Search

Partial characterization of digestive proteases in the three-spot cichlid Cichlasoma trimaculatum juveniles was conducted. It was determined that there is higher alkaline proteases activity (3.95 ± 0.32 IU mg−1 protein) compared to acidic proteases (2.01 ± 0.57 IU mg−1 protein). Optimal temperature for alkaline proteases is 60 °C which resulted in more thermostability to temperature changes. On the other hand, optimal temperature for acidic proteases is 50 °C. Optimal pH for acidic proteases was pH 2, while for alkaline proteases, it was pH 10, which resulted in more stability in relation to pH changes than acidic protease. The use of specific inhibitors and the SDS-PAGE electrophoresis analysis revealed seven types of bands for alkaline proteases, which make evident the main presence of serine proteases. In acidic proteases, more than 98 g kg−1 of the activity was inhibited with pepstatin A inhibitor. Therefore, it is evident that C. trimaculatum digestion is composed by acidic and alkaline proteases; thus, it should be considered an omnivorous fish.

A study was performed in order to understand the development of digestive enzymes during initial ontogeny of Cichlasoma trimaculatum, for which the activity of acidic and alkaline proteases, lipases, amylases and phosphatases was determined by means of biochemical and electrophoretic analysis. Our results showed that the activity of alkaline proteases, trypsin and chymotrypsin is present from day 6 after hatching (dah) during exogenous feeding with Artemia nauplii. The activities of carboxypeptidase A and leucine aminopeptidase are present from the first days, increasing at 6 dah and reaching their maximum activity at 9 dah while acid protease activity started at 9 dah. Furthermore, the lipase activity is detected on 6 dah and keeps increasing and decreasing on 17 dah. Amylase activity is detected on 3 dah, presenting fluctuations until 45 dah, where it reaches its maximum activity. Acid and alkaline phosphatases are detected from 3 dah and reach a maximum activity between 13 and 19 dah. The SDS-PAGE electrophoresis revealed six types of bands in the alkaline proteases, with molecular weight between 113.4 and 20.4 kDa. First three bands appear on 6 dah, but it is until 11 dah when all isoforms appear. Based on these results, it is considered that this species completes its digestive enzymatic machinery from day 9 after hatching, therefore is recommended to perform the transition from live feed to inert feed at 15 dah.

Summary In bay snook (Petenia splendida) larvae the histological development of the digestive system and swim bladder, and their relative timing of differentiation were studied from hatching to 45 days post-hatch (dph) at 29°C. Newly hatched larvae showed a simple digestive tract, which appeared as a straight undifferentiated tube lined by a single layer of columnar epithelial cells (future enterocytes). The anatomical and histological differentiation of the digestive tract and accessory glands was a very intense, asynchronous process, proceeding from the distal to the anterior part. The intestine was the first region to differentiate (9 days post-hatch – dph, 6.5 mm SL), and the oesophagus the last (21 dph, 8.4 mm SL). At the onset of feeding, the digestive system was organized into different functional and histologically differentiated sections, such as the buccopharynx, oesophagus, glandular stomach, and anterior and posterior intestine. This organization resembled that of the juveniles, with the exception of pharyngeal teeth and buccopharyngeal as well as oesophageal goblet cells, which proliferated later during the mixed feeding period. Histological observations revealed that bay snook larvae retained endogenous yolk reserves until 24 dph (8.9 ± 0.4 mm SL), which might be helpful for weaning this species onto a compound diet. The important lipidic accumulation observed in the intestinal mucosa, liver, and pancreas in fish fed a compound trout diet indicated that although fish were able to digest and absorb lipids, the diet formulation did not fit the nutritional requirements of early juveniles of this species. The ontogeny of the digestive system followed the same general pattern as in most cichlid species described to date. However, we detected species-specific differences in the timing of differentiation that were related to their reproductive guild. According to the histological results, some recommendations regarding the intensive culture of this species are also provided.

The development of digestive enzymes during the early ontogeny of the Mayan cichlid (Cichlasoma urophthalmus) was studied using biochemical and electrophoretic techniques. From yolk absorption (6 days after hatching: dah), larvae were fed Artemia nauplii until 15 dah, afterward they were fed with commercial microparticulated trout food (45% protein and 16% lipids) from 16 to 60 dah. Several samples were collected including yolk-sac larvae (considered as day 1 after hatching) and specimens up to 60 dah. Most digestive enzymes were present from yolk absorption (5-6 dah), except for the specific acid proteases activity (pepsin-like), which increase rapidly from 8 dah up to 20 dah. Three alkaline proteases isoforms (24.0, 24.8, 84.5 kDa) were detected at 8 dah using SDS-PAGE zymogram, corresponding to trypsin, chymotrypsin and probably leucine aminopeptidase enzymes, and only one isoform was detected (relative electromobility, Rf = 0.54) for acid proteases (pepsin-like) from 3 dah onwards using PAGE zymogram. We concluded that C. urophthamus is a precocious fish with a great capacity to digest all kinds of food items, including artificial diets provided from 13 dah.

espanolEl efecto antropico, la perdida de biodiversidad, el desplazamiento de especies, la introduccion de especies exoticas y el incremento demografico, han promovido aumentar el rendimiento de los cultivos, la recuperacion de sitios impactados, la reintroduccion de especies y la busqueda de alternativas para comprender, y evaluar la reproduccion de las especies icticas nativas. En ello, el conocimiento y el empleo de los gametos de hembras y machos de los peces, para la produccion masiva de larvas hacen necesario el uso de metodologias que permitan seleccionar todos aquellos reproductores que muestren cualidades geneticas y fisiologicas que den pauta a mejorar e incrementar la cantidad y la calidad de las larvas que seran empleadas para cultivo masivo, repoblacion, produccion de reproductores y para su empleo en experimentos de ciencia basica y aplicada. En esta revision bibliografica, se hace mencion de las particularidades de las celulas espermaticas de los peces y de las alternativas metodologicas para evaluar su calidad, mediante la comprension de la fisiologia de la activacion (porcentajes y tiempo de motilidad, tipo de movimiento y numero de espermatozoides) y hasta el analisis estructural en el nivel extracelular e intracelular (morfologia espermatica, dano a la membrana, ensayo cometa, celulas vivas/muertas/apoptoticas, mismas que estan en funcion de los objetivos de los programas de reproduccion o experimentacion). Con el desarrollo de equipos de citometria automatica y de genomica unicelular, se ha logrado mayor comprension del funcionamiento de las celulas espermaticas, de la expresion genetica ligada a las cualidades celulares y del significado biologico. EnglishAspects such as anthropic effects, loss of biodiversity, species displacement, introduction of exotic species, and demographic growth have caused an increase in crop yield, recovery of impacted sites, reintroduction of species and the search for alternatives to understand and evaluate reproduction of native fish species. In such context, knowing and using both female and male gametes for massive larvae production creates the need to use methodologies that would allow sorting the brood stock with genetic and physiological traits that would improve the quantity and quality of larvae to be used for massive culture, restocking, breeding production, and experiments on basic and applied sciences. This bibliographic review mentions the particularities of fish sperm cells and methodological alternatives to assess their quality, by understanding activation physiology (motility percentages and time of activity, type of movement and sperm number), including the structural analysis at extracellular and intracellular levels (spermatic morphology, damage to the membrane, comet assay, live/dead/apoptotic cells, which depend on the main objectives of reproduction or experimental programs). The development of equipment to automated cytometry and unicellular genomics has allowed for a better understanding of the functioning of spermatic cells, genetic expression linked to cellular qualities and their biological implication

Two experiments were conducted to evaluate the initial stocking density in larvae of Bay snook, Petenia splendida, and Mexican mojarra, Cichlasoma urophthalmus, using a recirculation system. Five initial stocking densities (0.5, 1, 5, 10, and 20 larvae/L) were evaluated by triplicate for 45 d. Weight and total length (TL) were measured every 15 d, and fish production was calculated for each density. The larvae stocked at the lowest densities (0.5 and 1 larvae/L) presented the highest growth for both species: C. urophthalmus (0.78 g and 45-mm TL, and 0.76 g and 45-mm TL, respectively) and P. splendida (0.80 g and 52-mm TL, and 0.79 g and 49-mm TL, respectively). However, lowest fish production was recorded (35 and 69 fish per tank, respectively, for C. urophthalmus and 34 and 70 fish per tank, respectively, for P. splendida) compared with those at densities of 5, 10, and 20 larvae/L (336, 584, and 604 fish per tank, respectively, for C. urophthalmus and 341, 679, and 912 fish per tank, respectively, for P. splendida). The polynomial model for biomass production related to the stocking density shows that the optimum stocking densities for C. urophthalmus and P. splendida are 12 and 14 larvae/L, respectively.

Changes in digestive enzyme activity and histology were studied in Atractosteus tropicus embryos, larvae and juvenile periods. Alkaline protease, chymotrypsin, carboxypeptidase A, lipase and α-amylase were detected in all periods and gradually increased until reaching the maximum peak in juveniles; meanwhile, acid protease was first detected at 5 days after hatching (dah) when first feeding started and trypsin and leucine aminopeptidase activities were detected from 19 dah, their values being increased gradually until reaching a maximum value at 31 dah. Acid and alkaline phosphatase activities increased from yolk-sac absorption (3 dah) until day 31 after hatching. Zymogram for acid protease showed two bands in active forms (0.4 and 0.5 Rfs) from day 5 after hatching and a third protease form (0.3 Rf) that appears at 31 dah. Two active forms (26.3 and 24.9 kDa) were detected using SDS-PAGE alkaline proteases zymogram at 5 dah, and an additional active form (44.1 kDa) was detected at 7 dah. Regarding the histological development of the digestive system, the exocrine pancreas containing zymogen granules was already visible at 3 dah, whereas at 5 dah first gastric glands were already detected in the stomach. Between 7 and 9 dah, the digestive tract of A. tropicus resembled that of a juvenile specimen with a well-developed and short oesophagus, stomach divided into a glandular and non-glandular (pyloric) stomach, folded intestine with pyloric caeca and a well-developed spiral valve (posterior intestine). Considering this, larvae of A. tropicus are capable of digesting several foods from yolk absorption (3 dah), maximizing its activities at 15 dah, age at which the organisms maximize its capability to absorb nutrients from diets provided.

Common snook (Centropomus undecimalis) is one of the most important marine species under commercial exploitation in the Gulf of Mexico; for this reason, interest in developing its culture is a priority. However, larviculture remains as the main bottleneck for massive production. In this sense, our objective was to determine the changes of digestive enzymes activities using biochemical and electrophoretic techniques during 36 days of Common snook larviculture fed with live preys (microalgae, rotifers, and Artemia). During larviculture, all digestive enzymatic activities were detected with low values since yolk absorption, 2 days after hatching (dah) onwards. However, the maximum values for alkaline protease (6,500 U mg protein(-1)), trypsin (0.053 mU × 10(-3) mg protein(-1)), and Leucine aminopeptidase (1.4 × 10(-3) mU mg protein(-1)) were detected at 12 dah; for chymotrypsin at 25 dah (3.8 × 10(-3) mU mg protein(-1)), for carboxypeptidase A (280 mU mg protein(-1)) and lipase at 36 dah (480 U mg protein(-1)), for α-amylase at 7 dah (1.5 U mg protein(-1)), for acid phosphatases at 34 dah (5.5 U mg protein(-1)), and finally for alkaline phosphatase at 25 dah (70 U mg protein(-1)). The alkaline protease zymogram showed two active bands, the first (26.3 kDa) at 25 dah onwards, and the second (51.6 kDa) at 36 dah. The acid protease zymogram showed two bands (RF = 0.32 and 0.51, respectively) at 34 dah. The digestive enzymatic ontogeny of C. undecimalis is very similar to other strictly marine carnivorous fish, and we suggest that weaning process should be started at 34 dah.

Several samples of P. splendida larvae were obtained from eggs until day 60 after hatching (dah) to determine acid and alkaline proteases, trypsin, chymotrypsin, leucine aminopeptidase, α-amylase, lipase, and acid and alkaline phosphatase activities using biochemical techniques. Additionally, SDS–PAGE alkaline protease zymogram and PAGE acid protease zymogram were carried out to identify active isoforms during larviculture. Alkaline protease and chymotrypsin were present at the moment of hatching, increased gradually reaching the maximum values at 35 dah. Trypsin and leucine aminopeptidase activities were low from hatching, increasing gradually as larvae grew. Alkaline protease zymogram showed four zymogens, which appears at different days, remaining present until the end of the larviculture (95.2 kDa at 11 dah, 26.4 kDa at 9 dah, 21.4 kDa at 3 dah, and 23.3 kDa at hatching). Pepsin activity was present at day 7 after hatching and increased progressively until the end of the larviculture. Acid protease zymogram only showed one zymogen (0.65 rf), which appear at 6 dah. Lipase was high at the time of hatching and increased until 15 dah, after which decreased gradually. Amylase was high from the beginning and until 15 dah and then decreased rapidly to almost nothing onward. Alkaline and acid phosphatases presented a high activity at the egg stage, fell slightly during the first feeding and increased again from 20 to 30 dah. Results obtained in this study show that larvae can be fed artificial diets starting on day 10 after hatching.