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Features of two populations of Anostraca (Crustacea: Branchiopoda) under cultivation in Southeastern Brazil.

(Resumo expandido apresentado e publicado no I Simpsio Internacional de

Ecologia 10/2011).

Marcelo Grombone de Vasconcellos (Federal University of So Carlos, Department of Ecology and Evolutionary Biology, DEBE-CCBS), Mara Alcntara Proena ((Federal University of So Carlos, Department of Ecology and Evolutionary Biology, DEBE-CCBS), Camila Pigo Soares (Federal University of So Carlos, Department of Genetics and Evolution Labolatory of Adaptive Biochemistry CCBS) Maria Carolina Vasconcellos Sanchez (Graduanda em cincias biolgicas, Unesp-Botucatu) Rodrigo Ferroni Passos (Federal University of So Carlos, Department of Hidrobiology. DHB-CCBS)

Resumo: As caractersticas da populao de Artemia franciscana Kellogg, 1906 e Dendrocephallus brasiliensis (Pesta, 1920), sob cultivo na estao de aqicultura da Universidade Federal de So Carlos, SP foram avaliadas quanto estrutura populacional, variveis fsicas e qumicas no meio de cultivo e valores nutricionais dos organismos cultivados, em termos de carboidratos, lipdeos e protenas totais. A reproduo da espcie Artemia franciscana reportada como partenogentica e tambm sexuada, havendo produo de cistos e nuplios continuamente, enquanto que a espcie aparentada D. brasiliensis, adaptada a lagoas temporrias, se reproduz apenas sexuadamente, produzindo cistos de resistncia que necessitam passar por um perodo de seca antes da ecloso. Para A. franciscana as densidades obtidas variaram entre 625.500 e 136.500 indivduos por metro cbico, em mdia, ao longo de um perodo de 60 dias, enquanto que para D. brasiliensis a densidade mdia variou entre 27.000 e 260 indvduos por metro cbico Em amostras populacionais, coletadas aps 60 dias, para o cultivo de A. salina, registrou-se 64,5 % de nauplios, 12,6 % de juvenis e 22,7 % de adultos. Destes 12,5 % eram machos e 10,2 % fmeas, denotando o recrutamento contnuo de novos indivduos populao. Para o cultivo de D. brasiliensis foram registrados, aps 60 dias, ocorreram apenas adultos, sendo 49,7 % fmeas e

50,4% machos, evidenciando que no h recrutamento em D. brasiliensis. Em relao composio bioqumica D. brasiliensis apresentou comparativamente maiores valores relativos de carboidratos, lipdeos e protenas totais do que A. franciscana, evidenciando um maior potencial para a sua utilizao em aqicultura, dado o maior valor nutricional. Em D. brasiliensis apenas uma gerao cresce e se reproduz a cada ciclo depositando os ovos de resistncia e assim o incremento populacional e o recrutamento s ocorrerem aps a secagem do corpo dgua e o novo enchimento, relembrando as condies na natureza e as foras seletivas que moldaram as estratgias reprodutivas desta espcie. Abstract: The characteristics of Artemia franciscana Kellogg, 1906 and Dendrocephallus brasiliensis (Pesta, 1920) populations cultured at the aquaculture station of the Federal University of So Carlos, SP, were evaluated regarding the population structure, physical and chemical variables in the culture tanks and the nutritional value of the organisms cultured, in terms of carbohydrates, lipids and total protein. The reproduction of Artemia franciscana is reported occurring by both systems, parthenogenetic and sexual, females continuously producing cysts and nauplii, while the related species D. brasiliensis which is highly adapted to temporary ponds just reproduces sexually, with resting cysts that require to pass through a dry period before hatching. For A. franciscana the recorded densities ranged from 625.500 to 136.500 individuals per cubic meter, on average, over a period of 60 days, whereas for D. brasiliensis the average density ranged from 27.000 to 260 individuals per cubic meter. In a population of A. salina sampled after 60 days cultivation, the relative proportion of 64.5 % nauplii, 12.6 % juveniles and 22.7 % (12.5 % males and 10.2 % females) was found, denoting continuous recruitment of new individuals to the population. In the population of D brasiliensis, after 60 days, only adults

were recorded, being 49.7 % females and 50.4 % males, thus evidencing that there is no egg hatching nor recruitment in D. brasiliensis. In relation to the biochemical

composition D. brasiliensis presented comparatively higher values for carbohydrate, lipid and total protein than A. franciscana, evidencing a greater potential for use in aquaculture, given its higher nutritional value. In D. brasiliensis only one generation grows and reproduces, in each life cycle. Resting eggs are deposited and the increase of population by young recruitment will only occur after the complete dry up of the water body and next season refilling, recalling the conditions in nature, thus responding to selective forces that shaped breeding strategies of this species. Introduction/ Aim The Branchiopoda-Anostraca are primitive forms of crustaceans, whose oldest fossil records go back to the Lower Cretaceous (Fryer, 1987 apud Weekers, et al, 2001). Known around the world as fairy shrimp due to its swimming in a reminiscent mannert of the graceful mythical beings, are widely spread over almost the entire globe, having been irradiated before the bone fishes, remaining in inaccessible or inhospitable

environments for the latter (Weekers et al, 2001). These species are characteristic of temporary, highly stochastic environments, such as temporary ponds, monsoon rivers and salt ponds, with species adapted to extremely high or low temperatures. His most famous representatives are the species of the cosmopolitan genus Artemia, an Anostraca that reaches about 1.2 cm, widely used in aquaculture word widely as a major subsidy for fishes and shrimps production (Atencio-Garcia, 2000). For these reasons D. brasiliensis, a branchiopod that reaches more of 2 cm in length, are being cultivated, mainly in the Northeast of Brazil since the last century (Lopes et al, 1998) as a live food in fishculture. This study aimed to characterize the populations of Dendrocephalus

brasiliensis and Artemia franciscana, grown in a controlled condictions, in the aquaculture station of the Federal University of So Carlos, So Paulo, Brazil, aiming to evaluate the conditions for its management and production, and to assess its nutritional values regarding carbohydrates, lipids and proteins contents of both species. Materials and Methods For the cultivation of A. salina 10 grams of cysts were inoculated into two circular tanks of 2,000 liters each, containing 30 grams of NaCl, per liter of water. For D. brasiliensis approximately 500 grams of sediment containing on average 220 cysts per gram were inoculated in 4 tanks of 10,000 liters each. Feeding was provided, after the cysts hatching, consisting of a mixture of 10 grams of dried yeast, dissolved in 15 liters of water, and a further addition of 10 grams of bird offspring food, ALCON trademark. This mixture was introduced in 2 cubic meters of water in A. franciscana culture tanks and in 10,000 liters tanks in the case of D. brasiliensis, on alternate days. After hatching the nauplii were provided with the same food used for A. franciscana cultivation. The populations were mantained under greenhouse in the aquaculture station at the Federal University of So Carlos, So Paulo, Brazil. Populations were sampled weekly by filtering 170 liters and 50 liters in the tanks with D.brasiliensis and Artemia respectively. After 60 days of cultivation, the number of individuals and the proportion of developmental stages and sexes were recorded.. Individuals considered adults by the presence of ovigerous sac in females, were treated according to the techniques of Dubois (1956) and Bradford (1976), to estimate of the percentage of carbohydrates and proteins in each species, for the food conditions reported above. The measurement of lipids was performed according to the techniques presented in Patarra (2008). Also, weekly, physical and chemical

parameters of the water in the cultures were monitored by using the multisensor Yellow Spring 500 probe. The data were processed through the software GraphPadInstat applying the variance test (ANOVA), parametric tests T, with or without Welchs correction and non-parametric Mann-Whitney test (Zar, 1999). Results/Discussion There were no significant differences between the sex ratios recorded in the cultures of A. franciscana (n=188, p=0,86) and D. brasiliensis (n= 165=0,88) (Figures 1 and 2) as mentioned in Grombone-Vasconcellos (in press) for Southeastern D.brasiliensis crops, unlike Northeast D.brasiliensis crops recorded by Lopes et al, (1998, 2007). For D. brasiliensis and A. franciscana higher rates of mortality during the first week were recorded (Figures 3 and 4). For D. brasiliensis, the sexual maturity was reached after

approximately 30 days, at the mean temperature of 20 C (Table 2), but no nauplii were recorded since then. This greatly differentiate the reproductive strategies of these two Anostaca because for A. franciscana, which reached sexual maturity at approximately 25 days, after 60 days nauplii were still the main developmental stage recorded, and probably responsible for the population density recuperation, corresponding to the slightly upward portion of the curve in the graph of population density (Figure 3). For D. brasiliensis, there no recruitment, therefore just one generation hatches, grows, spawns and dies at each cycle in the environment. So the population density drops sharply in the first week and continues decreasing less abruptly after the sexual maturity of individuals. Only the cysts or deposited eggs hatches, rising a new population after the complete drying of the pond and its refilling, in the raining season. This strategy was selected on

the temporary habitats without fishes, typical of arid regions of the Neotropics (Weekers et al, 2001; Grombone-de-Vasconcellos, 2010., Passos et al, in press).

70 60 50 40 30 20 10 0 Juveniles Nauplii Adult males Adult females

Figure 1 Population structure, expressed as relative abundances (%) for Artemia franciscana after 60 days of cultivation in the Aquaculture Station of The Federal University of So Carlos, SP.
15,6 15,5 15,4 15,3 15,2 15,1 15 14,9 Adult males Adult females

Figure 2 Population structure, in percentage, D. brasiliensis, after 60 days in the Aquaculture Station of The Federal University of So Carlos, SP.
750 700 650 600 550 500 450 400 350 300 250 200 150 100 50 0 21/jun 28/jun 05/jul 12/jul 19/jul 25/jul 02/ aug 09/aug 16/ aug 23/aug

R3 R4

Figure 3 Changes of density (ind/m3) recorded for A. franciscana grown in the tank of the R3 and R4 of Aquaculture Station of The Federal University of So Carlos, SP.
40 35 30 25 20 15 10 5 0 21/jun 28/jun 05/jul 12/jul 19/jul 25/jul 02/aug 09/aug 16/aug 23/aug

Q1 Q2 Q3 Q5 Q6

Figure 4 Curve of density (ind/m3) recorded for D. brasiliensis grown in the tank Q1, Q2, Q3, Q5 and Q6 of Aquaculture Station of The Federal University of So Carlos, SP.

In the conditions of Southeastern Brazil, especially during winter, low temperatures slow the growth and maturity of individuals, and thus biomass production of D. brasiliensis. In the summer, with temperatures averaging 32 degrees, the growth period is reduced to eight days and maturity can be reached within 5 days (Lopes et al 1998, 2007; Grombone-Vasconcellos, in press).The nutritional value of these organisms, grown under the above conditions are summarized in Table 3, which denote superiority in protein, lipids and carbohydrates for the species D. brasiliensis when compared with Artemia franciscana, a species widely used as a food resource in aquaculture.
Table 1 Mean values (%) of total carbohydrates, proteins and lipids, for individuals of A. franciscana and D. brasiliensis.
Species Carbohydrates for individuals (%),mean standard deviation 21,1 0,9 23,25 1,14 Total protein per individual (%),mean standard deviation 35,5 0,5** 51,7 2,06** Lipids per individual (%),mean standard deviation 15,5 4,3 16,8 4,8

A. franciscana D. brasiliensis

**Highly significant values p=0,00017 Table 2- Physical and chemical parameters of waters (average values + standard deviation) of D. brasiliensis and A. franciscana, culture tanks at the Aquaculture Station of the Federal University of So Carlos, SP.
Parameters pH Cond S/CM Q1 8,7851 75,225 Q2 8,2823 91,314 Q3 8,5348 50,328 Q5 8,4015 153,157 Q6 8,3026 104,636 R3 9,0112 89,912 (mS/cm) R4 8,001,12 86,418 (mS/cm)

OD mg/l 0 Temp C

7,11,2 213,8

6,01,5 21,53,5

6,01,2 202,4

6,11,8 233,5

6,01,2 23,53,2

6,661,8 22,83,2

5,582,03 22,53,1

Based in the results here obtained it can be concluded that the cultivation of these Anostraca of relatively high nutritional value, especially in the case of D. brasiliensis, have peculiarities that must be incorporated at the techniques for commercial exploitation of these species as food source in shrimp and fish farming. For Artemia the most obvious costs involved are related to the necessary salinization of the farm ponds and the first acquisition of cysts, but cultivation has the advantage of being continuous with controlled harvest of the biomass produced. For D. brasiliensis cultures population harvesting must be performed only after they have reached sexual maturity and produced the cysts, preferably in the summer, when production cycles are shortest (Grombone-Vasconcellos, in press). The requirement of renewing the water of tanks only after the total drying of the soil is the most costly part of the process due to the costs involved with the water resource. These drawbacks could be minimized with the acquisition of new cysts produced outside the culture tanks, for renewing the population by re-inoculation. Acknowledgements The authors thanks to FAPESP for funding the thematic project and the Federal University of So Carlos, its students and employees, by subsidies and facilities. Bibliography DUBOIS, M. ; GILLES, K. H. ; HAMILTON, J. K. ; ROBERTS, P. A. ; SMITH, F. Calorimetric method for determination of sugars an related substances. Anal. Chem., c. 28, p. 350-358, 1956.

GROMBONE-VASCONCELLOS, M. Caracteristicas populacionais, desenvolvimento e produo da branchoneta (Dendrocephalus brasiliensis pesta 1921), sob as condies climticas da regio sudeste do pas. Revista Brasileira de Zoocincias. No prelo.

LOPES, J. P.; DA SILVA, A. L. N.; DOS SANTOS, A. J. G & TENRIO, R. A. Branchoneta, uma notvel contribuio larvicultura e alevinagem de peixes carnvoros de gua doce. Panorama da aqicultura, p 31-34, 1998. LOPES, J. P.; GURGEL, H. C. B.; GLVEZ, A. O. & PONTES, C. S. Produo de cistos de branchoneta Dendrocephalus brasiliensis (Crustacea: Anostraca). Biotemas, 20 (2): 33-39, 2007 PATARRA, A. R. F. Pesquisa de cidos gordos em macroalgas marinhas do litoral de Aores. Dissertao (Mestrado em Cincias do Mar) Instituto de Cincias Biomdicas de Abel Salazar, Universidade do Porto, Porto, 2008. WEEKERS, P. H. H.; GOPAL, M. ; VANTEFLETEREN, J. R. BELK, D. & DUMONTA, H. J. A Phylogenetic analysis of anostracans (Branchiopoda: Anostraca) inferred from

nuclear 18S ribosomal DNA (18S rDNA) sequences. Molecular Phylogenetics and Evolution 25 (2002) 535544, 2001. ZAR, J. H. Biostatistical Analysis 4th ed. Prentice-Hall Inc., New Jersey, USA. 663p. 1999.

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