Parasites of Slipper-cupped Oyster Crassostrea iredalei from Pulau Betong, West Coast of Penang, Malaysia
M. Jasim Uddin,
Aileen Tan Shau- Hwai
Until now, knowledge about bivalve mollusk diseases in tropical waters of Asia is scarce. A histopathologic survey was performed on the slipper-cupped oyster, Crassostrea iredalei from the Malaysian waters for the first time to investigate the presence of parasites and their effects on host tissues. C. iredalei samples were collected from Pulau Betong, west coast of Penang, Malaysia. A total of 60 oysters were examined from October 2010 to January 2011. After shell height measurement, the samples were dissected. A transverse section (3 mm) was cut through the anterior part of the soft body containing gills, mantle, digestive glands, gonads and foot tissue. The sections were processed by standard histological techniques, stained with hematoxylene and eosin and examined under microscope. A protozoan parasite, Nematopsis sp. was detected from the connective tissue of digestive gland tubules, gills, mantle and gonads. The prevalence of infection of Nemaptosis sp. were 15% each during October and November and 30% during January. The maximum number of hemocytes containing Nematopsis sp. per histological section was 5. The oocysts were oval in shape and maximum no. of oocysts per hemocyte was 7. No pathological threat was detected from the histopathological survey performed. This preliminary survey will be useful for planning a health monitoring programme for the natural and cultivated slipper-cupped oyster and other bivalves of commercial importance in Malaysian waters.
July 19, 2011; Accepted: October 19, 2011;
Published: December 24, 2011
Aquaculture production is increasing at a faster rate to meet up the demand
of the rapidly increasing population. Undoubtedly, global aquaculture is playing
a vital socio-economic role in relation to nutrition, employment opportunities
and income generations (Berthe, 2008). In 2007, shellfish
contributed over 26% of the global aquaculture production. Asia contributed
over 80% of the global mollusk aquaculture production during this time and bivalves
including oysters, clams, mussels, cockles and scallops accounted for 90% of
such production (FAO, 2009). Although Asia is the main
contributor in mollusk aquaculture production in the world, the study of mollusk
diseases still remained a neglected field in most of the tropical parts of Asia.
There are many examples of bivalve disease outbreaks associated with mass mortalities
in many parts of Asia. Therefore, it is high time to focus on this field for
the protection of the increasingly active mollusk aquaculture sector in this
Aquaculture production in Malaysia has increased by 450% in the past 16 years
from 1993 to 2009 (DOF, 2011). Due to gradual decrease
in production from natural stocks over time, DOF introduced oyster culture in
Malaysia to fulfill the demand. Diseases and parasites affecting bivalves in
Malaysian waters have not been systematically investigated so far. However,
a number of parasites have been reported causing diseases in farmed and natural
bivalve stocks from neighboring countries (Tuntiwaranuruk
et al., 2008; Taveekijakarn et al., 2008;
Erazo-Pagador, 2010). Therefore, it is necessary to
investigate the presence of parasites of farmed Crassostrea iredalei
in Malaysian waters from their economic and epidemiological point of view. From
the above context, present survey was conducted to investigate the occurrence
of parasites in farmed slipper-cupped oyster, Crassostrea iredalei from
Pulau Betong, west coast of Penang, Malaysia using histopathology.
MATERIALS AND METHODS
Sampling efforts: Crassostrea iredalei samples were collected from Pulau Betong, west coast of Penang, Malaysia (5°31 N 100°18 E) (Fig. 1). Sampling was drawn from an oyster farm in October 2010, November 2010 and in early January 2011. Twenty C. iredalei were processed monthly for the current histopathology survey. Upon arrival, the shell height of each oyster was measured using a slide calipers and the soft body was exposed using an oyster opening knife. For histological preparations, a transverse section (3 mm) was cut through the anterior part of the soft body in such that it contained gills, mantle, digestive glands, gonad and foot tissue.
Histology: The transverse section of the oyster was fixed in Davidsons solution with proper labeling. The tissue was subsequently dehydrated and embedded in paraffin. The paraffin blocks were sectioned at 6 μm thickness and stained with Harriss haematoxylene and eosin Y. The histological preparations were examined under a light microscope to investigate the presence of parasites in different tissues. Parasite prevalence (i.e., number of animals infected divided by the number of animals examined multiplied by 100) and distribution in each host were evaluated for each month.
Statistical analysis: All the data were analyzed statistically using SPSS statistical package version 11.0 to calculate the mean and standard deviation. In all cases, differences were considered significant when p<0.05.
|| Sampling site at Pulau Betong, Penang, Malaysia (5°31
N 100°18' E)
Morphological measurements: Shell height was the only morphological measurement taken in the present investigation. Table 1 shows the mean shell height (±SD) of C. iredalei examined during the 3 sampling dates; October 2010, November 2010 and January 2011. The highest mean shell height was measured in October (105.49±10.57 mm) and the lowest mean value was noted in November (86.36±7.84 mm). The shell height ranged from 92.0-127.8 mm in October 2010, 70.8-103.5 mm in November 2010 and 73.7-110.8 mm in January 2011.
Gross examination: Gross examination of C. iredalei revealed the presence of barnacles, oyster spat and crab in the shells. Figure 2 shows the presence of a crab in the shell of the oyster. The screening of C. iredalei by gross observation did not reveal any evidence of disease or parasite in the samples.
Histopathology: This is the first report on histopathological survey
on oysters from Malaysian waters. Histological examination confirmed the presence
of protozoan parasite, Nemaptosis sp. with a very light intensity of
infection. The maximum number of hemocytes containing Nemaptosis per histological
section was 5. The prevalence of infection of Nemaptosis sp. oocysts
parasitized C. iredalei varied among sampling dates. The prevalences of
infection of Nematopsis sp. were 15% each in October and November 2010
while the values were 30% in January 2011 (Table 1). The parasite
was detected in connective tissue of digestive gland tubules, gills, mantle
and gonads (Fig. 3). Figure 3a displays
the occurrence of Nematopsis oocysts in the connective tissue of the
digestive gland tubules. No clear host tissue damage was detected due to the
presence of the parasites. No pathological damage was also evident due to the
presence of Nematopsis sp. sporocysts in the gill tissue of C.
iredalei investigated in the current study (Fig. 3b).
||Shell height (±SD) (range), prevalence (%) and site
of infection of Nematopsis sp. in Crassostrea iredalei collected
from Pulau Betong, west coast of Penang, Malaysia during October, November
2010 and January 2011
|| Presence of crab inside the shells reported during gross
||Oocysts of Nematopsis sp. in different tissues of Crassostrea
iredalei collected from Pulau Betong, west coast of Penang, Malaysia.
a. infection in connective tissue of digestive gland tubules; b. in gills;
c. infection in mantle tissue associated with hemocytic responses; and d.
in connective tissues of male gonad
Only some hemocytic responses were associated with Nematopsis sp. infection
in the mantle tissue (Fig. 3c). No other pathological threat
was detected from the current histopathology survey. In Fig. 3d,
the presence of parasites did not show any anomaly to the connective tissue
of the male gonads and no clear host response was also observed due to Nematopsis
The oocysts of this Gregarines protozoan appeared to be oval shape. Each phagocyte
contained variable number of oocysts with 3-4 being the most frequent condition.
The length of the phagocyte was within the range of 30 μm to 60 μm.
The maximum number of Nemaptosis oocyts per phagocyte was 7 (Fig.
3b). Each oocyst contained a single uninucleate sporozoite. The mean oocyst
length was 15.04 μm and the width was 12.22 μm.
Research concerning diseases of oyster in Malaysia is scarce. A number of parasites
have been reported causing diseases in farmed and natural bivalves from the
neighboring countries. In Thailand, Nemaptosis sp., Maeteilia sp.,
Perkinsus sp. and trematodes were detected in cultivated and locally
harvested bivalves from the Gulf of Thailand (Tuntiwaranuruk
et al., 2008; Taveekijakarn et al., 2008).
Nemaptosis sp., Tylocephalum sp., digenetic trematodes and
ciliates were reported from slipper-cupped oyster, Crassostrea iredalei
from the coast of the Philippines (Erazo-Pagador, 2010).
For the sustainable oyster farming in Malaysian waters, it is timely to monitor
the health status of farmed bivalves on regular basis.
This is the first report on the histological screening of farmed slipper-cupped
oyster, Crassostrea iredalei from Malaysian waters for the presence of
parasites. Histopathology confirmed the presence of intrahemolytic oocysts of
gregarines of the genus Nematopsis in the farmed stocks of C. iredalei
along the coast of Pulau Betong, west coast of Penang, Malaysia. The prevalence
and intensity of infection were low without any evidence of pathological damage
to the host. The Apicomplexan protozoan parasite, Nematopsis spp. has
been reported to infect several bivalve locations in tropical to temperate waters.
The genus Nematopsis was reported to infect severa species from many
l bivalve species from the upper Gulf of Thailand (Tuntiwaranuruk
et al., 2008). Nematopsis sp. was the most prevalent parasite
of farmed slipper-cupped oyster, Crassostrea iredalei from Philippines (Erazo-Pagador,
2010). Such infection was also reported to occur along the American and
European waters (Bower et al., 1994; Carballal
et al., 2001; Boehs et al., 2010).
Present findings are the confirmation of its distribution in Malaysian waters.
Nemaptosis sp. was the only parasite found in this research. The prevalence
of infection was very light and did not cause any pathological damage to the
farmed oyster, C. iredalei except some hemocytic responses. Gregarine
of Nemaptosis sp. use marine bivalves as their intermediate host and
completing their life cycle within marine arthropods gut. In tropical
waters, the prevalence of Nematopsis spp. infection reported were 65
to 71% in Crassostrea iredalei from the Philippines (Erazo-Pagador,
2010), 92% in A. arcuatala, 59% in A. granosa, 60% in Perna viridis and
70% in Paphia undulata from the Gulf of Thailand (Tuntiwaranuruk
et al., 2004) and 73% in Mytella guyanensis from the coast of South
Bahia, Brazil (Boehs et al., 2010). The prevalence
of Nematopsis sp. infection obtained in the present study is relatively lower
than the above reports which might be due to different geographic positions
and host species.
Variable number of oocysts of Nemaptosis sp. is usually present per
infected host cell (phagocyte). In the present study, we observed 1-7 oocysts
per Nematopsis sp. infected hemocytes with 3-4 at most cases. Carballal
et al. (2001) reported maximum four oocysts per phagocyte and 1-2
oocysts per infected phagocyte was the most common feature in Cerastoderma edule.
The number of Nematopsis oocysts per phagocyte in Mytella guyanensis
varied from 1 to 20, with 1-3 being the most frequent condition; whereas, only
one oocyst per Nematopsis sp. infected phagocyte was seen in Anomalocardia
brasiliana (Boehs et al., 2010). The shape and
size of the oocysts measured in the present study is comparable with the findings
reported by Tuntiwaranuruk et al. (2008), Boehs
et al. (2010) and Erazo-Pagador (2010).
In the current study, we observed light hemocytic infiltration in the mantle
tissue of some infected animals without any clear host tissue damage. Gregarines
of the genus Nematopsis could cause focal hemocytic infiltration at most,
without obvious pathogenic effects (Bower et al.,
1994) which are in consistent with our findings. In contrast, Carballal
et al. (2001) and Tuntiwaranuruk et al.
(2004) reported decreased filtering efficiency and food intake of the infected
animal due to the presence of large number of Nematopsis sp. containing
phagocytes in the gill lumen which resulted in the obstruction of the water-flow
between the inhalant and exhalent currents. Gills and mantle tissues were reported
as the most frequent locations for Nematopsis spp. (Carballal
et al., 2001; Sabry et al., 2007),
as we observed in the current investigation. Heavy infection of Nematopsis
sp. at gills and mantle of young bivalves may weaken the juveniles; however,
such infection was found harmless in adults (Sprague and
Orr, 1955; Sprague, 1970). The prevalence of Nematopsis
sp. infection often correlates with the salinity and temperature of the
seawater. In the present study, Nematopsis sp. exhibited lower prevalence
of infection during October and November 2010 than that during January 2011.
In west coast of Peninsular Malaysia, the northeast monsoon associated with
heavy rainfall commence in October-November. The observed least prevalence of
infection during October and November 2010 might be associated with the drop
of water salinity due to heavy rainfall during this time. This finding is in
consistent with Jimenez et al. (2002) who reported
higher intensity of Nematopsis sp. infection in Litopenaeus vannamei
during dry season and minor incidence during the rainy season. Sabry
et al. (2007) also reported slightly lower prevalence of Nemaptosis
sp. in mangrove oyster Crassostrea rhizophorae during rainy season
when both temperature and salinity decreased in that area. In agreement with
the above reports, the relatively higher prevalence of infection during January
might be associated with the little rainfall yielding higher salinity level
in the sea water in which Nematopsis can thrive well. However, an annual survey
is necessary to confirm such relationship in Malaysian waters.
A histopathological survey was conducted on farmed slipper-cupped oyster, Crassostrea iredalei from Pulau Betong, west coast of Penang, Malaysia. Only protozoan parasite, Nematopsis sp. was detected from C. iredalei during the study period having light prevalence and infection intensity. No threat was detected to C. iredalei from the histopathological survey performed. A yearly monitoring is necessary to obtain the seasonality of parasitic infection in farmed C. iredalei.
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