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For the commercial culture of the three‐spot cichlid (Amphilophus trimaculatus), it is important to have available a set of protein and lipid ingredients suitable for feed manufacturing, which allow to optimize the culture at affordable costs. The in vitro digestibility was evaluated with pH‐stat technique, using stomach and intestine multi‐enzymatic extracts from A. trimaculatus juveniles. The digestion of protein ingredients was taken place both in acid and in alkaline conditions, quantifying the total of free amino acids at the end of the hydrolysis, whereas the digestion of lipid ingredients was taken place only in alkaline conditions. The digestive enzymes of this species showed a better affinity to raw materials of animal origin such as beef meal, fish meal and premium chicken meal, which presented better digestibility. The best lipid ingredient that could be used is the fish oil. However, it is possible to use ingredients from the plant sources, such as soy or cereals, and vegetable oils such as olive, corn or soy oil. It is necessary to complement this study in vitro with the digestibility analysis in vivo, in order to determine the inclusion percentages in feeds and costs and therefore to develop an efficient feed formulation for the aquaculture of A. trimaculatus.

Determination of the main nutritional requirements at different developmental stages is a prerequisite in the formulation of feeds for newly cultured fish species. In the present study, the lipid and protein requirements of larvae of a native Mexican fish, the three-spot cichlid Cichlasoma trimaculatum, were assessed using a two-factor experimental design that considered four protein (35, 40, 45, and 50%) and two lipid levels (16 and 22%) on growth, survival, and digestive enzyme activities. The best growth and feed efficiency results were obtained when larvae were fed diets including 45% protein and 22% lipids. Comprehensive evaluation of the profile of digestive enzymes using multivariate analysis also demonstrated significant differences in nutritional condition generated by varying inclusion of nutrients. Thus, an increase in protein led to an increase in alkaline protease activity and a reduction in leucine aminopeptidase activity, and the reduction of dietary lipid content led to a significant increase in lipase and trypsin enzymatic activities. Based on our results, C. trimaculatum larvae have a high capacity to hydrolyze both nutrients (protein and lipids) for the high digestive enzyme activities and increase their growth, particularly with a diet containing 45% protein and 22% lipids.

The Holostei group occupies a critical phylogenetic position as the sister group of the Teleostei. However, little is known about holostean pituitary anatomy or brain distribution of important reproductive neuropeptides, such as the gonadotropin-inhibitory hormone (GnIH). Thus, the present study set out to characterize the structure of the pituitary and to localize GnIH-immunoreactive cells in the brain of Atractosteus tropicus from the viewpoint of comparative neuroanatomy. Juveniles of both sexes were processed for general histology and immunohistochemistry. Based on the differences in cell organization, morphology, and staining properties, the neurohypophysis and three regions in the adenohypophysis were identified: the rostral and proximal pars distalis (PPD) and the pars intermedia. This last region was found to be innervated by the neurohypophysis. This organization, together with the presence of a saccus vasculosus, resembles the general teleost pituitary organization. A vast number of blood vessels were also recognized between the infundibulum floor of the hypothalamus and the PPD, evidencing the characteristic presence of a median eminence and a portal system. However, this well-developed pituitary portal system resembles that of tetrapods. As regards the immunohistochemical localization of GnIH, we found four GnIH-immunoreactive (GnIH-ir) populations in three hypothalamic nuclei (suprachiasmatic, retrotuberal, and tuberal nuclei) and one in the diencephalon (prethalamic nucleus), as well as a few scattered neurons throughout the olfactory bulbs, the telencephalon, and the intersection between them. GnIH-ir fibers showed a widespread distribution over almost all brain regions, suggesting that GnIH function is not restricted to reproduction only. In conclusion, the present study describes, for the first time, the pituitary of A. tropicus and the neuroanatomical localization of GnIH in a holostean fish that exhibits a similar distribution pattern to that of teleosts and other vertebrates, suggesting a high degree of phylogenetic conservation of this system.

Implementation of massive sequencing techniques in aquaculture has led to increase genomic resources of different species and have generated useful specific markers to improve their management. In this regard, transcriptomic approaches have been widely used to easily obtain gene expression profiles and understand the regulation of molecular pathways of interest in aspects as development and nutrition. The three spot cichlid (Amphilophus trimaculatus) is a native fish from South Mexico, with commercial regional importance and aquaculture potential. However, there is no genomic information available for the species that could allow the analysis in physiological processes of metabolic pathways to contribute or improve its culture. Hence, a comprehensive transcriptome of three different developmental stages: early (3–5 dpf larvae), mid (8–13 dpf larvae) and late (16–25 dpf larvae), were obtained using Illumina Hi-seq 2000 platform. De novo assembly generated 237,164 transcripts grouped in 196,186 genes with an average contig length of 778 bp and an N50 value of 1546 bp. Conceptual translation yielded in 78,883 proteins, of which 35,033 are complete proteins. By analyzing the transcriptomic profile and differential expression among developmental stages, we identified nutritional markers involved in digestive enzyme, metabolic hormone, and organogenesis of the digestive system. Likewise, in order to confirm the transcriptomic results obtained, histological analyzes were performed using hematoxylin-eosin (H&E) staining, finding that the most important morphological changes occur in the mid stage (endo-exptrophic larvae, 8 dpf and exotrophic larvae, 13 dpf), coinciding with the maximum expressions of the candidate genes for nutritional markers.

RFamide peptides are expressed in the early stages of development in most vertebrates. Gonadotrophin-inhibitory hormone (GnIH) belongs to the RFamide family, and its role in reproduction has been widely studied in adult vertebrates, ranging from fish to mammals. Because only 3 reports have evaluated GnIH during development, the present study aimed to characterise the ontogeny of GnIH in the fish model Cichlasoma dimerus. We detected the presence of 2 GnIH-immunoreactive (-IR) cell clusters with spatial and temporal differences. One cluster was observed by 3 days post-hatching (dph) in the nucleus olfacto-retinalis (NOR) and the other in the nucleus posterioris periventricularis by 14 dph. The number of GnIH-IR neurones increased in both nuclei, whereas their size increased only in the NOR from hatchling to juvenile stages. These changes occurred from the moment that larvae started feeding exogenously and during development and differentiation of gonadal primordia. We showed by double-label immunofluorescence that only GnIH-IR neurones in the NOR co-expressed gonadotrophin-releasing hormone 3 associated peptide. In addition, GnIH-IR fibre density increased in all brain regions from 5 dph. GnIH-IR fibres were also detected in the retina, optic tract and optic tectum, suggesting that GnIH acts as a neuromodulator of photoreception and the integration of different sensory modalities. Furthermore, there were GnIH-IR fibres in the pituitary from 14 dph, which were in close association with somatotrophs. Moreover, GnIH-IR fibres were observed in the saccus vasculosus from 30 dph, suggesting a potential role of GnIH in the modulation of its function. Finally, we found that gnih was expressed from 1 dph, and that the pattern of variation of its transcript levels was in accordance with that of cell number. The results of the present study comprise a starting point for the study of new GnIH roles during development. Fil: Di Yorio, María Paula. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Biodiversidad y Biología Experimental y Aplicada. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Biodiversidad y Biología Experimental y Aplicada; Argentina Fil: Sallemi, Julieta Emilse. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Biodiversidad y Biología Experimental y Aplicada. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Biodiversidad y Biología Experimental y Aplicada; Argentina Fil: Toledo Solís, F.J.. Universidad Juárez Autónoma de Tabasco; México Fil: Pérez Sirkin, Daniela Irina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Biodiversidad y Biología Experimental y Aplicada. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Biodiversidad y Biología Experimental y Aplicada; Argentina Fil: Delgadin, Tomás Horacio. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Biodiversidad y Biología Experimental y Aplicada. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Biodiversidad y Biología Experimental y Aplicada; Argentina Fil: Tsutsui, K.. Waseda University; Japón Fil: Vissio, Paula Gabriela. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Biodiversidad y Biología Experimental y Aplicada. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Biodiversidad y Biología Experimental y Aplicada; Argentina

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.