Lília P. Souza-Santos

Development and population dynamics of Tisbe biminiensis (Copepoda: Harpacticoida) reared on different diets

Abstract The harpacticoid copepod Tisbe biminiensis was reared under controlled laboratory conditions. In order to study the effects of food on larval development and on population dynamics, three diets were tested: the microalgae Nitzschia closterium , Tetraselmis gracilis and a mixture of both algae. The life-cycle parameters were measured, and the demographic variables such as generation time, net reproductive rate, and exponential rate of increase were determined. Results showed that diet affected development and fecundity. Larval development was delayed in copepod fed T. gracilis . The mean number of nauplii per brood was significantly greater in copepod fed the N. closterium diet than those fed the other diets. Survival did not differ significantly between treatments. The exponential rate of population increase ( r ) was greater in copepod fed the N. closterium diet (0.49 day −1 ) and lower in those fed the T. gracilis diet (0.35 day −1 ). The results showed that although Tis. biminiensis seemed to be able to develop and reproduce on a T. gracilis diet, the diatom N. closterium promoted better results for this copepod in terms of development, fecundity and populational growth rates. The short generation time and high reproductive potential make the use of this copepod promising as live food in aquaculture.

Sensitivity of the marine benthic copepod Tisbe biminiensis (copepoda, harpacticoida) to potassium dichromate and sediment particle size

For the future use of the marine benthic copepod Tisbe biminiensis in solid-phase sediment toxicological bioassays, the present study investigated the effect of muddy sediment from the Maracaipe estuary (northeastern Brazil), sediment particle size and the reference toxicant potassium dichromate on the species. Muddy sediment from Maracaipe can be used as control sediment, since it does not interfere in the copepod life-cycle and has metal contamination levels that are unlikely to produce any detrimental biological effects on benthic invertebrates. Neither survival nor fecundity was affected by grain size, suggesting that this species can be used with any kind of sediment from muddy to sandy. The sensitivity of T. biminiensis to K2Cr2O7 in acute tests was similar to that of other organisms. The LC50 (lethal concentration to 50% of the test organisms) medium values for T. biminiensis were 7.51, 4.68 and 3.19 mg L-1 for Cr in 48, 72 and 96 h, respectively. These results suggest that T. biminiensis is a promising organism for use in solid-phase sediment toxicity assessments.

Spatio-Temporal Variations of Microphytobenthos in the Botafogo and Siri estuaries (norteast - Brazil)

Spatio-temporal variations of microphytobenthic biomass were investigated in the Botafogo and Siri estuaries, Pernambuco - Brazil. Samples were taken during low water (bi-monthly) from February to December, 2003. Three sampling stations were located in each estuary, in both midlittoral and sublittoral areas. Clear spatial and temporal variations were found for both chlorophyll-a and phaeopigment concentrations. In Botafogo, chlorophyll-a had 34% of total variation explained by MRA from nitrogen and phosphorus concentrations, while only 16% of total variation was thus explained from nitrogen concentration in the Siri estuary. Phaeopigments had 39% of total variation explained by MRA from nitrogen concentration and salinity in the Botafogo estuary and 58% of total variation was similarly explained from nitrogen concentration in the Siri estuary. Nitrogen was discriminated by the b coefficients as the most important factor in the explanation of the microphytobenthic pigments variations in the Botafogo estuary. The data of this study emphasize the role of nutrients and suggest that both estuaries show evidence of eutrophication.

Toxicity of sediments from a mangrove forest patch in an urban area in Pernambuco (Brazil).

Industrial and urban residues are discharged every day to the rivers and may arrive at the mangrove forest and prejudice the quality of the environment and the organisms present there. The mangrove forest patch studied is encircled by an urban area of the city of Recife (Brazil) that has approximate 1.5 million inhabitants and is one of the most industrialized centers in Northeast Brazil. The aim of this study was to assess the quality of the sediments of this mangrove patch in terms of metal contamination and ecotoxicology. Samples of surface sediment were collected in six stations for toxicological tests and trace metal determination (Cr, Zn, Mn, Fe, Cu, Pb, Co and Ni), in July and August, 2006 (rainy season); and in January and February 2007 (dry season). Toxicity tests with solid-phase sediments were carried out with the copepod Tisbe biminiensis in order to observe lethal and sub-lethal endpoints and correlate them with chemical data. In June, there were no observed lethal effect, but two stations presented sub-lethal effects. In January, lethal effect occurred in three stations and sub-lethal in one station. The levels for Zn and Cr were at higher levels than international proposed guidelines (NOAA). There was a negative significant correlation between the copepods׳ fecundity, and Zn and Cr concentrations. Therefore, the studied sediments can be considered to have potential toxic to benthos due to the high content of Zn and Cr.

EFFECT OF DIET ON THE FATTY ACID COMPOSITION OF THE COPEPOD TISBE BIMINIENSIS

Success in rearing fish depends on the nutritional content of food offered to the larvae. The aim of this study was to test the effect of diets on fatty acid composition of the copepod Tisbe biminiensis Volkmann-Rocco, 1973 as a food source. The copepods were reared in 5 l vessels with aeration at 29-31°C, salinity 30-35, and natural photoperiod of 13 hours light/11 hours dark. The copepods were fed two diets: a mixture of microalgae Thalassiosira weissflogii and commercial fish food, and only the fish food. Lipids were extracted with chloroform and methanol and esterified by acid catalysis with BF3. Three replicates were used in each treatment. Fatty acids were identified by comparison of retention times and co-injection with standard Mix C4-C24 using GC and GCMS apparatus. Thirty-one and 29 fatty acids were detected on copepods fed mixed diets and fish food, respectively. The most abundant fatty acids in copepod fed mixed a diet were C18, C16, and C12. In the other treatment, predominant fatty acids were C18, C16, and C20:2. Thirty-one and 30 fatty acids were detected in the fish food and the microalgae, respectively. The predominant fatty acids of microalgae were C18, C21, and C20:2. For fish food, the predominant fatty acids were C16, C18, and C18:1n9. Essential fatty acids were detected and EPA and DHA contents of copepods fed mixture were significantly higher compared to those fed only ration, indicating that the mixed diet provided better source of HUFA for T. biminiensis. However, its DHA level was lower than other harpacticoid copepods. This suggests that T. biminiensis either needs to be fed a better quality diet, or be enriched with some fatty acids before they are offered to the cobia larvae. The copepod T. biminiensis is capable of bio-converting fatty acid C18:0 into chains with 20 unsaturated carbons. T. biminiensis fed mixed a diet or the fish food alone showed DHA/EPA ratios of 1.3 and 3.3, respectively, suggesting that this copepod species may be suitable for use in fish larviculture.

A new protocol for ecotoxicological assessment of seawater using nauplii of Tisbe biminiensis (Copepoda:Harpacticoida).

Copepods are largely used in toxicity tests. The nauplii of these organisms are more sensitive to contaminants than the adult stage. The aim of the present study was to test a protocol for the use of nauplii of the copepod Tisbe biminiensis in the ecotoxicological assessment of seawater. The sensitivity of these organisms to zinc sulphate (ZnSO4·7H2O) was also determined. The following conditions were established for the protocol based on the best development of nauplii to copepodites: 72-h duration, the microalga Chaetocerus gracilis at 2.5×10(5)cellsmL(-1) as feed and incubation temperature of 28°C. In the zinc sulphate sensitivity tests, EC50-72 h and LC50/72 h were 3.25±0.59 mg L(-1) and 3.46±0.72 mg L(-1), respectively, as estimated by the final number of copepodites and total number of live animals in relation to the mean number of inoculated nauplii. The estimated NOEC was 2.0 mg L(-1). The test developed is fast and not labour intensive. T. biminiensis nauplii exhibit sensitivity to zinc sulphate similar to that of other species of copepods employed in water toxicity tests, demonstrating the usefulness of these organisms in ecotoxicological studies involving samples of environmental seawater.

Methyl esters from the copepod Tisbe biminiensis assayed by two transesterification methods

The present study assessed the efficiency of two transesterification methods for the identification of fatty acids in the copepod Tisbe biminiensis Volkmann-Rocco, 1973. In the first method, the samples were esterified by basic catalysis with KOH in methanol, and in the second method they were esterified by acid catalysis with BF3. Fatty acids were identified by comparison of retention times and co-injection with standard Mix C4-C24, using gas chromatography and gas chromatography-mass spectrometry. Twenty-nine fatty acids were detected in samples transesterified by basic catalysis or acid catalysis. The predominant fatty acids were C18 (19.18%), C16 (13.02%), and C12 (11.39%) for samples transesterified by acid catalysis; and C13 (25.24%), C21 (11.32%), and C18 (10.65%) for samples transesterified by basic catalysis. The results show that fatty acid analysis for copepods may be affected by the transesterification method. Sample transesterification by acid catalysis was efficient to identify monosaturated and polyunsaturated fatty acids, whereas transesterification by basic catalysis was efficient to identify saturated fatty acids. Essential fatty acids were present in the copepod, which suggests that T. biminiensis is a useful food to be used in larviculture.

PROTEASES FROM THE HARPACTICOID COPEPOD TISBE BIMINIENSIS: COMPARATIVE STUDY WITH ENZYMES FROM FARMED AQUATIC ANIMALS

Abstract The harpacticoid copepod Tisbe biminiensis has been tested as live prey in marine crustacean and fish larviculture. The aim of the present study was to characterize the proteinases in the crude extract of T. biminiensis. Trypsin activity was assayed in the crude extract prepared by the homogenization of specimens reared under controlled laboratory conditions and fed on diatoms and commercial fish food. The physical-chemical and kinectics parameters were determined using BApNA as substrate. Optimal pH and temperature were 9.0 and 55°C, respectively. This enzyme was thermostable until 50°C. Using BApNA as substrate, the Km was 0.59 mM and the proteolytic activity was strongly inhibited by specific trypsin inhibitors. However, the presence of other proteases was observed using substrate-SDS-PAGE. Eight caseinolytic bands were observed in the zymogram, four of them were not inhibited by classical mammalian trypsin inhibitors. Trypsin from T. biminiensis showed similar properties to those described for species used in commercial aquaculture. These results demonstrate that T. biminiensis may be a source of proteases, including trypsin-like enzymes.

Use of Tisbe biminiensis nauplii in ecotoxicological tests and geochemical analyses to assess the sediment quality of a tropical urban estuary in northeastern Brazil

An approach pooling geochemical analyses and ecotoxicological tests has been applied to assess the sediment quality of the Capibaribe River Estuary, Brazil. Toxicity tests were performed to compare a well-established, labor-intensive protocol using ovigerous females to a new, easier and faster protocol using nauplii of the epibenthic marine copepod Tisbe biminiensis. The endpoints of the nauplii toxicity test were comparable to those of the female test. Nauplii proved to be more sensitive than females as a biological model for indicating sediment toxicity. All sediments collected had at least one contaminant above the threshold effects level (TEL) proposed in the literature. Furthermore, more than one-third of samples exhibited contaminants above the probable effects level (PEL). The PCA revealed that nauplii mortality was associated with metals in October 2014, which was confirmed by the Spearman correlation factor. In contrast, no strong association among contaminants and toxicological endpoints in May 2015 was found.