Bacterial Diversity and Bioprospecting for Cold-Active Lipases,Amylases and Proteases,from Culturable Bacteria of Kongsfjorden and Ny-Ålesund,Svalbard, Arctic

Culturable bacterial diversity of seven marine sediment samples of Kongsfjorden and a sediment and a soil sample from Ny-Ålesund, Svalbard, Arctic was studied. The bacterial abundance in the marine sediments of Kongsfjorden varied marginally (0.5 × 10³–1.3 × 10⁴ cfu/g sediment) and the bacterial number in the two samples collected from the shore of Ny-Ålesund also was very similar (0.6 × 10⁴ and 3.4 × 10⁴, respectively). From the nine samples a total of 103 bacterial isolates were obtained and these isolates could be grouped in to 47 phylotypes based on the 16S rRNA gene sequence belonging to 4 phyla namely Actinobacteria, Bacilli, Bacteroidetes and Proteobacteria. Representatives of the 47 phylotypes varied in their growth temperature range (4–37°C), in their tolerance to NaCl (0.3–2 M NaCl) and growth pH range (2–11). Representatives of 26 phylotypes exhibited amylase and lipase activity either at 5 or 20°C or at both the temperatures. A few of the representatives exhibited amylase and/or lipase activity only at 5°C. None of the phylotypes exhibited protease activity. Most of the phylotypes (38) were pigmented. Fatty acid profile studies indicated that short chain fatty acids, unsaturated fatty acids, branched fatty acids, the cyclic and the cis fatty acids are predominant in the psychrophilic bacteria.     

Diversity of Chemotactic Heterotrophic Bacteria Associated with Arctic Cyanobacteria

The abundance and diversity of chemotactic heterotrophic bacteria associated with Arctic cyanobacteria was determined. The viable numbers ranged between 10⁴ and 10⁶ cell g^?1 cyanobacterial biomass. A total of 112 morphotypes, representing 22 phylotypes based on their 16S rRNA sequence similarity were isolated from the samples. All the phylotypes were Gram-negative with affiliation to the proteobacterial and bacteroidetes divisions. Among the 22 phylotypes, 14 were chemotactic to glucose. Majority of the phylotypes were psychrotolerant showing growth up to 30 °C. Representatives of Alphaproteobacteria, the genus Flavobacterium and the gammaproteobacterial Alcanivorax sp, were psychrophilic with growth at or below 18 °C. A significant percentage of phylotypes were pigmented (~68 %), rich in unsaturated membrane fatty acids and tolerated pH values and NaCl concentrations between 5.0–8.0 and 0.15–1.0 M, respectively. The percentages of phylotypes producing extracellular cold-active enzymes at 4 °C were amylase (18.18 %), lipase and urease (45.45 %), caseinase (59.09 %) and gelatinase (31.8 %).     

Formulation of the biocontrol agent <Emphasis Type="Italic">Bacillus amyloliquefaciens</Emphasis> CPA-8 using different approaches: liquid,freeze-drying and fluid-bed spray-drying

The present work focuses on the assessment and comparison of three different formulation technologies and the effect of protectants on cell viability, storage stability and antagonistic activity of the biocontrol agent Bacillus amyloliquefaciens CPA-8. Cultures were concentrated with different protective substances such as MgSO₄, sucrose and skimmed milk (SM) and subjected to liquid formulation, freeze-drying and fluid-bed spray-drying. Results showed that CPA-8 freeze-dried cells without protectants or amended with SM suffered the highest losses in cell viability (0.41?0.48 log). Moreover, the cell viability of the tested freeze-dried products decreased after four months of storage at both tested temperatures (4 and 22 °C). Otherwise, liquid and fluid-bed spray-dried products were stable for four months at 4 °C and for 12 months at 22, 4 and ?20 °C, respectively, and no effect of the protectants was observed. The most suitable CPA-8 products were then tested against Monilinia laxa and M. fructicola in artificially wounded nectarines and in all cases the antagonistic activity was maintained similar to fresh cells. The efficacy results revealed that the formulation process did not affect the biocontrol potential of CPA-8. This work led us to conclude that effective formulations with final concentrations ranging from 1.93 × 10⁹–2.98 × 10⁹ CFU ml^?1 and from 4.76 × 10⁹–1.03 × 10¹⁰ CFU g^?1 were obtained for liquid and dried products, respectively. Additionally, the suitability of the fluid-bed spray drying technology should be taken into account to develop a stable and effective CPA-8 product for practical applications to control brown rot in stone fruit.     

Cellulose Derivatives Enhanced Stability of Alginate-Based Beads Loaded with <Emphasis Type="Italic">Lactobacillus plantarum</Emphasis> LAB12 against Low pH,High Temperature and Prolonged Storage

The susceptibility of probiotics to low pH and high temperature has limited their use as nutraceuticals. In this study, enhanced protection of probiotics via microencapsulation was achieved. Lactobacillus plantarum LAB12 were immobilised within polymeric matrix comprised of alginate (Alg) with supplementation of cellulose derivatives (methylcellulose (MC), sodium carboxymethyl cellulose (NaCMC) or hydroxypropyl methylcellulose (HPMC)). L. plantarum LAB12 encapsulated in Alg-HPMC(1.0) and Alg-MC(1.0) elicited improved survivability (91%) in simulated gastric conditions and facilitated maximal release (～100%) in simulated intestinal condition. Alg-HPMC(1.0) and Alg-MC(1.0) significantly reduced (P < 0.05) the viability loss of LAB12 (viability loss <7%) when compared to Alg alone (viability loss <13%) under extreme temperatures (75 and 90 °C). Four-week storage of encapsulated LAB12 at 4 °C yielded viable counts >7 log CFU g^?1. Alg-MC and Alg-HPMC improved the survival of LAB12 against simulated gastric condition (9.24 and 9.55 log CFU g^?1, respectively), temperature up to 90 °C (9.54 and 9.86 log CFU g^?1, respectively) and 4-week of storage at 4 °C (8.61 and 9.23 log CFU g^?1, respectively) with sustained release of probiotic in intestinal condition (>9 log CFU g^?1). These findings strongly suggest the potential of cellulose derivatives supplemented Alg bead as protective micro-transport for probiotic strains. They can be safely incorporated into new functional food or nutraceutical products.     

Listeria monocytogenes Can Form Biofilms in Tap Water and Enter Into the Viable but Non-Cultivable State

Listeria monocytogenes is a foodborne pathogen that can be transmitted through contaminated raw food or by ready-to-eat products that have been in contact with contaminated surfaces. Tap water (TW) is used to wash produce, as a processed food constituent and to wash processing surfaces and floors. The main aim of this work was to investigate the formation and survival of L. monocytogenes biofilms on stainless steel (SS) coupons in TW at 4, 22, 30 and 37 °C. For that, coupons with biofilm were visualised in situ while other coupons were scraped to quantify total cells by SYTO 9, cultivable numbers by plating onto brain heart infusion agar and viable numbers by the direct viable count method. Results showed that L. monocytogenes can form biofilms on SS surfaces in TW at any temperature, including at 4 °C. The number of total cells was similar for all the conditions tested while cultivable numbers varied between the level of detection (<8.3 CFU cm^?2) and 3.5?×?10⁵ CFU cm^?2, meaning between 7.0?×?10⁴ and 1.1?×?10⁷ cells cm^?2 have entered the viable but non-cultivable (VBNC) state. This work clearly demonstrates that L. monocytogenes can form biofilms in TW and that sessile cells can remain viable and cultivable in some conditions for at least the 48 h investigated. On the other hand, VBNC adaptation suggests that the pathogen can remain undetectable using traditional culture recovery techniques, which may give a false indication of processing surface hygiene status, leading to potential cross-contamination of food products.     

Aerobiological study in the Mexico City subway system

Monitoring campaigns in two different seasons were carried out at two underground stations (Tacubaya and Azcapotzalco) of the subway of Mexico City, in order to assess airborne bacterial and fungi concentrations, as well as their relationship with several factors, such as depth of the station, sampling site, temperature, and relative humidity. Sixteen sampling sites were selected according to the depth of the lines and the transit of passengers in the corridors, concourses, and platforms. In addition, samples were also collected in the carriages when they were in movement. Outdoor samples were taken at the two stations for comparison. Two-stage multi-orifice cascade impactors were used to collect aerobiological particles, and with the aid of macroscopic and microscopic characterization were found 57 fungi and 61 bacteria colonies. Outdoor bacteria concentrations ranged from 1 to 68 CFU m^?3, while fungi concentrations varied from 6 to 80 CFU m^?3. The indoor concentration levels of bacteria and fungi ranged from 1 to 484 CFU m^?3 and from 51 to 715 CFU m^?3, respectively. Fungi and bacteria indoor concentrations in the subway were higher than outdoor, up to 8 times. Most of bacteria were identified as Gram-positive nonsporulating short bacillus, while the most abundant fungi genera identified were Aspergillus, Penicillium, and Alternaria in that order of frequency. Statistical analysis showed significant differences between the stations and the lines of different depths, showing a greater microorganisms’ concentration with a greater depth. Although bacteria and fungi concentrations were higher in the spring than in winter, this difference was not significant. Even if indoor microbiological pollution in underground stations was higher than outdoor, the concentrations found in this study were lower than indoor air international standards.     

Probiotic Effect of <Emphasis Type="Italic">Bacillus licheniformis fb11</Emphasis> on the Digestive Efficiency and Growth Performance in Juvenile <Emphasis Type="Italic">Chitala chitala</Emphasis> (Hamilton, 1822)

Five isocaloric (430 kcal 100 g^?1), isonitrogenous (40% CP) experimental diets were formulated with different concentrations of Bacillus licheniformis fb11 probionts (isolated from the gut of Chitala chitala) viz. Control (without probionts), 5 × 10⁴ CFU g^?1 (D₁), 5 × 10⁵ CFU g^?1 (D₂), 5 × 10⁶ CFU g^?1 (D₃), 5 × 10⁷ CFU g^?1 (D₄), 5 × 10⁸ CFU g^?1 (D₅) to evaluate its efficiency in C. chitala juvenile. The best growth performance, feed utilisation, specific α-amylase, total protease and lipase activity were observed with the diet D₃ (P < 0.05). The lowest Presumptive Pseudomonas Count, Motile Aeromonad Count, Total Coliform Count was observed for D₃ (P < 0.05) on 90th day of trial. Two uppermost values were achieved in case of crude protein for D₃ and D₂ (P > 0.05). The highest lipid content (12.12 ± 0.4 g 100 g^?1) was found for D₅ (P < 0.05). The highest gross energy (18.75 ± 0.21 MJ 100 g^?1) of carcass was recorded for D₃. Thus B. licheniformis fb11 at the concentration 5 × 10⁶ CFU g^?1 as probiotic supplement promoted growth, digestion in C. chitala juvenile significantly by modulating intestinal microflora.     

Biochemical composition, biological activities and toxicological effects of two non-nodularin producing strains of Nodularia spumigena Mertens in Jürgens

We tested two non-nodularin-producing strains of the cyanobacterium Nodularia spumigena, isolated from a marine (Kachelotplate) and a brackish water (Banter Sea, Wilhelmshaven) habitat in Lower Saxony, Germany, for allelochemical production (e.g. alkaloids, flavonoids) and allelopathic activities (e.g. algicidal, anti-microbial). The growth experiments showed for the marine strain the highest cell density at 10 and 20 °C for the brackish water isolate (80 μmol ?photons m^?2?s^?1). Phytochemical screening of the biomass extracts gave positive results for alkaloids, flavonoids, sterols and terpenoids in some of the tested assays. Most of these compounds were not present in supernatant extracts. Besides proalgal and anti-cyanobacterial properties of the high temperature treated marine strain, the supernatant extracts showed profungal and antibacterial activities in the 20 °C treated assays. In both, supernatant and biomass extracts, significant anti-oxidative activities were observed in the high-irradiance-treated marine and brackish water isolates. The highest toxicity was observed at the 5 and 20 °C brackish water isolates as well as 5 °C treated marine strain. With regard to fatty acid composition, both strains showed high levels of polyunsaturated fatty acids (PUFAs) and saturated fatty acids, with values of 36–54 % and 11–29 % of total fatty acids, respectively, whereas the levels of monounsaturated fatty acids were in general lower (8–16 %). Among PUFAs, linoleic (C18:2), α-linoleic (C18:3), γ-linoleic (C18:3) and arachidonic acid (C20:4) accounted 36.2 % of the total polyunsaturated fatty acids in the brackish water strain, while in the marine isolate, it was only 10.6 %.     

Association of Bacteria with Marine Invertebrates: Implications for Ballast Water Management

Bacteria associated with plankton are of importance in marine bioinvasions and the implementation of ship’s ballast water treatment technologies. In this study, epibiotic and endobiotic bacteria associated with zooplankton, including barnacle nauplii, veliger larvae, and adults of the copepod Oithona sp., were characterized and quantified. Barnacle nauplius and veliger larva harbored ~4.4 × 10⁵ cells ind^?1 whereas Oithona sp. had 8.8 × 10⁵ cells ind^?1. Computation of bacterial contribution based on biovolume indicated that despite being the smallest zooplankton tested, veliger larvae harbored the highest number of bacteria, while barnacle nauplii, the largest of the zooplankton, tested in terms of volume contributed the least. Pulverization of zooplankton led to an increase in bacterial numbers; for example, Vibrio cholerae, which was initially 3.5 × 10³, increased to 5.4 × 10⁵ CFU g^?1; Escherichia coli increased from 5.0 × 10² to 1.3 × 10⁴ CFU g^?1; and Streptococcus faecalis increased from 2.1 × 10² to 2.5 × 10⁵ CFU g^?1, respectively. Pulverized zooplankton was aged in the dark to assess the contribution of bacteria from decaying debris. Aging of pulverized zooplankton led to emergence of Chromobacterium violaceum, which is an opportunistic pathogen in animals and humans.     

Selection of indigenous isolates of entomopathogenic soil fungus Metarhizium anisopliae under laboratory conditions

Eight native isolates of the entomopathogenic fungus Metarhizium anisopliae (Metschnikoff) Sorokin were obtained by monitoring soils cultivated in a conventional manner. These isolates were compared in three areas: (a) conidial germination, (b) radial growth and sporulation and (c) ability of conidia to infect Tenebrio molitor larvae. All bioassays were carried out at constant temperatures of 10, 15, and 20 °C. Conidia of individual isolates demonstrated differences in germination after a 24-h long incubation at all evaluated temperatures. At 20 °C, the germination ranged from 67 to 100 % and at 15 °C from 5.33 to 46.67 %. At 10 °C, no germination was observed after 24 h; nevertheless, it was 8.67–44.67 % after 48 h. In terms of radial growth, the culture diameters and the associated production of spores of all isolates increased with increasing temperature. At 10 °C, sporulation was observed in three isolates while all remaining cultures appeared sterile. Three weeks post-inoculation, conidia of all assessed isolates caused 100 % cumulative mortality of treated larvae of T. molitor at 15 and 20 °C with the exception of isolate 110108 that induced 81.33 % mortality at 15 °C. At 10 °C, larval cumulative mortality ranged from 6.67 to 85.33 % depending on the isolate. Isolates 110108 and 110111 showed significantly slower outset and a much lower rate of infection at all temperatures compared to other tested isolates of M. anisopliae. The bioassays were carried out with the purpose to sort and select indigenous isolates of M. anisopliae useful as biocontrol agents in their original habitat.     

Production of microsclerotia by Brazilian strains of Metarhizium spp. using submerged liquid culture fermentation

We investigated the potential production and desiccation tolerance of microsclerotia (MS) by Brazilian strains of Metarhizium anisopliae (Ma), M. acridum (Mc) and M. robertsii (Mr). These fungi were grown in a liquid medium containing 16 g carbon l^?1 with a carbon:nitrogen ratio of 50:1. One hundred milliliters cultures were grown in 250 ml Erlenmeyer flasks in a rotary incubator shaker at 28 °C and 200 rpm for 5 days. Five-day-old MS were harvested, mixed with diatomaceous earth (DE) and air-dried for 2 days at 30 °C. The air-dried MS–DE granular preparations were milled by mortar + pestle and stored in centrifuged tubes at either 26 or ?20 °C. Desiccation tolerance and conidia production were assessed for dried MS granules by measuring hyphal germination after incubation for 2 days on water agar plates at 26 °C and for conidia production following 7 days incubation. Yields of MS by all strains of Metarhizium were 6.1–7.3 × 10⁶ l^?1 after 3 days growth with maximum MS yields (0.7–1.1 × 10⁷ l^?1) after 5 days growth. No differences in biomass accumulation were observed after 3 days growth, whereas Ma-CG168 showed the highest biomass accumulation after 5 days growth. Dried MS–DE preparations of all fungal strains were equally tolerant to desiccation (≥93 % germination) and the highest conidia production was obtained by MS granules of Mc-CG423 (4 × 10⁹ conidia g^?1). All MS granules showed similar stability after storage at either 26 or ?20 °C for 3.5 months.     

Thermal dependence of clearance and metabolic rates in slow- and fast-growing spats of manila clam Ruditapes philippinarum

Thermal dependence of clearance rate (CR: l h^?1), standard (SMR: J h^?1) and routine metabolic rates (RMR: J h^?1), were analyzed in fast (F)- and slow (S)-growing juveniles of the clam Ruditapes philippinarum. Physiological rates were measured at the maintenance temperature (17 °C), and compared with measurements performed at 10 and 24 °C after 16 h and 14 days to analyze acute and acclimated responses, respectively. Metabolic rates (both RMR and SMR) differed significantly between F and S seeds, irrespective of temperature. Mass-specific CRs were not different for F and S seeds but were significantly higher in F clams for rates standardized according to allometric size-scaling rules. Acute thermal dependency of CR was equal for F and S clams: mean Q ₁₀ were ≈3 and 2 in temperature ranges of 10–17 and 17–24 °C, respectively. CR did not change after 2 weeks of acclimation to temperatures. Acute thermal effects on SMR were similar in both groups (Q ₁₀ ≈ 1 and 1.6 in temperature ranges of 10–17 and 17–24 °C, respectively). Large differences between groups were found in the acute thermal dependence of RMR: Q ₁₀ in F clams (≈1.2 and 1.9 at temperature ranges of 10–17 and 17–24 °C, respectively) were similar to those found for SMR (Q ₁₀ = 1.0 and 1.7). In contrast, RMR of S clams exhibited maximum thermal dependence (Q ₁₀ = 3.1) at 10–17 °C and become depressed at higher temperatures (Q ₁₀ = 0.9 at 17–24 °C). A recovery of RMR in S clams was recorded upon acclimation to 24 °C. Contrasting metabolic patterns between fast and slow growers are interpreted as a consequence of differential thermal sensitivity of the fraction of metabolism associated to food processing and assimilation.     

Thermal mitigation of Pseudomonas aeruginosa biofilms

Bacterial biofilms infect 2–4% of medical devices upon implantation, resulting in multiple surgeries and increased recovery time due to the very great increase in antibiotic resistance in the biofilm phenotype. This work investigates the feasibility of thermal mitigation of biofilms at physiologically accessible temperatures. Pseudomonas aeruginosa biofilms were cultured to high bacterial density (1.7?×?10⁹ CFU cm^?2) and subjected to thermal shocks ranging from 50°C to 80°C for durations of 1–30 min. The decrease in viable bacteria was closely correlated with an Arrhenius temperature dependence and Weibull-style time dependence, demonstrating up to six orders of magnitude reduction in bacterial load. The bacterial load for films with more conventional initial bacterial densities dropped below quantifiable levels, indicating thermal mitigation as a viable approach to biofilm control.     

Phosphate solubilization potential and modeling of stress tolerance of rhizobacteria from rice paddy soil in northern Iran

The purposes of this study were to evaluate the phosphate solubilization activity of bacteria isolated from the rhizosphere of rice paddy soil in northern Iran, and to study the effect of temperature, NaCl and pH on the growth of these isolates by modeling. Three of the most effective strains from a total of 300 isolates were identified and a phylogenetic analysis was carried out by 16S rDNA sequencing. The isolates were identified as Pantoea ananatis (M36), Rahnella aquatilis (M100) and Enterobacter sp. (M183). These isolates showed multiple plant growth-promoting attributes such as phosphate solubilization activity and indole-3-acetic acid (IAA) production. The M36, M100 and M183 isolates were able to solubilize 172, 263 and 254 µg ml^?1 of Ca₃(PO₄)₂ after 5 days of growth at 28 °C and pH 7.5, and to produce 8.0, 2.0 and 3.0 μg ml^?1 of IAA when supplemented with l-tryptophan (1 mg ml^?1) for 72 h, at 28 °C and pH 7.0, respectively. The solubilization of insoluble phosphate was associated with a drop in the pH of the culture medium and there was an inverse relationship between pH and solubilized P (r = ?0.98, P < 0.0952). There were no significant differences among isolates in terms of acidity tolerance based on their confidence limits as assessed by segmented model analysis and all isolates were able to grow at pH 4.3–11 (with optimum at 7.0–7.5). Based on a sigmoidal trend of a three-parameter logistic model, the salt concentration required for 50 % inhibition was 8.15, 6.30 and 8.23 % NaCl for M36, M100 and M183 isolates, respectively. Moreover, the minimum and maximum growth temperatures estimated by the segmented model were 5.0 and 42.75 °C for M36, 12.76 and 40.32 °C for M100, and 10.63 and 43.66 °C for M183. The three selected isolates could be deployed as inoculants to promote plant growth in an agricultural environment.     

Degradation properties of various macromolecules of cultivable psychrophilic bacteria from the deep-sea water of the South Pacific Gyre

The deep-sea water of the South Pacific Gyre (SPG, 20°S–45°S) is a cold and ultra-oligotrophic environment that is the source of cold-adapted enzymes. However, the characteristic features of psychrophilic enzymes derived from culturable microbes in the SPG remained largely unknown. In this study, the degradation properties of 174 cultures from the deep water of the SPG were used to determine the diversity of cold-adapted enzymes. Thus, the abilities to degrade polysaccharides, proteins, lipids, and DNA at 4, 16, and 28 °C were investigated. Most of the isolates showed one or more extracellular enzyme activities, including amylase, chitinase, cellulase, lipase, lecithinase, caseinase, gelatinase, and DNase at 4, 16, and 28 °C. Moreover, nearly 85.6 % of the isolates produced cold-adapted enzymes at 4 °C. The psychrophilic enzyme-producing isolates distributed primarily in Alteromonas and Pseudoalteromonas genera of the Gammaproteobacteria. Pseudoalteromonas degraded 9 types of macromolecules but not cellulose, Alteromonas secreted 8 enzymes except for cellulase and chitinase. Interestingly, the enzymatic activities of Gammaproteobacteria isolates at 4 °C were higher than those observed at 16 or 28 °C. In addition, we cloned and expressed a gene encoding an α-amylase (Amy2235) from Luteimonas abyssi XH031^T, and examined the properties of the recombinant protein. These cold-active enzymes may have huge potential for academic research and industrial applications. In addition, the capacity of the isolates to degrade various types of organic matter may indicate their unique ecological roles in the elemental biogeochemical cycling of the deep biosphere.     

Molecular Detection of Anaerobic Ammonium-Oxidizing (Anammox) Bacteria in High-Temperature Petroleum Reservoirs

Anaerobic ammonium-oxidizing (anammox) process plays an important role in the nitrogen cycle of the worldwide anoxic and mesophilic habitats. Recently, the existence and activity of anammox bacteria have been detected in some thermophilic environments, but their existence in the geothermal subterranean oil reservoirs is still not reported. This study investigated the abundance, distribution and functional diversity of anammox bacteria in nine out of 17 high-temperature oil reservoirs by molecular ecology analysis. High concentration (5.31–39.2 mg l^?1) of ammonium was detected in the production water from these oilfields with temperatures between 55°C and 75°C. Both 16S rRNA and hzo molecular biomarkers indicated the occurrence of anammox bacteria in nine out of 17 samples. Most of 16S rRNA gene phylotypes are closely related to the known anammox bacterial genera Candidatus Brocadia, Candidatus Kuenenia, Candidatus Scalindua, and Candidatus Jettenia, while hzo gene phylotypes are closely related to the genera Candidatus Anammoxoglobus, Candidatus Kuenenia, Candidatus Scalindua, and Candidatus Jettenia. The total bacterial and anammox bacterial densities were 6.4?±?0.5?×?10³ to 2.0?±?0.18?×?10⁶ cells ml^?1 and 6.6?±?0.51?×?10² to 4.9?±?0.36?×?10⁴ cell ml^?1, respectively. The cluster I of 16S rRNA gene sequences showed distant identity (<92%) to the known Candidatus Scalindua species, inferring this cluster of anammox bacteria to be a new species, and a tentative name Candidatus “Scalindua sinooilfield” was proposed. The results extended the existence of anammox bacteria to the high-temperature oil reservoirs.     

Nitrogen biofiltration capacities and photosynthetic activity of Pyropia yezoensis Ueda (Bangiales,Rhodophyta): groundwork to validate its potential in integrated multi-trophic aquaculture (IMTA)

Porphyra spp. (currently Porphyra and Pyropia) are major sources of seafood globally. In this study, we investigated the effects of ammonium concentration, water temperature, and thallus stocking density on N-ammonium uptake rate (NUR), tissue nutrients content, N–NH₄ ⁺ filtration efficiency (NUE: nitrogen uptake efficiency %) of Pyropia yezoensis at a laboratory scale and in a mesoscale to evaluate the potential of this species as a biofilter. Additionally, photosynthetic activity was examined using Diving-PAM fluorometer to evaluate the health status. At a laboratory scale, the NUR and tissue nitrogen (N) content of P. yezoensis increased with increasing NH₄ ⁺ concentrations in the medium. The NUR at thallus stocking densities of 5 and 10 g fresh weight (FW) L^–1 were significantly higher than that at 20 g FW L^–1. Effective quantum yield (? F/F’ _m ) and tissue N content was significantly higher at all stocking densities than that at the beginning of experiment. The NUE was over 90 % at 10 and 17 °C, while all thalli cultured at 25 °C died after 5 days. In a mesoscale, the NUE at a thallus stocking density of 10.0 g FW L^–1 was significantly higher than that at a stocking density of 5.0 g FW L^–1. No differences in the NUE occurred between 10 °C and 17 °C. Photosynthetic activity (?F/F’_m and rETR_max) of P. yezoensis at optimal culture condition (10–12 °C and 10 g FW L^–1) increased over time through the experiment. This indicates that thallus was healthy during culture and chlorophyll a fluorescence can be as a monitoring tool for evaluating the physiological status of seaweeds in an integrated multi-trophic aquaculture.     

Halorubrum rubrum sp. nov., an extremely halophilic archaeon from a Chinese salt lake

Two halophilic archaeal strains, YC87^T and YCA11, were isolated from Yuncheng salt lake in Shanxi, China. Cells of the two strains were observed to be pleomorphic rod-shaped, stained Gram-negative and produced red-pigmented colonies. Strain YC87^T was able to grow at 20–50 °C (optimum 37 °C), at 1.4–4.8 M NaCl (optimum 2.1 M NaCl), at 0.05–1.0 M MgCl₂ (optimum 0.3 M MgCl₂) and at pH 6.0–9.0 (optimum pH 7.0) while strain YCA11 was able to grow at 20–50 °C (optimum 37 °C), at 2.1–4.8 M NaCl (optimum 3.1 M NaCl), at 0.01–0.7 M MgCl₂ (optimum 0.1 M MgCl₂) and at pH 6.0–9.0 (optimum pH 7.5). The cells of both isolates were observed to lyse in distilled water. The minimum NaCl concentrations that prevented cell lysis were determined to be 8 % (w/v) for strain YC87^T and 12 % (w/v) for strain YCA11. The major polar lipids of the two strains were identified as phosphatidylglycerol, phosphatidylglycerol phosphate methyl ester, phosphatidylglycerol sulfate and one major glycolipid chromatographically identical to sulfated mannosyl glucosyl diether; another major glycolipid and trace amounts of several unidentified lipids were also detected. The 16S rRNA gene sequences of the two strains were 99.8 % identical, showing 93.2–98.2 % similarity to members of the genus Halorubrum of the family Halobacteriaceae. The rpoB′ gene similarity between strains YC87^T and YCA11 was 99.3 % and showed 87.5–95.2 % similarity to the closest relative members of the genus Halorubrum. The DNA G+C content of strains YC87^T and YCA11 were determined to be 64.9 and 64.5 mol%, respectively. The DNA–DNA hybridization value between strain YC20^T and strain YC77 was 87 % and the two strains showed low DNA–DNA relatedness with Halorubrum cibi JCM 15757^T and Halorubrum aquaticum CGMCC 1.6377^T, the most related members of the genus Halorubrum. The phenotypic, chemotaxonomic and phylogenetic properties suggest that strains YC87^T and YCA11 represent a novel species of the genus Halorubrum, for which the name Halorubrum rubrum sp. nov. is proposed. The type strain is YC87^T (=CGMCC 1.12124^T = JCM 18365^T).     

The Tracing of Mycobacteria in Drinking Water Supply Systems by Culture, Conventional, and Real Time PCRs

Mycobacteria are widely present in diverse aquatic habitats, where they can survive for months or years while some species can even proliferate. The resistance of different mycobacterial species to disinfection methods like chlorination or ozonation could result in their presence in the final tap water of consumers. In this study, the culture method, Mycobacterium tuberculosis complex conventional duplex PCR for detection of non-tuberculous mycobacteria (NTM) and quantitative real-time PCR (qPCR) to detect three subspecies of M. avium species (M. a. avium, M. a. hominissuis, and M. a. paratuberculosis) were used to trace their possible path of transmission from the watershed through the reservoir and drinking water plant to raw drinking water and finally to households. A total of 124 samples from four drinking water supply systems in the Czech Republic, 52 dam sediments, 34 water treatment plant sludge samples, and 38 tap water household sediments, were analyzed. NTM of 11 different species were isolated by culture from 42 (33.9 %) samples; the most prevalent were M. gordonae (16.7 %), M. triplex (14.3 %), M. lentiflavum (9.5 %), M. a. avium (7.1 %), M. montefiorenase (7.1 %), and M. nonchromogenicum (7.1 %). NTM DNA was detected in 92 (76.7 %) samples. By qPCR analysis a statistically significant decrease (P < 0.01) was observed along the route from the reservoir (dam sediments), through water treatment sludge and finally to household sediments. The concentrations ranged from 10⁰ to 10⁴ DNA cells/g. It was confirmed that drinking water supply systems (watershed–reservoir–drinking water treatment plant–household) might be a potential transmission route for mycobacteria.     

Productivity and Temperature as Drivers of Seasonal and Spatial Variations of Dissolved Methane in the Southern Bight of the North Sea

Dissolved CH₄ concentrations in the Belgian coastal zone (North Sea) ranged between 670 nmol l^?1 nearshore and 4 nmol l^?1 offshore. Spatial variations of CH₄ were related to sediment organic matter (OM) content and gassy sediments. In nearshore stations with fine sand or muddy sediments, the CH₄ seasonal cycle followed water temperature, suggesting methanogenesis control by temperature in these OM-rich sediments. In offshore stations with permeable sediments, the CH₄ seasonal cycle showed a yearly peak following the chlorophyll-a spring peak, suggesting that in these OM-poor sediments, methanogenesis depended on freshly produced OM delivery. This does not exclude the possibility that some CH₄ might originate from dimethylsulfide (DMS) or dimethylsulfoniopropionate (DMSP) or methylphosphonate transformations in the most offshore stations. Yet, the average seasonal CH₄ cycle was unrelated to those of DMS(P), very abundant during the Phaeocystis bloom. The annual average CH₄ emission was 126 mmol m^?2 y^?1 in the most nearshore stations (~4 km from the coast) and 28 mmol m^?2 y^?1 in the most offshore stations (~23 km from the coast), 1260–280 times higher than the open ocean average value (0.1 mmol m^?2 y^?1). The strong control of CH₄ by sediment OM content and by temperature suggests that marine coastal CH₄ emissions, in particular in shallow areas, should respond to future eutrophication and warming of climate. This is supported by the comparison of CH₄ concentrations at five stations obtained in March 1990 and 2016, showing a decreasing trend consistent with alleviation of eutrophication in the area.