Association Between Environmental Microbiota and Indigenous Bacteria Found in Hemolymph, Extrapallial Fluid and Mucus of Anodonta cygnea (Linnaeus, 1758)

Bivalves filter and accumulate large numbers of microorganisms present in the harvesting water. A complete understanding of the balance between Anodonta cygnea and the microbiota present in their surrounding environment remains incomplete. Therefore, the aim of this study was to quantify and identify the indigenous bacteria in the biological fluids of A. cygnea collected from Mira Lagoon in northern Portugal. The results showed Vibrio metschnikovii and Aeromonas sobria as the dominant groups. The median for total bacteria from mucus was 3.1 × 10³ CFU g⁻¹, whereas the range in means from fluids was 1.5 × 10² to 6.5 × 10² CFU ml⁻¹. During the experimental work, Escherichia coli and enterococci were not detected in healthy A. cygnea. However, the periodic detection of E. coli and enterococci in Mira lagoon revealed its presence in the water. Our observations suggest that A. cygnea has the ability to filter and eliminate E. coli, present in the surrounding environment, through an active phagocytic process conducted by hemolymph circulating cells, the hemocytes.     

Production of recombinant herpes simplex virus protease in 10-L stirred vessels using a baculovirus–insect cell expression system

A gene expression system using recombinant Autographa californica nuclear polyhedrosis virus (baculovirus) and Sf-9 cells has been scaled up to the 10-L tank level and shown to be capable of producing herpes simplex virus (HSV) protease in serum-free media. High densities of Spodoptera frugiperda (Sf-9) cells were achieved by modifying two 10-L Biolafitte fermenters specifically for insect cell growth. The existing Rushton impellers were replaced by marine impellers to reduce shear and the aeration system was modified to allow external addition of air/O₂ mixtures at low flow rates through either the sparge line or into the head space of the fermenter. To inoculate the tanks, Sf-9 cells were adapted to grow to high cell densities (6–10 × 10⁶ cells ml⁻¹) in shake flasks in serum-free media. With these procedures, cell densities of 5 × 10⁶ cells ml⁻¹ were routinely achieved in the 10-L tanks. These cells were readily infected with recombinant baculovirus expressing the 247-amino acid catalytic domain of the HSV-1 strain 17 protease UL26 gene as a glutathione-S-transferase (GST) fusion protein (GST-247). Three days after infection at a multiplicity of infection (MOI) of 3 pfu cell⁻¹, the GST-247 fusion protein was purified from a cytoplasmic lysate by Glutathione Sepharose 4-B affinity chromatography with reproducible yields of 11–38 mg L⁻¹ of recombinant protein and ≥ 90% purity. Maximum production of this protein was observed at a cell density of 5.0 × 10⁶ cells ml⁻¹. Received 09 December 1996/ Accepted in revised form 13 April 1997     

Transmission electron microscope analysis of virus-like particles in the freshwater lakes of Signy Island, Antarctica

Water samples from a range of fresh-water Antarctic lakes on Signy Island (South Orkney Islands: 60°45′S, 45 °38′W) were examined for the presence of virus-like particles (VLPs) during the 1998/1999 field season. It was discovered that VLPs were ubiquitous, morphologically diverse and abundant, with high concentrations ranging from 4.9 × 10⁶ ml⁻¹ to 3.1 × 10⁷ ml⁻¹. Likely hosts include bacteria, cyanobacteria and eukaryotic algae. In addition, an unusually large virus morphotype was observed with a head diameter 370 × 330 nm and a tail 1.3 μm long. Accepted: 15 May 2000     

Modeling growth for predicting the contamination level of guava nectar by Candida pelliculosa under different conditions of pH and storage temperature

The combined effects of temperature (2–46°C) and pH (1.55–6.25) on the growth of Candida pelliculosa isolated from guava nectar produced in Cameroon were studied using a turbidity method, ie measurement of optical density at 630 nm. A quadratic polynomial model was constructed to predict the effects and interactions of these two environmental conditions on the maximal optical density obtained (r ² = 0.97). The relation between optical density and population density of C. pelliculosa (CFU ml⁻¹) was also established using an exponential regression (r ² = 0.99). According to the model, maximal growth conditions were 37°C and pH 6.25 for obtaining the maximal optical density of 1.25 corresponding to about 60 × 10⁶ CFU ml⁻¹. A good agreement of the model was found between the predicted values and the observed values of maximal optical density. The model was validated by the experimental values of maximal optical density obtained in the growth of C. pelliculosa in commercial guava nectar (pH 3.15). Received 01 December 1995/ Accepted in revised form 30 August 1996     

An alginate-layer technique for culture of Brassica oleracea L. protoplasts

Ten accessions belonging to the Brassica oleracea subspecies alba and rubra, and to B. oleracea var. sabauda were used in this study. Protoplasts were isolated from leaves and hypocotyls of in vitro grown plants. The influence of selected factors on the yield, viability, and mitotic activity of protoplasts immobilized in calcium alginate layers was investigated. The efficiency of protoplast isolation from hypocotyls was lower (0.7 ± 0.1 × 10⁶ ml⁻¹) than for protoplasts isolated from leaf mesophyll tissue (2 ± 0.1 × 10⁶ ml⁻¹). High (70–90%) viabilities of immobilized protoplasts were recorded, independent of the explant sources. The highest proportion of protoplasts undergoing divisions was noted for cv. Reball F1, both from mesophyll (29.8 ± 2.2%) and hypocotyl (17.5 ± 0.3%) tissues. Developed colonies of callus tissue were subjected to regeneration and as a result plants from six accessions were obtained.     

Distribution of viruses and their impact on bacterioplankton in mesotrophic and eutrophic reservoirs

Spatial distribution of planktonic viral particles (virioplankton) and mortality of heterotrophic bacteria caused by viral lysis were studied in the eutrophic Ivan’kovskoe and mesotrophic Uglichskoe reservoirs (the Upper Volga). During the summer peak of phytoplankton, the number of viral particles was higher in the Ivan’kovskoe Reservoir ((55.1 ± 9.5) × 10⁶ ml⁻¹ on average) than in the Uglichskoe Reservoir ((42.9 ± 5.1) × 10⁶ ml⁻¹ on average). The ratio of viral to bacterial abundances ranged from 2.5 to 7.0. The average number of mature phages in infected heterotrophic bacteria varied from 17 to 109 particles/cell. Most of the infected bacterial cells in the Ivan’kovskoe Reservoir were rod-shaped, and, in the Uglichskoe Reservoir, they were mainly vibrio-shaped. In the Ivan’kovskoe Reservoir, from 8.3 to 22.4% of planktonic bacteria were infected by phages, suggesting phage-induced mortality of bacterioplankton equal to 10.5–34.8% (19.1% on average) of the daily bacterial production. In the Uglichskoe Reservoir, from 9.4 to 33.5% of bacteria were phage-infected, suggesting phage-induced bacterial mortality of 13.7–40.2% (23.5% on average) of the daily bacterial production. The obtained results testify to an important role of autochthonous viruses in the regulation of bacterioplankton abundance and production in the reservoirs.     

Annual and interannual variability in phytoplankton at a permanent station off Kerguelen Islands, Southern Ocean

From November 1992 to February 1995 a quantitative and qualitative phytoplankton study was conducted at a permanent station (Kerfix) southwest off the Kerguelen Islands, in the vicinity of the Polar Front (50°40′S–68°25′E). Phytoplankton populations are low in this area both during summers and winters. They consist, in order of decreasing cell abundance, of pico- and nanoflagellates (1.5–20 μm), coccolithophorids (<10 μm), diatoms (5–80 μm) and dinoflagellates (6–60 μm). Flagellates form the dominant group throughout the year and attain the highest summer average of 3.0 × 10⁵ cells l⁻¹. Next in abundance year-round are coccolithophorids with the dominant Emiliania huxleyi (highest summer 1992 average 1.9 × 10⁵ cells l⁻¹), diatoms (summer 1992 average 1.0 × 10⁵ cells l⁻¹) and dinoflagellates (average 3.8 × 10⁴ cells l⁻¹). Winter mean numbers of flagellates and picoplankton do not exceed 8.4 × 10⁴ cells l⁻¹; those of the three remaining algal groups together attain 2 × 10⁴ cells l⁻¹. Summer peaks of diatoms and dinoflagellates are mainly due to the larger size species (>20 μm). The latter group contributes most to the total cell carbon biomass throughout the year. Dominant diatoms during summer seasons include: Fragilariopsis kerguelensis, Thalassionema nitzschioides, Chaetoceros dichaeta, C. atlanticus, Pseudonitzschia heimii, and P. barkleyi/lineola. This diatom dominance structure changes from summer to summer with only F. kerguelensis and T. nitzschioides retaining their first and second positions. Any one of the co-dominant species might be absent during some summer period. The variable diatom community structure may be due to southward meandering of the Polar Front bringing “warmer” species from the north, and to the mixing of the water masses in this area. The entire community structure characterized both during summer and winters by the dominance of flagellates can be related to deep mixing (ca. 40–200 m) of the water column as the probable controlling factor. Received: 13 November 1997 / Accepted: 11 May 1998     

The distribution of picoplankton and nanoplankton in Kongsfjorden,Svalbard during late summer 2006

Abundance and biomass of pico- (<2 μm) and nanoplankton (2–20 μm) were investigated in relation to hydrography in Kongsfjorden, Svalbard (79°N, 12°E) during late summer 2006. Autotrophic and heterotrophic picoplankton abundance ranged from 0.1 × 10⁶ to 35.2 × 10⁶ cells L⁻¹ and from 0.4 × 10⁶ to 20.3 × 10⁶ cells L⁻¹, respectively. The highest number of bacteria in the entire water column was recorded at station 2 at 10 m (22.3 × 10⁸ cells L⁻¹); the lowest concentration was observed at station 1 (6.0 × 10⁸ cells L⁻¹). The abundance of autotrophic and heterotrophic nanoplankton varied from 0.4 × 10⁵ cells L⁻¹ to 46 × 10⁵ cells L⁻¹ and from 0.3 × 10⁶ to 9.1 × 10⁶ cells L⁻¹, respectively. Our results demonstrated that heterotrophic nanoflagellates and bacteria in Kongsfjorden microbial community were relatively important. The structure of plankton communities integrated with environmental variables could act as indicators of the variability of the inflow of Atlantic Water into Kongsfjorden.     

Liquid media development for Heterorhabditis bacteriophora : lipid source and concentration

Lipids are important entomopathogenic nematode nutritional components because they are energy reserves and serve as indicators of nematode quality. The composition and concentration of the media lipid component determine bacterial and nematode yields. Heterorhabditis bacteriophora and its symbiont, Photorhabdus luminescens, were cultured in media containing various lipid sources. As lipid concentration increased from 2.5% to 8.0% (w/v), the final yield and productivity [calculated from the number of infective juveniles (IJ)] increased significantly from 2.1 × 10⁵IJ ml⁻¹ to 2.8 × 10⁵IJ ml⁻¹ (P < 0.05) and from 8.9 × 10⁵IJ l⁻¹ day⁻¹ to 11.8 × 10⁵ IJ l⁻¹day⁻¹ (P < 0.05), respectively. The nematode yield coefficient (IJ per gram of media), however, decreased from 2.8 × 10⁶ to 2.2 × 10⁶ (P < 0.05), while recovery increased from 45.3% to 58.0% (P < 0.05). Bacterial cell mass remained constant at 4.6 mg ml⁻¹ with changing lipid content (P > 0.05). The largest nematode yield (2.8 × 10⁵IJ ml⁻¹) was achieved within 8 days, using a medium containing an 8% (w/v) olive and canola oil (50:50 w/v) combination. Moreover, developmental synchrony was achieved in this medium with 96% infective juveniles. In short, lipid sources rich in mono-unsaturated fatty acids and poor in saturated fatty acids produced optimal nematode growth. Received: 1 May 2000 / Received revision: 17 July 2000 / Accepted: 27 July 2000     

Biodegradation of propanol and isopropanol by a mixed microbial consortium

The aerobic biodegradation of high concentrations of 1-propanol and 2-propanol (IPA) by a mixed microbial consortium was investigated. Solvent concentrations were one order of magnitude greater than any previously reported in the literature. The consortium utilized these solvents as their sole carbon source to a maximum cell density of 2.4 × 10⁹ cells ml⁻¹. Enrichment experiments with propanol or IPA as carbon sources were carried out in batch culture and maximum specific growth rates (μ_max) calculated. At 20 °C, μ _max values were calculated to be 0.0305 h⁻¹ and 0.1093 h⁻¹ on 1% (v/v) IPA and 1-propanol, respectively. Growth on propanol and IPA was carried out between temperatures of 10 °C and 45 °C. Temperature shock responses by the microbial consortium at temperatures above 45 °C were demonstrated by considerable cell flocculation. An increase in propanol substrate concentration from 1% (v/v) to 2% (v/v) decreased the μ _max from 0.1093 h⁻¹ to 0.0715 h⁻¹. Maximum achievable biodegradation rates of propanol and IPA were 6.11 × 10⁻³% (v/v) h⁻¹ and 2.72 × 10⁻³% (v/v) h⁻¹, respectively. Generation of acetone during IPA biodegradation commenced at 264 h and reached a maximum concentration of 0.4% (v/v). The results demonstrate the potential of mixed microbial consortia in the bioremediation of solvent-containing waste streams. Received: 14 December 1999 / Received revision: 3 April 2000 / Accepted: 7 April 2000     

Degradation of naphthalene by cells of Pseudomonas sp. strain NGK 1 immobilized in alginate, agar and polyacrylamide

A Pseudomonas sp. strain NGK 1 (NCIM 5120) was immobilized in various matrices, namely, alginate, agar (1.8 × 10¹¹ cfu g⁻¹ beads) and polyacrylamide (1.6 × 10¹¹ cfu g⁻¹ beads). The degradation of naphthalene was studied, by freely suspended cells (4 × 10¹⁰ cfu ml⁻¹) and immobilized cells in batches, with shaken culture and continuous degradation in a packed-bed reactor. Free cells brought about the complete degradation of 25 mmol naphthalene after 3 days of incubation, whereas, a maximum of 30 mmol naphthalene was degraded by the bacteria after 3–4 days of incubation with 50 mmol and 75 mmol naphthalene, and no further degradation was observed even after 15 days of incubation. Alginate-entrapped cells had degraded 25 mmol naphthalene after 3.5 days of incubation, whereas agar- and polyacrylamide-entrapped cells took 2.5 days; 50 mmol naphthalene was completely degraded by the immobilized cells after 6–7 days of incubation. Maximum amounts of 55 mmol, 70 mmol and 67 mmol naphthalene were degraded, from an initial 75 mmol naphthalene, by the alginate-, agar- and polyacrylamide-entrapped cells after 15 days of incubation. When the cell concentrations were doubled, 25 mmol and 50 mmol naphthalene were degraded after 2 and 5.5 days of incubation by the immobilized cells. Complete degradation of 75 mmol naphthalene occurred after 10 days incubation with agar- and polyacrylamide-entrapped␣cells, whereas only 60 mmol naphthalene was degraded by alginate-entrapped cells after 15 days of␣incubation. Further, with 25 mmol naphthalene, alginate-, agar- and polyacrylamide-entrapped cells (1.8 × 10¹¹ cfu g⁻¹ beads) could be reused 18, 12 and 23 times respectively. During continuous degradation in a packed-bed reactor, 80 mmol naphthalene 100 ml⁻¹ h⁻¹ was degraded by alginate- and polyacrylamide-entrapped cells whereas 80 mmol naphthalene 125 ml⁻¹␣h⁻¹ was degraded by agar-entrapped cells. Received: 21 October 1997 / Received revision: 15 January 1998 / Accepted: 18 January 1998     

Changes in bacterial and ciliate densities with trophic status in Mediterranean shallow lakes

Ciliate and bacterial densities and their link with eutrophication were studied in fourteen shallow lakes in northwest Spain. Total phosphorus (TP) in these lakes varied between 30 μg l⁻¹ and 925 μg l⁻¹ and chlorophyll a concentration (chla) between 0.5 μg l⁻¹ and 107 μg l⁻¹. Bacterial abundance ranged from 1 × 10⁶ to 14 × 10⁶ cells ml⁻¹, while ciliate abundance ranged from 0.6 cells ml⁻¹ to 229 cells ml⁻¹. Lakes were classified into three trophic types from their TP and chla concentrations. Bacterial abundance was significantly correlated with trophic type, as well as with TP and with chla separately, whereas ciliate abundance was only correlated with chla. No significant relationship could be established between bacterial and ciliate abundance across the trophic gradient. A general pattern was observed in the ratios of bacterial abundance to TP and chla concentrations, of decreasing ratios with increases in the nutrient loading. This pattern was not found for ciliates. The dominant zooplankton group in 13 of the 14 lakes studied was Rotifera, which accounted for a mean of 71% of total zooplankton abundance (41% of zooplankton biomass). The positive correlation between bacteria and ciliates with this group, and the absence of any relationship with Cladocera suggest that top down control by cladocerans was weaker in our lakes than previously shown in northern European shallow lakes. Rotifers could be important predators of bacteria in the high-nutrient lakes of our study. Higher slopes of regressions on bacterial abundance towards the hypertrophic range indicate that top-down control was weaker in our lakes than in northern European shallow lakes.     

Enhancement of Glutaraldehyde Biocidal Efficacy by the Application of an Electric Field. Effect on Sessile Cells and on Cells Released by the Biofilm

Summary The aim of this paper was to evaluate the possible enhancement of the biocidal efficacy of glutaraldehyde against Pseudomonas fluorescens biofilms by the application of an electric field. The behaviour of sessile cells and cells released by the biofilms was assed. Biofilms were formed on thin stainless steel coupons immersed in culture media inoculated with Pseudomonas fluorescens. Treatments using glutaraldehyde (TGA) and both glutaraldehyde and electric field application (TGAEF) were carried out with the samples with biofilms. TGA: samples with biofilms were immersed in glass cells containing a buffer solution with different glutaraldehyde concentrations in the 25–500 ppm range. TGAEF: samples with biofilms were immersed in an electrochemical cell containing glutaraldehyde solution where a direct electric current (4 × 10⁻⁴ A cm⁻²) was delivered to the chamber. The evolution of biofilms was observed through optical microscopy at real time. Results show that the electric field enhanced glutaraldehyde efficacy reducing the number of surviving cells in the range of one to four orders with respect to those with TGA treatment. The sensitivity of the cells to the treatments decreased in the following order: planktonic cells > cells released by the biofilm > sessile cells.     

Relevance of bacterioplankton abundance and production in the oligotrophic equatorial Indian Ocean

Bacterioplankton abundance and production, chlorophyll a (Chl a) concentrations and primary production (PP) were measured from the equatorial Indian Ocean (EIO) during northeast (NEM), southwest (SWM) and spring intermonsoon (SpIM) seasons from 1°N to 5°S along 83°E. The average bacterial abundance was 0.52 ± 0.29, 0.62 ± 0.33 and 0.46 ± 0.19 (× 10⁸ cells l⁻¹), respectively during NEM, SWM and SpIM in the top 100 m. In the deep waters (200 m and below), the bacterial counts averaged ∼0.35 ± 0.14 × 10⁸ cells l⁻¹ in SWM and 0.39 ± 0.16 × 10⁸ cells l⁻¹ in SpIM. The 0–120 m column integrated bacterial production (BP) ranged from 19 to 115 and from 10 to 51 mg C m⁻² d⁻¹ during NEM and SWM, respectively. Compared with many open ocean locations, bacterial abundance and production in this region are lower. The bacterial carbon production, however, is notably higher than that of phytoplankton PP (BP:PP ratio 102% in SWM and 188% in NEM). With perpetually low PP (NEM: 20, SWM: 18 and SpIM: 12 mg C m⁻² d⁻¹) and Chl a concentration (NEM: 16.5, SWM: 15.0 and SpIM: 20.9 mg m⁻²), the observed bacterial abundance and production are pivotal in the trophodynamics of the EIO. Efficient assimilation and mineralization of available organics by bacteria in the euphotic zone might serve a dual role in the ultra-oligotrophic regions including EIO. Thus, bacteria probably sustain microheterotrophs (micro- and meso-zooplankton) through microbial loop. Further, rapid mineralization by bacteria will make essential nutrients available to autotrophs.     

Placental Alpha Hemoglobin Stabilizing Protein (AHSP) and recurrent miscarriage

AHSP inhibits cellular production of the reactive oxygen species. Reduced AHSP indicates reduced protection against oxidative stressors. Our objective was to investigate AHSP levels in recurrent miscarriage (RM). Trophoblast was collected from women of 10 weeks gestation: voluntary abortion controls (VA, n = 10); spontaneous first miscarriage with subsequent normal pregnancy (SMSN, n = 15) or with subsequent miscarriage (SMSM, n = 5); RM previously investigated (RMPS, n = 5) or not previously investigated (RM, n = 5). AHSP mRNA and protein were determined using real-time quantitative polymerase chain reaction (PCR) and Western blot, respectively. One-way ANOVA was performed to assess statistical significance (p < 0.05). ahsp mRNA levels were maximally reduced in RM and RMPS (8.0 × 10⁻⁶ ± 1.3 and 8.1 × 10⁻⁶ ± 0.7, respectively) compared with SMSN and VA (16.1 × 10⁻⁶ ± 2.3 and 26.1 × 10⁻⁶ ± 2.7, respectively). SMSM showed levels significantly reduced as well (9.0 × 10⁻⁶ ± 2.3). In RM, a reduced defense from oxidative stressors is evident at first miscarriage, identifying women at high risk for subsequent eventful pregnancy. Reduced AHSP may identify women at risk of experiencing further miscarriages. Monica Emanuelli and Monia Cecati contributed equally to this paper.     

Combined effects of zinc and algal food on the competition between planktonic rotifers, <Emphasis Type="Italic">Anuraeopsis fissa</Emphasis> and <Emphasis Type="Italic">Brachionus rubens</Emphasis> (Rotifera)

We evaluated the combined effects of algal (Chlorella vulgaris) food levels (low, 0.5 × 10⁶ (or 2.9 μg C ml⁻¹); and high, 1 × 10⁶ cells ml⁻¹ (or 5.8 μg C ml⁻¹)) and zinc concentrations (0, 0.125, and 0.250 mg l⁻¹ of ZnCl₂) on the competition between two common planktonic rotifers Anuraeopsis fissa and Brachionus rubens using their population growth. Median lethal concentration data (LC₅₀) (mean ± 95% confidence intervals) showed that B. rubens was more resistant to zinc (0.554 ± 0.08 mg l⁻¹) than A. fissa (0.315 ± 0.07 mg l⁻¹). A. fissa when grown alone or with Zn was always numerically more abundant than B. rubens. When grown in the absence of zinc, under low- and high-food levels, the peak abundances of A. fissa varied from 251 ± 24 to 661 ± 77 ind. ml⁻¹, respectively, and the corresponding maxima for B. rubens were 52 ± 3 and 102 ± 18 ind. ml⁻¹. At a given food level, competition for food reduced the peak abundances of both rotifers considerably. Increase in Zn concentration also lowered the rotifer abundances. The impact of zinc on competition between the two-rotifer species was evident at low-food level, mainly for A. fissa. At zinc concentrations of 0 and 0.125 mg l⁻¹, the populations of both rotifers continued to grow for about 10 days, but thereafter B. rubens began to decline. Role of zinc on the competitive outcome of the two species is discussed in relation to the changing algal densities in natural water bodies.     

Effects of Contact Time, Pressure, Percent Relative Humidity (%RH), and Material Type on Listeria Biofilm Adhesive Strength at a Cellular Level Using Atomic Force Microscopy (AFM)

Whereas the transfer of Listeria from surfaces to foods and vice versa has been well documented, little is known about the mechanism of bacterial transfer. The objective of this work is to gain a better understanding of the forces involved in listerial biofilms adhesion using atomic force microscopy (AFM). L. monocytogenes Scott A was grown as biofilms on stainless steel surfaces by inoculating stainless steel coupons with Listeria and incubating the coupons for 48 h at 32 °C with a diluted 1:20 tryptic soy broth. After growth, biofilms were equilibrated over saturated salt solutions at a constant relative humidity (%RH) before measurement of adhesion forces using AFM. The effects of contact time, loading force, and biofilm relative humidity (%RH) suggested that neither contact time, loading force nor biofilm %RH had a significant effect on biofilm adhesiveness at a cellular level (P > 0.05). In a second set of experiments, the influence of material type on biofilm adhesiveness was evaluated using two different colloidal probes (SiO₂ and polyethylene). Results showed that the maximum pull-off force and retraction work needed to retract the cantilever for glass (−85.42 nN and 1.610⁻¹⁵ J, respectively) were significantly lower than those of polyethylene (−113.38 nN and 2.7 × 10^–15 J, respectively; P < 0.001). The results of this study suggest that Listeria biofilms adhere more strongly to hydrophobic surfaces than hydrophilic surfaces when measured at a cellular level. These results provide important insights that could lead to new ways to remediate and avoid listerial biofilm formation in the food industry.     

Change in ammonia-oxidizing microorganisms in enriched nitrifying activated sludge

In this study, sludge was taken from a municipal wastewater treatment plant that contained a nearly equal number of archaeal amoA genes (5.70 × 10⁶ ± 3.30 × 10⁵ copies mg sludge⁻¹) to bacterial amoA genes (8.60 × 10⁶ ± 7.64 × 10⁵ copies mg sludge⁻¹) and enriched in three continuous-flow reactors receiving an inorganic medium containing different ammonium concentrations: 2, 10, and 30 mM NH₄⁺–N (28, 140, and 420 mg N l⁻¹). The abundance and communities of ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) in enriched nitrifying activated sludge (NAS) were monitored at days 60 and 360 of the operation. Early on, between day 0 and day 60 of reactor operation, comparative abundance of AOA amoA genes to AOB amoA genes varied among the reactors depending on the ammonium levels found in the reactors. As compared to the seed sludge, the number of AOA amoA genes was unchanged in the reactor with lower ammonium level (0.06 ± 0.04 mgN l⁻¹), while in the reactors with higher ammonium levels (0.51 ± 0.33 and 0.25 ± 0.10 mgN l⁻¹), the numbers of AOA amoA genes were deteriorated. By day 360, AOA disappeared from the ammonia-oxidizing consortiums in all reactors. The majority of the AOA sequences from all NASs at each sampling period fell into a single AOA cluster, however, suggesting that the ammonium did not affect the AOA communities under this operational condition. This result is contradictory to the case of AOB, where the communities varied significantly among the NASs. AOB with a high affinity for ammonia were present in the reactors with lower ammonium levels, whereas AOB with a low affinity to ammonia existed in the reactors with higher ammonium levels.     

Studies on water transport through the sweet cherry fruit surface: characterizing conductance of the cuticular membrane using pericarp segments

Water conductance of the cuticular membrane (CM) of mature sweet cherry fruit (Prunus avium L. cv. Sam) was investigated by monitoring water loss from segments of the outer pericarp excised from the cheek of the fruit. Segments consisted of epidermis, hypodermis and several cell layers of the mesocarp. Segments were mounted in stainless-steel diffusion cells with the mesocarp surface in contact with water, while the outer cuticular surface was exposed to dry silica (22 ± 1 °C). Conductance was calculated by dividing the amount of water transpired per unit area and time by the difference in water vapour concentration across the segment. Conductance values had a log normal distribution with a median of 1.15 × 10⁻⁴ m s⁻¹ (n=357). Transpiration increased linearly with time. Conductance remained constant and was not affected by metabolic inhibitors (1 mM NaN₃ or 0.1 mM carbonylcyanide m-chlorophenylhydrazone) or thickness of segments (range 0.8–2.8 mm). Storing fruit (up to 42 d, 1 °C) used as a source of segments had no consistent effect on conductance. Conductance of the CM increased from cheek (1.16 ± 0.10 × 10⁻⁴ m s⁻¹) to ventral suture (1.32 ± 0.07 × 10⁻⁴ m s⁻¹) and to stylar end (2.53 ± 0.17 × 10⁻⁴ m s⁻¹). There was a positive relationship (r²=0.066**; n=108) between conductance and stomatal density. From this relationship the cuticular conductance of a hypothetical astomatous CM was estimated to be 0.97 ± 0.09 × 10⁻⁴ m s⁻¹. Removal of epicuticular wax by stripping with cellulose acetate or extracting epicuticular plus cuticular wax by dipping in CHCl₃/methanol increased conductance 3.6- and 48.6-fold, respectively. Water fluxes increased with increasing temperature (range 10–39 °C) and energies of activation, calculated for the temperature range from 10 to 30 °C, were 64.8 ± 5.8 and 22.2 ± 5.0 kJ mol⁻¹ for flux and vapour-concentration-based conductance, respectively. Received: 23 March 2000 / Accepted: 28 July 2000     

Antarctic sea ice viral dynamics over an annual cycle

Viral abundance, burst sizes, lytic production and temperate phage were investigated in land-fast ice at two sites in Prydz Bay Antarctica (68°S, 77°E) between April and November 2008. Both ice cores and brine were collected. There was no seasonal pattern in viral or bacterial numbers. Across the two sites virus abundances ranged between 0.5 × 10⁵ and 5.1 × 10⁵ viruses ml⁻¹ in melted ice cores and 0.6 × 10⁵–3.5 × 10⁵ viruses ml⁻¹ in brine, and bacterial abundances between 2.7 × 10⁴ and 17.3 × 10⁴ cells ml⁻¹ in melted ice cores and 3.9 × 10⁴–32.5 × 10⁴ cells ml⁻¹ in brine. Virus to bacterium ratios (VBR) showed a clear seasonal pattern in ice cores with lowest values in winter (range 1.2–20.8), while VBRs in brine were lower (0.2–4.9). Lytic viral production range from undetectable to 2.0 × 10⁴ viruses ml⁻¹ h⁻¹ in ice cores with maximum rates in September and November. In brine maximum, lytic viral production occurred in November (1.18 × 10⁴ viruses ml⁻¹ h⁻¹). Low burst sizes were typical (3.94–4.03 viruses per bacterium in ice cores and 3.16–4.0 viruses per bacterium in brine) with unusually high levels of visibly infected cells—range 40–50%. This long-term investigation revealed that viral activity was apparent within the sea ice throughout its annual cycle. The findings are discussed within the context of limited data available on viruses in sea ice.