Histology Gonad Based on Morphochromatically

GSJ: Volume 7, Issue 7, July 2019 ISSN 2320-9186 189

1 2 GSJ: Volume 7, Issue 7, July 2019, Online: ISSN 2320-9186 3 www.globalscientificjournal.com

4 HISTOLOGY GONAD BASED ON 5 MORPHOCHROMATICALLY-DEFINED OF GONADAL 6 MATURATION STAGES ABALONE (HALIOTIS SQUAMATA 7 REEVE, 1846), IN BANTEN COASTAL, INDONESIA 8 SYAMSUL BACHRY1 , DEDY DURYADI SOLIHIN1 RUDHY GUSTIANO2, KADARWAN 9 SOEWARDI3, NURLISA A. BUTET3 10 1Department of Biology, Faculty of Mathematics and Science, IPB 16680, Indonesia 11 2Institute for Freshwater Aquaculture and Fisheries Extension, 16151, Indonesia 12 3Department of Aquatic Resources Management, FPIK-IPB 16680, Indonesia 13 *E-mail: [email protected] 14 15 Abstract 16 The identification of the gonadal maturity stage in the specimen is very important

17 for determining the spawning season, this is because it can facilitate the process

18 of sustainable fisheries management such as abalone H. squamata. This study

19 aims to identify, histologically, gonads through morphochromatically-defined of

20 gonadal maturation. The results showed that four stages of gonadal development

21 in male and female abalone H. squamata were found where the four stages

22 included pre-proliferation, maturation, maturity, and consumption. Therefore

23 morphochromatic definitions can be used as a practical tool for identification of

24 gonad abalone H. squamata.

25 26 27 Introduction

28 Abalone is one type of marine mollusk that is of exotic and economical

29 value because the abalone shell is used for decoration and the meat is very

30 popular as one of the delicious and popular foods (Setyono, 2006). Abalone is

31 one of the commodities of marine products with demand in large quantities,

32 especially Asian countries including Japan, China, Taiwan, Indonesia, and others

33 (Sales & Britz, 2001; Gordon & Cook, 2013).

34 Abalone is a gastropod that broadcast spawners (Takami & Kawamura,

35 2017). Abalone gonads can be observed through the left side of the shell by

36 lifting the legs and epipodium (Cox,1962; Setyono, 2006). The gonads lining the

37 digestive gland are called canonical appendage, which is known as

38 hepatogonadal complex (Velez-Arellano et al. 2015). Generally, females gonads

39 have a greenish color and males are beige in color (Sobhon et al. 1999;

40 Singhakaew et al. 2003; Hadijah et al., 2013; Roux et al. 2013).

41 Abalone size during gonadal maturation differs according to type,

42 geography, and water quality conditions where the abalone lives (Capinpin et al.

43 1998). Some sizes of abalone when first gonad mature include of H. midae from

44 the east coast of South Africa showing a shell size of 20-25 mm (Wood & Buxton,

45 1996), H. asinina from the waters of Tanakeke Island in South Sulawesi has a

46 shell length > 60 mm (Hadijah et al., 2013) other abalone species such as the

47 Haliotis type hannai disc that is kept in hatchery have a shell length of 30 mm

48 (Awaji & Hamano, 2004). Abalone in the subtropical region is different at the

49 beginning of gonadal maturity. The H. cracherodii species get a shell size of 140

50 mm (Del Proo, 1992). In addition, histological studies of gonadal development in

51 the adult phase have been carried out in various species (Hooker & Creese,

52 1995; Wood & Buxton, 1996; Capinpin et al. 1998; Setyono, 2004). However,

53 scientific information regarding the morphology and histology of gonads in

54 abalone H. squamata from Indonesia waters has not been widely reported.

55 Therefore, the purpose of this study was to identify the stages of gonadal

56 development through histological descriptions the morphochromatically-defined

57 of gonad abalone H. squamata on the coastal of Banten, Indonesia.

59 Materials and Method

60 About 113 specimens were collected in February 2017. Specimens were

61 collected on the coastal of Banten, Indonesia (105º 53’630’ E, 06º 49, 87,4’ S).

62 Specimens are surgically removed from the hypogonadal complex. Each

63 hepatogonadal complex is photographed and examined to describe the color and

64 consistency of the gonads, and the size relative to the size of the

65 hepatopancreas. Male and female abalone gonads from several abalone

66 samples were fixed in 10% formalin solution and processed following standard

67 histological techniques: gonad specimens were inserted in paraffin, and 3-5 µm

68 thick sections were obtained and stained with hematoxylin-eosin (Humason,

69 1979). Criteria for determining the level of gonad maturity in H. squamata follow

70 (modification of Wood & Buxton 1996; Capinpin et al. 1998; Setyono, 2006,

71 Velez-Arellano et al. 2015). Sex determination of surviving abalone can be

72 observed visually by opening the side of the abalone shell. The color of the male

73 gonad is characterized by beige and female gonads characterized by green.

75 Result and Discussion

76 A total of 113 H. squamata specimens were examined, observations were

77 made based on gonad maturity in morphochromatic and histological features.

78 The gonadal characteristics found include four stages (Pre-proliferative,

79 development, maturation and total spawning) for the ovaries and testes (Table 1,

80 Figure 1 and Figure 2). Size of abalone H. squamata shell morphology found at

81 the time of observation was 4-7 cm.

82 Gonad maturation cycle in abalone occurs throughout the year. However,

83 abalone has its own image for the level of gonad maturity based on geography

84 and abalone living conditions. Thus the image of gonads can be used as a tool to

85 differentiate the stages of abalone gonad maturity based on the same level.

86 Several studies that have explained the stage of gonadal maturity of the Haliotis

87 genus based on histology (Wood & Buxton, 1996; Capinpin et al. 1998; Bilbao et

88 al. 2012).

89 Based on the observation of macroscopic features, histological

90 characteristics generally showed an increase in the number of stable cells in the

91 process of gonadal maturation. Thus an increase in the size of the gonad is

92 easily observed visually. This is also seen in the size of the hepatopancreas

93 which has decreased, because of the presence of nutrients transported to the

94 gonad. The color of the gonad changes when the cells mature inside, this is

95 because it shows the development of various levels of gonadal maturity based on

96 macroscopic features.

97 Observations based on this description have been successfully used in fish

98 species such as Labeo cylindricus (Booth & Weyl, 2000), where macroscopic

99 patterns have also been used to see ovarian development in crustaceans such

100 as Callinectes danae (Zara et al. 2013), which has been explained in terms of the

101 main macroscopic relationship and histological structure of the oviductal gland

102 morphology that correlates well in the macroscopic evaluation of molluscs such

103 as Octopus hubbsorum (Alejo-Plata & Gómez-Márquez, 2015), where

104 macroscopic features have been described for various stages of maturation in

105 gastropods of Hexaplex trunculus (Elhasni et al. 2010).

106 Each species has a variety of different stages of gonadal maturity, this is in

107 accordance with environmental conditions. But results revealed in this study

108 indicate a practical and accurate way of identification in determining the stages of

109 gonadal maturation, without the need to do a long and time-consuming

110 histological analysis.

111 Conculsion

112 Based on morphochromatic observations on gonad abalone H. squamata, four

113 stages of gonadal development were found including the pre-proliferation,

114 maturation, maturing and spent stages. In addition, the Criteria for TKG II (the

115 initial development stage) is a criterion that is mostly found in the observation of

116 male and female gonadal morphology in February 2017 in the waters of

117 Binuangeun, Banten Province.

118 Acknowledgment

119 This research was funded by Indonesian Government through Directorate

120 General of Higher Education (DIKTI), Ministry of Research, Technology and

121 Higher Education. We would like to thank for the Center for Research and

122 Development of Cultivation Sea from Gondol-Bali (CRDMAGB), fishermen and

123 collectors of abalon for their information.

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194

195

196 Table 1. Description stage of gonad maturity in H. squamata species Pre-proliferative Description Immature gonads are characterized by little or no germinal epithelium between the outer epidermis and the digestive gland (Fig. 1A and 1B). Female gonad has a greenish color and male gonad by beige (Fig. 2A and 2B). Maturing Gonads enter the gametogenesis process. Histology analysis revealed a number of oogonia began to associate with trabeculae (Fig. 1C); in males, trabeculae begin to form with the attachment of spermatogonia and spermatocytes; (Fig. 1D). Externally, this organ is still soft and the base of the hepatopancreas has not been surrounded by gonads. Gonad tissue covers the digestive gland 25% to 50% of the cross-sectional area. Female gonad has a greenish color while the male gonad by beige colored (Fig. 2C and 2D). Ripe Gonad maturity fully contains gametes. Histology reveals female gonads are almost full of mature oocytes (Gambar. 1E), while the male gonad shows that the trabeculae have begun to be covered by spermatozoa (Gambar. 1F). Female gonads show full ripe by covering 90% of digestive organs, as well as male gonads showing full development 85% envelop the digestive gland. While the gonadal organ ends appear enlarged for each gonad. The ovary shows a

bright green color, with the appearance of mature oocytes. Testicles show a bright beige color, with a viscous fluid containing spermatozoa (Fig. 2E and 2F).

Total spawning Gonads show evidence of gamete release (total spawning); histological observations showed that ovaries containing mature gametes were reduced and began to empty. Likewise with the testicular segment visible around the tubules, where the spermatozoa empty themselves, the lumen of the gonad is partially destroyed and the trabecula folds (Fig. 1H and 1G). Morphologically, the gonads look soft and inconsistent. Hepatogonad looks soft; the color is opaque green in females, while males gonad display with opaque beige color (Fig. 2H and 2G). 197

198

199

200

201 202 Figure 1. Histology images of morphochromatic differences in stage of 203 gonadal development in H. squamata: A) pre-proliferative female, B) pre- 204 proliferative male, C) maturing female, D) maturing male, E) ripe female, 205 F) ripe male, G) total spawning female, H) total spawning male. 206 Hepatopancreas (Hp), Spermatozoa (Sz), Trabeculae (Tr), Oocytes (Oc), 207 Nucleus (N), Nucleolus (NL), female gonads filled with oocytes (Ro). 208 Hematoxylin-eosin technique, were photographed with an Olympus 209 microscope at a 10× magnification.

210

211 Female Male

Pre-proliferative

Maturing

Ripe

Total spawning

212

213

214 Figure 2. Morphological features of various stages of maturation of the 215 gonads of Haliotis squamata: A) pre-proliferative female, B) pre- 216 proliferative male, C) maturing female, D) maturing male, E) ripe female, 217 F) ripe male, G) total spawning female, H) total spawning male. 218