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In this study, a pyruvate carboxylase gene (PYC1) from a marine fungus Penicillium rubens I607 was cloned and characterized. ORF of the gene (accession number: KM397349.1) had 3534 bp encoding 1177 amino acids with a molecular weight of 127.531 kDa and a PI of 6.20. The promoter of the gene was located at −1200 bp and contained a TATAA box, several CAAT boxes and a sequence 5′-SYGGRG-3′. The PYC1 deduced from the gene had no signal peptide, was a homotetramer (α4), and had the four functional domains. After expression of the PYC1 gene from the marine fungus in the marine-derived yeast Yarrowia lipolytica SWJ-1b, the transformant PR32 obtained had much higher specific pyruvate carboxylase activity (0.53 U/mg) than Y. lipolytica SWJ-1b (0.07 U/mg), and the PYC1 gene expression (133.8 %) and citric acid production (70.2 g/l) by the transformant PR32 were also greatly enhanced compared to those (100 % and 27.3 g/l) by Y. lipolytica SWJ-1b. When glucose concentration in the medium was 60.0 g/l, citric acid (CA) concentration formed by the transformant PR32 was 36.1 g/l, leading to conversion of 62.1 % of glucose into CA. During a 10-l fed-batch fermentation, the final concentration of CA was 111.1 ± 1.3 g/l, the yield was 0.93 g/g, the productivity was 0.46 g/l/h, and only 1.72 g/l reducing sugar was left in the fermented medium within 240 h. HPLC analysis showed that most of the fermentation products were CA. However, minor malic acid and other unknown products also existed in the culture.

Biotechnological research on marine organisms, such as ex situ or in situ aquaculture and in vitro cell culture, is being conducted to produce bioactive metabolites for biomedical and industrial uses. The Caribbean marine sponge Discodermia dissoluta is the source of (+)-discodermolide, a potent antitumoural polyketide that has reached clinical trials. This sponge usually lives at depths greater than 30 m, but at Santa Marta (Colombia) there is a shallower population, which has made it logistically possible to investigate for the first time, on ways to supply discodermolide. We thus performed in situ, 6-month fragment culture trials to assess the performance of this sponge in terms of growth and additional discodermolide production and studied possible factors that influence the variability of discodermolide concentrations in the wild. Sponge fragments cultured in soft mesh bags suspended from horizontal lines showed high survivorship (93 %), moderate growth (28 % increase in volume) and an overall rise (33 %) in the discodermolide concentration, equivalent to average additional production of 8 μg of compound per millilitre of sponge. The concentration of discodermolide in wild sponges ranged from 8 to 40 μg mL−1. Locality was the only factor related to discodermolide variation in the wild, and there were greater concentrations in peripheral vs. basal portions of the sponge, and in clean vs. fouled individuals. As natural growth and regeneration rates can be higher than culture growth rates, there is room for improving techniques to sustainably produce discodermolide.

Chitooligosaccharides (COS) has many biological activities, such as antitumor activity and hepatoprotective effect. Herein, we investigated the protective effect of COS against hydrogen peroxide (H2O2)-induced oxidative stress on human embryonic hepatocytes (L02 cells) and its scavenging activity against the 1,1-diphenyl-2-picrylhydrazyl radical in vitro. The results showed that the lost cell viability induced by H2O2 was markedly restored after 24 h pre-incubation with COS (0.1–0.4 mg/ml). This rescue effect could be related to the antioxidant property of COS, in which we showed that the radical scavenging activity of COS reached 80% at concentration of 2 mg/ml. In addition, COS could prevent cell apoptosis induced by H2O2, as shown by the inhibition of the cleavage of poly (adenosine diphosphate-ribose) polymerase and increased expression of the anti-apoptotic protein Bcl-xL. Furthermore, we have utilized confocal laser microscopy to observe cellular uptake of COS, an important step for COS to exert its effects on target cells. Taken together, our findings suggested that COS could effectively protect L02 cells against oxidative stress, which might be useful in clinical setting during the treatment of oxidative stress-related liver damages.

We describe a strategy that identifies molecular biomarkers and links the study of abiotic stress to evolutionary history. By utilizing the moon jellyfish Aurelia spp. as a model, we identified genes differentially regulated in response to the chemical stressor tributyltin by means of complementary DNA subtraction analyses. Expression of 3 out of 25 identified candidate genes, one oxidative stress gene, one heat shock (hsp70) gene, and one GTP-binding gene, was quantified under laboratory conditions and in field tests using semiquantitative reverse transcriptase polymerase chain reaction. Differential expression patterns were found following exposure to tributyltin and temperature treatments. The findings suggest that the identified genes are involved in response to chemical as well as heat- induced stress and may serve as biomarkers for monitoring marine habitats. Gene regulatory patterns combined with phylogenetic inferences of the hsp70 gene support a possible role of ecologically driven divergence within the genus Aurelia. We show that added information on genetic variability can raise the predictive power of molecular biomarkers in studies of individual stress response.

The toxic flatworm, Planocera multitentaculata, possesses highly concentrated tetrodotoxin (TTX), also known as pufferfish toxin, throughout its life cycle, including the egg and larval stages. Additionally, TTX analogues, 5,6,11-trideoxyTTX and 11-norTTX-6(S)-ol, have also been detected in the flatworm. The high concentration of TTX in the eggs and larvae appears to be for protection against predation, and 11-norTTX-6(S)-ol in the pharyngeal tissue in the adults is likely used to sedate or kill prey during predation. However, information on the role of 5,6,11-trideoxyTTX, a potential important biosynthetic intermediate of TTX, in the toxic flatworm is lacking. Here, we aimed to determine the region of localization of TTX and its analogues in the flatworm body, understand their pharmacokinetics during maturation, and speculate on their function. Flatworm specimens in four stages of maturity, namely juvenile, mating, spawning, and late spawning, were subjected to LC–MS/MS analysis, using the pharyngeal tissue, oocytes in seminal receptacle, sperm, and tissue from 12 other sites. Although TTX was consistently high in the pharyngeal tissue throughout maturation, it was extremely high in the oocytes during the spawning period. Meanwhile, 5,6,11-trideoxyTTX was almost undetectable in the pharyngeal part throughout the maturation but was very abundant in the oocytes during spawning. 11-norTTX-6(S)-ol consistently localized in the pharyngeal tissue. Although the localization of TTX and its analogues was approximately consistent with the MS imaging data, TTX and 11-norTTX-6(S)-ol were found to be highly localized in the parenchyma surrounding the pharynx, which suggests the parenchyma is involved in the accumulation and production of TTXs.

With the draft sequence of the human genome available and an increasing number of organisms being sequenced, attention is becoming focused on sequence interpretation and functional analysis. Comparative genomics will play an important role in evaluating these data. At the molecular level, roles for uncharacterized proteins can be hypothesized by identifying conserved protein domains and putative noncoding regulatory elements can be defined from direct sequence comparisons of evolutionarily distant organisms. At a higher level, questions, such as the importance of gene order positioning, conservation of linkage, and genome evolution, can begin to be answered by collecting map data from different organisms. This minireview, centering on Fugu regions sharing synteny with human chromosomes 11p, 20q, and 6p21.3, details some of the ways in which the Japanese pufferfish can contribute to the study of comparative genomics and evaluation of sequence data from the genome programs.

The principal fatty acids from the lipid profiles of two autochthonous dinoflagellates (Alexandrium minutum and Karlodinium veneficum) and one raphidophyte (Heterosigma akashiwo) maintained in bubble column photobioreactors under outdoor culture conditions are described for the first time. The biomass production, lipid content and lipid productivity of these three species were determined and the results compared to those obtained when the strains were cultured indoors. Under the latter condition, the biotic values did not significantly differ among species, whereas under outdoor conditions, differences in both duplication time and fatty acids content were observed. Specifically, A. minutum had higher biomass productivity (0.35 g·L−1 day−1), lipid productivity (80.7 mg lipid·L−1 day−1) and lipid concentration (252 mg lipid·L−1) at harvest time (stationary phase) in outdoor conditions. In all three strains, the growth rate and physiological response to the light and temperature fluctuations of outdoor conditions greatly impacted the production parameters. Nonetheless, the species could be successfully grown in an outdoor photobioreactor and were of sufficient robustness to enable the establishment of long-term cultures yielding consistent biomass and lipid production.

To identify muscle-related protein isoforms expressed in the white muscle of the mandarin fish Siniperca chuatsi, we analyzed 5,063 high-quality expressed sequence tags (ESTs) from white muscle cDNA library and predicted the integrity of the clusters annotated to these genes and the physiochemical properties of the putative polypeptides with full length. Up to about 33% of total ESTs were annotated to muscle-related proteins: myosin, actin, tropomyosin/troponin complex, parvalbumin, and Sarcoplasmic/endoplasmic reticulum calcium ATPase (SERCa). Thirty-two isoforms were identified and more than one isoform existed in each of these proteins. Among these isoforms, 14 putative polypeptides were with full length. In addition, about 2% of total ESTs were significantly homologous to “glue” molecules such as alpha-actinins, myosin-binding proteins, myomesin, tropomodulin, cofilin, profilin, twinfilins, coronin-1, and nebulin, which were required for the integrity and maintenance of the muscle sarcomere. The results demonstrated that multiple isoforms of major muscle-related proteins were expressed in S. chuatsi white muscle. The analysis on these isoforms and other proteins sequences will greatly aid our systematic understanding of the high flexibility of mandarin fish white muscle at molecular level and expand the utility of fish systems as models for the muscle genetic control and function.

The acclimation of a nitrifying biofilter is a crucial and time-consuming task for setting up a recirculating aquaculture system (RAS). Gaining a better understanding of the dynamics of the microbial community during the acclimation period in the system could be useful for the development of mature nitrifying biofilters. In this study, high-throughput DNA sequencing was applied to monitor the microbial communities on a biofilter during the acclimation period (7 weeks) in high (100 mg N/L) and low (5 mg N/L) total ammonia nitrogen (TAN) treatments. Both treatments were successful for developing a mature nitrifying biofilter, dominated by Proteobacteria, Bacteroidetes, and Nitrospirae. Complete nitrification was found after 7 days of biofilter acclimation as indicated by decreasing TAN concentration, increasing nitrate concentration, and high abundances of the nitrifying bacteria, Nitrosomonadaceae and Nitrospiraceae. The beta diversity analysis of microbial communities showed different clustering of the samples between high and low TAN treatment groups. A greater abundance of nitrifying bacteria was found in the high TAN treatments (27–51%) than in the low TAN treatment (15–29%). The bacterial diversity in biofilters acclimated at high TAN concentration (Shannon’s index 5.40–6.15) were lower than those found at low TAN treatment levels (Shannon’s index 6.40–7.01). The higher diversity in biofilters acclimated at low TAN concentrations, consisting of Planctomycetes and Archaea, might benefit the nutrient recycling in the system. Although nitrification activity was observed from the first week of the acclimation period, the acclimation period should be taken as at least 6 weeks for full development of nitrifying biofilm. Moreover, the reduction of potentially pathogenic Vibrio on biofilters was found at that period.