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Journal of Applied Microbiology Jun 2018The objective was to determine the effects of dietary substitution of fishmeal (FM) with live yeast and increasing water temperature on the diversity and composition of...
AIMS
The objective was to determine the effects of dietary substitution of fishmeal (FM) with live yeast and increasing water temperature on the diversity and composition of gut microbiota in rainbow trout.
METHODS AND RESULTS
Fish were fed either FM or yeast (Saccharomyces cerevisiae) and reared in water temperatures of either 11°C (cold) or 18°C (warm) for 6 weeks. Luminal content and mucosa were collected from the distal gut and the load, diversity and species abundance of yeast and bacteria were analysed using agar plating, MALDI-TOF and rRNA gene amplicon sequencing. Yeast in the gut of fish fed FM were represented by S. cerevisiae, Rhodotorula spp. and Debaryomyces hansenii, while fish fed yeast contained 4-5 log higher CFU per g of yeast that were entirely represented by S. cerevisiae. For gut bacteria, sequencing of 16S rRNA gene amplicons using Illumina MiSeq showed lower bacterial diversity and abundance of lactic acid bacteria, especially Lactobacillus, in fish reared in warm rather than cold water. Fish fed yeast had similar bacterial diversity and lower abundance of Leuconostocaceae and Photobacterium compared with fish fed FM.
CONCLUSIONS
Feeding live yeast mainly increased yeast load in the gut, while increased water temperature significantly altered the gut microbiota of rainbow trout in terms of bacterial diversity and abundance.
SIGNIFICANCE AND IMPACT OF THE STUDY
Live yeast can replace 40% of FM without disrupting bacteria communities in the gut of rainbow trout, while increased water temperature due to seasonal fluctuations and/or climate change may result in a gut dysbiosis that may jeopardize the health of farmed fish.
Topics: Animal Feed; Animals; Bacteria; Gastrointestinal Microbiome; Gastrointestinal Tract; Oncorhynchus mykiss; RNA, Ribosomal, 16S; Saccharomyces cerevisiae; Temperature; Water
PubMed: 29464844
DOI: 10.1111/jam.13738 -
International Journal of Molecular... Mar 2023subsp. () is a Gram-negative fish pathogen with worldwide distribution and broad host specificity that causes heavy economic losses in aquaculture. Although was first...
subsp. () is a Gram-negative fish pathogen with worldwide distribution and broad host specificity that causes heavy economic losses in aquaculture. Although was first identified more than 50 years ago, its pathogenicity mechanisms are not completely understood. In this work, we report that secretes large amounts of outer membrane vesicles (OMVs) when cultured in vitro and during in vivo infection. These OMVs were morphologically characterized and the most abundant vesicle-associated proteins were identified. We also demonstrate that OMVs protect cells from the bactericidal activity of fish antimicrobial peptides, suggesting that secretion of OMVs is part of the strategy used by to evade host defense mechanisms. Importantly, the vaccination of sea bass () with adjuvant-free crude OMVs induced the production of anti- antibodies and resulted in partial protection against infection. These findings reveal new aspects of biology and may provide a basis for developing new vaccines against this pathogen.
Topics: Animals; Bass; Photobacterium; Virulence; Vaccines; Fish Diseases; Gram-Negative Bacterial Infections
PubMed: 36982212
DOI: 10.3390/ijms24065138 -
MicrobiologyOpen Mar 2021Polyhydroxyalkanoates (PHAs) are biodegradable bioplastics that can be manufactured sustainably and represent a promising green alternative to petrochemical-based...
Polyhydroxyalkanoates (PHAs) are biodegradable bioplastics that can be manufactured sustainably and represent a promising green alternative to petrochemical-based plastics. Here, we describe the complete genome of a new marine PHA-producing bacterium-Photobacterium ganghwense (strain C2.2), which we have isolated from the Black Sea seashore. This new isolate is psychrotolerant and accumulates PHA when glycerol is provided as the main carbon source. Transmission electron microscopy, specific staining with Nile Red visualized via epifluorescence microscopy and gas chromatography analysis confirmed the accumulation of PHA. This is the only PHA-producing Photobacterium for which we now have a complete genome sequence, allowing us to investigate the pathways for PHA production and other secondary metabolite synthesis pathways. The de novo assembly genome, obtained using open-source tools, comprises two chromosomes (3.5, 2 Mbp) and a megaplasmid (202 kbp). We identify the entire PHA synthesis gene cluster that encodes a class I PHA synthase, a phasin, a 3-ketothiolase, and an acetoacetyl-CoA reductase. No conventional PHA depolymerase was identified in strain C2.2, but a putative lipase with extracellular amorphous PHA depolymerase activity was annotated, suggesting that C2.2 is unable to degrade intracellular PHA. A complete pathway for the conversion of glycerol to acetyl-CoA was annotated, in accordance with its ability to convert glycerol to PHA. Several secondary metabolite biosynthetic gene clusters and a low number of genes involved in antibiotic resistance and virulence were also identified, indicating the strain's suitability for biotechnological applications.
Topics: Acetyl Coenzyme A; Acetyl-CoA C-Acyltransferase; Acyltransferases; Alcohol Oxidoreductases; Aquatic Organisms; Biosynthetic Pathways; Drug Resistance, Bacterial; Genome, Bacterial; Glycerol; Photobacterium; Plant Lectins; Plasmids; Polyhydroxyalkanoates; Soil Microbiology; Virulence; Whole Genome Sequencing
PubMed: 33970538
DOI: 10.1002/mbo3.1182 -
Microbiome Mar 2020Recently, increasing evidence supports that some complex diseases are not attributed to a given pathogen, but dysbiosis in the host intestinal microbiota (IM). The full...
BACKGROUND
Recently, increasing evidence supports that some complex diseases are not attributed to a given pathogen, but dysbiosis in the host intestinal microbiota (IM). The full intestinal ecosystem alterations, rather than a single pathogen, are associated with white feces syndrome (WFS), a globally severe non-infectious shrimp disease, while no experimental evidence to explore the causality. Herein, we conducted comprehensive metagenomic and metabolomic analysis, and intestinal microbiota transplantation (IMT) to investigate the causal relationship between IM dysbiosis and WFS.
RESULTS
Compared to the Control shrimp, we found dramatically decreased microbial richness and diversity in WFS shrimp. Ten genera, such as Vibrio, Candidatus Bacilloplasma, Photobacterium, and Aeromonas, were overrepresented in WFS, whereas 11 genera, including Shewanella, Chitinibacter, and Rhodobacter were enriched in control. The divergent changes in these populations might contribute the observation that a decline of pathways conferring lipoic acid metabolism and mineral absorption in WFS. Meanwhile, some sorts of metabolites, especially lipids and organic acids, were found to be related to the IM alteration in WFS. Integrated with multiomics and IMT, we demonstrated that significant alterations in the community composition, functional potentials, and metabolites of IM were closely linked to shrimp WFS. The distinguished metabolites which were attributed to the IM dysbiosis were validated by feed-supplementary challenge. Both homogenous selection and heterogeneous selection process were less pronounced in WFS microbial community assembly. Notably, IMT shrimp from WFS donors eventually developed WFS clinical signs, while the dysbiotic IM can be recharacterized in recipient shrimp.
CONCLUSIONS
Collectively, our findings offer solid evidence of the causality between IM dysbiosis and shrimp WFS, which exemplify the 'microecological Koch's postulates' (an intestinal microbiota dysbiosis, a disease) in disease etiology, and inspire our cogitation on etiology from an ecological perspective. Video abstract.
Topics: Animals; Bacteria; Dysbiosis; Fecal Microbiota Transplantation; Feces; Gastrointestinal Microbiome; Genetic Variation; Intestines; Penaeidae
PubMed: 32156316
DOI: 10.1186/s40168-020-00802-3 -
Frontiers in Nutrition 2023Valorising waste from the processing of fishery and aquaculture products into functional additives, and subsequent use in aquafeed as supplements could be a novel...
Valorising waste from the processing of fishery and aquaculture products into functional additives, and subsequent use in aquafeed as supplements could be a novel approach to promoting sustainability in the aquaculture industry. The present study supplemented 10% of various fish protein hydrolysates (FPHs), obtained from the hydrolysis of kingfish (KH), carp (CH) and tuna (TH) waste, with 90% of poultry by-product meal (PBM) protein to replace fishmeal (FM) completely from the barramundi diet. At the end of the trial, intestinal mucosal barriers damage, quantified by villus area (VA), lamina propria area (LPA), LPA ratio, villus length (VL), villus width (VW), and neutral mucin (NM) in barramundi fed a PBM-based diet was repaired when PBM was supplemented with various FPHs ( < 0.05, 0.01, and 0.001). PBM-TH diet further improved these barrier functions in the intestine of fish ( < 0.05 and 0.001). Similarly, FPHs supplementation suppressed PBM-induced intestinal inflammation by controlling the expression of inflammatory cytokines ( and ; < 0.05 and 0.001) and a mucin-relevant production gene (; < 0.001). The 16S rRNA data showed that a PBM-based diet resulted in dysbiosis of intestinal bacteria, supported by a lower abundance of microbial diversity ( < 0.001) aligned with a prevalence of Photobacterium. PBM-FPHs restored intestine homeostasis by enhancing microbial diversity compared to those fed a PBM diet ( < 0.001). PBM-TH improved the diversity ( < 0.001) further by elevating the Firmicutes phylum and the , , and genera. Muscle atrophy, evaluated by fiber density, hyperplasia and hypertrophy and associated genes (, , and ), occurred in barramundi fed PBM diet but was repaired after supplementation of FPHs with the PBM ( < 0.05, 0.01, and 0.001). Similarly, creatine kinase, calcium, phosphorous, and haptoglobin were impacted by PBM-based diet ( < 0.05) but were restored in barramundi fed FPHs supplemented diets ( < 0.05 and 0.01). Hence, using circular economy principles, functional FPHs could be recovered from the fish waste applied in aquafeed formulations and could prevent PBM-induced intestinal dysbiosis and muscular atrophy. GRAPHICAL ABSTRACT.
PubMed: 37057066
DOI: 10.3389/fnut.2023.1145068 -
ISME Communications Jun 2021Fish microbiota are intrinsically linked to health and fitness, but they are highly variable and influenced by both biotic and abiotic factors. Water temperature...
Fish microbiota are intrinsically linked to health and fitness, but they are highly variable and influenced by both biotic and abiotic factors. Water temperature particularly limits bacterial adhesion and growth, impacting microbial diversity and bacterial infections on the skin and gills. Aquaculture is heavily affected by infectious diseases, especially in warmer months, and industry practices often promote stress and microbial dysbiosis, leading to an increased abundance of potentially pathogenic bacteria. In this regard, fish mucosa health is extremely important because it provides a primary barrier against pathogens. We used 16 rRNA V4 metataxonomics to characterize the skin and gill microbiota of the European seabass, Dicentrarchus labrax, and the surrounding water over 12 months, assessing the impact of water temperature on microbial diversity and function. We show that the microbiota of external mucosae are highly dynamic with consistent longitudinal trends in taxon diversity. Several potentially pathogenic genera (Aliivibrio, Photobacterium, Pseudomonas, and Vibrio) were highly abundant, showing complex interactions with other bacterial genera, some of which with recognized probiotic activity, and were also significantly impacted by changes in temperature. The surrounding water temperature influenced fish microbial composition, structure and function over time (days and months). Additionally, dysbiosis was more frequent in warmer months and during transitions between cold/warm months. We also detected a strong seasonal effect in the fish microbiota, which is likely to result from the compound action of several unmeasured environmental factors (e.g., pH, nutrient availability) beyond temperature. Our results highlight the importance of performing longitudinal studies to assess the impact of environmental factors on fish microbiotas.
PubMed: 36739461
DOI: 10.1038/s43705-021-00019-x -
Nature Communications Feb 2023In bacteria and archaea, tripartite ATP-independent periplasmic (TRAP) transporters uptake essential nutrients. TRAP transporters receive their substrates via a secreted...
In bacteria and archaea, tripartite ATP-independent periplasmic (TRAP) transporters uptake essential nutrients. TRAP transporters receive their substrates via a secreted soluble substrate-binding protein. How a sodium ion-driven secondary active transporter is strictly coupled to a substrate-binding protein is poorly understood. Here we report the cryo-EM structure of the sialic acid TRAP transporter SiaQM from Photobacterium profundum at 2.97 Å resolution. SiaM comprises a "transport" domain and a "scaffold" domain, with the transport domain consisting of helical hairpins as seen in the sodium ion-coupled elevator transporter VcINDY. The SiaQ protein forms intimate contacts with SiaM to extend the size of the scaffold domain, suggesting that TRAP transporters may operate as monomers, rather than the typically observed oligomers for elevator-type transporters. We identify the Na and sialic acid binding sites in SiaM and demonstrate a strict dependence on the substrate-binding protein SiaP for uptake. We report the SiaP crystal structure that, together with docking studies, suggest the molecular basis for how sialic acid is delivered to the SiaQM transporter complex. We thus propose a model for substrate transport by TRAP proteins, which we describe herein as an 'elevator-with-an-operator' mechanism.
Topics: N-Acetylneuraminic Acid; Membrane Transport Proteins; Biological Transport; Archaea; Adenosine Triphosphate
PubMed: 36849793
DOI: 10.1038/s41467-023-36590-1 -
MicrobiologyOpen Aug 2023Gene inactivation studies are critical in pathogenic bacteria, where insights into species biology can guide the development of vaccines and treatments. Allelic exchange...
Transforming the untransformable with knockout minicircles: High-efficiency transformation and vector-free allelic exchange knockout in the fish pathogen Photobacterium damselae.
Gene inactivation studies are critical in pathogenic bacteria, where insights into species biology can guide the development of vaccines and treatments. Allelic exchange via homologous recombination is a generic method of targeted gene editing in bacteria. However, generally applicable protocols are lacking, and suboptimal approaches are often used for nonstandard but epidemiologically important species. Photobacterium damselae subsp. piscicida (Pdp) is a primary pathogen of fish in aquaculture and has been considered hard to transform since the mid-1990s. Consequently, conjugative transfer of RK2/RP4 suicide vectors from Escherichia coli S17-1/SM10 donor strains, a system prone to off-target mutagenesis, was used to deliver the allelic exchange DNA in previous studies. Here we have achieved efficient electrotransformation in Pdp using a salt-free highly concentrated sucrose solution, which performs as a hypertonic wash buffer, cryoprotectant, and electroporation buffer. High-efficiency transformation has enabled vector-free mutagenesis for which we have employed circular minimalistic constructs (knockout minicircles) containing only allelic exchange essentials that were generated by Gibson assembly. Preparation of competent cells using sucrose and electroporation/integration of minicircles had virtually no detectable off-target promutagenic effect. In contrast, a downstream sacB selection apparently induced several large deletions via mobilization of transposable elements. Electroporation of minicircles into sucrose-treated cells is a versatile broadly applicable approach that may facilitate allelic exchange in a wide range of microbial species. The method permitted inactivation of a primary virulence factor unique to Pdp, apoptogenic toxin AIP56, demonstrating the efficacy of minicircles for difficult KO targets located on the high copy number of small plasmids.
Topics: Animals; Photobacterium; Electroporation; Fishes
PubMed: 37642481
DOI: 10.1002/mbo3.1374 -
Angewandte Chemie (International Ed. in... Dec 2021Recombinant 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...
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.
Topics: Erythropoietin; Glycosylation; Humans; Molecular Structure; N-Acetylneuraminic Acid; Photobacterium; Sialyltransferases; beta-D-Galactoside alpha 2-6-Sialyltransferase
PubMed: 34523784
DOI: 10.1002/anie.202110013 -
International Journal of Food... Nov 2016The processing environment of salmon processing plants represents a potential major source of bacteria causing spoilage of fresh salmon. In this study, we have...
The processing environment of salmon processing plants represents a potential major source of bacteria causing spoilage of fresh salmon. In this study, we have identified major contamination routes of important spoilage associated species within the genera Pseudomonas, Shewanella and Photobacterium in pre-rigor processing of salmon. Bacterial counts and culture-independent 16S rRNA gene analysis on salmon fillet from seven processing plants showed higher levels of Pseudomonas spp. and Shewanella spp. in industrially processed fillets compared to salmon processed under strict hygienic conditions. Higher levels of Pseudomonas spp. and Shewanella spp. were found on fillets produced early on the production day compared to later processed fillets. The levels of Photobacterium spp. were not dependent on the processing method or time of processing. In follow-up studies of two plants, bacterial isolates (n=2101) from the in-plant processing environments (sanitized equipment/machines and seawater) and from salmon collected at different sites in the production were identified by partial 16S rRNA gene sequencing. Pseudomonas spp. dominated in equipment/machines after sanitation with 72 and 91% of samples from the two plants being Pseudomonas-positive. The phylogenetic analyses, based on partial 16S rRNA gene sequencing, showed 48 unique sequence profiles of Pseudomonas of which two were dominant. Only six profiles were found on both machines and in fillets in both plants. Shewanella spp. were found on machines after sanitation in the slaughter department while Photobacterium spp. were not detected after sanitation in any parts of the plants. Shewanella spp. and Photobacterium spp. were found on salmon in the slaughter departments. Shewanella was frequently present in seawater tanks used for bleeding/short term storage. In conclusion, this study provides new knowledge on the processing environment as a source of contamination of salmon fillets with Pseudomonas spp. and Shewanella spp., while Photobacterium spp. most likely originate from the live fish and seawater. The study show that strict hygiene during processing is a prerequisite for optimal shelf life of salmon fillets and that about 90% reductions in the initial levels of bacteria on salmon fillets can be obtained using optimal hygienic conditions.
Topics: Animals; Bacterial Load; Fish Products; Food Contamination; Food Handling; Food Industry; Food Microbiology; Food Preservation; Norway; Photobacterium; Phylogeny; Pseudomonas; RNA, Ribosomal, 16S; Salmon; Seawater; Shewanella
PubMed: 27552347
DOI: 10.1016/j.ijfoodmicro.2016.08.016