Table Of ContentGreatBasin Nahiralist55(3),© 1995,pp.258-266
MORPHOLOGICAL AND HOST-SYMBIONT STUDIES OF TRICHODINA
TENUIFORMIS ANDAPIOSOMA CAMPANULATUM INFESTING MOTTLED
SCULPIN {COTTUS BAIRDI) FROM PROVO RIVER, UTAH
YingQi' and KicluirdA. Heckmannl'2
—
Abstiuct. TrichocliiKi teintijonnis Stein, 1979 andApiosomci cciinpanitlatiiin Tiinoteev, 1962 were found on j^ills of
mottled sculpin {Cotttis bairdi) from two locations in the Provo River, UT. They were studied by light and electron
optics. Dimensions and morphologyoftheadhesive discanddenticles ofT. tenuifonnis were differentiated from other
Trichodmaspecies.A. campanulatumwascharacterizedbyitsspindle-shapedcellbody. Finefeaturesexaminedbyscan-
ningelectron microscopyincludedbodyshape,pellicle, elements oftheadhesivedisc, aboralciliaiycomplex, andado-
ral ciliary spiral. Histopathological studies suggested that the organisms are ectocommensals. Ecological aspects of
organism infestation between two areas were also investigated. This report establishes a new host and distribution
recordforthesetwospeciesinmottledsculpinfromtheProvoRiver, UT.
Keywords:Trichodinatenuiformis,Apiosomacampanulatum,Cottusbairdi,morphology, host-syinbiontrelationship,
ecologicalaspects. ProvoRiver
High numbers of two ciliated protozoa, may cause to the host, and (3) evaluate the sea-
Trichodina and Apiosoma, were encountered sonal infestation rate to provide ecological infor-
on the gills of mottled sculpin {Cottus bairdi) mation forthe listed ciliates and theirhost.
during a study of ectoparasites of fishes from
the Provo River Materials and Methods
Trichodina is a mobile ciliate belonging to
the subclass Feritrichia, family Trichodinidae Studies were earned out in late summerand
(Lorn and Dykova 1992). This protozoan has fall (August, October 1993), late winter and
an adhesive disc characterized by very promi- spring (March, May 1994). Watertemperatures
nent and taxonomically significant denticles in the Frovo River ranged from 14°C to 4°C
(Van As and Basson 1987). More than 140 and 6°C to 10°C, respectively. One hundred
species ofTrichodina have been reported from si.xty sculpin were collected from two sites:
wild, cultured, and laboratoiy fishes in many one in the city ofFrovo (Utah County) munici-
parts oftheworld (Rand 1993). pal area, the second in a relatively pristine
Sessile peritrich ciliates ofthe genus Apio- region near the Jordanelle Reservoir (Wasatch
soma (syn. GlossateUa) belong to the subclass County). Sculpin were collected using elec-
Feritrichia, family Epistylididae (Lom and trofishing, placed in buckets containing river
Dykova 1992). They are generally attached to water, transported to the laboratory, and exam-
fish by a scopula (Lom 1973). They have been inedwithin 24 h aftercapture.
largely neglected by fish parasitologists until For light microscopy, air-dried smears of
recently, when more attention has been given gill filament scrapings were prepared from
to this group. freshlykilled fish and treatedby Klein's diy sil-
Many species ofthese two ciliated protozoa ver impregnation technique (Clark and Heck-
have been investigated (Arthur and Margolis mann 1984) to examine components of the
1984, Cone and Odense 1987, Rand 1993); adhesive disc. Other smears were prepared,
however, a detailed study on mottled sculpin fixed, air-dried, and stained with iron hema-
has never been reported. Objectives of this toxylin (Carcia and Bruckner 1988) to obsen'e
studywere to (1) incoiporate differentlevels of the position and structure of the macro- and
microscopy to study ciliate structure, (2) ob- micronuclei. Sections ofinfested gills from the
sei-ve histopathological changes these protozoa spring sample were fixed, blocked, cut, and
'Ofpartmcntol'Zoology',BrighamYoungUniversity,Provo,UT84602.
^Autliortowhomcorrespondenceshouldbeaddressed.
258
1995] Ciliated Protozoa in Mottled Sculpin 259
stained with hematoxylin-eosin (Garcia and Micronucleus in -Y position (Lom 1958)
3x2
Bruckner 1988) forhistopathological studies. observed in six specimens, dimension
For scanning electron microscopy, gills of (Fig. 3). —
freshly killed fish were fixed in 2% buffered Scanning electron microscopy. Body
glutaraldehyde, followed by repeated washes of T. tenuiformis circular in aboral view and
in asodium cacodylate bufferandpost-fixed in aboral surface relatively flat (Fig. 4). Body
a 1% solution of osmium tetroxide. After that bell-shaped or domed when viewed from the
they were washed in the same buffer system. side (Fig. 5).
Specimens were dehydrated through a graded The aboral ciliaiy complex consists ofthree
alcohol series and critical-point-dried and distinct ciliary bands: the basal ciliary ring,
sputter-coated with gold for examination with locomotor ciliaiy wreath, and marginal ciliary
a Joel-840 high-resolution scanning electron ring. The basal ciliary ring, adjacent to the
microscope. border membrane, has a single row of fine,
For transmission electron microscopy, after distally tapering cilia 1-2 /xm long. Separated
fixation and dehydration, gills were embedded from the basal ciliary ring by the basal septum
in Spurrresin and sectionedwith aglass knife. is the locomotor ciliaiy wreath, which is com-
Each section was stained with lead citrate and posed of numerous rows of well-developed,
examined with a Philip EM400 transmission powerful cilia 2-3 fim long whose primary
electron microscope. function is locomotion. The precise number of
Terminology and methods ofmeasurement ciliaiy rows composing this wreath could not
follow those given bv Lom (1958), Lorn and be ascertained. It is separated anteriorly from
Dykova (1992), Wellborn (1967), Arthur and the marginal ciliary ringby apoorly developed
Margolis (1984). Measurements are in micro- anterior septum that is evident only when the
meters (/xm) and are based on 30 specimens aboral ciliary complex is uncovered by the
for each species from each of the four sam- velum. The marginal ciliaiy ring is difficult to
pling periods; range is followed by the mean distinguish from the locomotor ciliary wreath
and±standard deviation in parentheses. in T. tenniformis. The velum is a thick, well-
developed structure covering the bases ofthe
Results
cilia ofthe aboral ciliaiy complex and separat-
Morphology ing this complex from the adoral ciliar)' spiral
(Figs. 5, 6).
Trichodina temiifonnis Stein, 1979 The adhesive disc has a smooth pellicular
— surface beneath which the outline ofthe den-
Host. Co—ttushairdi (Pisces: Cottidae). ticles can be clearly seen. The disc is sur-
LOCALITY'. Provo River, Utah and Wasatch rounded peripherally by a 2-)U,m-wide border
counties, Utah. — membrane, which functions to seal the margin
Site of infestation.— Gill filaments. ofthe disc during adherence and contains fine
Light microscopy. Body 39-53 (44.2 ± vertical striae over its entire surface. These
4.0) dia (diameter). Adhesive disc 19-30 (26.3 striae on the internal surface of the border
± 2.8) dia, surrounded by a border membrane membrane are the radial pins that give the
2—3 (2.5 ± 0.4) wide, with fine transverse stri- membrane rigidity while retaining its ability
ae. Various-sized light forms present in center
of adhesive disc when silver-impregnated. to conform to the host's surface (Fig. 7).
Denticular ring 13.5-20 (17.2 ± 1.8) dia, con- The adoral ciliature forms a counterclock-
sisting of 20-26 (23.7 ± 1.3) denticles with wise spiral ofabout270°. The base ofeach cil-
6-10 (7.8 ± 0.8) radial pins per denticle. ium is inserted into a deep furrow and hidden
Denticle with conical central portions 0.7-1 from viewwhen SEM is u—sed (Fig. 8).
(0.99 ± 0.06) from which a thorn 2.5-4 (2.9 ± Deposition of slides. One slide (HWML
0.4) extends externally with broadly rounded 37721) of silver-impregnated specimens and
(HWML
lobes, tapered slightly to a blunt tip and blade another slide 37724) of iron-hema-
2-3 (2.3 ± 0.3) attached to central region, toxylin-stained specimens are deposited in
somewith rounded ends (Figs. 1, 2). the Harold W. Manter Laboratory, University
Macronucleus horseshoe-shaped 27-48 (39 ofNebraska State Museum. The senior author
± 5.7) dia and approximately 10 ^tm thick. has additional slides in hercollection.
260 Great Basin Natur.\list [Volume 55
Figs. 1-3. Light micrographs ot Triclioclhm tcuuijurmis: 1-2. Silver-impregnated specimens showingbody shape and
arrangement ofcomponents oftheadhesivedisc. BM, bordermembrane; D, denticle; RR radial pins. Bar = 10/im. 3.
Iron-hemato.xylin-stained specimen showing horseshoe-shaped macronucleus (MA); arrow points to the micronucleus
(MI). Bar = 10/Lim.
Apiosoma campanulatum Timofeev, 1962 ture ofthis species. Circular striations ofpelli-
—
cle conspicuous. Pellicle wrinkled into longi-
Host. Co—ttus bairdi (Pisces: Cottidae). tudinal furrows. Upper part ofbody bears the
Locality. Provo River, Utah and W^isatch
adoral zone, consisting ofatuft of1-2-^tm-long
counLStiiitgeesh,otfUmTii.ncfreosstcatoipoyn..——BGoidllyficlaammpenatnsu.late. cciolniat.raMcotsetd psepreicsitmoemnessvainedwecdonwtirtachteSdEpMerihsatvoe-
Macronucleus round or slightly conical. Size mial lips (Fig. 10). —
ofstainedspecimens 3L0-66.0 (47.8± 7.2) long Deposition of slides. A representative
by 25.0-45.0 (35.6 ± 4.2) wide. Macronucleus slide ofApiosoma campanulatum (silver stain)
lLO-20.0 (15.6 ± 2.4). Micronucleus not ob- is deposited in the Harold W. Manter Labora-
served (Fig. 9). — tory, University of Nebraska State Museum
Scanning electron microscopy. The (HWML 37722). The senior author has addi-
spindle-shaped body is the characteristic fea- tional slides inhercollection.
1995] Ciliated Protozoa in Mottled Sculpin 261
Figs. -J-7. Scaumug electron iiueiugraplis ofthe surface ofI temiiformis: 4. Aboral view ofentire specnnen ofI
tenuifonnis. B,bacteria. Bar = 10fxm.5. LateralviewofentirespecimenofI temiifonnis. ACS,adoralciliarvspiral; BS,
basalseptum; BCR,basilciliaiyring;GE,gillepithelium; LCW,locomotorciliaryweath;VEL,velum. Bar = 10fim. 6.
HighermagnificationofFigure4showingthe structureofaboral ciliaiycomplex.ACS, adoralciliaryspiral; BCR,basal
ciliaryring; K, kinetosomes; LCW, locomotorciliarywreath; MCR, marginalcilian,-ring. Bar = 1^tm. 7. Adhesive disc
ofT tenuifonnis. D,denticle;PRperipheralpins;RRradialpins.Bar= 1^tm.
262 Great Basin Natur.\list [Volume 55
Fig. 8. AdoralviewofI tenuifonnis showinghowtheadoralcihatiire (ACS) forms acounterclockwise spiral ofabout
270°. Bar = 10/xm. Fig. 9. Lightmicrograph ofApiosomacampanulatum. Noteconicle-shapedbody. MA, macronucle-
us. Bar = 1/xm. Fig. 10. Scanningelectron micrograph ofA. campanulatum attached to the gill epithelium (GE). Note
transversestriations ofpellicleandits longitudinal furrows (aiTOw). PL, peristomallip; PS, peristome. Bar = 5^im. Fig.
11. LightmicrographsshowingThchodina(T)andApiosomu(A)infestedgillepithelium. Bar = 20/xm.
Host-Symbiont Relationships lium, and particles dispersed in the water
Light microscopy.—Sections of mottled were on the surface ofT! tenuifonnis (Fig. 13).
No ultrastructural damagewas observedfor
sculpin gills had no apparent pathological
A. campanulatum. Presence of this ciliate in-
damage. The conical body ofsomeA. campan-
ulatum appeared to be attached to host gill wflaicstssnoomesercihoaunsgdeaimnagneumtboetrheofhomsittoceclhl.onTdhreirae,
surfaces by the scopula, while others were
with cristae showingmajorchanges (Fig. 14).
freely distributed over the epithelial surflice.
Most T. tenuifonnis glide over the surface; Ecological Aspects ofInfestation
only afewciliates adhere to the host epithelial
In the Provo Rivernearthe Provoresidential
cells (Fig. 11). —
Transmission electron microscopy. area, T. tenuifonnis reached the highest infes-
Sections of the interface between the host tation rate in April and May. Itwas uncommon
epithelial cell and T. tenuifonnis were pre- during summer and autumn and appeared to
pared. No permanent or temporary structure be absent in the winter With the increase of
could be detected between the adhesive disc, water temperature in spring, ciliates reinfest-
adoral zone of cilia, and gill epithelial cells ed the fish. Apiosoma campanulatum at this
(Fig. 12). However, injury to the epithelium site maintained an average of35% infestation
due to 7^ tenuifonnis can be detected by the rate (no. ofinfested fish vs. no. oftotal exam-
number of mitochondria, which decrease and ined fish) forall seasons.
disappear in the immediate host cell. Host In the upper Provo Riverthe tendencyofin-
necrotic tissue, mucous layers from gill epithe- festation ofT. tenuifonnis corresponded closely
1995] Ciliated Protozoa in Mottled Sculpin 263
Y 4^
^m
••••
fife'
^^.
^^
Figs. 12-13. Transmissionelectron micrographs ofgill epithelium infestedby T. teniiiformis. 12. Hostnecrotic tissue
(arrows) sloughs offforparasite's food. ACC, aboral cilian' complex. Bar - 1/j,m. 13. Interfacebetween T. teniiiformis
andmucouslayer(ML)ofepithelialcells. Notedamagetomitochondria(M).C,cilia.Bar — 1fim.
264 Great Basin Naturalist [Volume 55
14
y^^;:^^!^^
Nu
\-
''«^' i^y/fon^t. \
=^!-,
r
Fig. 14.TransmissionelectronmicrographofgillepitheliuminfestedhyA. campanulatum.A. campanulatwn(A)caus-
es numberofmitochondria(M) to decrease and cristae to disappear. M, mitonchondria; Nu, nucleus ofepithelial cell.
Bar= Ifim.
to that ofthe lower area. The highest infesta- represents a new host record for Trichodina
tion rate occurred in May and then decreased tenuiformis andApiosoma campanulatum.
until the next spring. Percentage offish infest-
Comparative Morphology
ed by T. tenuiformis in the lower river area
was 20.5% vs. 12.5% in the upper Provo River. At the LM level comparison ofthe adhesive
Similar to that ofthe lower river, A. campanu- disc ofr tenuiformis with that ofother species
latum at the upper site had an average of37% oiTrichodina reveals afew similarities. Tricho-
infestation in all four seasons. In general,Apio- dina reticulata Hirschmann and Partsch, 1955
soma did not showmeasurable fluctuations with described from Carassius auratus has denti-
seasons. cles similar to T tenuiformis (Bauer 1984). The
adhesive disc ofthe former has a central light
Discussion zone separated into reticulated structures. But
T. reticulata differs in having larger overall
Taxonomy and host-symbiont studies of dimensions (average adhesive disc diameter is
Trichodina and Apiosoma infesting fishes in 60/xmvs. 25^tm forourmaterial). T tenuiformis
the United States have received surprisingly has a close affinity to T elegans described by
little attention considering the frequency with Stein (1979) from fish in Russia. The latter is
which these organisms have been associated characterizedby an unbroken light zone in the
with fish diseases (Khan et al. 1974, Cone and adhesive disc. Our specimens have various-
Odense 1987, Khan 1991). Wellborn (1967) sized light forms in the center ofthe adhesive
described 13 species of Trichodina in south- disc. To a lesser extent T tenuiformis is similar
eastern United States, but few reports have to T puijtoraci Lom, 1962 and T domerguei
been published for this ciliate west of the Dogel, 1940; however, denticle shape and
Mississippi River(Hechmann etal. 1987). Little structure of the adhesive disc clearly distin-
information is available on Apiosoma studies guish T tenuiformis from these species.
in this country, which is not the case in the Surface features of the adhesive disc and
former Soviet Union (Bauer 1984). Cottushairdi arrangement ofthe aboral ciliaiy complex ofT
1995] Ciliated Protozoa in Mottled Sculpin 265
tenuifonnis seen by SEM were generally simi- Host-Symbiont Relationships
lar to those described for T! truttae, an ecto- Trichodina tenuifonnis is an ectocommensal
parasite on pacific salmon {Oncorliyncluis spp.)
with a tendency to be parasitic in mottled
and steelhead trout {OncoHiynclius inykiss;
sculpin. There were no visible pathological
Arthur and Margolis 1984), and T. labrisomi, symptoms with light microscopy; however,
an ectoparasite on hairy blenny {Labrisomas
electron microscopy disclosed changes in the
nuchipinnis; Rand 1993). However, in T. tenui- organelles ofhost epithelial cells infested by T.
fonnis, aboral cilia length is generally shorter tenuifonnis. Mitochondria decreased in num-
than in those previously described. Further-
ber and disappeared, which might indicate
more, comparison of the aboral ciliature of T. respiratory blockage due to lack of oxygen.
tenuifonnis with these species of Trichodina
showed some differences in the extent of This change in mitochondria was obsei-ved in
Trichophrya infesting other fish (Heckmann
development of the anterior and basal septa,
in velum structure, and in the degree of evi- and Carroll 1985). Necrotic host epithelial tis-
dence ofthe marginal ciliary ring. The anteri- sue sloughs off following organelle loss, sup-
plying sustenance forthe parasite.
or septum is relatively large and the basal one No serious damage to mottled sculpin could
itshesmbaalslalinsTe.pttnvimtitaies,pwrhoemrienaesnti.n TT.hteenvueilfounmniiss bseugogbesseti-evdedthfaortAt.hicsamspiamnpulleateucmt.ocLoommme(n19s7a3l)
well developed in both T. labrisomi and T.
relationship could change to parasitism in case
tenuifonnis, but T. tenuifonnis lacks anyprotu-
berances (Rand 1993). Similar to T. labrisomi, omfuchehalveyssipnvraosnioonusn,ceadltthhoaunghintthriischtoednidniednsc.y is
the marginal ciliary ring of T. tenuijormis is
poorly developed and cannot be distinguished Ecological Aspects ofInfestation
from the locomotor ciliaiy ring, whereas in T.
This study shows that the infestation ofTri-
truttae the marginal ciliary ring is well devel-
chodina has both seasonal and regional fluctu-
oped (Arthur and Margolis 1984). Rand (1993)
ations. The higher infestation rate on fish came
has suggested these marginal ciliature are sen-
from the Provo residential area during the
sory structures associated with feeding and
spring sampling period. Heavy impact from
odT.reintetenintcualitefisoonna.nnidUsnthlhaieskepneoTl!lpilecalubllriacirusloramiridgpaeosnrdeosnT.btethrteuwttoeareean,l tthhieslioncfaelsthautimoan.nApfotpeurlastuimomnerm,aythceonnturmibbuetre toof
surface, which might be a species-specific T. tenuiformis gradually reduces with the
decrease of water temperature and reaches
characteristic for these two species respective-
ly (Arthurand Margolis 1984, Rand 1993). the highest number the following spring. This
Over 50 species ofApiosoma have been maybe related to the ciliate life cycle.
recorded from fishes, the majority of which Unlike T. tenuifonnis in this study, A. cam-
have been describedby Russian authors (Bauer panulatum maintained a fairly constant infes-
1984). Although some are common fish para- tation on mottled sculpin from the two sites on
sites in some parts ofthe world, only one ref- the Provo Riverin all four seasons.
erence concerningApiosomapiscicola on Salve-
Acknowledgments
linusfontinalis was reported in North America
(Cone and Odense 1987). There is a paucity of The authors thank the Utah Fish and Game
data pertaining to Apiosoma over the last two
Department for their cooperation on this study.
decades, likely reflecting taxonomic difficul-
Dr. Dennis K. Shiozowa provided help with
ties due to variability in ciliaiy structure and fish collections. Technical assistance from Dr
lack ofstricthost-specificity.
Apiosoma conica has a body shape similar John Gardner and staffmembers ofthe Elec-
to A. campanulatum. But our specimens com- tron Optics Laboratoiy, BrighamYoung Univer-
pared more closely to the original description sity, was greatly appreciated.
ofA. campanulatum.
The species identifications were based on Literature Cited
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