The light microclimate of phototrophic endoliths growing within the scleractinian corals Porites cylindrica and Montipora monasteriata was described by scalar irradiance microprobe measurements within different layers of the coral skeleton. Characterisation of the pigments in individual layers was done by reflectance spectroscopy with fibre-optic radiance microprobes. The spectral measurements showed the presence of an endolithic community largely comprised of the green alga Ostreobium sp. within a 1 to 2 mm thick green band 2 to 6 mm below the coral surface. Additionally, spectral signatures of cyanobacteria and anoxygenic phototrophic bacteria were detected both in the coral tissue-containing top layer and within the skeleton matrix. The light microclimate within the coral skeleton was extremely poor in visible light but enriched in far-red wavelengths. Only a fraction of the incident photosynthetically available radiation (PAR, 400 to 700 nm) penetrated the coral tissue-containing layer, wherein 90 to 99% of the incident irradiance was attenuated due to intense scattering and absorption. Near-infrared radiation (NIR, >700 to 1000 nm) was mainly scattered in the tissue-skeleton matrix and penetrated much deeper into the skeleton. Multiple scattering and light-trapping effects caused high NIR scalar irradiance levels in the topmost layers of the coral. Our data show that the endolithic community in healthy corals is strongly light-limited with respect to PAR, but not with respect to NIR in shallow waters where water absorption of NIR is not limiting. Light limitation of PAR is mainly imposed by the tissue-containing part of the coral, and could thus be alleviated during coral bleaching, resulting in blooming of the phototrophic endoliths.
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To examine whether there is a relationship between the degree of Campylobacter contamination observed in product lots of retail Icelandic broiler chicken carcasses and the incidence of human disease, 1,617 isolates from 327 individual product lots were genetically matched (using the flaA short variable region [SVR[) to 289 isolates from cases of human campylobacteriosis whose onset was within approximately 2 weeks from the date of processing. When there was genetic identity between broiler isolates and human isolates within the appropriate time frame, a retail product lot was classified as implicated in human disease. According to the results of this analysis, there were multiple clusters of human disease linked to the same process lot or lots. Implicated and nonimplicated retail product lots were compared for four lot descriptors: lot size, prevalence, mean contamination, and maximum contamination (as characterized by direct rinse plating). For retail product distributed fresh, Mann-Whitney U tests showed that implicated product lots had significantly (P = 0.0055) higher mean contamination than nonimplicated lots. The corresponding median values were 3.56 log CFU/carcass for implicated lots and 2.72 log CFU/carcass for nonimplicated lots. For frozen retail product, implicated lots were significantly (P = 0.0281) larger than nonimplicated lots. When the time frame was removed, retail product lots containing Campylobacter flaA SVR genotypes also seen in human disease had significantly higher mean and maximum contamination numbers than lots containing no genotypes seen in human disease for both fresh and frozen product. Our results suggest that cases of broiler-borne campylobacteriosis may occur in clusters and that the differences in mean contamination levels may provide a basis for regulatory action that is more specific than a presence-absence standard.
Frequency and numbers of Campylobacter spp. were assessed per freshly processed, contaminated broiler carcass. Campylobacter-positive flocks were identified by cecal sample analysis at slaughter. These flocks had been tested as Campylobacter negative at 4.1 ± 0.9 d prior to slaughter. Levels of contamination were estimated using 2 sampling approaches per carcass: (1) free weep fluids and (2) whole-carcass, 100 mL of distilled water rinses. Estimations of counts were determined by directly plating dilutions of weeps and rinses onto Campy-Cefex agar and incubating the plates at 41.5°C under microaerobic atmosphere. Confirmation was provided by latex agglutination to quantify levels per milliliter of weep and per 100 mL of rinse. Thirty-two slaughter groups (∼20 carcasses per group) were compared from 2003 to 2004. The Campylobacter-positive weep frequency was 84.8%, whereas the frequency for rinse samples was 74.4% (P < 0.001). Enumeration of Campylobacter spp. on positive samples ranged from 0.70 to 6.13 log10 cfu/mL of weep (geometric mean of 2.84) and from 2.30 to 7.72 log10 cfu/100 mL of rinse (geometric mean of 4.38). The correlations between weep and rinse were 0.814 with 0.5 mL of rinse and 0.6294 when applying 0.1 mL of rinse The quantitative regression analyses for these 2 corresponding tests were log10 rinse (for 0.5 mL of inoculum) = 1.1965 log10 weep + 0.4979, and log10 rinse (for 0.1 mL of inoculum) = 1.322 log10 weep − 0.1521. FlaA SVR sequencing of isolates indicated that the same genotypes were found in weep and rinse samples. Weep and rinse sampling led to different proportions of Campylobacter-positive carcasses detection, but we demonstrated that this difference was reduced by increasing the amount of rinse fluid used for plating.
Whole meat of Blue marlin (Makaira mazara) was used to prepare edible films. Protein solubility in film-forming solutions was high at acidic and alkaline pHs, while that at neutral pH was close to zero. Acidic and alkaline pHs improved the tensile strength while the effects of pH, on elongation at break, water vapour permeability and light transmission of the films, were not significant. From the film solubility in various protein denaturants it was revealed that the main interaction responsible for the formation of acidic and alkaline pH films was hydrophobic interaction, while that for neutral pH films was ionic bonding.
Fish muscle protein films were prepared from blue marlin (Makaira mazara Jordan & Evermann) meat which had been stored at 30°C to intentionally lower the meat quality. In this study, the effects of meat quality and pH on the formation of these protein films were investigated. Moreover, ε-polylysine was added to the film-forming solutions to reduce the microbial population of films. The mechanical properties of the films were slightly affected by acidic and alkaline pHs. However, the water vapor permeability of muscle protein films was not affected by either the quality of the fish meat or the pH of the film-forming solutions. SDS-PAGE (sodium dodecyl sulfate- polyacrylamide gel electrophoresis) showed the degradation of myosin heavy chain in acidic films and polymerization in alkaline films. It was revealed that biodegradable films can be produced even from very low quality fish meat, and that the bacterial population of films could be drastically reduced by the addition of ε-polylysine.
Atlantic cod of initial mean weight ∼220 g were fed a control diet and three diets in which fish meal (FM) was replaced with increasing levels of full-fat soybean meal (FFS) supplied at 12, 24, and 36% of dry diet, for 12 weeks. There were no significant differences in final weights, but the specific growth rate (SGR) was significantly higher in fish fed the control (FFS0) diet compared to fish fed the FFS12 and FFS36 diets, and the feed conversion ratio (FCR) was significantly lower in fish fed the FFS0 diet compared to the other three treatments. The fatty acid (FA) compositions of the cod muscle and liver were highly affected by dietary treatment, and linear relationships between dietary and tissue FA concentrations were shown for some of these. Moreover, selective utilization or accumulation in the tissues of specific FA was suggested by the results.
The biochemical and genetic search for a bc 1 complex in Rhodothermus marinus was always fruitless; however, a functional equivalent, i.e. having quinol:cytochrome c oxidoreductase activity was characterized. Now, with the sequencing of R. marinus genome, it was possible to assign the N-terminal sequences of several proteins of this complex to its coding genes. The alternative complex III from R. marinus has the same genomic organization of the so-called MFIcc complexes, proposed to be oxidoreductases of the respiratory and photosynthetic electron transfer chains. In this report, we establish undoubtedly the existence of an alternative complex III, a functional substitute of the bc 1 complex, by its identification at both the biochemical and genomic level.
Trehalases play a central role in the metabolism of trehalose and can be found in a wide variety of organisms. A periplasmic trehalase (a,a-trehalose glucohydrolase, EC 3.2.1.28) from the thermophilic bacterium Rhodothermus marinus was purified and the respective encoding gene was identified, cloned and overexpressed in Escherichia coli. The recombinant trehalase is a monomeric protein with a molecular mass of 59 kDa.
Maximum activity was observed at 88C and pH 6.5. The recombinant trehalase exhibited a Km of 0.16 mM and a Vmax of 81 lmol of trehalose (min)1 (mg of protein)1 at the optimal temperature for growth of R. marinus (65C) and pH 6.5. The enzyme was highly specific for
trehalose and was inhibited by glucose with a Ki of 7 mM. This is the most thermostable trehalase ever characterized. Moreover, this is the first report on the identification and characterization of a trehalase from a
thermophilic bacterium.
The aerobic respiratory chain of the thermohalophilic bacterium Rhodothermus marinus has been extensively studied. In this study the isolation and characterization of a third oxygen reductase expressed in this organism are described. This newly isolated enzyme is a typical member of the type B family of haem-copper oxygen reductases, showing 43% amino acid sequence identity and 63% similarity with the ba3 oxygen reductase from Thermus thermophilus. It constitutes two subunits with apparent molecular masses of 42 and 38 kDa. It contains a low-spin B-type haem and a high-spin A-type haem. A stoichiometry of 1B: 1A haem per protein was obtained by spectral integration of UV-visible spectra. Metal analysis showed the presence of two iron and three copper ions, which is in agreement with the existence of a CuA centre.
Taking advantage of having two spectroscopically distinct haems, the redox behaviour of the ba3 oxygen reductase was analysed and discussed in the framework of a model with interacting centres. Both haems, B and A, present two transitions, have unusually low reduction potentials of −65 mV and an interaction potential of −52.5 mV.
An α-l-rhamnosidase (E.C. 3.2.1.40) from a newly discovered thermophilic bacterium was expressed in Escherichia coli BL21 DE3 pRIL cells. The cells were immobilized in Ca2+-alginate beads. The temperature of 50 °C used in reactions, appeared to be sufficient for making the mesophilic strain porous enough for the substrate to access the cloned thermostable enzyme. Pretreatment of cells with heat or lysozyme prior to bead formation did not improve the results. The best cell concentration (w/w) for bead preparation was found to be 0.0192 g ml−1 and stability of beads increased if CaCl2 concentration in buffers and substrate was kept at 50 mM. In a 60 min assay, the optimal pH of the entrapped cells was found to be 7.8 and the optimal temperature 60 °C. By packing the beads in a column, a bioreactor for production of l-rhamnose from naringin was created. Full degradation of 7.9 mM naringin could be reached by running the reactor at 1 ml min−1 at 50 °C. The optimal running temperature of the reactor was found to be 50 °C and the reactor was fully stable over 3 days at that temperature. On the fourth day, substrate degradation capacity had decreased by 10–15%.