Ocean Sci. J. (2017) 52(1):7589 Available online at http://link.springer.

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http://dx.doi.org/10.1007/s12601-017-0010-y

Article

pISSN 1738-5261
eISSN 2005-7172

Reproductive Biology of Blood Cockle Anadara granosa (Bivalvia: Arcidae)

in the Northern Region of the Strait of Malacca
Munawar Khalil1*, Zulfigar Yasin2, and Tan Shau Hwai2
1
Department of Aquaculture, Universitas Malikussaleh, Aceh 24351, Indonesia
2
Marine Sciences Laboratory, School of Biological Science, Universiti Sains Malaysia, Gelugor 11800, Malaysia

Received 19 February 2016; Revised 4 August 2016; Accepted 31 October 2016

 KSO, KIOST and Springer 2017

Abstract  A study on the reproductive cycle of the blood cockle (Gosling 2003; Arapov et al. 2010). Their common name
Anadara granosa (Bivalvia: Arcidae) was conducted at three refers to the hemoglobin and hemocyanin pigments in their
different areas in the northern region of the Strait of Malacca. A blood and tissue cells, giving their blood a dark red color
total of 1,920 samples of adult A. granosa (38–71 mm length) were (Ruppert and Barnes 1994) which has allowed this species
collected from June 2009 until September 2010. Qualitative techniques to live in oxygen-critical habitats (Broom 1985; Terwilliger
(gonadal microscopic fresh smear test and histology analysis) as
and Terwilliger 1985; Cilenti et al. 2010). The species is
well as quantitative techniques (analysis of condition index and
gonadal index) were used to predict monthly gonadal development indigenous to the intertidal mudflats of many Southeast
stages of A. granosa. The gonadal index of A. granosa from Banda Asian countries, particularly Indonesia, Malaysia and Thailand.
Aceh (Indonesia) (r = 0.469, P > 0.05) and Pulau Pinang (Malaysia) Anadara granosa are mainly distributed in mangrove forests,
(r = 0.123, P > 0.05) did not show any correlation to their condition muddy vegetation and mixed areas. The intertidal species
index, whereas the gonadal index of A. granosa from Lhokseumawe A. granosa is known as a keystone species in mangrove
(Indonesia) (r = 0.609, P < 0.05) showed moderate positive correlation
habitats in several areas in the northern region of the Strait
to the condition index. During the 16 month sampling period, four
reproductive cycles were observed: each from three to six months. of Malacca. This species has also been one of the most
The process of releasing gametes is termed dribble spawning, and important fisheries commodities in Southeast Asia for many
is the same in all populations. The principle component analysis years (Borrero 1986; Broom 1985; Suwanjarat et al. 1990,
(PCA) indicated that A. granosa reproduction was affected by 2009).
interaction between internal physiological factors and indigenous The northern Strait of Malacca is an important nursery
environmental factors. In all sampling areas, phytoplankton density
area for many intertidal organisms and a feeding area for
played a key role in the reproductive cycle in A. granosa. Information on
the reproductive biology of this species is essential for species migrating species. Being the most important species in
management and to improve the sustainability practices of the terms of fisheries production, this cockle has become the
fisheries industry. These findings will provide basic information on target of an extensive culturing operation in West Malaysia
the biology of the blood cockle A. granosa for stock management (Broom 1983). At the same time, harvesting of wild stock of
in the region. cockles in Sumatra and Java (Indonesia) is at an all-time
Key words  blood cockle, reproductive cycle, gametogenesis, high to meet the demand for shellfish. In Malaysia, the
gonadal index, condition index annual production of blood cockles in 2009 exceeded metric
65,000 tonnes, which is valued at US $36.60 million (DOF
1. Introduction 2010). The main blood cockle production areas in Malaysia
are concentrated in Kedah (Merbok), Pulau Pinang (Juru),
Anadara granosa is one of 7500 bivalve species in the Perak (Kuala Gula, Kula Sangga-Matang, Kuala Trong,
family Arcidae, often called “blood arks” or “blood cockles” Sungai Jarum), Selangor (Kuala Selangor) and Johor (Muar).
*Corresponding author. E-mail: [email protected] In Indonesia, this species can be found in abundance on the
76 Khalil, M. et al.

coast of West Sumatra, Central and South Java, East and This data is lacking for the blood cockle Anadara granosa
West Kalimantan and other muddy bottoms in Sulawesi, but is essential to optimize aquaculture of this species. This
Maluku and Papua (Khalil et al. 2009). The most recent data bivalve species can be managed more effectively after evaluating
available on annual cockle production in Indonesia is from the regeneration capabilities of natural stocks and interpreting
2009 when it reached 47,437 metric tonnes, or equal to US$ growth patterns. Detailed and comprehensive information
23.72 million (DKP 2010). on gonadal development is also important for economic
The Northern Straits of Malacca is an important area for management of this species (Gribben et al. 2004; Peharda et
the harvesting and culture of the blood cockle A. granosa al. 2006). This study aimed to investigate the seasonal gonadal
due to the suitability of the habitats for spawning and growth cycle of the cockle A. granosa by using quantitative techniques
(Mirzaei and Hwai 2016). However, annual production (gonadal index and condition index) through gonadal fresh
statistics indicate a decrease in stocks in the last decade. This smear test and gonad histology (a qualitative technique)
situation may be due to inadequate management of wild from specimens collected from the northern region of the
cockle populations. Fisheries management is needed to Strait of Malacca.
improve policies for the sustainability of the fisheries industry. In
depth information on reproductive cycles is necessary for 2. Materials and Methods
predicting annual recruitment, as well as interpreting growth,
mortality, and survival data in the marine culture of species Collecting of samples
(Shaw 1965; Manzi et al. 1985; Sbrenna and Campioni 1994). A total of 120 samples of adult A. granosa were collected

Fig. 1. Sampling location

 Reproductive Biology of Anadara granosa 77

monthly from June 2009 till September 2010 from the natural microtome was used to cut 5–7 µm thick tissue sections
habitat in Banda Aceh (532'34.67"N–9517'2.54"E), Lhokseumawe which were mounted on a glass microscope slide. The light
(0509'35.3"N–09708'29.4"E) in Aceh, Indonesia and Pulau compound microscope was used to analyze the gonad structure
Pinang (516'9.66"N–10023'27.37"E) in Malaysia (Fig. 1). to recognize the sex and gametogenesis stages (divided into:
The total number of specimens sampled was 1,920 and the (+1) indeterminate, (+2) developing, (+3) developed and
adult cockle sizes ranged from 38–71 mm in length. The (+4) spawned).
sampling area was characterized by muddy substrate which
was surrounded by mangroves, no wave action and high Quantitative method
salinity. The specimens were collected at a depth of 5–30 cm Analysis of condition index (CI)
and salinity ranged from 10–33 ppt. Sampling activity in the The water displacement method was used to determine
field was conducted once a month over the specified time the condition index. A total of 30 specimens (size range: 38–
frame during low tides. The live specimens were collected 71 mm in length) from each sampling station were examined
manually with the aid of a harrow, which was run through from June 2009 to September 2010. Each specimen was
the muddy area to pull bivalves to the surface. After collecting, measured for the following: dry flesh weight, wet weight of
the specimens were stored in isotherm containers and shell in grams (g) and internal cavity volume (ml). Fresh cockle
immediately transported to the laboratory. The samples were tissue including its shell was weighed using digital balances.
cleaned to fully remove all fouling organisms and other The flesh was dried at 105°C for 72 hours to a constant
adherences. weight. The volume of the shell internal cavity was calculated by
means of subtracting the volume of the shell (ml) from the
Qualitative technique total wet volume (ml). These data were used to calculate the
Gonadal microscopic fresh smear test condition index using the formula described by Lawrence
A total of 40 specimens per sampling site were randomly and Scott (1982):
allocated for the gonadal microscopic fresh smear test each
Condition index = dry flesh weight × 100 / shell internal
month. All the specimens were dissected with a dissecting
cavity volume (cm3)
needle and pipette. The fresh smear procedure was adopted
to observe the gonad content under a compound light Analysis of gonadal index (GI)
microscope (magnification = 100 x) to analyze the stages of Gonadal index was calculated based on the formula proposed
the gonadal development. The sex and gametogenesis stages by Gosling (2003) and Kim and Lee (2008):
were identified using image analysis, which included 4 stages:
(+1) indeterminate, (+2) developing, (+3) developed and (+4) Gonadal index =  n individual from each stage level
spawned (Rajagopal et al. 2006). gonad stage / n total specimen for each sampling batch
The gonadal index (GI) was calculated for each sampling
Histology analysis month through the gonadal microscopic fresh smear test and
A total of ten gonad specimens from each of the three histological analysis to estimate the proportion of the gonadal
sampling sites were allocated for histological analysis each stages (indeterminate, developing, developed and spawned).
month. Slides were prepared through the process of embedding The GI value was ranked to: 1 (all individuals’ gonads in
paraffin wax into the tissue. Haematoxyline and Eosin the samples were in spawned stage), 2 (all individuals’
coloration were used for tissue coloring (Howard et al. 2004). gonads in the samples were in indeterminate stage), 3 (all
The initial process requires dehydration of the specimen individuals’ gonads in the samples were in developing
tissue. Dehydration was done through a series of steps of stage) and 4 (all individuals’ gonads in the samples were in
immersing the sample into varying concentrations of alcohol. developed stage).
The sample was embedded into a mold of wax next and kept
in a refrigerator overnight before preparing it for HE coloration. Statistical analysis
The solutions used for histology included bouins, alcohol Raw data was compiled and entered into Microsoft Office
(50%, 70%, 80%, 90%, 95% and absolute alcohol), xylene, Excel 2011 (Macintosh version) for processing and analyzing
liquid wax, histosolve, HE solution and 1.5% ammonia. A of minimum and maximum values, averages, and the standard
78 Khalil, M. et al.

deviation as well as to generate graphs. One-Way ANOVA oocytes. Previtellogenic oocytes can still be found in small
statistical analysis and post hoc test were used to determine amounts.
significance level (P < 0.05 and P < 0.01) in the values of Stage 4 (spawned).
each data cluster. Pearson correlation test was also utilized to Male: Gonadal compound reduced drastically. Spermatozoa
determine and understand the relationship between differing have diminished. Unused residual spermatozoa can be found
variables (CI and GI). The principle component analysis (PCA) inside the lumen.
was used to analyze the correlation between parameters which Female: Gonadal compound turned bright orange due to
were affected by reproductive activities in each sampling the lowest concentration of oocytes. Mature oocytes were
area. These statistical analyses were conducted using SPSS found in small amounts, but these are expected to be
(Statistical Package for Social Science) release 20.0 for residue or absorbed as phagocytes. Most of the oocytes had
Macintosh. no shape and the nuclei appeared to have shrunk and
disappeared.
3. Results
Gonadal histology analysis
Gonadal structure of Anadara granosa Stage 1 (indeterminate).
Gonadal microscopic fresh smear analysis Male and female: The stage is also called the dormant stage;
The description of gonad structure of A. granosa based on the sexes cannot be distinguished. Undeveloped gonads’
microscopic fresh smear analysis was categorized as shown content during this stage only consisted of connecting tissues
below: and a handful of residual gametes leftover from the previous
Stage 1 (indeterminate). spawned stage (stage 4) (Fig. 2).
Male and female: Determination of sex cannot possibly be Stage 2 (developing).
determined. Gonadal compound appeared to be empty and Male: Gonad was gradually filled up with spermatogonia,
filled up only by network of connecting tissues. Unused residual spermatocyte, and a small quantity of spermatozoa. The
of gametes can be found. average diameter of the follicles at this stage was 117.77 ±
Stage 2 (developing). 19.58 μm in size (Fig. 3a).
Male: The gonadal compound turned cream in color. Gametes Female: Oocytes occur in a range of sizes and were
have been very active and the testis was filled with spermatogonia generally not the same shape (irregular). Gonad was gradually
and spermatid. Spermatozoa were also found in limited numbers filled up with oogonia as well as vitellogonia oocyte and
and sometimes found in tailed form and actively swim. vitellogenic oocytes, the nuclei have uneven shapes. The
Female: The gonadal compound turned orange in color. average diameter of the follicles at this stage was 136.21 ±
Gametes in ovary have begun to appear, which are previtellogenic 22.12 μm, whereas the average diameter of oocytes was
oogonia, oocytes and a limited number of oocytes vitellogenic. 24.81 ± 6.19 μm in size (Fig. 4a).
Oocytes were scattered and filled inside the follicle. Nuclei
in oocytes vitellogenic have started forming and are clearly
visible. Oocytes have uneven sizes.
Stage 3 (developed).
Male: The gonadal compound turned a more concentrated
cream color as a result of highly condensed developed
spermatozoa. The spermatozoa have already developed their
tail and are swimming actively. Sometimes, spermatids can
still be found in small numbers.
Female: Gonadal compound turned an intense, concentrated
orange due to formation of highly condensed oocytes. Gametes
were generally mature oocytes. Oocytes are in polyhedral
Fig. 2. Gonadal structure of Anadara granosa based on histology
form. The nuclei within the oocytes have matured and grown analysis at indeterminate stage. FW: follicle wall; Lu:
larger in size. The yolks were found in most of the mature Lumen; EL: empty lumen; Ct: connective tissue
 Reproductive Biology of Anadara granosa 79

Fig. 3. Gonadal structure of male Anadara granosa based on histology Fig. 4. Gonadal structure of female Anadara granosa based on
analysis: (a) Stage 2 (Developing); (b) Stage 3 (Developed); histology analysis: (a) Stage 2 (Developing); (b) Stage 3
(c) Stage 4 (Spawned). FW: follicle wall; Lu: lumen; Spz: (Developed); (c) Stage 4 (Spawned). FW: follicle wall;
spermatozoa; MS: mature spermatozoa; SD: sperm ductus; Lu: Lumen; EVO: early stage of vitellogenic oocyte; LVO:
St: spermatid; DS: degenerative space; DSz: degenerative late stage of vitellogenic oocyte; MO: mature oocyte Nl:
spermatozoa; FF: follicle fragment; EF: empty follicle; nucleus; FF: follicle fragment; EF: empty follicle; RO:
Ct: connective tissue residual oocyte; IS: interfolicular space; YG: yolk granule

Stage 3 (developed). Stage 4 (spawned).

Male: Gonad was mainly dominated by spermatozoa content. Male: Spermatozoa seemed to be reduced, as the follicle
Interfollicular space at this stage was seen to be experiencing appeared almost empty. Spermatogonia were not found (Fig. 3c).
constriction due to the growing follicle size. Spematogonia Female: Residual oocytes were present. The follicles’ wall
were still found in limited number and typically found on the seemed to be damaged and unfilled. Phagocytes were found
side wall of the follicle. The average diameter of the follicles around the residue oocytes (Fig. 4c).
was 186.16 ± 14.47 μm in size (Fig. 3b).
Female: Gonad was characterized by the dominance of Gonadal development cycle
vitellogenic oocytes with a visibly large nucleus. Lumen The gonad percentage (for each stage) was compared between
space was dominated by the polyhedral oocyte vitellogenic the three sampling locations: Banda Aceh (Indonesia),
shape which was untouched or free from the follicle wall. Lhokseumawe (Indonesia) and Pulau Pinang (Malaysia).
The cytoplasm of mature oocytes had been filled by a Figures 5a, 6a, 7a, as well as 5b, 6b and 7b depict the computation
number of yolk granules. The average diameter of the of gonad percentages per month for all the 4 phases discussed
follicles was 215.13 ± 38.40 μm and oocytes were 30.01 covering a span of 16 months, from June 2009 until September
6.80 μm in size (Fig. 4b). 2010, through gonadal microscopic fresh smear analysis
80 Khalil, M. et al.

Fig. 5. Anadara granosa gonadal development pattern from Banda Aceh, Indonesia (June 2009–September 2010)

and gonadal histology analysis, respectively. Figures 5c, 6c index (CI), and monthly gonadal index (GI), respectively,
and 7c, as well as 5d, 6d and 7d depict the monthly condition covering the same 16 months.
 Reproductive Biology of Anadara granosa 81

Fig. 6. Anadara granosa gonadal development pattern from Lhokseumawe, Indonesia (June 2009–September 2010)

Environmental variable other environmental parameters. The principle component

Monthly seasonal variations of environmental parameters analysis (PCA) was conducted to evaluate the comprehensive
in the three different sampling areas are reported in Table 1. relationship between environmental factors and reproduction
During the study period, water temperature, salinity and variables in the A. granosa populations in Banda Aceh,
phytoplankton density fluctuated significantly compared to Lhokseumawe, and Pulau Pinang (Fig. 8).
82 Khalil, M. et al.

Fig. 7. Anadara granosa gonadal development pattern from Pulau Pinang, Indonesia (June 2009–September 2010)

4. Discussion year. A high CI value implies the gonad has already reached
maturity. However, CI is not always linearly correlated to its
Gonad development for Anadara granosa breeding pattern. This can be shown from the comparison of
The recorded CI values for the samples indicated significant the monthly CI vs GI values. The GI value is an assumed
varying values every month for samples from the same indication of the breeding status. A sudden drop in GI value
sampling location as well as those from different sampling signifies the occurrence of spawning activities. From this
locations. The difference in the trend of CI value indicated analysis, there was no linear correlation between CI and GI
the status of the population of blood cockles throughout the values for samples from Banda Aceh and Penang. However,
Table 1. Mean monthly seasonal environmental parameter at the sampling areas from June 2009 to September 2010
Jun. Jul. Aug. Sep. Oct. Nov. Dec. Jan. Feb. Mar. Apr. May Jun. Jul. Aug. Sep.
Environmental parameter Average
2009 2009 2009 2009 2009 2009 2009 2010 2010 2010 2010 2010 2010 2010 2010 2010
Temperature (oC)
Banda Aceh
Minimum 26.32 24.27 24.29 25.43 26.90 25.00 25.39 25.79 22.96 22.84 25.47 21.87 25.90 22.82 25.27 26.00 24.78
Maximum 30.97 30.44 30.45 29.88 32.61 31.93 31.95 32.48 32.93 32.10 31.40 31.03 30.93 32.11 30.98 30.38 31.41
Lhokseumawe
Minimum 28.82 28.82 26.81 28.07 27.71 27.03 28.45 28.06 28.96 28.48 28.60 28.23 27.95 28.23 27.23 27.40 28.05
Maximum 31.08 31.71 30.06 31.17 30.87 30.33 30.81 30.68 30.75 31.10 30.90 31.65 31.27 31.39 30.84 31.17 30.99
Pulau Pinang
Minimum 27.23 27.52 26.65 27.10 25.87 27.93 26.05 23.68 25.61 26.74 26.37 26.29 26.67 26.74 26.65 27.37 26.53
Maximum 31.63 31.10 31.45 30.60 30.90 31.43 30.58 28.35 30.46 31.71 31.13 30.68 31.53 31.32 31.45 31.30 30.98
Salinity (ppt)
Banda Aceh 32.27 31.35 29.98 27.47 30.06 27.20 26.45 29.68 31.50 31.16 29.30 30.71 30.85 31.29 30.45 28.27 29.87
Lhokseumawe 31.00 30.97 31.16 31.20 31.03 29.07 30.94 31.16 31.46 31.84 30.57 31.26 30.90 30.81 30.65 31.27 30.95
Pulau Pinang 29.33 28.52 26.39 26.87 27.35 26.13 25.23 28.06 29.00 28.94 28.67 26.48 29.70 29.32 31.06 30.40 28.22
pH
Banda Aceh 7.65 8.02 8.03 8.17 7.80 8.02 7.91 8.17 8.08 8.06 8.02 7.97 7.74 8.27 7.94 8.23 8.01
Lhokseumawe 8.13 7.88 8.04 8.06 8.17 8.13 7.98 8.21 7.89 8.18 7.91 7.84 8.08 7.93 7.99 8.13 8.03
Pulau Pinang 8.02 7.49 7.85 8.14 8.04 8.07 7.79 8.02 7.95 7.86 8.08 8.21 7.91 7.86 7.33 7.54 7.89
Dissolved oxygen (mg/L)
Reproductive Biology of Anadara granosa

Banda Aceh 6.53 6.81 6.96 6.05 5.97 6.12 6.05 5.95 6.10 5.84 6.23 5.86 6.68 5.84 6.32 5.47 6.17
Lhokseumawe 6.01 6.38 6.47 6.14 6.28 6.04 6.07 6.10 6.97 6.02 6.17 5.98 6.02 5.89 6.28 6.13 6.18
Pulau Pinang 7.20 5.20 5.20 4.90 5.13 5.21 5.09 5.29 5.20 5.64 5.87 5.39 7.67 5.64 6.29 5.87 5.67
Turbidity (NTU)
Banda Aceh 17.40 29.30 8.61 9.02 10.86 9.12 19.16 14.09 16.03 10.27 18.98 13.83 10.27 34.29 9.74 10.48 15.09
Lhokseumawe 43.20 30.50 36.50 66.90 31.60 15.18 103.00 93.67 29.13 37.30 64.92 38.95 35.30 49.98 46.90 98.30 51.33
Pulau Pinang 29.30 17.36 15.11 13.09 17.27 74.30 57.80 77.10 109.67 107.00 98.00 76.00 103.40 107.00 76.65 93.12 67.01
Orthophospate (mg/L)
Banda Aceh 0.05 0.03 0.04 0.13 0.03 0.02 0.00 0.40 0.00 0.07 0.13 0.07 0.03 0.09 0.08 0.53 0.11
Lhokseumawe 0.05 0.01 0.02 0.07 0.01 0.01 0.01 0.70 0.00 0.04 0.06 0.04 0.04 0.08 0.07 0.01 0.08
Pulau Pinang 0.10 0.05 0.08 0.06 0.03 0.01 1.00 0.01 0.16 0.13 0.09 0.08 0.52 0.13 0.15 0.81 0.21
83
 84

Table 1. Continued
Jun. Jul. Aug. Sep. Oct. Nov. Dec. Jan. Feb. Mar. Apr. May Jun. Jul. Aug. Sep.
Environmental parameter Average
2009 2009 2009 2009 2009 2009 2009 2010 2010 2010 2010 2010 2010 2010 2010 2010
Nitrate (mg/L)
Banda Aceh 0.71 0.03 0.11 0.73 0.75 0.01 0.04 0.05 0.05 0.09 0.23 0.63 0.18 0.05 0.53 0.65 0.30
Lhokseumawe 0.68 0.14 0.03 0.03 0.20 0.01 0.00 0.03 0.01 0.10 0.77 0.58 0.10 0.17 0.07 0.98 0.24
Pulau Pinang 0.80 0.02 1.30 0.73 0.03 2.02 0.03 1.02 0.11 0.14 0.61 0.42 0.64 0.14 0.65 1.76 0.65
Nitrite (mg/L)
Banda Aceh 0.05 0.02 0.02 0.03 0.75 0.01 0.03 0.03 0.03 0.03 0.05 0.05 0.05 0.03 0.04 0.05 0.08
Lhokseumawe 0.03 0.03 0.04 0.04 0.05 0.03 0.44 0.03 0.43 0.02 0.08 0.07 0.02 0.08 0.03 0.04 0.09
Pulau Pinang 0.03 0.03 0.04 0.06 1.09 1.68 0.18 1.00 0.13 0.12 0.09 0.03 0.14 0.12 0.39 0.87 0.37
Ammonia (mg/L)
Khalil, M. et al.

Banda Aceh 0.87 0.20 0.16 0.15 0.11 0.68 0.06 0.13 0.08 0.09 0.19 0.79 0.18 0.06 0.59 0.13 0.28
Lhokseumawe 0.19 0.17 0.25 0.25 0.14 0.35 0.23 0.30 0.19 0.27 0.39 0.49 0.27 0.21 0.43 0.24 0.27
Pulau Pinang 0.24 0.18 0.14 0.25 0.42 0.11 0.61 0.15 0.68 0.54 0.65 0.98 0.65 0.54 0.82 0.65 0.48
Phytoplankton density (cell/L)
Banda Aceh 1831.67 1446.67 851.67 1178.33 630.00 991.67 1201.67 385.00 1773.33 1388.33 1516.67 1785.00 1738.33 1283.33 1341.67 1108.33 1278.23
Lhokseumawe 1656.67 1365.00 711.67 2601.67 3010.00 1435.00 4001.67 1365.00 2986.67 2415.00 2333.33 2298.33 2415.00 1050.00 1003.33 2310.00 2059.90
Pulau Pinang 4340.00 4001.67 1470.00 11713.33 4340.00 4281.67 2636.67 4561.67 4235.00 5751.67 5693.33 6090.00 5728.33 5751.65 2905.00 7910.00 5088.12
 Reproductive Biology of Anadara granosa 85

at P > 0.05) has no significant correlation to their respective

GI, but there is a mild correlation for samples from Lhokseumawe
(r = 0.609 at P < 0.05). A negative correlation has also been
reported in studies of other bivalve species. Herrmann et al.
(2009) reported a negative correlation between CI and
gametogenesis cycle for Amarilladesma mactroides (Reeve,
1854). Mladineo et al. (2007) also reported zero correlation
between CI and GI for the bivalve Modiolus barbatus (Linnaeus,
1758). The same applies to Mercenaria mercenaria (Linnaeus,
1758) from the Gulf of Narragensett in the United States, as
reported by Marroquin-Mora and Rice (2008).
The GI values obtained throughout the year indicate high
diversity in reproductive patterns among the three sampling
locations. This is expected due to the differences in habitat
conditions as well as breeding seasons. Blood cockles from
all three sampling locations showed a rapid transition from
gonad development to maturation phase. GI analysis shows
that spawning activity happened every month throughout
the year with varying intensity. The GI value increases during
gametogenesis and decreases after spawning. The fast-paced
transition could be a strategy for the blood cockles to increase
the amount of gamete released whilst favorable environmental
conditions are present. This behavior is characteristic of the
reproduction of invertebrates in tropical regions. Species have
been shown to adopt opportunistic strategies to develop the
gonadal matter from energy available from food rather than
from energy stored inside somatic parts (Cárdenas and Aranda
2000). Freites et al. (2010) found that Anadara notabilis
exhibits a continuous reproductive cycle throughout the year
and that particulate organic matter, temperature and food
availability were regulating factors of the reproduction of A.
notabilis.
This study of blood cockles’ GI shows that it has a breeding
cycle lasting an average of 3~6 months across the three
sampling locations (Banda Aceh, Lhokseumawe and Pulau
Pinang). During the 16 month sampling period, four reproductive
cycles were observed. For the A. granosa population from
Banda Aceh (Indonesia), cycle I occurred from June to October
Fig. 8. Principle component analysis (PCA) plot for the reproductive 2009, cycle II from November 2009 to January 2010, cycle
factor component on Anadara granosa population. (a) III from February to April 2010, and cycle IV from April to
Banda Aceh, Indonesia; (b) Lhokseumawe, Indonesia; (c) September 2010. In Lhokseumawe (Indonesia), cycle I started
Pulau Pinang, Malaysia
from June to August 2009, cycle II from September 2009 to
a linear correlation between these values was noted for samples January 2010, cycle III from February to June 2010 and cycle
from Lhokseumawe. These were tested with the Pearson IV from July to September 2010. For the A. granosa population
correlation test, which indicated that CI values for samples in Pulau Pinang (Malaysia), cycle I started from June to
from Banda Aceh (r = 0.469 at P > 0.05) and Penang (r = 0.123 October 2009, cycle II from November 2009 to February
86 Khalil, M. et al.

2010, cycle III from February 2010 to April 2010 and cycle IV spawning period was determined to be parallel between the
from April 2010 to September 2010. All three populations two opposing sexes, a scenario known as synchrony. According
started the first cycle around June 2009 and ended the fourth to Levitan (1993), synchrony in gonad development of bivalves
cycle also around the same time, September 2010. The population is crucial to increase the possibility of effective mating.
from Lhokseumawe (Indonesia) showed a tendency to spawn Extended spawning durations from one to two months is a
faster compared to the other two populations. However, during common breeding strategy for bivalve species. Such a strategy
the third cycle, populations from Banda Aceh (Indonesia) is essential to maintain the cockle population over time within
and Penang (Malaysia) exhibited a more rapid and shorter cycle its habitat. Generally, sporadic gamete mating will happen
lasting approximately 2~3 months, compared to Lhokseumawe concurrently under suitable surrounding conditions. Blood
(Indonesia) which took about 5 months. cockles for all three sampling locations, and in general, exhibit
a tendency to be characterized as bivalve brachidictics, which
Breeding pattern of Anadara granosa means they are capable of undergoing a continual breeding
Generally, the bivalve breeding process is characterized cycle throughout the year, with varying spawning intensity
by a continual and seasonal pattern (Ceballos-Vazquez et al. every month. Pathansali (1966), Narasimham (1988) and Broom
2000), and is iteroparous in nature, continually and repeatedly (1983) reported that A. granosa in Peninsular Malaysia and
breeding throughout its entire life span (Dame 1996). Bivalves India has a spawning season throughout the year with no
give birth to their young by means of gametogenesis. This apparent seasonal pattern. In comparison, the spawning season
process is then followed by the release of one or several of Archidae (genus Anadara) is presented in Table 2.
gametes. The process of rearranging the empty gonad with The information on the reproductive cycle of A. granosa
new gametes for the next cycle always is the signal for the provided by this study is crucial for initiating its commercial
beginning of a new breeding cycle (Gosling 2003). Variation in aquaculture value, as well as for the sustainable management
the breeding trend amongst cockle populations of different of wild stocks. In the future, data on spawning periodicity
geographical locations makes it difficult to determine a might be used to identify trochophore or veliger larvae in
pattern of gonad development. A well balanced distribution wild habitats and for seed collection activities. When bivalve
of males to females in blood cockles is supported by the sex culture production depends on natural seed supply, the timing
ratio analysis done in this study. Gonad development and of seed collection is critical since the potential brood stock is

Table 2. Comparison of spawning period with the highest intensity of releasing gamete in genus Anadara
Species Location Spawning period Sources
A. granosa Banda Aceh, Indonesia September and October Present study
A. granosa Lhokseumawe, Indonesia Jun and September Present study
A. granosa Pulau Pinang, Malaysia October, February to March Present study
A. granosa Perak December Pathansali (1966)
A. granosa Phuket, Thailand October to November Boonruang and Janekarn (1983)
A. granosa Pulau Pinang, Malaysia August to September Broom (1983)
A. granosa Selangor, Malaysia September and November Broom (1983)
A. granosa West coast, Thailand August and November Suwanjawat and Parnrong (1990)
A. granosa Pattani bay, Thailand September, December and July Suwanjarat et al. (2009)
A. trapezia Sydney, Australia end of summer Sullivan (1961)
A. broughtoni Japan Jun to August Kanno and Kikuchi (1962)
A. senilis Nigeria coast October and November Yoloye (1974)
A. broughtoni Great Bay, Japan July to September Dzyuba and Maslennikova (1982)
A. trigonopsis New Zealand winter and summer Booth (1983)
A. trapezia Sydney water, Australia winter Hadfield and Anderson (1988)
A. descripta Sydney water, Australia winter and autumn Hadfield and Anderson (1988)
A. inaequivalvis Black sea, Turkey Mei and November Sahin et al. (2006)
Anadara/Scapharca kagoshimensis Ariake bay, Japan autumn to early of summer Yurimoto et al. (2008)
A. notabilis Northeastern of Venezuela Jun and October Freites et al. (2010)
 Reproductive Biology of Anadara granosa 87

primed for a short period of time. Information presented here strongly correlated with A. granosa reproduction is phytoplankton
indicates that quantitative methods (condition index and density. This variable is known to interact with and affect
gonadal index) are a precise indicator in A. granosa brood gonadal index and condition index as determined by gonadal
stock. development stage in the three A. granosa populations. Lodeiros
and Himmelman (1999) conducted a statistical analysis,
Factors that affected reproduction cycle of Anadara granosa namely multiple regression analysis, to see the relationship
in the northern region of the Strait of Malacca between environmental factors and reproduction of the bivalve
Gametogenesis is affected by the change and interaction Lima scabra. The conclusion of that study found that
of exogenous (temperature, salinity, light, food), and endogenous phytoplankton density was the only primary factor positively
factors (nervous system, hormones) that could determine correlated to the reproduction of L. scabra. Phytoplankton
the reproductive strategy of bivalve species (Ram et al. 1996; density is the principle factor influencing the reproduction
Utting and Millican 1997; Louro et al. 2003; Barber and of bivalves (Wacker and von Elert 2003; Villalejo-Fuerte et
Blake 2006; Magnesen and Cristophersen 2008). The principle al. 2005; Kang et al. 2006; Liu et al. 2006; Hernández-Olalde
component analysis (PCA) has shown that the principle et al. 2007; Calderon-Aguilera et al. 2010). Phytoplankton are
components which affected the reproductive cycle of the A. also known to be the main source of diet to anadarinid
granosa population in Banda Aceh were gonadal index, animals (Kasigwa and Mahika 1991).
condition index, phytoplankton density, orthophosphate, Gonadal maturation and the fertilization activities of A.
salinity, and water temperature. The principle component granosa that correspond with the high level of phytoplankton
analysis for Lhokseumawe showed that there were five density is a strategy to increase planktotrophic larval autonomy
variables affecting A. granosa reproduction, namely interaction by increasing the larvae growth rate. The duration of the
among gonadal index, condition index, phytoplankton density, planktonic phase can be reduced through optimal utilization
ammonia, and pH. The reproduction of the A. granosa population of the food source (phytoplankton). Himmelman (1975)
in Pulau Pinang also revealed the complex interaction of showed that a high content of phytoplankton in the aquatic
seven principle variables, namely interaction between gonadal environment will stimulate the reproductive period of invertebrate
index and environmental factors such as salinity, nitrite, organisms, particularly species that have pelagic larvae.
ammonia, phytoplankton density, turbidity and dissolved Ram et al. (1992) found that phytoplankton release a type of
oxygen. chemical substance that could stimulate the nervous system
Principle component analysis indicated that the environment of bivalves to make them release gametes.
parameters modifying the reproduction of A. granosa
populations were diverse and complex. This analysis also Acknowledgements
indicated that reproduction of A. granosa populations is
significantly affected by the interaction of local environment This project was supported by Universiti Sains Malaysia-
parameters. For example, water temperature was found to Postgraduate Research Grant Scheme. The authors would
be modestly interacting with and affecting components of like to thanks to Marine Sciences Laboratory Universiti Sains
reproduction in A. granosa in Banda Aceh. Dissolved oxygen, Malaysia, Ministry of Research Technology and Higher
nitrite, ammonia, and turbidity variables were only found to Education of the Republic of Indonesia, Universitas Malikussaleh
be specifically interacting and affecting the components of Indonesia, Malaysia Quarantine and Inspection Services
reproduction in the A. granosa population in Pulau Pinang, (MAQIS) Malaysia and Indonesia Fisheries Quarantine Service
however these variables were not the factors affecting for their continuous support in making this project a success.
reproduction in the A. granosa population in Banda Aceh
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