Design and optimization of an experimental maintenance system for yellow clam broodstock Amarilladesma mactroides (Reeve, 1854) / Desenvolvimento e otimização de um sistema de manutenção experimental para reprodutores de marisco branco Amarilladesma mactroides (Reeve, 1854)

The yellow clam is a sand-burrowing bivalve that inhabits the dissipative beaches from southern Brazil to the north coast of Argentina. In the last decades, populations of this species have been impacted by mass mortality events, overfishing and other anthropogenic activities. The production of juveniles in captivity would allow feasibility studies to be carried out to restore the natural stock as well as the production in aquaculture systems. Given the scarcity of studies on the maintenance of this species in captivity, a culture system and a management protocol were developed and tested. Wild-caught clams (total length ≥50 mm) were used in a series of 14 day-long trials. Survival was higher in clams that were allowed to bury into the sand. A permanent ink marker covered with a thin layer of a quick-hardening adhesive proved to be a reliable method to tag clams. The maintenance of yellow clams in this system resulted in high survival and growth, increases in the condition factor and oocyte diameter, and a relative advancement of gonadal development.

O marisco branco é um bivalve de areia que habita as praias dissipativas do sul do Brasil até a costa norte da Argentina. Nas últimas décadas, as populações desta espécie têm sido afetadas por eventos de mortalidade maciça, sobrepesca e outras atividades antropogênicas. A produção de juvenis em cativeiro permitiria a realização de estudos de viabilidade para restaurar o estoque natural, assim como a produção em sistemas de aquicultura. Dada a escassez de estudos sobre a manutenção desta espécie em cativeiro, um sistema de cultivo e um protocolo de manejo foram desenvolvidos e testados. Mariscos branco selvagens (comprimento total ≥50 mm) foram utilizados em uma série de ensaios de 14 dias de duração. A sobrevivência foi maior nos mariscos que podiam ser enterrados na areia. Um marcador de tinta permanente coberto com uma fina camada de adesivo de endurecimento rápido provou ser um método confiável para marcar os mariscos. A manutenção dos mariscos neste sistema resultou em alta sobrevivência e crescimento, aumento do fator de condição e do diâmetro do ovócito, e um relativo avanço do desenvolvimento gonadal.

Design and optimization of an experimental maintenance system for yellow clam broodstock Amarilladesma mactroides (Reeve, 1854) / Desenvolvimento e otimização de um sistema de manutenção experimental para reprodutores de marisco branco Amarilladesma mactroides (Reeve, 1854)

The yellow clam is a sand-burrowing bivalve that inhabits the dissipative beaches from southern Brazil to the north coast of Argentina. In the last decades, populations of this species have been impacted by mass mortality events, overfishing and other anthropogenic activities. The production of juveniles in captivity would allow feasibility studies to be carried out to restore the natural stock as well as the production in aquaculture systems. Given the scarcity of studies on the maintenance of this species in captivity, a culture system and a management protocol were developed and tested. Wild-caught clams (total length ≥50 mm) were used in a series of 14 day-long trials. Survival was higher in clams that were allowed to bury into the sand. A permanent ink marker covered with a thin layer of a quick-hardening adhesive proved to be a reliable method to tag clams. The maintenance of yellow clams in this system resulted in high survival and growth, increases in the condition factor and oocyte diameter, and a relative advancement of gonadal development.

O marisco branco é um bivalve de areia que habita as praias dissipativas do sul do Brasil até a costa norte da Argentina. Nas últimas décadas, as populações desta espécie têm sido afetadas por eventos de mortalidade maciça, sobrepesca e outras atividades antropogênicas. A produção de juvenis em cativeiro permitiria a realização de estudos de viabilidade para restaurar o estoque natural, assim como a produção em sistemas de aquicultura. Dada a escassez de estudos sobre a manutenção desta espécie em cativeiro, um sistema de cultivo e um protocolo de manejo foram desenvolvidos e testados. Mariscos branco selvagens (comprimento total ≥50 mm) foram utilizados em uma série de ensaios de 14 dias de duração. A sobrevivência foi maior nos mariscos que podiam ser enterrados na areia. Um marcador de tinta permanente coberto com uma fina camada de adesivo de endurecimento rápido provou ser um método confiável para marcar os mariscos. A manutenção dos mariscos neste sistema resultou em alta sobrevivência e crescimento, aumento do fator de condição e do diâmetro do ovócito, e um relativo avanço do desenvolvimento gonadal.

Design and optimization of an experimental maintenance system for yellow clam broodstock Amarilladesma mactroides (Reeve, 1854)

Abstract The yellow clam is a sand-burrowing bivalve that inhabits the dissipative beaches from southern Brazil to the north coast of Argentina. In the last decades, populations of this species have been impacted by mass mortality events, overfishing and other anthropogenic activities. The production of juveniles in captivity would allow feasibility studies to be carried out to restore the natural stock as well as the production in aquaculture systems. Given the scarcity of studies on the maintenance of this species in captivity, a culture system and a management protocol were developed and tested. Wild-caught clams (total length 50 mm) were used in a series of 14 day-long trials. Survival was higher in clams that were allowed to bury into the sand. A permanent ink marker covered with a thin layer of a quick-hardening adhesive proved to be a reliable method to tag clams. The maintenance of yellow clams in this system resulted in high survival and growth, increases in the condition factor and oocyte diameter, and a relative advancement of gonadal development.

Resumo O marisco branco é um bivalve de areia que habita as praias dissipativas do sul do Brasil até a costa norte da Argentina. Nas últimas décadas, as populações desta espécie têm sido afetadas por eventos de mortalidade maciça, sobrepesca e outras atividades antropogênicas. A produção de juvenis em cativeiro permitiria a realização de estudos de viabilidade para restaurar o estoque natural, assim como a produção em sistemas de aquicultura. Dada a escassez de estudos sobre a manutenção desta espécie em cativeiro, um sistema de cultivo e um protocolo de manejo foram desenvolvidos e testados. Mariscos branco selvagens (comprimento total 50 mm) foram utilizados em uma série de ensaios de 14 dias de duração. A sobrevivência foi maior nos mariscos que podiam ser enterrados na areia. Um marcador de tinta permanente coberto com uma fina camada de adesivo de endurecimento rápido provou ser um método confiável para marcar os mariscos. A manutenção dos mariscos neste sistema resultou em alta sobrevivência e crescimento, aumento do fator de condição e do diâmetro do ovócito, e um relativo avanço do desenvolvimento gonadal.

Preliminary investigation of multiple antibiotic-resistant bacteria isolated from blood cockles (Anadara granosa) and green lipped mussel (Perna viridis)

Aims@#Bivalve molluscs are filter feeders that tend to accumulate and concentrate any suspended particles or bacteria from the water environment. Although they have been proven to accumulate heavy metals, investigation on the presence of antibiotic-resistant bacteria is still lacking. Therefore, this study examines the occurrence of antibiotic-resistant bacteria isolated from bivalve shellfish, namely, blood cockles (Anadara granosa) and green-lipped mussel (Perna viridis) purchased from markets in Johor, Malaysia.@*Methodology and results@#Samples purchased were homogenized and then diluted. Viable cell count and bacterial isolation were performed using diluted samples followed by an antibiotic susceptibility test (ampicillin, ciprofloxacin and tetracycline) conducted on the pure isolates. The total viable count of bacterial colonies for cockle and mussel samples ranged from 1.1 × 107 to 4.4 × 109 CFU/mL and 1.2 × 109 to 4.3 × 109 CFU/mL. The numbers of colonies isolated from respective bivalves were 11 and 6. Generally, cockles isolates revealed higher resistance towards all three antibiotics at or above the Clinical and Laboratory Standard Institute (CLSI) threshold value. Meanwhile, mussel isolates showed full susceptibility to any ciprofloxacin concentration and tetracycline but exhibited resistance to ampicillin at a concentration exceeding the CLSI value. The number of drug resistance isolates in cockle and mussel samples decreased with increasing antibiotic concentration. The maximum number of antibiotics the mussel isolates were resistant to was two, whereas cockle isolates achieved three.@*Conclusion, significance and impact of study@#The outcome of this study concludes that some isolates from cockle and mussel samples can resist antibiotic concentration above the CLSI threshold value. Resistance of more than the CLSI threshold level revealed that these isolates could pose significant health risks especially when the bivalves are ingested raw or undercooked.

Qualidade microbiológica da água e dos mexilhões Perna perna (Linnaeus, 1758) cultivados em Piúma, Espírito Santo, Brasil / Microbiological quality of water and Perna perna (Linnaeus, 1758) mussels cultivated in Piúma, Espírito Santo, Brazil

RESUMO Este estudo objetivou verificar a qualidade microbiológica da água e dos mexilhões cultivados pela Associação dos Maricultores de Piúma (AMPI), Espírito Santo, Brasil. Foram realizadas sete coletas de água e mexilhões, mensalmente, entre outubro de 2016 e maio de 2017. Os mexilhões foram coletados nos long lines da AMPI, e em cada mês foram coletados 40 mexilhões Perna perna e 100 mL de água do local. O material coletado foi destinado ao laboratório para a realização das análises microbiológicas em duplicata, número mais provável de coliformes totais (CT) e termotolerantes (Ctt), presença ou ausência de Salmonella sp e número de unidades formadoras de colônias de Staphylococcus aureus. Os resultados mostraram que o número de Ctt nas amostras de água no mês de janeiro estavam acima do permitido pela Resolução n° 357 do Conselho Nacional do Meio Ambiente (CONAMA). Já os níveis de Ctt e Staphylococcus aureus na carne dos mexilhões mostraram-se dentro do limite aceitável pela RDC n° 12 da Agência Nacional de Vigilância Sanitária (ANVISA). Entretanto, foram encontradas bactérias com características do gênero Salmonella sp nos meses de dezembro e março nos mexilhões, impossibilitando sua comercialização e seu consumo. Durante esses meses, a cidade tem alto fluxo de turistas. Por fim, recomenda-se a realização das análises microbiológicas continuamente, principalmente no período do verão, época que tem grande fluxo de turistas no município de Piúma e que registrou presença de Salmonella na carne dos mexilhões e níveis de Ctt na água acima do permitido pelas legislações vigentes.

ABSTRACT This study aimed to verify the microbiological quality of water and mussels cultivated by the Piúma Farmers Association (Associação dos Maricultores de Piúma — AMPI), Espírito Santo, Brazil. Seven samples of water and mussels were collected monthly from October 2016 to May 2017. The mussels were collected from the long lines of the AMPI, and in each month 40 mussels Perna perna and 100 mL of water were sampled from the site. The collected material was sent to the laboratory for duplicate microbiological analysis, Most Probable Number of Total (CT) and Thermotolerant (Ctt) coliforms, presence or absence of Salmonella sp and number of Staphylococcus aureus colony forming units. The results showed that the number of Ctt in the water samples in January was higher than that allowed by Resolution 357 of the National Environment Council (Conselho Nacional do Meio Ambiente — CONAMA). The levels of Ctt and Staphylococcus aureus in mussel meat were within the acceptable range by the Brazilian National Environment Council (Agência Nacional de Vigilância Sanitária — ANVISA) Resolution RDC No. 12. However, colonies with characteristics of Salmonella sp were found in December and March in the mussels, making it impossible to sell and consume. This period coincides with a high flow of tourists in the municipality. Finally, it is recommended to perform microbiological analyzes continuously, especially in the summer, where there are a lot of tourists in the city of Piúma, period that showed the presence of Salmonella in the meat of mussels and Ctt levels in water above the allowed current legislation.

I See Golden Mussel! They are Everywhere! Environmental DNA Supports Widespread Dissemination of Limnoperna fortunei in Hydrographic Basins in the Paraná State, Brazil

Abstract The golden mussel is an aquatic invasive species that was introduced in South America in the '90s, including Brazil, and was firstly registered in 2001 in the Paraná state. This bivalve causes macrofouling issues because adhere in substrates and form dense aggregations, affecting especially hydroelectric power plants, and water treatment facilities, apart from the native biodiversity. The present research aimed to diagnose the distribution of Limnoperna fortunei in the Paraná state using environmental DNA (eDNA) from 174 sites from 12 basins, and scientific and technical previous records. L. fortunei eDNA was found in 90 sites from 11 hydrographic basins sampled. Most of the positive samples were detected in Iguaçu (42), Tibagi (15), and Ivaí (10) rivers basins. We also registered the first occurrence for five basins: Cinzas, Itararé, Ivaí, Pirapó e Ribeira rivers. Together, our and previous data recorded L. fortunei in 118 sites, between adult, larvae, and eDNA detection. Moreover, the results evidenced that eDNA is a low coast and reliable tool, and it may be very recommended for L. fortunei early detection and diagnosis. The present research was the greatest and most widespread survey for golden mussel prospection in a state in Brazil, supported a worrying scenario for Parana, with the urgent need for intense and continuous monitoring and prevention actions for controlling and mitigation of the L. fortunei impacts.

First report of detection of Toxoplasma gondii DNA in oysters (Crassostrea sp.) in the state of Maranhão / Primeiro relato da detecção de DNA de Toxoplasma gondii em ostras (Crassostrea sp.) no Estado do Maranhão

Abstract The aim of this study was to report on detection of Toxoplasma gondii DNA in oysters (Crassostrea sp.) in the state of Maranhão. To conduct this study, 200 farmed oysters were acquired in the municipality of Raposa and 100 in Paço do Lumiar; and a further 100 oysters were taken from the natural stock in the municipality of Primeira Cruz. This total of 400 specimens sampled was divided into 80 pools composed of five animals each. The gills and visceral mass of each oyster were removed for DNA extraction (per pool of oysters), using a commercial kit. The nested PCR technique (with the primer SAG-1) was then used to investigate any presence of protozoa. This molecular technique demonstrated the presence of DNA of T. gondii in 2.5% of the pools of oysters (n = 2/80): these oysters were exclusively from farms. The results from this study allow the conclusion that oysters of the genus Crassostrea that are farmed in the state of Maranhão are capable of filtering oocysts of T. gondii and maintaining them in their tissues. They are therefore potential sources of contamination for humans and other animals.

Resumo: Objetivou-se com este estudo relatar a detecção do DNA de Toxoplasma gondii em ostras (Crassostrea sp.) no estado do Maranhão. Para a realização do estudo foram adquiridas 200 ostras de cultivo do município de Raposa, e 100 de Paço do Lumiar, além de 100 ostras extraídas de estoque natural do município de Primeira Cruz. Do total de 400 exemplares amostrados, formaram-se 80 pools em que cada pool foi constituído por cinco animais. De cada ostra foi procedida à retirada das brânquias e massa visceral, seguido da extração de DNA de cada pool de ostras, com a utilização de kit comercial. Posteriormente, realizou-se a pesquisa do protozoário por meio da técnica de nested PCR (primer SAG-1). Com a técnica molecular utilizada, foi diagnosticado o DNA do protozoário pesquisado em 2,5% (n=2/80) pools de ostras oriundas exclusivamente de cultivo. Com os resultados obtidos neste estudo, conclui-se que ostras do gênero Crassostrea sp., cultivadas no estado do Maranhão, são capazes de filtrar e manter nos seus tecidos oocistos de T. gondii, sendo, portanto, fontes potenciais de contaminação para seres humanos e outros animais.

Perkinsus sp. infecting three important mollusks from Jaguaribe River estuary, Ceará, Brazil / Perkinsus sp. infectando três importantes moluscos do estuário do Rio Jaguaribe, Ceará, Brasil

This work investigated the occurrence of Perkinsus sp. in clam Anomalocardia brasiliana, oyster Crassostrea sp. and mussel Mytella falcata from the Jaguaribe River estuary, northeastern Brazil. The collection of clam (N = 300), oysters (N = 300) and mussels (N = 300) were carried out in the estuary of the Jaguaribe River, Ceará, in March and April (rainy season) and October (dry season) in 2017. The mollusks were measured in their major axis, open, and had their tissues submitted to tissue incubation techniques in Ray's fluid thioglycollate medium (RFTM), histology, real-time polymerase chain reaction (qPCR), PCR and sequencing. The RFTM assays showed Perkinsus sp. infecting the three mollusks investigated. The prevalence of infected clams was 1.33% in both sampling periods, oysters ranged from 2.66 (rainy season) to 8% (dry period), and mussels from 0% (dry period) to 51.33% (rainy season). The intensity of infection was very light to light in clams, very soft to severe in oysters and very soft to moderate in mussels. Histological analyses showed cells of Perkinsus sp. infecting the gills and connective tissue around the digestive gland of some individuals. The qPCR generated amplicons in all positive samples in RFTM, confirming the presence of Perkinsus sp., while the sequencing evidenced high similarity (99%) with the species P. beihaiensis. In conclusion, the results obtained contribute to increasing knowledge about the occurrence of Perkinsus sp. in bivalve mollusks from northeastern Brazil.(AU)

Foi investigada a ocorrência da infecção pelo protozoário Perkinsus sp. em berbigões Anomalocardia brasiliana, ostras Crassostrea sp. e mexilhões Mytella falcata do estuário do Rio Jaguaribe, Nordeste do Brasil. As colheitas dos berbigões (N = 300), ostras (N = 300) e mexilhões (N = 300) foram realizadas no estuário do Rio Jaguaribe, Ceará, nos meses de março e abril (período chuvoso) e outubro (período seco) de 2017. Os moluscos foram medidos em seu maior eixo, abertos e os seus tecidos foram submetidos às técnicas de incubação de tecidos em meio fluido de tioglicolato de Ray (RFTM), histologia, reação em cadeia da polimerase em tempo real (qPCR), PCR e sequenciamento. Os ensaios de RFTM evidenciaram Perkinsus sp. infectando os três moluscos investigados. A prevalência de berbigões infectados foi de 1,33% em ambos os períodos de amostragem, a de ostras variou de 2,66 (período chuvoso) a 8% (período seco) e a de mexilhões de 0% (período seco) a 51,33% (período chuvoso). A intensidade de infecção apresentou-se muito leve a leve em berbigões, muito leve à severa nas ostras e muito leve à moderada nos mexilhões. As análises histológicas mostraram células de Perkinsus sp. infectando as brânquias e tecido conjuntivo em torno da glândula digestiva de alguns indivíduos. A qPCR gerou amplicons em todas as amostras positivas em RFTM, confirmando a presença de Perkinsus sp., enquanto o sequenciamento mostrou alta similaridade (99%) com a espécie P. beihaiensis. Em conclusão, os resultados do presente estudo contribuem para ampliar o conhecimento sobre a ocorrência de Perkinsus sp. em moluscos bivalves do Nordeste do Brasil.(AU)

Virulence genes in Escherichia coli isolates from commercialized saltwater mussels Mytella guyanensis (Lamarck, 1819) / Presença de genes de virulência em isolados de Escherichia coli provenientes de sururu Mytella guyanensis (Lamarck, 1819) comercializado

Abstract The isolation of Escherichia coli from food is a major concern. Pathogenic strains of these bacteria cause diseases which range from diarrhea to hemolytic-uremic syndrome. Therefore the virulence genes in E. coli isolates from the mussel ( Mytella guyanensis) commercialized in Cachoeira, Bahia, Brazil were investigated. Samples were purchased from four vendors: two from supermarkets and two from fair outlets. They were conditioned into isothermal boxes with reusable ice and transported to the laboratory for analysis. E. coli strains were isolated in eosin methylene blue agar, preserved in brain-heart infusion medium with 15% glycerol and stored at -20 °C, after microbiological analysis. Virulence genes in the isolated strains were identified by specific primers, with Polymerase Chain Reaction. Twenty-four isolates were obtained, with a prevalence of elt gene, typical from enterotoxigenic infection, in 75% of the isolates. The stx and bfpA genes, prevalent in enterohemorragic and enteropathogenic E. coli, respectively, were not detected. The occurrence of elt virulence-related gene in the E. coli isolates of Mytella guyanensis reveals urgent improvement in food processing, including good handling practices, adequate storage and cooking before consumption, to ensure consumer's health.

Resumo O isolamento de Escherichia coli a partir de alimentos é uma grande preocupação, pois cepas patogênicas desta bactéria podem causar desde diarreia até síndrome hemolítico-urêmica. Diante do exposto, o objetivo do trabalho foi pesquisar genes de virulência em isolados de Escherichia coli provenientes do sururu Mytella guyanensis comercializado na cidade de Cachoeira, Bahia, Brasil. As amostras foram adquiridas de quatro comerciantes, sendo duas de mercados e duas em pontos de venda na feira livre da cidade de Cachoeira, acondicionadas em caixas isotérmicas com gelo reutilizável e transportadas até o laboratório para a análise. Após a análise microbiológica, as cepas de Escherichia coli foram isoladas em ágar Eosina Azul de Metileno e preservadas em caldo Brian Heart Infusion e glicerol a 15% e mantidas a - 20° C. A identificação dos genes de virulência nas cepas isoladas foi realizada utilizando primers específicos, por meio da Reação em Cadeia da Polimerase. Foram obtidos 24 isolados de Escherichia coli, destes a prevalência do gene elt , característico de Escherichia coli enterotoxigênica, foi de 75% dos isolados. Não houve a detecção dos genes stx e bfpA nos isolados, os quais são prevalentes nas cepas de Escherichia coli enterohemorrágica e Escherichia coli enteropatogênica, respectivamente. A presença do gene elt relacionado à virulência de Escherichia coli nos isolados de Mytella guyanensis revela a necessidade da melhoria no processamento, incluindo boas práticas de manipulação, armazenamento adequado e cocção previa ao consumo, visando a garantia da saúde do consumidor.

Bioluminescence of Vibrio fischeri: A novel application for PSP quantification

Paralytic Shellfish Poison (PSP) are the most harmful neurotoxins create a serious public health problem. It is important to assess PSP in Shellfish destined for human consumption. However, recommended methods have some limitations for example in the case of Mouse Bioassay (MBA) showed a low sensitivity and reproducibility, and undesirability for ethical reasons; while physico-chemical techniques rest expensive and time-consuming. The main objective of this study, after discovered that PSP inhibited the luminescence of Vibrio fischeri, was the quantification of PSP by using Bioluminescence Inhibition Assay (BIA), and comparing the results obtained with those determined by MBA and LC-MS. Bivalve used were collected from Corniche Martil, Kabila and Oued Laou, along the Mediterranean coast of Morocco in Mars-2015. Results showed a weak correlation between LC-MS and MBA with r = 0.11, while, the correlation between LC-MS and BIA was very strong with r = 0.97, which suggests that, BIA could offer an interesting additional assessment of PSP risk. In addition, after seen its rapidity, ease, reliability, sensitivity, reproducibility and cost effectiveness, it would be eligible to use for monitoring in surveillance programs.

Reproductive cycle of Mytella guyanensis (Lamarck, 1819) in a Marine Reserve (RESEX Bay of Iguape), Bahia, Brazil / Ciclo reprodutivo de Mytella guyanensis (Lamarck, 1819) na Reserva Extrativista Marinha Baía do Iguape, Bahia, Brasil

Abstract Mytella guyanensis, consumed and commercialized in coastal regions of Brazil, is one of several bivalve species of socioeconomic interest for coastal communities. Besides serving as a source of income and subsistence for these communities, it also contributes to their food security as it is a source of proteins and micronutrients. Thus, the reproductive cycle of this species was studied aiming to contribute to food security and its preservation. Samples were collected monthly, between March 2014 and March 2015, in a natural stock (12°38'50"S; 38°51'43"W) in a Marine Reserve (RESEX Bay of Iguape) (community Engenho da Ponte), Bahia, Brazil. Mytella guyanensis is collected by women on site, where the artisanal fishing of this resource is performed without following any specific handling procedure. Also, empirical evidence indicates overexploitation. The specimens collected were measured along the anterior-posterior axis (length), and after macroscopic analysis they were fixed in Davidson solution, processed by routine histology techniques and stained with Harris haematoxylin and eosin (H&E). The macroscopic analysis showed sexual dimorphism, with the male and female gonads presenting a milky-white and orange colour, respectively. A 1:1 sex ratio (M: F) was observed and reproduction of the species was continuous all year round. March, April, July and August were the months with highest values of gamete elimination. We suggest that a M. guyanensis management plan should restrict capture during these months, in order to sustainably regulate exploitation of this food resource in this reserve.

Resumo Mytella guyanensis, consumida e comercializada em regiões litorâneas do Brasil, é uma das diversas espécies de bivalves de interesse socioeconômico para comunidades litorâneas. Além de servir como fonte de renda e subsistência para essas comunidades, esta contribui para a sua segurança alimentar, por ser fonte de proteínas e micronutrientes. Assim, o ciclo reprodutivo desta espécie foi estudado visando contribuir com a segurança alimentar e a preservação da mesma. As amostragens foram realizadas mensalmente, entre março de 2014 e março de 2015 em um estoque natural (12°38'50"S e 38°51'43"W) na Reserva Extrativista Marinha Baía do Iguape (comunidade Engenho da Ponte), Bahia. Mytella guyanensis é coletada no local por mulheres, onde a pesca artesanal desse recurso é realizada sem seguir nenhum procedimento específico de manejo e evidência empírica indica sobrexplotação. Os espécimes coletados foram medidos ao longo do eixo ântero-posterior (comprimento) e após a análise macroscópica, foram fixados em solução de Davidson, processados por técnicas rotineiras de histologia e coradas em hematoxilina de Harris e eosina (HE). A análise macroscópica evidenciou dimorfismo sexual, com as gônadas de machos e fêmeas apresentando coloração branco leitosa e alaranjada, respectivamente. Uma proporção sexual (M: F) de 1:1 foi observada e a reprodução foi contínua ao longo do ano. Março, abril, julho e agosto apresentaram os maiores valores de eliminação de gametas. Sugerimos que um plano de manejo de M. guyanensis restrinja a captura deste durante esses meses, a fim de regular de forma sustentável a exploração desse recurso nesta reserva.

Molecular cloning and characterisation of scavenger receptor class B in pearl oyster Pinctada fuctada martensii

Background: Molluscs can accumulate carotenoids in their body tissues by predominantly feeding on aquatic plant sources. Carotenoid transport and absorption are determined by the regulation of various proteins such as Scavenger receptor class B(SR-BI). We report the identification and characterisation of pearl oyster Pinctada fuctada martensii SR-BI (PmSR-BI). The correlation between total carotenoid content (TCC) and gene expression was also estimated. Results: The full-length cDNA of PmSR-BI was 1828 bp, including an open-reading frame encoding of 1518 bp with a pI value of 5.83. PmSR-BI protein contains a hydrophobic CD36 domain and four centrally clustered cysteine residues for the arrangement of disulphide bridges. The deduced amino acid sequence had an identity of 30% to 60% with the SR-B of other organisms. Reverse transcription polymerase chain reaction analysis showed that mRNA transcripts were expressed in multiple tissues of adult pearl oyster. A higher expression of PmSR-BI gene was observed in the hepatopancreas than in the adductor muscle, gill and mantle. The TCC and gene expression of PmSR-BI were significantly correlated (P b 0.05), with a correlation coefficient of 0.978. Conclusions: The results suggested that PmSR-BI is involved in the absorption of carotenoids in the pearl oyster P. fuctada martensii.

Asociación de floraciones de algas nocivas y Vibrio spp. en áreas de pesca y acuicultura de bivalvos de moluscos en las bahías de Sechura y Pisco, Perú

Entre febrero de 2010 y mayo de 2014 se realizaron 22 evaluaciones en Pisco y 16 en Sechura; ambas áreas importantes en Perú para la extracción de moluscos y maricultura. La incidencia de Vibrio en el agua de mar fue monitoreada durante las floraciones de algas y en épocas normales. Se midieron parámetros ambientales tales como temperatura y nutrientes. En Sechura, Pseudo-nitzschia seriata y Protoperidinium depressum causaron floraciones de algas y fueron dominantes durante todo el período de evaluación. Las temperaturas en esta zona oscilaron entre 21.8 y 25.3 °C. En Pisco, la concentración Akashiwo sanguinea, Messodinium rubrum y Prorocentrum mínimum y el dinoflagelado Cochlodinium polikrykoides fueron los más frecuentes. Las floraciones de algas nocivas ocurrieron cuando las temperaturas estaban entre 17.1 y 23.3 °C, concentraciones de fosfatos que oscilaban entre 1.22 - 6.85 μM y nitratos 0.15 - 7.85 μM. En mayo de 2012, el dinoflagelado Alexandrium peruvianum causó una floración algal, bajo temperaturas de 18.0 a 23.2 °C, valores de fosfato de 0.73 a 11.56 μM y nitratos de 0.76 a 9.81 μM. Los coliformes fueron bajos < 2 - 23 MPN/100 mL, en ambas bahías durante todo el período de estudio. Vibrio alginolyticus fue la especie dominante. Mientras que V. vulnificus y V. parahaemolyticus se detectaron en Pisco, donde las temperaturas del mar comúnmente son mayores y los casos de infecciones graves por ingestión de mariscos se han asociado con el patógeno V. parahaemolyticus.

Between February 2010 and May 2014, 22 surveys in Pisco and 16 in Sechura were conducted; both are major areas for shellfish production and mariculture in Peru. The incidence of Vibrio in seawater was monitored during algal blooms and in their absence. Environmental parameters such as temperature and nutrients were measured. In Sechura, Pseudo-nitzschia seriata and Protoperidinium depressum caused algal blooms and were dominant throughout the evaluation period. The temperatures in this area ranged from 21.8 to 25.3 °C. In Pisco, the harmful algal bloom-forming Akashiwo sanguinea, Messodinium rubrum, and Prorocentrum minimum and the dinoflagellate Cochlodinium polikrykoides were most prevalent. Harmful algal blooms occurred when temperatures were between 17.1 and 23.3 °C, with phosphates ranging 1.22 - 6.85 μM and nitrates 0.15 - 7.85 μM. In May 2012, the dinoflagellate Alexandrium peruvianum caused an algal bloom, with temperatures ranging 18.0 to 23.2 °C, phosphate values from 0.73 to 11.56 μM, and nitrates from 0.76 to 9.81 μM. Coliforms were low, < 2 - 23 MPN/100 ml, in both bays throughout the study period. Vibrio alginolyticus was the dominant Vibrio spp. predominated in both bays, while V. vulnificus and V. parahaemolyticus were detected in Pisco, where warmer sea temperatures are common and severe infections cases by seafood ingestion has been associated with a pathogen V. parahaemolyticus.

Freshwater mussels from South America: state of the art of Unionida, specially Rhipidodontini / Bivalves de água doce da América do Sul: estado da arte de Unionida, especialmente Rhipidodontini

Abstract Unionida is the most diverse clade of freshwater bivalves. Among the groups occurring in South America, one with the highest number of species is Rhipidodontini (Hyriidae, Unionida, Paleoheterodonta, Bivalvia). However several issues remains on taxonomy and systematic of this group, leading to problems on species identification, description, as also as a limiting factor to other type of studies (e.g., ecology, conservation,…). In this paper is presented a synthesis of available knowledge about Diplodon Spix in Wagner, 1827 and Rhipidodonta Mörch, 1853 in South America, as a first step in order to a better understating of Rhipidodontini. The evaluation of different authors exposes the little agreement between them that resulted in a sort of divergent taxonomical opinions. Some comments on ecology, conservation and habitat preferences were made. This work can also encourage future research on taxonomy, systematic, ecology and conservation of freshwater mussels in South America.

Resumo Unionida é o clado mais diverso de bivalves de água doce. Entre os grupos que ocorrem na América do Sul, um dos com maior número de espécies é Rhipidodontini (Hyriidae, Unionida, Paleoheterodonta, Bivalvia). Porém, diversas questões taxonômicas e sistemáticas ainda incidem sob este grupo, levando a problemas de identificação de espécies, descrição, entre outros, como também tem atuado como limitador de outros tipos de estudos (e.g., ecologia, conservação,..). Neste trabalho é apresentada uma revisão do conhecimento acerca dos gêneros Diplodon Spix in Wagner, 1827 e Rhipidodonta Mörch, 1853 na América do Sul como um primeiro passo para a melhor compreensão de Rhipidodontini. Avaliando-se diferentes autores, se torna claro a pouca concordância entre eles, resultando em opiniões taxonômicas divergentes. São feitos também alguns comentários sobre ecologia, conservação e preferências ambientais. Este trabalho também deve encorajar futuros trabalhos sobre a taxonomia, sistemática, ecologia e conservação de bivalves de água na América do Sul.

Enterococcus spp. Resistant to Multiple Antimicrobial Drugs and Determination of Fecal Contamination Levels in Mangrove Oysters (Crassostrea rhizophorae)

ABSTRACT The aim of this study was to determine and compare the Most Probable Number (MPN) of Total Coliforms (TC), Escherichia coli and Enterococcus spp. and to characterize the antimicrobial resistance profiles of Enterococcus spp. isolated from oysters collected in the Barra de Guaratiba Mangrove, Rio de Janeiro, Brazil. The enumeration of E. coli has been used to indicate fecal contamination and hygienic-sanitary conditions of bivalve molluscs. Enterococci are capable to transfer several antimicrobial resistance genes to pathogenic bacteria, including those from Gram-negative group. The oysters were bought from local fishermen and a total of 123 individuals were analyzed. The TC, E. coli and Enterococcus spp. MPN mean were 26,300/100 g, 3,260/100 g and 2,820/100 g, respectively. The only correlation found was between TC and E. coli. Two strains of Enterococcus spp. were resistant to three different antimicrobial categories, including a high level resistance to streptomycin. One strain presented intermediate resistance to vancomycin. The E. coli levels exceeded the limits established by international legislation. This microbiological contamination in oysters reflects the water pollution and indicates a probable contamination of other seafood species from this mangrove, which can represent a risk for consumers and a threat to the environment and public health.

Reproductive cycle of Mytella guyanensis (Lamarck, 1819) in a Marine Reserve (RESEX Bay of Iguape), Bahia, Brazil

Abstract Mytella guyanensis, consumed and commercialized in coastal regions of Brazil, is one of several bivalve species of socioeconomic interest for coastal communities. Besides serving as a source of income and subsistence for these communities, it also contributes to their food security as it is a source of proteins and micronutrients. Thus, the reproductive cycle of this species was studied aiming to contribute to food security and its preservation. Samples were collected monthly, between March 2014 and March 2015, in a natural stock (12°38'50S; 38°51'43W) in a Marine Reserve (RESEX Bay of Iguape) (community Engenho da Ponte), Bahia, Brazil. Mytella guyanensis is collected by women on site, where the artisanal fishing of this resource is performed without following any specific handling procedure. Also, empirical evidence indicates overexploitation. The specimens collected were measured along the anterior-posterior axis (length), and after macroscopic analysis they were fixed in Davidson solution, processed by routine histology techniques and stained with Harris haematoxylin and eosin (H&E). The macroscopic analysis showed sexual dimorphism, with the male and female gonads presenting a milky-white and orange colour, respectively. A 1:1 sex ratio (M: F) was observed and reproduction of the species was continuous all year round. March, April, July and August were the months with highest values of gamete elimination. We suggest that a M. guyanensis management plan should restrict capture during these months, in order to sustainably regulate exploitation of this food resource in this reserve.

Resumo Mytella guyanensis, consumida e comercializada em regiões litorâneas do Brasil, é uma das diversas espécies de bivalves de interesse socioeconômico para comunidades litorâneas. Além de servir como fonte de renda e subsistência para essas comunidades, esta contribui para a sua segurança alimentar, por ser fonte de proteínas e micronutrientes. Assim, o ciclo reprodutivo desta espécie foi estudado visando contribuir com a segurança alimentar e a preservação da mesma. As amostragens foram realizadas mensalmente, entre março de 2014 e março de 2015 em um estoque natural (12°38'50S e 38°51'43W) na Reserva Extrativista Marinha Baía do Iguape (comunidade Engenho da Ponte), Bahia. Mytella guyanensis é coletada no local por mulheres, onde a pesca artesanal desse recurso é realizada sem seguir nenhum procedimento específico de manejo e evidência empírica indica sobrexplotação. Os espécimes coletados foram medidos ao longo do eixo ântero-posterior (comprimento) e após a análise macroscópica, foram fixados em solução de Davidson, processados por técnicas rotineiras de histologia e coradas em hematoxilina de Harris e eosina (HE). A análise macroscópica evidenciou dimorfismo sexual, com as gônadas de machos e fêmeas apresentando coloração branco leitosa e alaranjada, respectivamente. Uma proporção sexual (M: F) de 1:1 foi observada e a reprodução foi contínua ao longo do ano. Março, abril, julho e agosto apresentaram os maiores valores de eliminação de gametas. Sugerimos que um plano de manejo de M. guyanensis restrinja a captura deste durante esses meses, a fim de regular de forma sustentável a exploração desse recurso nesta reserva.

Virulence genes in Escherichia coli isolates from commercialized saltwater mussels Mytella guyanensis (Lamarck, 1819)

Abstract The isolation of Escherichia coli from food is a major concern. Pathogenic strains of these bacteria cause diseases which range from diarrhea to hemolytic-uremic syndrome. Therefore the virulence genes in E. coli isolates from the mussel ( Mytella guyanensis) commercialized in Cachoeira, Bahia, Brazil were investigated. Samples were purchased from four vendors: two from supermarkets and two from fair outlets. They were conditioned into isothermal boxes with reusable ice and transported to the laboratory for analysis. E. coli strains were isolated in eosin methylene blue agar, preserved in brain-heart infusion medium with 15% glycerol and stored at -20 °C, after microbiological analysis. Virulence genes in the isolated strains were identified by specific primers, with Polymerase Chain Reaction. Twenty-four isolates were obtained, with a prevalence of elt gene, typical from enterotoxigenic infection, in 75% of the isolates. The stx and bfpA genes, prevalent in enterohemorragic and enteropathogenic E. coli, respectively, were not detected. The occurrence of elt virulence-related gene in the E. coli isolates of Mytella guyanensis reveals urgent improvement in food processing, including good handling practices, adequate storage and cooking before consumption, to ensure consumers health.

Resumo O isolamento de Escherichia coli a partir de alimentos é uma grande preocupação, pois cepas patogênicas desta bactéria podem causar desde diarreia até síndrome hemolítico-urêmica. Diante do exposto, o objetivo do trabalho foi pesquisar genes de virulência em isolados de Escherichia coli provenientes do sururu Mytella guyanensis comercializado na cidade de Cachoeira, Bahia, Brasil. As amostras foram adquiridas de quatro comerciantes, sendo duas de mercados e duas em pontos de venda na feira livre da cidade de Cachoeira, acondicionadas em caixas isotérmicas com gelo reutilizável e transportadas até o laboratório para a análise. Após a análise microbiológica, as cepas de Escherichia coli foram isoladas em ágar Eosina Azul de Metileno e preservadas em caldo Brian Heart Infusion e glicerol a 15% e mantidas a - 20° C. A identificação dos genes de virulência nas cepas isoladas foi realizada utilizando primers específicos, por meio da Reação em Cadeia da Polimerase. Foram obtidos 24 isolados de Escherichia coli, destes a prevalência do gene elt , característico de Escherichia coli enterotoxigênica, foi de 75% dos isolados. Não houve a detecção dos genes stx e bfpA nos isolados, os quais são prevalentes nas cepas de Escherichia coli enterohemorrágica e Escherichia coli enteropatogênica, respectivamente. A presença do gene elt relacionado à virulência de Escherichia coli nos isolados de Mytella guyanensis revela a necessidade da melhoria no processamento, incluindo boas práticas de manipulação, armazenamento adequado e cocção previa ao consumo, visando a garantia da saúde do consumidor.

Perfil socioeconômico dos marisqueiros e condições higiênicas adotadas na cadeia produtiva de ostra (mollusca, bivalvia) / Socio economic profile of shellfish collector and hygienic conditions in the production chain of oysters (mollusca, bivalvia) / Perfil socioeconómico de marisqueros y condiciones higiénicas adoptadas en la cadena productiva de ostra (mollusca, bivalvia)

O consumo de ostras na forma in natura exige muito cuidado e práticas higiênicas e sanitárias corretas pelos marisqueiros e demais manipuladores desse alimento durante o processo de beneficiamento à comercialização, para que não haja contaminação por microrganismos patogênicos e deteriorantes. O presente estudo teve como objetivo avaliar o perfil socioeconômico de marisqueiros e as condições higiênicas e sanitárias praticadas na cadeia produtiva de ostras do gênero Crassostrea desde a extração até a comercialização. No período de janeiro a abril de 2014, por meio da técnica de amostragem "Bola de Neve" foram selecionados 40 marisqueiros na Ilha de São Luís - MA, os quais foram entrevistados utilizando-se questionário semiestruturado. De acordo com os resultados, a maioria dos marisqueiros, possui ensino fundamental incompleto, faixa etária entre 21 e 30 anos e encontram-se em união estável ou casados. Parte das ostras são extraídas em bancos naturais na área de estudo, enquanto outra parte é proveniente de atravessadores oriundos de outros municípios maranhenses.Após a aquisição, pode demorar até três dias para serem comercializadas. Concluiu-se que as condições higiênicos sanitárias adotadas na atividade de extração de ostras até a comercialização são insatisfatórias, indicando falta de boas práticas em todas as etapas ao longo do processo de produção, o que pode representar riscos à saúde da população apreciadora do consumo in natura desse bivalve. Além disso, indica a necessidade de fiscalização dessa atividade, já que os organismos são transportados de forma inadequada e passam muito tempo até chegar ao consumidor final.

Consumption of fresh oysters requires very careful and correct hygienic and sanitary practices by shellfish collectors and other food handlers during the handling process to marketing, in order to avoid contamination by pathogenic microorganisms or deterioration. This study aimed to evaluate the socio-economic profile of shellfish collectors and the hygienic conditions applied in the production chain of Crassostrea oysters from extraction to marketing. In the period from January to April 2014, through a snowball sampling technique, a total of 40 shellfish collector were selected in Ilha de São Luís ­ MA. These collectors were interviewed using a semi-structured questionnaire. According to the results, most shellfish collector had not completed elementary school, aged between 21 and 30 years and married or co-habiting. Part of the oysters is extracted from natural stocks in the study area, while other oysters come from middlemen who bring them from other cities in Maranhão. After acquisition, they can take up to three days to be marketed. Therefore, it was concluded that although it is an important source of income, hygienic and sanitary conditions adopted in the oyster extraction activity are unsatisfactory, indicating a lack of good practices throughout the production process, which can pose risks to the population who consume this bivalve fresh. It also shows the need for supervision of the activity, since the organisms are improperly transported, spending much time until reaching the end consumer.

El consumo de ostras en la forma fresca, in natura, requiere gran cuidado y prácticas higiénicas sanitarias correctas por los marisqueros y otros manipuladores de ese alimento durante el proceso de beneficiación a la comercialización, para que no haya contaminación por microorganismos patógenos y de deterioración. Este estudio tuvo como objetivo evaluar el perfil socioeconómico de marisqueros y las condiciones higiénicas sanitarias practicadas en la cadena productiva de ostras del género Crassostrea, desde la extracción hasta la comercialización. En el período de enero a abril de 2014, a través de la técnica de muestreo "Bola de Nieve" se seleccionaron 40 marisqueros en la Isla de São Luís - MA, que fueron entrevistados mediante cuestionario semiestructurado. De acuerdo con los resultados, la mayoría de los marisqueros tiene enseñanza primaria incompleta, con edades comprendidas entre 21 y 30 años y se encuentran en unión estable o casados. Parte de las ostras son extraídas en bancos naturales en el área del estudio, mientras otra parte proviene de los intermediarios de otros municipios de Maranhão. Después de la adquisición, puede tardar hasta tres días para que sean comercializadas. Se concluye que las condiciones higiénicas sanitarias adoptadas en la actividad de extracción de ostras hasta la comercialización son insatisfactorias, indicando falta de buenas prácticas en todo el proceso de producción, lo que puede plantear riesgos a la salud de la población aficionada al consumo in natura de ese bivalvo. Además, indica la necesidad de supervisar esta actividad, ya que los organismos son transportados inadecuadamente y pasan mucho tiempo para llegar al consumidor final.

Study on Sterility Test after Using Non-PVC Bivalve Soft-bag Injection in PIVAS / 中国药房

OBJECTIVE:To study sterility test after using Non-PVC bivalve soft-bag injection in PIVAS. METHODS:The test was divided into 3 groups according to the type of transfusion solution packaging and dispensing environment. Group 1 received Glucose solution using bivalve soft-bag,dispensed in PIVAS;group 2 received Glucose solution using bivalve soft-bag,dispensed in wards area;group 3 received Glucose solution using plastic bottle,dispensed in wards area. After puncturing 1,3,6,9 times (n=80),finished products placed in ward for 0,2,4,6 h(n=20),and then sterility test was conducted with membrane filtra-tion method stated in second part of Chinese Pharmacopoeia (2010 edition). Infusion contamination of 3 groups was analyzed at 9th puncture. RESULTS:The growth of bacteria was not found in group 1;the positive detection rate of group 2 and 3 were 2.5%and 3.8%(n=320). The total positive detection rates after puncturing 1,3,6,9 times were 0,0.4%,0.4%,7.5%(n=240);the positive detection rates of group 1 were all 0,those of group 2 were 0,1.25%,0,8.75%and those of group 3 were 0,0,1.25%, 13.75%(n=80). After 9 times of puncture,the positive detection rates of group 1 after placing 0,2,4,6 h were all 0,those of group 2 were 25%,5%,0,5%;those of group 3 were 5%,15%,5%,30%(n=20). CONCLUSIONS:The use of the Non-PVC bi-valve soft-bag injection in PIVAS can effectively prevent microbial contamination.

Impact of marine pollution in green mussel Perna viridis from four coastal sites in Karachi, Pakistan, North Arabian Sea: Histopathological observations.

Pathological changes are regarded as a standard technique to monitor the effects of pollutants in marine animals. Histopathological examination of the population of green mussel Perna viridis (L.) from four sites in Pakistan, namely, Manora Channel, Rehri Creek, Sandspit Backwaters and Bhanbore was conducted. The first three sites are on the Karachi coast, whereas the fourth one, Bhanbore is situated outside Karachi, and is considered to be less polluted. Two types of parasites, Rickettsia-like organisms and metacestode were found in the mussels studied. In the present study, we observed various pathological lesions, such as inflammatory responses, granulocytomas, lipofuscin pigments, vacuolation in the digestive gland and gonads, lamellar fusion and dilated hemolymphatic sinus in the gills of P. viridis. These observations indicate the extent of environmental pollution in the studied areas. Although, Bhanbore is considered to be relatively less polluted compared to other three sites, the present results have revealed that the waters of Bhanbore are also polluted as evidenced by the pathological changes observed in the mussels collected from there.