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A status assessment of perkinsiosis, bonamiosis, and mateiliosis in commercial marine bivalves from Southern Brazil.

ABSTRACT The protozoans Perkinsus marinus, Perkinsus olseni, Bonamia ostreae, Bonamia exitiosa, and Marteilia refringens are responsible for some of the most detrimental diseases in the production of cultivated shellfish worldwide and are classified as notifiable diseases by the World Organization for Animal Health (OIE). This study examined the general health status of wild and cultured bivalves from southern Brazil and included diagnostic tests for the presence of Perkinsus sp., Bonamia sp., and Marteilia sp. Cultured bivalves included the mangrove oyster Crassostrea gasar (syn. Crassostrea brasiliand), the brown mussel Perna perna, the lion's paw scallop Nodipecten nodosus, and wing pearl oyster Pteria hirundo. The mangrove oyster Crassostrea rhizophorae and the carib pointed venus clam Anomalocardia brasiliana (syn. Anomalocardia flexuosa) were collected from wild populations. A variety of parasitic or commensal organisms were detected by histology including Ancistrocoma-like and Spenophrya-like organisms; Bucephalus genus, Nematopsis sp., Steinhausia sp., and Tylocephalum sp.; unidentified trematode; unknown protozoan and metazoans; and an amoeba parasite. Hemocytic infiltration was most commonly associated with parasitized animals. Histological, culture and molecular diagnostic tests did not find any evidence of the presence of OIE-listed pathogens or related species in this study. Although the current survey did not identify any pathogens or diseases of concern, it provides baseline health assessment data for these species against which any future disease developments or significant changes in population health can be compared. These data are also valuable with respect to the development and implementation of public policies related to aquatic animal health.

KEY WORDS: OIE diseases, shellfish, pathology

INTRODUCTION

Historically bivalve molluscs such as the zigzag scallop Euvola (Pecten) ziczac (Linnaeus, 1758), carib pointed venus clam Anomalocardia brasiliana (Gmelin, 1791) [syn. Anomalocardia flexuosa (Linnaeus, 1767)], brown mussel Perna perna (Linnaeus, 1758), and the mangrove oysters Crassostrea rhizophorae (Guilding, 1828) and Crassostrea gasar (Adanson, 1757) [syn. Crassostrea brasiliana (Lamark, 1819)] have been important in Brazil as both food and income sources for many coastal communities. Over the past 20 y, the demand for bivalves has increased, which coupled with declining wild harvests has increased interest in bivalve aquaculture, including the introduction and culture of nonnative species such as the Japanese oyster Crassostrea gigas (Thunberg, 1793) (Poli 2004).

Bivalve culture in Brazil began in the 1970s, and since 1988 activities have increased to a scale that provides an important alternative source of income to small enterprise fisherman, who often operate as members of associations or cooperatives. Brazilian shellfish culture is dominated by the culture of the brown mussel, followed by the Japanese oyster. There is additional production of native species including the lion's paw scallop Nodipecten nodosus (Linnaeus, 1758) and the mangrove oysters Crassostrea rhizophorae and Crassostrea gasar (FAO 2004). Bivalves are primarily grown in the southern states of Brazil, with Santa Catarina responsible for more than 95% of the total mollusc production (EPAGRI 2014).

Relatively little is known about the health status, pathogens, and parasites of bivalve populations in Brazil (Table 1). The World Organization for Animal Health (OIE)-listed diseases of bivalve molluscs are those which are caused by Bonamia ostreae, Bonamia exitiosa, Marteilia refringens, Perkinsus marinus, and Perkinsus olseni (OIE 2011). Species of Perkinsus have been reported from oysters in the states of Ceara, Paraiba, Bahia, and Sergipe (Sabry et al. 2009, 2013, Brandao et al. 2013b, Da Silva et al. 2013, Queiroga et al. 2013, Da Silva et al. 2014, Da Silva et al. 2015, Queiroga et al. 2015) and in clams from Ceara (Ferreira et al. 2015). Two of these cases represent reports of an OIE-listed Perkinsus parasite. Da Silva et al. (2013) reported the presence of P. marinus based on rDNA region internal transcribed spacer and large subunit sequences in oysters Crassostrea rhizophorae, and Da Silva et al. (2014) and Queiroga et al. (2015) reported P. marinus and P. olseni by molecular analysis in Crassostrea gasar. Both oyster species were collected from Paraiba, northern Brazil. None of the OIE-listed bivalve diseases have been reported from other states in Brazil.

This study examined wild and cultured populations of economically important bivalves from the state of Santa Catarina, southern Brazil, for general health using histology techniques and for the presence of pathogens responsible for OIE-listed bivalve diseases using culture and molecular methods. Improved understanding of the baseline health status of Brazilian bivalve populations is important and is essential to ensure the continued successful development of the industry and to support the development of appropriate regulatory policies.

MATERIALS AND METHODS

Collection Sites and Processing

A total of 870 bivalves were collected in winter 2009 and summer 2010 from four locations in Santa Catarina, southern Brazil. Wild oysters, Crassostrea rhizophorae, were randomly selected from one rocky shore site (approximately 100 [m.sup.2]) at Praia da Ponta do Sambaqui, Florianopolis (27[degrees] 29' 26.49" S and 48[degrees] 32' 17.88" W). This site is approximately 500 m from areas where molluscs are cultured. Carib pointed venus clams Anomalocardia brasiliana were collected randomly from one site (approximately 200 [m.sup.2]) within the Reserva Marinha Extrativista do Pirajubae (Marine Extractive Reserve of Pirajubae), Florianopolis (27[degrees] 29' 18.8" S and 48[degrees] 32' 12.9" W). This reserve, managed by the Instituto Brasileiro do Meio Ambiente e dos Recursos Naturais Renovaveis, is one of the largest clam harvesting areas in Santa Catarina.

Samples of brown mussels Perna perna, mangrove oysters Crassostrea gasar, and winged pearl oysters Pteria hirundo (Linnaeus, 1758) were obtained from culture systems within the experimental area of the Federal University of Santa Catarina, Ponta do Sambaqui beach(27[degrees] 29' 18.8" Sand48[degrees] 32' 12.9" W), Florianopolis. This is the most important area of oyster seed production in southern Brazil. Samples of lantern net-cultured scallops Nodipecten nodosus were collected from the experimental area of Empresa de Pesquisa Agropecuaria e Extensao Rural de Santa Catarina (27[degrees] 12' 5.39" S and 48[degrees] 30' 47.7" W). This northern location is an important shellfish production area, especially for P. perna.

Live animals were transported on ice to the Nucleus for Aquatic Pathology Study laboratory in the Federal University of Santa Catarina for necropsy and tissue sampling. At necropsy, shell lengths were obtained, and each animal was examined for gross tissue lesions or other signs of disease. Tissue samples of the bivalves for diagnostic testing were removed using sterile techniques and treated as described below.

The number and size of each species examined is provided in Table 2. In the case of wild-collected Crassostrea rhizophorae and Anomalocardia brasiliana, subsamples of 40 of the 150 individuals examined for gross signs of disease were randomly selected for analysis by hemolymph smears, tissue imprints, and histology (Table 2). A random subset of animals previously sampled for diagnostic testing was also sampled for molecular analysis.

Microscopic Analysis

Ray's Fluid Thioglycollate Culture Method

Rectum and gill tissues were dissected into Falcon tubes containing Ray's fluid thioglycollate culture media (Ray 1954, 1966), to which antibiotics (500 U/ml penicillin, 500 U/ml streptomycin) and an antifungal (nystatin: 10,000 U/ml) were added. Tubes were incubated at room temperature (24.0[degrees]C [+ or -] 1.5[degrees]C) for 5 days in the dark. After incubation, the tissues were removed, macerated onto glass slides, and stained with Lugol's solution. Slides were examined first at 5x objective, then at 100x objective for the detection of hypnospores of Perkinsus spp. as recommended by the OIE (OIE 2011).

Tissue Imprints

Small pieces (2 x 2 mm) of heart, gonad, gill, and digestive gland were dried on absorbent paper, and four imprints of each tissue were made onto glass slides. Slides were air-dried, fixed in methanol (1 min), stained with Giemsa (2 min), washed in distilled water (1 min), air-dried, and then mounted with a coverslip using Evermount. Slides were examined first at 40x objective and then under oil immersion at 100x objective for the detection of Bonamia spp. and Marteilia spp. (OIE 2011).

Hemolymph Smears

Hemolymph samples were obtained from the adductor muscle using 21-gauge (for scallops, mussels, pearl oysters, and true oysters) and 26-gauge needles (for clams) attached to 1-mL sterile syringes. Samples were smeared onto glass slides to form a monolayer, air-dried, and stained with Giemsa as described in section Tissue Imprints. Slides were examined first at 10x, then at 40x objective, and finally under oil immersion at 100x objective for the presence of Bonamia spp. and Perkinsus spp. (OIE 2011).

Histopathology

To obtain a representative sampling of the visceral mass (which includes gills, gonad, digestive gland, foot, and mantle), a single transverse cross-section (approximately 5 mm thick) was excised from each individual following the procedures outlined in the works of Howard and Smith (1983) and Howard et al. (2004). Tissues were placed in cassettes, fixed for 48 h in Davidson's fixative (Shaw & Battle 1957), dehydrated, and embedded in paraffin. Then 5-pm sections were mounted on slides, deparaffinized, and stained with hematoxylin and eosin. Slides were examined using light microscopy at 10-100x objective for the presence of Bonamia spp., Perkinsus spp., and Marteilia spp. (OIE 2011), as well as signs of disease and/or other pathogens and parasites. The percent prevalence of each parasite was calculated as the number of parasitized animals divided by the total number of each bivalve species multiplied by 100 for each sampling (Bush et al. 1997).

Molecular Analysis: Polymerase Chain Reaction

For the detection of Bonamia spp. and Marteilia spp. by polymerase chain reaction (PCR), a small piece of gill and digestive gland tissue was preserved in 95% undenatured ethanol. Molecular diagnostic testing was conducted on 60 individuals each of Perna perna, Crassostrea gasar, Nodipecten nodosus, and Anomalocaria brasiliana (Table 2).

For each individual a pool consisting of approximately equal amounts of gill and digestive gland was created, and DNA was extracted using DNeasy Blood & Tissue Kit (QIAGEN) following the manufacturer's instructions. Deoxyribose nucleic acid concentration and purity was measured using a NanoVue spectrophotometer.

For the Marteilia refringens analysis, the PCR was performed according to the protocol by Le Roux et al. (2001) and OIE (2011), using the primer set M2A 5'-CCG-CAC-ACG-TTC-TTC-ACT-CC-3' and M3AS 5'-CTC-GCG-AGT-TTC-GAC-AGA-CG-3'. Each PCR reaction consisted of 48 [micro]l of PCR SuperMix (Applied Biosystems) and 2 [micro]l of DNA at a concentration of 100 ng/[micro]l. Amplification was carried out in a Mastercycler Gradient (Eppendorf) under the following conditions: initial denaturation at 94[degrees]C for 10 min, followed by 30 amplification cycles at 94[degrees]C for 1 min, 55[degrees]C for 1 min, and 72[degrees]C for 1 min, with a final elongation step at 72[degrees]C for 10 min. The PCR products were run on 1.5% agarose gels at 100 V for 40 min, and the presence or absence of amplified DNA of the expected product size (412 bp) was assessed by comparison with products obtained for positive controls (plasmid ligations of Marteilia type M DNA supplied by Isabelle Arzul, Laboratory for Mollusc Diseases and OIE Reference Laboratory for bonamiosis and marteiliosis, IFREMER, France).

For the detection of Bonamia spp., the PCR was performed according to the protocol by Cochennec et al. (2000) and OIE (2011), using the primer set BO 5'-CAT-TTA-ATT-GG-TCGG-GCC-GC-3' and BOAS 5'-CTG-ATC-GTC-TTC-GAT-CCC-CC-3'. The PCR reaction consisted of 48 [micro]l of PCR SuperMix and 2 [micro]l of DNA at a concentration of 100 ng/[micro]l. Amplification was carried as follows: initial denaturation at 94[degrees]C for 5 min, followed by 30 amplification cycles at 94[degrees]C for 1 min, 55[degrees]C for 1 min, and 72[degrees]C for 1 min, followed by a final elongation at 72[degrees]C for 10 min. The PCR products were run on a 1.5% agarose gel at 100 V for 40 min, and the presence or absence of amplified DNA of the expected product size (300 bp) was assessed by comparison with products obtained from the amplification of positive controls for Bonamia exitiosa and Bonamia ostreae (plasmid ligations of parasite DNA from each species again supplied as above by Isabelle Arzul).

RESULTS

The animals collected from aquaculture sites ranged in age from 1 to 3 years. The precise age of the wild collected bivalves were unknown, but their size and level of gonad development suggested that they were all adults (Table 2). During necropsy, orange lesions on the gonad of the brown mussel, Perna perna, were observed in 25% of the animals collected in both winter and summer. These lesions were not observed in the other studied species.

Ostracoblabe-Yike condition was observed in the summer 2010 in cultured oysters Crassostrea rhizophorae and Pteria hirundo, in 1 and 5% of samples respectively (Table 3). There was no evidence of these infections impeding valve closure and there was no damage evident in adjacent tissues.

Polychaetes, genus Polydora (Spionidae), were present in all of the bivalve species that were examined, occurring at highest prevalence in Crassostrea rhizophorae and Crassostrea gasar in both winter and summer samples. Polychaetes were also found albeit at lower prevalences in Perna perna, Nodipecten nodosus and Anomalocardia brasiliana in both seasons, and only occurred in Pteria hirundo in low numbers in the summer season (Table 3). Some individuals with the highest infestations also had yellow abscesses in the gonadal tissue and the adductor muscle in association with the mud blisters.

No other shell diseases were observed.

Microscopic Observations and PCR

A high prevalence of haemocytic infiltration (HI) was observed in many of the species examined (Table 3). The highest prevalences occurred in Anomalocardia brasiliana, Crassostrea rhizophorae, Crassostrea gasar and Perna perna with 28% to 98% of the animals displaying this condition. In A. brasiliana haemocytic infiltrations were associated with Bucephalus infections, which were commonly found in the digestive gland, gonad and mantle tissues. These were also associated with unidentified trematode infection in the connective tissues and with an unknown protozoan infection in the gill of A. brasiliana. In P. perna the haemocytic infiltration was again associated with Bucephalus infections, and was observed in 20% of the heavily infected animals (i.e., those with the parasite in more than three different organs) in the winter and 70% of the heavy infected animals in the summer.

During the analysis two ovarian parasites were observed an amoeba parasite (Figure 1 A) and a microsporidian (Figure 1B). The amoeba was found in the oocyte and gonad connective tissues of both oyster species, Crassostrea rhizophorae and Crassostrea gasar. Full descriptions of this parasite can be found in Suhnel et al. (2014). Da Silva et al. (2012) and Ivachuck (2013) observed a similar parasite described as unknown ovarian parasite in the oyster Crassostrea gigas, from the same region (Sambaqui Beach, Florianopolis). The highest prevalence of this amoeba was observed in winter samples of C. gasar (Table 3). This amoeba parasite may affect the oocyte viability, impacting reproduction performance, although more analysis would be necessary to fully elucidate the impact of this parasite on the reproductive viability of the host. The other ovarian parasite was a Steinhausia-like microsporidian (Figure IB), which was observed at low prevalence in summer Anomalocardia brasiliana clam samples, where it infected the oocyte cytoplasm (Table 3). No host reaction was observed in the infected animals. Sporocysts (diameter: 14.91 [+ or -] 0.08 [micro]m) contained spores at different development stages.

The bucephalid infecting the mussel Perna perna was identified as Bucephalus margaritae (Figure 1C). The species from Anomalocardia brasiliana could only be identified to genus as Bucephalus sp. (Figure 1D). In both cases the parasite was observed as sporocyst and cercariae stages, according to the description in Marchiori et al. (2010). In both summer and winter samples prevalence was low in P. perna mussel samples but higher in A. brasiliana clams. 50% of individuals in winter P. perna samples had heavy infections (parasite found in more than three organs; i.e. gill, gonad and digestive gland). All summer samples had heavy infections. This was not the case for A. brasiliana where 100% of the infected animals showed low-intensity infections in the connective tissue in the winter and summer. Whilst 100% of infections were associated with characteristic orange lesions in P. perna, such lesions were not observed in A. brasiliana necropsies.

An unidentified trematode (Figure 1E) was observed in the connective tissue of the clam Anomalocardia brasiliana. Nematopsis sp. (Apicomplexa: Eugregarinida) (Figure 1F) were present in 80% and 75% of Crassostrea rhizophorae, collected in the winter and summer samples respectively (Table 3), and was most commonly found in digestive glands, with occurrence to a lesser extent in gonad, gill and mantle tissues. No other bivalve species examined in the present study was infected by Nematopsis sp.

Ciliates (genus Sphenophrya, Figure 1G) were found at low infection rates in gills of summer Perna perna samples, and in winter and summer samples of Crassostrea gasar. Another ciliate species, genus Ancistrocoma (Figure 1H), was found at low prevalence in the digestive gland of Crassostrea rhizophorae (summer) and C. gasar (winter, Table 3).

Platyhelminth infections (Tylocephalum sp.; Figure 11; mean [+ or -] standard deviation sizes 97.66 [+ or -] 0.04 [micro]m by 80.34 [+ or -] 0.08 [micro]m) were observed in the connective tissues of Anomalocardia brasiliana and Crassostrea rhizophorae winter samples, with a higher prevalence seen in the latter (Table 3).

An unknown metazoan (Figure 1J) was observed in the gill filaments of Anomalocardia brasiliana in an encapsulated form, with higher winter prevalence than in summer samples. This parasite was also observed in low levels in the gill filaments of Perna perna winter samples (Table 3), but at a different development stage, with the diameter ranging from 135.36 to 190.15 [micro]m.

Three other unidentified metazoans were observed at low prevalence in different species in this study. In Pteria hirundo an unknown metazoan (99.47 [+ or -] 2.49 [micro]m diameter, Figure 1L) was found encapsulated in the digestive gland. In Perna perna, an unknown metazoan (93.00 [+ or -] 8.11 [micro]m diameter; Figure 1M) was observed encapsulated in the connective tissue of the mussel. An unknown metazoan (Figure 1N) in the lumen of the intestine and stomach of Nodipecten nodosus was found.

RFTM assays (Perkinsus spp.), tissue imprints (Bonamia spp. and Marteilia spp.), haemolymph smears (Bonamia spp. and Perkinsus spp.) and PCR analysis (Bonamia exitiosa, Bonamia ostreae and Marteilia refringens) showed all negative results for the studied bivalves.

DISCUSSION

Ostracoblabe-like condition observed in the present study showed similar characteristics described by Bower (2001), where nacre covered bumps which protrude from the shell into the adductor muscle are a typical sign of infection with the fungus Ostracoblabe implexa. Other shell diseases observed in the present study, Poly dor a, was observed by Sabry and Magalhaes (2005) causing mud blisters in the bivalve shells, with 100% of the infected animals, and are known to be associated with polychaete infections.

A high prevalence of haemocytic infiltration (HI) was observed in many of the species examined (Table 3). The highest prevalences occurred in Anomalocardia brasiliana, Crassostrea rhizophorae, Crassostrea gasar and Perna perna with 28% to 98% of the animals displaying this condition. The high haemocytic infiltration observed in C. rhizophorae and C. gasar could be related to gonad parasites infection as observed by Suhnel et al. (2014). In P. perna, infiltration was linked to unknown metazoans, but was also observed in unparasitized mussels, and may be related to reproductive physiology or to an unidentified potential agent.

Ovarian parasites observed in the present study as similar those observed by Da Silva et al. (2012) and Ivachuck (2012), described as unknown ovarian parasite in the oyster Crassostrea gigas, from the same region (Sambaqui Beach, Florianopolis). The other ovarian parasite, Steinhausia-like, has been observed in Brazil in the oysters C. gigas (Pontinha 2009, Sabry et al. 2011) and Crassostrea rhizophorea (Sabry et al. 2011), in the mussel Mytella guyanensis (Matos et al. 2005), and in the clam Anomalocardia brasiliana (Da Silva et al. 2012).

As bucephalids are often highly specific for their host species (Lauckner 1983), more studies are required to determine which species is associated with Anomalocardia brasiliana infections. Buchephalosis has been reported in the A. brasiliana clam in Brazil (Narchi 1966, Araujo & Rocha-Barreira 2004, Boehs & Magalhaes 2004, Boehs et al. 2010, Da Silva et al. 2012), as well as in the mussel Perna perna (Umiji et al. 1976, Magalhaes 1998, Lima et al. 2001, Da Silva et al. 2002, Galvao et al. 2006, Costa 2007, Marchiori et al. 2010, Da Silva et al. 2012). In this study bucephalosis was not observed in oysters (Crassostrea rhizophorae, Crassostrea gasar, Pteria hirundo) or the scallop (Nodipecten nodosus) (Table 3), but it has been reported in C. rhizophorae in more northerly sample sites by Nascimento et al. 1986 (Sao Paulo State). Bucephalid parasites have also been found in two commercially important fishery species, Mytella guyanensis (Boehs et al. 2010, Ceuta & Boehs 2012) and Lucinapectinata (Oliveira 2008) in Bahia. Bucephalids are known to cause parasitic castration in molluscs (Shelley 1988), which can affect the reproductive capacity of individuals in naturally recruiting wild populations.

The unidentified trematode observed in Anomalocardia brasiliana as similar those registered by Da Silva et al. (2012), with encysted phases in the kidney and mantle of A. brasiliana.

Histological signs of infection with Nematopsis sp., as registered in the present study, include oocysts with an irregular outline that are bound by a single thickened membrane (Padovan et al. 2003). These protozoa use bivalves as intermediate hosts for gametogonic and sporogonic life stages and complete their life cycle in the digestive tract of crustaceans. In the present study, there was little evidence of histopathological changes in any infected tissues, even for animals with more intense haemocytic infiltration. The infection of Crassostrea rhizophorae with Nematopsis sp. is not unusual as previous reports have found oocysts of this species ranging from 60% to 100% (Sabry et al. 2007) in a Jaguaribe River Estuary population (mainly in mantle and gill tissues), and 14% in subepithelial gill tissues in a northeastern population in Recife (Azevedo et al. 2005).

This is the first time that these Sphenophrya-like ciliate have been reported in the Crassostrea gasar oyster. While ciliates are commensal organisms, feeding from suspended particles in the gill and mantle of bivalves in this case (Kinne 1983), their presence in high abundance or during periods of environmental and/or physiological stress can cause damage to the host organism (Lauckner 1983).

An unknown metazoan (Fig. 1J) was observed in the gill filaments of Anomalocardia brasiliana in an encapsulated form, with higher winter prevalance than in summer samples. This parasite was also observed in low levels in the gill filaments of Perna perna winter samples (Table 3), but at a different development stage, with the diameter ranging from 135.36 to 190.15 [micro]m.

The unknown metazoan infecting Anomalocardia brasiliana observed, is morphologically identified as a copepod, but other authors studying gastropod gill lamellae suggest a similar encapsulated organism may be a metacercarian cyst (De Vico & Carella 2012). This is the first time that this unknown metazoan in A. brasiliana has been observed parasitizing the bivalves A. brasiliana and Perna perna. This study is also the first reported case of this unknown protozoan in the connective tissue and gill filaments of A. brasiliana, although a similar unidentified protozoan has been reported in the gill filaments of the mussel P. perna by Da Silva et al. (2012). Also the unknown metazoan observed in Nodipecten nodosus is the first known occurrence.

In general the parasites and commensal species found in this study have been previously reported in bivalves from Brazil and elsewhere, and are generally considered benign (Bower et al. 1992, Magalhaes & Ferreira 2006). The exceptions for this are the ovarian parasites and Bucephalus infections, which are known to affect the bivalve reproduction. More research into the unknown parasites observed in this study is needed to identify possible host relationships, using specific techniques such as in situ hybridization, molecular analysis and electron microscopy.

About OIE diseases, other studies have examined bivalves collected in the vicinity of the sample sites for the presence of Perkinsus species. Crassostrea rhizophorae collected from Sambaqui Beach and Ribeirao da Ilha, Florianopolis (SC) were reported to be negative for the parasites Perkinsus spp. and a Perkinsus beihaiensis-like species using the RFTM assay, whereas oysters collected in the estuary of the River Package, Fortaleza (CE), in northeastern Brazil tested positive (Sabry et al. 2009, Sabry et al. 2013). Also in Bahia, Brandao et al. (2013b) confirmed the presence of Perkinsus spp., by RFTM and PCR assays, which had infected the rectum, intestine and stomach of C. rhizophorae oysters. Da Silva et al. (2012) did not find Perkinsus spp. using RFTM assays in a range of the present studied bivalve species from Santa Catarina (Perna perna, C. rhizophorae, Crassostrea gigas and Anomalocardia brasiliana). In addition, these authors also examined two specimens of Crassostrea gigas for the presence of Marteilioides chungmuensis, both of which tested negative for this pathogen. Based on the histological and tissue imprint examinations, no Marteilioides refringens or Marteilia-related species were present in the bivalves tested. Furthermore the M. refringens-specific PCR results were all negative. Although species of Marteilia have been reported from a wide variety of bivalves and geographical locations, this genus has never been reported in South American waters (Table 1; Berthe et al. 2004).

The present study found no evidence of Bonamia species in the samples following examination of haemolymph samples, tissue imprints and histological sections. PCR results using a primer set specific for Bonamia ostreae and Bonamia exitiosa were also all negative. The OIE lists all susceptible hosts of B. exitiosa and B. ostreae as all species within the genus Ostrea. Bonamiosis has been reported in Ostrea chilensis from New Zealand (Dinamani et al. 1987, Doonan et al. 1994, Hine and Jones 1994, Hine 1996), in Ostrea angasi from Australia (Corbeil et al. 2006) Ostrea stentina from Tunisia (Hill et al. 2010) and in Ostrea edulis from Europe (Abollo et al. 2008, Carrasco et al. 2012). Also, bonamiosis has been reported in the genus Crassostrea, where Crassostrea ariakensis was infected by Bonamia exitiosa (Dungan et al. 2012). Although Bonamia have not been reported in Brazilian bivalves, it is important to note that oysters within the genus Ostrea have not been examined in Brazil. The puelche oyster, Ostrea puelchana, is found in the waters of southern Brazil, has been reported to be a host of Bonamia spp. based on collections made in Northern Argentina (Kroeck & Montes 2005, Kroeck 2010). Bonamia spp. has also been reported from the cultured Chilean oysters, Ostrea chilensis, in their native Chile (Campalans & Lohrmann 2009).

The results of this study support other survey work conducted over the past several years that have examined the health and disease status of bivalves in Santa Catarina State (Lima et al. 2001, Da Silva et al. 2002, Sabry 2003, Boehs & Magalhaes 2004, Sabry & Magalhaes 2005, Garcia & Magalhaes 2008, Pontinha 2009, Sabry et al. 2009, Marchiori et al. 2010, Sabry et al. 2011, Da Silva et al. 2012, Ivachuk 2013). This work contributes to baseline health assessments of C. gasar (syn. C. brasiliana), providing first reports of Sphenophrya-like ciliate infections. It has also provided health data on unknown and unidentified parasites or commensal organisms in a range of bivalve species in Brazil, including new reports as for Anomalocardia brasiliana, Crassostrea gasar and Pteria hirundo.

Based on the wild and cultured populations studied here, no OIE-notifiable bivalve diseases were found in the state of Santa Catarina in southern Brazil in the following species: Anomalocardia brasiliana (syn. A.flexuosa), Crassostrea gasar (syn. brasiliana), Nodipecten nodosus and Perna perna. Examination of the wild populations found no OIE-notifiable diseases, suggesting that they may be unlikely to be a harbouring source of disease transmission to cultured populations. Although the current survey did not identify any pathogens or diseases of concern, it provides important baseline health assessment data for these species against which any future disease developments or significant changes in population health can be compared. The results of the present study support the Ministry of Fisheries and Aquaculture in Brazil (Ministerio da Pesca e Aquicultura--MPA) in the development and implementation of public policies related to aquatic animal health. Currently reference laboratories are being developed for the diagnosis of OIE diseases for molluscs, as part of a National Network of Laboratories of the MPA (Rede Nacional de Laboratories do Ministerio da Pesca e Aquicultura--RENAQUA), which was created in April of 2012 in Brasilia.

Continued monitoring along the Brazilian coast (which would include the development of sampling programs covering broader geographical regions, times of the year, and also be extended to include other bivalve species) would lead to a more comprehensive overview of the health of bivalve populations. This information could then be used to inform managers and producers to enable the environmentally responsible aquaculture development of these economically and socially important species.

ACKNOWLEDGMENTS

We would like to thank to the Ministry of Fisheries and Aquaculture, Brazil, for the research support provided to the Fundacao de Amparo a Pesquisa do Estado de Santa Catarina (FAPESC), Brazil; the National Council of Technological and Scientific Development, Brazil (CNPq-558222/2008-0), for supporting the macro- and microscopical analysis and for providing a grant (CNPq-301072/2007-8); the Coordenajao de Aperfeifoamento de Pessoal de Nivel Superior (CAPES), Brazil, for post-doctoral scholarship support for SS; the Pacific Biologic Station and Vancouver Island University (VIU) in Canada for supporting the molecular studies; and the Laboratory for Molluscs Diseases and OIE Reference Laboratory, IFREMER, France, for providing molecular diagnostic protocols and positive control material for the OIE-listed pathogens. We would also like to thank the undergraduate students from Federal University of Santa Catarina (UFSC), Brazil, and Angeline de Bruyns and Nathan Stefani from Vancouver Island University for support in the diagnostic procedure, and the World Fisheries Trust, Canada, for facilitating the partnership between the VIU and the UFSC.

LITERATURE CITED

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Azevedo, C. & E. Matos. 1999. Description of Nematopsis mytella n.sp. (Apicomplexa), parasite of the mussel Mytella guyanensis (Mytilidae) from the Amazon Estuary and description of its oocysts. Eur. J. Protistol. 35:427-433.

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Boehs, G. & A. R. M. Magalhaes. 2004. Simbiontes associados com Anomalocardia brasiliana (Gmelin) (Mollusca, Bivalvia, Veneridae) na Ilha de Santa Catarina e regiao continental adjacente, Santa Catarina, Brasil. Rev. Bras. Zool. 21:865-869.

Boehs, G., A. Villalba, L. O. Ceuta & J. R. Luz. 2010. Parasites of three commercially exploited bivalve mollusc species of the estuarine region of the Cachoeira river (Ilheus, Bahia, Brazil). J. Invertebr. Pathol. 103:43-47.

Bower, S. M. 2001. Synopsis of infectious diseases and parasites of commercially exploited shellfish: Ostracoblabe implexa (shell disease) of oysters. Accessed October 2012. Available at: http://www. pac.dfo-mpo.gc.ca/science/species-especes/shellfish-coquillages/ diseases-maladies/pages/oioy-eng.htm.

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Ceuta, L. O. & G. Boehs. 2012. Parasites of the mangrove mussel Mytella guyanensis (Bivalvia: Mytilidae) in Camamu Bay, Bahia, Brazil. Braz. J. Biol. 72:421-427.

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Howard, D. W. & C. S. Smith. 1983. Histological techniques for marine bivalve mollusks. NOAA Technical Memorandum NMFS-F/NEC-25. Woods Hole, MA: Oxford Lab., National Marine Fisheries Serv. 97 pp.

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Kinne, O. 1983. Diseases of marine animals. Volume II. Introduction, Bivalvia to Scaphopoda. Hamburg, Germany: Biologische Anstalt Helgoland, pp. 477-879.

Kroeck, M. A. 2010. Gross signs and histopathology of Ostrea puelchana infected by a Bonamia exitiosa-like parasite (Haplosporidia). Dis. Aquat. Organ. 89:229-236.

Kroeck, M. A. & J. Montes. 2005. Occurrence of the haemocyte parasite Bonamia sp. in flat oysters Ostrea puelchana farmed in San Antonio Bay (Argentina). Dis. Aquat. Organ. 63:231-235.

Lauckner, G. 1983. Diseases of mollusca: Bivalvia. In: Kinne, O., editor. Diseases of marine animals: introduction, Bivalvia to Scaphopoda. Hamburg, Germany: Biologische Anstalt Helgoland, pp. 477-961.

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Lima, F. C., M. G. Abreu & E. F. M. Mesquita. 2001. Monitoramento histopatologico de mexilhao Perna perna da Lagoa de Itaipu, Niteroi, RJ. Arq. Bras. Med. Vet. Zootec. 53:203-206.

Magalhaes, A. R. M. 1998. Efeito da parasitose por Trematoda Bucephalidae na reproducao, composigao bioquimica e indice de condigao de mexilhoes Perna perna (L.). PhD thesis, Federal University of Santa Catarina, Florianopolis, Brazil.

Magalhaes, A. R. M. & J. F. Ferreira. 2006. Patologias e manejo em malacocultura. In: Siva-Souza, A. T., editor. Sanidade de organismos aquaticos no Brasil. Abrapoa, Maringa, Brazil.

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Matos, E., P. Matos & C. Azevedo. 2005. Observations on the intracytoplasmic microsporidian Steinhausia mytilovum, a parasite of mussel (Mytella guyanensis) oocytes from the Amazon River estuary. Braz. J. Morphol. Sci. 22:183-186.

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Narchi, W. 1966. Encontro de Bucephalopsis haimeana (Lacaze-Duthiers) no Brasil. Cienc. Cult. 18:22-24.

Nascimento, I. A., D. H. Smith, L. F. Kern & S. A. Pereira. 1986. Pathological findings in Crassostrea rhizophorae from Todos os Santos Bay, Bahia, Brazil. J. Invertebr. Pathol. 47:340-349.

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Suhnel, S., C. S. Ivachuk, A. L. C. Schaefer, V. A. Pontinha, M. L. Martins, A. Figueras, G. R. Meyer, S. R. M. Jones, J. C. Stewart, H. J. Gurney-Smith, A. R. M. Magalhaes & S. M. Bower. 2014. Detection of a parasitic amoeba (order Dactylopodida) in the female gonad of oysters in Brazil. Dis. Aquat. Organ. 109:241-250.

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SIMONE SUHNEL, (1,2) * STEWART C. JOHNSON, (3) HELEN J. GURNEY-SMITH, (2) CELENE DA S. IVACHUK, (1) ANA L. C. SCHAEFER, (1) CATHERINE A. THOMSON, (2) MARIA LUIZA T. MACIEL, (4) MAURICIO L. MARTINS, (1) RAQUEL ARANGUREN, (5) ANTONIO FIGUERAS (5) AND AIME RACHEL M. MAGALHAES (1)

(1) Nucleus for Aquatic Pathology Studies, Center of Agrarian Sciences, Federal University of Santa Catarina, Rodovia Admar Gonzaga, 1346, 88040-900, Florianopolis, Santa Catarina, Brazil; (2) Centre for Shellfish Research, Vancouver Island University, 900 Fifth Street, Nanaimo, British Columbia, V9R 5S5 Canada; (3) Pacific Biological Station, Fisheries and Oceans Canada, 3190 Hammond Bay Road, Nanaimo, British Columbia, V9T 6N7, Canada; (4) Ministry of Agriculture, Avenida Loureiro da Silva, 515, 90010420, Porto Alegre, Rio Grande do Sul, Brazil; (5) Spanish National Reference Laboratory for Mollusc Diseases, Instituto de Investigaciones Marinas, Consejo Superior de Investigaciones Cientificas (CSIC), Eduardo Cabello, 6, 36208 Vigo, Spain

* Corresponding author. E-mail: ssuhnel@gmail.com

DOI:10.2983/035.035.0116

TABLE 1.
Parasites, pathological conditions, and commensals reported to
occur in Brazilian bivalves.

                 Parasite,
                 pathological
Bivalve          condition or
species          commensal          Author             Site

Anomalocardia    Bucephalus sp.     Araujo and         Ceara, Santa
brasiliana                          Rocha-Barreira     Catarina
(syn.                               (2004), present
                                    study

                 Bucephalopsis      Narchi (1966)      Sao Paulo
                 haimeana

                 Cestoda            Boehs and          Santa Catarina
                                    Magalhaes (2004)

Anomalocardia    Encapsulation in   Da Silva et al.    Santa Catarina
flexuosa)        gills              (2012)

                 Encysted           Da Silva et al.    Santa Catarina
                 organisms          (2012)

                 Holothuriophilus   Boehs and          Santa Catarina
                 tomentosus         Magalhaes (2004)

                 Metacercaria       Da Silva et al.    Santa Catarina
                                    (2012)

                 Nematopsis sp.     Boehs et al.       Bahia
                                    (2010)

                 Perkinsus          Ferreira et al.    Ceara
                 beihaiensis        (2015)

                 Polydora           Boehs and          Santa Catarina
                 (Spionidae)        Magalhaes
                                    (2004), present
                                    study

                 Sphenia            Boehs and          Santa Catarina
                 antillensis        Magalhaes (2004)

                 Steinhausia sp.    Da Silva et al.    Santa Catarina
                                    (2012), present
                                    study

                 Tylocephalum sp.   Boehs et al.       Bahia, Santa
                                    (2010), present    Catarina
                                    study

                 Trichodina         Boehs and          Santa Catarina
                 (genus)            Magalhaes
                                    (2004), Da Silva
                                    et al. (2012)

                 Trematode          Boehs and          Bahia
                                    Magalhaes
                                    (2004), Boehs et
                                    al. (2010), Da
                                    Silva et al.
                                    (2012)

                 Turbellarians      Da Silva et al.    Santa Catarina
                                    (2012)

                 Unidentified       Boehs et al.       Bahia
                 metazoans          (2010)

                 Unidentified       Da Silva et al.    Santa Catarina
                 trematode          (2012), present
                                    study

                 Unknown ciliates   Boehs and          Santa Catarina
                                    Magalhaes
                                    (2004), Da Silva
                                    et al. (2012)

                 Unknown metazoan   Present study      Santa Catarina

                 Unknown            Present study      Santa Catarina
                 protozoan

Crassostrea      Ancistrocoma sp.   Pontinha (2009),   Santa Catarina
gigas                               Sabry et al.

                 Encapsulation in   (2011), Da Silva   Santa Catarina
                 gills              et al. (2012),
                                    Ivachuck (2012)
                                    Da Silva et al.
                                    (2012)

                 Nematopsis sp.     Sabry and          Santa Catarina
                                    Magalhaes
                                    (2005), Sabry et
                                    al. (2011)

                 Ostracoblabe       Sabry and          Santa Catarina
                 implexa            Magalhaes
                                    (2005), Pontinha
                                    (2009)

                 Polydora sp.       Sabry et al.       Santa Catarina
                                    (2011), Da Silva
                                    et al. (2012)

                 Polydora websteri  Sabry and          Santa Catarina
                                    Magalhaes
                                    (2005), Pontinha
                                    (2009), Da Silva
                                    et al. (2012)

                 Rickettsia-like    Pontinha (2009),   Santa Catarina
                                    Sabry et al.
                                    (2011), Da Silva
                                    et al. (2012)

                 Steinhausia sp.    Pontinha (2009),   Santa Catarina
                                    Sabry et al.
                                    (2011)

                 Sphenophrya sp.    Pontinha (2009),   Santa Catarina
                                    Sabry et al.
                                    (2011), Da Silva
                                    et al. (2012),
                                    Ivachuck (2012)

                 Tylocephalum sp.   Sabry and          Santa Catarina
                                    Magalhaes
                                    (2005), Sabry et
                                    al. (2011), Da
                                    Silva et al.
                                    (2012)

                 Trichodina sp.     Pontinha (2009),   Santa Catarina
                                    Sabry et al.
                                    (2011), Da Silva
                                    et al. (2012)

                 Turbellarians      Pontinha (2009),   Santa Catarina
                                    Sabry et al.
                                    (2011), Da Silva
                                    et al. (2012)

                 Unknown ovarian    Da Silva et al.    Santa Catarina
                 parasite           (2012), Ivachuck
                                    (2012)

                 Unknown ciliates   Da Silva et al.    Santa Catarina
                                    (2012)

                 Unknown            Pontinha (2009),   Santa Catarina
                 metazoans          Da Silva et al.
                                    (2012)

                 Urastoma sp.       Sabry et al.       Santa Catarina
                                    (2011)

                 Viral              Pontinha (2009),   Santa Catarina
                 gametocytic        Sabry et al.
                 hypertrophy        (2011), Ivachuck
                                    (2012)

Crassostrea      Amoeba (order      Suhnel et al.      Santa Catarina
rhizophorae      Dactylopodia)      (2014)

                 Ancistrocoma sp.   Nascimento et      Bahia, Ceara,
                                    al. (1986),        Santa Catarina
                                    Sabry et al.
                                    (2011), Zeidan
                                    et al. (2012),
                                    Sabry et al.
                                    (2013), Brandao
                                    et al. (2013a),
                                    present study

                 Bucephalus sp.     Nascimento et      Bahia
                                    al. (1986),
                                    Zeidan et al.
                                    (2012), Brandao
                                    et al. (2013a)

                 Nematopsis sp.     Nascimento et      Bahia, Ceara,
                                    al. (1986),        Santa Catarina,
                                    Sabry and          Sao Paulo
                                    Magalhaes
                                    (2005), Sabry et
                                    al. (2007),
                                    Sabry et al.
                                    (2011), Da Silva
                                    et al. (2012),
                                    Sabry et al.
                                    (2013), Brandao
                                    et al. (2013a),
                                    present study

                 Ostracoblabe-      Sabry and          Santa Catarina
                 like               Magalhaes
                                    (2005), present
                                    study

                 Polydora sp.       Sabry et al.       Ceara, Santa
                                    (2011), Da Silva   Catarina
                                    et al. (2012),
                                    Sabry et al.
                                    (2013), present
                                    study

                 P. websteri        Sabry and          Santa Catarina
                                    Magalhaes (2005)

                 Perkinsus sp.      Sabry et al.       Bahia, Ceara
                                    (2009), Sabry et
                                    al. (2013),
                                    Brandao et al.
                                    (2013b)

                 Perkinsus          Da Silva et al.    Paraiba
                 marinus            (2013)

                 P. beihaiensis     Da Silva et al.    Ceara, Paraiba
                                    (2013), Sabry et
                                    al. (2013)

                 Perkinsus          Da Silva et al.    Paraiba
                 olseni-Wkz         (2013)

                 Rickettsia-like    Sabry et al.       Santa Catarina
                                    (2011), Da Silva
                                    et al. (2012)

                 Steinhausia sp.    Sabry et al.       Ceara, Santa
                                    (2011), Sabry et   Catarina
                                    al. (2013)

                 Sphenophrya sp.    Nascimento et      Bahia, Santa
                                    al. (1986),        Catarina
                                    Sabry et al.
                                    (2011), Zeidan
                                    et al. (2012),
                                    Brandao et al.
                                    (2013a)

                 Tylocephalum sp.   Nascimento et      Ceara, Bahia
                                    al. (1986),
                                    Sabry et al.
                                    (2007), Zeidan
                                    et al. (2012),
                                    Sabry et al.
                                    (2013), Brandao
                                    et al. (2013a),
                                    present study

                 Trichodina sp.     Sabry and          Bahia, Ceara
                                    Magalhaes          Santa, Catarina
                                    (2005), Sabry et
                                    al. (2011),
                                    Sabry et al.
                                    (2013), Brandao
                                    et al. (2013a)

                 Unidentified       Da Silva et al.    Paraiba
                 Perkinsus sp.      (2012)

                 Unknown copepod    Sabry et al.       Santa Catarina
                                    (2011)

                 Unknown            Nascimento et      Santa Catarina
                 metazoans          al. (1986),
                                    Sabry and
                                    Magalhaes
                                    (2005), Da Silva
                                    et al. (2012),
                                    Brandao et al.
                                    (2013a)

                 Unknown ciliates   Da Silva et al.    Santa Catarina
                                    (2012)

                 Urastoma sp.       Sabry et al.       Bahia, Ceara,
                                    (2011), Zeidan     Santa Catarina
                                    et al. (2012),
                                    Brandao et al.
                                    (2013a), Sabry
                                    et al. (2013)

                 Viral              Sabry et al.       Santa Catarina
                 gametocytic        (2011), Da Silva
                 hypertrophy        et al. (2012)

                 Xenomas            Boehs et al.       Bahia
                                    (2009), Zeidan
                                    et al. (2012)

Crassostrea      Amoeba (order      Suhnel et al.      Santa Catarina
gasar (syn.      Dactylopodia)      (2014), present
                                    study

Crassostrea      Ancistrocoma sp.   Queiroga et al.    Paraiba, Santa
brasiliana)                         (2015), present    Catarina
                                    study

                 Copepod unknown    Da Silva et al.    Sergipe
                                    (2015)

                 Maladie du pied    Da Silva et al.    Sergipe
                 shell disease      (2015)

                 Nematopsis sp.     Queiroga et al.    Paraiba, Sergipe
                                    (2015), Da Silva
                                    et al. (2015)

                 P. marinus         Da Silva et al.    Paraiba
                                    (2014), Queiroga
                                    et al. (2015)

                 P. beihaiensis     Queiroga et al.    Paraiba
                                    (2015)

                 P. olseni          Da Silva et al.    Paraiba
                                    (2014), Queiroga
                                    et al. (2015)

                 Perkinsus sp.      Queiroga et al.    Paraiba, Sergipe
                                    (2013), Da Silva
                                    et al. (2015)

                 Polydora sp.       Da Silva et al.    Sergipe, Santa
                                    (2015), present    Catarina
                                    study

                 Prokaryote-like    Queiroga et al.    Paraiba
                 colonies           (2015)

                 Steinhausia sp.    Da Silva et al.    Sergipe
                                    (2015)

                 Sphenophrya-like   Present study      Santa Catarina

                 Turbellarians      Da Silva et al.    Sergipe
                                    (2015)

                 Tylocephalum sp.   Queiroga et al.    Paraiba
                                    (2015)

                 Viral              Queiroga et al.    Paraiba
                 gametocytic        (2015)
                 hypertrophy

                 Urastoma sp.       Queiroga et al.    Paraiba
                                    (2015)

Iphigenia        Unidentified       Boehs et al.       Bahia
brasiliana       metacercariae      (2010)

                 Unidentified       Boehs et al.       Bahia
                 metazoan           (2010)

                 Tylocephalum sp.   Boehs et al.       Bahia
                                    (2010)

Lucina           No parasite        Zeidan et al.      Bahia
pectinata        observed           (2012)

Mytella          Bucephalus sp.     Ceuta and Boehs    Bahia
guyanensis                          (2012), Zeidan
                                    et al. (2012)

                 Nematopsis sp.     Azevedo and        Amazon Estuary,
                                    Matos (1999),      Bahia
                                    Pinto and Boehs
                                    (2008), Boehs et
                                    al. (2010),
                                    Ceuta and Boehs
                                    (2012), Zeidan
                                    et al. (2012)

                 Rickettsia-like    Boehs et al.       Bahia
                                    (2010), Ceuta
                                    and Boehs
                                    (2012), Zeidan
                                    et al. (2012)

                 Sphenophrya sp.    Zeidan et al.      Bahia
                                    (2012)

                 Steinhausia        Matos et al.       Amazon Estuary
                 mytilovum          (2005)

                 Trematode          Boehs et al.       Bahia
                                    (2010),

                 Unidentified       Boehs et al.       Bahia
                 metazoans          (2010), Zeidan
                                    et al. (2012),
                                    Ceuta and Boehs
                                    (2012)

                 Urastoma sp.       Zeidan et al.      Bahia
                                    (2012)

Nodipecten       Polydora sp.       Present study      Santa Catarina
nodosus
                 Unknown            Present study      Santa Catarina
                 metazoans

Perna perna      Bucephalus sp.     Umiji et al.       Rio de Janeiro,
                                    (1976),            Santa Catarina e
                                    Magalhaes          Sao Paulo
                                    (1998), Lima et
                                    al. (2001), Da
                                    Silva et al.
                                    (2002), Galvao
                                    et al. (2006),
                                    Costa (2007),

                 Bucephalus         Marchiori et al.   Santa Catarina
                 margaritae         (2010),
                                    Suarez-Morales
                                    et al. (2010),
                                    Da Silva et al.
                                    (2012), Medeiros
                                    (2013), present
                                    study

                 Didemnum           Suarez-Morales     Santa Catarina
                 psammathodes       et al. (2010)

                 Copepoda           Suarez-Morales     Santa Catarina
                 (Monstrilla sp.)   et al. (2010)

                 Nematopsis sp.     Lima et al.        Rio de Janeiro,
                                    (2001), Da Silva   Santa Catarina
                                    et al. (2012),
                                    present study

                 Polydora sp.       Sabry and          Santa Catarina
                                    Magalhaes
                                    (2005), Costa
                                    (2007),
                                    Suarez-Morales
                                    et al. (2010),
                                    Da Silva et al.
                                    (2012), present
                                    study

                 Sphenophrya-Yike   Present study      Santa Catarina

                 Tylocephalum sp.   Da Silva et al.    Santa Catarina
                                    (2012)

                 Trichodina sp.     Sabry and          Santa Catarina
                                    Magalhaes (2005)

                 Unknown metazoan   Present study      Santa Catarina

                 Unknown            Da Silva et al.    Santa Catarina
                 protozoan in       (2012)
                 gills

                 Unknown ciliates   Da Silva et al.    Santa Catarina
                                    (2012)

                 Urastoma           Suarez-Morales     Santa Catarina
                 cyprinae           et al. (2010)

Pteria           Ostracoblabe-      Present study      Santa Catarina
hirundo          like

                 Polydora sp.       Present study      Santa Catarina

                 Unknown            Present study      Santa Catarina
                 metazoans

Tagelus          Parvatrema sp.     Da Silva et al.    Santa Catarina
plebeius                            (2009)

TABLE 2.
Sampling details and observations of bivalve molluscs collected
in Santa Catarina State, Brazil, during the winter (August 2009)
and summer (January 2010).

                                                      Mean shell
                     Age       Site sampling      length [+ or -] SD
Species            (years)        location               (mm)

From culture
  Pteria hirundo      3      Sambaqui Beach,      60.18 [+ or -] 1.60
  Perna perna         1        Florianopolis/SC   71.11 [+ or -] 4.03
                               (27[degrees]29'    70.53 [+ or -] 3.63
                               17.73" Sand
                               48[degrees]32'
                               23.88" W)
  Crassostrea         1                           52.01 [+ or -] 8.35
    gasar                                         52.67 [+ or -] 4.52
  Nodipecten         1.5     Enterprise for       67.08 [+ or -] 5.70
    nodosus                    Agricultural       66.42 [+ or -] 3.85
                               Research and
                               Extension,
                               Bombinhas/SC
                               (27[degrees]12'
                               5.39" S and
                               48[degrees]30'
                               47.7" N)
From wild
  Crassostrea        --      Ponta do Sambaqui    60.55 [+ or -] 7.90
    rhizophorae                Beach,             61.34 [+ or -] 4.54
                               Florianopolis/SC
                               (27[degrees]29'
                               26.49" S and
                               48[degrees]32'
                               17.88" W)
  Anomalocardia      --      Marine Extractive    21.63 [+ or -] 4.29
    brasiliana                 Reserve of         22.43 [+ or -] 5.31
                               Pirajubae,
                               Florianopolis/SC
                               (27[degrees]38'
                               32.63" Sand
                               48[degrees]32'
                               01.47" W)

                                            Hemolymph
                                          smears, tissue   Gross signs,
                                          imprints, and     RFTM, and
                   Length     Season      histopathology   tissue assay
Species             (n)      sampling          (n)             (n)

From culture
  Pteria hirundo     30     Summer/2010         30              30
  Perna perna        40     Winter/2009         40              40
                     40     Summer/2010         40              40
  Crassostrea        40     Winter/2009         40              40
    gasar            40     Summer/2010         40              40
  Nodipecten         40     Winter/2009         40              40
    nodosus          40     Summer/2010         40              40

From wild
  Crassostrea       150     Winter/2009         40             150
    rhizophorae     150     Summer/2010         40             150
  Anomalocardia     150     Winter/2009         40             150
    brasiliana      150     Summer/2010         40             150

Species            PCR (n)

From culture
  Pteria hirundo     ns
  Perna perna        30
                     30
  Crassostrea        30
    gasar            30
  Nodipecten         30
    nodosus          30

From wild
  Crassostrea        ns
    rhizophorae
  Anomalocardia      30
    brasiliana       30

Species included the cultured bivalves: winged pearl oyster P.
hirundo, the brown mussel P.perna, the mangrove oyster C. gasar
(syn. C. brasiliana), and the lion's paw scallop A. nodosus', and
bivalves from wild: the mangrove oyster C. rhizophorae and the
carib pointed venus A. brasiliana clam (syn. Anomalocardia
flexuosa). ns = not sampled.

TABLE 3.
Percent prevalence of pathology and parasites in bivalves
collected from Santa Catarina State, southern Brazil, during
the winter 2009 and summer 2010.

                                          Winter/2009

                                                   Wild-collected
    Pathological           Cultured bivalves          bivalves
      condition
    and organisms        P.p.     c.g.     N.n      C.r.     A.b.

HI                      42.5      2.5     2.5      27.5     97.5
Amoeba                   0       45.0     0         0        0
Ancistrocoma-like        0        2.5     0         0        0
Bucephalus sp.           0        0       0         0       97.5
Bucephalus margaritae   25.0      0       0         0        0
Nematopsis sp.           0        0       0        80.0      0
Ostracobla.be-like       0        0       0         0        0
Polydora sp.            12.5     75.0     5.0      84.0      0.3
Sphenophrya-like         2.5 *    5.0 *   0         0        0
Steinhausia sp.          0        0       0         0        0
Tylocephalum sp.         0        0       0        15.0      2.5
Unidentified             0        0       0         0        2.5
  trematode
Unknown metazoan         2.5 *    0       0         0       10.0 *
Unknown metazoans        5.0      0       7.5 *     0        0
Unknown protozoan        0        0       0         0        2.5 *

                                        Summer/2010

                                                          Wild-collected
    Pathological               Cultured bivalves             bivalves
      condition
    and organisms        P.h.    P.p.    C.g.     N.n      C.r    A.b.

HI                       5.0     67.5   42.5      2.5     32.5   57.5
Amoeba                   0        0      2.5      0       10.0    0
Ancistrocoma-like        0        0      0        0        2.5    0
Bucephalus sp.           0        0      0        0        0     57.5
Bucephalus margaritae    0       25.0    0        0        0      0
Nematopsis sp.           0        0      0        0       75.0    0
Ostracobla.be-like       5.0      0      0        0        1.0    0
Polydora sp.            33.3     22.5   97.5     10.5     96.6    0.5
Sphenophrya-like         0        0     22.5 *    0        0      0
Steinhausia sp.          0        0      0        0        0      2.5
Tylocephalum sp.         0        0      0        0        0      0
Unidentified             0        0      0        0        0      2.5
  trematode
Unknown metazoan         0        0      0        0        0      2.5 *
Unknown metazoans        7.5 *    7.5    0       15.0 *    0      0
Unknown protozoan        0        0      0        0        0      0

Cultured bivalves included brown mussels Perna perna (P.p.),
mangrove oysters Crassostrea gasar (C.g.) (syn. brasiliana),
lion's paw scallops Nodipecten nodosus (N.n.), and wing pearl
oysters Pteria hirundo (P.h.). Wild-collected bivalves were
mangrove oysters Crassostrea rhizophorae (C.r.) and carib pointed
venus Anomalocardia brasiliana clams (A.b.) (syn. A.flexuosa).

* Parasite or commensal first time reported for the species.
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Author:Suhnel, Simone; Johnson, Stewart C.; Gurney-Smith, Helen J.; Ivachuk, Celene Da S.; Schaefer, Ana L.
Publication:Journal of Shellfish Research
Article Type:Report
Geographic Code:3BRAZ
Date:Apr 1, 2016
Words:9657
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