Antimicrobial effect of Moringa oleifera seed powder against Vibrio cholerae isolated from the rearing water of shrimp (Penaeus vannamei) postlarvae

Shrimp farming has experienced rising costs as a result of disease outbreaks associated with Vibrio spp. Suitable strategies for disease prevention and control are therefore urgently needed. This study aimed to evaluate the antimicrobial effect of Moringa oleifera seed powder against Vibrio cholerae in the rearing water of Pacific white shrimp (Penaeus vannamei) postlarvae. In vitro assays included the determination of minimum inhibitory concentration (MIC) of M. oleifera seed powder against V. cholerae, whereas in vivo assays included the effect of M. oleifera seed powder on bacterial load and water quality parameters in the rearing tanks, as well as its effect on shrimp postlarvae survival. M. oleifera seed powder inhibited the growth of V. cholerae with MIC values of 62·5 µg ml⁻¹. Moreover, seawater pH of treated tanks (8·66) was significantly lower (P < 0·01) than pH of the control tanks (9·02), whereas the visibility of treated tanks (37·08 cm) was significantly higher (P < 0·01) as compared to control tanks (35·37 cm). Likewise, V. cholerae load was significantly reduced (P < 0·01) from 4·7 × 10⁴ to 3·1 × 10³ CFU per ml in tanks treated with M. oleifera seed powder. Altogether, this study demonstrates the antimicrobial activity of M. oleifera against V. cholerae in shrimp culture.     

The effects of copper on photosynthesis and biomolecules yield in Chlorolobion braunii

Our knowledge of the effects of copper on microalgal physiology is largely based on studies conducted with high copper concentrations; much less is known when environmentally relevant copper levels come into question. Here, we evaluated the physiology of Chlorolobion braunii exposed to free copper ion concentrations between 5.7 × 10^?9 and 5.0 × 10^?6 mol · L^?1, thus including environmentally relevant values. Population growth and maximum photosynthetic quantum yield of PSII were determined daily during the 96 h laboratory controlled experiment. Exponentially‐growing cells (48 h) were analyzed for effective quantum yield and rapid light curves (RLC), and total lipids, proteins, carbohydrates, chlorophyll a and carotenoids were determined. The results showed that growth rates and population density decreased gradually as copper increased in experiment, but the photosynthetic parameters (maximum and effective quantum yields) and photochemical quenching (qP) decreased only at the highest free copper concentration tested (5.0 × 10^?6 mol · L^?1); nonphotochemical quenching (NPQ) increased gradually with copper increase. The RLC parameters Ek and rETRmax were inversely proportional to copper concentration, while α and Im decreased only at 5.0 × 10^?6 mol · L^?1. The effects of copper in biomolecules yield (mg · L^?1) varied depending on the biomolecule. Lipid yield increased at free copper concentration as low as 2.5 × 10^?8 mol · L^?1, but proteins and carbohydrates were constant throughout.     

E. coli Nissle microencapsulation in alginate-chitosan nanoparticles and its effect on Campylobacter jejuni in vitro

Microencapsulation enhances the oral delivery of probiotic bacteria. In this study, the probiotic Escherichia coli Nissle 1917 (EcN) was microencapsulated using alginate and chitosan nanoparticles. The result showed 90% encapsulation yield of EcN, and the encapsulated EcN displayed significantly (P < 0.05) increased survival in low pH (1.5), high bile salt concentration (4%), and high temperature (70 °C). The most effective cryopreservatives of EcN during freezing and thawing was skim milk and sucrose. Exposure to microencapsulated EcN significantly (P < 0.05) reduced the Campylobacter jejuni growth by 2 log CFU. The rate of EcN release from microcapsule was 9.2 × 10⁵ cell min⁻¹, and the appropriate model to describe its release kinetics was zero order. Importantly, the entrapment of EcN inside the microcapsule did not eliminate the exterior diffusion of EcN produced antioxidant compounds. In addition, the EcN microcapsule efficiently adhered to intestinal HT-29 cells and the pre-treatment of HT-29 cells with EcN-microcapsule for 4 h significantly (P < 0.05) reduced the invasion (1.9 log) of C. jejuni; whereas, completely abolished the intracellular survival. Furthermore, HT-29 cells pre-treated with encapsulated EcN in PCR array showed decreased expression (> 1.5-fold) of genes encoding chemokines, toll-like receptors, interleukins, and tumor necrosis factors. In conclusion, the alginate-chitosan microcapsule can provide effectual platform to deliver probiotic EcN and thereby can reduce the Campylobacter infection in chickens and humans.

     

Validation and identification of molecular markers linked to the leaf rust resistance gene <Emphasis Type="Italic">Lr</Emphasis>28 in wheat

Leaf rust resistance gene Lr28 controls one of the important resistances in the Indian subcontinent against the most prevalent Puccinia triticina pathotype 77-5. Pyramiding Lr28 with other resistance genes would therefore, provide durable resistance against rust, a process that can be facilitated by DNA markers. A microsatellite marker wmc313 linked to Lr28 at a distance of 5.0 cM was identified in the population HD2329 × HW2037. The marker was validated in another population developed from WL711× CS + Lr28: 2D/2M 3/8 (acc. 2956) as well as in a few near- isogenic lines (NILs) carrying gene Lr28. Compared to the previously reported marker TPSCAR SCS421₅₇₀, wmc313 is more closely linked to Lr28. Both these markers flanking the gene should be useful in the deployment of Lr28 into the breeding program using marker-assisted selection allowing pyramiding with other effective genes to confer durable resistance.     

Reconstitution conditions for dried probiotic powders represent a critical step in determining cell viability

Aims: Resuscitation of dried cultures represents a critical control point in obtaining active and effective probiotic strains. This study examined the effects of various rehydration conditions on the viability of Bifidobacterium longum NCC3001 and Lactobacillus johnsonii La1. Methods and Results: Reconstitution conditions for these strains were optimized using a multivariate experimental design approach. Furthermore, using flow cytometry, the cell integrity was followed during reconstitution. By adjusting the pH, availability of a metabolizable sugar, reconstitution duration, powder matrix and ratio of powder to reconstitution solution, the recovery of Bif. longum NCC3001 and Lact. johnsonii La1 following reconstitution was increased eight‐ and two‐fold, respectively, over standard reconstitution in maximum recovery diluent. It was shown that pH had a significant effect on the recovery of Bif. longum NCC3001 and Lact. johnsonii La1. Conclusions: The recovery of dried probiotic cultures is greatly dependent on the reconstitution conditions. The maximum recovery of 11·7 ¹⁰log CFU g^?1Bif. longum NCC3001 was achieved at 30‐min reconstitution at pH 8, in the presence of 2%l ‐arabinose and a ratio of 1 : 100 of powder to diluent. Lact. johnsonii La1 showed highest recovery (9·3 ¹⁰log CFU g^?1) after reconstitution, when mixed with maltodextrin at pH 4. Significance and Impact of the Study: To achieve accurate viable probiotic numbers from dried probiotic cultures, the reconstitution conditions should be optimized for the strain used.     

Beneficial activity of Enterococcus faecalis CECT7121 in the anti‐lymphoma protective response

Aims: To study the anti‐tumour effects of Enterococcus faecalis CECT7121 on LBC cells, an aggressive murine T‐cell lymphoma that kills the host in 18 days when is intraperitoneally (i.p.) administrated. Methods and Results: In vitro studies have shown that LBC cell proliferation was inhibited by Ent. faecalis CECT7121 stimulus in a dose‐dependent manner, inducing apoptosis. The production of ceramide was involved in the latter effect. To undertake in vivo studies, syngeneic BALB/c mice pre‐treated i.p. with Ent. faecalis CECT7121 (2·5 × 10⁸ CFU) were challenged i.p. with LBC cells (1·0 × 10⁶ cells) the day after. On day 30 post‐inoculation of LBC cells, 70% of Ent. faecalis CECT7121 pre‐treated mice survived, whereas no survivals were recorded in the control group. A group of surviving mice was re‐challenged with LBC cells, and 89% of them survived. Upon stimulation with irradiated LBC cells, spleen cell proliferation, high IFNγ, IL‐12 and IL‐10 levels were observed in surviving animals. Conclusions: Enterococcus faecalis CECT7121 affected multiple factors of the tumour establishment by the following methods: down‐regulating the LBC cell proliferation and inducing apoptosis in these cells; and enhancing the immune response that protects animals from lymphoma challenge and re‐challenge. Significance and Impact of the Study: This study demonstrate that Ent. faecalis CECT7121 has potential as a probiotic that could facilitate the development of novel complements to therapeutic strategies against oncological diseases.     

In vitro lung delivery of bacteriophages KS4‐M and ΦKZ using dry powder inhalers for treatment of Burkholderia cepacia complex and Pseudomonas aeruginosa infections in cystic fibrosis

Aims: To determine the feasibility of formulating and aerosolizing powders containing bacteriophages KS4‐M and ΦKZ for lung delivery and treatment of pulmonary Burkholderia cepacia complex and Pseudomonas aeruginosa infections. Methods and Results: Endotoxin‐removed bacteriophages KS4‐M and ΦKZ were lyophilized in lactose/lactoferrin 60 : 40 w/w matrix and deagglomerated in a mixer mill (without beads) to formulate respirable powders. The powders were then aerosolized using an Aerolizer^® capsule inhaler. Mass median aerodynamic diameter (MMAD) of this inhalable aerosol was determined using Andersen cascade impactor at 60 l min^?1. Measured MMAD for both types of powders was 3·4 μm, and geometric standard deviation was 1·9–2·0. Viability of bacteriophages delivered distal to an idealized mouth‐throat replica was determined from bioassays of samples collected on filters placed after the idealized replica. As a percentage of inhaler load, amount of powder delivered distal to the mouth‐throat replica, which is a measure of lung delivery, was 33·7 ± 0·3% for KS4‐M and 32·7 ± 0·9% for ΦKZ. Titres collected downstream of the mouth throat were (3·4 ± 2·5) × 10⁶ PFU for KS4‐M with an Aerolizer capsule load of (9·8 ± 4·8) × 10⁶ and (1·9 ± 0·6) × 10⁷ for ΦKZ with an Aerolizer capsule load of (6·5 ± 1·9) × 10⁷. Conclusions: Bacteriophages KS4‐M and ΦKZ can be lyophilized without significant loss of viability in a lactose/lactoferrin 60 : 40 w/w matrix. The resulting powders can be aerosolized to deliver viable bacteriophages to the lungs. Significance and Impact of the Study: Development of lactoferrin‐based bacteriophage aerosol powders solidifies the ground for future research on developing novel formulations as an alternative to inhaled antibiotic therapy in patients with cystic fibrosis.     

Postprocessing microflora of commercial attieke (a fermented cassava product) produced in the south of Côte d'Ivoire

The distribution and presence of hygiene indicator and pathogenic micro‐organisms in 375 samples of attieke marketed in Côte d'Ivoire, and their roles in the food poisoning were evaluated. Microbiological analyses were carried out, which included the total viable bacteria, coliforms, Escherichia coli, Staphylococcus aureus, Salmonella, Bacillus spores, fungi and Clostridium perfringens. The results revealed that the viable bacteria counts ranged from 2·2 ± 1·2 × 10⁵ to 3·4 ± 1·4 × 10⁶ CFU g^?1, while the yeasts and the moulds counts ranged, respectively, from 2·4 ± 0·12 × 10⁴ to 9·8 ± 0·4 × 10⁵ CFU g^?1 and 1·3 ± 0·7 × 10¹ to 1·7 ± 0·7 × 10² CFU g^?1. Bacillus subtilis, Bacillus cereus, Staphylococcus aureus, Citrobacter freundi, Enterobacter amnigenus, Citrobacter youngae, Enterobacter aerogenes, Klebsiella pneumoniae, Serratia marcescens, Enterobacter agglomerans and Klebsiella oxytoca were the bacteria isolated, and Rhizopus spp., Mucor spp., Thamnidium spp., Fusarium spp., Moniliella spp. the fungi. Escherichia coli, Clostridium perfringens and Salmonella spp. were not found. The occurrence of some bacteria and fungi illustrate that attieke collected in Côte d'Ivoire markets may act as a reservoir of pathogenic micro‐organisms for human.

Significance and Impact of Study

This study demonstrates the great need to carry out microbiological tests frequently on attieke and even more the need to apply correct HACCP system during the production. Attieke is especially a well‐known product in West Africa; hence, it is extremely important to ensure an adequate microbiological quality to guarantee consumers health. Overall, the study highlighted the need for effective communication on microbiological food risks, proper instruction and supervision in food‐handling procedures, greater education on food safety risks.     

Dietary Administration of Probiotic Aeromonas veronii V03 on the Modulation of Innate Immunity,Expression of Immune-Related Genes and Disease Resistance Against Aeromonas hydrophila Infection in Common Carp (Cyprinus carpio)

This study investigates the effects of dietary Aeromonas veronii V03 supplementation on growth performances, innate immunity, and expression of immune-related genes in lymphoid organs of Cyprinus carpio and resistance to Aeromonas hydrophila infection. Fish were fed for 4 weeks with basal diet (BD; without probiotic), and experiment diet containing different doses of A. veronii V03 at 3.2 × 10⁷ (DI) and 3.5 × 10⁹ (DII) CFU g⁻¹ of diet. At the end of the probiotic feeding trial, fish were challenged with A. hydrophila, and the percentage of survival rates was recorded over 7 days. Results revealed that fish fed with A. veronii V03 demonstrated a significant improvement in growth and enhancement of innate immunity, including respiratory burst, myeloperoxidase, and lysozyme activities, and total immunoglobulin level compared with BD fed to fish. Relatively, expression of cytokines (MyD88, IL-1β1, IL-8, and IL-10) and c- and g-type lysozymes were significantly up- and downregulated in lymphoid organs of fish. Moreover, dietary supplementation of A. veronii V03 exhibited significantly (p < 0.001) higher survival rates of DI (90%) and DII (96.66%) compared with BD (53.33%) fed fish against A. hydrophila infection. These findings help to understand the effects of probiotic A. veronii V03 administrated feed influences on growth and ailment resistance to A. hydrophila infection by regulating innate and systemic immunity in common carp fish.

     

Cloning and characterization of a thermostable and halo-tolerant endoglucanase from <Emphasis Type="Italic">Thermoanaerobacter tengcongensis</Emphasis> MB4

A β-1,4-endoglucanase (Cel5A) was cloned from the genomic DNA of saccharolytic thermophilic eubacterium Thermoanaerobacter tengcongensis MB4 and functionally expressed in Escherichia coli. Substrate specificity analysis revealed that Cel5A cleaves specifically the β-1,4-glycosidic linkage in cellulose with high activity (294 U mg⁻¹; carboxymethyl cellulose sodium (CMC)). On CMC, kinetics of Cel5A was determined (K _m 1.39 ± 0.12 g l⁻¹; k _cat/K _m 1.41 ± 0.13 g⁻¹ s⁻¹). Cel5A displays an activity optimum between 75 and 80 °C. Residues Glu187 and Glu289 were identified as key catalytic amino acids by sequence alignment. Interestingly, derived from a non-halophilic bacterium, Cel5A exhibits high residual activities in molar concentration of NaCl (3 M, 49.3%) and KCl (4 M, 48.6%). In 1 M NaCl, 82% of Cel5A activity is retained after 24 h incubation. Molecular Dynamics studies performed at 0 and 3 M NaCl, correlate the Cel5A stability to the formation of R-COO⁻···Na⁺ ···⁻OOC-R salt bridges within the Cel5A tertiary structure, while activity possibly relates to the number of Na⁺ ions trapped into the negatively charged active site, involving a competition mechanism between substrate and Na⁺. Additionally, Cel5A is remarkably resistant in ionic liquids 1-butyl-3-methyllimidazolium chloride (1 M, 54.4%) and 1-allyl-3-methylimidazolium chloride (1 M, 65.1%) which are promising solvents for cellulose degradation and making Cel5A an attractive candidate for industrial applications.     

Absolute configuration and biological profile of pyrazoline enantiomers as MAO inhibitory activity

A new racemic pyrazoline derivative was synthesized and resolved to its enantiomers using analytic and semipreparative high‐pressure liquid chromatography. The absolute configuration of both fractions was established using vibrational circular dichroism. The in vitro monoamine oxidase (MAO) inhibitory profiles were evaluated for the racemate and both enantiomers separately for the two isoforms of the enzyme. The racemic compound and both enantiomers were found to inhibit hMAO‐A selectively and competitively. In particular, the R enantiomer was detected as an exceptionally potent and a selective MAO‐A inhibitor (K_i = 0.85 × 10^?3 ± 0.05 × 10^?3 μM and SI: 2.35 × 10^?5), whereas S was determined as poorer compound than R in terms of K_i and SI (0.184 ± 0.007 and 0.001). The selectivity of the enantiomers was explained by molecular modeling docking studies based on the PDB enzymatic models of MAO isoforms.     

Resistance to glyphosate in the cyanobacterium <Emphasis Type="Italic">Microcystis aeruginosa</Emphasis> as result of pre-selective mutations

Adaptation of Microcystis aeruginosa (Cyanobacteria) to resist the herbicide glyphosate was analysed by using an experimental model. Growth of wild-type, glyphosate-sensitive (G^s) cells was inhibited when they were cultured with 120 ppm glyphosate, but after further incubation for several weeks, occasionally the growth of rare cells resistant (G^r) to the herbicide was found. A fluctuation analysis was carried out to distinguish between resistant cells arising from rare spontaneous mutations and resistant cells arising from other mechanisms of adaptation. Resistant cells arose by rare spontaneous mutations prior to the addition of glyphosate, with a rate ranging from 3.1 × 10⁻⁷ to 3.6 × 10⁻⁷ mutants per cell per generation in two strains of M. aeruginosa; the frequency of the G^r allele ranged from 6.14 × 10⁻⁴ to 6.54 × 10⁻⁴. The G^r mutants are slightly elliptical in outline, whereas the G^s cells are spherical. Since G^r mutants have a diminished growth rate, they may be maintained in uncontaminated waters as the result of a balance between new resistants arising from spontaneous mutation and resistants eliminated by natural selection. Thus, rare spontaneous pre-selective mutations may allow the survival of M. aeruginosa in glyphosate-polluted waters via G^r clone selection.     

Implication of storage conditions on luminescence response from Photobacterium phosphoreum in free and immobilized form

Bioluminescent bacteria in the form of a cell suspension for on-site hazard analysis are not suitable as in vivo luminescence in free cells fluctuates and may lead to erroneous results. Furthermore, the culture broth cannot be stored for long durations to continue sensing analytes as the luminescence ceases over time. Factors that affect luminescence response include growth dynamism, and ambient environmental conditions. The present study investigated the effect of storage conditions such as temperature (25 ± 2°C, room temperature; 4°C; and −20°C) and ambient aqueous environment (M1: sucrose, 1.02 M; M2, bioluminescent media [tryptone, 10 g L⁻¹; NaCl, 28.5 g L⁻¹; MgCl₂.7H₂O, 4.5 g L⁻¹; CaCl₂, 0.5 g L⁻¹; KCl 0.5 g L⁻¹; yeast extract, 1 g L⁻¹; H₂O, 1 L]; M3, bioluminescent media and 95% glycerol, 1:1 ratio) on the luminescence emission from the calcium alginate-immobilized Photobacterium phosphoreum (S_b) against the cells in free suspension for an extended period. The results indicated that both the parameters that were undertaken markedly affected the luminescence. In the study, S_b showed an enhanced luminescence emission than the control up to 18.5-fold and for a prolonged period which can be efficiently utilized for rapid biosensing of hazardous materials.     

Production and regeneration of protoplasts from <Emphasis Type="Italic">Grateloupia turuturu</Emphasis> Yamada (Rhodophyta)

Protoplasts were isolated enzymatically from the carrageenophyte red alga Grateloupia turuturu (Halymeniales, Rhodophyta) that occurs along the coast of the French Channel in Normandy. Effects of the main factors on the protoplast yield were identified to improve the isolation protocol. The optimal enzyme composition for cell wall digestion and protoplast viability consisted of 2% cellulase Onozuka R-10, 0.5% macerozyme R-10, 2% crude extract from viscera of Haliotis tuberculata, 0.8 M mannitol, 20 mM sodium citrate, 0.3% bovine serum albumin at 25°C, and 4-h incubation period. The protoplasts were approximately 5–15 μm in diameter, liberated mainly from the surface cell layers. Maximum yield was 1.5 × 10⁷ protoplasts g^-1 fresh tissue. The protoplasts underwent initial division after 14 days with a high density level of 1 × 10⁶ cells mL^-1 in culture medium and developed into microthalli of a line of two to six cells.     

Toxicity assessment of paraverine hydrochloride and papaverine-derived metabolites in primary cultures of rat hepatocytes

Summary The present study was undertaken to assess and compare the toxic effects of papaverine hydrochloride and its metabolites. Primary cell cultures of rat hepatocytes were treated with papavarine (papaver), 3′-O-desmethyl (3′-OH), 4′-O-desmethyl (4′-OH), and 6-O-desmethyl (6-OH) papaverine at 1×10⁻⁵, 1×10⁻⁴, and 1×10⁻³ M for 4,8, 12, and 24-h periods. Cell injury was determined by: a) cell viability using the trypan blue exclusion test; b) cytosolic enzyme leakage of lactate dehydrogenase and aspartate aminotransferase; c) morphologic alterations; and d) lactate: pyruvate (L:P) ratios. Cell cultures showed concentration-and time-dependent responses. For example, a decrease in cell viability and an increase in enzyme leakage were observed after cell treatment with 1×10⁻⁴ and 1×10⁻³ M papaver for 8 h; 1×10⁻³ M 6-OH papaverine for 8 h and 1×10⁻⁴ M for 24 h; and 1×10⁻³ M 4′-OH papaverine for 24 h (P<0.05). Furthermore, changes in morphology correlated to cell viability and enzyme release in those cultures treated with papaver, 4′-OH and 6-OH papaverine. Some of these changes included size deformation, cell detachment from the dishes, and cell necrosis. On the other hand, an increase in L:P ratios (P<0.05) was detected with papaver as early as 8 h with 1×10⁻⁴ and 1×10⁻³ M and 12 h with 1×10⁻⁵ M; 6-OH showed an increase, in L:P ratios at 8 h with 1×10⁻³ M and 12 h with 1×10⁻⁴ M; these changes were evident with 4′-OH at 12 h with 1×10⁻³ M. In contrast, cells treated with 3′-OH papaverine did not show significant damage with any time period and concentration used in this study. The results of this study indicate that papaverine-derived metabolites are less cytotoxic than its parent compound, papaver. The toxicity was ranked as follows: papaver>6-OH>4′-OH>−3′-OH. This work was supported in part by grant ES04200-02 from the National Institute of Environmental Health Sciences, Bethesda, MD. Presented in part at the fall ASPET meeting in Salt Lake City, August, 1989. Daniel Acosta is a Burroughs Wellcome Scholar in Toxicology.     

Disruption of glpF gene encoding the glycerol facilitator improves 1,3-propanediol production from glucose via glycerol in Escherichia coli

Engineered Escherichia coli has recently been applied to produce 1,3-propanediol (1,3-PDO) from glucose. A metabolic intermediate in the production pathway, glycerol, is partially secreted into the extracellular of E. coli through a glycerol facilitator encoded by glpF, and this secretion consequently decreases 1,3-PDO production. Therefore, we aimed to determine whether disrupting the glpF gene would improve 1,3-PDO production in E. coli. The intracellular glycerol concentration in a glpF-disruptant was 7·5 times higher than in a non-disruptant. The glpF-disrupted and non-disrupted E. coli strains produced 0·26 and 0·09 g l⁻¹ of 1,3-PDO, respectively, from 1% glucose after 72 h of cultivation. The specific growth rate (μ) and the 1,3-PDO yield from glucose (Y_P/S) in the disruptant were higher than those in the non-disruptant (ΔglpF, μ = 0·08 ± 0·00 h⁻¹, Y_P/S = 0·06 mol mol-glucose⁻¹; BW25113, μ = 0·06 ± 0·00 h⁻¹, Y_P/S = 0·02 mol mol-glucose⁻¹). Disruption of the glpF gene decreased the production of the by-product, acetic acid. These results indicated that disruption of glpF increased the intracellular concentration of glycerol and consequently increased 1,3-PDO production in E. coli.     

Cross‐effects of dietary probiotic supplementation and rearing temperature on antioxidant responses in European seabass [Dicentrarchus labrax (Linnaeus, 1758)] juveniles

Temperature modulates the metabolism in both fish and bacteria and therefore the effect of probiotic bacteria on its host may vary accordingly. The current study aim was to evaluate the effect of probiotic supplementation (Bacillus sp., Lactobacillus sp., Enterococcus sp., Pediococcus sp.) in juvenile seabass, Dicentrarchus labrax, when reared under different temperatures (17, 20 and 23°C). A control diet was tested against a probiotic‐supplemented diet, with a concentration of 3 × 10⁹ CFU probiotic/kg diet. Antioxidant responses (TG, GSH, GSSG, GR, CAT and GSTs) and lipid peroxidation (LPO) were evaluated after 70 days of dietary probiotic supplementation. An effect of temperature was observed on LPO, which increased significantly in fish reared at 17°C (p < .05) compared to the 20 and 23°C groups. Total glutathione (TG) was significantly higher in the probiotic treatments in fish reared at 17 and 20°C (p < .05). In addition, a probiotic temperature interaction was observed for TG, reduced glutathione (GSH) levels, and for reduction of the oxidized glutathione ratio (GSH/GSSG; p < .05). In conclusion, the current study showed a strong temperature effect on oxidative stress responses, with an anti‐oxidant role of dietary probiotic supplementation at different rearing temperatures.     

Kocuria SM1 controls vibriosis in rainbow trout (Oncorhynchus mykiss,Walbaum)

Aims: To develop probiotics for the control of vibriosis caused by Vibrio anguillarum and Vibrio ordalii in finfish. Methods and Results: Kocuria SM1, isolated from the digestive tract of rainbow trout, was administered orally to rainbow trout (Oncorhynchus mykiss) for 2 weeks at a dose equivalent to c. 10⁸ cells per g of feed and then challenged intraperitoneally with V. anguillarum and V. ordalii. Use of SM1 led to a reduction in mortalities to 15–20% compared to 74–80% mortalities in the controls. SM1 stimulated both cellular and humoral immune responses in rainbow trout, by elevation of leucocytes (5·5 ± 0·8 × 10⁶ ml^?1 from 3·7 ± 0·8 × 10⁶ ml^?1), erythrocytes (1·2 ± 0·1 × 10⁸ ml^?1 from 0·8 ± 0·1 × 10⁸ ml^?1), protein (23 ± 4·4 mg ml^?1 from 16 ± 1·3 mg ml^?1), globulin (15·7 ± 0·2 mg ml^?1 from 9·9 ± 0·1 mg ml^?1) and albumin (7·3 ± 0·2 mg ml^?1 from 6·1 ± 0·1 mg ml^?1) levels, upregulation of respiratory burst (0·05 ± 0·01 from 0·02 ± 0·01), complement (56 ± 7·2 units ml^?1 from 40 ± 8·0 units ml^?1), lysozyme (920 ± 128·8 units ml^?1 from 760 ± 115·3 units ml^?1) and bacterial killing activities. Conclusions: Kocuria SM1 successfully controlled vibriosis in rainbow trout, and the mode of action reflected stimulation of the host innate immune system. Significance and Impact of the Study: Probiotics can contribute a significant role in fish disease control strategies, and their use may replace some of the inhibitory chemicals currently used in fish farms.     

Use of Real-Time PCR Technique in Studying Rumen Cellulolytic Bacteria Population as Affected by Level of Roughage in Swamp Buffalo

A real-time polymerase chain reaction approach was used in this study to determine the population of major ruminal bacterial species (Fibrobacter succinogenes, Ruminococcus albus, and Ruminococcus flavefaciens) in digesta and rumen fluid of swamp buffalo (Bubalus bubalis). Four rumen-fistulated, male swamp buffalo were randomly assigned according to a 4 × 4 Latin square design to evaluate the effect of the urea-treated rice straw (roughage source)-to-concentrate ratio on cellulolytic bacterial distribution. Animals were fed roughage-to-concentrate (R:C) ratios of 100:0, 75:25, 50:50, and 25:75, respectively. At the end of each period, rumen fluid and digesta were collected at 0 h and 4 h post-morning-feeding. It was found that feeding urea-treated rice straw solely increased these three cellulolytic bacteria numbers up to 2.65 × 10⁹ and 3.54 × 10⁹ copies per milliliter for F. succinogenes, 5.10 × 10⁷ and 7.40 × 10⁷ copies per millilter for R. Flavefaciens, and 4.00 × 10⁶ and 6.00 × 10⁶ copies per milliliter for R. albus in rumen fluid and digesta, respectively. The distribution of the three cellulolytic bacteria species in digesta were highest at 3.21 × 10⁹, 4.55 × 10⁷, and 4.56 × 10⁶ copies per milliliter for F. succinogenes, R. flavefaciens, and R. albus, respectively. Moreover, at 4 h post-morning-feeding, the populations of the three cellulolytic bacteria were higher than found at 0 h post-morning-feeding. It is most notable that F. succinogenes were the highest in population in the rumen of swamp buffalo and cellulolytic bacteria mostly adhered to feed digesta in the rumen.