Isolamento de uma quitinase extraída do cefalotórax do camarão marinho Litopenaeus schmitti (BURKENROAD, 1936) : avaliação de suas atividades antimicrobiana e larvicida

Chitinases are enzymes involved in degradation of chitin and are present in a range of organisms, including those that do not contain chitin, such as bacteria, viruses, plants and animals, and play important physiological and ecological roles. Chitin is hydrolyzed by a chitinolytic system classified as: endo-chitinases, exo-chitinases and N-acetyl-b-D-glucosaminidases. In this study a Litochitinase1 extracted from the cephalotorax of the shrimp Litopenaeus Schmitt was purified 987.32 times using ionexchange chromatography DEAE-Biogel and molecular exclusion Sephacryl S-200. These enzyme presented a molecular mass of about 28.5 kDa. The results, after kinetic assay with the Litochitinase1 using as substrate p-nitrophenyl-N-acetyl-b-Dglucosaminideo, showed apparent Km of 0.51 mM, optimal activity at pH ranging from 5.0 to 6.0, optimum temperature at 55°C and stability when pre-incubated at temperatures of 25, 37, 45, 50 and 55°C. The enzyme showed a range of stability at pH 4.0 to 5.5. HgCl2 inhibited Litochitinase1 while MgCl2 enhances its activity. Antimicrobial tests showed that Litochitinase1 present activity against gram-negative bacterium Escherichia coli in the 800 μg/mL concentration. The larvicidal activity against Aedes aegypti was investigated using crude extracts, F-III (50-80%) and Litochitinase1 at 24 and 48 hours. The results showed larvicidal activity in all these samples with EC50 values of 6.59 mg/mL for crude extract, 5.36 mg/mL for F-III and 0.71 mg/mL for Litochitinase1 at 24 hours and 3.22 and 0.49 mg/mL for the F-III and Litochitinase1 at 48 hours, respectively. Other experiments confirmed the presence of chitin in the midgut of Aedes aegypti larvae, which may be suffering the action of Litochitinase1 killing the larvae, but also the absence of contaminating proteins as serine proteinase inhibitors and lectins in the crude extract, F-III and Litochitinase1, indicating that the death of the larvae is by action of the Litochitinase1. We also observed that the enzymes extracted from intestinal homogenate of the larvae no have activity on Litochitinase1. These results indicate that the enzyme can be used as an alternative to control of infections caused by Escherichia coli and reducing the infestation of the mosquito vector of dengue.

Ecological distribution of the shrimp Litopenaeus schmitti (Burkenroad, 1936) (Decapoda, Penaeoidea) in Ubatuba Bay, São Paulo, Brazil

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Distribuição diária das atividades comportamentais dos camarões marinhos Farfantepenaeus subtilis (Pérez Farfante, 1967) e Litopenaeus schmitti (Burkenroad, 1938) em condições laboratoriais

Marine aquaculture has developed in the last decades all over the world, especially related to shrimp management. In Brazil, the introduction of the species Litopenaeus vannamei has contributed to the success of the activity, even if there are problems associated with the management of a exotic species, such as new diseases and ecological pressure on native species. It has been emphasized the need of research for developing new methodology that will allow native species management, being the most important Farfantepenaeus subtilis and Litopenaeus schmitti. Most knowledge obtained from research with those species has generally used a technical approach and mainly focused feeding process. There are no specific behavioral data on their activity pattern which should be of great importance for the use of native species on commercial culture farms. So, it was our objective to study and compare the daily distribution of behavioral activities of the marine shrimp species Litopenaeus schmitti and Farfantepenaeus subtilis. Forty animals of each species, 5 individuals per aquarium, were maintained in aquaria containing 200L of sea water under continuous aeration and filtration. They were marked individually and were observed by the instantaneous focal time sampling, along 10 continuous days, in 6 daily 15 min observation windows, every two hour. In each window, behaviors and location position of the animals in the aquarium were registered at 1 min intervals. Food was offered 3 times a day, representing 10% of each aquarium biomass. Aquaria were maintained in artificial photoperiod, 12hour light/l2 hour dark, 4 aquaria in light cycle equivalent to the environmental one (light from 06:00 to 17:59 h and dark from 18:00 to 05:59 h) and the other 4 in the reverse light cycle (light from 18:00 to 05:59 h and dark from 06:00 to 17:59 h) to allow sequential behavioral observation in both phases of the 24 hour cycle. There was a clear distinction between the distribution of behavioral activities of F. subtilis and L. schmitti in the two phases. The activity pattern of Farfantepenaeus subtilis demonstrates that species has prominently night habits and a burying pattern during the light cycle. Exploration, inactivity and swimming were the most common activities. The behavioral pattern of Litopenaeus schmitti indicates that species is active along both phases of the light cycle, and the most evident behaviors were exploration, inactivity and swimming

Distribuição ecológica e estrutura populacional em escala espacial, temporal e anual do camarão-branco Litopenaeus schmitti (Burkenroad, 1936) (Dendrobranchiata: Penaeidae) na enseada de Ubatuba: 4 anos de estudo

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Distribuição e dinâmica populacional dos camarões-rosa, Farfantepenaeus brasiliensis (Latreille, 1817) e F. paulensis (Pérez-Farfante, 1967) e do camarão-branco Litopenaeus schmitti (Burkenroad, 1936) (Decapoda: Dendrobranchiata: Penaeidae) no complexo baía-estuário de Santos-São Vicente, São Paulo, Brasil: subsídios científicos para a averiguação do período ideal de defeso

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Temporal and spatial distribution of the commercial shrimp Litopenaeus schmitti (Dendrobranchiata: Penaeidae) in the south-eastern Brazilian coast

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Ecological distribution of the shrimp Litopenaeus schmitti (Burkenroad, 1936) (Decapoda, Penaeoidea) in Ubatuba Bay, Sao Paulo, Brazil

The penaeidean Litopenaeus schmitti, popularly known as white shrimp, is a species of great economic importance, being a target of fishing fleets in the southeast region of Brazil. It is distributed through the western Atlantic, from Cuba to Brazil, until Rio Grande do Sul. Adults are found from shallow depths up to 30 m and have been found to depths of 47 m in the state of Rio de Janeiro, while juveniles are located in bays and estuaries. The studied species is seasonally distributed in the region of Ubatuba. The objectives of this study are to analyze the abundance and ecological distribution of L. schmitti and to assess if and when juveniles use the Indaia estuary during their life cycle. Furthermore, the hypothesis was tested that the main period of recruitment in the bay coincides with the period of closure of fisheries defined by the Instruction of IBAMA. To that end, samples were taken monthly from July 2005 through June 2007, both in Ubatuba Bay and in the estuary formed by the Rio Indaia. At each sampling station, salinity, temperature (bottom), depth, organic matter content (%), and grain size of sediments were measured. We found that the largest catches in the estuary were in late spring and early summer. In Ubatuba Bay, peak catches occurred during winter and early spring, whereas in the second year, already in May, there was a high peak capture. The variation in the number of individuals was correlated with some environmental factors both in the estuary and in the inlet (p < 0.05). In the estuary, abundance was positively correlated with temperature (p = 0.008) and organic matter (p = 0.025) and negatively with depth (p = 0.009). Regarding the Ubatuba Bay, only temperature (p = 0.034) and depth (p = 0.013) were significantly associated with the distribution of the shrimp, both being negative relations. The shrimp L. schmitti uses both the estuarine as well the shore environment, particularly the Ubatuba estuary and its adjacent bay, to complete its life cycle. The proposed period of fisheries closure (between March to May in the state of Sao Paulo) for this and other shrimps coincides with individuals capable of reproduction entering the inlet and thus are being protected.

Isolamento de uma quitinase extraída do cefalotórax do camarão marinho Litopenaeus schmitti (BURKENROAD, 1936) : avaliação de suas atividades antimicrobiana e larvicida

Chitinases are enzymes involved in degradation of chitin and are present in a range of organisms, including those that do not contain chitin, such as bacteria, viruses, plants and animals, and play important physiological and ecological roles. Chitin is hydrolyzed by a chitinolytic system classified as: endo-chitinases, exo-chitinases and N-acetyl-b-D-glucosaminidases. In this study a Litochitinase1 extracted from the cephalotorax of the shrimp Litopenaeus Schmitt was purified 987.32 times using ionexchange chromatography DEAE-Biogel and molecular exclusion Sephacryl S-200. These enzyme presented a molecular mass of about 28.5 kDa. The results, after kinetic assay with the Litochitinase1 using as substrate p-nitrophenyl-N-acetyl-b-Dglucosaminideo, showed apparent Km of 0.51 mM, optimal activity at pH ranging from 5.0 to 6.0, optimum temperature at 55°C and stability when pre-incubated at temperatures of 25, 37, 45, 50 and 55°C. The enzyme showed a range of stability at pH 4.0 to 5.5. HgCl2 inhibited Litochitinase1 while MgCl2 enhances its activity. Antimicrobial tests showed that Litochitinase1 present activity against gram-negative bacterium Escherichia coli in the 800 μg/mL concentration. The larvicidal activity against Aedes aegypti was investigated using crude extracts, F-III (50-80%) and Litochitinase1 at 24 and 48 hours. The results showed larvicidal activity in all these samples with EC50 values of 6.59 mg/mL for crude extract, 5.36 mg/mL for F-III and 0.71 mg/mL for Litochitinase1 at 24 hours and 3.22 and 0.49 mg/mL for the F-III and Litochitinase1 at 48 hours, respectively. Other experiments confirmed the presence of chitin in the midgut of Aedes aegypti larvae, which may be suffering the action of Litochitinase1 killing the larvae, but also the absence of contaminating proteins as serine proteinase inhibitors and lectins in the crude extract, F-III and Litochitinase1, indicating that the death of the larvae is by action of the Litochitinase1. We also observed that the enzymes extracted from intestinal homogenate of the larvae no have activity on Litochitinase1. These results indicate that the enzyme can be used as an alternative to control of infections caused by Escherichia coli and reducing the infestation of the mosquito vector of dengue.

Abundância e distribuição sazonal de espécies autóctones de camarões peneídeos no canal de abastecimento do sistema primar de aquicultura orgânica (Laguna de Guaraíras, Tibau do Sul, Rio Grande do Norte, Brasil)

Guaraíras lagoon, located in Tibau do Sul in the eastern littoral of Rio Grande do Norte (Brazil), presents a permanent connection to the sea, which guarantees the occurrence of a rich biodiversity, which includes the autochthonous shrimp species Litopenaeus schmitti, Farfantepenaeus subtilis and Farfantepenaeus brasiliensis. In spite of being subject to a strong human intervention in the last decade, mainly related to the installation of shrimp (Litopenaeus vannamei) farms, the lagoon is still scarcely studied. The present study aims at characterizing the populations of the three autochthonous penaeid shrimp species inhabiting Guaraíras, taking into consideration their abundance and seasonal distribution in the inflow channel of Primar System of Organic Aquaculture (Tibau do Sul, Rio Grande do Norte, Brazil). Twelve monthly samples were carried out from May 2005 to April 2006 with the aid of a circular cast net in the inflow channel, which is daily supplied with water from Guaraíras. Sampling months were grouped in trimesters according to the total pluviosity, thus comprising four trimesters. Water salinity was monitored twice a week and temperature values registered on a daily basis at noon, during the study period. The daily pluviosity data from the municipality of Tibau do Sul were supplied by Empresa de Pesquisa Agropecuária do Rio Grande do Norte (EMPARN). Collected shrimp were identified, weighted, measured and sexed. L. schmitti specimens (0.2 g to 17.8 g) were distributed in 1.3 g weight classes intervals. From the eighth sampling month (December 2005) onwards, males were classified into three categories, in accordance with the development of their petasm: (a) rudimentary petasm, (b) partially formed petasm, and (c) completely formed petasm. Among the ecological variables, rainfall showed the greatest dispersion (s.d.=187.74Rainfall and abundance of L. schmitti were negatively correlated (r = -0.85) whereas its abundance and water salinity were positively correlated (r = 0.63). Among 1,144 collected individuals, 1,127 were L. schmitti, 13 were F. subtilis and 4 were F. brasiliensis, which corresponded to 98.51%, 1.14% and 0.35% of the total of collected individuals. L. schmitti occurred in 100 % of all samples. Differently, the presence of F. subtilis and F. brasiliensis was restricted to 33% and 17% of the collected samples, respectively. The present study confirmed the occurrence of L. schmitti, F. brasiliensis and F. subtilis in Guaraíras. However, this lagoon seems to be primarily inhabited by juvenile Litopenaeus schmitti. The population of L. schmitti analysed showed a seasonal pattern of distribution. In general, in the months of high salinity and absence of rain, the number of individuals was higher than in the wet months. Further studies on the reproductive biology and ecology of L. schmitti, F. brasiliensis and F. subtilis may elucidate questions referring to the abundance, period, and phase of occurrence of these shrimp genera in Guaraíras. Finally, the risks associated to the establishment of L. vannamei in the lagoon provide a novel outlet for studies in this biotope

The invasion of five alien species in the Delta do Parnaiba Environmental Protection Area, Northeastern Brazil

Marine biological invasions have been regarded as one of the major causes of native biodiversity loss, with shipping and aquaculture being the leading contributors for the introductions of alien species in aquatic ecosystems. In the present study, five aquatic alien species (one mollusk, three crustaceans and one fish species) were detected during dives, shore searches and from the fisheries on the coast of the Delta do Parnaiba Environmental Protection Area, in the States of Piaui and Maranhao, Northeastern Brazil. The species were the bicolor purse-oyster Isognomon bicolor, the whiteleg shrimp Litopenaeus vannamei, the giant river prawn Macrobrachium rosenbergii, the Indo-Pacific swimming crab Charybdis hellerii and, the muzzled blenny Omobranchus punctatus. Ballast water (I. bicolor, C. hellerii, and O. punctatus) and aquaculture activities (L. vannamei and M. rosenbergii) in adjacent areas are the most likely vectors of introduction. All exotic species found have potential impact risks to the environment because they are able to compete against native species for resources (food and habitat). Isognomon bicolor share the same habitat and food items with the native bivalve species of mussels and barnacles. Litopenaeus vannamei share the same habitat and food items with the native penaeids such as the pinkspot shrimp Farfantepenaeus brasiliensis, the Southern brown shrimp Farfantepenaeus subtilis, and the Southern white shrimp Litopenaeus schmitti, and in the past few years L. vannamei was responsible for a viral epidemics in the cultivation tanks that could be transmitted to native penaeid shrimps. Charybdis hellerii is also able to cause impacts on the local fisheries as the species can decrease the populations of native portunid crabs which are commercialized in the studied region. Macrobrachium rosenbergii may be sharing natural resources with the Amazon River prawn Macrobrachium amazonicum. Omobranchus punctatus shares habit with the native redlip blenny Ophioblennius atlanticus and other fishes, such as the frillfin goby Bathigobius soporator. Some immediate remedial measures to prevent further introductions from ballast water and shrimp farm ponds should be: (i) to prevent the release of ballast water by ship/vessels in the region; (ii) to reroute all effluent waters from shrimp rearing facilities through an underground or above-ground dry well; (iii) to install adequate sand and gravel filter which will allow passage of water but not livestock; (iv) outdoor shrimp pounds located on floodable land should be diked, and; (v) to promote environmental awareness of those directly involved with ballast water (crews of ship/vessels) and shrimp farms in the region. Rev. Biol. Trop. 58 (3): 909-923. Epub 2010 September 01.

Chave ilustrada para identificação dos camarões dendrobranchiata do litoral norte do estado de São Paulo, Brasil

O presente estudo teve como objetivo a elaboração de uma chave de identificação ilustrada para diferenciar as espécies de camarões marinhos Dendrobranchiata, com ocorrência no litoral do Estado de São Paulo até a profundidade de 40m. As 13 espécies apresentadas neste trabalho foram obtidas mediante coletas mensais durante os anos de 1995 a 2000 na região de Ubatuba, SP . Nesta chave estão incluídas as espécies de interesse econômico, como os peneídeos Xiphopenaeus kroyeri, Farfantepenaeus brasiliensis, F. paulensis, Litopenaeus schmitti, Artemesia longinaris e o solenocerídeo Pleoticus muelleri. Além destas espécies, também foram adicionados os camarões que não são alvos das frotas pesqueiras, entre eles, Rimapenaeus constrictus, Acetes americanus, Peisos petrunkevitchi e os sicionídeos Sicyonia dorsalis, S. typica, S. laevigata e S. parri. A chave proposta servirá como uma ferramenta no auxílio da identificação dos camarões Dendrobranchiata, quer seja por pesquisadores ligados à área científica, como também por pessoas relacionadas aos órgãos responsáveis pelo controle da pesca, principalmente, na época do defeso.

Biodiversidade e variação espaço-temporal da abundância das populações de camarões e estrutura populacional de Farfantepenaeus subtilis (Pérez-Farfante, 1967) (Penaeidae) em um estuário do litoral amazônico brasileiro

Este estudo investigou a biodiversidade e a distribuição espaço-temporal da abundância das populações de camarões e descreveu a estrutura populacional de Farfantepenaeus subtilis em um estuário amazônico. Os camarões foram coletados mensalmente de agosto de 2006 a julho de 2007 nos períodos chuvoso (janeiro a junho) e seco (julho a dezembro) com arrasto-de-fundo e arrasto-de-praia nos setores Médio-Superior (MS), Médio (M) e Inferior (I). Em cada setor dois locais foram amostrados, nos quais dois arrastos de fundo de cinco minutos cada e, três arrastos praiais de 150 m² cada um foram realizados. As variáveis: temperatura, pH e salinidade da água e granulometria e matéria orgânica do sedimento foram analisadas. Para cada camarão foram anotados os Comprimentos Total (CT) e do Cefalotórax (CC), peso total, sexo e estádio de maturação gonadal. A salinidade diferiu significativamente entre os setores I e MS (p<0,05) e o M apresentou baixo teor de matéria orgânica. Em todos os setores os grãos arenosos foram predominantes no substrato, destacando-se a areia fina no setor M. Foram coletados 11.939 camarões, distribuídos em doze espécies e seis famílias. Palaemonidae e Penaeidae tiveram maior riqueza com cinco e três espécies, respectivamente. Penaeidae, Sergestidae e Palaemonidae apresentaram maior abundância, com Xiphopenaeus kroyeri, F. subtilis, Acetes marinus e Nematopalaemon schmitti, contribuindo com 43, 31, 21 e 1,3% do total capturado, respectivamente. As espécies F. subtilis, X. kroyeri e Litopenaeus schmitti contribuíram com 97% da biomassa total. A densidade e a biomassa de F. subtilis diferiram entre o chuvoso e seco (p<0,05), sendo maior no chuvoso. No chuvoso, a densidade de X. kroyeri foi mais elevada nos setores M e I e no seco nos setores MS e M, com maior biomassa no I em ambos períodos sazonais. A densidade e a biomassa de A. marinus foi elevada no setor I (p<0,05). A salinidade, areia, silte e argila influenciaram significativamente a distribuição espaço-temporal da abundância dos camarões (p<0,05). O CC das fêmeas de F. subtilis foi superior aos machos com 13,7 mm e 12,8 mm, respectivamente, e diferiu em março (p<0,05). As fêmeas foram mais abundantes e a razão sexual foi de 0,8:1 com destaque para janeiro, fevereiro, maio e o total analisado (p<0,05). Camarões desenvolvidos ocorreram no chuvoso e a abundância de juvenis foi maior de janeiro a junho quando ocorre o pico mais intenso de desova em mar aberto.

Estrutura da comunidade e biologia reprodutiva dos camarões marinhos (Penaeidea e Caridea), no complexo Baía-Estuário de Santos e São Vicente/SP, Brasil

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Ecologia das populações e comunidades dos camarôes Decapoda no litoral norte de Santa Catarina

Pós-graduação em Ciências Biológicas (Zoologia) - IBB

Abundância e distribuição sazonal de espécies autóctones de camarões peneídeos no canal de abastecimento do sistema primar de aquicultura orgânica (Laguna de Guaraíras, Tibau do Sul, Rio Grande do Norte, Brasil)

Guaraíras lagoon, located in Tibau do Sul in the eastern littoral of Rio Grande do Norte (Brazil), presents a permanent connection to the sea, which guarantees the occurrence of a rich biodiversity, which includes the autochthonous shrimp species Litopenaeus schmitti, Farfantepenaeus subtilis and Farfantepenaeus brasiliensis. In spite of being subject to a strong human intervention in the last decade, mainly related to the installation of shrimp (Litopenaeus vannamei) farms, the lagoon is still scarcely studied. The present study aims at characterizing the populations of the three autochthonous penaeid shrimp species inhabiting Guaraíras, taking into consideration their abundance and seasonal distribution in the inflow channel of Primar System of Organic Aquaculture (Tibau do Sul, Rio Grande do Norte, Brazil). Twelve monthly samples were carried out from May 2005 to April 2006 with the aid of a circular cast net in the inflow channel, which is daily supplied with water from Guaraíras. Sampling months were grouped in trimesters according to the total pluviosity, thus comprising four trimesters. Water salinity was monitored twice a week and temperature values registered on a daily basis at noon, during the study period. The daily pluviosity data from the municipality of Tibau do Sul were supplied by Empresa de Pesquisa Agropecuária do Rio Grande do Norte (EMPARN). Collected shrimp were identified, weighted, measured and sexed. L. schmitti specimens (0.2 g to 17.8 g) were distributed in 1.3 g weight classes intervals. From the eighth sampling month (December 2005) onwards, males were classified into three categories, in accordance with the development of their petasm: (a) rudimentary petasm, (b) partially formed petasm, and (c) completely formed petasm. Among the ecological variables, rainfall showed the greatest dispersion (s.d.=187.74Rainfall and abundance of L. schmitti were negatively correlated (r = -0.85) whereas its abundance and water salinity were positively correlated (r = 0.63). Among 1,144 collected individuals, 1,127 were L. schmitti, 13 were F. subtilis and 4 were F. brasiliensis, which corresponded to 98.51%, 1.14% and 0.35% of the total of collected individuals. L. schmitti occurred in 100 % of all samples. Differently, the presence of F. subtilis and F. brasiliensis was restricted to 33% and 17% of the collected samples, respectively. The present study confirmed the occurrence of L. schmitti, F. brasiliensis and F. subtilis in Guaraíras. However, this lagoon seems to be primarily inhabited by juvenile Litopenaeus schmitti. The population of L. schmitti analysed showed a seasonal pattern of distribution. In general, in the months of high salinity and absence of rain, the number of individuals was higher than in the wet months. Further studies on the reproductive biology and ecology of L. schmitti, F. brasiliensis and F. subtilis may elucidate questions referring to the abundance, period, and phase of occurrence of these shrimp genera in Guaraíras. Finally, the risks associated to the establishment of L. vannamei in the lagoon provide a novel outlet for studies in this biotope