Expression and activity of trypsin and pepsin during larval development of the spotted rose snapper Lutjanus guttatus

The present study aimed to describe and understand the development of the digestive system in spotted rose snapper (Lutjanus guttatus) larvae from hatching to 40 days post-hatch (dph). The mouth opened between 2 and 3 dph, at that moment the digestive tract was barely differentiated into the anterior and posterior intestine, although the liver and pancreas were already present. Gastric glands were observed until 20 dph, followed by the differentiation of the stomach between 20 and 25 dph. Trypsinogen expression and trypsin activity were detected at hatching, increasing concomitantly to larval development and the change in the type of food. Maximum levels of trypsinogen expression were observed at 25 dph, when animals were fed with Artemia nauplii, and maximum trypsin activity was detected at 35 dph, when larvae were fed with an artificial diet. On the other hand, pepsinogen gene expression was detected at 18 dph, two days before pepsin enzymatic activity and appearance of gastric glands. Maximum pepsin activity was also observed at 35 dph. These results suggest that in this species weaning could be initiated at an earlier age than is currently practiced (between 28 and 30 dph), since larvae of spotted rose snapper develop a functional stomach between days 20 and 25 post-hatch.     

Vibrios of the spotted rose snapper Lutjanus guttatus Steindachner, 1869 from northwestern Mexico

AIM: To characterize and identify vibrios present in wild and cultured juvenile snappers (Lutjanus guttatus) in northwestern Mexico. METHODS AND RESULTS: Spotted rose snapper juveniles were collected at four localities in northwestern Mexico. Bacteria were isolated from external lesions, kidney, liver, and spleen from cultured and wild caught organisms. In total, 280 isolates were obtained and fingerprinted with rep-PCR (GTG5). Nearly 93.2% of the strains belonged to the Vibrionaceae family. Species and genera identified were Photobacterium damselae subsp. damselae (76 strains), Photobacterium leiognathi (13), Vibrio sp. (24), Vibrio alginolyticus (12), Vibrio campbellii (19), Vibrio fortis (9), Vibrio harveyi (49), Vibrio ichthyoenteri (4), Vibrio mediterranei (4), Vibrio parahaemolyticus (1), Vibrio ponticus (2), Vibrio rotiferianus (22), and four potential new species. Conclusions: A wide diversity of vibrios was found in the external lesions and internal organs of both wild and cultured spotted rose snapper juveniles. In total, 12 species of vibrios and four potential new species were identified. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first study on the vibrios present in the spotted rose snapper and therefore might serve as a basis for future studies and diagnosis.     

Allometric growth and larval development in Pacific red snapper Lutjanus peru under culture conditions

Allometric growth is a common feature during fish larval development. It has been proposed as a growth strategy to prioritize the development of body segments related to primordial functions like feeding and swimming to increase the probability of survival during this critical period. In the present study we evaluated the allometric growth patterns of body segments associated to swimming and feeding during the larval stages of Pacific red snapper Lutjanus peru. The larvae were kept under intensive culture conditions and sampled every day from hatching until day 33 after hatching. Each larva was classified according to its developmental stage into yolk-sac larva, preflexion larva, flexion larva or postflexion larva, measured and the allometric growth coefficient of different body segments was evaluated using the potential model. Based on the results we can infer the presence of different ontogenetic priorities during the first developmental stages associated with vital functions like swimming during the yolk-sac stage [total length (TL) interval = 2.27–3.005 mm] and feeding during the preflexion stage (TL interval = 3.007–5.60 mm) by promoting the accelerated growth of tail (post anal) and head, respectively. In the flexion stage (TL interval = 5.61–7.62 mm) a change in growth coefficients of most body segments compared to the previous stage was detected, suggesting a shift in growth priorities. Finally, in the postflexion stage (TL interval = 7.60–15.48 mm) a clear tendency to isometry in most body segments was observed, suggesting that growth priorities have been fulfilled and the larvae will initiate with the transformation into a juvenile. These results provide a framework of the larval growth of L. peru in culture conditions which can be useful for comparative studies with other species or in aquaculture to evaluate the changes in larval growth due to new conditions or feeding protocols.     

Post-transcriptional silencing of myostatin-1 in the spotted rose snapper (Lutjanus guttatus) promotes muscle hypertrophy

Molecular Biology Reports - Muscle growth is regulated by several factors including the growth differentiation factor 8, known as myostatin, an inhibitor of myocyte differentiation and...     

Hematological characteristics of the spotted rose snapper Lutjanus guttatus (Steindachner, 1869) healthy and naturally infected by dactylogyrid monogeneans

The aim of the present study was to obtain a basic knowledge of the hematology in order to determine changes in blood parameters of the spotted rose snapper Lutjanus guttatus. The morphological features of blood cells were described according to the observations made by light microscopy of Wright-Giemsa-stained blood films. The reference intervals and the mean value were determined for each hematological parameter evaluated in healthy fish and data were compared to those of naturally infected, with dactylogyrid monogeneans fish.Infected fish showed a prevalence of 100% and a mean intensity of 246.6 parasites per fish. Mean values of HCT, WBC, thrombocytes percentage and eosinophils percentage were significantly higher (P < 0.05) in the infected fish. In addition, lymphocytes percentage and total protein were significantly lower (P < 0.05) in the infected fish compared to healthy fish. Only total WBC count, lymphocytes percentage and eosinophils percentage in infected fish were outside reference interval. The hematology of the spotted rose snapper of this study might serve as a basis for future studies and diagnosis. Changes observed in blood parameters in infected fish suggest that the immune system of L. guttatus was affected by the presence of the parasites.     

Effects of feed restriction on growth performance,lipid mobilization,and gene expression in rose spotted snapper (Lutjanus guttatus)

The effects of feed deprivation were evaluated for 1 week and 2 weeks in Lutjanus guttatus juveniles. A significant reduction in body mass was observed in both feed deprivation schemes, as well as in hepatosomatic, viscerosomatic and mesenteric fat indexes. The composition of fasted fish was characterized by a decrease in lipid content; the liver displayed an intense reduction of lipid reserves in both fasted groups, and increased expression of the lysosomal acid lipase. 1 week after re-feeding, both experimental groups showed an increase in specific growth rate, feed intake and feed conversion ratio. A recovery in hepatic lipid reserves was observed, and the expression of the lysosomal acid lipase decreased, although lipid content in both groups was still significantly lower than in control groups. Hepatic expression of the growth hormone receptor decreased after fasting, and remained low even after the fish were fed, whereas the expression of insulin-like growth factor 1 and 2 increased after fasting and decreased in both groups when fish were fed again. Altogether, these results showed a partial compensatory growth response in L. guttatus juveniles after fasting, with enhanced growth rates and improved feed efficiency. Fish used stored lipid reserves as the main energy source, and the expression of growth-related and lipid mobilization marker genes in the liver showed similar patterns in both fasting schemes. Based on the results, we suggest as much as 1-week fasting intervals during grow-out programmes to reduce visceral fat and increase growth rate in this species.

     

Parasite communities of the spotted rose snapper Lutjanus guttatus (Perciformes: Lutjanidae) off the Mexican Pacific coasts: Spatial and long-term inter-annual variations

A total of 802 individuals of Lutjanus guttatus (Steindachner, 1869) specimens were collected over a 10-year period (August 2012 to February 2021) from four locations on the south-central Pacific coast of Mexico. Their parasite communities were quantified and analyzed to determine if they experience significant spatial and inter-annual variations.Thirty-two taxa of metazoan parasite were recovered and identified: four species of Digenea, four Monogenea, one Cestoda, two Acanthocephala, seven Nematoda, one Hirudinea, and nine of Crustacea (six Copepoda, and three Isopoda). Species richness was greatest among the digenean group, which represented 25% of the total species recovered, followed by the nematodes (22% of total species). Species richness at the component community level (10 to 20 species) was similar to reported richness in other Lutjanus spp. The component communities and infracommunities exhibited a similar pattern: low species richness and diversity, and dominance by a single species, mainly the monogenean Haliotrematoides guttati. Parasite community structure and species composition varied through the years, as well as between sampling locations. Variations were possibly caused by a combination of biotic and abiotic factors which generated notable changes in the infection levels of several component species. However, the similarity in the parasite species composition was high locally for short-term periods (one or two years). This result, therefore, suggests that parasite communities of L. guttatus may be more predictable locally, but only for short-term periods.     

Haploinsufficiency of myostatin protects against aging‐related declines in muscle function and enhances the longevity of mice

The molecular mechanisms behind aging-related declines in muscle function are not well understood, but the growth factor myostatin (MSTN) appears to play an important role in this process. Additionally, epidemiological studies have identified a positive correlation between skeletal muscle mass and longevity. Given the role of myostatin in regulating muscle size, and the correlation between muscle mass and longevity, we tested the hypotheses that the deficiency of myostatin would protect oldest-old mice (28–30 months old) from an aging-related loss in muscle size and contractility, and would extend the maximum lifespan of mice. We found that MSTN^+/− and MSTN^−/− mice were protected from aging-related declines in muscle mass and contractility. While no differences were detected between MSTN^+/+ and MSTN^−/− mice, MSTN^+/− mice had an approximately 15% increase in maximal lifespan. These results suggest that targeting myostatin may protect against aging-related changes in skeletal muscle and contribute to enhanced longevity.     

A frameshift mutation in the coding region of the myostatin gene (MSTN) affects carcass conformation and fatness in Norwegian White Sheep (Ovis aries)

Mutations in the coding region of the myostatin gene ( MSTN ) are known to cause an increased muscle mass (IMM) phenotype in several mammals, including mice, dogs, cattle and humans. In sheep, a mutation in the 3'-UTR region introducing a microRNA target site has been reported to cause an IMM-like phenotype because of downregulation of translation. Here we report a novel single base deletion in the coding region of the myostatin gene causing an IMM phenotype in Norwegian White Sheep, characterized by a high carcass conformation class and low fat class (EUROP classification system). The deletion disrupts the reading frame from amino acid (aa) position 320, ending in a premature stop codon in aa position 359. In our material, these MSTN mutations segregated in a pattern showing that they reside in two different haplotypes. The phenotypic effect of the single base deletion is more profound than that of the 3'-UTR mutation.     

Structure of the human myostatin precursor and determinants of growth factor latency

Myostatin, a key regulator of muscle mass in vertebrates, is biosynthesised as a latent precursor in muscle and is activated by sequential proteolysis of the pro‐domain. To investigate the molecular mechanism by which pro‐myostatin remains latent, we have determined the structure of unprocessed pro‐myostatin and analysed the properties of the protein in its different forms. Crystal structures and SAXS analyses show that pro‐myostatin adopts an open, V‐shaped structure with a domain‐swapped arrangement. The pro‐mature complex, after cleavage of the furin site, has significantly reduced activity compared with the mature growth factor and persists as a stable complex that is resistant to the natural antagonist follistatin. The latency appears to be conferred by a number of distinct features that collectively stabilise the interaction of the pro‐domains with the mature growth factor, enabling a regulated stepwise activation process, distinct from the prototypical pro‐TGF‐β1. These results provide a basis for understanding the effect of missense mutations in pro‐myostatin and pave the way for the design of novel myostatin inhibitors.     

Characterization and expression of a myosin heavy–chain isoform in juvenile walleye Sander vitreus

In this study, myosin, the major component of myofibrillar protein in the skeletal muscle, was characterized and its expression was monitored during growth in juvenile walleye Sander vitreus. First, the coding region of myosin heavy chain (MyHC) from the fast skeletal muscle of walleye was amplified by long-distance PCR using a full-length cDNA. Phylogenetic analysis was used to determine the evolutionary relationship of this S. vitreus myosin sequence to other vertebrate myosin sequences. Next, it was established that the myosin isoform was most prevalent in the white muscle, compared with the red and cardiac muscle. Myosin expression was monitored over a series of experiments designed to influence growth. Specifically, change in MyHC mRNA was monitored after acute changes in feeding. Fish exposed to a one-week fasting period showed significant decreases in MyHC mRNA levels by the end of the fast. The effect of feeding was also examined more closely over a 24 h period after feeding, but results showed no significant change in myosin expression levels through this time period. Finally, fish with higher growth rates had higher MyHC mRNA and protein expression levels. This study indicates that MyHC mRNA expression is sensitive to the factors that may influence growth in juvenile S. vitreus .     

Morphological and biochemical effects of food deprivation during the early development of Pacific red snapper Lutjanus peru

We report the effects of food deprivation on the early development of Pacific red snapper Lutjanus peru during the first days of development. The point of no return (PNR) was determined using the feeding incidence after a delay in first feeding. The gradual deterioration of the larvae during food deprivation was recorded using morphometric, histological, enzymatic and biochemical analysis. The time to reach the PNR was 120 h after hatching. Morphologically, the total length, muscle height, head length, tail length and pectoral angle showed the biggest reductions and their growth coefficients changed significantly during food deprivation. Histologically, enterocyte height also was reduced significantly. The protein concentration and activities of the digestive enzymes trypsin, cathepsin-like and lipase showed a significant decrease; meanwhile, amylase activity remained constant during food deprivation. The concentration of total essential free amino acids (EFAAs) decreased significantly while that of the nonessential free amino acids (NEFAAs) remain stable during food deprivation. The most abundant EFAAs were lysine, leucine, isoleucine and valine; the most abundant NEFAAs were alanine, glycine and glutamate, suggesting a more prominent role as energy substrates. At the time of the PNR the concentration of almost all the free amino acids showed a significant decrease. Early food deprivation has a significant impact on the morphology and biochemical characteristics of L. peru. These results suggest that initial feeding of L. peru should begin within 3 days of yolk sac depletion to avoid the PNR. Further studies are necessary to confirm and validate the characters identified in this study as biomarkers of starvation under culture conditions and evaluate their possible utility in ichthyoplankton surveys.     

Cloning and expression of gene encoding the thermostable direct hemolysin from Vibrio alginolyticus strain HY9901, the causative agent of vibriosis of crimson snapper (Lutjanus erythopterus)

AIMS: The main aims of this study were to clone and express complete open reading frame (ORF) of thermostable direct haemolysin gene (tdh) from Vibrio alginolyticus strain HY9901 in Escherichia coli, and further evaluate the virulence of expressed TDH on mouse and crimson snapper. METHODS AND RESULTS: A 410 bp internal fragment of the tdh gene was amplified by touchdown PCR with designed primers. Then its unknown flanking sequences of the 5'- and 3'-ends were finally characterized by inverse PCR and nested PCR. Sequence analysis showed that the tdh gene contain 570 bp ORF which encoded 189 amino acids. The deduced amino acid sequence of the ORF was in significant homology with several Vibrio TDH. The product that the tdh gene expressed in E. coli was purified by Ni(2+)-IDA Sepharose affinity column. The activity of purified TDH was 4651 U mg(-1) protein by hide powder azure digestion. The lethal toxicity test showed that LD(50) values of the purified TDH were 5.68 and 8.34 microg TDH g(-1) body weight for mouse and crimson snapper, respectively. CONCLUSIONS: The complete ORF of tdh gene was obtained by touchdown PCR, inverse PCR and nested PCR. The ORF was perfectly expressed in E. coli. The activity and toxicity assays showed that the N-terminal signal peptide was essential to autocatalyse and fold correctly to obtain the activity and toxicity in the purified TDH. The Native-PAGE analysis showed that the activated tdh gene expressed in E. coli was a dimer with two identical subunits. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrates that the expressed activated TDH can produce the toxicity protein determined on mouse and fish, which will lead to better understandings of the identifying virulence factor that could be considered as a candidate antigen for vaccine and a diagnostic tool for vibriosis. Its use as an immunizing antigen might prevent the ability of V. alginolyticus to infect the marine aquatic animals, as a complementary measure to tick control and appropriate management in countries affected by vibriosis.     

Expression profiles of sugarcane under drought conditions: Variation in gene regulation

Drought is a major factor in decreased sugarcane productivity because of the resulting morphophysiological effects that it causes. Gene expression studies that have examined the influence of water stress in sugarcane have yielded divergent results, indicating the absence of a fixed pattern of changes in gene expression. In this work, we investigated the expression profiles of 12 genes in the leaves of a drought-tolerant genotype (RB72910) of sugarcane and compared the results with those of other studies. The genotype was subjected to 80–100% water availability (control condition) and 0–20% water availability (simulated drought). To analyze the physiological status, the SPAD index, Fv/Fm ratio, net photosynthesis (A), stomatal conductance (g _s) and stomatal transpiration (E) were measured. Total RNA was extracted from leaves and the expression of SAMDC, ZmPIP2-1 protein, ZmTIP4-2 protein, WIP protein, LTP protein, histone H3, DNAj, ferredoxin I, β-tubulin, photosystem I, gene 1 and gene 2 was analyzed by quantitative real-time PCR (RT-PCR). Important differences in the expression profiles of these genes were observed when compared with other genotypes, suggesting that complex defense mechanisms are activated in response to water stress. However, there was no recognizable pattern for the changes in expression of the different proteins associated with tolerance to drought stress.     

Expression of genetically distinct superoxide dismutases in the maize seedling during development

Immunoassays for the cytosolic and mitochondrial superoxide dismutases (SOD) of maize were developed and used to study the expression of these proteins in the maize seedling. The genetically distinct proteins, SOD-3 and SOD-4, are preferentially expressed in the scutellum, comprising approximately 1% of the total water-soluble protein of that tissue. SOD-2, SOD-3, and SOD-4 are synthesized in the scutellum during early sporophytic development, probably on cytosolic ribosomes. Two-dimensional gel electrophoresis of crude scutellar extracts indicates that significant changes occur in the protein composition of the maize scutellum following seed imbibition. Using the immunoassays, a maize line exhibiting a significant reduction in cyanide-sensitive SOD protein was identified.