Unveiling the microbiome of hydroponically cultivated lettuce: impact of Phytophthora cryptogea infection on plant-associated microorganisms.

Understanding the complex interactions between plants and their associated microorganisms is crucial for optimizing plant health and productivity. While microbiomes of soil-bound cultivated crops are extensively studied, microbiomes of hydroponically cultivated crops have received limited attention. To address this knowledge gap, we investigated the rhizosphere and root endosphere of hydroponically cultivated lettuce. Additionally, we sought to explore the potential impact of the oomycete pathogen Phytophthora cryptogea on these microbiomes. Root samples were collected from symptomatic and nonsymptomatic plants in three different greenhouses. Amplicon sequencing of the bacterial 16S rRNA gene revealed significant alterations in the bacterial community upon P. cryptogea infection, particularly in the rhizosphere. Permutational multivariate analysis of variance (perMANOVA) revealed significant differences in microbial communities between plants from the three greenhouses, and between symptomatic and nonsymptomatic plants. Further analysis uncovered differentially abundant zero-radius operational taxonomic units (zOTUs) between symptomatic and nonsymptomatic plants. Interestingly, members of Pseudomonas and Flavobacterium were positively associated with symptomatic plants. Overall, this study provides valuable insights into the microbiome of hydroponically cultivated plants and highlights the influence of pathogen invasion on plant-associated microbial communities. Further research is required to elucidate the potential role of Pseudomonas and Flavobacterium spp. in controlling P. cryptogea infections within hydroponically cultivated lettuce greenhouses.

Total petroleum hydrocarbons and influencing factors in co-composting of rural sewage sludge and organic solid wastes.

Co-composting is an efficient strategy for collaborative disposal of multiple organic wastes in rural areas. In this study, we explored the co-composting of rural sewage sludge and other organic solid wastes (corn stalks and kitchen waste), with a focus on the variation of total petroleum hydrocarbons (TPH) during this process. 12% corn-derived biochar was applied in the composting (BC), with no additives applied as the control treatment (CK). The TPH contents of piles after composting ranged from 0.70 to 0.74 mg/g, with overall removal efficiencies of 35.6% and 61.1% for CK and BC, respectively. The results indicate that the addition of 12% biochar increased the rate of TPH degradation and accelerated the degradation process. 16s rDNA high-throughput sequencing was applied to investigate the biodiversity and bacterial community succession during the composting process. Diverse bacterial communities with TPH degradation functions were observed in the composting process, including Acinetobacter, Flavobacterium, Paenibacillus, Pseudomonas, and Bacillus spp. These functional bacteria synergistically degraded TPH, with cooperative behavior dominating during composting. Biochar amendment enhanced the microbial activity and effectively promoted the biodegradation of TPH. The physicochemical properties of the compost piles, including environmental factors (pH and temperature), nutrients (nitrogen, phosphorus, potassium), and humic substances produced in composting (humic acids and fulvic acids), directly and indirectly affected the variation in TPH contents. In conclusion, this work illustrates the variation in TPH content and associated influencing factors during co-composting of rural organic solid wastes, providing valuable guidance toward the further optimization of rural organic waste management.

Insufficient or excessive dietary carbohydrates affect gut health through change in gut microbiota and regulation of gene expression of gut epithelial cells in grass carp (Ctenopharyngodon idella).

Dietary carbohydrate levels can affect gut health, but the roles played by gut microbiota and gut epithelial cells, and their interactions remain unclear. In this experiment, we investigated gut health, gut microbiota, and the gene expression profiles of gut epithelial cells in grass carp consuming diets with different carbohydrate levels. Compared to the moderate-carbohydrate diet, low-carbohydrate diet significantly increased the relative abundance of pathogenic bacteria (Ralstonia and Elizabethkingia) and decreased the abundance of metabolism in cofactors and vitamins, implying a dysregulated gut microbiota and compromised metabolic function. Moreover, low-carbohydrate diet inhibited the expression levels of key genes in autophagy-related pathways in gut epithelial cells, which might directly lead to reduced clearance of defective organelles and pathogenic microorganisms. These aforementioned factors may be responsible for the imperfect organization of the intestinal tract. High-carbohydrate diet also significantly increased the abundance of pathogenic bacteria (Flavobacterium), which directly contributed to a decrease in the abundance of immune system of the microbiota. Furthermore, the active pathways of staphylococcus aureus infection and complement and coagulation cascades, as well as the inhibition of the glutathione metabolism pathway were observed. Above results implied that high-carbohydrate diet might ultimately cause severe gut damage by affecting immune function of microbiota, mentioned immune-related pathways, and the antioxidant capacity. Finally, the correlation network diagram revealed strong correlations of the differentially immune-related gene major histocompatibility complex class I antigen (MR1) with Enhydrobacter and Ruminococcus_gnavus_group in low-carbohydrate diet group, and Arenimonas in high-carbohydrate diet group, respectively, suggesting that MR1 might be a central target for immune responses in gut epithelial cells induced by gut microbiota at different levels of dietary carbohydrate. All these results provided insight in the development of antagonistic probiotics and target genes to improve the utilization of carbohydrate.

Variations in antibiotic resistance genes and microbial community in sludges passing through biological nutrient removal and anaerobic digestion processes in municipal wastewater treatment plants.

Antimicrobial resistance (AMR) represents a relentless, silent pandemic. Contributing to this are wastewater treatment plants (WWTPs), a potential source of antibiotic resistance genes' (ARGs) transmission to the environment, threatening public health. The presence of ARGs in pathogenic bacteria and their release into the environment by WWTPs threatens the public health. The current study investigated changes in ARGs' abundance in biological nutrient removal (BNR) processes and anaerobic digestion (AD) reactors of two WWTPs. Also, microbial community structure, which is known to shape the distribution and abundance of ARGs, was also analyzed. The relative abundance of eight ARGs (tetX, tetA, tetM, TEM, sul1, sul2, ermB and qnrD) was quantified as ARGs' copies/16 S rRNA gene copies using quantitative polymerase chain reaction (qPCR). Microbial community composition was assessed by 16 S rRNA microbiome sequencing analysis. TetX was prevalent among the eight ARGs, followed by TEM and sul1. However, its abundance was decreased in the AD sludges compared to BNR sludges. Proteobacteria was the major bacterial phylum found in all the sludge samples, while Arcobacter, 12up and Acidovorax were the predominant genera. Acinetobacter and Flavobacterium were significantly more abundant in the BNR sludges, while 12up and Aeromonas were predominant in AD sludges. Principal component analysis (PCA) revealed a clear difference in dominant ARGs and bacteria between the sludges in the processes of BNR and AD of the two WWTPs. Clinically relevant bacterial genera, Klebsiella and Enterococcus, found in both the BNR and AD sludges, were significantly correlated with the tetX gene. Throughout this study, the relationship between microbial communities and specific ARGs was revealed, illustrating that the composition of the microbial community could play a vital role in the abundance of ARGs. These results will better inform future studies aimed at controlling the spread of ARGs and their potential hosts from WWTPs.

New Insight on the Immune Modulation and Physical Barrier Protection Caused by Vitamin A in Fish Gills Infected With Flavobacterium columnare.

In this study, we have investigated the influence of vitamin A on gill barrier function of grass carp (Ctenopharyngodon idella) infected with Flavobacterium columnare. The fish were fed different concentrations of vitamin A diets for 10 weeks and then infected with F. columnare by immersion. We observed that optimal vitamin A significantly prevented gill rot morbidity in fish infected with F. columnare. Further investigations revealed that vitamin A boosted the gill immunity by increasing the contents of complements (C3 and C4), activities of acid phosphatase (ACP) and lysozyme, mRNAs of ß-defensin-1, liver-expressed antimicrobial peptide 2A and 2B (LEAP-2A and LEAP-2B), hepcidin, and anti-inflammatory cytokines like transforming growth factor ß1 (TGF-ß1), TGF-ß2, interleukin-10 (IL-10), and IL-11. It also enhanced the levels of various related signaling molecules including inhibitor protein κBα (IκBα), target of rapamycin (TOR), and ribosome protein S6 kinase 1 (S6K1) but downregulated the expression of pro-inflammatory cytokines including IL-1ß, IL-8, tumor necrosis factor α (TNF-α), and interferon γ2 (IFN-γ2) and related signaling molecules including nuclear factor κB p65 (NF-κB p65) (rather than NF-κB p52), IκB kinase ß (IKKß), IKKγ (rather than IKKα), eIF4E-binding protein 1 (4E-BP1), and 4E-BP2 mRNA levels in fish gills. In addition, dietary vitamin A markedly lowered the concentrations of reactive oxygen species (ROS), malondialdehyde (MDA), and protein carbonyl (PC), increased both the activities and mRNAs of copper/zinc superoxide dismutase (Cu/ZnSOD), MnSOD, glutathione transferases (GSTs), glutathione peroxidase (GPx), and glutathione reductase (GR) associated with upregulation of NF-E2-related factor 2 (Nrf2) mRNAs and downregulation of Kelch-like-ECH-associated protein (Keap1a) and Keap1b mRNAs. Moreover, vitamin A decreased the mRNAs of different apoptotic mediators [caspases 8, 9, 3 (rather than 7)] associated with downregulation of signaling molecule p38 mitogen-activated protein kinase (p38MAPK) mRNAs in fish gills. Besides, vitamin A promoted tight junction (TJ) complex mRNAs [including claudin-b, -c, -3, -7, -12, occludin, and zonula occludens-1 (ZO-1)] that have been linked to the downregulation of myosin light chain kinase (MLCK) signaling. Taken together, the current study demonstrated for the first time that vitamin A markedly enhanced gill health associated with immune modulation and physical barrier protection. Based on protecting fish against gill rot morbidity, ACP activity, and against lipid peroxidation, optimum vitamin A concentrations in on-growing grass carp (262-997 g) were found to be 1,991, 2,188, and 2,934 IU/kg diet, respectively.

A widely distributed phosphate-insensitive phosphatase presents a route for rapid organophosphorus remineralization in the biosphere.

The regeneration of bioavailable phosphate from immobilized organophosphorus represents a key process in the global phosphorus cycle and is facilitated by enzymes known as phosphatases. Most bacteria possess at least one of three phosphatases with broad substrate specificity, known as PhoA, PhoX, and PhoD, whose activity is optimal under alkaline conditions. The production and activity of these phosphatases is repressed by phosphate availability. Therefore, they are only fully functional when bacteria experience phosphorus-limiting growth conditions. Here, we reveal a previously overlooked phosphate-insensitive phosphatase, PafA, prevalent in Bacteroidetes, which is highly abundant in nature and represents a major route for the regeneration of environmental phosphate. Using the enzyme from Flavobacterium johnsoniae, we show that PafA is highly active toward phosphomonoesters, is fully functional in the presence of excess phosphate, and is essential for growth on phosphorylated carbohydrates as a sole carbon source. These distinct properties of PafA may expand the metabolic niche of Bacteroidetes by enabling the utilization of abundant organophosphorus substrates as C and P sources, providing a competitive advantage when inhabiting zones of high microbial activity and nutrient demand. PafA, which is constitutively synthesized by soil and marine flavobacteria, rapidly remineralizes phosphomonoesters releasing bioavailable phosphate that can be acquired by neighboring cells. The pafA gene is highly diverse in plant rhizospheres and is abundant in the global ocean, where it is expressed independently of phosphate availability. PafA therefore represents an important enzyme in the context of global biogeochemical cycling and has potential applications in sustainable agriculture.

Efficacy of ulvan on immune response and immuno-antioxidant gene modulation in Labeo rohita against columnaris disease.

This study reports the effect of ulvan enriched diet on the influence of growth, changes in hemato-biochemical indices, improvement of antioxidant system, enhancement of innate-adaptive immunity and modification of immuno-antioxidant genes expression in Labeo rohita against Flavobacterium columnaris. The weight gain (WG) was significantly high (P > 0.05) in unchallenged normal and challenged fish fed with diets enriched with 25 and 50 mg kg-1 ulvan; the FCR was better (P > 0.05) when fed with 50 mg kg-1 enriched diet. In normal fish fed with or without ulvan supplementation was noted 100% survival rate (SR). In both groups, the red blood cell (RBC) and while blood cell (WBC) counts increased significantly (P > 0.05) when fed with 50 mg kg-1 ulvan diet whereas the hemoglobin (Hb) level increased significantly on being fed with 25 and 50 mg kg-1 ulvan diets. The SOD activity was enhanced significantly in both groups fed with any dose of ulvan diets whereas the MDA and GPx activity increased only with 25 and 50 mg kg-1 ulvan diets. The phagocytic (PC) activity significantly increased with any enriched diet and control diet groups while the respiratory burst (RB) activity increased only with 50 mg kg-1 ulvan diet. The alternate complement pathway (ACP), activity of lysozyme (Lyz), and immunoglobuline M (IgM) were better in both groups fed with 50 mg kg-1 ulvan diet. The SOD and GPx antioxidant gene expression were significantly high in both groups fed with any ulvan diet while the Nrf2 gene expression was high with 50 mg kg-1 ulvan diet. The IL-1ß, TNFα, hepcidin, Lyz, and IgM cytokines or proteins mRNA expression were significant in both groups fed with all ulvan supplement diet whereas the ß-2M expression was significant only with 50 mg kg-1 ulvan diet. The present research indicates that both L. rohita groups fed with 50 mg kg-1 ulvan diet significantly improved growth, antioxidant system, immune defense system, and immuno-antioxidant related gene expression against F. columnaris.

Growth of Flavobacterium columnare Genomovars in the Presence or Lack of Supplemental Cations.

Flavobacterium columnare is a problematic pathogen for the aquaculture industry where isolates are classified by genomovars. Suspended growth in a low nutrient media, like tryptone yeast extract salts, is a common method used for laboratory study. The presence of calcium and magnesium is the factor contributing to growth, virulence, and biofilm formation for F. columnare. Exponential growth occurs within 24 h for F. columnare when grown in complete tryptone yeast extract salts medium at 30 ºC. Withholding CaCl2 and MgSO4 components from a complete TYES formulation reduced or completely inhibited growth of genomovar I isolates but not the growth of genomovar II, IIB, or III isolates. Only 3 of 20 genomovar I isolates, MS-FC-4, FC-CSF-53, and 023-08-3, could achieve O.D. 540 readings ≥ 0.3 but only after 48-h incubation in cation-restricted TYES. Independently adding CaCl2 or MgSO4 to tryptone and yeast extracts did not result in a genomovar-specific growth phenotype, but generally demonstrated increased clumping with individual isolates presenting abnormal growth. Clumping formed filamentous strings that migrated to the top of the culture tube when isolates were grown in TYE+CaCl2. Several of the F. columnare isolates from all the genomovars exhibited delayed growth when a single cation source was provided. This study demonstrates phenotypic differences between and within genomovars of a single bacterial species when grown under different TYES media conditions.

16S rRNA gene-based microbiome analysis identifies candidate bacterial strains that increase the storage time of potato tubers.

In the past, the potato plant microbiota and rhizosphere have been studied in detail to improve plant growth and fitness. However, less is known about the postharvest potato tuber microbiome and its role in storage stability. The storage stability of potatoes depends on genotype and storage conditions, but the soil in which tubers were grown could also play a role. To understand the ecology and functional role of the postharvest potato microbiota, we planted four potato varieties in five soil types and monitored them until the tubers started sprouting. During storage, the bacterial community of tubers was analysed by next-generation sequencing of the 16S rRNA gene amplicons. The potato tubers exhibited soil-dependent differences in sprouting behaviour. The statistical analysis revealed a strong shift of the tuber-associated bacterial community from harvest to dormancy break. By combining indicator species analysis and a correlation matrix, we predicted associations between members of the bacterial community and tuber sprouting behaviour. Based on this, we identified Flavobacterium sp. isolates, which were able to influence sprouting behaviour by inhibiting potato bud outgrowth.

Impact of grape pomace flour (GPF) on immunity and immune-antioxidant-anti-inflammatory genes expression in Labeo rohita against Flavobacterium columnaris.

This study evaluates the effects of dietary inclusion of grape pomace flour (GPF) on growth, antioxidant, anti-inflammatory, innate-adaptive immunity, and immune genes expression in Labeo rohita against Flavobacterium columnaris. In both normal and challenged fish the growth rate, hematology and biochemical parameters significantly increased when fed with 200 and 300 mg GPF enriched diets; similarly the activities of antioxidants and innate-adaptive immune parameters, such as malondialdehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase (GPx), glutathione (GSH), phagocytic (PC), respiratory burst (RB), alternative pathway complement (ACP), lysozyme (Lyz), and total immunoglobulin M (IgM) significantly increased in both groups. Similarly, the immune, antioxidant, and anti-inflammatory-related gene mRNA expression was significantly up-regulated in head kidney (HK) tissues. The challenged fish fed without GPF always exhibited lower values of all the studied parameters. The results indicate that both normal and challenged fish treated with 200 mg GPF inclusion diet had significantly enhanced growth rate, antioxidant status, and immune defense mechanisms than with 300 mg GPF diet in L. rohita against F. columnaris.

The regulatory effects of pyridoxine deficiency on the grass carp (Ctenopharyngodon idella) gill barriers immunity, apoptosis, antioxidant, and tight junction challenged with Flavobacterium columnar.

The effects of dietary pyridoxine (PN) on the gill immunity, apoptosis, antioxidant and tight junction of grass cap (Ctenopharyngodon idella) were investigated in this study. Fish were fed semi-purified diets containing graded levels of PN for 10 weeks, and then challenged with Flavobacterium columnare by bath immersion exposure for 3 days. The results indicated that compared with the optimal PN level, PN deficiency resulted in a decline in the antimicrobial compound production of gill. In addition, PN deficiency up-regulated the pro-inflammatory cytokines and down-regulated the anti-inflammatory cytokines gene expression, which might be associated with the enhanced nuclear factor κB p65 and the inhibited target of rapamycin signalling pathways, respectively, suggesting that PN deficiency could impair gill immune barrier function. Furthermore, PN deficiency (1) induced cell apoptosis, which may be partly associated with the (apoptotic protease activating factor-1, Bcl-2 associated X protein)/caspase-9 and c-Rel/tumor necrosis factor α (rather than FasL)/caspase-8 mediated apoptosis pathway. (2) Inhibited Kelch-like ECH-associating protein 1a/NF-E2-related factor 2 mRNA expression, decreased the mRNA expression and activities of antioxidant enzymes, increased the levels of reactive oxygen species, protein carbonyl and malondialdehyde. (3) Increased the mRNA expression level of myosin light chain kinase, which may be result in the down-regulation of tight junction complexes such as zonula occludens 1, occludin and claudins (expect claudin-12 and claudin-15). These results suggest that PN deficiency could impair gill physical barrier function. In summary, dietary PN deficiency could cause the impairment of gill barrier function associated with immunity, apoptosis, antioxidant and tight junction, which may result in the increased the susceptibility of fish to pathogenic bacteria. Moreover, based on the gill rot morbidity, LZ activity and MDA content, the dietary PN requirements for grass cap were estimated to be 4.85, 4.78 and 4.77 mg kg-1 diet, respectively.

Administration of dietary recombinant hepcidin on grass carp (Ctenopharyngodon idella) against Flavobacterium columnare infection under cage aquaculture conditions.

Hepcidin links iron metabolism with innate immunity during the inhibition of bacterial infection. Our previous studies had shown that recombinant hepcidin can significantly reduce the mortality rate of Ctenopharyngodon idella infected with Flavobacterium columnare under laboratory conditions. Here, we studied the preventive and therapeutic effects of feed supplemented with different doses of recombinant hepcidin on F. columnare-challenged C. idella reared in a cage culture environment. The results showed that in the prevention groups, 30 and 90 mg/kg of added purified and unpurified hepcidin respectively resulted in a higher survival rate in the early post-infection period, while 60 mg/kg of purified hepcidin significantly improved the survival rate in the therapy group (all compared to the control group). In the hepatopancreas, the expression of hepcidin and ferritin was significantly up-regulated, and the levels of ferroportin and serum iron were significantly decreased, especially in the therapy group. In addition, the expression of iron-related genes in spleen and intestine exhibited a similar trend to that in hepatopancreas. Meanwhile, immune genes were up-regulated to varying degrees, and the therapy group exhibited a significantly improved expression of pro-inflammatory cytokines and specific immunity. In summary, our study shows that different doses of recombinant hepcidin had protective effects against bacterial infection by regulating the iron distribution and immune gene expression, which provides a strong foundation for the application of recombinant hepcidin in aquaculture.

Dietary Glycyrrhiza uralensis extracts supplementation elevated growth performance, immune responses and disease resistance against Flavobacterium columnare in yellow catfish (Pelteobagrus fulvidraco).

The present study was conducted to evaluate the influence of Glycyrrhiza uralensis (G. uralensis) extracts on the growth performance, histological structure, immune response and disease resistance against Flavobacterium columnare (F. columnare) of yellow catfish. Fish were fed with two different diets, i.e., basal diet as control group (CG) and diet containing G. uralensis extracts as experimental group (GG). After 60 days feeding, growth performance of GG fish was significantly improved, with increased WG and SGR but decreased FCR compared to CG fish. Fish were then challenged with F. columnare for two times, as fish showed rare mortality after the first infection. GG fish showed significantly lower cumulative mortality during F. cloumnare infection than CG fish after 21 days infection (dpi). Epithelial cell exfoliation and obvious cellular vacuolization in the skin and congestion of gill lamellae were detected in CG fish, while GG fish showed increased width of epidermis and mucous cells number in skin, and increased length of secondary lamina in gill. GG fish also exhibited higher enzyme activity of lysozyme in serum and mRNA expression of lysozyme in head kidney than CG fish at most time points post infection. G. uralensis extracts supplementation also induced earlier serum anti-oxidative responses, with increased superoxide dismutase activity and total antioxidant capacity in GG fish at 1 dpi. Compared to CG fish, GG fish showed increased expression level of genes involved in TLRs-NFκB signaling (TLR2, TLR3, TLR5, TLR9, Myd88, and p65NFκB), resulting in higher expression levels of pro-inflammatory cytokines (IL-1ß and IL-8) in the head kidney post infection. However, these genes showed deviation in the gill of GG fish, which increased at some time points but decreased at other time points. Moreover, G. uralensis extracts supplementation also significantly unregulated the expression levels of IgM and IgD in head kidney, and the expression levels of IgM in the gill of yellow catfish, suggesting the elevated humoral immune response during F. columnare infection. All these results contributed to the elevated disease resistance ability against F. cloumnare infection of yellow catfish after dietary G. uralensis extracts supplementation.

Pathogen-induced activation of disease-suppressive functions in the endophytic root microbiome.

Microorganisms living inside plants can promote plant growth and health, but their genomic and functional diversity remain largely elusive. Here, metagenomics and network inference show that fungal infection of plant roots enriched for Chitinophagaceae and Flavobacteriaceae in the root endosphere and for chitinase genes and various unknown biosynthetic gene clusters encoding the production of nonribosomal peptide synthetases (NRPSs) and polyketide synthases (PKSs). After strain-level genome reconstruction, a consortium of Chitinophaga and Flavobacterium was designed that consistently suppressed fungal root disease. Site-directed mutagenesis then revealed that a previously unidentified NRPS-PKS gene cluster from Flavobacterium was essential for disease suppression by the endophytic consortium. Our results highlight that endophytic root microbiomes harbor a wealth of as yet unknown functional traits that, in concert, can protect the plant inside out.

Transcriptome analysis of the fish pathogen Flavobacterium columnare in biofilm suggests calcium role in pathogenesis.

BACKGROUND: Flavobacterium columnare is the causative agent of columnaris disease that affects cultured freshwater fishes worldwide. F. columnare easily colonizes surfaces by forming biofilm, which helps the pathogen resist antibiotic and disinfectant treatments. Previously, we had shown that increasing concentrations of calcium (Ca2+) promoted biofilm formation by F. columnare. The objective of this study was to further characterize the role of Ca2+ on biofilm formation and to compare the transcriptome profiles of planktonic and biofilm cells. RESULTS: RNA-Seq analysis was conducted to identify genes that were differentially expressed between the following states: i) planktonic cells in control medium (P), ii) planktonic cells in calcium-enriched medium (P/Ca), and iii) biofilm cells in calcium-enriched medium (B/Ca). Overall, we identified 441 significant (FDR-adjusted p < 0.05, fold change > 2) differentially expressed genes (DEGs) between P and B/Ca samples; 112 significant DEGs between P/Ca and B/Ca samples, and 175 significant DEGs between P/Ca and P samples, corresponding to 15.87, 4.03 and 6.30% of the total protein-coding sequences, respectively. The significant DEGs fell into different functional categories including iron acquisition, oxidative stress response, extracellular protein secretion, and respiratory metabolism. CONCLUSIONS: Our results posit Ca2+ as a critical signal in regulating bacterial surface adhesion and biofilm formation in F. columnare. Living in biofilm elicited a shift in several metabolic pathways that allowed the cells to cope with oxidative stress and nutrient starvation. In addition, Ca2+ supplementation induced the expression of putative virulence factors in F. columnare, such as extracellular protein secretion and iron acquisition.

Erythromycin and florfenicol treatment of rainbow trout Oncorhynchus mykiss (Walbaum) experimentally infected with Flavobacterium psychrophilum.

Flavobacterium psychrophilum is responsible for significant economic losses in rainbow trout aquaculture. Antimicrobial treatment remains the primary means of control; however, there are limited choices available for use. The objectives of the study were therefore to determine the minimum inhibitory concentrations for erythromycin and florfenicol in selected F. psychrophilum isolates and to evaluate their clinical treatment efficacy in experimentally infected rainbow trout. All isolates tested had moderate susceptibility to florfenicol and erythromycin except one isolate, which had low susceptibility to erythromycin. Two isolates (one with moderate and one with low susceptibility to erythromycin) were used in an experimental infection trial. Rainbow trout juveniles were injected intraperitoneally with 108 cfu/fish and after mortality had begun, fish were given erythromycin- and florfenicol-medicated feed at a rate of 75 mg kg- 1 day- 1 and 10 mg kg- 1  day- 1 fish body weight, respectively, for 10 consecutive days. The splenic F. psychrophilum load was determined using an rpoC quantitative PCR throughout the 30-day trial. Relative to antibiotic-free controls, erythromycin treatment significantly (p < 0.05) reduced mortality of rainbow trout juveniles infected with FPG101, even when treatment was initiated after clinical signs developed.

Efficacy and safety of a non-mineral oil adjuvanted injectable vaccine for the protection of Atlantic salmon (Salmo salar L.) against Flavobacterium psychrophilum.

Flavobacterium psychrophilum is the causative agent of Rainbow Trout Fry Syndrome which has had a major impact on global salmonid aquaculture. Recent outbreaks in Atlantic salmon in Scotland and Chile have added to the need for a vaccine to protect both salmon and trout. At present no licensed vaccines are available in Europe, leaving antibiotics as the only course of action to contain disease outbreaks. Outbreaks generally occur in fry at temperatures between 10 and 15 °C. Recently outbreaks in larger fish have given added impetus to the development of a vaccine which can provide long term protection from this highly heterogeneous pathogen. Most fish injectable vaccines are formulated with oil emulsion adjuvants to induce strong and long lasting immunity, but which are known to cause side effects. Alternative adjuvants are currently sought to minimise these adverse effects. The current study was performed to assess the efficacy of a polyvalent, whole cell vaccine containing formalin-inactivated F. psychrophilum to induce protective immunity in Atlantic salmon. The vaccine was formulated with an adjuvant containing squalene and aluminium hydroxide, and was compared to a vaccine formulated with a traditional oil adjuvant, Montanide ISA 760VG, and a non-adjuvanted vaccine. Duplicate groups of salmon (23.5 ± 6.8 g) were vaccinated with each of the vaccine formulations or phosphate buffered saline by intraperitoneal injection. Fish were challenged by intramuscular injection with F. psychrophilum six weeks post-vaccination to test the efficacy of the vaccines. Cumulative mortality reached 70% in the control salmon, while the groups of salmon that received vaccine had significantly lower mortality than the controls (p = 0.0001), with no significant difference in survival between vaccinated groups. The squalene/alum adjuvant was safe, more readily metabolised by the fish and induced less histopathological changes than the traditional oil adjuvant.

Biotransformation of ginsenoside Rb1 to Gyp-XVII and minor ginsenoside Rg3 by endophytic bacterium Flavobacterium sp. GE 32 isolated from Panax ginseng.

The rare ginsenoside Rg3 is attracting more attention because of its good physiological activity and urgent need. There are many pathways to obtain ginsenoside Rg3, including chemical and biological methods. Among these, the conversion of the protopanaxadiol-type ginsenosides by microbial hydrolysis is a trend due to its high efficiency and mild conditions. For effectively extracting from the other panaxadiol saponins, the conversion process for ginsenoside Rg3 was investigated using ß-glycosidase-producing endophytic fungus in Panax ginseng in this study. The metabolic pathways are as follows: ginsenoside Rb1 â†’ Gyp-XVII and ginsenoside Rb1 â†’ ginsenoside Rd â†’ ginsenoside Rg3. Phylogenetic analysis of 16S rDNA gene sequence, showed that GE 32 strain belonged to Flavobacterium species. These results suggest that the process of rare ginsenoside Rg3 production by endophytic bacteria GE 32 is efficient for the industrial production and application. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report on cultivable ß-glycosidase-producing endophytic bacteria from Panax ginseng. Flavobacterium sp. GE32 could convert major ginsenoside Rb1 into Gyp-XVII and minor ginsenoside Rg3. Strain GE 32 has potential to be applied on the preparation for minor ginsenoside Rg3 in pharmaceutical industry.

Antibacterial Activities of Metabolites from Vitis rotundifolia (Muscadine) Roots against Fish Pathogenic Bacteria.

Enteric septicemia of catfish, columnaris disease and streptococcosis, caused by Edwardsiella ictaluri, Flavobacterium columnare and Streptococcus iniae, respectively, are the most common bacterial diseases of economic significance to the pond-raised channel catfish Ictalurus punctatus industry. Certain management practices are used by catfish farmers to prevent large financial losses from these diseases such as the use of commercial antibiotics. In order to discover environmentally benign alternatives, using a rapid bioassay, we evaluated a crude extract from the roots of muscadine Vitis rotundifolia against these fish pathogenic bacteria and determined that the extract was most active against F. columnare. Subsequently, several isolated compounds from the root extract were isolated. Among these isolated compounds, (+)-hopeaphenol (2) and (+)-vitisin A (3) were found to be the most active (bacteriostatic activity only) against F. columnare, with 24-h 50% inhibition concentrations of 4.0 ± 0.7 and 7.7 ± 0.6 mg/L, respectively, and minimum inhibitory concentrations of 9.1 ± 0 mg/L for each compound which were approximately 25X less active than the drug control florfenicol. Efficacy testing of 2 and 3 is necessary to further evaluate the potential for these compounds to be used as antibacterial agents for managing columnaris disease.

Antibacterial Activity of Constituents from Mangosteen Garcinia mangostana Fruit Pericarp against Several Channel Catfish Pathogens.

Bacterial diseases cause major financial damage to the producers of Channel Catfish Ictalurus punctatus in the southeastern USA. The two most common bacterial diseases among pond-raised Channel Catfish are enteric septicemia of catfish, caused by the gram-negative bacterium Edwardsiella ictaluri, and columnaris disease, caused by the rod-shaped, gram-negative bacterium Flavobacterium columnare. Streptococcosis is another, less-common bacterial disease in catfish and is caused by the gram-positive coccus Streptococcus iniae. Catfish farmers typically rely on commercial antibiotics and other chemicals to prevent the economic damage from these diseases. Environmentally benign and efficacious alternatives to the currently used antibiotics and chemicals will tremendously help the catfish aquaculture industry. As part of our ongoing efforts in the search for such novel compounds, we investigated ethyl acetate and methanol extracts of mangosteen Garcinia mangostana fruit pericarp via bioassay-guided fractionation. Gamma-mangostin (γ-mangostin) was isolated and identified as the most promising active metabolite against F. columnare. One of the constituents in the mangosteen fruit pericarp, alpha-mangostin (α-mangostin), is the major xanthone; α-mangostin was found to be 10-fold less active than γ-mangostin when minimum inhibitory concentration values were compared.