CHROMOSOME STUDIES OF ICHTHYOCOTYLURUS PLATYCEPHALUS (CREPLIN, 1825) ODENING 1 9 6 9 WITH DESCRIPTION OF TRIPLOID VARIANT
AND COMPARATIVE KARYOLOGY OF THE GENUS ICHTHYOCOTYLURUS
STANEVIČIŨTÉ G.* & KISELIENÉ V.*
S u m m a r y :
This paper reports the first karyological study of trematode Ichthyocotylurus platycephalus (Creplin, 1825) Odening 1 9 6 9 . Chromosome number and morphology were studied in somatic cells of parthenites from intermediate host - mollusc Valvata piscinalis using an air drying technique and Giemsa staining. The karyotype consisted of 2 0 chromosomes comprising five pairs of subtelo-acrocentric (1st to 5t h), two pairs of submeta-subtelocentric (6t h-7t h), one submetacentric (9t h) and two pairs of metacentric chromosomes (8t h, 10t h). A supernumerary chromosome variation (presenceIabsence of B-chromosome) and occurrence of triploidy in one population of I. platycephalus are reported. In addition, the karyological analysis of Lithuanian population of l. variegatus was carried out. In the light of presented and previously obtained data comparative karyology of genus Ichthyocotylurus is discussed.
KEY WORDS : karyotype, Tremafoda, Ichthyocotylurus, triploidy, B- chromosomes.
Resume : Etude chromosomique de ICHTHYOCOTYLURUS PLATYCEPHALUS (creplin, 1825) odening 1969 avec la description de la forme triploïde et la caryologie comparée du genre ICHTHYOCOTYLURUS
Cette étude présente la première description du caryotype d'une espèce de Trématode, Ichthyocotylurus platycephalus (Creplin,
1825) Odening 1969 ainsi que le caryotype de I. variegatus non connu auparavant pour les populations lituaniennes. Le nombre et la morphologie des chromosomes ont été étudiés sur les cellules somatiques des formes larvaires obtenues à partir des hôtes intermédiaires, les mollusques Valvata piscinalis, par une méthode de suspension cellulaire et coloration au Giemsa. L'analyse effectuée a établi que le nombre diploïde de I. platycephalus varie de 20 à 21, à cause de la présence chez certains individus d'un chromosome surnuméraire, ou chromosome 6. La garniture chromosomique basique (les chromosomes A) comprend 10 paires dont cinq sont subtélo-acrocentriques (I à 5), deux subméta- subtélocentriques (6 et 7), une submétacentrique (9) et deux métacentriques (8 et 10). En outre, un cas de triploïde (2N = 3X
= 30) a été détecté chez un spécimen de I. platycephalus. Les données obtenues ont été comparées et discutées aves celles disponibles pour le genre Ichthyocotylurus.
MOTS CLÉS : coryotype, Trémolodes, Ichthyocotylurus, triploidie, chromosomes surnuméraires.
I N T R O D U C T I O N
T he s p e c i e s o f t h e g e n u s Ichthyocotylurus for a
l
o n g time h a v e b e e n c o n s i d e r e d as b e l o n g i n g t o t h e g e n u s Cotylurus (Szidat, 1 9 2 8 ) . M o r e r e c e n t investigations o n t h e life c y c l e s o f s p e c i e s o f Cotylurus s h o w e d that this g e n u s includes t w o groups o f s p e c i e s , differing from e a c h o t h e r in t h e b i o l o g y , life c y c l e a n d m o r p h o l o g y o f larval stages a n d t h e adults. O d e n i n g et. al. ( 1 9 6 9 ) created within it t w o subg e n e r a : Cotylurus (set o f hosts involved: P u l m o n a t a - snails a n d l e e c h e s - anatine bir ds) a n d Ichthyocoty-
lurus ( P r o s o b r a n c h i a - fishes - p i s c i v o r o u s b i r d s ) . N i e w i a d o m s k a ( 1 9 7 1 ) c o m p a r e d differences b e t w e e n t h e s e t w o s u b g e n e r a with differences existing a m o n g
* Institute o f Ecology, Lithuanian Academy of Sciences, Akade- mijos 2, Vilnius LT 2600, Lithuania.
Correspondence: Grazina Stanevičiũte.
Tel.: (370 2) 72 95 95 - Fax: (370 2) 72 92 57 - E-mail: helmi@ekoi.lt
the o t h e r g e n e r a within t h e family Strigeidae a n d e l e vated t h e m t o t h e rank o f valid g e n e r a .
T h e t a x o n o m i c status o f s p e c i e s within t h e g e n u s Ich- thyocotylurus h a s provided c o n s i d e r a b l e discussion. It is difficult t o define t h e set o f diagnostic features, w h i c h c o u l d well separate t h e s p e c i e s in various stage o f d e v e l o p m e n t . If validity o f Ichthyocotylurus (Coty- lurus) pileatus ( R u d o l p h i , 1 8 0 2 ) , I. (C.) erraticus (Rudolphi, 1 8 0 9 ) , I. (C.) platycephalus (Creplin, 1 8 2 5 ) w a s undoubtfull for m a n y systematisists ( O d e n i n g et.
al, 1 9 6 9 ; O l s o n , 1 9 7 0 ; Sudarikov, 1 9 8 4 ) , it w a s other
w i s e with I variegatus. D u b o i s ( 1 9 3 8 ) included I. (C.) variegatus as s y n o n y m o f I. (C.) pileatus. O d e n i n g &
B o c k h a r d t ( 1 9 7 1 ) after t h e elaborating o f c o m p l i c a t e d history a n d s y n o n y m y o f metacercaria a n d adult stated that "C. variegatus (= cumulitestis) is neither identical with C. platycephalus (= cuculus), n o r with C. pileatus just as little as with I. erraticus. I. variegatus is a s p e c i e s o f its o w n , differing from I. platycephalus".
O n e o f t h e r e c e n t efforts to elucidate the specific struc
ture o f genus w a s m a d e by D u b o i s ( 1 9 7 8 ) . O n the b a s e
Parasite, 2001, 8, 137-145
Mémoire 137
Article available athttp://www.parasite-journal.orgorhttp://dx.doi.org/10.1051/parasite/2001082137
STANEVICIUTÉ G. & KISELIENÉ V,
o f original material and m u s e u m samples h e stated the identity o f I. platycephalus and I. variegatus owing to insufficiency o f the differences recorded in cercariae o f I. platycephalus and I. variegatus for g o o d identifica
tion o f them. O n the contrary, Odening ( 1 9 7 9 ) affirmed the s p e c i e s o f I. platycephalus and I. variegatus as valid b a s e d o n e s e l f o n structure o f adults, cercariae, metacercariae and localization o f adults in the defini
tive hosts. But it is clear that morphological differences among the various stages o f species o f Ichthyocotylurus are rather vaguely distinctive. In many digenean spe
cies the larvae lack distinctive morphological features.
Recent studies have emphasised the value o f a multi- disciplinary approach in resolving t a x o n o m i c problems for various group o f organisms. Molecular methods can offer a n e w tool for larval-stage identification (Jousson et al, 1998). T h e karyotype is o n e o f the basic genetic characteristics o f eukaryotic species. In recent years, more than o n c e cytogenetic studies w e r e a g o o d sup
plement for morphological, biological and other cha
racteristics used for systematic analysis (Barsiene, 1993;
Insua etal, 1994). Cytogenetic parameters such as chro
m o s o m e number and morphology provide investigatory tools to define phylogenetic patterns and genetic struc
ture o f species and population (White, 1 9 7 3 ) .
Until n o w , c h r o m o s o m a l data w e r e restricted to three s p e c i e s o f Ichthyocotylurus-I. erraticus, I. pileatus and I. variegatus presenting different karyotypes with 2 n = 20 ( B a r s i e n e et al, 1 9 9 0 ; Bell et al, 1 9 9 8 ) .
I. platycephalus (Creplin, 1 8 2 5 ) is type s p e c i e s o f genus Ichthyocotylurus. This species as well as I. varie
gatus are fairly c o m m o n parasites o f fish in Lithuanian water bodies (Rauckis, 1 9 8 8 ) . T h e r e are recorded meta
cercariae o f four Ichthyocotylurus spp. as parasites o f fish in Lithuania. So far there are poorly investigated adult forms, parasiting birds o f Lithuania. Only adults o f I. platycephalus and I. pileatus are r e c o r d e d in Lithuanian water birds (Volskis & Pilipaviciute, 1 9 7 0 ) . Herein, w e present the karyotypical data o n I. platy
cephalus after conventional staining. A supernumerary c h r o m o s o m e variation and fact o f triploidy in p o p u lation o f I. platycephalus are reported. In the c o u r s e of a comparative investigation w e report also the karyo- logical analysis o f Lithuanian population o f I. variegatus.
MATERIALS AND METHODS
C
ercariae o f two s p e c i e s o f Ichthycotylurus w e r e identified in molluscs o f Lithuanian water bodies - I. platycephalus (Creplin, 1 8 2 5 ) O d e n i n g 1 9 6 9 and I. variegatus (Creplin, 1 8 2 5 ) O d e n i n g 1 9 6 9 . Naturally infected first intermediate hosts - the mollusc Val- vata piscinalis ( P r o s o b r a n c h i a ) w e r e c o l l e c t e d from May to October on 1994-1998. Parasite identification was
confirmed from the m o r p h o l o g y o f cercariae. Cercariae w e r e e x a m i n e d alive. Four s p e c i m e n s o f V. piscinalis with parthenites o f I. platycephalus out o f 3 2 8 (exten- siveness o f invasion - 1.22 % ) e x a m i n e d in the samples, w e r e collected from river Vilnele and w e r e used as a source o f the material for investigation. Also 3 2 2 spe
cimens o f molluscs V. piscinalis w e r e collected from the lake Asveja. Intramolluscan larval stages o f the Ich
thyocotylurus spp. isolated from 22 (extensiveness o f invasion - 6.83 % ) naturally infected molluscs were used for the cytogenetic analysis. T w o species o f Ichthyoco
tylurus have b e e n identified here - 1 , platycephalus and I. variegatus. 2 0 s p e c i m e n s w e r e infected with parthe
nites o f I. platycephalus ( 6 . 2 1 % ) and only two speci
m e n s ( 0 . 6 2 % ) - with parthenites o f I. variegatus.
Infected molluscs w e r e treated with 0.01 % c o l c h i c i n e in well water at 2 2 ° C for three to 15 hours. D i s s e c t e d digestive glands, containing sporocysts and developing cercariae w e r e transferred to distilled water for 4 0 - 5 0 minutes for h y p o t o n i c treatment, then fixed in freshly- prepared and cold solution o f e t h a n o l - acetic acid (3:1). Fixation involved three c h a n g e s ( 3 0 minutes, o n e hour and 24 hours) with storing at 4° C.
Each slide w a s m a d e from a single snail using an air- drying t e c h n i q u e (Kligerman & B l o o m , 1 9 7 7 ) with s o m e modification. Small pieces o f tissue w e r e removed from the fixative o n pre-cleaned slides, briefly s o a k e d in s o m e drops o f 4 5 % acetic acid, s m e a r e d and flame- dried by holding the slide over a b u n s e n burner. T h e n slides w e r e stained with Giemsa ( 4 %, pH 6.8) for 4 0 minutes. Phothomicrograps o f well spread meta- phases w e r e taken with an Amplival p h o t o m i c r o s c o p e under an oil-immersion system. For karyotyping, chro
m o s o m e s w e r e cut out o f the photomicrograps and paired o n the basis o f their size and centromeric posi
tion. Morphometric measurements o f c h r o m o s o m e s in the karyotypes o f 10 metaphasic cells o f e a c h species studied w e r e made. Absolute length in pm with stan
dard deviation w e r e calculated. T h e n relative length or percent total c o m p l e m e n t length w a s e x p r e s s e d as 100 times the absolute c h r o m o s o m e pair length divided by the total length o f the haploid c o m p l e m e n t . T h e length o f the B - c h r o m o s o m e was not included in the total l e n g t h o f h a p l o i d c o m p l e m e n t . C e n t r o m e r i c i n d e x e s w e r e calculated by dividing 100 times the length o f the short arm b y the total c h r o m o s o m e length.
C h r o m o s o m e m o r p h o l o g y was defined in a c c o r d a n c e with c h r o m o s o m e nomenclature o f Levan et al. ( 1 9 6 4 ) .
R E S U L T S
C
h r o m o s o m e data w e r e obtained from parthenites o f I. platycepbalus taken from 14 s p e c i m e n s o fV. piscinalis. T h e investigated s p e c i m e n s pré
s e n t e d diploid n u m b e r varying from 2 n = 2 0 to 21 d u e to p r e s e n c e o f extra small o r medium-sized sub- telocentric or submetacentric c h r o m o s o m e s considered as supernumeraries ( B s ) .
T h e b a s i c karyotype with 2 n = 2 0 was described from mitotic m e t a p h a s e s o f somatic cells o f I. platycephalus taken from eight s p e c i m e n s o f V. piscinalis (Fig. 1 ) . A diploid number was scored in 8 9 somatic cells (89-9 %).
Eight cells had an aneuploid n u m b e r o f c h r o m o s o m e s , two cells had a tetraploid set. T h e total length o f the haploid g e n o m e - 31-85 urn. C h r o m o s o m e s range in size from 1.72 to 5-19 ( m ( T a b l e I ) . T h e ls t- 5t h pairs o f c o m p a r a t i v e l y large c h r o m o s o m e s ( 6 6 - 6 7 % o f the total g e n o m e c o m p l e m e n t l e n g t h ) r e p r e s e n t e d the single-armed - subtelo-acrocentric g e n o m e units. Accor
ding to the c e n t r o m e r e position, h o m o l o g u e s o f pair 6t h and 7t h w e r e o b s e r v e d as submeta-subtelocentric, and those o f pair 9t h as submetacentric. T h e pairs 8t h
and 1 0t h are a clearly r e c o g n i z a b l e metacentric.
Six snails c o l l e c t e d in Asveja w e r e infected with par- thenitae o f I. platycephalus containing 21 c h r o m o s o m e s in the diploid sets o f 2 1 7 cells out o f 2 8 5 studied.
T h e m o r p h o l o g y o f this supernumerary c h r o m o s o m e w a s different in m e t a p h a s e plates from various indivi
duals (Fig. 1 ) . T h e small subtelocentric to submeta
centric supernumerary c h r o m o s o m e ( B s ) was found in c h r o m o s o m e sets o f I. platycephalus from four snails.
T h e medium-sized submetacentric supernumerary chro
m o s o m e w a s found in e x a m i n e d mitotic m e t a p h a s e s
o f I. platycephalus from two snails. T h e B s were clearly distinguishable from the basic c h r o m o s o m e s in the G i e m s a staining m e t a p h a s e s ( n o n - h o m o l o g o u s to the regular c h r o m o s o m e s ) . T h e m e a s u r e m e n t s o f chro
m o s o m e s o f basic set and additional c h r o m o s o m e s are given in T a b l e I.
W e have o b s e r v e d c h r o m o s o m a l preparation o f the intramolluscan stages o f Icbthyocotylurus prepared from o n e snail collected in Vilnele in which discovered that many o f cells c o n t a i n e d 3 0 instead o f the normal 20 c h r o m o s o m e s . C h r o m o s o m e numbers w e r e deter
m i n e d only from cells in w h i c h the entire cell was visible as in Figure 2. Thus, the karyotype was iden
tified as b e l o n g i n g to o n e cell and not due to an arte
fact o f the preparation. T h e 3 0 c h r o m o s o m e s w e r e c o m p a r e d with diploid (2 n = 2 0 ) set o f Icbthyocoty
lurus - the karyotype was similar to that o f I. platy
cephalus. T h e 3 0 c h r o m o s o m e s a p p e a r e d to consist o f three sets o f haploid (n = 1 0 ) I. platycephalus. 3 n = 3 0 was found in 2 3 ( 6 3 . 8 9 % ) mitotic metaphases and the remaining 13 ( 3 6 , 1 1 % ) being aneuploid - 3 n • 26-29- Parthenites o f the triploid forms was m o r p h o logically similar to that o f diploid.
C h r o m o s o m e s o f 116 mitotic metaphases o f somatic cells o f I. variegatus taken from two s p e c i m e n s o f V. piscinalis were examined (Fig. 3 ) . Counts o f 100 cells ( 8 6 . 2 1 % ) resulted in the diploid n u m b e r 2 n = 2 0 , 15 cells ( 1 2 . 9 3 % ) resulted as hypoploid - 2 n - 17- 19 and only o n e cell has a hiperploid n u m b e r o f chro-
Fig. 1. - Mitotic metaphases and karyo
types of karyomorphs of Icbthyocoty- hirus platycephalus with 2 n = 20 (a) and 2n=21 (b, c ) . Scale bar = 10 um.
C H R O M O S O M E S O F ICHTHYOCOTYLURUS PIATYCEPHALUS
Parasite, 2001, 8, 137-145
Mémoire 139
STANEVICIUTÉ G. & KISELIENÉ V.
Chromosome Aboslute length Relative length
number (um) (%) Centrometric indices Classification
1 A 5.19 ± 1.02 16.22 ± 0.96 13.39 ± 1.71 st-a
B 5.49 ± 0.84 17.05 ± 0.80 12.03 ± 1.36 st-a
2 A 4.72 + 0.89 14.79 ± 0.64 13.04 ± 2.08 st-a
B 4.81 ± 0.81 14.93 ± 1.00 12.39 ± 3.16 st-a
3 A 4.22 ± 0.78 13.22 ± 0.74 12.63 ± 2.56 st-a
4.20 ± 0.45 13.09 ± 0.52 13.05 ± 1.86 st-a
4 A 3.88 ± 0.70 11.90 ± 0.81 12.55 ± 2.89 st-a
B 3.86 ± 0.55 12.01 ± 0.44 13.64 ± 2.45 st-a
5 A 3.44 ± 0.79 10.69 ± 0.85 12.79 ± 2.96 st-a
B 3.56 ± 0.48 11.06 ± 0.42 11.39 ± 2.24 a-st
6 A 2.61 ± 0.42 8.23 ± 0.32 28.52 ± 4.20 sm-st
B 2.31 ± 0.28 7.21 + 0.39 26.15 + 5.52 sm-st
7 A 2.12 ± 0.31 6.73 ± 0.76 28.50 ± 4.03 sm-st
B 2.10 ± 0.23 6.64 ± 0.47 27.87 ± 3.02 sm-st
H A 2.02 ± 0.33 6.37 ± 0.40 45.74 ± 1.73 M
B 1.99 ± 0.17 6.23 + 0.35 45.01 ± 1.89 m
9 A 1.93 ± 0.23 6.12 + 0.52 32.23 ± 3.25 sm
B 1.96 ± 0.16 6.16 ± 0.31 34.77 ± 2.88 sm
m A 1.72 ± 0.18 5.47 ± 0.61 46.60 ± 2.53 m
B 1.81 ± 0.19 5.68 - 0.46 47.41 ± 1.20 m
Bs(b) 2.97 ± 0.18 37.66 ± 5.21 sm-m
Bs(c) 1.91 ± 0.19 23.33 4.17 st-sm
Abbreviations: A, measurements o f karyomorph with 2 n = 20; B, measurements of karyomorph with 2 n = 21; Bs(b), Bs(c), variation of supernumerary chromosomes; m, metacentric; sm, submetacentric; st, subtelocentric; a, acrocentric chromosomes.
Table I. - Measurements (means ± SD) and classification of two karyomorhs of Ichthyocotylurusplatycephalus.
Fig. 2. - Mitotic metaphase and karyotype of triplod form of Ichthyocotylurus platy
cephalus with 3 n = 29. Scale bar = 10 um.
C h r o m o s o m e Absolute length Relative length
n u m b e r ( u m ) (%) C e n t r o m e t r i c Indices Classification
1 5.88 ± 1.56 16.23 ± 0.82 47.35 ± 2.09 m
2 5.48 ± 1.50 15.09 ± 0.48 12.56 ± 3 1 1 st-a
3 4.88 ± 1.32 13.46 ± 0.65 11.90 ± 2.66 a-st
4 4.29 ± 1.10 11.86 ± 0.48 13.66 ± 3.48 st-a
5 3.85 ± 0.88 10.53 ± 0.78 12.91 ± 2.23 st-a
6 2.83 ± 0.75 7.81 ± 0.35 28.46 ± 4.38 S i l l
7 2.52 ± 0.62 6.95 ± 0.49 26.54 ± 3.21 sm-st
8 2.36 ± 0 . 6 1 6.51 ± 0.39 35.96 ± 3.41 sm-m
9 2.14 ± 0.47 5.96 ± 0.45 46.39 ± 2.73 m
10 1.95 ± 0.45 5.42 ± 0.37 45.52 ± 1.89 m
Abbreviations: m, metacentric; sm, submetacentric; st, subtelocentric; a, acrocentric chromosomes Table II. - Measurements (means ± SD) and classification of chromosomes of Ichthyocotylurus variegatus.
Fig. 3- - Mitotic metaphase and karyotype of Lithuanian population of Ichthyocotylurus variegatus. Scale bar = 10 um.
m o s o m e s . T h e total length o f the haploid g e n o m e - 3 6 . 1 8 pm. C h r o m o s o m e s range in size from 1.95 to 5.88 pm (Table II). T h e karyotype consisted o f o n e ( 1s t) pair o f large and two small ( 9t h and 1 0t h) pairs o f meta
centric c h r o m o s o m e s ; o n e pair ( 6t h) o f submetacentric a n d ( 8t h) s u b m e t a - m e t a c e n t r i c c h r o m o s o m e s ; four m o n o - a r m e d (pairs 2-5 represent subtelo-acrocentic units). C h r o m o s o m e s o f 7t h pair represented b y sub- meta-subtelocentic c h r o m o s o m e s .
D I S C U S S I O N
O
ur current knowledge on cytogenetics o f various s p e c i e s o f family Strigeidae reveals o n e type o f c h r o m o s o m e c o m p l e m e n t with a clear distinction b e t w e e n five comparatively large and five smaller c h r o m o s o m e pairs (Barsiene, 1993; Petkeviciute
& Staneviciute, 1 9 9 9 ) . T h e s a m e pattern o f karyotype is characteristic also for most o f karyologically studied representatives o f family Diplostomidae ( B a r s i e n e &
Staneviciute, 1 9 9 1 ; B a r s i e n e , 1 9 9 3 ; Staneviciute et al., 1 9 9 8 ) . In recent years karyologically investigated three species o f Ichthyocotylurus presented the same diploid n u m b e r 2 n = 2 0 , but s h o w e d three different karyo
types ( B a r s i e n e et al, 1 9 9 0 ; Bell et al, 1 9 9 8 ) . For the first time reported here karyotype o f I. platycephalus display a typical strigeid pattern with 20 c h r o m o s o m e s , arranging into two distinct size groups. For the c o m parative karyology the idiograms o f all k n o w n karyo
types o f g e n u s Ichthyocotylurus w e r e c o n s t r u c t e d (Fig. 4 ) , b a s e d o n the m e a n values o f relative length and centromeric indices presented in T a b l e s I and II, as well as previously published data ( B a r s i e n e et al., 1990; Bell et al, 1 9 9 8 ) . T h e subtelocentric type is dominant in the first group o f c h r o m o s o m e s . Groups o f the small g e n o m e units are c o m p o s e d dominantly o f submeta- and metacentric c h r o m o s o m e s . Similarity o f c h r o m o s o m e relative length a m o n g the s p e c i e s s h o w e d that this parameter cannot well discriminate inside the genus. In all s p e c i e s studied, a m o r e signi
ficant difference was found for centromeric index bet
w e e n corresponding c h r o m o s o m e pairs. T h e conside
rable interspecific differences w e r e o b s e r v e d in the m o r p h o l o g y o f the large elements. T h e large pericen
tric inversions o c c u r r e d in the process o f speciation o f Ichthyocotylurus rather frequently ( 1s t pair o f I. varie
gatus, I. erraticus, 2n d pair o f I. erraticus). Such large pericentric inversion in the process o f speciation within
CHROMOSOMES OF ICHTHYOCOTYLURUS PLATYCEPHALUS
Parasite, 2001, 8, 137-145
Mémoire 141
Chromosome pair number
Fig.4. - Idiograms representing the haploid sets of genus Ichthyocotylurus: a, I. platycephalus; b. I. pileatus (data of BarSiené et al, 1990);
c, I. variegatus; d, I. variegatus (data of Bell et al., 1998); e, I. erraticus (data of BarSiené et al, 1990); f, I. erraticus (data o f Bell et al, 1998).
the g e n u s are registered in the families Diplostomidae (Diplostomum spathaceum), Strigeidae (Apatemon gra
cilis) (Barsiene & Staneviciute, 1991; Petkeviciute & Sta- neviciute, 1999). C h r o m o s o m e complements o f I. varie
gatus and I. erraticus possess a pair o f large metacentric c h r o m o s o m e s , at that time I. platycephalus and I. pilea
tus have not such pair o f c h r o m o s o m e s . S o , c h r o m o s o m e sets o f I. variegatus - I. erraticus and I. platy
cephalus - I. pileatus s e e m respectively closer. Species I. platycephalus and I. pileatus s h o w slight interspecific differences in the relative length. T h e difference bet
w e e n the length o f the longest and the smallest pair in I. platycephalus r e a c h e s three times, m e a n w h i l e in I. pileatus this difference r e a c h e s four times. T h e first five pairs o f c h r o m o s o m e s o f I. platycephalus consti
tute 66-67 %, corresponding c h r o m o s o m e s o f I. pileatus - 7 0 - 7 1 % o f the total c o m p l e m e n t length. Compara
tive cytological analysis ( b y Student's t-test, P < 0 . 0 5 ) o f these s p e c i e s demonstrated statistically significant differences in the Ic values in c h r o m o s o m e pairs 5, 7 and 9.
Geographical karyotypic variation w a s found in karyo
type o f I. variegatus. Bell et al. ( 1 9 9 8 ) has studied the k a r y o t y p e o f I. variegatus population in Scotland.
Although n o differences w e r e revealed in the measu
rements o f the relative length o f c h r o m o s o m e s , the t w o populations differ in the position o f c e n t r o m e r e in c h r o m o s o m e pair 7 ( Ic= 2 6 . 5 4 according o u r data and Ic = 3 9 . 4 according data from S c o t l a n d ) . S u c h remar
kable difference m a y b e c a u s e d b y small c h r o m o s o m e rearrangement, most p r o b a b l y pericentric inver
sion. Slight differences w e r e found in the Ic values o f three, four and five pairs o f subtelocentric c h r o m o s o m e s , w h i c h w e associated with the variability o f the short arms and with their small dimensions exactly like subjectivity in the metrical evaluation o f c h r o m o s o m a l characteristics. Small c h a n g e s m a y arise in the c e n t r o - meric index values due to probable differential conden
sation o f c h r o m o s o m e s . C h r o m o s o m e c o n d e n s a t i o n is a d y n a m i c p r o c e s s that p r o c e e d s at different rates along the length o f e a c h c h r o m o s o m e ( F r a n c k e &
Oliver, 1 9 7 8 ) . T h e variation in the c e n t r o m e r i c i n d e x values m a y b e greater in c h r o m o s o m e s with unequal arms. R e c o r d e d distinct difference o f Ic values o f 2n d
pair b e t w e e n I. eiraticus from North-West Chukotka (Barsiene et al, 1 9 9 0 ) and Scotland populations w a s interpreted b y authors as interpopulation variation from geographically distant samples or i n a d e q u a n c y and not conspecificity o f investigated s p e c i e s (Bell et al.. 1 9 9 8 ) .
STANEVICIUTE G. & KISELIENE V.
CHROMOSOMES OF ICHTHYOCOTYLURUS PLATYCEPHALUS
Since o n l y recently g e n u s Ichthyocotylurus was e l e vated to the rank o f valid g e n e r a ( N i e w i a d o m s k a , 1 9 7 1 ) it is interesting to c o m p a r e c y t o g e n e t i c data o f this g e n u s and genus Cotylurus. S o far, c h r o m o s o m e n u m b e r s are reported for four s p e c i e s o f genus Coty
lurus (Petkeviciute, 1992; Barsiene, 1 9 9 3 ) . All s p e c i e s p r e s e n t e d the typical strigeid karyotype with diploid c h r o m o s o m e n u m b e r 2 n = 2 0 . T h e uniform generic marker has not b e e n singled out in the c h r o m o s o m e set structure among investigated species o f genus Coty
lurus and Ichthyocotylurus. T h e karyotypes o f genus Cotylurus m o r e frequently are notable for the preva
l e n c e o f uniarmed c h r o m o s o m e s in b o t h groups o f g e n o m e e l e m e n t s . With data n o w available it can b e stated that trematodes are karyologically conservative and their karyotypes tend to have the s a m e n u m b e r and closely related gross c h r o m o s o m e m o r p h o l o g y o n the genus and e v e n o n family t a x o n o m i c level (Bar
siene, 1 9 9 3 ; Staneviciute, 1 9 9 4 ) .
It should b e m e n t i o n e d an interesting and rather rare fact o f triploidy in the metaphase plates from o n e snail infected with parthenites o f I. platycephalus. In the Tre- matoda there are few reports o f triploid organism. Tri
ploidy have b e e n reported for Schistosomatium dou- thitti - 3 n = 21 (Short & Menzel, 1 9 5 9 ) , Fasciola sp. - 3 n = 3 0 (Sakaguchi & Nakagawa, 1 9 7 5 ) , Paragoni- mus westermani - 3 n = 3 3 ( S a k a g u c h i & Tada, 1976;
Hirai et al, 1 9 8 5 ) , Allocreadium fasciatusi- 3 n = 21 (Ramanjaneyulu & Madhavi, 1 9 8 4 ) , Apatemon minor - 3 n = 3 0 (Petkeviciute, 1 9 9 2 ) , Diplostomum sp. - 3 n
= 3 0 (Staneviciute, 1 9 9 4 ) , Schistosoma mansoni - 2 n = 24 (Hirai & LoVerde, 1989). T h e presented data sugges
ted that natural triploids might not b e so u n c o m m o n . T h e observation o f C-banding patterns (Hirai et al..
1 9 8 5 ) suggested that the triploid P. westermani form was allotriploid, which means composed o f two g e n o m e set introduced from P. westermani ( 2 n ) and another unidentified set. Triploidy in A. fasciatusi (Ramanja
neyulu & Madhavi, 1 9 8 4 ) w a s a result o f duplication o f the c h r o m o s o m e sets rather than fragmentation o f the c h r o m o s o m e s . Short & M e n z e l ( 1 9 5 9 ) suggested that the p r e s e n c e o f triploid g e n o m e s implied that a diploid c o m p l e m e n t was present in the egg before fer
tilization and argued strongly in favour o f diploid virgin eggs as a result o f meiotic irregularity. In the report o f Hirai & LoVerde ( 1 9 8 9 ) triploid S. mansoni s e e m s to b e determined by either an abnormal egg or s p e r m resulted from nondisjunction during meiotic division and formed a zygote with either a normal sperm or egg. S o the origin o f triploidy in most c a s e s it is c o n s i d e r e d to b e the result o f fertilization (diploid egg o r s p e r m ) , then a miracidial e m b r y o with triploid constitution could b e produced. Successful infection o f a snail with such a miracidium would result in the pro
duction through asexual division o f thousands o f cer- carial e m b r y o s (Hirai & LoVerde, 1 9 8 9 ) .
B - c h r o m o s o m e s are e l e m e n t s extra to the normal c o m p l e m e n t (A c h r o m o s o m e s ) w h i c h h a v e b e e n found in all major groups o f plants and animals and vary in n u m b e r s b e t w e e n individuals o f a s p e c i e s ( C a m a c h o et al, 1 9 9 7 ) B e u k e b o o m etal, 1 9 9 8 ) . It has b e e n estimated that they o c c u r in 1 0 - 1 5 % o f euka- ryotes ( B e l l & Burt, 1 9 9 0 ) . T h u s , it is likely that many m o r e s p e c i e s , w h e n analysed with sufficient intensity, will b e found to p o s s e s s B s . T h e function and c o m position o f B s is still a controversial question. Their frequencies in populations are determined b y their transmission rates and effects o n host fitness. In most cases, they appear to have negative effects o n the phe- notypic fitness o f their host w h e n they o c c u r in high number. There are s o m e models concerning the effects o f B s at l o w n u m b e r s ( B e u k e b o o m et al., 1 9 9 8 ) . Under the heterotic m o d e l B s are c o n s i d e r e d to h a v e positive fitness effects (White, 1 9 7 3 ) . T h e parasitic m o d e l a s s u m e s that B s are g e n o m i c parasites ( B e l l &
Burt, 1 9 9 0 ; B e u k e b o o m et al, 1 9 9 8 ) . C a m a c h o et al.
( 1 9 9 7 ) considers the B s in the g r a s s h o p p e r Eyprepoc- nemis plorans to b e c l o s e to the selective neutrality ("near-neutral" m o d e l ) .
Reports on the o c c u r r e n c e o f B - c h r o m o s o m e s ( B s ) in T r e m a t o d a are still scarce. Until n o w s p e c i e s with occuring B s are c o m p o s e d aproximately 5 % o f all cytogenetically k n o w n s p e c i e s o f Trematoda. T h e B s w e r e recently described in three studied s p e c i e s o f the genus Apatemon (Barsiene, 1 9 9 3 ; Petkeviciute & Sta
neviciute, 1 9 9 9 ) , also in s o m e s p e c i e s o f g e n e r a Noto- cotylus (Petkeviciute & Barsiene, 1 9 8 8 ; Petkeviciute et al., 1989a; Barsiene, 1 9 9 3 ) , Diplodiscus (Petkeviciute et al., 1 9 8 9 b ) , Echinostoma (Barsiene, 1 9 9 3 ) . T h e diploid n u m b e r variation in s p e c i e s o f Trematoda w e r e deter
mined b y including m o r e frequently o n e or two (rare m o r e ) B s o f different m o r p h o l o g i e s and sizes (from rather large metacentric to small a c r o c e n t r i c ) in the cells. In most c a s e s B s w e r e mitotically stable.
In summary, exhibited distinct differences in the chro
m o s o m e m o r p h o l o g y o f the investigated Ichthyocoty
lurus spp. s h o w e d these as s p e c i e s specific. Species differ mainly by centromeric index b e t w e e n corres
p o n d i n g c h r o m o s o m e pairs and such differences let support the assumption about c h r o m o s o m e rearran
g e m e n t involved in their karyotypic evolution, i.e.
m e c h a n i s m s not affecting the c h r o m o s o m e n u m b e r - pericentric inversions, small translocations and dele
tions. Presented data o f cytological investigation o f Ich
thyocotylurus spp. obviously negate the posibility o f s y n o n y m y of I. variegatus and I. pileatus as well as I. variegatus and I. platycephalus and contradict with sta
tement o f Dubois concerning identity o f them (Dubois, 1938, 1 9 7 8 ) . At that time these data support the point o f v i e w o f O d e n i n g c o n c e r n i n g the validity o f these s p e c i e s ( O d e n i n g & Bockhardt, 1 9 7 1 ; Odening, 1 9 7 9 ) . Our data could serve as a useful start to further inves-
Parasite, 2001. 8. 137-145
Mémoire 143
STANEVICIUTE G. & KISELIENE V.
tigations together with molecular, m o r p h o l o g i c a l a n d e c o l o g i c a l analyses o f g e n u s Ichthyocotylurus.
ACKNOWLEDGEMENTS
W
e wish t o thank Dr. R. Petkeviciute (Institute of E c o l o g y ) for assistance in t h e collection of material a n d for critically reviewing t h e manuscript a n d Dr. V. Stunzenas (Institute o f E c o l o g y ) for helping in preparing t h e manuscript.R E F E R E N C E S
BARSIEXÉ J . The karyotypes of trematodes. Academia, Vilnius, 1993.
BARSIEXÉ J . , PETKEMCILTÉ R . , STAXEVICIÛTÉ G . & Orlovskaya O.M. Karyological investigations of trematodes of the families Notocotylidae, Echinostomatidae and Strigeidae of North West Chukotka. Parazitologiya, 1990, 24, 3-10.
BARSIENÉ. J . & STANEVICIÙTÉ G . A comparative karyological study of trematodes within the genus Diplostomum. Hel-
minthologia, 1991, 28, 31-36.
BELL A . S . , SOMMERYILLE C. & GIBSON D . I . Karyological studies on three strigeid digeneans: Ichthyocotylurus erraticus (Rudolphi. 1809). Ivariegatus (Creplin, 1825) and Apa- temon gracilis (Rudolphi, 1819). Systematic Parasitology, 1998, 41, 169-178.
BELL G. & BURT A. B-chromosomes: germ-line parasites which include changes in host recombination. Parasitology, 1990, 100, S19-S26.
BEUKEBOOM L.W., SEIF M., PLOWMAN A . B . , DE RIDDER F . &
MICHIELS X.K. Phenotypic fitness effects of B chromo
somes in the pseudogamous parthenogenetic planarian Polycelis nigra. Heredity, 1998, 80, 594-603.
CAMACHO J.P.M., SHAW M.W., LOPEZ-LEON M.D., PARDO M.C. &
CABRERO J. Population dynamics of a selfish B chromosome neutralized by the standard genome in the grasshopper Eyprepocnemis plorans. The American Naturalist, 1997, 149, 6, 1030-1050.
DUBOIS G. Monographie des Strigeid (Trematoda). Mémoires de la Société Neuchateloise des Sciences Naturelles, 1938, 6, 1-535.
DUBOIS G. A propos des matériels originaux de Cotylurus pla- tycephalus et de Cotylurus variegatus (Creplin, 1825) (Tre
matoda: Strigeidae). Annales de Parasitologie Humaine et Comparée. 1978, 53. 53-62.
FRANCKE U. & OLIVER N. Quantitative analysis of high-reso
lution trypsin-Giemsa bands on human prometaphase chromosomes. Human Genetics, 1978, 45, 137-165.
HIRAI H . , SAKAGUCHI Y . , HABE S. & IMAI H.T. C-banding ana
lysis of six species of lung flukes, Paragonimus spp. (Tre
matoda: Platyhelminthes) from Japan and Korea. Zeischrift fur Parasitenkunde, 1985, 71, 617-629.
HIRAI H. & LOVERDF. P. Triploid cells found in intramolluscan stages of Schistosoma mansoni. The Journal of Parasito- logy, 1989, 75, 5, 800-802.
INSUA A., LABAT J . P . & THIRIOT-QLTEVREUX C. Comparative ana
lysis of karyotypes and nucleolar organizer regions in dif
ferent populations of Mytilus trossulus, Mytilus edulis and Mytilus galloprovincialis. Journal of Molluscan Studies, 1994, 60, 359-370.
JOUSSON O . , BARTOLI P . , ZANINETTI L. & PAWLOWSKI J . Use of ITS rDNA for elucidation of some life-cycles of Mesome- tridae (Trematoda, Digenea). International Journal for Parasitology, 1998, 28, 1403-1411.
KLIGERMAX A.D. & BLOOM E. Rapid chromosome preparations from solid tissues o f fishes, fournal of the Fisheries Research Board of Canada, 1977, 34, 266-269.
LEVAN A., FREDGA K . & SANDBERG A.A. Nomenclature for cen- tromeric position on chromosomes. Hereditas, 1964, 52, 201-220.
NIEWIADOMSKA K . On the necessity o f dividing the genus Cotylurus Szidat, 1928 into to valid genera Cotylurus Szidat, 1928 and Ichthyocotylurus Odening, 1 9 6 9 (Stri
geidae). Acta Parasitológica Polonica, 1971, 19, 8, 113- 120.
ODENING K . À propos de la validité d'Ichthyocotylurus varie- gatus (Creplin, 1825) et de la position spécifique de Tetra- cotyle ovata (v. Linstow, 1877) (Trematoda: Strigeidae).
Annales de Parasitologie Humaine et Comparée, 1979, 54, 2, 171-183.
ODENING K . , MATTHEIS T. & BOCKHARDT I. Status und lebens- zyklus des trematoden Cotylurus platycephalus. Ange-
wandte Parasitologie, 1969, 10, 2, 76-80.
ODENING K. & BOCKHARDT I. Der lebenszyklus des trematoden Cotylurus variegatus im Spree-Havel-Seengebiet. Biolo- gistes Zentralblatt, 1971, 90, 49-84.
OLSON R . E . The life cycle of Cotylurus erraticus (Rudolphi, 1809) Szidat, 1928 (Trematoda: Strigeidae). The fournal of Parasitology, 1970, 56, 1, 55-63.
PETKEVICIÛTÉ R. Karyological investigations of some species of trematodes (taxonomical and phylogenetical aspects).
PhD Thesis, University of St. Petersburg, 1992.
PETKEVICIÛTÉ R. & BARSIEXÉ J . The comparative karyological analysis of three species o f trematodes of genus Notoco-
tylus. Parazitologiya, 1988, 22, 21-28.
PETKEVICIÛTÉ R . & BARSIEXÉ J . & MAZEIKA V. Cytogenetic cha
racteristics o f Notocotylus noyeri Joyeux, 1922 (Trema
toda, Notocotylidae). Acta Parasitológica lituanica, 1989a, 23, 9 3 - 9 8 .
PETKEVICIÛTÉ R . , KISELIEXÉ V. & STEXKO R . P . Cytogenetic ana
lysis of two populations of Diplodiscus subclavatus (Tre
matoda, Diplodiscidae). Parazitologiya. 1989b. 489-495.
PETKEVICIÛTÉ R. & STANEVICIÙTÉ G. Karyotypic characterization of Apatemon gracilis, fournal of 'Helmitithology, 1999, 73, 73-77.
RAMANJANEYULU J.V. & MADHAVI R. Cytological investigations of two species of Allocreadiid trematodes with reference to the occurrence of triploidy and parthenogenesis in Allocreadium fasciatusi. International fournal for Parasi- tology, 1984, 14, 309-316.
RAUCKIS E . Fish parasites in Lithuanian waters. Mokslas, Vil
nius, 1988, 208 p.
S A K A G U C H I Y. & N A K A G A W A C. A note on the chromosome of the common liver fluke (Fasciola sp.) from Japan. Chro- mosome Information Service, 1975, 19, 20-21.
S A K A G U C H I Y. & T A D A I. Chromosomes of a lung fluke, Para- gonimus westermani. Chromosome Information Service,
1976, 20, 23-24.
S H O R T R.B. & M E N Z E L M.Y. Chromosomes in parthenogenetic
miracidia and embryonic cercariae of Schistosomatium douthitti. Experimental Parasitology, 1959, 8, 249-264.
S T A N E V I C I Û T É G. Karyological investigations of trematodes of
order Strigeiformes La Rue, 1926. PhD Thesis, Institute of Ecology, Vilnius, 1994.
S T A N E V I C I Û T É G., P E T K E V I C I Û T É R. & K I S E L I E N É V. Karyological analysis of two species of genus Posthodiplostomum Dubois, 1936 (Trematoda: Diplostomidae) with remarks on the karyological evolution of Diplostomidae. Acta Zooló- gica Lituanica. Parasitología, 1998, 8, 1, Al-Al.
S u D A R i K O v V.E. Trematodes of fauna of USSR. Strigeids.
Nauka, Moscow, 1984.
V o L S K i s G. & P I L I P A V I C I Ü T É J . On the infection of the Lithua
nian water birds by Strigeidae (Trematoda). Acta Parasi- tológica Lituanica, 1970, 10, 47-58.
W H I T E M.J.D. Animal Cytology and Evolution. Cambridge Uni
versity Press, Cambridge, 1973.
Reçu le 27 octobre 2000 Accepté le 15 janvier 2001
CHROMOSOMES OF ICHTHYOCOTYLURUS PLATYCEPHALUS
Parasite, 2001, 8, 137-145
Mémoire 145
