’ ’ Family Cyathocotylidae Poche, 1925 . • Diagnosis: Strigeoidea with body small, either oval and LurSiform, or ovoid to piriform and massive, or more dolioform, either elliptical to linguiform or foliiform « ‘ to spathaceous, and more or less concave ventrally# • . Tribocytic organ either cyathoid and very prominent, or circular or elliptical, into which the genital organs do . ■ not penetrate or only in part. • Ovary opposite the cirrus •' • pouch, testes' ovid to ellipsical, spherical or reniform, sometimes virguliform or more or less bi-or trilobed, never multilobed. Vitelline follicles generally large or • > • average# • Parasites of reptiles, birds and of mammals# . Subfamily Prohemistomatinae Lutz, 1935 . ‘ Syn. Szidatinae Dubois, 1938 and .Gogatinae Mehra, 19h7. • Diagnosis: ‘Cyathocotylidae with body oval, piri«-or linguiform, undivided or provided with a small caudal , appendage not containing genital glands, or even more or less bisegmented and whose well developed posterior, segment contains all or part of the glands. Trib&cytic * » * '# 77 « *. organ relatively little developed, opening by a median slit and around from which the vitelline follicles, con fined behind the ventral sucker equator or pre-equator, u are gathered in arrangement almost always in a crown (exc, Szidatih). Genus Mesostephanus > Diagnosis: Prohemistomatinae with body long, foli'i-or linguiform becoming slightly concave behind by the fact of the ventral incurvature of the lateral posterior- border, with a small dorso-subterminal appendage (vis* in toto), Cirrus sac well or even very developed, • extending to 1/3 - 2/$• or even more than 1/2 the length of the worm and obtaining or passing the zone of the first testis. Ventral sucker well developed, more often ' post-equator,- or on the other hand, feeble or very , reduced. Anterior limit of the vitelline follicles comprising typically between 50 - ^8/100 of the length of the body, exceptionally more in advance, . Testes subglobular to ovoid (sometimes grooved). Presence of a vaginal sphincter, Metacercaria harbored by fish, ■ Natural hosts of adults: Pelecaniformes (Secondarily or accidentally in dogs). Species Mesostephanus kentuckiensis (Cable, 1935) comb, nov, •Diagnosis: See page o 0 SUMMARY • , . ' . The life history of the cyathocotyli'd trematode, Mesostephanus kentuckiensis (Cable, 1935) comb, nov., is elucidated. The cercaria, Cercaria kentuckiensis, is redescribed and the daughter-sporocysts, metacercaria, and adult are described. These stages are illustrated by camera lucida drawings and photomicrographs, and are compared to • similar stages of closely related specieso ‘ . The cercaria of Mesostephanus kentuckiensis was found in 3*18 percent of the prosobranch snails, Goniobasis livescans, collected from‘streams in central Qhio0 Daughter, sporocysts, but no mother '' ‘ sporocysts, were recovered from crushed snails from which.cercariae. • were emerging. The cercariae Were, found to encyst and develop experimentally ’ • as metaeercariae in-the common guppy, -Lebistes reticulatus, tadpoles of Rana clamitans and R. catesbiana, and adult specimens of R,..-' pipiens, „ ' ' Fish collected'from the Olentangy River near The Ohio State ■ University that were found to be naturally infected with the metacercariae of Mesostephanus kentuckiensis were Ictalurus natalis, Campostoma anomaluiy Cyprinus carpio, Notropis cornutus, Pimephales notatus, Semotilus atromaculatus, Amploplites rupestris, Lepomis macrochirus, Micropterus dolomieui, and Pomoxis annularis, A tadpole of Rana clamitans and R. catesbiana, and a specimen of the fish, * Micropterus dolomieui, collected from Big Walnut Greek in Pickaway 0 0 • • , 79 • * County* Ohio* were also found naturally infected with this meta cercaria* . . • ' , Metacercariae were fed to the following experimental definitive hosts: Matrix sipedon, Natrix septemvittata, Procyon lotor, as well as laboratory mice* and one-day-old chicks. Only.chicks became in fected with M. kentuckiensis.’ The metacercaria first became infective for chicks after having been in the guppy from twenty-two to twenty- '' eight days* . ■ ' Eggs of Mesostephanus kentuckiensis first, appeared in the feces' of experamentally fed chicks after, seven days.. In the chick, the life span of the adult trematode is between fourteen and seventeen days. .. . . ' ' _ . : • • •; '• Eggs recovered from the feces of infected chicks were Incubated . in either tap water or 0.5.percent formalin; however,'eggs did not develop for more than a few days. .'.••• The possible synonomy of Mesostephanus kentuckiensis and. Prohemistomum chandleri based upon the similarity of their’ cercariae '' and immature adults is discussed* The classification of cyathocotylid cercariae is reviewed and .two new cercarial groups are proposed for the reception of cercariae not assignable to groups erected previous to their descriptions* CercarTa yankapinensis Goodman* 1951 is assigned to the new Tetis (b) subgroup which is separated from the Tetis (a) subgroup in that the former possesses a ventral sucker. A- new subgroup, Kasenyi,® is erected for cercaria having six pairs of flame cells In the body., and *• ... |( two pairs of flame cells" in the tail-stern. To this subgroup are * % © * V-J ' _ r;» -.«• ® ® 0 & ' 00 a# n® n ® ®0 ® A © ® ® ® ® @ © 0 6 a assigned the unnamed cercaria of Maxon and Pequegnat, 19^9 as well* as Cercaria kasenyi Fain, 1953, Cercaria schoutedeni Fain, 1.953, and Cercaria multiplicata Premvati, 1955* “ The relationships of the family Cyathocotylidae to' other . families of the suborder Strigeata, based upon larval'characters,, are discussed# . . ■ . • • ' LITERATURE CITED Anderson, D. J, 19UU. Studies on C. szidati sp. nov., a new furcocercous. cercaria of the Viva* type. J. Parasit. 30:' ' 26U-268. , ’. V ■ ... ’* , * . *\; 1 ■». ■* . Anderson, D. J, and Cable, R. M. 1950. ‘ Studies on the life *•__ I - * » history of Linstowiella szidati (Anderson) (Trematoda: . * . Strigeatoidea) (Cyathocotylidae'). J. Parasit. 3&: 295-klO. •• • Azim, M. A. 1933# On Prohemistomum viva?: (Sonsino, 1892) and its , development from Cercaria* vivax Sonsino,-1892. Ztschr. » m ■ - -■ ■ ■■■■ ■ * . , 0 , «**■*' Par'asitenk. 3>: U32 —U36• 8 ’ * „ * « « v* * , ■ % . , . * ° * Baker, F. G. 1928. The fresh water mollusca of Wisconsin# Part** 1. Gastropoda. Wise. Geol. and Nat. Hist, Survey. • Bull. * , . 70s 1-570. * 1 J .i * < , * * . , ,_ ° Balozet, L* 1953# Trematodes larvaires de l*afriq\ie du Nords 1 <»• '« * t> t * * * „ larves de strigeida. Arch'. Inst, Pasteur Algerie 31: * 381-396 Banks, tf. 19U9. 'A study of cercariae from snail hosts of the * * ' . • * * Columbus, Ohio area. I The sis,-.Department of Zoology and Entomb * * • logy, The Ohio State University, Cclurfcus,' Ohio. . Borror, D, J. 1990. A check list of the birds of Ohio, with the. © « * 0 ® , ; migration dates for the birds of central Ohio. Ohio J. Sci. . , . ® • . • . ® 50:. 1-32 €. I ®o o 0 e % » * " . * © © © * Caballero, S., Grocott. and Zerecero. 1993. Kelmintos de la © . a ® ® Republics deePanama IX algunos trematodos de aves marinas ! •* ® * ** , * , ® del Oceano I^acifico del Norte. Anal, Inst. Biol. Mexico Cable, R. M. 1935. Ceroaria kentuckiensis n. sp., first representa ’ o ; tive of the Vivax group known to occur in the United States. ’ - o J. Parasit. 21: 1*36. Cable, R. M, 1937# Cercaria of Indiana. A preliminary not°e on larval trematodes from McCormicks* Creek with a description , of three new species. . «J. Indiana Acad. Sci. 1*7:' 227-228* Cable, R. H. 1938* Studies on larval trematodes from Kentucky , with a summary of known related species'. Am, Midi, Nat. 19! Iib04*61i. ^ . Cable, R, M. • 1956., Marine cercariae of Puerto Rico, Scienti fic Survey of Porto Rico, and the Virgin; Islands 16 (U): U91-577 - 8 .(N, I. Acad. Sci,). t *1 » -Callot,’ J, 1936, Trematodes du suo tunisien et en particuler \ J'’ du Hefzacua. Ann. Parasit.- lh:. 130-11*9, Chandler, A," 1930, Mesostephanus longlsaecus, a new cyathoco- . ; * ** - . , • ■ * % tylid. trematode of a dog, J.* Parasit. 36:. 9 e * * * * * , Chatterji, R. C, 191*0, Helminth parasites of the snakes of Burma^ °I * . ‘ 0 I,. Trematoda. • Philip.,J. Sci. 71: 3814*01, 3 0 ‘ * ® ° o . ® • Siurea, I»* 1916* Prohemistomum appendlculatura, eine neue HoXosio*-* * ° ® ® © s ® ® • s jndien-Art’aus Hunden-und- Katzendarm, dessen Infektionsquelle / e in den Susswasserfischen zu suchen 1st, Mebst einer Benerkung0 o * ® de Arbeit Prof, Kaisurada "Studien© uber Trematodenlarven © © _ o • e . ’ - >0 bei Susswasserfischeh, mit hesonderer Berucksiehtigung der . s . ° ® °0 $lb- und Alsterfische,’* eZtsehr* Infek't. tv* Berlin X ft 309-28 o ® • • ® a © , .^ollfus, 'Zm I V 1951* ‘QuelqueS trematode3$ @estodesg, and &ea>nthe«> P © © <§. ® cephales# *$Fch« Jnst# Pasteur Maro© 1*: XOl|-230g o » Dollfus, S. P. 1953. Mis0cellanea helminthologica maroccana VII o « les Szidatia®de Natrix viperina. Arch. Inst. Pasteur Maroe O © 0 Us 505-512. ' e o Dubois, G. 1938. Honographie des Strigeida (Trematoda)# Hem, Soc, Neuchateloise Sci, Nat, 6;- 1-535, . Dubois, G, 19UU. A propos de la speeifieiie carasitaire des Strigeida, Bull, Soc, HeuchAteloise Sci. Mat, 69: 5*103• • ’ Dubois, G. 1951* Wouvelle cle de determination des groups systezna- . tiques et des genres de Strigeida Poche (Trematoda), Rev, .Suisse Zool, 58: 639-691* • ° ° ’ V> & r- ’’ r> ~ O Dubois, S, 1953, Syst6matique des Strigeida. Complement de la ° o ° ° ■ • Monographic, Hem, Soc* Heuchateloise Sci. Hat, 8t 1-lUl. !!-, ca O Pain, A, 1953, Contribution a I 1 etude des formes larvaires des w (- , o trematodes au Congo Beige et epecialement de la larve de ° !>-■ 0 ••• ■' * cO i (D Schistosoma mansoni-. Mem. Inst. Royal Colon,’Beige 22 (5) ? * O ° O Q G 6 ° Q° 1-321, 3U pi. • o ° ° o„ o 0 * O o 0 ° a ° o » On o 0 C 0o faust, E. C, 1919* The excretory- system in Digenea II. Biol. "o 0 ,, . ® Bull.'36: 5 322-339® • o 0 ° ° o , o * o o jteust* £• C* 1922.) Hotes m larval flukes fromChina* Parasitology ■T) t • Uit 2U8-267# G 0 ° ’ ’ • % ° °0O o° ° • 0 ° ° Oo ° c ° s fiust, E* C. 1930# larval flutes associated with the cercariae of * e • <9 0 jSlonorehls sinensis Is® Mttsrawid snails in China and adjacent territory® parasitology 22$ Ui5-15j!?® ° 0 • ° o ° ® 1|ocdrna% J)(i) 19511; §t>udi@a» €>$> tfematodf ©epeaplaeat Reelfoof ° o ' o ° -©• # ieWS» of® ffcMl® Jt,esJ® • 0 o ° Johnston, T. H. and Angel, L. M. 19b0Larval trematodes /rom '* * ... Australian fresh-water molluscs. VII, Trans’, Soy. Soc, South Australia Ski 331-339* Johnston, I, H» and Beckwith, A* C, 19h$» Larval trematodes from 'Australian fresh-water molluscs* ’X# Trans* Roy, Soc,‘South Australia 69: 229«2l}2# . » ® * . .Joyeux, Ch. and Baer, J. G* 193 U. Sur un Trematode de Couleuvre# * ‘ * ’ ; Rev. Suisse Zool© hit 203~l£# *; >•> !0 0 ?1 • ® Joyeux, Ch. and Baer, J, G, " I9I4I* Le cycle evolutif de Szidatia o 'n 3 '•r’ • B 18 joyeuxi. Arch* Inst. Pasteur Tunis 30s 279-286, 0 0 =• * * ", " ’ a Katsurada, F, 191U* Studien uber Trematoden Larven bei Sftsswasser- •S' O 41 . * » 0 fischen, nit besonderer berucksichtigung der Elb- und Alster- o » • ' ^ fische. Central bl, Bakt, und Parasit* I Abt, orig* .73: “ s 9 ‘30U-31U. *. /-.• . .*;• • 0 \ .••* . ’ ° 8 » 9 O ’ © • . ‘ Komiya, Y* 193?* Lie Entwicklung des Exkretionssystems einiger » © • * a •* © . . * • ' * ° . Trematodenlarven.aus Alster und Elbe, nebsfc Bemerkungen uber o0 o . • • Ihren’Entwicklungszyklus, Ztschr, Parasitenk 10: 3hO-385>« O is ® * . 8 . » e Langercn, M, 192!;* Reeherches sur les cercaires des piscines o » * o o G * ® # 0 © a o o de Gafsa et enqueto sur la Bilharsiose tunisienne* Arch* o 0 0 Inst* Pasteur Tunis 13? 19-67* 0 ° G e*e ® 0 ® • • , e © © © 0 0 % © Xa Rue, .G* R* 1937* Classification of the digenetic trematodes® § 191$® Obfemtfons on the apreal . * o of Asiatic geMstssomlaiii® Irlt# Sfed* 3>g gOL-203® . © 0 © ©° a ® & * %oB0sS) I® £8f6. ^Reeherehes *uf la taunt parasltaSra 1'ggyptfe. 0 o ° o |a?°ei|ier@ fart I®®0 | ? e % tngst® fgypC ||° © Lutz, A« 1933. Beobachtmgen tteber Brasillanische Dicranocer- carlen, .Mem, Inst. Osw. Cruz, 27s 377-2*03, Lutz, A* 1935* 'Beobachtungen und betrachtungen ueber Cyathoco- •tylinen und Prohemistominen. Mem® Inst, Osw* Cruz 30(2) s ’S ,.j V • 169-180. • 1 a , * o ' * . Mathias, P, 1935?. Cycle evolutif d ’urt Ir&natode Holostomide■ (Cyathocotyls gravieri n. sp,) C. R, Acad* Sc, Paris 200s 1786-1788. • . ° ° ° : , O f! Ci o * . ° r-. v} . . a Haxon, M. 0, and Pequegnat,- W. E.1 9h9» Cercariae from Upper *• * * « m o Newport Bay. * J, Ent. Zool.* his 1-28.,a ° “s °o ° ° o * * * * * McCoy," 0. R«. 19.28., Further life history studies on marine ® ® * *, * « ■** , < ,o . ■ 8 , * . • * ° * “ trematodes. Carneg. Inst,. Hash. Yearbook 28s 290-292.’ ® ■ „ * * ' * , Mehra, H. P.* 19h7• Studies on the family Cyathocotylidae Poche * «. e „ / . » * o o 0 "I Part 2 ,t Proc. Mat. Acad.’Sci. India 17 s 1-52, % s „ 0 ® ' ® ' < *5 ^ e **’■' * Peter, C. T. ...1955.' Studies on the-c'ercarial fauna in Madras I. 63 « ® ® w * * o e 5 a a ® « © * 0 e . The furcocercous cercariae. Indian J. Yet. Sci. and Anim. Husb.;25:* 121-127. * 0 s ° ° a o s 9j s © e Peterson, H. * 1932. Cercarien der Niederelbe^. (Vorlaufige Mittei- ° ® <„ o lung). Zooi. Anz. 97: 13-27. ° 0 ’ °. >-03 * 0 ®*o . . ' . O • ° ffi © ® ® Porter.* A. 1938. The- larval trematode fo’und in certain *> South 0 © © ® ® 0 African mollusca with special reference to schistosomiasis# e 0 0 0 . 0 ® o South African Inst# Med# &es» 8s 11-1*92# ® . 0 ' G = s • fyemvati# 19^5# 'Cercaria multiplicata n* so* from the snail 0 . 0 ° -a & e ©• ■ Melanoldes tubereulatuso [Muller]# Jm Zool. Soe« India 7® o Prendel, J\.. R. 1930. Ein Reitrag zum Stadium der Helminthenfauna der Hunde in der U., Si S. R. (Sudliehe Ukraine). Zool Anz. 89s 323-326, * '• ; « ' ' J ■ ■ Price, E. W. 1928. Occurrence of Prohemistomum appendiculatum in the United States,1 J. Parasit. 15* 58- . : ■f > ’■ . " Price, E. If. 193k. Hew trematode parasites of birds.. Smithsonian ’ Misc. Coll. 91 (6): 1-6. e S .:,° •Sewell, H. B. S. 1922.* Cercariae Indicae. Indian .J. Med, Res, ’ 70: *1-370. 0 ° ° . l a \ ° ° 0 - ® , £ «> - ,t © ■T ' ® •'* , * Sewell, H. B. S. .1930, The evolution of the excretory system in • • ” " * * , certain groups ©f the furcocercbus cercaria. ‘Rec. Indian Mus. °o o * * f> Szidat, L. 1933. Ueber drei neue monostcme Gabelschwanzcercarien ‘ O ,— * * * » * " * 4 * der ostpretissichen Fauna. Ztsehr. Parasitenk. 5 (3—it) s V 0 ’ * 0 * »' 9 s * „ *> “ • Ut3-!i59. \ . oe' ° . C) • \. 3 3 / / - 8 * ,° ® ° ® *■, i* 0 4 * m ’ e» 'v <•> * 0 ® ft © Szidat, L. 1936. Parasiten atis Seeschvalbe’n. l. Uber neuo B « » , • * C). 0 0 cyathocotyliden aus dem barm von Sterna hirundo L» und> • « \ ° , 9 J ■ Sterna paradises. Ztsehr. Parasitenk. 6: 285-316. / ° * s •* „ * ■* 0 Stunkard, H. „ 1955. Freedom, Bondage, and the Welfare*State, “s 0° Science 121? 811-816. ^ , ° 0 °0 » ;o O G 0 ,s 0 ^ Ss 0 0 Tang, *C, C* I9kl# Morphologgr end life history of Prososfcephanus © » o Industrlus (Tubangui, 1922) lutz, l935*o Peking Hat. Hist* gun. .i s - 19-1,3.• •••• ° 3 ° ° ° @ SS ° ® ” . Tubangui, M. l9?8§ f,arvai trematodes from fhillippine snails.’8 o Phillip. ' Phillippine*Vertebrates IX. Philip. J. Sci. 75 s 131-li|2. ® Vernberg, W. B. 1952. Studies on the trematode family Cyathocb- tylidae Poche, 1926, with a description of a new species of * . Holostephanus from fish and the life history of Prohemistomum chandler! n. sp* J* Parasit. 38: 327-UO* Vfesenberg-Lund, C* 193k* Contributions to the Development of the Trematoda Digenea* Part II« The Biology of the freshwater o , ... o ’ o cercariae in Danish freshwaters. Mem* Acad. Roy* Sci, et o schwanzes. Zool* Jahrb. Abt. Anat. 1*6: 303-3^2* Yamaguti, S.' 1939* Studies on the helminth fauna of Japan. Part i; o 25* Trematodes of Birds IV. Jap. J. Zool. 8: 129?210# . * 0 ■ = Yamaguti,- S. 1?U0. 2ur Entwicklungsgeschichte von Cyathocotyle © ' © " “ ^ <•.) w ® v orientalis Faust, 1921* 'Ztsehr* Para“sitenk* 12: 78-83* o •® O . * * » . « PLATE I Fig. 1, The egg of M. kentuckiensis Fig. 2 Daughter' sporocyst of M. kentuckiensis containing germ balls and cercariae in various stages of development Fig. 3 Developing cercaria with three pair flame cells- Fig. U Developing cercaria with six pair flame dells in body, two pair in the tail Fig. $ The. cercaria of M. kentuckiensis. Dorsal view Fig. 6 Variations which occur in the canals anterior'to the. ascending trunks Fig. 7 View of posterior surface of furca showing furcal flagellets 88 © ® • ft * 8? « PLATE I & m * # o o PLATE II Fig. 1 The.metacercaria of M. kentuckiensis. Ventral view Fig. 2 Immature worm after four days in the chick. Ventral view Fig. 3 Adult_ of M. kentuckiensis. Dorsal view © © 0 90 * • • o o o O 0 * * * • • 91 PLATE E * U: ,v ® * m m m « ‘ * * PLATE III Fig. 1 Longitudinal section through hindbody of adult of M, kentuckiensis showing uterus and vaginal sphincter. X 3^0 . 92 © O o o PLATE III PLATE IV a • * Fig. 1 Cross section through the pharyngeal region of the adult • laterally. X 1$0 . Fig. 2 Cross section through the acetabular region of the adult of « ■* M. kentuckiensis. X l£0 9k Q Q © © o ® o PLATE IV o o PLATE V * * Fig. 1 Cross section through the region of the tribocytic organ of # the adult*of M. kentuckiensis showing the glandular tribocytic organ, some vitellaria, and the intestinal caeca. X 1^0 Fig. 2 Cross section through the region of the anterior testis of the adult of M. kentuckiensis showing the ascending and descending portions of the uterus ventral to the testis* X 1^0 o o o o 96 O O PLATE V o o © o O o 0 PLATE VI i# O 0 Fig. 1 Cross lection through the region of the anterior testis and »« * o the ovary of the adult of M. kentuckiensis. In addition, « O p ventral to the testis’is the anterior portion of the cirrus ' ® « • 0 sac, and ventral to the ovary is the ascending and descending liftibs of the uterus. The ascending limb contains an egg. « o ° X 1^0 * * 8 Fig. 2 Cross section through region of the’posterior testis of the ' ,, . * adult of M. kentuckiensis.1 Ventral to the testis are the muscular cirru6 sac and the descending limb of the uterus, X l£0 * ' 98 O PLATE VI o ° o AUTOBIOGRAPHY I, Donal Gene Myer, was born at Toledo, Ohio, on May U, 1930. . * I was graduated from Warren G. Harding High School in Warren, Ohio, * , * * in June, 19^7. Both my undergraduate and graduate training were received at The Ohio State University. I received the Bachelor of Science in 1951 and the degree Master of Science in 1953. In the * d Department of Zoology and Entomology, I held the positions of Graduate Assistant from 1952 to 1955, Assistant from 1955 to 1957, and Assistant Instructor from 1957 to 1958. I was admitted to candidacy for the Ph. D. in May, 1956. 100 o O 0 o of M. kentuckiensis. The cavity of the pharynx is flattened