Evidence for spawning aggregations of the endangered atlantic goliath grouper by Athila Bertoncini

Journal of Fish Biology (2016) doi:10.1111/jfb.13028, available online at wileyonlinelibrary.com

Evidence for spawning aggregations of the endangered Atlantic goliath grouper Epinephelus itajara in southern Brazil L. S. Bueno*†‡§, A. A. Bertoncini‡‖, C. C. Koenig¶, F. C. Coleman¶, M. O. Freitas‡**, J. R. Leite‡**, T. F. De Souza† and M. Hostim-Silva‡†† *Programa de Pós-Graduação em Oceanografia Ambiental, Universidade Federal do Espírito, Santo – Base Oceanográfica – UFES, 565 Rodovia ES 010, km 16, Aracruz, ES 29199-970, Brazil, †Instituto COMAR – Conservação Marinha do Brasil, 104 Helena Degelmann St, Joinville, SC 89218-580, Brazil, ‡Instituto Meros do Brasil, 67 Benjamin Constant St, Curitiba, PR 80060-020, Brazil, ‖Laboratório de Ictiologia Teórica e Aplicada (LICTA), Universidade Federal do Estado do Rio de Janeiro – UNIRIO, 296 Pasteur Av., Urca, RJ 22290-240, Brazil, ¶The Florida State University Coastal and Marine Laboratory, 3618 Coastal Highway, St Teresa, FL 32358, U.S.A., **Rede Abrolhos, Jardim Botânico St, 920, Rio de Janeiro, RJ 22.460-000, Brazil and ††Centro Universitário Norte do Espírito, Santo/Universidade Federal do Espírito Santo, Rodovia BR 101 Norte, Km. 60, São Mateus, ES 29932-540, Brazil In this study, seasonal numerical abundance of the critically endangered Atlantic goliath grouper Epinephelus itajara was estimated by conducting scuba dive surveys and calculating sightings-per-unit-effort (SPUE) at three sites in southern Brazil. Seasonal differences in size and reproductive condition of captured or confiscated specimens were compared. The SPUE differed significantly with season, increasing in late spring and peaking during the austral summer months. A significant effect was observed in the number of fish relative to the lunar cycle. All females sampled during the summer were spawning capable, while all those sampled during other seasons were either regressing or regenerating. What these data strongly infer is that the E. itajara spawning aggregation sites have been located in the southern state of Paraná and the northern state of Santa Catarina and summer is the most likely spawning season. Size frequency distributions, abundance and reproductive state were estimated and correlated with environmental variables. © 2016 The Fisheries Society of the British Isles

Key words: artificial reefs; endangered species; Epinephelidae; reef fish; South Atlantic.

INTRODUCTION Atlantic goliath grouper Epinephelus itajara (Lichtenstein 1822), the largest reef fish in the western Atlantic Ocean, is considered critically endangered throughout its range (IUCN, 2013). In the western Atlantic Ocean, it ranges from North Carolina to southern Brazil, including the Gulf of Mexico and the Caribbean Sea. In the eastern Atlantic Ocean, its distribution extends from Senegal to Congo, although it is rare in the Canary Islands (Ferreira et al., 2012) and is believed to be extinct in the eastern Atlantic Ocean §Author to whom correspondence should be addressed. Tel.: +55 47 96346873; email: lecobueno@gmail.com

L . S . B U E N O E T A L.

from Senegal to Congo (Craig et al., 2009). Epinephelus itajara can reach over 2 m in total length (LT ) and weigh up to 400 kg with late maturation at around 6–8 years (males between 110 and 115 cm LT , and females between 120 and 135 cm LT ) with longevity of at least 37 years (Bullock et al., 1992; Sadovy & Eklund, 1999). Epinephelus itajara form reproductive aggregations in shallow water (<50 m) (Ferreira et al., 2012; C. C. Koenig & F. C. Coleman, unpubl. data), with a preference for high-relief rocky and artificial reefs (C. C. Koenig & F. C. Coleman, unpubl. data). Their spawning aggregations may consist of >100 individuals (Bullock et al., 1992; Sadovy & Eklund, 1999; Koenig et al., 2007, 2011; C. C. Koenig & F. C. Coleman, unpubl. data). Domeier (2012) defined reproductive aggregations as single species that gather at specific times at specific locations at densities or numbers that are significantly higher than those found at the same site during non-reproductive times. These aggregations generally occur at the same time and same site annually (Johannes, 1978; Carter & Perrine, 1994; Sadovy et al., 1994; Domeier & Colin, 1997). Gerhardinger et al. (2009) using fishermen’s local ecological knowledge and Freitas et al. (2015), based on biological evidences suggested that the E. itajara spawn during the summer (January to March) in south Atlantic, similar to summer spawning (July to September) in the northern hemisphere (Bullock et al., 1992). Knowledge and protection of these spawning aggregations are key factors for the species’ persistence (Sadovy de Mitcheson & Colin, 2012). Fishers threaten species persistence by targeting spawning aggregations to increase catch per effort. In Brazil, E. itajara are protected by a fishing moratorium set in place in 2002 for 5 years (2002–2007), renewed in 2007 for another 5 years (2008–2012) and again in 2012 for additional 3 years (2012–2015). Illegal catches, however, continue (Giglio et al., 2014) which stifles stock recovery and reduces any ability to understand their ecology. During the processing of this manuscript the moratorium was renewed in October of 2015, protecting goliath grouper for more 8 years in Brazil (2015–2023). This study was initiated to provide critical data relevant to the effective management and conservation of E. itajara in Brazil. The main objective was to locate and verify E. itajara spawning aggregations in southern Brazil and to describe the physical and biological characteristics of spawning sites as well as seasonal timing of spawning.

MATERIALS AND METHODS S T U DY A R E A The study area was located in southern Brazil, between 25∘ and 27∘ south latitude in the western Atlantic Ocean (Fig. 1). The region has distinct seasonality with summer (late December through to late March) being the warmest months. During this time, moderate east and north-east winds predominate, bringing warm (up to 27∘ C) clear waters from the east. The weather is more variable in autumn (late March through to late June), and spring (late September through to late December) with an increase in large eastern and south-eastern swells and a coincident decrease in underwater visibility. The winter (late June through to late September) is dominated by cold fronts that bring very large swells from the south and south-east, decreasing water temperature (18∘ C) and increasing turbidity in coastal waters. The surface currents 3·6 m deep in this region in summer and spring, direct currents towards the coast and parallel to the coast in the autumn and winter (E. A. M. Stein & M. A. Noernberg, unpubl. data). Three artificial reefs were chosen as sampling sites: two in the state of Paraná–Balsa Norte (BN) and Marine Artificial Reefs (RAM), and one offshore in the state of Santa

R E P R O D U C T I O N O F E P I N E P H E L U S I TA J A R A

25° 30' S

Paranagua

25° 45' S

Guaratuba

RAM

Balsa Norte

26° 0' S

Monobóia 26° 15' S

Säo Francisco do Sul

26° 30' S 48° 30' W

48° 15' W

Fig. 1. Map of Brazil indicating the study area on the left and an expanded view of the study sites on the right including Balsa Norte (BN), Marine Artificial Reefs (RAM) and Monobóia (MB).

Catarina–Monobóia (MB). These three sites were selected after 7 years of dive samples at eight different reef areas. The selection criteria were frequency and abundance of E. itajara. The shipwreck Balsa Norte was intentionally sunk in January 2001 to form an artificial reef and is located 38 km offshore at a depth of 27 m. The ship is 76·3 m long × 11·4 m wide × 5 m high with large features and dark crevices (Fig. 2). The RAM site is composed of two reef areas, separated by 1 km, each composed of 30 concrete structures, block forms and reef balls, sunk in June 2000. The structures are 1·5 m high, spread on the sand in an area of 30 m × 30 m (Fig. 2). These structures occur 12 km offshore at a depth of 18 m over sand bottom. Since both areas compose one single site, they were sampled on the same day. The Monobóia is an artificial reef, formed by pipelines, manifolds and some concrete and metallic structures, installed in the 1970s. This reef is attached to a single point mooring buoy located 8 km offshore, where tankers moor to unload oil. The main artificial reef area is 50 m × 50 m and is composed of diverse materials spread on the sand at 25 m deep. A protective structure, located on top of the manifold, is 9 m long, 8 m wide and 2 m high. Flexible pipelines extend from the manifold to the surface buoy, and six heavy chains, arranged in a radial pattern, support the entire structure which extends from the bottom to the surface (Fig. 2).

D ATA C O L L E C T I O N Data were collected from: (1) underwater visual census (UVC) using scuba, (2) photographs of E. itajara taken during scuba dives, (3) live specimens obtained by hook-and-line and (4) dead specimens donated by law enforcement officers. All activities were authorized by licenses: SISBIO 15080-2 and SISBIO 31719-1. U V C S U RV E Y S Between 2007 and 2014, UVC surveys were conducted for E. itajara at all three study sites during each season, with greater emphasis during summer. The roving diver technique (RDT)

L . S . B U E N O E T A L.

RAM

Monobóia

Balsa norte

Fig. 2. Sketches and photographs of artificial reefs study sites: RAM, Monobóia and Balsa Norte. Drawings from R. Schlögel and photographs from L. S. Bueno.

(Jones & Thompson, 1978) was used, with the surveys conducted only when visibility exceeded 3 m. During UVC, each site was thoroughly surveyed including all crevices and the surrounding perimeter. Temperature, visibility and number of E. itajara encountered were recorded. Epinephelus itajara were photographed to determine size, with size estimations conducted using two parallel laser beams, 25 cm apart, when fish were perpendicular to the beams. In the photograph, laser dots appearing on the fish are used to determine the fish’s LT in cm size classes (Koenig et al., 2011).

C AT C H S U RV E Y S In addition to the diving surveys, E. itajara were captured with hook-and-line. After the fish was hooked, it was slowly brought to the surface, gently pulled onto a stretcher on the deck of the boat and strapped down. The fish was then vented using a trocar and cannula, eyes were covered with a wet towel (to avoid eye damage from the sun) and gills were irrigated with sea water from a pump. The fish were measured (cm LT ) and tagged using conventional dart tags for the purpose of identifying individuals during subsequent UVC or recapture. Gonad biopsies were obtained by inserting one end of a plastic tube (0·7 cm ID) through the gonoduct into the gonad; the other end was attached to a manually operated vacuum pump. The pump was used to suck gonad tissue into an in-line vial (135 ml). The contents were immediately preserved in 10% formalin for 24 h, and then transferred to 70% ethyl alcohol. After sampling, fish were released at the same site. Gonad samples were also obtained from dead fish donated by law enforcement officials. These samples were processed in the same manner as those from captured fish. Gonad tissue samples were embedded in paraffin, sectioned to 4–6 um, stained in haematoxylin-eosin and then examined under a compound microscope to determine sex and reproductive condition. For gonad analysis, five developmental phases were used following Brown-Peterson et al. (2011): immature (IM), developing (DV), spawning capable (SC), regressing (RG) and regenerating (RT). The IM phase corresponds to fish that have never spawned, characterized histologically in females by the presence of oogonia and primary growth oocytes through the perinuclear stage (Grier et al., 2009), as well as little space among oocytes in the lamellae and ovarian wall generally thin. In DV females, the ovary is beginning to develop, but not ready to spawn. The SC fish are developmentally and physiologically able

R E P R O D U C T I O N O F E P I N E P H E L U S I TA J A R A

to spawn; RG represents the cessation of spawning, and the RT phase corresponds to a sexually mature, but reproductively inactive individual.

D ATA A N A LY S I S Sightings data obtained during diving surveys were transformed into sightings-per-unit-effort SPUE (equation 1), thus taking into account the different dive sampling times (in min) for each survey. For the purpose of this index, 30 min was the standard effort time because this was the average time of surveys. y = nTt−1 (1) where y is SPUE, n the number of fish observed, T the duration in min of each survey and t is the 30 min to standardization efforts. A three-factor PERMANOVA (season × site × lunar phase) design was used to evaluate differences in SPUE by season. A one-way PERMANOVA was applied to evaluate differences in sightings per unit effort by months. Linear regression was used to evaluate the relationship between water temperature and SPUE, as well as the relationship between underwater visibility and SPUE. Approximately, a four-fold increase in spawning season abundance over average monthly abundance was considered as strong evidence for spawning aggregation formation (Domeier, 2012). The seasons were defined by the Gregorian calendar. Lunar phases were defined as new moon, first quarter, full moon and third quarter. Each phase consisted of the peak ± 3 days (=7 days total). The LT classes were created using the Sturges’s formula (Vieira, 2003).

RESULTS D I V I N G S U RV E Y

Between 2007 and 2014, 107 RDT surveys were distributed over the three study sites totalling 3040 min (50·7 h sampling effort; Table I). Epinephelus itajara were sighted more frequently during surveys at Balsa Norte (at least one present at 100% of the samples) followed by Monobóia (96·2%) and RAM (74%). Monobóia was also the site where the highest number of E. itajara was observed during a single survey (n = 54) (32 for RAM and 30 for Balsa Norte). SPUE of E. itajara was higher during summer at Monobóia, RAM and Balsa Norte. High values were also observed during spring at Monobóia and RAM. Comparison of seasonal mean abundance for spring and summer showed mean values (15 and 12) more than four times higher than autumn and winter (1·5 and 2·9) (Table II).

Table I. Characteristics of three primary study sites off southern Brazil Characteristics Site area (m2 ) Height of site (m) Number of surveys Total time (min) Mean ± s.d. effort time

Balsa Norte

RAM

Monobóia

870 5 25 635 29·2 ± 5·8

900 1·5 30 761 26·5 ± 8·7

2500 25 52 1644 28·8 ± 8·4

L . S . B U E N O E T A L.

Table II. Seasonal differences in sightings-per-unit-effort (SPUE; unit effort = 30 min survey) for Epinephelus itajara on artificial habitats in southern Brazil, showing number of dive surveys (n samples), and maximum abundance (max), minimum abundance (min) and mean ± s.d. SPUE Season

Samples (n)

Max

Min

Mean ± s.d.

Spring Summer Autumn Winter

23 47 27 10

22 54 4 5

8 0 0 1

15·2 ± 3·1 12·6 ± 12·8 1·5 ± 1·4 2·9 ± 1·5

PERMANOVAs of E. itajara SPUE showed significant differences among seasons (F 3,107 = 33·53, P < 0·05). Pair-wise comparisons showed differences among all seasons, except for winter and autumn (Table III). Furthermore, summer had many extreme values with high abundances exceeding other seasons. Spring showed high abundances in mid-December, a few days before the summer. Months that show greatest maximum values in abundance were February (n = 54) followed by January (n = 42), December (n = 34) and November (n = 20). Comparing sightings among months, January had the highest mean SPUE followed by February, December and November (Fig. 3). A sudden drop in abundances occurred in March, reaching a low value in July. The PERMANOVA one-way test of SPUE by months showed that November, December, January and February, each showed significant differences from the other months (P < 0·05). Size-class distributions measured by lasers (n = 126) and catches (n = 10) by season (Fig. 4) were examined. LT ranged from 50 to 230 cm. The most abundant LT classes in summer were 98–121, 146–169 and 170–193 cm, summer being the only season with specimens >194 cm. While the mean size of fish appeared to differ little among seasons (Table IV), the greatest range in LT and the largest fish were recorded during summer (Fig. 4) while the smallest (50 cm) was observed during autumn. G O NA D S A M P L E S

Gonad samples were obtained from 17 E. itajara (15 females and two males): 10 collected from the study sites, one that was found dead and six obtained from law Table III. Pair-wise a posteriori comparison of seasonal differences in Epinephelus itajara sightings-per-unit-effort (SPUE) (t-statistic on pseudo-F values) Season Summer × winter Summer × autumn Summer × spring Winter × autumn Winter × spring Autumn × spring

4·42 3·19 2·53 7·79 20·75 14·65

<0·05 <0·05 <0·05 >0·05 <0·05 <0·05

R E P R O D U C T I O N O F E P I N E P H E L U S I TA J A R A

SPUE

January March May July September November August October December February June April

Fig. 3. Monthly average SPUE (sightings-per-unit-effort) of Epinephelus itajara determined by roving diver surveys conducted from 2007–2014 at Monobóia, RAM and Balsa Norte artificial reefs in southern Brazil. Error bars represent the maximum values observed and black dots represent means.

enforcement officers. Confiscated specimens (n = 6) obtained during winter (16 July 2011) from areas close to the study sites were also sampled. Four of these were females (132, 144, 148 and 180 cm LT ) and two were males (136 and 147 cm). All females were in the RT stage [Fig. 5(a)], indicating that they were not reproductively active. Among the E. itajara collected, seven females (100–195 cm) obtained from RAM during December and January of 2013 and January and February of

Number of fish

25 20 15 10 5

21 8– 24 1

7· 9

3· 9

21 19 4–

9· 9 17 0–

5· 9

16 14 6–

1·

9 12 2–

98 –1 2

7· 74 –9

50 –7

3·

LT classes (cm) Fig. 4. Total length (LT ) class distributions of Epinephelus itajara from south Brazil relative to seasons ( , autumn; , spring; , summer; , winter), measured in situ using laser metrics, hook-and-line samples and confiscated specimens from 2007 to 2014. The dashed vertical line marks size at 50% maturity, according to Bullock et al. (1992) and Freitas et al. (2015).

L . S . B U E N O E T A L.

Table IV. Seasonal differences in the mean ± s.e. total length (LT ) of all Epinephelus itajara measured with lasers and caught from study sites off southern Brazil (pooled Balsa Norte, RAM and Monobóia) Season

LT (cm)

Autumn Spring Summer Winter Total

7 18 87 14 126

121·4 ± 12·0 125·0 ± 5·7 137·2 ± 4·6 126·4 ± 6·4 133·4

n, sample size.

2014 were at SC phase [Fig. 5(b)]. A partially decayed E. itajara found on 27 December 2012 was sampled revealing a female (230 cm LT ), also at SC phase. A female (159 cm LT ) captured on 4 February 2014 at RAM was in SC phase, at the actively spawning sub-phase with hydrated oocytes [Fig. 5(c)]. Another two females captured at the same site on 14 February 2013 (119 cm LT ) and 10 January 2014 (205 cm LT ) were in post-spawning condition with post-ovulatory follicles (POFs) [Fig. 5(d)]. C O R R E L AT I O N W I T H E N V I R O N M E N TA L VA R I A B L E S AND MOON PHASES

Assuming that November to February is the spawning season, the factors that may influence aggregating behaviour in this period were analysed. SPUE of E. itajara showed no correlation with either water temperature (r2 = 0·02, P > 0·05) or visibility (r 2 = 0·003, P > 0·05). Comparing the abundance among lunar phases, the full and new moons had higher SPUE mean and maximum values (Fig. 6; peak of error bar) when compared to other phases. The maximum value of SPUE was associated with the new moon phase (Fig. 6; peak of error bar). The PERMANOVA for SPUE showed that moon phase has a significant effect on E. itajara aggregation (P < 0·05). The pair-wise test showed that new moon is significantly more important (P < 0·05) than the full moon and second quarter.

DISCUSSION According to Sadovy de Mitcheson et al. (2008), spawning aggregations of many species have been severely disrupted by overexploitation and loss of habitat to the point of disappearing from traditional sites. This is a global phenomenon that brings a sense of urgency to the need to better understand how aggregations function wherever they occur (Nemeth, 2009). In southern Brazil in the 1950s, for instance, E. itajara aggregations were quite large and also heavily fished (Souza, 2000; Gerhardinger et al., 2006). Nowadays, most of the aggregations known from anecdotal references have disappeared without being even documented. Historically, little is known about the dynamics of reproductive aggregations of reef fish or the locations and timing of spawning in the South Atlantic Ocean. This study is the first to describe E. itajara

R E P R O D U C T I O N O F E P I N E P H E L U S I TA J A R A

Fig. 5. Photomicrographs of histological sections of ovarian biopsies of Epinephelus itajara sampled in south Brazil (Ă&#x2014;10 magnification): (a) female, 144 cm total length (LT ), sampled in July 2011, in regenerating (RT) phase (MB, muscle bundle; PG, primary growth oocyte), (b) female, 230 cm LT , sampled on 27 December 2012, showing spawning-capable reproductive phase (CA, cortical alveolar; PG, primary growth; Vtg, primary vitellogenesis oocyte; Vtg3, tertiary vitellogenic oocyte; GVM, germinal vesicle migration), (c) female, 159 cm LT showing capable of spawning (SC) phase in active spawning condition (H, hydrated oocytes) and (d) female 205 cm LT , sampled on 10 January 2014, in the post-spawning reproductive phase (POF, post-ovulatory follicle complex).

spawning aggregations in south Brazil, based on histological data and seasonal abundance recorded from diver surveys. Based on the definition of Domeier (2012), the abundance data indicate that spawning aggregations of E. itajara are formed near artificial reefs. Moreover, data also suggests that there is a seasonal component for the aggregation, with highest abundance values in summer months. Therefore, the extreme values and significant differences observed between the seasons are also strong evidences for the occurrence of spawning aggregations. According to Colin et al. (2003), to identify a spawning aggregation site, the area must meet two main criteria: (1) a sudden increase in the number of individuals in a certain location and certain time and (2) that the physical characteristics of the fish suggest imminent reproduction including changes in colour patterns, distended abdomens or the presence of hydrated eggs, post-ovulatory follicles or viewing the release of gametes in the water column. These two criteria were satisfied in this study: the number of E. itajara increased significantly during the summer, November to February, and fish were reproductively active (spawning capable, SC and RG phases, with hydration and POFs oocytes) during this time. Thus, with both of these criteria

ÂŠ 2016 The Fisheries Society of the British Isles, Journal of Fish Biology 2016, doi:10.1111/jfb.13028

L . S . B U E N O E T A L.

SPUE

New moon

First quarter

Full moon

Second quarter

Fig. 6. Mean values of SPUE (sightings-per-unit-effort) of Epinephelus itajara among lunar phases, for pooled months (November, December, January and February) from 2007–2014 in south Brazil. Peak of error bars indicate maximum number observed.

satisfied, it is likely that the spawning sites and seasons for E. itajara have been correctly identified. Summer spawning has also been confirmed in the south-eastern U.S.A. (Colin, 1990; Bullock et al., 1992; Eklund & Schull, 2001; Koenig et al., 2011). Spawning of E. itajara occurs at night according to Mann et al. (2009). Spawning was not observed in this study because the surveys were conducted during the day, but the observation of night-time spawning is difficult even in the best of conditions (C. C. Koenig & F. C. Coleman, unpubl. data). Nevertheless, behaviours were seen that are believed to be related to courtship and spawning such as ‘stacking behaviour’ and colouration changes were observed, both of which were reported by Colin (1990) in his description of E. itajara on spawning sites in south-western Florida. The three study sites are located near large estuarine areas (Paranaguá, Guaratuba and Babitonga Bay). This proximity may be a factor for E. itajara choosing these as spawning areas, since juveniles are mangrove-dependent during their first 5–6 years (Koenig et al., 2007). This hypothesis is reinforced by the fact that a surface current flows towards the coast in this region during spring and summer (E. A. M. Stein & M. A. Noernberg, unpubl. data) possibly transporting eggs and larvae to suitable mangrove habitats. Among the sites, Monobóia had the highest abundances of E. itajara during aggregation time and throughout the year, this may be related to two characteristics: (1) the site has the most complex structure, including many artificial reefs, anchors, chains, pipelines and concrete and (2) there may be an advantage to the fish in having the structure extend from the bottom to the surface. Epinephelus itajara avoid very cold water [temperatures below 15∘ C may be lethal (Sadovy & Eklund, 1999)], so the near-surface structure may provide refuge from cold water brought onto the shelf by upwelling events, known to occur in the area. Supporting this hypothesis, E. itajara were observed occupying the higher vertical structures during the presence of strong thermoclines.

R E P R O D U C T I O N O F E P I N E P H E L U S I TA J A R A

Fifty-five per cent of individuals observed during the dives were >130 cm LT . According to Bullock et al. (1992) and Koenig et al. (2011), this size is probably adult, but the size range of smaller individuals extended from 98 to 121 cm LT , suggesting either that all the individuals at the purported spawning sites did not actively participate in spawning or that size at maturity is lower in southern Brazil than it is in Florida (Bullock et al., 1992) and in Abrolhos Bank Brazil (Freitas et al. 2015), that all of the individuals present participated in spawning. The latter is likely because histological samples of ovaries from two females, 100 and 119 cm LT , were mature (spawning capable and post-spawning phases). Smaller sizes at maturity may be related to overfishing (Dayton et al., 2003) which occurred in this area. These findings, combined with the estimates of size structure, provide data required for stock assessment models, while replication of the visual surveys across sites and over time could provide an index of abundance within the study area (Porch & Eklund, 2004; Porch et al., 2006). Several studies (Gerhardinger et al., 2006, 2009; Mann et al., 2009; C. C. Koenig & F. C. Coleman, unpubl. data) have shown a lunar pattern to spawning of E. itajara. Similar lunar patterns of aggregation activity have been observed with the fish here, but acoustic recordings, such as those used by Mann et al. (2009) are needed to confirm that night-time sounds in Brazil are similar to those observed in the Gulf of Mexico. New technologies such acoustic telemetry and passive acoustic monitoring are needed to improve the knowledge of the location and nature of E. itajara spawning aggregations in Brazil. Protection of these aggregation locations should be a high priority, but the condition of mangroves, the primary juvenile habitat, should also be improved once the most productive habitats are found. Continued research will provide the necessary impetus for the conservation and production of this species. Based on the present experience and findings, long-term and intensive short-term monitoring strategies are recommended to fully characterize trends in seasonal abundance and habitat use for E. itajara. On the other hand, during this study, diving surveys at the MonobĂłia, the most important spawning site after the summer of 2012, were prohibited. This restriction, imposed by the organization maintaining the structure, seriously compromised ongoing studies of E. itajara spawning in south Brazil. N E E D S F O R M A NAG E M E N T A N D E N F O R C E M E N T

Illegal fishing of E. itajara in Brazil (Giglio et al., 2014), and more specifically in the study area, is very common. Unfortunately, apprehensions and punishment, such as the one carried out by Federal Maritime Police Special Core-NEPOM/SDF during the present research, are rare. As enforcement operations are uncommon, illegal fishing is not being controlled in the region. Two actions that could prove to be effective are the establishment of marine protected areas (MPA) around the presumed spawning areas and increased enforcement during the spawning season. An MPA called the National Marine Park of Currais Islands was established on 20 June 2013, this was an important first step because it protects one of the studied sites (RAM). Such protection should be expanded to include other spawning sites as well as increases in surveillance and enforcement. Enforcement is an important deterrent to illegal fishing, but raising awareness of the public about the value of E. itajara aggregations for dive tourism may be equally effective. Monetary benefits of ecotourism dives on Nassau grouper Epinephelus striatus

ÂŠ 2016 The Fisheries Society of the British Isles, Journal of Fish Biology 2016, doi:10.1111/jfb.13028

L . S . B U E N O E T A L.

(Bloch 1792) aggregations were described by Sala et al. (2003), and are also being used by dive boat operators in south-eastern Florida since multiple dive boats are ferrying divers out to see E. itajara aggregations nearly every day and the business is expanding (C. C. Koenig, pers. obs.). When government agencies realize that healthy populations and aggregations of E. itajara can increase revenues through ecotourism, there will be increased incentive for protection. In addition to illegal fishing, pollution and mangrove habitat destruction threaten the survival of E. itajara populations in Brazil. Koenig et al. (2007) have clearly demonstrated the importance of mangrove habitat to juvenile E. itajara in the south-eastern U.S.A. High water quality standards in mangrove and coastal habitats must be maintained or the consequences could be dire, not only for E. itajara, but also for many other estuary-dependent species as well. This study shows evidence for the formation of spawning aggregations of E. itajara in southern Brazil, and provides a starting point for additional research into the ecology and behaviour of this endangered species over a broader area. It is intended to raise awareness of the importance of the areas described here as highly significant to the recovery of E. itajara populations throughout southern Brazil. Through this awareness, management agencies must continue to take effective conservation measures that will lead to population recovery and therefore benefit both a limited fishery and an ecotourism dive industry. We are grateful to J. A. Alves, T. F. Souza, F. Darros, R. L. Velo and L. F. Machado for their active participation in this research. We also thank all Meros of Brazil Project team, which is sponsored by Petrobras, and all the team of Fish Ecology Lab (FSUCML). L.S.B. has received a scholarship from FAPES (Fundação de Amparo a Pesquisa do Espírito Santo) and Sandwich scholarship by CAPES Foundation, Ministry of Education of Brazil from PDSE programme. Thanks to P. C. Pinheiro, A. Cattani, V. Abilhoa and D. A. Moreira. We would like to acknowledge the support from COMAR Institute, Lancha Furacão and Submarine Serviços for providing important technical help for dive operations.

References Brown-Peterson, N. J., Wyanski, D. M., Saborido-Rey, F., Macewicz, B. J. & Lowerre-Barbieri, S. K. (2011). A standardized terminology for describing reproductive development in fishes. Marine and Coastal Fisheries: Dynamics, Management, and Ecosystem Science 3, 52–70. Bullock, L. H., Murphy, M. D., Godcharles, M. F. & Mitchell, M. E. (1992). Age, growth, and reproduction of jewfish, Epinephelus itajara, in eastern Gulf of México. Fishery Bulletin 90, 243–249. Carter, J. & Perrine, D. (1994). A spawning aggregation of dog snapper, Lutjanus jocu (Pisces: Lutjanidae) in Belize, Central America. Bulletin of Marine Science 55, 228–234. Colin P. L. (1990). Preliminary investigations of reproductive activity of the Jewfish, Epinephelus itajara (Pisces: Serranidae). In Proceedings of the 43rd Gulf and Caribbean Fisheries Institute (Goodwin, M. H. & Waugh G. T., eds). Miami, FL: Gulf and Caribbean Fisheries Institute. Colin, P. L., Sadovy, Y. J. & Domeier, M. L. (2003). Manual for the study and conservation of reef fish spawning aggregations. Society for the Conservation of Reef Fish Aggregations Special Publication 1, 98. Craig, M. T., Graham, R. T., Torres, R. A., Hyde, J. R., Freitas, M. O., Ferreira, B. P., Hostim-Silva, M., Gerhardinger, L. C., Bertoncini, A. A. & Robertson, D. R. (2009). How many species of goliath grouper are there? Cryptic genetic divergence in a threatened marine fish and the resurrection of a geopolitical species. Endangered Species Research 7, 167–174.

R E P R O D U C T I O N O F E P I N E P H E L U S I TA J A R A

Dayton, P. K., Thrush, S. & Coleman, F. C. (2003). Ecological Effects of Fishing in Marine Ecosystems of the United States. Arlington, VA: Pew Oceans Commission. Domeier, M. L. (2012). Revisiting spawning aggregations: definitions and challenges. In Reef Fish Spawning Aggregations: Biology, Research and Management (Sadovy de Mitcheson, Y. & Colin, P. L., eds), pp. 1–20. Dordrecht: Springer. doi: 10.1007/978-94-0071980-4_12 Domeier, M. L. & Colin, P. L. (1997). Tropical reef fish spawning aggregations: defined and re-viewed. Bulletin of Marine Science 60, 698–726. Eklund, A. & Schull, J. (2001). A stepwise approach to investigating the movement patterns and habitat utilization of goliath grouper, Epinephelus itajara, using conventional tagging, acoustic telemetry and satellite tracking. In Electronic Tagging and Tracking in Marine Fisheries (Sibert, J. R. & Nielsen, J. L., eds), pp. 189–216. Dordrecht: Kluwer Academic Publishers. Ferreira, B. P., Hostim-Silva, M., Bertoncini, A. A., Coleman, F. C. & Koenig, C. C. (2012). Atlantic goliath grouper – Epinephelus itajara. In Reef Fish Spawning Aggregations: Biology, Research and Management (Sadovy de Mitcheson, Y. & Colin, P. L., eds), pp. 417–422. Dordrecht: Springer. doi: 10.1007/978-94-007-1980-4_12 Freitas, M. O., Abilhoa, V., Giglio, V. J., Hostim-Silva, M., Moura, R. L., Francini-Filho, R. B. & Minte-Vera, C. V. (2015). Diet and reproduction of the goliath grouper, Epinephelus itajara (Actinopterygii: Perciformes: Serranidae), in eastern Brazil. Acta Ichthyologica et Piscatoria 45, 1–11. Gerhardinger, L. C., Medeiros, R., Marenzi, R. C., Bertoncini, A. A. & Hostim-Silva, M. (2006). Local ecological knowledge on the goliath grouper Epinephelus itajara. Neotropical Ichthyology 4, 441–450. Gerhardinger, L. C., Hostim-Silva, M., Medeiros, R. P., Matarezi, J., Bertoncini, A. A., Freitas, M. O. & Ferreira, B. P. (2009). Fishers’ resource mapping and goliath grouper Epinephelus itajara (Serranidae) conservation in Brazil. Neotropical Ichthyology 7, 93–102. Giglio, V. J., Bertoncini, A. A., Ferreira, B. P., Hostim-Silva, M. & Freitas, M. O. (2014). Landings of goliath grouper, Epinephelus itajara, in Brazil: despite prohibited over ten years, fishing continues. Natureza & Conservação 12, 118–123. Grier, H. J., Uribe-Aranzábal, U. C. & Patiño, R. (2009). The ovary, folliculogenesis, and oogenesis in teleosts. In Reproductive Biology and Phylogeny of Fishes (Jamieson, B. G. M., ed), pp. 25–84. Enfield, NH: Science Publishers. Johannes, R. E. (1978). Reproductive strategies of coastal marine fishes in the tropics. Environmental Biology of Fishes 3, 65–84. Jones, R. S. & Thompson, M. J. (1978). Comparison of Florida reef fish assemblages using a rapid visual technique. Bulletin of Marine Science 28, 159–172. Koenig, C. C., Coleman, F. C., Eklund, A. M., Schull, J. & Ueland, J. (2007). Mangroves as essential nursery habitat for goliath grouper (Epinephelus itajara). Bulletin of Marine Science 80, 567–586. Koenig, C. C., Coleman, F. C. & Kingon, K. (2011). Pattern of recovery of the goliath grouper Epinephelus itajara population in the southeastern US. Bulletin of Marine Science 87, 1–20. doi: 10.5343/bms.2010.1056 Mann, D. A., Locascio, J. V., Coleman, F. C. & Koenig, C. C. (2009). Goliath grouper (Epinephelus itajara) sound production and movement patterns on aggregation sites. Endangered Species Research 7, 229–236. Nemeth, R. S. (2009). Dynamics of reef fish and decapod crustacean spawning aggregations: underlying mechanisms, habitat linkages, and trophic interactions. In Ecological Connectivity among Tropical Coastal Ecosystems (Nagelkerken, I., ed), pp. 73–134. New York, NY: Springer. Porch, C. E. & Eklund, A. M. (2004). Standardized visual counts of goliath grouper off south Florida and their possible use as indices of abundance. Gulf of Mexico Science 2, 155–163. Porch, C. E., Eklund, A. M. & Scott, G. P. (2006). A catch-free stock assessment model with application to goliath grouper (Epinephelus itajara) off southern Florida. Fishery Bulletin 104, 89–101.

L . S . B U E N O E T A L.

Sadovy de Mitcheson, Y. & Colin, P. L. (Eds) (2012). Reef Fish Spawning Aggregations: Biology, Research and Management. Dordrecht: Springer. doi: 10.1007/978-94-0071980-4_12 Sadovy, Y. & Eklund, A.M. (1999). Synopsis of biological data on the Nassau grouper, Epinephelus striatus (Bloch, 1792), and the jewfish, E. itajara (Lichtenstein, 1822). NOAA Technical Report NMFS 146. Sadovy, Y., Colin, P. L. & Domeier, M. L. (1994). Aggregation and spawning in the tiger grouper, Mycteroperca tigris (Pisces: Serranidae). Copeia 1994, 511–516. Sadovy de Mitcheson, Y., Cornish, A., Domeier, M. L., Colin, P., Russell, M. & Lindeman, K. (2008). A global baseline for spawning aggregations of reef fishes. Conservation Biology 22, 1233–1244. Sala, E., Arbuto-Oropeza, P. G. & Thompson, G. (2003). Spawning aggregations and reproductive behavior of reef fishes in the Gulf of California. Bulletin of Marine Science 72, 103–201. Souza, H. S. (2000). O homem da ilha e os pioneiros da caça submarina, 2nd edn. Florianópolis: Editora Dehon. Vieira, S. (2003). Bioestatística: tópicos avançados. Rio de Janeiro: Elsevier.

Electronic Reference IUCN (International Union for the Conservation of Nature) (2013). IUCN Red List of Threatened Species. Available at www.iucnredlist.org (last accessed 25 August 2013).