Marine Biotechnology最新文献

The present study aims to characterize and to evaluate the biological effects of a skin dressing manufactured with the organic part of the Chondrilla caribensis marine sponge (called spongin-like collagen (SC)) associated or not to photobiomodulation (PBM) on the skin wound healing of rats. Skin dressings were manufactured with SC and it was characterized using scanning electron microscopy (SEM) and a tensile assay. In order to evaluate its biological effects, an experimental model of cutaneous wounds was surgically performed. Eighteen rats were randomly distributed into three experimental groups: control group (CG): animals with skin wounds but without any treatment; marine collagen dressing group (DG): animals with skin wounds treated with marine collagen dressing; and the marine collagen dressing + PBM group (DPG): animals with skin wounds treated with marine collagen dressing and PBM. Histopathological, histomorphometric, and immunohistochemical evaluations (qualitative and semiquantitative) of COX2, TGFβ, FGF, and VEGF were done. SEM demonstrates that the marine collagen dressing presented pores and interconnected fibers and adequate mechanical strength. Furthermore, in the microscopic analysis, an incomplete reepithelialization and the presence of granulation tissue with inflammatory infiltrate were observed in all experimental groups. In addition, foreign body was identified in the DG and DPG. COX2, TGFβ, FGF, and VEGF immunostaining was observed predominantly in the wound area of all experimental groups, with a statistically significant difference for FGF immunostaining score of DPG in relation to CG. The marine collagen dressing presented adequate physical characteristics and its association with PBM presented favorable biological effects to the skin repair process.

Pneumococcal infections caused by Streptococcus pneumoniae are a leading cause of morbidity and mortality globally, particularly among children. The ability of S. pneumoniae to form enduring biofilms makes treatment inherently difficult, and options are further limited by emerging antibiotic resistance. The discovery of new antibiotics, particularly those with antibiofilm activity, is therefore increasingly important. Antimicrobial proteins and peptides (AMPs) from marine invertebrates are recognised as promising pharmacological leads. This study determined the in vitro antibacterial activity of hemolymph and unique protein fractions from an Australian oyster (Saccostrea glomerata) against multi-drug-resistant S. pneumoniae. We developed a successful method for hemolymph extraction and separation into 16 fractions by preparative HPLC. The strongest activity was observed in fraction 7: at 42 µg/mL protein, this fraction was bactericidal to S. pneumoniae and inhibited biofilm formation. Proteomic analysis showed that fraction 7 contained relatively high abundance of carbonic anhydrase, cofilin, cystatin B–like, and gelsolin-like proteins, while surrounding fractions, which showed lower or no antibacterial activity, contained these proteins in lower abundance or not at all. This work supports traditional medicinal uses of oysters and contributes to further research and development of novel hemolymph/AMP-based treatments for pneumococcal infections.

Seaweed from the genus Ulva (Ulvales, Chlorophyta) has a worldwide distribution and represents a potential biomass source for biotechnological applications. In the present study, we investigated the ulvan polysaccharide-rich fraction (UPRF) isolated from two Ulva species (U. rigida and U. pseudorotundata), naturally occurring on the Spanish Mediterranean coast. Chemical characterization of UPRFs was performed in order to explore the polysaccharides’ composition. Biological assessments of UPRFs were compared by antioxidant activity and in vitro toxicity tests in the human cell lines: HCT-116 (colon cancer), G-361 (malignant melanoma), U-937 (leukemia), and HaCaT cells (immortalized keratinocytes). Chemical analysis revealed that both UPRFs presented rhamnose as the major relative sugar constituent, followed by glucose in U. rigida and xylose in U. pseudorotundata. Both also presented glucuronic acid, galactose, ribose, and mannose as the remaining monosaccharides. Similar antioxidant activity was obtained, where we observed increased activity in response to increased polysaccharide concentrations. Both UPRFs presented moderate toxicity against HCT-116 cell lines and a selectivity index ≥ 3, suggesting a good potential for use in pharmaceutical products.

Vibrio harveyi, a recently discovered pathogenic bacterium isolated from American eels (Anguilla rostrata), poses uncertainties regarding its pathogenesis in American eel and the molecular mechanisms underlying host defense against V. harveyi infection. This study aimed to determine the LD₅₀ of V. harveyi in American eel and assess the bacterial load in the liver, spleen, and kidney post-infection with the LD50 dose. The results showed that the LD₅₀ of V. harveyi via intraperitoneal injection in American eels over a 14d period was determined to be 1.24 × 10³ cfu/g body weight (6.2 × 10⁴ cfu/fish). The peak bacterial load occurred at 36 h post-infection (hpi) in all three organs examined. Histopathology analysis revealed hepatic vein congestion and thrombi, tubular vacuolar degeneration, and splenic bleeding. Moreover, quantitative reverse transcription polymerase chain reaction (qRT-PCR) results indicated significant up or downregulation of 18 host immune- or anti-infection–related genes post 12 to 60 hpi following the infection. Additionally, RNA sequencing (RNA-seq) unveiled 7 hub differentially expressed genes (DEGs) and 11 encoded proteins play crucial roles in the anti-V. harveyi response in American eels. This study firstly represents the comprehensive report on the pathogenicity of V. harveyi to American eels and RNA-seq of host’s response to V. harveyi infection. These findings provide valuable insights into V. harveyi pathogenesis and the strategies employed by the host’s immune system at the transcriptomic level to combat V. harveyi infection.

The role of hepcidins, antimicrobial peptides involved in iron metabolism, immunity, and inflammation, is studied. First, gilthead seabream (Sparus aurata L.) head-kidney leucocytes (HKLs) were incubated with λ-carrageenin to study the expression of hepcidin and iron metabolism-related genes. While the expression of most of the genes studied was upregulated, the expression of ferroportin gene (slc40a) was downregulated. In the second part of the study, seabream specimens were injected intramuscularly with λ-carrageenin or buffer (control). The expression of the same genes was evaluated in the head kidney, liver, and skin at different time points after injection. The expression of Hamp1m, ferritin b, and ferroportin genes (hamp1, fthb, and slc40a) was upregulated in the head kidney of fish from the λ-carrageenin-injected group, while the expression of Hamp2C and Hamp2E genes (hamp2.3 and hamp2.7) was downregulated. In the liver, the expression of hamp1, ferritin a (ftha), slc40a, Hamp2J, and Hamp2D (hamp2.5/6) genes was downregulated in the λ-carrageenin-injected group. In the skin, the expression of hamp1 and (Hamp2A Hamp2C) hamp2.1/3/4 genes was upregulated in the λ-carrageenin-injected group. A bioinformatic analysis was performed to predict the presence of transcription factor binding sites in the promoter region of hepcidins. The primary sequence of hepcidin was conserved among the different mature peptides, although changes in specific amino acid residues were identified. These changes affected the charge, hydrophobicity, and probability of hepcidins being antimicrobial peptides. This study sheds light on the poorly understood roles of hepcidins in fish. The results provide insight into the regulatory mechanisms of inflammation in fish and could contribute to the development of new strategies for treat inflammation in farm animals.

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Reef-building corals are a fundamental pillar of coral reef ecosystems in tropical and subtropical shallow environments. Corals harbor symbiotic dinoflagellates belonging to the family Symbiodiniaceae, commonly known as zooxanthellae. Extensive research has been conducted on this symbiotic relationship, yet the fundamental information about the distribution and localization of Symbiodiniaceae cells in corals is still limited. This information is crucial to understanding the mechanism underlying the metabolite exchange between corals and their algal symbionts, as well as the metabolic flow within holobionts. To examine the distribution of Symbiodiniaceae cells within corals, in this study, we used fluorescence imaging and matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MS-Imaging) on branches of the Acropora tenuis coral. We successfully prepared frozen sections of the coral for molecular imaging without fixing or decalcifying the coral branches. By combining the results of MS-Imaging with that of the fluorescence imaging, we determined that the algal Symbiodiniaceae symbionts were not only localized in the tentacle and surface region of the coral branches but also inhabited the in inner parts. Therefore, the molecular imaging technique used in this study could be valuable to further investigate the molecular dynamics between corals and their symbionts.

The starfish Asterias amurensis, a well-known predator of molluscan species in intertidal ecosystems, has caused substantial ecological and economic losses in North China such as offshore Qingdao. Effective monitoring and prevention measures are urged to minimize its negative impacts. Compared with traditional biomonitoring methods, environmental DNA technology has emerged as a powerful and cost-efficient tool for inferring species’ presence and abundance. In this study, we developed a pair of species-specific primers (i.e., Ast-F and Ast-R) for the A. amurensis mitochondrial COI gene and tested its utility in amplifying and quantifying the DNA fragments from environmental samples under both laboratory and field conditions. The results of controlled water tank experiments demonstrated that the amount of eDNA released by A. amurensis was positively related to its biomass; after the removal of the starfish, the eDNA degraded significantly in 24 h and remained detectable for 8 days. The number of eDNA copies enriched tended to increase with smaller pore size of filter membrane and larger volume of filtered water. For field tests, we confirmed the validation of our approach in six locations in Qingdao by filtering 1000 ml water per sample with a 0.45-µm pore size filtration. All the amplification products generated a single and bright band via gel electrophoresis, and the quantitative PCR results unveiled significant differences in eDNA copies. This study provided an eDNA-based approach for investigating the distribution and biomass of A. amurensis, which may help to formulate early warning and management strategies in coastal Qingdao and other regions.

The caligid ectoparasite, Caligus rogercresseyi, is one of the main concerns in the Chilean salmon industry. The molecular mechanisms displayed by the parasite during the reproductive process represent an opportunity for developing novel control strategies. Vitellogenin is a multifunctional protein recognized as a critical player in several crustaceans’ biological processes, including reproduction, embryonic development, and immune response. This study aimed to characterize the C. rogercresseyi vitellogenins, including discovering novel transcripts and regulatory mechanisms associated with microRNAs. Herein, vitellogenin genes were identified by homology analysis using the reference sea louse genome, transcriptome database, and arthropods vitellogenin-protein database. The validation of expression transcripts was conducted by RNA nanopore sequencing technology. Moreover, fusion gene profiling, miRNA target analysis, and functional validation were performed using luciferase assay. Six putative vitellogenin genes were identified in the C. rogercresseyi genome with high homology with other copepods vitellogenins. Furthermore, miR-996 showed a putative role in regulating the Cr_Vitellogenin1 gene, which is highly expressed in females. Moreover, vitellogenin-fusion genes were identified in adult stages and highly regulated in males, demonstrating sex-related expression patterns. In females, the identified fusion genes merged with several non-vitellogenin genes involved in biological processes of ribosome assembly, BMP signaling pathway, and biosynthetic processes. This study reports the genome array of vitellogenins in C. rogercresseyi for the first time, revealing the putative role of fusion genes and miRNA regulation in sea lice biology.

Natural substances are strategic candidates for drug development in cancer research. Marine-derived molecules are of special interest due to their wide range of biological activities and sustainable large-scale production. Melanoma is a type of skin cancer that originates from genetic mutations in melanocytes. BRAF, RAS, and NF1 mutations are described as the major melanoma drivers, but approximately 20% of patients lack these mutations and are included in the triple wild-type (tripleWT) classification. Recent advances in targeted therapy directed at driver mutations along with immunotherapy have only partially improved patients’ overall survival, and consequently, melanoma remains deadly when in advanced stages. Fucose-containing sulfated polysaccharides (FCSP) are potential candidates to treat melanoma; therefore, we investigated Fucan A, a FCSP from Spatoglossum schröederi brown seaweed, in vitro in human melanoma cell lines presenting different mutations. Up to 72 h Fucan A treatment was not cytotoxic either to normal melanocytes or melanoma cell lines. Interestingly, it was able to impair the tripleWT CHL-1 cell proliferation (57%), comparable to the chemotherapeutic cytotoxic drug cisplatin results, with the advantage of not causing cytotoxicity. Fucan A increased CHL-1 doubling time, an effect attributed to cell cycle arrest. Vascular mimicry, a close related angiogenesis process, was also impaired (73%). Fucan A mode of action could be related to gene expression modulation, in special β-catenin downregulation, a molecule with protagonist roles in important signaling pathways. Taken together, results indicate that Fucan A is a potential anticancer molecule and, therefore, deserves further investigation.

Bisphenol A is one of the most used components of the polycarbonate plastic industry in the word. This contaminant has disrupting effect in cells in in vitro and in vivo in fish. This study evaluated for the first time the cytotoxicity, oxidative stress and apoptosis induced by bisphenol A (BPA) in head-kidney and spleen leukocytes isolated from Pacific red snapper Lutjanus peru. Head-kidney and spleen leukocytes were exposed to 100, 1000 and 10,000 µg/mL of BPA at 2 and 24 h. Results showed cytotoxicity of BPA at 1000 and 10,000 µg/mL. Cell viability > 80% was observed in leukocytes exposed to 100 µg/mL for 2 h; thus, this concentration was selected for the remainder of the study. Reactive oxygen species (ROS) production, analyzed by DCF-DA and NBT assays, significantly increased in those leukocytes exposed to BPA compared to controls after 2 or 24 h. Superoxide dismutase and catalase activities increased in head-kidney leukocytes after 24 h of BPA exposure. Apoptosis was inferred from caspase (casp-1 and casp-3), granzyme A (granz-A) and perforin 1 (perf-1) gene expression, which was significantly up-regulated, at 2 h BPA exposure in head-kidney leukocytes, and from granz-A and perf-1, which were up-regulated, after 24 h BPA exposure in spleen leukocytes. Short cytoplasmic prolongations and membrane blebs, suggestive of apoptosis, were observed by scanning electron microscopy. These data suggest that BPA at 100 µg/mL induces cytotoxicity, oxidative stress, apoptosis in Pacific red snapper head-kidney and spleen leukocytes.

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