reproduction of the sea urchin pseudechinus magellanicus ... - UMAR

BULLETIN OF MARINE SCIENCE, 79(1): 127–136, 2006

REPRODUCTION OF THE SEA URCHIN PSEUDECHINUS MAGELLANICUS ECHINOIDEA: TEMNOPLEURIDAE FROM GOLFO NUEVO, ARGENTINA Ezequiel M. Marzinelli, Gregorio Bigatti, Juliana Giménez, and Pablo E. Penchaszadeh ABSTRACT Pseudechinus magellanicus (Philippi, 1857) is a common and abundant sea urchin along the Argentinean coast. Gonad index cycle was studied for this species in Golfo Nuevo, Patagonia, from May 2001 to March 2003. Comparative studies of gonad index and gonad histology were performed for the year 2002. Gonad index reflects gonadal stages, showing two spawning peaks for the studied period. Major spawning occurred in winter and a second spawning event occurred during summer. Males and females spawned synchronously. Male gonads recovered faster than females after spawning, suggesting that females invest more time and energy in gametogenesis. Gonads were resorbed by both sexes. Gonad index was negatively correlated with photoperiod and water temperature with a lag period of 3 mo.

Reproductive cycles of temperate echinoids are typically annual or semiannual (Lawrence, 1987; Pearse and Cameron, 1991). Gametogenesis is considered to be influenced by factors such as seasonal changes in photoperiod (Pearse et al., 1986; BaySchmith and Pearse, 1987), water temperature (Yamamoto et al., 1988; Pearse and Cameron, 1991), and nutrition (Lawrence, 1987; Lamare et al., 2002). There are very few studies on reproduction of echinoids from Argentina (Brögger et al., 2003). The small urchin Pseudechinus magellanicus (Philippi, 1857) is one of the most abundant echinoids in Argentinean waters. It is distributed around South America from off Rio de la Plata (35°S) in the Atlantic Ocean to Puerto Montt (41°S), Chile in the Pacific Ocean; it is also found on islands of the Antarctic Sea (Bernasconi, 1953). In Golfo Nuevo, Patagonia individuals of this species live on mixed gravel and sandy bottoms and amongst Macrocystis blades and holdfasts. Usually cryptic, they occur at depths of 5–20 m in Golfo Nuevo. Sea surface temperature (SST) ranges from 9 to 18 °C (Esteves and De Vido, 1980) and day length varies from 9 to 16 hrs at this locality. Pseudechinus magellanicus is an omnivorous species depending on the habitat (Penchaszadeh et al., 2004). Guisado (1995) described the larval development and settlement of P. magellanicus in Chile and Orler (1992) reported on gonadic development, morphology, and structure of the gonads, and sexual maturity of P. magellanicus in southern Argentina. The spawning patterns of P. magellanicus differed between localities (Orler, 1992; Guisado, 1995). Although most sea urchins have a defined annual reproductive cycle, the exact pattern of spawning for a species can vary among localities and years (Byrne et al., 1998). We aimed to determine the annual reproductive cycle of P. magellanicus from Golfo Nuevo, Patagonia Argentina through the histological examination of gonad development, identification of gonad index and sex ratio, as well as the relationship between environmental factors and gametogenesis.

Bulletin of Marine Science © 2006 Rosenstiel School of Marine and Atmospheric Science of the University of Miami

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Figure 1. Sampling site in Golfo Nuevo, Patagonia, South Atlantic Ocean.

Materials and Methods Samples of P. magellanicus (N = 341, size range: 9.3–24.1 mm test diameter) were collected monthly by SCUBA divers at depths between 5 and 10 m off Golfo Nuevo, Puerto Madryn (42°S, 65°W; Fig. 1) from May 2001 to March 2003. Mature specimens (test diameter >12 mm; Orler, 1992) (n = 9–20 per sample) were fixed in Bouin’s solution over 2 d and then preserved in 70% alcohol. The test diameter of the sea urchins was measured with a Vernier calliper. Total body wet weight and gonad wet weight were measured with a Mettler precision balance (0.0001 g) to calculate gonad index [GI = (gonad wet weight/total wet weight) = 100] (Grant and Tyler, 1983; Pearse and Cameron, 1991) discriminating sex by gonad squash (n varies per month, Table 1). To corroborate gonad index reliability, fixed gonads of individuals (n = 112) sampled during 2002 were embedded in paraplast, sectioned at 5 +m, and stained with hematoxylin and eosin. Sections were examined microscopically with each individual assigned to one of seven male and female gametogenic stages using standard methods (Pearse and Cameron, 1991): (a) recovery, (b) proliferation and growth, (c) partially mature, (d) mature, (e) spawning, (f) spent, (g) resorption (Table 2). Size-frequency distribution of oocytes and ova was examined per sample. We considered ova > 67.5μm to be mature when lacking a distinctive nucleus, detached from the ascinal wall, and lying centrally in the lumen of the ovary. The maximum diameter of 20–40 oocytes/ova with distinct nucleolus found in sectioned ovaries of five females per month was measured with an optical microscope. Histological images were taken with a Zeiss Axiostar microscope equipped with a Sound Vision 2.0 digital camera. Mean monthly water temperatures were obtained from daily measurement of the SST at Golfo Nuevo by Navy Coastal Ocean Model–Naval Research Laboratory (NCOM), U.S. Navy. Photoperiod was obtained from the U.S. Navy Observatory. Data Analysis.—Male to female sex ratio was examined using a Chi-square test. GI was correlated with photoperiod and surface water temperature to examine the factors influencTable 1. Number of males and females analyzed in the gonad index per month. Month M J Males 7 7 Females 10 5

J A S O N D J F M A M J J A S O N D J F M 9 5 7 8 8 7 7 11 9 9 6 11 6 8 4 3 1 8 6 3 4 9 13 13 9 6 7 10 8 5 5 5 6 9 6 9 8 8 11 12 11 2

MARZINELLI ET AL.: REPRODUCTION OF PSEUDECHINUS MAGELLANICUS

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Figure 2. Mean monthly gonad index (GI) ((SE) of males and females (≥ 12 mm test diameters). Surface water temperature from Golfo Nuevo taken from NCOM (Navy Coastal Ocean Model– Naval Research Laboratory), U.S. Navy. Photoperiod obtained from the U.S. Navy Observatory. Table 2. Description of male and female gametogenic stages used in this study. Gametogenic stages Recovery

Proliferation and growth

Partially mature

Mature

Spawning Spent Resorption

Females Ovary contains primarily darker stained oocytes attached to the ascinal wall ( 0.05) from 1:1. Gonad index data indicates that P. magellanicus undergoes an annual reproductive cycle in Golfo Nuevo (Fig. 2) with a pre-spawning peak during the austral winter months (June–July 2001, May–June 2002), and a post-spawning minimum in the late winter and spring months (September 2001, August 2002; Fig. 2). A second pre-spawning peak was observed during summer (January and December 2002). Female gonad index decreased faster than male gonad index after the winter spawning peak. Minimum (21 June) and maximum photoperiod (21 December) were followed by corresponding changes in water temperature (Fig. 2). Gonad index was negatively correlated with photoperiod (no lag; P < 0.05, r = −0.66) and water temperature (3-mo lag; P < 0.05, r = −0.58). There was no relationship between gonad index and test diameter (P = 0.19).


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Figure 4. Sectioned ovaries of Pseudechinus magellanicus: (A) proliferation and growth (scale bar = 50 +m), (B) mature (scale bar = 100 +m), (C) spent (scale bar = 50 +m) and (D) proliferation and growth (scale bar = 100 +m). m = oocytes, O = ova, f = nutritive phagocytes, g = germinal cells, L = lumen.

Gonad Histology.—In total, 112 individuals sampled during 2002 were examined microscopically. Sectioned testes and ovaries of P. magellanicus in different gonad stages (Figs. 3, 4) are similar to other sea urchins. Histology indicated two spawning events, one during winter and another in summer. No hermaphrodite individuals were found. The gametogenic cycle of male and female P. magellanicus is annual in this population (Figs. 5, 6). Mean oocyte/ova diameter differed significantly among months (ANOVA: P < 0.001), reaching minimum values in February and August–September, and maximum values in April–June and December (Fig. 6). The maximum period of vitellogenesis occurred from February to June. Mean oocyte/ova diameter differed significantly among these months (Tukey: P = 0.003). There was also a significant difference in mean oocyte/ova diameter between June and August (Tukey: P = 0.0001), corresponding to the main spawning event. Finally, mean oocyte/ova diameter differed significantly between August and December (Tukey: P = 0.015), reflecting a period of gonad recovery and vitellogenesis. No significant differences in mean oocyte/ova diameter were found between June and December (Tukey: P = 0.27), corresponding to spawning peaks. In general, proliferation and differentiation of gametes occurred in late summer and autumn (February–May) and in spring (October–December). The presence of mature gonads was synchronous among males and females during the winter pre-

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Figure 5. Monthly oocyte/ova size-frequency distribution histograms from 2002. Arrows indicate mean oocyte/ova diameter per month. n = number of oocytes/ova measured per month.

spawning peak (April–June) (Fig. 7). From the histological analysis of sectioned ovaries we found that the variability in gonad stages between specimens collected during the summer spawning peak was higher than the one between specimens collected during the winter spawning peak. In summer, spawning in females occurred from partially mature ovaries, whereas male gonads were fully mature. Resorption of gametes was observed in both males and females after spawning (February and August). Discussion Pseudechinus magellanicus from Golfo Nuevo has an annual cycle with two spawning events: major spawning occurred in winter and a second smaller spawning peak was recorded in summer. Gonad index reflects this seasonal change in gametogenesis. In the smaller summer spawning, peak spawning in females occurred from partially mature ovaries. Recovery of gonads occurs more rapidly in males than in fe-


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Figure 6. Mean monthly oocyte/ova diameter (( SE) from females (n = 4–7) collected during 2002.

males. Females apparently spawn before males as indicated by the gonad index analysis and gonad histology. This difference could also be explained by high variability among individuals during the period of gamete maturity, or by males continuing gametogenesis longer than females so they have plenty of sperm available throughout the spawning period. In future studies, more individuals should be sampled per month. Nevertheless, we maintain that the gonad index is a good indicator of the gonad development in this species. Orler (1992) found a similar spawning pattern of P. magellanicus farther south at Ushuaia, Argentina (54°S, 68°W), but the cycle was delayed by 1 mo relative to Golfo Nuevo. This author also observed successive partial spawning events during summer. It is unusual to observe both winter and summer spawning peaks in the reproductive cycle of echinoids (Pearse and Cameron, 1991). In another study of the same species in Chile (41°S, 72°W) the first spawning peak was recorded in approximately the same winter months (Guisado, 1995). However, the second spawning event was recorded in spring, almost 2 mo before the one recorded in Golfo Nuevo during summer. McClary and Barker (1998) described the reproductive cycle of three sympatric species of the echinoid genus Pseudechinus (Mortensen, 1921) from New Zealand (45°S). They found that Pseudechinus huttoni (Benham, 1908) reproduce primarily during early summer (Dec–Jan), but mature gametes were present in the gonad throughout much of the year. Conversely, P. novaezealandiae (Mortensen, 1921) also from New Zealand, reproduce primarily during autumn and early winter (May–Jul). Pseudechinus albocinctus (Hutton, 1872) has an extended reproductive period, which peaked in early summer (Dec–Jan) followed by a gradual spawn-out to late autumn (May–Jun) (McClary and Barker, 1998). For all three species, males tended to be in spawning condition for a longer period than females. A remarkable similarity is observed in gametogenic patterns of the same genus from the southwestern Atlantic and south Pacific. On the Argentinean and New Zealand coasts, Pseudechinus male sea urchins are ready to spawn almost throughout the year, while females have restricted spawning periods. Phytoplankton blooms occur during the spring and summer months in the northern gulfs of Patagonia (Esteves et al., 1981; 1992) coinciding with an increase in pho-

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Figure 7. Monthly changes in the percentage of (A) female (n = 4–7) and (B) male (n = 2–7) P. magellanicus in each of the seven gametogenic stages from Golfo Nuevo, Patagonia, Argentina during 2002.

toperiod and water temperature in these temperate environments. These factors may be favorable for larval feeding and development. Correlative evidence from the present study suggests that photoperiod influences the reproductive cycle of P. magellanicus. Giese (1959) proposed that seasonal changes in photoperiod influence gametogenesis in marine invertebrates. In echinoids, this was experimentally confirmed by Pearse et al. (1986) with the sea urchin Strongylocentrotus purpuratus (Stimpson, 1857). Such experimental studies might corroborate the influence of photoperiod in the reproductive cycle of P. magellanicus, however, since gametogenesis occurs during both increasing and decreasing photoperiod, and mature animals spawn during both long and short days, it is difficult to determine how change in photoperiod or absolute photoperiod influences reproduction in this species. Seasonally changing water temperature may also regulate gametogenesis (Pearse and Cameron, 1991), and there was a significant correlation between gonad index and water temperature with a lag period of three months in this study. Other factors that could influence spawning include food availability (Lawrence, 1987; Pearse and Cameron, 1991; Lamare et al., 2002) and lunar cycle (Pearse, 1972; Pearse, 1975; Iliffe and Pearse, 1982).


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Acknowledgments Special thanks to V. Zavattieri, E. Zavattieri, and O. Wheeler for field support. We also thank J. Pearse and M. Brögger for literature supply, and J. Pearse and two anonymous reviewers for improving this paper. This work was partially supported by PICT 10975, PIP 02193, UBACyT X316 and The Explorers Club (E.M.M.).

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Date Accepted: 16 March, 2006.

Address: CONICET – Museo Argentino de Ciencias Naturales Bernardino Rivadavia, Av. Ángel Gallardo 470, (1045) Buenos Aires, Argentina. Present Address: (E.M.M.) Centre for Research on Ecological Impacts of Coastal Cities, Marine Ecology Laboratories, A 11, University of Sydney, NSW 2006, Australia. E-mail: Telephone: 61-2-9351 4931 (G.B.) Centro Nacional Patagónico CENPAT – CONCET, Bvd. Brown s/n, U9120ACV Puerto Madryn, Chubut, Argentina.