Cytotoxic and radical activities of metal-organic framework modified with iron oxide: Biological and physico-chemical analyses.

Metal-organic framework (MOF) modified with iron oxide, Fe3O4-MOF, is a perspective drug delivery agent, enabling magnetic control and production of active hydroxyl radicals, •OH, via the Fenton reaction. This paper studies cytotoxic and radical activities of Fe-containing nanoparticles (NPs): Fe3O4-MOF and its components - bare Fe3O4 and MOF (MIL-88B). Luminous marine bacteria Photobacteriumphosphoreum were used as a model cellular system to monitor bioeffects of the NPs. Neither the NPs of Fe3O4-MOF nor MOF showed cytotoxic effects in a wide range of concentrations (<10 mg/L); while Fe3O4 was toxic at >3·10-3 mg/L. The NPs of Fe3O4 did not affect the bacterial bioluminescence enzymatic system; their toxic effect was attributed to cellular membrane processes. The integral content of reactive oxygen species (ROS) was determined using a chemiluminescence luminol assay. Bacteria mitigated excess of ROS in water suspensions of Fe3O4-MOF and MOF, maintaining bioluminescence intensity closer to the control; this resulted in low toxicity of these NPs. We estimated the activity of •OH radicals in the NPs samples with physical and chemical methods - spin capture technology (using electron paramagnetic resonance spectroscopy) and methylene blue degradation. Physico-chemical interpretation of cellular responses is provided in terms of iron content, iron ions release and •OH radical production.

Photobacterium pectinilyticum sp. nov., a novel bacterium isolated from surface seawater of Qingdao offshore.

A Gram-staining-negative, facultative aerobic, motile strain, designated strain ZSDE20T, was isolated from the surface seawater of Qingdao offshore. Phylogenetic analysis of the 16S rRNA gene of strain ZSDE20T, affiliated it to the genus Photobacterium. It was closely related to Photobacterium lutimaris DF-42 T (98.92% 16S rRNA gene sequence similarity). Growth occurred at 4-28ºC (optimum 28ºC), pH 1.0-7.0 (optimum 7.0) and in the presence of 1-7% (w/v) NaCl (optimum 3%). The dominant fatty acids were summed feature 3 (C16:1 ω7c or/and C16:1 ω6c, 34.23%), summed feature 8 (C18:1 ω7c and C18:1 ω6c, 10.36%) and C16:0 (20.05%). The polar lipids of strain ZSDE20T comprised phosphatidylethanolamine, phosphatidylcholine, lyso-phosphatidylglycerol, phosphatidylethanolamine, phosphatidylinositol dimannoside, phosphatidylinositol mannosides and two unknown lipids. The major respiratory quinone was ubiquinone-8 (Q-8). The DNA G + C content of strain ZSDE20T was 45.6 mol%. Average nucleotide identity (ANI) values between ZSDE20T and its reference species were lower than the threshold for species delineation (95-96%); in silico DNA-DNA hybridization further showed that strain ZSDE20T had less than 70% similarity to its relatives. Based on the polyphasic evidences, strain ZSDE20T is proposed as representing a novel species of the genus Photobacterium, for which the name Photobacterium pectinilyticum sp. nov. is proposed. The type strain is ZSDE20T (= MCCC 1K06283T = KCTC 82885 T).

Photobacterium obscurum sp. nov., a marine bacterium isolated from the coast of Qingdao.

A Gram-stain-negative, facultative anaerobic, rod-shaped strain, named SDRW27T, was isolated from offshore seawater collected near Qingdao. Strain SDRW27T was able to grow at 16-37 °C (optimum, 28 °C), pH 6.0-9.0 (optimum, pH 6.0) and in the presence of 1-7 % (w/v) NaCl (optimum, 3 %). Phylogenetic analysis using 16S rRNA gene sequences indicated that strain SDRW27T was most closely related to Photobacterium toruni H01100410BT (97.89 % sequence similarity), Photobacterium andalusiense H01100409BT (97.89 %) and Photobacterium leiognathi ATCC 25521T (97.82 %). The predominant fatty acids were summed feature 3 (C16 : 1 ω7c and/or iso-C15 : 0 2-OH), summed feature 8 (C18 : 1 ω7c and/or C18 : 1 ω6c) and C16 : 0. The polar lipids of strain SDRW27T comprised phosphatidylglycerol, phosphatidylinositol dimannoside, phosphatidylcholine, phosphatidylethanolamine and three unidentified lipids. The major respiratory quinone was ubiquinone-8. The G+C content was 47.71 mol%. The genome size was 5.84 Mbp, including 85 contigs with an N50 value of 223 542. The average nucleotide identity (ANI) values of SDRW27T with its three most similar strains, P. toruni H01100410BT, P. andalusiense H01100409BT and P. leiognathi ATCC 25521T, were 71.36, 71.58 and 72.23 %, respectively (all lower than the 95-96 % ANI threshold), and the DNA-DNA hybridization (DDH) values were 20.4, 20.8 and 20.4 % (all lower than the 70 % DDH threshold). The obtained results of polyphasic analysis demonstrate that strain SDRW27T represents a novel species, for which the name Photobacterium obscurum sp. nov. is proposed. The type strain is SDRW27T (=MCCC 1K06286T=KCTC 82892T).

Susceptibility of Sea Bream (Sparus aurata) to AIP56, an AB-Type Toxin Secreted by Photobacterium damselae subsp. piscicida.

Photobacterium damselae subsp. piscicida (Phdp) is a Gram-negative bacterium that infects a large number of marine fish species in Europe, Asia, and America, both in aquacultures and in the natural environment. Among the affected hosts are economically important cultured fish, such as sea bream (Sparus aurata), sea bass (Dicentrarchus labrax), yellowtail (Seriola quinqueradiata), and cobia (Rachycentron canadum). The best characterized virulence factor of Phdp is the Apoptosis-Inducing Protein of 56 kDa (AIP56), a secreted AB-type toxin that has been shown to induce apoptosis of sea bass phagocytes during infection. AIP56 has an A subunit that displays metalloprotease activity against NF-kB p65 and a B subunit that mediates binding and internalization of the A subunit in susceptible cells. Despite the fact that the aip56 gene is highly prevalent in Phdp isolates from different fish species, the toxicity of AIP56 has only been studied in sea bass. In the present study, the toxicity of AIP56 for sea bream was evaluated. Ex vivo assays showed that sea bream phagocytes are resistant to AIP56 cytotoxicity and that resistance was associated with an inefficient internalization of the toxin by those cells. Accordingly, in vivo intoxication assays revealed that sea bream is much more resistant to AIP56-induced lethality than sea bass. These findings, showing that the effect of AIP56 is different in sea bass and sea bream, set the basis for future studies to characterize the effects of AIP56 and to fully elucidate its virulence role in different Phdp susceptible hosts.

Structural insights into the enhanced thermostability of cysteine substitution mutants of L-histidine decarboxylase from Photobacterium phosphoreum.

Enzymatic amino acid assays are important in physiological research and clinical diagnostics because abnormal amino acid concentrations in biofluids are associated with various diseases. L-histidine decarboxylase from Photobacterium phosphoreum (PpHDC) is a pyridoxal 5'-phosphate-dependent enzyme and a candidate for use in an L-histidine quantitation assay. Previous cysteine substitution experiments demonstrated that the PpHDC C57S mutant displayed improved long-term storage stability and thermostability when compared with those of the wild-type enzyme. In this study, combinational mutation experiments of single cysteine substitution mutants of PpHDC were performed, revealing that the PpHDC C57S/C101V/C282V mutant possessed the highest thermostability. The stabilizing mechanism of these mutations was elucidated by solving the structures of PpHDC C57S and C57S/C101V/C282V mutants by X-ray crystallography. In the crystal structures, two symmetry-related PpHDC molecules form a domain-swapped homodimer. The side chain of S57 is solvent exposed in the structure, indicating that the C57S mutation eliminates chemical oxidation or disulfide bond formation with a free thiol group, thereby providing greater stability. Residues 101 and 282 form hydrophobic interactions with neighboring hydrophobic residues. Mutations C101V and C282V enhanced thermostability of PpHDC by filling a cavity present in the hydrophobic core (C101V) and increasing hydrophobic interactions.

Photobacterium damsela Necrotizing Fasciitis of the Arm.

Photobacterium damsela is a gram-negative bacterium that is known to cause infection, most commonly in marine animals and rarely in humans. Photobacterium damsela typically results in skin and soft tissue infections in humans and has been associated with necrotizing fasciitis in several case reports. After an initially benign presentation, P damsela necrotizing fasciitis often progresses rapidly to multiorgan failure and death. Here, we present a fatal case of P damsela necrotizing fasciitis after a minor cut while scaling a fish with a discussion of current literature and treatment strategies.

The Era of Antimicrobial Peptides: Use of Hepcidins to Prevent or Treat Bacterial Infections and Iron Disorders.

The current treatments applied in aquaculture to limit disease dissemination are mostly based on the use of antibiotics, either as prophylactic or therapeutic agents, with vaccines being available for a limited number of fish species and pathogens. Antimicrobial peptides are considered as promising novel substances to be used in aquaculture, due to their antimicrobial and immunomodulatory activities. Hepcidin, the major iron metabolism regulator, is found as a single gene in most mammals, but in certain fish species, including the European sea bass (Dicentrarchus labrax), two different hepcidin types are found, with specialized roles: the single type 1 hepcidin is involved in iron homeostasis trough the regulation of ferroportin, the only known iron exporter; and the various type 2 hepcidins present antimicrobial activity against a number of different pathogens. In this study, we tested the administration of sea bass derived hepcidins in models of infection and iron overload. Administration with hamp2 substantially reduced fish mortalities and bacterial loads, presenting itself as a viable alternative to the use of antibiotics. On the other hand, hamp1 seems to attenuate the effects of iron overload. Further studies are necessary to test the potential protective effects of hamp2 against other pathogens, as well as to understand how hamp2 stimulate the inflammatory responses, leading to an increased fish survival upon infection.

Chemical and Enzymatic Synthesis of Sialylated Glycoforms of Human Erythropoietin.

Recombinant human erythropoietin (EPO) is the main therapeutic glycoprotein for the treatment of anemia in cancer and kidney patients. The in-vivo activity of EPO is carbohydrate-dependent with the number of sialic acid residues regulating its circulatory half-life. EPO carries three N-glycans and thus obtaining pure glycoforms provides a major challenge. We have developed a robust and reproducible chemoenzymatic approach to glycoforms of EPO with and without sialic acids. EPO was assembled by sequential native chemical ligation of two peptide and three glycopeptide segments. The glycopeptides were obtained by pseudoproline-assisted Lansbury aspartylation. Enzymatic introduction of the sialic acids was readily accomplished at the level of the glycopeptide segments but even more efficiently on the refolded glycoprotein. Biological recognition of the synthetic EPOs was shown by formation of 1:1 complexes with recombinant EPO receptor.

Intestinal Explant Cultures from Gilthead Seabream (Sparus aurata, L.) Allowed the Determination of Mucosal Sensitivity to Bacterial Pathogens and the Impact of a Plant Protein Diet.

The interaction between diet and intestinal health has been widely discussed, although in vivo approaches have reported limitations. The intestine explant culture system developed provides an advantage since it reduces the number of experimental fish and increases the time of incubation compared to similar methods, becoming a valuable tool in the study of the interactions between pathogenic bacteria, rearing conditions, or dietary components and fish gut immune response. The objective of this study was to determine the influence of the total substitution of fish meal by plants on the immune intestinal status of seabream using an ex vivo bacterial challenge. For this aim, two growth stages of fish were assayed (12 g): phase I (90 days), up to 68 g, and phase II (305 days), up to 250 g. Additionally, in phase II, the effects of long term and short term exposure (15 days) to a plant protein (PP) diet were determined. PP diet altered the mucosal immune homeostasis, the younger fish being more sensitive, and the intestine from fish fed short-term plant diets showed a higher immune response than with long-term feeding. Vibrio alginolyticus (V. alginolyticus) triggered the highest immune and inflammatory response, while COX-2 expression was significantly induced by Photobacterium damselae subsp. Piscicida (P. damselae subsp. Piscicida), showing a positive high correlation between the pro-inflammatory genes encoding interleukin 1ß (IL1-ß), interleukin 6 (IL-6) and cyclooxygenase 2(COX-2).

Molecular and histopathological characterization of Photobacterium damselae in naturally and experimentally infected Nile tilapia (Oreochromis niloticus).

Mass mortality has occurred among cultured Nile tilapia, Oreochromis niloticus, on fish farms in Manzala, Dakahlia province, Egypt, in the summer season, 2019. Moribund fish were reported with deep ulcers, septicaemic lesions and sampled for bacterial isolation. In this study, most isolates were subjected to bacteriological examination, antibiotic sensitivity test, 16S rRNA gene sequencing and histopathological examination. Following isolate identification, intraperitoneal challenge of Nile tilapia with a bacterial suspension 2 × 106  CFU/ml was performed. Samples from liver, spleen and kidney were collected for histological and biochemical analysis. The results showed a high similarity (99%) to Photobacterium damselae strains using phylogenetic analysis of 16S rRNA. P. damselae exhibited resistance to amoxicillin and erythromycin, as well it was highly sensitive to chloramphenicol and doxycycline. Moreover, haemorrhage, oedema, hemosiderosis and melanomacrophage activation in the liver and head kidney of infected fish were detected by light and electron microscopy. Also, significant higher levels of CAT and SOD in the spleen and head kidney, as well as the serum levels of NO were observed in experimentally challenged O. niloticus, compared to the control fish. Our data identified P. damselae for the first time from infected Nile tilapia, describing its sensitivity to a variety of antibiotics, histopathological alterations and oxidative stress impact, and it could be useful indicators for understanding P. damselae pathogenesis, which might provide a preventive efficacy for P. damselae.

Effects of aqueous and ethanolic leaf extracts from drumstick tree (Moringa oleifera) on gilthead seabream (Sparus aurata L.) leucocytes, and their cytotoxic, antitumor, bactericidal and antioxidant activities.

Aqueous and ethanolic extracts of drumstick, Moringa oleifera, leaves were evaluated in vitro to ascertain their principal active components and determine their immunostimulant, cytotoxic, antitumoral, bactericidal and antioxidant activities. Phytochemical screening of M. oleifera leaf extracts showed a greater abundance of phenolic and cyanogenic glycosides in aqueous than in ethanolic extracts, characterized by several flavonoids, condensed tannins and saponins. No significant effects on gilthead seabream (Sparus aurata) head-kidney leucocyte activities (phagocytic ability and capacity, respiratory burst and peroxidase) were detected after incubation for 24 h with different concentrations (0.001/1 mg mL-1) of either extract. In addition, the aqueous extract showed a marked cytotoxic effect on both SAF-1 (at doses above 0.01 mg mL-1) and PLHC-1 (at doses above 0.25 mg mL-1) cell lines. The ethanolic extract improved the viability of SAF-1 cells and decreased the viability of PLHC-1 cells when used at higher concentrations. Both the ethanolic and, particularly, the aqueous extracts showed significant bactericidal activity on pathogenic Vibrio anguillarum and Photobacterium damselae strains. The antiradical activity of M. oleifera, as determined by the ABTS assay, increased in a linear dose-response with increasing extract concentrations. The results as a whole for the cytotoxic, bactericidal and antioxidant activities of M. oleifera leaf extracts point to their possible use as additives in functional diets for farmed fish.

Rapid destruction of triclosan by Iron(III)-Tetraamidomacrocyclic ligand/hydrogen peroxide system.

Iron(III)-tetraamidomacrocyclic ligand (Fe(III)-TAML) activators can activate hydrogen peroxide to oxidize many kinds of organic pollutants. In this study, we investigated the degradation of triclosan, a widely used broad-spectrum bactericide, under the treatment of Fe(III)-TAML/H2O2 system at different pH conditions. We also studied the influence of natural organic matter (NOM) on the degradation process. Our results showed that complete removal of triclosan could be obtained within several minutes under the optimal conditions. The degradation of triclosan by Fe(III)-TAML/H2O2 system exhibited strong pH-dependence and the degradation rate increased with the increase in pH level from 7.0 to 10.0. When adding fulvic acid (FA) or humic acid (HA) in the reaction system, the degradation of triclosan could be suppressed slightly, and HA exhibited stronger inhibition than FA. Based on the analysis of reaction intermediates, phenoxyl radical reaction and ring open reaction were involved in the decomposition of triclosan. Significant inhibition of overall toxicity to Photobacterium phosphoreum further confirmed the high efficiency of Fe(III)-TAML/H2O2 system for the removal of antibiotic activities resulting from the parent triclosan molecule and its degradation products.

The interplay between iron limitation, light and carbon in the proteorhodopsin-containing Photobacterium angustum S14.

Iron (Fe) limitation is known to affect heterotrophic bacteria within the respiratory electron transport chain, therefore strongly impacting the overall intracellular energy production. We investigated whether the gene expression pattern of the light-sensitive proton pump, proteorhodopsin (PR), is influenced by varying light, carbon and Fe concentrations in the marine bacterium Photobacterium angustum S14 and whether PR can alleviate the physiological processes associated with Fe starvation. Our results show that the gene expression of PR increases as cells enter the stationary phase, irrespective of Fe-replete or Fe-limiting conditions. This upregulation is coupled to a reduction in cell size, indicating that PR gene regulation is associated with a specific starvation-stress response. We provide experimental evidence that PR gene expression does not result in an increased growth rate, cell abundance, enhanced survival or ATP concentration within the cell in either Fe-replete or Fe-limiting conditions. However, independent of PR gene expression, the presence of light did influence bacterial growth rates and maximum cell abundances under varying Fe regimes. Our observations support previous results indicating that PR phototrophy seems to play an important role within the stationary phase for several members of the Vibrionaceae family, but that the exact role of PR in Fe limitation remains to be further explored.

The strains of bioluminescent bacteria isolated from the White Sea finfishes: genera Photobacterium, Aliivibrio, Vibrio, Shewanella, and first luminous Kosakonia.

Bioluminescence is a spectacular feature of some prokaryotes. In the present work, we address the distribution of bioluminescence among bacteria isolated from the White Sea finfishes. Luminous bacteria are widely distributed throughout the World Ocean. Many strains have been isolated and described for tropical latitudes, while Nordic seas still remain quite a white spot in studying bioluminescence of bacteria. We describe the strains related to the two main genera of luminous bacteria, Photobacterium and Aliivibrio, as well as Shewanella and Vibrio. They are related to families Vibrionaceae and Shewanellaceae of the Gammaproteobacteria class. Here, we at the first time, report the bioluminescence of the Enterobacteriaceae Kosakonia cowanii. Moreover, we applied the polyphasic approach to identify and describe the isolated microorganisms. The data on sequencing, diversity of cell fine structure, and light emission spectra at room temperature on the solid medium are discussed. The bacteria are characterized by features in their light emission spectra. It may reflect possible molecular mechanisms of bioluminescence as well as features of bacterial composition. The obtained data expands the existing body of knowledge about the bioluminescence spread among the bacteria of Nordic latitudes and provides complex information that is crucial for their precise identification.

Ecotoxicity and interacting mechanism of anionic surfactant sodium dodecyl sulfate (SDS) and its mixtures with nonionic surfactant fatty alcohol-polyoxyethlene ether (AEO).

This paper reports the proportion-dependent toxicity of binary surfactant mixtures containing anionic sodium dodecyl sulfate (SDS) and nonionic fatty alcohol-polyoxyethlene ether (AEO) toward Photobacterium phosphoreum. The crucial role of toxicity interactions was elucidated by spectroscopic probing the refolding of the unfolded bovine serum albumin (BSA) induced by SDS and theoretical calculating the interaction parameter of mixed surfactants based on Rubingh's model from the critical micelle concentrations. The SDS/AEO mixtures can be divided into two groups based on the toxicity response to the proportion of AEO in the mixtures: Group I contained low mass proportions of AEO, that is, SDS:AEO = 4:1, 3:1; Group II featured high AEO proportions, that is, SDS:AEO = 3:2, 1:1, 2:3, 1:4. The toxicity of SDS/AEO mixtures decreased with the enhanced proportion of AEO in Group I and then fluctuated slightly when the AEO proportion increased to that of Group II. The mixture with the mass ratio of 1:1 showed a slightly higher toxicity than the others in Group II. Scanning electron microscopy (SEM) images illustrated that the addition of AEO hindered the action of SDS against the cell membrane. Fluorescence measurement indicated that AEO could extract SDS molecules embedded in the BSA matrix, except for those bound to the highly active sites of BSA, and refold stepwise the unfolded protein. The results were in excellent analogy to the proportion-dependent toxicity of SDS/AEO mixture, indicating the formation of mixed micelles playing a key role. The interaction parameter further revealed that antagonism led to the mixture with equal mass ratio (1:1) showing higher toxicity than other mass ratios in Group II. These results can be useful for compounding SDS/AEO mixtures in application efficiently and eco-friendly.

Inclusion of dietary Ulva ohnoi 5% modulates Solea senegalensis immune response during Photobacterium damselae subsp. piscicida infection.

Macroalgae represent valuable sources of functional ingredients for fish diets, and the influence of supplemented aquafeeds on growth performance has been studied for some fish and seaweed species. In the present work, the potential immunomodulation exerted by U. ohnoi (5%) as dietary ingredient was investigated in Senegalese sole. After feeding with the experimental diets for 90 d, fish immune response before and after challenge with Photobacterium damselae subsp. piscicida (Phdp) was assessed. In absence of infection, systemic immune response was not modified by 5% U. ohnoi dietary inclusion for 90 d. Thus, no differences in liver and head kidney immune gene transcription or serum lysozyme, peroxidase, antiprotease and complement activities were observed based on the diet received by Senegalese sole specimens. Regarding mucosal immune parameters, no changes in gene transcription were detected in the skin and gills, whilst only tnf, cd4 and cd8 were significantly up-regulated in the intestine of fish fed with U. ohnoi, compared to the values obtained with control diet. On the contrary, when S. senegalensis specimens were challenged with Phdp, modulation of the immune response consisting in increased transcription of genes encoding complement (c1q4, c3, c9), lysozyme g (lysg), tumor necrosis factor alpha (tnfα) as well as those involved in the antioxidant response (gpx, sodmn) and iron metabolism (ferrm, hamp-1) was observed in the liver of fish fed with U. ohnoi. In parallel, decreased inflammatory cytokine and complement encoding gene transcription was displayed by the spleen of fish receiving the algal diet. Though mortality rates due to Phdp challenge were not affected by the diet received, lower pathogen loads were detected in the liver of soles receiving U. ohnoi diet. Further research to investigate the effects of higher inclusion levels of this seaweed in fish diets, feeding during short periods as wells as to assess the response against other pathogens needs to be carried out.

Functional characterization of NK-lysin in golden pompano Trachinotus ovatus (Linnaeus 1758).

NK-lysin is an important part of the innate immune defence system and plays an important role in resisting the invasion of pathogenic microorganisms. In this study, NK-lysin from golden pompano (Trachinotus ovatus) was characterized and its expression in response to Photobacterium damselae was investigated. The full-length NK-lysin cDNA was 731 bp, which comprised a 5'-UTR of 63 bp, an ORF of 444 bp, and a 3'-UTR of 224 bp, and encoded 147 amino acids; NK-lysin consisted of a conserved saposin B domain and six conserved cysteines that formed three pairs of disulfide bonds. The genomic organization of NK-lysin was also determined and the gene consisted of four introns and five exons. The predicted promoter region of ToNK-lysin contained several putative transcription factor binding sites. Quantitative real-time (qRT-PCR) analysis indicated that ToNK-lysin was ubiquitously expressed in all examined tissues; the highest mRNA levels were observed in the skin, kidney and intestine, while the lowest expression level was detected in the stomach. After P. damselae stimulation, the expression level of NK-lysin mRNA was significantly upregulated in various tissues of golden pompano. In addition, SDS-PAGE showed that the molecular mass of recombinant NK-lysin expressed in pGEX-6P-1 was approximately 37 kDa. The purified recombinant protein showed antibacterial activity against gram-positive and gram-negative bacteria. The results indicate that golden pompano NK-lysin has potential antimicrobial roles in fish innate immunity.

Role of AIP56 disulphide bond and its reduction by cytosolic redox systems for efficient intoxication.

Apoptosis-inducing protein of 56 kDa (AIP56) is a major virulence factor of Photobacterium damselae subsp. piscicida, a gram-negative pathogen that infects warm water fish species worldwide and causes serious economic losses in aquacultures. AIP56 is a single-chain AB toxin composed by two domains connected by an unstructured linker peptide flanked by two cysteine residues that form a disulphide bond. The A domain comprises a zinc-metalloprotease moiety that cleaves the NF-kB p65, and the B domain is involved in binding and internalisation of the toxin into susceptible cells. Previous experiments suggested that disruption of AIP56 disulphide bond partially compromised toxicity, but conclusive evidences supporting the importance of that bond in intoxication were lacking. Here, we show that although the disulphide bond of AIP56 is dispensable for receptor recognition, endocytosis, and membrane interaction, it needs to be intact for efficient translocation of the toxin into the cytosol. We also show that the host cell thioredoxin reductase-thioredoxin system is involved in AIP56 intoxication by reducing the disulphide bond of the toxin at the cytosol. The present study contributes to a better understanding of the molecular mechanisms operating during AIP56 intoxication and reveals common features shared with other AB toxins.

Comparison of toxicity induced by EDTA-Cu after UV/H2O2 and UV/persulfate treatment: Species-specific and technology-dependent toxicity.

Advanced oxidation processes (AOPs) can degrade heavy metal complexes in wastewater to improve the removal efficiency of metals. However, the influences of AOP treatments on toxicity induced by metal complexes are not well understood. This study compared the toxicity induced by EDTA-copper (Cu) after UV/persulfate (PS) and UV/H2O2 treatments on luminescent bacteria and human HepG2 cells. The results showed that EDTA-Cu complexes decreased Cu toxicity in luminescent bacteria but increased the cytotoxicity in HepG2 cells, indicating species-specific toxicity. The UV/PS and UV/H2O2 treatments under most pH values and [oxidant]/[EDTA-Cu] conditions decreased the toxicity of EDTA-Cu in HepG2 cells but increased the toxicity in luminescent bacteria. When the ratio of [oxidant] to [EDTA-Cu] was 10, low toxicity in treated solutions was observed in both UV treatment processes. The alkaline precipitation treatment had a significant influence on toxicity reduction after UV/PS treatment; however, it had minimal influence on the UV/H2O2 treatment system. The Cu and total organic carbon (TOC) removal efficiency cannot completely explain the results of toxicity assays. EDTA-Cu intermediates might play important roles in changing the toxicity of EDTA-Cu after both UV treatments. This study provides insights into evaluating the treatment efficiency of UV/PS and UV/H2O2 on EDTA-Cu decomplexation.

Protective effects of seaweed supplemented diet on antioxidant and immune responses in European seabass (Dicentrarchus labrax) subjected to bacterial infection.

European seabass (Dicentrarchus labrax) production is often hampered by bacterial infections such as photobacteriosis caused by Photobacterium damselae subsp. piscicida (Phdp). Since diet can impact fish immunity, this work investigated the effect of dietary supplementation of 5% Gracilaria sp. aqueous extract (GRA) on seabass antioxidant capacity and resistance against Phdp. After infection, mortality was delayed in fish fed GRA, which also revealed increased lysozyme activity levels, as well as decreased lipid peroxidation, suggesting higher antioxidant capacity than in fish fed a control diet. Dietary GRA induced a down-regulation of hepatic stress-responsive heat shock proteins (grp-78, grp-170, grp-94, grp-75), while bacterial infection caused a down-regulation in antioxidant genes (prdx4 and mn-sod). Diet and infection interaction down-regulated the transcription levels of genes associated with oxidative stress response (prdx5 and gpx4) in liver. In head-kidney, GRA led to an up-regulation of genes associated with inflammation (il34, ccr9, cd33) and a down-regulation of genes related to cytokine signalling (mif, il1b, defb, a2m, myd88). Additionally, bacterial infection up-regulated immunoglobulins production (IgMs) and down-regulated the transcription of the antimicrobial peptide leap2 in head kidney. Overall, we found that GRA supplementation modulated seabass resistance to Phdp infection.