Gametogenic patterns of the larviparous oyster Ostrea nomades from Karachi, Pakistan (northern Arabian Sea)
Abstract
The larviparous oyster Ostrea nomades from two different localities was examined histologically and found to be mostly hermaphroditic. However, pure males, pure females and indifferent oysters were also found. The hermaphrodites of O. nomades were classified into three categories: ambisexual, predominantly female and predominantly male. Gonadal development appears to take place throughout the year with increasing activity and ripening in the winter months, leading to immediate spawning and later resorption. The number of brooding larvae of O. nomades was low at both collection sites.
Introduction
Knowledge of the reproductive biology of larviparous oyster species of the genus Ostrea is mainly based on the studies of two commercially important species, the European flat oyster Ostrea edulis (Orton 1922, 1927, 1933; Orton & Amirthalingham 1931; Sato 1951; Loosanoff 1962; Millar 1962; Perusko 1967, 1969, 1970; Wilson & Simons 1985; Shpigel 1989; Ruiz, Martinez, Mosquera, Abad & Sanchez 1992) and the native or Olympic oyster Ostrea lurida (Coe 1931a, b, 1932, 1934; Cole 1942; Roman 1992; Santos, Downing & Chew 1993). All these studies provide information from the colder waters of Europe and North America respectively. However, Shpigel (1989) carried out studies on the gametogenesis of cultured O. edulis from the warm waters of Israel. Some information is also available on the reproductive biology of Ostrea puelchana (Morricono & Calvo 1978, 1980, 1983; Fernandez Castro 1987; Pascual, Iribarne, Zampatti & Bocca 1989), Ostrea angasi (Hickman, O'Meley & Sause 1988) from Australia and Ostrea chilensis from southern New Zealand (Jeffs & Hickman 2000).
Information on the biology of species of Ostrea from the tropical and subtropical waters of India and Pakistan is virtually non-existent. Ahmed (1971) reported the presence of one species of Ostrea, Ostrea folium (now identified as Ostrea nomades) from Pakistan and also gave its chromosomes counts (Ahmed 1973). There is no information available on the biology and distribution of this species with the exception of Iredale (1939) and Thomson (1954), who identified and reported this species from Australia. Siddiqui & Ahmed (2001a,b,c) have recently worked on the allometry, epibionts and systematics of this species. The present study provides for the first time information on the gametogenesis and seasonal gonadal development of O. nomades from the coast of Pakistan.
Materials and methods
Monthly sampling of O. nomades was carried out at low tides from two sites in Karachi, namely Buleji and Keamari (Fig. 1). A total of 505 specimens of O. nomades were collected from July 1992 to December 1993 and in April 1994 (to compensate for the small sample size in April 1993) from Buleji rocky ledge, which is a triangular platform extending into the open Arabian Sea. At this site, oysters are present in the small shallow pools in the low tide area facing maximum wave action.
Map showing collection sites. 1, Buleji; 2, Keamari Seawall.
From Keamari, 249 oysters were collected during October 1992 to December 1993, except for July and November 1993. Keamari seawall is an artificial breakwater, about 1.5 km long, extending into the open Arabian Sea from Karachi Harbour.
The oysters were fixed in Davidson's fluid (Shaw & Battle 1957). A 5-mm-thick section of oyster gonad was cut with a razor blade along a line extending from the lower corner of the labial palps across the stomach to the posterior end of the body. The gonad was then dehydrated, wax embedded, cut at 7 µm and stained in Delafield haematoxylin and eosin. The sexes were identified on the basis of histological slides.
Results
Sex categories
Depending on the amount and type of gonadal material present, the oysters were assigned to the following sex categories:
Unisexuals
Photomicrographs showing gonads of Ostrea nomades:(a) male;(b) female;(c) ambisexual;(d) predominantly male;(e) predominantly female. CT, connective tissue; F, follicle; Oc, oocyte; N, nucleus; n, nucleolus; Sg, spermatogonia; Sc, spermatocytes; St, spermatids; Sz, spermatozoa.
Hermaphrodites
- 3
Ambisexual (AMB): follicles containing almost 50% male and 50% female gonadal tissue (Fig. 2c).
- 4
Predominantly male (PM): follicles containing more than 50% male gonadal tissue, the rest being female gonadal tissue (Fig. 2d).
- 5
Predominantly female (PF): follicles containing more than 50% female gonadal tissue, the rest being male gonadal tissue (Fig. 2e).
Unsexed
- 6
Indifferent: collapsed or empty follicles devoid of any gonadal tissue, male or female (Fig. 2f).
Stages of gametogenesis
The male and female gonadal tissue in different sex categories was termed as male phase and female phase respectively. The stages of gametogenesis were characterized as stage 1 (resting), stage 2 (developing), stage 3 (ripe), stage 4 (partially spawned), stage 5 (fully spawned) and stage 6 (resorbing) (Table 1).
| Stages | Sexual phase | Description |
|---|---|---|
| Stage 1 (resting) | Indifferent | Follicles completely collapsed and had abundant connective tissue, but no trace of gonadal material |
| Stage 2 (developing) | Male | Spermatogonial and spermatocyte balls abundant, few spermatids and sperm balls present |
| Female | Mostly oogonia and attached oocyte and occasional free oocyte | |
| Stage 3 (ripe) | Male | Mostly spermatid balls, few spermatocyte balls and occasional spermatogonial balls present |
| Female | Free oocytes with distinct nucleus and nucleoli, few attached oocytes and oogonia present | |
| Stage 4 (partially spawned) | Male | Follicles partly empty, few sperm balls present |
| Female | Follicles partly empty, few free oocytes present | |
| Stage 5 (fully spawned) | Male | Follicles nearly empty containing residual sperm balls. In some cases, contracted follicles and connective tissue was evident. Phagocytes present |
| Female | Follicles nearly empty containing few mature ova. In some cases, contracted follicles and connective tissue was evident. Phagocytes present | |
| Stage 6 (resorbing) | Male | Follicles contracted, few in number, occasional residual sperm balls are present, connective tissue is abundant |
| Female | Follicles contracted, few in number, few residual free oocytes are present, connective tissue is abundant |
A comparison of the different gonadal stages in different sex categories of O. nomades from Buleji and Keamari indicated that gonadal development in the male and female phases of the same oyster was asynchronous in most of the samples. In the AMB and PF sex categories, the female phase was dominated by stage 3 and the male phase by stages 4 and 6. In PM oysters, the female phase was mostly in stage 6, whereas the male phase was dominated by stages 2 and 4. Among the unisexual females, stage 2 dominated, whereas in unisexual males, all the stages were found in almost equal proportion. No specimen was present in stage 5 in either sex (Fig. 3).
Percentages of different stages of gametogenesis in male and female phases of different sex categories (see text) of O. nomades.
Seasonal gonadal development
The seasonal gonadal development of male phase and female phase O. nomades was assessed by combing the male and female phases of all the sex categories. The oysters in stage 1 (resting) were termed indifferent.
Indifferent
At Buleji, indifferent oysters exhibiting stage 1 (resting) appeared during 7 months of the sampling period (Fig. 4). Their highest percentage (10.71) was observed in July 1993. At Keamari, they were found during October and November 1992 and from April to June. Their maximum number (25%) occurred in June 1993.
Seasonal gonadal development in O. nomades from Buleji. M, male phase; F, female phase; I, indifferent.
Male phase
At Buleji, no specimen in stage 2 (developing) was found in July 1992. This process resumed in August and continued until December. The maximum development (42.86%) was observed in January 1993 and lasted until June 1993. Stage 3 (ripe) specimens were evident in significant numbers in February 1993 (30.43%) and reached their peak in September 1993 (47.62%) followed by December (42.86%). The frequency of stage 4 (partially spawned) specimens started to increase in February 1993, and the maximum number was encountered in May 1993 (56.67%). Stage 5 (fully spawned) specimens were found from October 1992 to March 1993 and then in June to July 1993, November and April, the highest percentage (20) in July 1993. Stage 6 (resorption) occurred throughout the year except August 1993 with maximum activity (57.14%) in July 1992 (Fig. 4).
In O. nomades from Keamari, development (stage 2) in the male phase was noted in 8 months between January and December 1993. Its highest percentage (25) was observed in May 1993 and the lowest (8.33) in September 1993. Stage 3 (ripe) was found throughout the study period except for March and June 1993. The highest percentage (50) of such oysters was found in January 1993 and the lowest (6.67) in August. Male gonads in stage 4 (partially spawned) were found during the entire study period except June 1993. Their highest percentage (42.86) was observed in October 1993 and the lowest (12.90) in December 1993. The fully spawned stage (stage 5) was observed in January, February, March, October and December. Stage 6 (resorption) occurred throughout the study period, with the highest percentage (58.33) in September (Fig. 5).
Seasonal gonadal development in O. nomades from Keamari. M, male phase; F, female phase; I, indifferent.
Female phase
At Buleji, the female phase in stage 2 (developing) was found from August 1992 to January 1993. This stage was absent from February to April, reappeared in May 1993 and persisted until December 1993. Stage 3 (ripe) was present throughout the study period with a maximum (91.3%) in August 1992. Oysters in stage 4 (partially spawned) female phase were found in July 1992, May and July 1993. Their highest number (16.67%) was observed in July 1992. The fully spawned stage (stage 5) was evident during 7 months of the study period with percentages ranging from 2.86 in January to 16.67 in July 1993. The female resorbing stage (stage 6) occurred throughout the study period except August 1993. Its highest percentage (38.46) was observed in April (Fig. 4).
At Keamari, the female phase in stage 2 (developing) was observed in March, April, August, September and December 1993. Its highest percentage (19.35) was found in December and the lowest (5.41%) in April 1993. Stage 3 (ripe) was present during the entire study period. Their highest percentage (100) was observed in November, February, May and June 1993 (Fig. 5). Very few females in stage 4 (partially spawned) were found, and these occurred in January, April and October 1993. In no oyster was the female gonadal phase found in stage 4 (fully spawned). Female phase in stage 6 (resorbing) was present in October and December 1992, April, October and December 1993. Its highest percentage (27.78) was found in October 1992 (Fig. 5).
Brooding larvae
At Buleji, the brooding larvae in the mantle cavity of O. nomades were found from September to November 1992, in January, February, June, July, September, October and December 1993 and April 1994. Their highest percentage (41.17) was observed in June 1993 (Fig. 6).
Percentages of brooding larvae of O. nomades in different months from Buleji and Keamari.
In oysters brought from Keamari, the brooding larvae appeared only in March, September and October 1993. Such oysters constituted 3.03%, 2.5% and 3.44%, respectively, of the population (Fig. 5). During the entire study period at both sites, the brooding larvae occurred in very small numbers. They were white in colour.
Discussion
During the present study, most of the O. nomades examined histologically were hermaphrodites being AMB, PM or PF in character. Very few unisexuals, i.e. pure males and females, and indifferent oysters were found. Similar sex categories were also reported in the European flat oyster O. edulis by Orton (1927), who observed changes in sex from pure male to pure female and from these to hermaphrodites. Cole (1942) found alternative sex changes in O. edulis and O. lurida. Loosanoff (1962) examined O. edulis from Maine (USA) and found them to be hermaphrodites and classified them as strongly ambisexuals, predominantly males and predominantly females. He was hesitant in accepting the pure male and pure female categories of O. edulis identified by Orton (1927) on the basis of gonadal smears. Subsequently, Galtsoff (1964) drew attention to three species of Ostrea, namely O. edulis, O. lurida and Ostrea equestris, which undergo rhythmical changes in sexuality. Wilson & Simons (1985) reported the presence of both sexes in each gonad of O. edulis. Morriconi & Calvo (1978) reported Ostrea puelchana from Argentina to be a hermaphrodite, with predominantly male or predominantly female gonads in which developing or resting cells of the other sex were present. Pascual et al. (1989) found O. puelchana to be a protandric hermaphrodite with consecutive rhythmic sexuality.
The presence of a large number of hermaphrodites but a small percentage of males, females and indifferent oysters in the populations of O. nomades from the Karachi area may be indicative of the fact that, in warm subtropical environments, one gametogenic phase immediately follows the next phase without undergoing complete resorption. Hofmann, Klinck & Powell (1992) indicated that, in oysters residing in the higher latitudes, the reproductive pattern showed discrete spawning compared with oysters occurring at the lower latitudes, which show continuous spawning.
The occurrence of a greater proportion of ripe (stage 2) tissue in the female phase compared with the male phase, and the presence of a greater percentage of partially spawned (stage 4) and fully spawned (stage 5) oysters in the male phase, indicated that the male stages are completed more rapidly than the female stages, and that the change from female to male phase is more rapid than that from male to female phase. Yonge (1960) reported in O. edulis that change from female to male phase is very rapid, whereas the process of change from functional male to functional female is very slow. Morriconi & Calvo (1978) observed in O. puelchana that ovulation was followed by rapid maturation of males and sperm evacuation. This would suggest that, in PM O. nomades from Karachi, the change is taking place from female to male and, in PF and AMB oysters, the change is taking place from male to female. The transition from male to female and vice versa has also been reported by Cole (1942) and Mann (1979) in O. edulis.
During the present study, in O. nomades in both male and female phases, development (stage 2) took place almost throughout the year with increasing activity in the winter months, followed by immediate ripening and later spawning and resorption throughout the year. Similar results have been reported by several authors in the case of O. edulis (Orton 1933; Cole 1942; Loosanoff 1962; Wilson & Simons 1985), in which development was initiated in the colder months while the ripe stage occurred in the warmer months.
Brooding larvae were found in O. nomades in the present study in small numbers almost throughout the year. Therefore, the breeding season of the Pakistani O. nomades is a prolonged one, with ripe (stage 3) females found throughout the year. From this observation, it can be inferred that the larvae must be continuously released in small numbers. Ostrea puelchana, which has a narrow breeding season, was reported to produce a greater number of small embryos (Morriconi & Calvo 1980) in Argentinean waters. This study shows that the fecundity of O. nomades from Buleji and Keamari waters (in the Greater Karachi area) is low compared with that of the cold-water species of Ostrea, such as O. edulis and O. lurida, in which the mantle cavities are full of a large number of brooding larvae (as witnessed by one of us, M.A.), which makes those oysters ‘white sick’, ‘grey sick’ or ‘black sick’ in their respective breeding seasons. In contrast, O. nomades from Karachi waters were only seen to have white-coloured larvae in small numbers. It must be because of this small number of larvae that O. nomades does not develop large populations on the Karachi coast and occurs individually at some distance apart from each other in its habitat. The spawning season of this species lasts throughout the year with gaps in between, but it is certain that spawning occurs in both summer and winter, never restricting itself to the summer period alone as in some species of Crassostrea found in Pakistani waters (Ahmed 1980). In Ahmed's (1980) terminology, O. nomades would be included in the category of year-round spawners.
