尼罗罗非鱼traf6基因表达谱及信号转导

肿瘤坏死因子受体相关因子6(TRAF6)是一种重要的接头蛋白,在Toll样受体/白细胞介素-1受体(TLR/IL-1R)超家族所触发的信号通路中起重要作用,与先天性免疫密切相关.文章研究了尼罗罗非鱼(Oreo-chromis niloticus)traf6的表达模式和初步的功能.在健康鱼中,traf6转录本广泛表达于所...     

Expression profiles of growth-related genes in two Nile tilapia strains and their crossbred provide insights into introgressive breeding effects

The Nile tilapia (Oreochromis niloticus) is a prominent farmed fish in aquaculture worldwide. Crossbreeding has recently been carried out between the Red-Stirling and the wt Chitralada strains of Nile tilapia, producing a heterotic hybrid (7/8 Chitralada and 1/8 Red-Stirling) that combines the superior growth performance of the Chitralada with the reddish coloration of the Red-Stirling strain. While classical selective breeding and crossbreeding strategies are well known, the molecular mechanisms underlying the phenotypic expression of economically advantageous traits in tilapia remain largely unknown. Molecular investigations have shown that variable expression of growth hormone (gh), insulin-like growth factors (igf1 and 2) and somatolactin (smtla) – components of the growth hormone/insulin-like growth factor (GH/IGF) axis – and myostatin (mstn) genes can affect traits of economic relevance in farmed animals. The aim of this study was to assess and compare the gene expression signature among Chitralada, Red-Stirling and their backcross hybrid in order to gain insights into the effects of introgressive breeding in modulation of the GH/IGF axis. Gene expression analyses in distinct tissues showed that most genes of the GH/IGF axis were up-regulated and mstn was down-regulated in backcross animals in comparison with Red-Stirling and Chitralada animals. These gene expression profiles revealed that backcross animals displayed a distinctive expression signature, which attests to the effectiveness of the introgressive breeding technique. Our findings also suggest that the GH/IGF axis and mstn genes might be candidate markers for fish performance and prove useful within genetic improvement programs aimed at the production of superior-quality tilapia strains using introgressive breeding.     

Establishment and growth responses of Nile tilapia embryonic stem‐like cell lines under feeder‐free condition

Embryonic stem (ES) cells provide an invaluable tool for molecular analysis of vertebrate development and a bridge linking genomic manipulations in vitro and functional analysis of target genes in vivo. Work towards fish ES cells so far has focused on zebrafish (Danio renio) and medaka (Oryzias latipes). Here we describe the derivation, pluripotency, differentiation and growth responses of ES cell lines from Nile tilapia (Oreochromis niloticus), a world‐wide commercial farmed fish. These cell lines, designated as TES1‐3, were initiated from blastomeres of Nile tilapia middle blastula embryos (MBE). One representative line, TES1, showed stable growth and phenotypic characteristics of ES cells over 200 days of culture with more than 59 passages under feeder‐free conditions. They exhibited high alkaline phosphatase activity and expression of pluripotency genes including pou5f3 (the pou5f1/oct4 homologue), sox2, myc and klf4. In suspension culture together with retinoic acid treatment, TES1 cells formed embryoid bodies, which exhibited expression profile of differentiation genes characteristics of all three germ cell layers. Notably, PKH26‐labeled TES1 cells introduced into Nile tilapia MBE could contribute to body compartment development and led to hatched chimera formation with an efficacy of 13%. These results suggest that TES1 cells have pluripotency and differentiation potential in vitro and in vivo. In the conditioned DMEM, all of the supplements including the fetal bovine serum, fish embryonic extract, fish serum, basic fibroblast growth factor and non‐protein supplement combination 5N were mitogenic for TES1 cell growth. This study will promote ES‐based biotechnology in commercial fish.     

Foraging in Non-Native Environments: Comparison of Nile Tilapia and Three Co-Occurring Native Centrarchids in Invaded Coastal Mississippi Watersheds

We examined the diet of the alien Nile tilapia and bluegill, redear sunfish, and largemouth bass over a two-year period in coastal Mississippi. Nile tilapia diet was visually separated from the three natives based on group-average linkage cluster analysis. Sequential two-way nested analysis of similarities indicted there was no season effect (Global R = 0.026, P = 24.3%), but there was a moderate size class effect (Global R = 0.457, P = 0.1%) and a strong species effect (Global R = 0.876, P = 0.1%). Pairwise tests indicated species fed on different components of and locations within the environment, with bluegill, redear sunfish and largemouth bass (all R ≤ 0.683, P = 0.1%) having the most similar dietary components and Nile tilapia (all R ≥ 0.953, P = 0.1%) having the most distinct. Multivariate dispersion indicated that largemouth bass (1.425) and bluegill (1.394) had the most diverse diets compared to redear sunfish (0.906) and Nile tilapia (0.918). Similarities of percentages indicated that diets were separated based on prey: bluegill and redear sunfish consumed chironomids and insects; largemouth bass consumed fish and insects; and Nile tilapia fed most often on sediment resources such as nematodes, rotifers, bryozoans and hydrozoans. Nile tilapia had the highest frequency of mud, sand and detritus in their stomachs, suggesting they fed directly on bottom sediments. These data and the fact that Nile tilapia has a 1.3–7.6 times longer intestine on average than its body length, support our contention that this alien species feeds at the base of the food web and is well adapted to survive and proliferate in non-native environments.     

Nonadditive expression of lipid metabolism pathway-related genes in intestine of hybrids of Nile tilapia females (Oreochromis niloticus) and blue tilapia males (Oreochromis aureus)

Nonadditive expression contributes to heterosis in hybrids. In this study, the expression profiles of twelve lipid metabolism pathway-related genes were investigated in the intestine of Nile tilapia (Oreochromis niloticus) ♀?×?blue tilapia (Oreochromis aureus) ♂ hybrid. The expression of genes from the hybrid were assigned to nonadditive and additive expression pattern groups and compared with expression patterns from Nile tilapia and blue tilapia. In the intestine of the hybrid, apoA4B was expressed at intermediate levels, but apoB and MTP were assigned to ELD-B and ELD-N categories, respectively. The LPL and LRP1 showed transgressive up-regulation in the hybrid, but LDLR was assigned to the ELD-B category. For fatty acid uptake related genes, only FABP11a was categorized as nonadditive expression with transgressive up-regulation, while CD36 and FABP3 were categorized as additive expression in the intestine of the hybrid. Two genes in triacylglycerol metabolism, namely, FAS and DGAT2, showed transgressive up-regulation in the hybrid. Most of the genes analyzed in the present study showed nonadditive expression (8 in 12), and five genes showed transgressive up-regulation. These results indicated that the stimulation of lipid metabolism in the hybrid compared to that of its parents. The hyperactive expression of these genes in the hybrid may be associated with the growth and lipid usage vigor.

     

Microsatellites from genes show polymorphism in two related Oreochromis species

Six microsatellites were detected in the IGF‐II, GH2, prolactin I and insulin genes of Mozambique tilapia (Oreochromis mossambicus) and Nile tilapia (Oreochromis niloticus) by screening DNA sequences deposited to GenBank. Genotyping of 24 individuals from each species revealed that all six microsatellites were polymorphic in both species. In Mozambique tilapia the number of alleles ranged from 3 to 17 (average 9.8), whereas in Nile tilapia the range was 4–21 (average 10.5). The range of expected heterozygosity values were 0.44–0.95 (average 0.79) in Mozambique tilapia, and 0.52–0.96 (average 0.73) in Nile tilapia.     

Molecular characterization and immunohistochemical localization of tilapia piscidin 3 in response to Aeromonas hydrophila infection in Nile tilapia

The antimicrobial activity of tilapia piscidin 3 (TP3) was determined in vitro against a locally isolated Aeromonas hydrophila. A 388 bp fragment was amplified from the TP3 cDNA and sequenced. The coding sequence (CDS) of TP3 was estimated to be 231 bp codes for 76 amino acids long and stop codon. In silico analysis was performed to detect both the signal peptide and the prodomain cleavage sites to follow the amino acids number 22 and 70, respectively. Based on this, a peptide 23 amino acids long with a remarkably high computed antimicrobial probability was synthesized and used in the subsequent experiments. The antimicrobial activity of TP3 was determined with minimum inhibitory concentration (MIC) and minim um bactericidal concentration (MBC) methods. TP3 exhibited relatively weak antimicrobial activities against the tested bacteria. A challenge experiment was then performed in Nile tilapia with low and high doses of A. hydrophila, followed by timely recognition; after 3, 6, 24 h, and 7 days of the specific TP3 gene expression, immunohistochemical localization was also performed. Histopathological examination revealed provoked inflammatory responses and congestion in the same organs of TP3 expression. Immunohistochemical localization showed that A. hydrophila induced tilapia fish to express TP3 after 24 h within the gills, intestine, hepatopancreas, spleen, and posterior kidney. In quantitative real time (RT)‐polymerase chain reaction analysis, the high dose showed higher mRNA expression levels than the low dose, and its expression levels increased in the A. hydrophila‐infected fish. It was therefore concluded that TP3 plays an essential role in fish immunity.     

Comparative age and growth of Nile tilapia (<Emphasis Type="Italic">Oreochromis niloticus</Emphasis> L.) in lakes Nabugabo and Wamala,Uganda

Age and growth of Nile tilapia (Oreochromis niloticus) from Lake Nabugabo and Lake Wamala, Uganda, were determined using cross-sectioned sagittal otoliths. Marginal-increment and edge analyses of Nile tilapia otoliths from Lake Nabugabo indicated formation of two annuli per 12-month period. Opaque zones associated with faster growth were observed between April and June and between September and December, coincident with the two rainy seasons of the year. Within both lakes, males were larger at age than females. Nile tilapia from Lake Nabugabo, however, had faster growth rates than Nile tilapia from Lake Wamala, and fish >3 years old from Lake Nabugabo were larger at age than those from Lake Wamala. Ages ranged from 0 to 8.0 years for Nile tilapia from Lake Nabugabo, and from 0.5 to 6.5 years for tilapia from Lake Wamala. Differences in the patterns of growth in Nile tilapia between lakes may reflect, at least in part, the relatively energy-rich omnivorous diet of Nile tilapia in Lake Nabugabo versus a phytoplanktivorous diet in Lake Wamala. Diet differences of Nile tilapia between the two lakes are ascribed to trophic changes in the lakes due to the introduction of Nile perch (Lates niloticus) into Lake Nabugabo but not Lake Wamala. Alternatively, the greater exploitation of Nile tilapia in Lake Nabugabo may have resulted in increased growth rates, whereas Nile tilapia in Lake Wamala may be subject to slower, density-dependent growth. Handling editor: J. Cambray     

Evaluation of the genetic diversity of microsatellite markers among four strains of Oreochromis niloticus

Different strains of Nile tilapia can be found worldwide. To successfully use them in breeding programs, they must be genetically characterized. In this study, four strains of Nile tilapia – UFLA, GIFT, Chitralada and Red‐Stirling – were genetically characterized using 10 noncoding microsatellite loci and two microsatellites located in the promoter and first intron of the growth hormone gene (GH). The two microsatellites in the GH gene were identified at positions ?693 to ?679 in the promoter [motif (ATTCT)₈] and in intron 1 at positions +140 to +168 [motif (CTGT)₇]. Genetic diversity was measured as mean numbers of alleles and expected heterozygosity, which were 4 and 0.60 (GIFT), 3.5 and 0.71 (UFLA), 4.5 and 0.57 (Chitralada) and 2.5 and 0.42 (Red‐Stirling) respectively. Genetic differentiation was estimated both separately and in combination for noncoding and GH microsatellites markers using Jost's D_EST index. The UFLA and GIFT strains were the least genetically divergent (D_EST = 0.10), and Chitralada and Red‐Stirling were the most (D_EST = 0.58). The UFLA strain was genetically characterized for the first time and, because of its unique origin and genetic distinctness, may prove to be an important resource for genetic improvement of Nile tilapia. This study shows that polymorphisms found in coding gene regions might be useful for assessing genetic differentiation among strains.     

Endosulfan increases seric interleukin-2 like (IL-2L) factor and immunoglobulin M (IgM) of Nile tilapia (Oreochromis niloticus) challenged with Aeromona hydrophila

Endosulfan is a persistent organochlorine insecticide which is extremely toxic to fish. It is known to induce immunological alterations in juvenile Nile tilapia (Oreochromis niloticus) such as increases in phagocytic activity and reactive oxygen species production of spleen macrophages. The purpose of the present study was to demonstrate the effects of acute exposure to a sublethal concentration of endosulfan (7 ppb, 96 h) on parameters of the adaptive humoral immune response of the aforementioned aquatic organism. The effect of endosulfan on the capacity of immune cells to produce interleukin-2 like (IL-2L) factor and immunoglobulin M (IgM) in response to a challenge with ½ LD50 of the infectious bacteria Aeromonas hydrophila was evaluated.Experimental results indicate that short, sublethal, endosulfan exposure triggers a succession of events beginning with non-specific activation of macrophages followed by an exacerbated synthesis of the IL-2L factor by activated B cells. This leads to significantly increased secretion of IgM and could in turn facilitate autoantibody production and the development of autoimmune pathologies.     

Cyto-histopathological and protein polymorphism alterations in five populations of Nile tilapia (Oreochromis niloticus) as biomonitor for water heavy metal pollution

Cytological, histopathological and sodium dodecyl sulphate polyacrylamide gel analyses were carried out on five populations of common Nile tilapia fish (Oreochromis niloticus) occurring in five sites – River Nile (reference site), Bahr Yusef canal, Ibrahimia canal, Irrigation drain and El Moheet drain of El Minia Province, Egypt – to evaluate the usability of Nile tilapia as a biomonitor for water heavy metal contaminants. Water surface samples were collected from the five sites, and lead (Pb) concentration was shown to surpass the limits defined by WHO. Ni and Cd levels were shown to be elevated in Ibrahimia canal samples. Moreover, the concentration of heavy metals in fish muscles collected from Bahr Yusef canal and El Moheet drain was the highest in comparison with those of the other water sites. Cytological examinations of blood smears showed not only a significant percentage of micronuclei in Irrigation drain population but also a significant percentage of binucleated cells in Ibrahimia canal and El Moheet drain populations. In addition, pathological alteration was observed in blood cells, especially in samples collected from Irrigation drain and El Moheet drain. Histopathological changes were strongly observed in the liver and the kidneys of El Moheet and Irrigation drain population. Moreover, total protein band pattern profiles showed extra bands in both Ibrahimia canal and Irrigation drain more than that recorded for the River Nile population. In conclusion, cyto-histopathological and total protein band pattern results confirmed that O. niloticus responds sensitively to the excess of heavy metals present in the water.