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Parasites of the grouper fish Epinephelus
coioides (Serranidae) as potential
environmental indicators in Indonesian
coastal ecosystems
S. Kleinertz
1,2
* and H.W. Palm
2,3
1
Institute of Parasitology, Justus Liebig University Giessen, D-35392
Giessen, Germany:
2
Aquaculture and Sea-Ranching, Faculty of
Agricultural and Environmental Sciences, University of Rostock,
Justus-von-Liebig-Weg 6, D-18059 Rostock, Germany:
3
Faculty of
Veterinary Medicine, UDAYANA University, Denpasar, Bali, Indonesia
(Received 11 February 2013; Accepted 3 August 2013)
Abstract
A total of 195 Epinephelus coioides (Hamilton, 1822) were studied for fish
parasites from Javanese (Segara Anakan lagoon) and Balinese waters. Up to 25
different parasite species belonging to the following taxa: one Ciliata, one
Microsporea, five Digenea, one Monogenea, four Cestoda, four Nematoda, one
Acanthocephala, one Hirudinea and seven Crustacea were identified with four
new host and locality records. The dominant parasites included the monogenean
Pseudorhabdosynochus lantauensis (53.3–97.1%), the nematode Spirophilometra
endangae (23.3–42.9%), the digenean Didymodiclinus sp. (2.9–40.0%), the
nematodes Philometra sp. (22.6–34.3%) and Raphidascaris sp. (2.9–28.6%), and
the isopod Alcirona sp. (6.7–31.4%). Regional differences for E. coioides were
found in terms of endoparasite diversity, total diversity according to Shannon–
Wiener, Simpson index and Evenness. A comparison with published data from
Sumatera revealed highest endoparasite diversity (Shannon–Wiener: 1.86/1.67–
2.04) and lowest ectoparasite/endoparasite ratio (0.73/0.57–0.88) off the
Balinese coast, followed by Lampung Bay, Sumatera (1.84; 0.67), off the coast
of Segara Anakan lagoon (1.71; 0.71), and in the lagoon (0.30/0.19–0.66;
0.85/0.67–1.00). The presented data demonstrate the natural range of these
parameters and parasite prevalences according to habitat and region, allowing
adjustment of the scale that has been used in the visual integration of the parasite
parameters into a star graph. The parasite fauna of E. coioides in Segara Anakan
lagoon ‘improved’ from 2004 until 2008/09, possibly related to earlier oil spill
events in 2002 and 2004. The use of grouper fish parasites as an early warning
system for environmental change in Indonesian coastal ecosystems is discussed.
Introduction
Coastal marine ecosystems experience a variety of
environmental stressors, such as anthropogenic induced
pollution, environmental degradation and change
(Cooper et al., 2009; Dsikowitzky et al., 2011). Heavy
exploitation of the coastal resources leads to overfished
fish stocks, altered population sizes and species compo-
sition, as well as changed habitats. By 2025, 2.75 billion
people worldwide are expected to live close to the coast
(Palm et al., 2011), increasing the urgent need for a better
*Fax: þ 49 (0)641 99 38469
E-mail: sonja.kleinertz@vetmed.uni-giessen.de
Journal of Helminthology, page 1 of 14 doi:10.1017/S0022149X1300062X
q Cambridge University Press 2013
and sustainable use of the coastal resources. This requires
increased understanding and the development of new
methodologies to assess and visualize regional environ-
mental conditions and change.
It has been difficult to demonstrate the environmental
status of any coastal marine habitat, due to the complexity
and natural variability of such systems. According
to Kurtz et al. (2001), monitoring systems are required,
as it is impossible to measure and interpret all the
various influencing factors within an ecosystem. So far,
many environmental assessment studies have focused
on descriptive methodologies with no clear purpose
and using uncorrelated methods (Downs et al., 2005),
without further potential for practical applications.
As summarized by Palm & Ru
¨
ckert (2009) and reviewed
by Palm (2011), the status of a marine environment and
environmental change can either be studied directly, by
using, for example, water quality parameters such as
phosphate, nitrate and dissolved organic carbon (DOC),
or indirectly by using bioindicators. Such indicator
organisms react sensitively to specific environmental
conditions. Their occurrence or abundance can be used to
describe the current status of the environment, and even
environmental change.
Because of the direct linkage and dependence of para-
sites with multiple-host life cycles to the surrounding
animal communities (Hechinger et al., 2007), these
organisms have been considered as sensitive bioindica-
tors for aquatic ecosystem health (Overstreet, 1997;
Dzikowski et al., 2003). Fish parasites have been used as
biological and environmental indicators (for a review see
Palm, 2011), especially for environmental change and
pollution (Diamant et al., 1999; Dzikowski et al., 2003; Palm
&Ru
¨
ckert, 2009) or environmental stress (Landsberg et al.,
1998). Sures (2001, 2003) used acanthocephalan parasites
to detect heavy metal pollution, because acanthocepha-
lans accumulate 1000 times higher amounts of heavy
metals in contrast to their host tissues. Sasal et al. (2007)
utilised fish parasites to detect anthropogenic influences
(urban and industrial pollution) in coral reef lagoons, and
Lafferty et al. (2008b) suggested that they are a convenient
method to assess spatial variation of their final host
distribution. Heteroxenous fish parasites (multiple hosts)
with complex life cycles can be used to indicate food-web
relationships in unaffected marine habitats (e.g. Palm,
1999; Klimpel et al., 2006; Lafferty et al., 2008a). While the
occurrence of endoparasites often decreases in polluted
waters (Nematoda: Kiceniuk & Khan, 1983), ectoparasitic
parasites such as monogeneans can increase (Monogenea:
Khan & Kiceniuk, 1988; Trichodina: Khan, 1990; Palm &
Dobberstein, 1999; Ogut & Palm, 2005). Ectocommensals
with direct life cycles, such as trichodinid ciliates,
Operculi
Gills
Surface
Caligus cf. epinepheli (Cr), a
Pennelidae gen. et sp. indet. I, II* (Cr), 1
Pseudorhabdosynochus lantauensis (M), a, 1
Gnathiidae gen. et sp. indet.* (Cr), 1 (praniza
Hatschekia sp. (Cr), a
Sagum epinepheli (Cr), a
Trichodina sp. I*(P), a
Zeylanicobdella arugamensis (H), a
Microsporea gen. et sp. indet. (MI), sp
Allopodocotyle epinepheli (D), a
Prosorhynchus luzonicus*(D), a
Scolex pleuronectis* (Ce), 1
Callitetrarhynchus gracilis*(Ce), 1 (pl)
Southwellina hispida*(A), 1 (cystacanth)
Prosorhynchus sp. I, II*(D), a,1
Bothriocephalus sp.*(Ce), a
Raphidascaris sp. *(N), 1 (L3)
Camallanus carangis (N), a
Cainocraedium epinepheli *(D), a
Parotobothrium balli (Ce), 1(pl)
Spirophilometra endangae (N), 1
P: Protozoa
MI: Microsporea
D: Digenea
M: Monogenea
Ce: Cestoda
N: Nematoda
A: Acanthocephala
H: Hirudinea
Cr: Crustacea
* in different organs
a: adult
1: larval
pl: plerocercoid
sp: spore
Didymodiclinus sp. (D), a
Philometra sp. *(N), a
Alcirona sp. *(Cr), a, 1
Ectoparasites
Endoparasites
Intestine
Pyloric caeca
Stomach
Fins
Operculi
Mouth/Nostrils
Fig. 1. The occurrence of ectoparasites and endoparasites from the grouper fish Epinephelus coioides from Indonesian waters during
2007–2009.
2 S. Kleinertz and H.W. Palm
favour polluted waters and can indicate high bacterial
load (Palm & Dobberstein, 1999; Ogut & Palm, 2005),
in contrast to many endoparasites with complex life cycles
that favour stable and non-polluted waters, where the
full range of their required hosts is present (Lafferty
et al., 2008b).
The Indonesian coastal marine habitat has one of the
highest aquatic biodiversities on Earth (Yuniar et al., 2007;
Palm, 2011). This includes fish species as well as their
parasite fauna, though not more than about 4% of the
estimated fish parasite fauna in Indonesia has been
explored (Jakob & Palm, 2006). Palm & Ru
¨
ckert (2009)
applied a method to visualize environmental differences
by using fish parasites. They used the star-graph method
according to Bell & Morse (2003). The authors also
sampled Epinephelus coioides, from the wild and coastal
mariculture in Lampung Bay, Sumatera, and from inside
the anthropogenic influenced Segara Anakan lagoon
in Central Java. As exemplified by Palm et al. (2011) from
a mariculture facility in the Thousand Islands, six
different parasite metrics from Epinephelus fuscoguttatus
demonstrated a significant change in parasite compo-
sition and abundance over six consecutive years. The
authors suggested that groupers might also be useful
biomarkers to monitor environmental change in the wild.
Kleinertz et al. (2012) have shown regional differences
in the parasite composition of free-living Epinephelus
areolatus from Indonesian waters using the same
methodology.
Fish parasites of groupers (e.g. Cromileptes altivelis,
E. areolatus, E. fuscoguttatus) from tropical marine waters
have been of special interest in recent years. The groupers
are of high commercial value and, consequently, of
importance for fisheries as well as finfish mariculture
(Rimmer et al., 2004). This steadily growing business
is also playing a significant role in the Indonesian
economy, ensuring food availability and improving
the living standards of the coastal communities (Ru
¨
ckert
et al., 2010). Grouper (Epinephelus spp.) mariculture
production in Indonesia has increased 340% from 2004
to 2009 (DJPB, 2009).
Taxonomical and ecological studies on fish parasites
from Indonesia have been intensified in recent years (e.g.
Palm et al., 2007, 2008, 2011; Yuniar et al., 2007; Palm, 2008,
2011; Bray & Palm, 2009; Kuchta et al., 2009; Ru
¨
ckert et al.,
2009a, b, 2010; Kleinertz, 2010, Kleinertz et al., 2012, Dewi
& Palm, 2013; Kuhn et al., 2013), taking into account the
high parasite biodiversity at this tropical location.
The purpose of the present study is an assessment of
the fish parasite fauna of E. coioides, a widely distributed
and rapidly developing mariculture species in Indonesia,
from additional sampling sites. We have correlated the
observed parasite fauna with regional differences in
the sampled regions. Being aware that limited sample
replications of theoretically ‘impacted’ versus ‘healthy’
environments can be tested in Indonesia, we herewith
apply ecological and parasitological parameters that were
used to monitor regional differences and environmental
change by Palm & Ru
¨
ckert (2009) and Palm et al. (2011).
The use of grouper fish parasites as an early warning
system for environmental change in Indonesian coastal
ecosystems is discussed.
Materials and methods
Collection and examination of fish
Samples were taken within the framework of the SPICE
project (Science for the Protection of Indonesian Coastal
Marine Ecosystems) during the rainy season 2007/08 and
2008/09, and dry seasons 2008 and 2009. A total of 195
E. coioides (Hamilton, 1822) were studied from Javanese
(Segara Anakan lagoon) (108846
0
–109803
0
E; 08835
0
–
08848
0
S) and Balinese waters (114825
0
53
00
–115842
0
400
0
E;
8830
0
40
00
–08850
0
48
00
S) in Indonesia (fig. 1, table 1). Addi-
tional data were calculated based on Yuniar (2005; dry
season 2004) and Ru
¨
ckert (2006; dry season 2003) and
Ru
¨
ckert et al. (2009a) for comparison (also see table 1).
Table 1. The mean body length (cm) and body weight (g) of wild Epinephelus coioides sampled from
Indonesian waters in the rainy and dry seasons from 2007 to 2009 for comparison with *Yuniar (2005) and
**Ru
¨
ckert (2006); measurements of body length and weight shown in brackets.
Locality Year No. of fish Body length Body weight
Rainy season
Segara Anakan 2007/08 35 29 345.4
(25.1–38.8) (242.0–753.0)
Segara Anakan 2008/09 35 29.3 374.3
(25.1–36.4) (284.0–715.0)
Off the coast of Segara Anakan 2008/09 30 27.7 344.1
(20.3–40.3) (182.0–902.0)
Dry season
Off the coast of Segara Anakan 2008 30 27.3 294.2
(21.5–34.2) (140.0–579.0)
Bali 2008 35 29.7 355.6
(23.3–40.2) (201.0–746.0)
Bali 2009 30 33.1 529
(27.3–46.4) (340.0–1400.0)
Segara Anakan* 2004 21 17.3 83.2
(10.0–28.0) (13.0–250.0)
Ringgung** 2003 35 23.9 185.1
(19.5–34.5) (100.9–495.0)
Parasites of grouper fish as environmental indicators 3
Live fish were obtained from local fishermen using fish
traps in Segara Anakan lagoon and from Balinese waters.
Groupers from the coastal zone off Segara Anakan were
bought at the fish market and were separated into plastic
bags directly after the catch. Fish were transported
immediately to the laboratory, or kept on ice and then
frozen (, 2 208C) until subsequently dissected at the
Faculty of Biology, Jenderal Soedirman University,
Purwokerto (UNSOED) and the Faculty of Veterinary
Medicine, Udayana University, Jimbaran, Bali. Total fish
length (L
T
), weight (W
T
) and liver weight (W
L
) were
measured to the nearest 1.0 cm and 1.0 g (table 1) prior to
the parasitological examination (see Ru
¨
ckert et al., 2009a).
Smears were taken from the gills, surface and the inner
opercula of the living fish. The skin, fins, eyes, gills,
nostrils, mouth- and gill-cavity were examined for
ectoparasites. Inner organs such as the digestive tract,
liver, gall bladder, spleen, kidneys, gonads, heart and
swim bladder were separated and transferred into saline
solution for microscopical examination under the stereo-
microscope (Zeiss Stemi DV4; Carl Zeiss, Oberkochen,
Germany) in order to allow a quantitative parasitological
examination of each organ; belly flaps and musculature
(fillets) were examined on a candling table. Isolated
parasites were fixed in 4% borax-buffered formalin and
preserved in 70% ethanol. Smears from the gills, surface
and opercula were stained using silver nitrate (AgNO
3
)
impregnation, after Klein (1926, 1958): slides were rinsed
and covered with 5% silver nitrate solution and
impregnated for 30 min in the dark; the AgNO
3
was
removed and the slides were covered with distilled water
and exposed to ultraviolet light for 40–50 min. Smears
were dried after exposure. Finally, the musculature was
sliced into 0.5- to 1-cm-thick fillets and pressed between
two Petri dishes to identify and isolate parasites from
the musculature. Nematoda were dehydrated in a gra-
duated ethanol series and transferred to 100% glycerine
(Riemann, 1988). Digeneans, monogeneans and cestodes
were stained with acetic carmine, dehydrated, cleared
with eugenol and mounted in Canada balsam, whereas
crustaceans were dehydrated and transferred directly
into balsam. The identification of parasites was based
on original descriptions given in Palm et al. (2011).
Parasitological parameters
A variety of ecological parameters were evaluated
to indicate regional differences, such as the different
diversity indices (Shannon–Wiener, Evenness and
Simpson index), fish ecological indices (such as the
hepatosomatic index) and parasitological parameters
(such as ectoparasite/endoparasite ratio and different
prevalences of infection of metazoan parasites) (see Palm
&Ru
¨
ckert, 2009; Palm, 2011; Palm et al., 2011).
Parasitological calculations were made according to
Bush et al. (1997). The present study applies the method
by Palm & Ru
¨
ckert (2009) and Palm et al. (2011) to monitor
the parasite community of groupers from Indonesia.
This is based on the assumption that certain parasite
prevalence data and parameters are characteristic for
undisturbed environmental conditions with scenarios
of high parasite diversity. The Berger–Parker index
characterizes the dominance of a respective parasite
Table 2. The prevalence (%), intensity (I), mean intensity (MI) and mean abundance (MA) of ectoparasites from Epinephelus coioides in Javanese (in and off the coast of the Segara
Anakan) and Balinese waters.
Locality Segara Anakan Off the coast of Segara Anakan Bali
Year of sampling 2007/08 2008/09 2008 2008/09 2008 2009
Parasite species/-taxa (%) MI (I) MA (%) MI (I) MA (%) MI (I) MA (%) MI (I) MA (%) MI (I) MA (%) MI (I) MA
Trichodina sp. I 51.4 3.1 (1 –9) 1.57 40.0 2.6 (1 –11) 1.03 nc nc 17.1 11.5 (2 –32) 2.00 nc
Pseudorhabdosynochus
lantauensis
91.4 79.5 (3–433) 72.73 97.1 37.4 (1 –150) 36.31 86.7 39.2 (1 –259) 34.00 80.0 33.3 (1–225) 26.57 80.0 43.1 (1–246) 34.5 53.3 8.4 (1 –35) 4.50
Zeylanicobdella arugamensis 8.6 2.0 (2) 0.17 – – 16.7 1.2 (1–2) 0.20 40.0 2.3 (1–11) 0.93 – – – –
Alcirona sp. 31.4 19.6 (1–194) 6.17 – – 23.3 5.7 (1–33) 1.33 13.3 1.3 (1 –2) 0.17 8.6 1.0 (1) 0.09 6.7 2.5 (1 –4) 0.17
Gnathiidae gen.
et sp. indet.
8.6 1.0 (1) 0.08 40.0 1.7 (1–4) 0.69 6.7 11.5 (1–22) 0.77 13.3 1.8 (1 –4) 0.23 5.7 1.5 (1–3) 0.09 10.0 2.7 (1–6) 0.27
Caligus cf. epinepheli ––––––––5.71.0(1)0.60––
Caligidaegen.etsp.indet.––––––––60.025.9(2–89)15.66.71.0(1)0.07
Hatschekia sp.* ––––––––14.38.0(1–16)1.14––
Sagum epinepheli ––––––––2.91.0(1)0.03––
Pennelidae gen.
et sp. indet. I
28.6 5.2 (1–28) 1.49 – – – – – – – – – –
Pennelidae gen.
et sp. indet. II
20.0 4.3 (1 –11) 0.86 2.9 1.0 (1) 0.03 56.7 2.5 (1 –8) 1.43 23.3 94.6 (1 –636) 22.10 – – 33.3 7.0 (1 –21) 2.33
Ectoparasite species 7 4 5 5 7 4
* New host record. nc, not calculated.
4 S. Kleinertz and H.W. Palm
Table 3. The prevalence (%), intensity (I), mean intensity (MI) and mean abundance (MA) of endoparasites from Epinephelus coioides in Javanese (in and off the coast of the Segara
Anakan) and Balinese waters.
Locality Segara Anakan lagoon Off the coast of Segara Anakan lagoon Bali
Year of sampling 2007/08 2008/09 2008 2008/09 2008 2009
Parasite species/-taxa (%) MI (I) MA (%) MI (I) MA (%) MI (I) MA (%) MI (I) MA (%) MI (I) MA (%) MI (I) MA
Microsporea gen. et sp. indet. – – 17.1 5.3 (1–15) 0.91 6.7 4.5 (1–8) 0.60 – – – – – –
Didymodiclinus sp. 17.1 4.5 (2 – 9) 0.77 31.4 4.5 (1–32) 1.43 – – 40.0 2.1 (1–7) 0.83 2.9 5.0 (5) 0.14 13.3 2.8 (2–4) 0.37
Cainocraedium epinepheli* – – – – – – – – 11.4 4.5 (2–8) 0.51 6.7 2.0 (1 – 3) 0.13
Prosorhynchus luzonicus – – – – 6.7 31.0 (1–61) 0.60 – – – – – –
Prosorhynchus sp. I – – – – – – 16.7 8.8 (1–29) 1.47 11.4 7.3 (1 – 23) 0.83 26.7 2.6 (1–7) 0.70
Prosorhynchus sp. II 5.7 6.0 (1 –11) 0.34 – – – – – – – – – –
Bucephalidae gen. et sp. indet. – – – – – – – – – – 3.3 2.0 (2) 0.07
Digenea gen. et sp. indet. – – – – – – – – – – 3.3 1.0 (1) 0.03
Bothriocephalus sp. 8.6 2.0 (1–3) 0.17 – – 20.0 1.3 (1 –2) 0.27 26.7 2.5 (1–6) 0.67 – – – –
Scolex pleuronectis 2.9 2.0 (2) 0.06 – – – – – – 42.9 4.9 (1 – 32) 2.10 – –
Callitetrarhynchus gracilis* – – – – – – – – – – 3.3 1.0 (1) 0.03
Parotobothrium balli – – – – – – – – 8.6 1.7 (1–3) 0.14 – –
Raphidascaris sp. 2.9 1.0 (1) 0.03 8.6 2.3 (2–3) 0.20 – – 3.3 1.0 (1) 0.03 28.6 2.9 (1–14) 0.83 20.0 1.3 (1–2) 0.27
Camallanus carangis – – – – – – – – 2.9 1.0 (1) 0.03 – –
Philometra sp. 22.6 1.8 (1–3) 0.40 34.3 2.4 (1–7) 0.83 – – 23.3 1.0 (1) 0.23 34.3 2.8 (1–10) 0.94 33.3 1.3 (1–2) 0.43
Spirophilometra endangae 42.9 3.5 (1 – 13) 1.51 31.4 3.9 (1–17) 1.23 36.7 2.3 (1–7) 0.83 26.7 3.5 (1–10) 0.93 – – 23.3 2.9 (1–7) 0.67
Nematoda gen. et sp. indet. I 22.9 2.6 (1–8) 0.60 – – 3.3 1.0 (1) 0.03 – – 2.9 2.0 (2) 0.06 3.3 1.0 (1) 0.03
Nematoda gen. et sp. indet. II 2.9 9.0 (9) 0.26 – – – – – – – – – –
Southwellina hispida* 8.6 1.0 (1) 0.08 2.9 1.0 (1) 0.03 16.7 3.6 (1–7) 0.60 20.0 3.5 (1–13) 0.70 – – – –
Endoparasite species 8 6 5 7 8 7
Ecto-/endoparasite ratio 0.88 0.67 1.00 0.71 0.88 0.57
* New host record.
Parasites of grouper fish as environmental indicators 5
species within the sample BP ¼ N
max
/N, with N
max
being
the number of specimens of the most dominant species in
relation to the total number of parasites within the sample
(N) (Munkittrik et al., 1994). The diversity of the collected
metazoan endoparasite fauna of each fish species was
determined by using the Shannon–Wiener diversity
index (H
0
) and, according to Kleinertz et al. (2012), the
Evenness index (E) of Pielou (Magurran, 1988) and other
parameters were tested (see below). Microsporean
parasites were not considered because it was not possible
to calculate their intensity. In the case of trichodinid
ciliates, the calculations given in table 2 refer to the
density, based on counts from slides with mucous smears
obtained from about 1 cm
2
of gill surface area. The ratio
of ecto- to endoparasites was calculated (Ec/En ratio
(R) ¼ number of ectoparasite species/number of endo-
parasite species), with trichodinid ciliates treated as
present or absent in this calculation. Species groups
(higher taxa such as Nematoda indet.) that could not be
further identified and might represent other recorded
taxa were omitted from the calculations (see Palm et al.,
2011). The hepatosomatic index was calculated as a
descriptor of a possible pollution impact to the fish host,
which may affect increasing liver weights (W
L
) in relation
to the total weight (W
T
) of the host (HSI ¼ W
L
/W
T
£ 100)
(Munkittrik et al., 1994). The Simpson diversity index was
also considered as a bioindicator D ¼ 1=
P
s
i¼1
ðn
i
=NÞ
2
,
excluding the data for the trichodinids (see the
explanation above, only density was recorded), with
s ¼ the total number of parasite species collected within
the sample (ecto- and endoparasites included), N ¼ the
total number of parasite individuals collected within the
sample, n
i
¼ number of specimens of a single species i.
Visual integration
The visual integration of the calculated ecological
indicators follows Palm & Ru
¨
ckert (2009) for the
prevalence of trichodinids, ectoparasite/endoparasite
ratio and endoparasite diversity after Shannon–Wiener.
The Simpson diversity index, Evenness index and
hepatosomatic index were added according to Kleinertz
et al. (2012). In addition, the prevalences of five different
parasite species were used to distinguish among the
sampling sites: Scolex pleuronectis and Terranova sp.
(according to Lafferty et al., 2008b; Palm et al., 2011),
Raphidascaris sp. (Nematoda: Kiceniuk & Khan 1983; Palm
et al., 2011), Zeylanicobdella arugamensis (Grosser et al.,
2001) and Trichodina sp. (Khan, 1990; Palm & Dobberstein,
1999; Ogut & Palm, 2005). Values that indicate unnatural
environmental conditions are orientated towards the
centre of the star graph. Values representing natural and
unaffected environmental conditions are arranged
towards the frame of the star graph. Based on Palm &
Ru
¨
ckert (2009), Palm et al. (2011) and Kleinertz et al.
(2012), we adjusted the parameter ranges to values that
represent all available samples of E. coioides.
Data analysis
Univariate and multivariate statistical analyses were
conducted with the programs STATISTICA (release 6,
StatSoft Inc., Tulsa, Oklahoma, USA) and PRIMER
(release 6, Primer-E Ltd. 6.1.11, Ivybridge, Devon, UK),
respectively. Homogeneously distributed (Levene’s test)
and normally distributed data (Shapiro test) were tested
for significant differences with the t-test or with one- or
two-factorial analyses of variances (ANOVA), using
Tukey’s HSD test for post-hoc comparisons. The chi-
square test was used to compare each year and sampling
site with another for all parameters showing parasite
prevalences and ectoparasite/endoparasite ratios (see
Palm et al., 2011). All tests were considered statistically
significant at P , 0.05.
In order to compare the parasite communities, abun-
dance data were square-root transformed. A similarity
matrix was constructed using the Bray –Curtis similarity
measure. The relation between samples based on the
comparison of similarity matrices was displayed using
cluster analysis and multi-dimensional scaling (MDS)
with stress value estimation: , 0.05, excellent; , 0.2,
reliable; . 0.2, start of loss of accuracy. One-way analyses
of similarity were applied to identify the differences in
parasite species composition between the sampling sites
(routine ANOSIM, values close to 1 indicate high
differences and close to 0 indicate high similarity between
species compositions). Routine SIMPER analysis was
applied to test which parasite species contributed most to
the shown differences between the sampling sites
(Clarke & Warwick, 1994; see also Nordhaus et al., 2009).
SIMPER analysis was used to determine which
species was most responsible for the differences that
have been seen between sites with Bray–Curtis analysis
(according to Bell & Barnes, 2003; see also Kleinertz
et al., 2012).
Results
In both years, during rainy season 2007/08, 2008/09
and dry seasons 2008 and 2009, fish parasitological
studies on E. coioides in Segara Anakan lagoon, off the
coastal zone of Segara Anakan lagoon and Balinese
waters revealed 25 different parasite species, belonging
to the following taxa: one Ciliata, one Microsporea, five
Digenea, one Monogenea, four Cestoda, four Nematoda,
one Acanthocephala, one Hirudinea and seven Crustacea
(fig. 1, tables 2 and 3). Four new host and locality records
were established for E. coioides (tables 2 and 3) mainly in
Balinese waters. Information on prevalence, intensity,
mean intensity and mean abundance of the collected
parasite species is summarized in tables 2 and 3.
To analyse the parasite composition and ecological status
at the respective sampling sites, the ecological parameters
as suggested by Palm & Ru
¨
ckert (2009) and Palm et al.
(2011) were considered as given below (table 4). Regional
differences for E. coioides were found in terms of endo-
parasite diversity, total diversity (Shannon–Wiener),
Simpson index and Evenness between Bali and in the
Segara Anakan lagoon (table 4, figs 2, 3 and 4; for regional
comparison see fig. 5).
Parasite diversity and infection levels
The parasite species richness in Bali (up to 17 taxa,
calculated and pooled in the fish samples for both years)
6
S. Kleinertz and H.W. Palm
was higher than that in fish off the coast of the Segara
Anakan lagoon (14 taxa). For each single sample, the
highest species richness of 15 taxa was recorded from
both Bali in 2008 and Segara Anakan lagoon in 2007/08
during the rainy season. The lowest species richness
of 10 taxa was recorded in fish from both the Segara
Anakan lagoon 2008/09 during the rainy season and off
the coast of this lagoon in 2008 during the dry season
(tables 2 and 3).
The lowest ectoparasite richness (four taxa) was found
in the second year (2008/09) of samples from both Segara
Anakan lagoon and Bali, and highest (seven taxa) for
the same samples in the first year (2007/08) (table 2). The
endoparasite richness was highest (eight taxa) in fish
from Segara Anakan lagoon and Bali in the first year
(2007/08), and lowest (five taxa) in the first sample from
off the coast of Segara Anakan lagoon (table 3).
Ectoparasite/endoparasite ratios, calculated by using
the numbers of ectoparasite species vs. the numbers of
endoparasite species, ranged from 0.6 to 1.0 (table 3).
Regional differences of the ectoparasite/endoparasite
ratio were not significant.
The endoparasite diversity (Shannon–Wiener index) of
E. coioides of the present study ranged from 0.19 in Segara
Anakan lagoon to 2.04 in Bali (table 4). The Simpson
diversity index for the whole parasite community was
lower for grouper parasites in Segara Anakan lagoon
(1.39) compared with Bali (3.73) (table 4). The highest
Evenness value (1.00) for endoparasites was recorded for
Bali, compared with the lowest value (0.09) in Segara
Anakan lagoon. The Berger–Parker index was lowest in
Balinese waters (0.45) and highest in Segara Anakan
lagoon (0.87) (table 4). The hepatosomatic index ranged
from 0.77 off the coast of Segara Anakan to 1.37 in Segara
Anakan lagoon (table 4), with a significant difference
(ANOVA: F ¼ 3.74, P , 0.001).
The most predominant parasites, occurring at all
sampling sites, were the monogenean Pseudorhabdosyno-
chus lantauensis 53.3 –97.1%, the nematode Spirophilometra
endangae 23.3–42.9%, the digenean Didymodiclinus sp.
2.9–40.0%, the nematodes Philometra sp. 22.6–34.3% and
Raphidascaris sp. 2.9–28.6%, and the isopod Alcirona sp.
6.7–31.4%. The prevalence of infection of the larval
tetraphyllidean cestode Scolex pleuronectis as well as the
larval nematode Raphidascaris sp. was different between
the different regions during the first year (2007/08). The
prevalence for both parasite taxa was significantly higher
in Balinese waters compared to Segara Anakan lagoon:
42.9 versus 2.9% and 28.6 versus 2.9%, P ¼ 0.000 and
0.003 (table 3). The larval nematode Terranova sp. could
only be isolated from E. coioides from Ringgung (Ru
¨
ckert,
2006; Palm & Ru
¨
ckert, 2009) at a prevalence of 14.3%,
resulting in significant regional differences between all
sampled groupers of the present study with n ¼ 30–35
fish per location and year (see table 1) in contrast to those
from Ringgung with n ¼ 35 (P ¼ 0.020–0.025). The
prevalence of infection of the leech Z. arugamensis was
significantly different during the second year of investi-
gation (2008/09) between Segara Anakan lagoon and off
the coast of Segara Anakan, with 0% versus 40.0% in
2008/09, P ¼ 0.000. The same trend was observed in the
first year (2007/08), with no significant difference (8.6%
versus 16.7%, P. 0.05) (table 2). The prevalence of
Table 4. Mean values (^ SD) of hepatosomatic index and condition factor for the free-living Epinephelus coioides together with parasite species diversity in Javanese (in and off the
coast of the Segara Anakan) and Balinese waters from 2007 to 2009 for comparison with modified data from *Yuniar (2005), **Ru
¨
ckert (2006) and Ru
¨
ckert et al. (2009a).
Locality Segara Anakan Off the coast of Segara Anakan Bali Ringgung
Year of sampling 2004* 2007/08 2008/09 2008 2008/09 2008 2009 2003**
Host/parasite parameters
Hepatosomatic index nc 1.03 (0.04) 1.37 (0.07) 0.77 (0.05) 0.96 (0.07) 1.11 (0.07) 1.02 (0.06) 1.58 (0.12)
Condition factor 1.34 (0.29) 1.38 (0.03) 1.47 (0.03) 1.38 (0.02) 1.57 (0.04) 1.34 (0.03) 1.41 (0.04) 1.29 (0.22)
Shannon–Wiener (endoparasites) 0.66 0.19 0.33 0.43 1.71 1.67 2.04 1.84
Shannon–Wiener (total) 0.92 0.65 0.51 0.81 1.19 0.79 1.67 1.14
Evenness (endoparasites) 0.41 0.09 0.21 0.27 0.88 0.80 1.00 0.71
Evenness (total) 0.42 0.25 0.25 0.37 0.48 0.30 0.70 0.38
Ec/En ratio 1.00 0.88 0.67 1.00 0.71 0.88 0.57 0.67
Simpson index 2.00 1.39 1.31 1.50 2.51 2.26 3.73 2.05
Berger–Parker index 0.67 0.85 0.87 0.81 0.48 0.60 0.45 0.69
Ec/En ratio, ectoparasite/endoparasite ratio; nc, not calculated.
Parasites of grouper fish as environmental indicators 7
infection with the ciliate Trichodina sp. was significantly
higher for fish in Segara Anakan lagoon compared to
Balinese coastal waters, with 51.4% and 40.0% versus
17.1%, P ¼ 0.003 for 2007/08 and P ¼ 0.034 for 2008/09
(table 2).
Regional parasite composition and visual integration
Significant regional differences in species composition
were not found between the sampled E. coioides from
Balinese and Javanese coastal waters in either year.
Highest differences regarding to ANOSIM analyses were
found between samples from Bali in 2008 and off the coast
of Segara Anakan in 2008, with ANOSIM: R ¼ 0.399,
P ¼ 0.01, and between samples from Bali in 2008 and
Segara Anakan lagoon in 2007/08, with ANOSIM:
R ¼ 0.342, P ¼ 0.01. There was a distinct separation
between the parasite composition of the sampled
E. coioides from Segara Anakan region (both sites)
compared to those from Balinese waters in the first
sample with ANOSIM: R ¼ 0.332, P ¼ 0.01 (fig. 6a). In the
following year, the parasite composition was different,
without an obvious regional separation, with ANOSIM:
R ¼ 0.217, P ¼ 0.01 (Fig. 6b). With regard to SIMPER
analysis, the parasite species contributing most to the
regional differences in Segara Anakan lagoon in 2007/08
were P. lantauensis, 76.80%; S. endangae, 7.71%; Alcirona
sp., 4.19%; and Pennelidae gen. et sp. indet. I, 3.26%,
being also present in the samples from the other sampling
sites. Off the coast of Segara Anakan lagoon in 2008, the
species contributing most were P. lantauensis, 67.68%;
Pennelidae gen. et sp. indet. II, 20.70%; and S. endangae,
5.55%. Those from Bali in 2008 were P. lantauensis, 53.36%;
Caligidae gen. et sp. indet., 22.49%; S. pleuronectis, 11.91%;
and Philometra sp., 6.59%.
Ten parasite bioindicators are visualized within a star
graph according to Bell & Morse (2003), Palm & Ru
¨
ckert
(2009) and Palm et al. (2011), to illustrate regional
differences between the sampling sites. The presented
data demonstrate the natural range of these parameters
Evenness (endoparasites)
Hepatosomatic index
Scolex pleuronectis
Terranova sp.
Raphidascaris sp.
Zeylanicobdella arugamensis
Trichodina sp.
Ecto-/ endoparasite ratio
Simpson index
0.2
0.4
0.6
0.8
1.0
2.0
3.0
4.0
5.0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
90
90
80
80
70
70
60
60
50
50
40
40
30
30
20
20
10
10
90
90
90
80
80
80
70
70
70
60
60
60
50
50
50
40
40
40
30
30
30
20
20
20
10
10
10
0.5
0.25
0.25
0
0.5
0.5
0.75
0.75
1
1
1.25
1.25
1.5
1.5
1.75
1.75
2
2
2.25
1
1.5
2
2.5
3
3.5
4
4.5
Evenness (endoparasites)
Hepatosomatic index
Endoparasite
biodiversity
Evenness (endoparasites)
Hepatosomatic index
Scolex pleuronectis
Terranova sp.
Raphidascaris sp.
Zeylanicobdella arugamensis
Trichodina sp.
Ecto-/ endoparasite ratio
Simpson index
0.4
0.5
0.7
0.6
0.8
0.9
1.0
2.0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
90
80
70
60
50
40
30
20
10
90
80
70
60
50
40
30
20
10
50
50
50
40
40
40
30
30
30
20
20
20
10
10
10
0.25
0.5
0.5
0.75
0.75
1
1
1.25
1.25
1.5
1.5
1.75
1.75
2
2.25
1
1.5
2
2.5
3
3.5
4
Endoparasite
biodiversity
Evenness (endoparasites)
Hepatosomatic index
Scolex pleuronectis
Terranova sp.
Raphidascaris sp.
Zeylanicobdella arugamensis
[trichodinids]
no data
Ecto-/ endoparasite ratio
Simpson index
0.4
0.5
0.7
0.6
0.8
0.9
1.0
2.0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
90
80
70
60
50
50
40
40
30
30
20
20
10
10
50
50
50
40
40
40
30
30
30
20
20
20
10
10
10
0.25
0.5
0.5
0.75
0.75
1
1
1.25
1.25
1.5
1.5
1.75
1.75
2
2.25
1
1.5
2
2.5
3
3.5
4
Endoparasite
biodiversity
Endoparasite
biodiversity
Scolex pleuronectis
Terranova sp.
Ra
p
hidascaris s
p
.
Zeylanicobdella arugamensis
[trichodinids]
no data
Ecto-/ endoparasite ratio
Simpson index
0.2
0.4
0.6
0.8
1.0
2.0
3.0
4.0
5.0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
90
90
80
80
70
70
60
60
50
50
40
40
30
30
20
20
10
10
90
90
90
80
80
80
70
70
70
60
60
60
50
50
50
40
40
40
30
30
30
20
20
20
10
10
10
0.5
0.25
0.25
0
0.5
0.5
0.75
0.75
1
1
1.25
1.25
1.5
1.5
1.75
1.75
2
2
2.25
1
1.5
2
2.5
3
3.5
4
4.5
a
1
b
1
a
2
b
2
Fig. 2. Visual integration of environmental indicators for free-living Epinephelus coioides from Javanese waters: (a
1
,a
2
) Segara Anakan
during the rainy season 2007/08 and (b
1
,b
2
) off the coast of Segara Anakan lagoon during the dry season 2008, with normal integration
(1)
and adjusted parameter range
(2)
. Host/parasite parameters are given in the upper half and prevalences (%) of parasite species in the
lower half of each star graph.
8 S. Kleinertz and H.W. Palm
and parasite prevalences according to habitat and region,
allowing an adjustment of the scale to be utilized in the
visual integration of the parasite parameters. According
to the newly collected and already published data of
E. coioides parasites from Indonesia, the hepatosomatic
index among the sampling sites ranged from 0.77 to 1.58,
the Evenness from 0.09 to 1.00, the ectoparasite/endo-
parasite ratio from 0.57 to 1.00, the Simpson index from
1.31 to 3.73 and the endoparasite diversity according to
Shannon–Wiener from 0.19 to 2.04. The prevalence of
infection for S. pleuronectis was 0–42.9%; for Terranova sp.,
0–14.3%, Raphidascaris sp., 0–31.4%; Z. arugamensis,
0–40.0% and for Trichodina sp. 14.3–52.4%. Figure 2a
1
,
b
1
and fig. 3a, b illustrate the parasite parameters by
utilizing a prevalence range from 0 to 100% and the range
for the ecological parasite parameters according to Palm
&Ru
¨
ckert (2009) and Palm et al. (2011), with most of the
indicators oriented towards the centre of the star graph
in Segara Anakan lagoon and towards the middle in
the sample off the coast of Segara Anakan lagoon.
According to the parasitological data of E. coioides from
Indonesia recorded here, the star graphs with adjusted
parameter range are given in fig. 2a
2
,b
2
. The regional
differences in the parasite infection of E. coioides between
inside Segara Anakan lagoon in 2004 (Yuniar, 2005;
Ru
¨
ckert et al., 2009a), Bali (present study) and Ringgung
2003 (Ru
¨
ckert, 2006) are given in fig. 4a
1
,a
2
,b
1
,b
2
and
fig. 5a
1
,a
2
with and without adjustment of the parameter
range, respectively.
Discussion
Grouper parasites
To our knowledge, up to 2009, 28 parasitological studies
had been recorded for E. coioides worldwide, revealing a
total of 57 different parasite species/taxa, belonging to the
Ciliata (4), Microsporidia (1), Myxozoa (1), Digenea (7),
Monogenea (13), Cestoda (6), Nematoda (13), Acanthoce-
phala (2), Hirudinea (1) and Crustacea (9) (Kleinertz,
2010). Of these records, 77% originate from Indonesian
waters; with the present study adding four new host
records (see tables 2 and 3). The 25 different parasite
species recorded here cover 57% of all previous records
from Indonesian waters and 44% of the worldwide
records for this host. Kuchta et al. (2009) stated that only
four bothriocephalideans have been reported so far from
Indonesia. Palm & Ru
¨
ckert (2009) added Botriocephalus
sp. from E. coioides from Segara Anakan lagoon; it was
also recorded within the present study but so far not
identified to the species level. This provides further
evidence for the high parasite biodiversity in Indonesian
waters (Palm et al., 1999; Palm, 2000; Carpenter & Springer,
2005; Yuniar et al., 2007), encouraging further parasito-
logical studies within the region. Most recently, Justine
et al. (2010) added one more parasite record, Argathona
rhinoceros, for E. coioides from New Caledonian waters.
Parasite infection according to region and year of sampling
As already stated by Williams et al. (1992) and Arthur
(1997), the parasite species composition of distinct
fish species reflects differences in food sources, feeding
preferences and habitats. Consequently, fish parasites
are useful for a range of different applications, such
as biological-, accumulation-, effect- and ecosystem-
indicators (Palm, 2011). According to the selected parasite
parameters, the infracommunity of E. coioides parasites in
Segara Anakan lagoon was significantly different from
the other regions studied so far in Indonesia. Segara
Anakan can be considered an extreme habitat, with a low
stability within the lagoon on the ecosystem and
biogeochemical level (Jennerjahn et al., 2009, see below).
In addition, it is an area with a high load of organic
contaminants (Dsikowitzky et al., 2011). High water mass
exchange rates between Segara Anakan lagoon and the
coastal region, regularly changing salinities depending on
seasons, and possibly natural migration of the sampled
Evenness (endoparasites)
Hepatosomatic index
Scolex pleuronectis
Terranova sp.
Raphidascaris sp.
Zeylanicobdella arugamensis
Trichodina sp.
Ecto-/ endoparasite ratio
Simpson index
0.2
0.4
0.6
0.8
1.0
2.0
3.0
4.0
5.0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
90
90
80
80
70
70
60
60
50
50
40
40
30
30
20
20
10
10
90
90
90
80
80
80
70
70
70
60
60
60
50
50
50
40
40
40
30
30
30
20
20
20
10
10
10
0.5
0.25
0.25
0
0.5
0.5
0.75
0.75
1
1
1.25
1.25
1.5
1.5
1.75
1.75
2
2
2.25
1
1.5
2
2.5
3
3.5
4
4.5
Endoparasite
biodiversity
Evenness (endoparasites)
Hepatosomatic index
Scolex pleuronectis
Terranova sp.
Raphidascaris sp.
Zeylanicobdella arugamensis
Ecto-/ endoparasite ratio
Simpson index
0.2
0.4
0.6
0.8
1.0
2.0
3.0
4.0
5.0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
90
90
80
80
70
70
60
60
50
50
40
40
30
30
20
20
10
10
90
90
90
80
80
80
70
70
70
60
60
60
50
50
50
40
40
40
30
30
30
20
20
20
10
10
10
0.5
0.25
0.25
0
0.5
0.5
0.75
0.75
1
1
1.25
1.25
1.5
1.5
1.75
1.75
2
2
2.25
1
1.5
2
2.5
3
3.5
4
4.5
Endoparasite
biodiversity
[trichodinids]
no data
(a) (b)
Fig. 3. Visual integration of environmental indicators for free-living Epinephelus coioides from Javanese waters: (a) Segara Anakan during
the rainy season 2008/09 and (b) off the coast of Segara Anakan during the rainy season 2008/09. Host/parasite parameters are given in
the upper half and prevalences (%) of parasite species in the lower half of each star graph.
Parasites of grouper fish as environmental indicators 9
fish result in a low parasite load, especially of the
endohelminths in E. coioides (see data for the first year,
2007/08). This is clearly visualized in the resulting star
graphs, with most parasite parameters oriented towards
the centre (figs 2a
1
,a
2
,b
1
,b
2
, 3a, 4a
1
,a
2
).
A comparison of parasite data revealed the highest
richness in 2007/08 with 15 species in the lagoon
compared to 12 species during rainy season 2008/09 off
the coast. Both values were higher compared to the data
from the dry season 2008 off the coast of Segara Anakan
and during the rainy season 2008/09 in the lagoon.
Groupers off the coast of Segara Anakan were bought on
the fish market as dead specimens, and it is possible that
the fish from the first sample might have originated as
living specimens from inside the lagoon. According to
ANOSIM, in the second year, the parasite fauna was
different from samples in the lagoon, representing the
situation of coastal fish in other regions. A comparison
with published data from Ringgung, Lampung Bay,
Sumatera in 2003 (Ru
¨
ckert, 2006; Palm & Ru
¨
ckert, 2009)
revealed a low endoparasite diversity according to
Shannon–Wiener: 0.30/0.19–0.66 and high ectoparasite/
endoparasite ratio, with 0.85/0.67–1.00 in the lagoon,
followed off the coast of Segara Anakan lagoon at
Teluk Bay with 1.71; 0.71 and Lampung Bay with 1.84;
0.67. Highest endoparasite diversity according to
Shannon–Wiener: 1.86/1.67–2.04 and a low ectoparasite/
endoparasite ratio with 0.73/0.57–0.88 were recorded
for E. coioides off the Balinese coast, a region that was
considered of high environmental quality by Kleinertz
et al. (2012). The cluster analyses and multi-dimensional
scaling plots likewise illustrated these differences;
however, they are far less sensitive than the applied star
graph method (compare fig. 6 with figs 2, 3, 4 and 5).
During the present study there was only small
variability, without any significance, between both years
in Segara Anakan lagoon (2007/08 versus 2008/09).
However, according to Khrycheva (2009), two fatal oil
Evenness (endoparasites)
Hepatosomatic index
Calculation not
possible
Scolex pleuronectis
Terranova sp.
Raphidascaris sp.
Zeylanicobdella
arugamensis
Trichodina sp.
Ecto-/ endoparasite ratio
Simpson index
0.2
0.4
0.6
0.8
1.0
2.0
3.0
4.0
5.0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
90
90
80
80
70
70
60
60
50
50
40
40
30
30
20
20
10
10
90
90
90
80
80
80
70
70
70
60
60
60
50
50
50
40
40
40
30
30
30
20
20
20
10
10
10
0.5
0.25
0.25
0
0.5
0.5
0.75
0.75
1
1
1.25
1.25
1.5
1.5
1.75
1.75
2
2
2.25
1
1.5
2
2.5
3
3.5
4
4.5
Endoparasite
biodiversity
a
1
Evenness (endoparasites)
Hepatosomatic index
Scolex pleuronectis
Terranova sp.
Raphidascaris sp.
Zeylanicobdella
arugamensis
Trichodina sp.
Ecto-/ endoparasite ratio
Simpson index
0.4
0.5
0.6
0.8
0.7
0.9
1.0
2.0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
90
90
80
80
70
70
60
60
50
50
40
40
30
30
20
20
10
10
50
50
50
40
40
40
30
30
30
20
20
20
10
10
10
0.25
0.5
0.5
0.75
0.75
1
1
1.25
1.25
1.5
1.5
1.75
1.75
2
2.25
1
1.5
2
2.5
3
3.5
4
Endoparasite
biodiversity
a
2
Evenness (endoparasites)
Hepatosomatic index
Calculation not
possible
Scolex pleuronectis
Terranova sp.
Raphidascaris sp.
[Zeylanicobdella
arugamensis]
no data
[Zeylanicobdella
arugamensis]
no data
Trichodina sp.
Ecto-/ endoparasite ratio
Simpson index
0.2
0.4
0.6
0.8
1.0
2.0
3.0
4.0
5.0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
90
90
80
80
70
70
60
60
50
50
40
40
30
30
20
20
10
10
90
90
90
80
80
80
70
70
70
60
60
60
50
50
50
40
40
40
30
30
30
20
20
20
10
10
10
0.5
0.25
0.25
0
0.5
0.5
0.75
0.75
1
1
1.25
1.25
1.5
1.5
1.75
1.75
2
2
2.25
1
1.5
2
2.5
3
3.5
4
4.5
Endoparasite
biodiversity
b
1
Evenness (endoparasites)
Hepatosomatic index
Scolex pleuronectis
Terranova sp.
Raphidascaris sp.
Trichodina sp.
Ecto-/ endoparasite ratio
Simpson index
0.4
0.5
0.6
0.8
0.7
0.9
1.0
2.0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
90
90
80
80
70
70
60
60
50
50
40
40
30
30
20
20
10
10
50
50
50
40
40
40
30
30
30
20
20
20
10
10
10
0.25
0.5
0.5
0.75
0.75
1
1
1.251.25
1.5
1.5
1.75
1.75
2
2.25
1
1.5
2
2.5
3
3.5
4
Endoparasite
biodiversity
b
2
Fig. 4. Visual integration of environmental indicators for free-living Epinephelus coioides from Javanese and Balinese waters: (a
1
,a
2
) Segara
Anakan during the dry season 2004 (data modified after Yuniar, 2005) and (b
1
,b
2
) off the coast of Bali during the dry season 2008, with
normal integration
(1)
and adjusted parameter range
(2)
. Host/parasite parameters are given in the upper half and prevalences (%) of
parasite species in the lower half of each star graph.
10 S. Kleinertz and H.W. Palm
tanker accidents happened in 2002 and 2004 within the
lagoon. Our first data of E. coioides from the lagoon
originated from the dry season 2004 (fig. 4a
1
,a
2
), the rainy
season 2004/05 and the dry season 2006 (Yuniar, 2005;
Palm & Ru
¨
ckert, 2009). The diversity based on the
Shannon–Wiener and Simpson indices, as well as the
Evenness, were higher during the dry season 2004
compared to 4 years later. However, the ectoparasite/
endoparasite ratio changed slightly throughout the
different samples, from 1.00 in the dry season 2004 and
rainy season 2004/05, to 0.80 in the dry season 2006, 0.88
in the rainy season 2007/08 and 0.67 in the rainy season
2008/09. Having similar parasite species throughout the
years might indicate a potential recovery towards the
natural parasite fauna of E. coioides in the lagoon after
both pollution events.
Visual integration
By using the star graph method to integrate different
fish parasitological parameters into the same figure,
Palm & Ru
¨
ckert (2009) and Kleinertz et al. (2012)
visualized regional differences within Indonesian waters,
and Palm et al. (2011) visualized annual changes. Ten
different parameters were chosen to describe the parasite
communities of E. coioides at the sampling sites. The
hepatosomatic index describes a possible pollution
impact to the fish host (Munkittrik et al., 1994). The
Evenness for endoparasites, Simpson index and endopar-
asite biodiversity according to Shannon–Wiener are used
in order to describe natural environmental conditions
(Palm & Ru
¨
ckert, 2009; Ru
¨
ckert et al., 2009a; Palm, 2011;
Palm et al., 2011; Kleinertz et al., 2012). The prevalence of
trichodinid ciliates describes bacteria-enriched waters
(Palm & Ru
¨
ckert, 2009). Different leeches have been used
as a kind of substandard sensitive marker for definite
chemical parameters (Grosser et al., 2001). The authors
noted a high sensitivity of these organisms, especially to
hypoxic water conditions, high phosphate, heavy metal
and organic pollutant concentrations. In the case of our
model, we utilized the leach Z. arugamensis, a regular
parasite in our samples. We are aware that leeches may
fall off the fish host during sampling and might not be
considered a good bioindicator in all cases. However,
Z. arugamensis is firmly attached to the groupers, needs to
be pulled off with the help of forceps and can be counted,
Evenness (endoparasites)
Hepatosomatic index
Scolex pleuronectis
Terranova sp.
Raphidascaris sp.
Trichodina sp.
Ecto-/ endoparasite ratio
Simpson index
0.2
0.4
0.6
0.8
1.0
2.0
3.0
4.0
5.0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
90
90
80
80
70
70
60
60
50
50
40
40
30
30
20
20
10
10
90
90
90
80
80
80
70
70
70
60
60
60
50
50
50
40
40
40
30
30
30
20
20
20
10
10
10
0.5
0.25
0.25
0
0.5
0.5
0.75
0.75
1
1
1.25
1.25
1.5
1.5
1.75
1.75
2
2
2.25
1
1.5
2
2.5
3
3.5
4
4.5
Endoparasite
biodiversity
a
1
Evenness (endoparasites)
Hepatosomatic index
Scolex pleuronectis
Terranova sp.
Raphidascaris sp.
Trichodina sp.
Ecto-/ endoparasites ratio
Simpson index
0.4
0.5
0.6
0.8
0.7
0.9
1.0
2.0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
90
90
80
80
70
70
60
60
50
50
40
40
30
30
20
20
10
10
50
50
50
40
40
40
30
30
30
20
20
20
10
10
10
0.25
0.5
0.5
0.75
0.75
1
1
1.25
1.25
1.5
1.5
1.75
1.75
2
2.25
1
1.5
2
2.5
3
3.5
4
Endoparasite
biodiversity
a
2
[Zeylanicobdella
arugamensis]
no data
[Zeylanicobdella
arugamensis]
no data
Fig. 5. Visual integration of environmental indicators for free-living Epinephelus coioides from Sumatera waters: (a
1
,a
2
) for cultured
Epinephelus coioides from Balai Budidaya Laut during the dry season 2003 (data modified after Ru
¨
ckert (2006) and Ru
¨
ckert et al. (2009a)),
with normal integration
(1)
and adjusted parameter range
(2)
. Host/parasite parameters are given in the upper half and prevalences (%) of
parasite species in the lower half of each star graph.
Fig. 6. A multi-dimensional scaling plot of the parasite
community of Epinephelus coioides from Javanese and Balinese
waters in (a) 2007/08 and (b) 2008/09.
, Segara Anakan; P,off
the coast of Segara Anakan;
, Bali.
Parasites of grouper fish as environmental indicators 11
especially after collecting the fish in separate plastic bags.
The cestode S. pleuronectis and the nematode Raphidascaris
sp. are also common in Indonesian waters, and have
been recorded for E. coioides in Segara Anakan lagoon
(Yuniar, 2005; Ru
¨
ckert et al., 2009b). Groupers represent
intermediate hosts in the life cycle of Raphidascaris
sp., becoming infected via abundant amphipods as
first intermediate hosts. Ru
¨
ckert (2006) concluded that
epinephelids can also be final hosts for these nematodes.
Terranova sp., with a possible zoogeographical restriction,
has to be considered for the Segara Anakan region (Palm
et al., 2011).
Segara Anakan lagoon can be considered an extreme
habitat for the fish as well as the parasite fauna. The
lagoon has high freshwater influx, mostly from Citanduy
River (Holtermann et al., 2009), is governed by tides
(Jennerjahn et al., 2009) and can be divided into two
major water bodies, mainly connected via a single water-
exchange channel. Each of the parts has a direct
connection to the ocean (Holtermann et al., 2009).
Jennerjahn et al. (2009) observed spatio-temporal vari-
ations in the distribution of dissolved nutrients in Segara
Anakan lagoon, probably the result of seasonally varying
interactions of natural (hydrology, geomorphology, soils,
vegetation) and anthropogenic (land use, urbanization)
factors. The lagoon has been facing a number of
environmental problems for decades, because of resource
exploitation (Jennerjahn et al., 2009) such as overfishing,
logging of mangrove wood, high sediment input by the
Citanduy River because of poor upland agricultural
practices, agricultural runoff, potential pesticide and oil
pollution (White et al., 1989; Jennerjahn et al., 2009;
Dsikowitzky et al., 2011). Due to all those facts, we can
expect that the high hydrological variability in Segara
Anakan lagoon has an important impact on the associated
biotics. Consequently, the observed parasite parameters
of E. coioides in the lagoon, with the characteristic shape of
the star graph (figs 2a
1
,a
2
,b
1
,b
2
, 3a), represent a heavily
disturbed ‘natural habitat’. This is in contrast to the
coastal zones of Bali, Lampung Bay and even off the coast
of Segara Anakan at Teluk Bay, with stable hydrological
conditions and less disturbed environments. Thus, our
samples represent the greatest possible range of the
respective parasite parameters under natural conditions
in Indonesia. This leads to the adjustment of the scales
that have been used to place the observed parasite
parameters into the star graph system for E. coioides (see
figs 2 and 3). One open question still remains, on how the
recorded parasite species react to defined polluted
conditions. This will allow final adjustment of the still
theoretical range of parameters that we have applied so
far for E. coioides parasites as environmental indicators
in Indonesian coastal ecosystems.
It can be concluded that the presented methodology to
visualize fish parasite parameters can distinguish
definitive environmental conditions in Indonesian waters
under high biodiversity scenarios. So far, regional
differences (Palm & Ru
¨
ckert, 2009, Kleinertz et al., 2012;
the present study) and long-term annual changes inside a
mariculture farm in the Thousand Islands (Palm et al.,
2011) and inside the heavily disturbed ‘natural habitat’ of
Segara Anakan have been found. According to these data,
free-living E. coioides had a high parasite load, similar
to those of E. fuscoguttatus (Ru
¨
ckert et al., 2010) and
E. areolatus (Kleinertz et al., 2012). Regular parasitological
monitoring of these commercially important fish species
will be able to detect environmental conditions and
change, possibly serving as an early warning system in
Indonesian coastal habitats. We are aware that it is
difficult to link directly all observed parasite parameters,
without replicates and experiments, to define environ-
mental or anthropogenic factors at all sampling sites and
times. However, the star graph system allows direct
statements to be made about otherwise highly complex
biological scenarios, supporting decision making on the
future use of the Indonesian coastal ecosystems.
Acknowledgements
We are thankful to the Indonesian State Ministry of
Research and Technology (RISTEK) for the support and
research permit (No. Surat Izin: 0037/FRP/SM/II/09).
We are thankful for institutional support to the Leibniz
Center for Tropical Marine Ecology, GmbH, Bremen,
Germany, and the Jenderal Soedirman University
(UNSOED), Purwokerto, Java (Professor Dr E. Yuwono).
Special thanks to Mr Andih Rinto Suncoko and Mr Edwin
Hermawaran from UNSOED for their personal initiative
and organizational support during fieldwork. This is
publication No. 4 under the Memorandum of Under-
standing between the Faculty of Veterinary Medicine,
UDAYANA University, Bali, and the Faculty of Agri-
cultural and Environmental Sciences, Aquaculture and
Sea-Ranching, University Rostock, Germany, in order
to promote fish parasite and biodiversity research in
Indonesia.
Financial support
Financial support was provided by the German Federal
Ministry for Education and Science (BMBF Grant
Nos. 03F0471 A and 03F0641D) (S.K., H.W.P.) within the
framework of the joint Indonesian–German research
programme SPICE II and III-MABICO (Science for the
Protection of Indonesian Coastal Marine Ecosystems).
Conflict of interest
None.
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