Evaluation of VOCs from fungal strains, building insulation materials and indoor air by solid phase microextraction arrow, thermal desorption-gas chromatography-mass spectrometry and machine learning approaches.

Solid phase microextraction Arrow and thermal desorption-gas chromatography-mass spectrometry allowed the collection and evaluation of volatile organic compounds (VOCs) emitted by fungal cultures from building insulation materials and in indoor air. Principal component analysis, linear discriminant analysis and supported vector machine were used for visualization and statistical assessment of differences between samples. In addition, a screening tool based on the soft independent modelling of class analogies (SIMCA) was developed for identification of fungal contamination of indoor air. Ten different fungal strains, incubated under ambient and microaerophilic conditions, were analyzed for time period ranging from 5 to 29 days after inoculation resulting in a total of 140 samples. In addition, the effect of additives on the fungal growing media was studied. The total number of compounds and concentration values were used for the evaluation of the results. Clear differences were observed for VOC profiles emitted by different fungal strains by exploiting long chain alcohols (3-octanol, 1-hexanol and 2-octen-1-ol) and sesquiterpenes (farnesene, cuprene). The analysis of glass-wool and cellulose based building insulation materials (3 samples) gave clear differences, mainly for oxygenated compounds (ethyl acetate and hexanal) and benzenoids (benzaldehyde). Moreover, the comparison of indoor air and insulation materials collected from a house with fungal indoor air problems indicated that 42% of the VOCs were found in both samples. The analysis of 52 indoor air samples demonstrated clear differences in their VOC profiles, especially for hydrocarbons, and between control (44 samples) and indoor air problem houses (8 samples). Finally, the SIMCA model enabled to recognize differences between control and fungi contaminated houses with a prediction capacity over 84%.

Newtonian boreal forest ecology: The Scots pine ecosystem as an example.

Isaac Newton's approach to developing theories in his book Principia Mathematica proceeds in four steps. First, he defines various concepts, second, he formulates axioms utilising the concepts, third, he mathematically analyses the behaviour of the system defined by the concepts and axioms obtaining predictions and fourth, he tests the predictions with measurements. In this study, we formulated our theory of boreal forest ecosystems, called NewtonForest, following the four steps introduced by Newton. The forest ecosystem is a complicated entity and hence we needed altogether 27 concepts to describe the material and energy flows in the metabolism of trees, ground vegetation and microbes in the soil, and to describe the regularities in tree structure. Thirtyfour axioms described the most important features in the behaviour of the forest ecosystem. We utilised numerical simulations in the analysis of the behaviour of the system resulting in clear predictions that could be tested with field data. We collected retrospective time series of diameters and heights for test material from 6 stands in southern Finland and five stands in Estonia. The numerical simulations succeeded to predict the measured diameters and heights, providing clear corroboration with our theory.

Streptomyces strains producing mitochondriotoxic antimycin A found in cereal grains.

Reasons for mammalian cell toxicity observed in barley and spring wheat grains were sought. Streptomyces sp. isolates from wheat and barley produced heat-stable methanol-soluble substances which inhibited the motility of exposed porcine spermatozoa used as a toxicity indicator. Several barley isolates produced antimycin A (2 to 5 ng/mg wet wt of biomass), a macrolide antibiotic known to block oxygen utilization in mitochondria. The antimycin-producing isolates were members of the Streptomyces albidoflavus group. In in vitro assays with porcine kidney tubular epithelial cells, the specific toxicity of antimycin A towards mitochondria was higher than that of the mycotoxin enniatin B but lower than that of the mitochondriotoxins cereulide and paenilide, produced by food-related Bacillus cereus and Paenibacillus tundrae, respectively. The toxic wheat isolates, related to Streptomyces sedi, did not produce antimycin A and or any other known toxin. Our results suggest that the presence of toxin-producing streptomycetes in stored cereal grains may pose a thus far unrecognized threat for food and feed safety.

Toxic indole alkaloids avrainvillamide and stephacidin B produced by a biocide tolerant indoor mold Aspergillus westerdijkiae.

Toxic Aspergillus westerdijkiae were present in house dust and indoor air fall-out from a residence and a kindergarten where the occupants suffered from building related ill health. The A. westerdijkiae isolates produced indole alkaloids avrainvillamide (445 Da) and its dimer stephacidin B (890 Da). It grew and sporulated in presence of high concentrations of boron or polyguanidine (PHMB, PHMG) based antimicrobial biocides used to remediate mold infested buildings. The boar sperm cells were used as sensor cells to purify toxins from HPLC fractions of the fungal biomass. Submicromolar concentrations (EC50 0.3-0.4 µM) blocked boar spermatozoan motility and killed porcine kidney tubular epithelial cells (PK-15). Plate grown hyphal mass of the A. westerdijkiae isolates contained 300-750 ng of avrainvillamide and 30-300 ng of stephacidin B per mg (wet weight). The toxins induced rapid (30 min) loss of boar sperm motility, followed (24 h) by loss of mitochondrial membrane potential (ΔΨm). Apoptotic cell death was observed in PK-15 cell monolayers, prior to cessation of glucose uptake or loss of ΔΨm. Avrainvillamide and stephacidin B were 100-fold more potent towards the porcine cells than the mycotoxins stephacidin A, ochratoxin A, sterigmatocystin and citrinin. The high toxicity of stephacidin B indicates a role of nitrone group in the mechanism of toxicity. Avrainvillamide and stephacidin B represent a new class of toxins with possible a threat to human health in buildings. Furthermore, the use of biocides highly enhanced the growth of toxigenic A. westerdijkiae.

The peptide toxin amylosin of Bacillus amyloliquefaciens from moisture-damaged buildings is immunotoxic, induces potassium efflux from mammalian cells, and has antimicrobial activity.

Amylosin, a heat-stable channel-forming non-ribosomally synthesized peptide toxin produced by strains of Bacillus amyloliquefaciens isolated from moisture-damaged buildings, is shown in this paper to have immunotoxic and cytotoxic effects on human cells as well as antagonistic effects on microbes. Human macrophages exposed to 50 ng of amylosin ml(-1) secreted high levels of cytokines interleukin-1ß (IL-1ß) and IL-18 within 2 h, indicating activation of the NLRP3 inflammasome, an integral part of the innate immune system. At the same exposure level, expression of IL-1ß and IL-18 mRNA increased. Amylosin caused dose-dependent potassium ion efflux from all tested mammalian cells (human monocytes and keratinocytes and porcine sperm cells) at 1 to 2 µM exposure. Amylosin also inhibited the motility of porcine sperm cells and depolarized the mitochondria of human keratinocytes. Amylosin may thus trigger the activation of the NLRP3 inflammasome and subsequently cytokine release by causing potassium efflux from exposed cells. The results of this study indicate that exposure to amylosin activates the innate immune system, which could offer an explanation for the inflammatory symptoms experienced by occupants of moisture-damaged buildings. In addition, the amylosin-producing B. amyloliquefaciens inhibited the growth of both prokaryotic and eukaryotic indoor microbes, and purified amylosin also had an antimicrobial effect. These antimicrobial effects could make amylosin producers dominant and therefore significant causal agents of health problems in some moisture-damaged sites.

Mitochondrial toxicity of triclosan on mammalian cells.

Effects of triclosan (5-chloro-2'-(2,4-dichlorophenoxy)phenol) on mammalian cells were investigated using human peripheral blood mono nuclear cells (PBMC), keratinocytes (HaCaT), porcine spermatozoa and kidney tubular epithelial cells (PK-15), murine pancreatic islets (MIN-6) and neuroblastoma cells (MNA) as targets. We show that triclosan (1-10 µg ml-1) depolarised the mitochondria, upshifted the rate of glucose consumption in PMBC, HaCaT, PK-15 and MNA, and subsequently induced metabolic acidosis. Triclosan induced a regression of insulin producing pancreatic islets into tiny pycnotic cells and necrotic death. Short exposure to low concentrations of triclosan (30 min, ≤1 µg/ml) paralyzed the high amplitude tail beating and progressive motility of spermatozoa, within 30 min exposure, depolarized the spermatozoan mitochondria and hyperpolarised the acrosome region of the sperm head and the flagellar fibrous sheath (distal part of the flagellum). Experiments with isolated rat liver mitochondria showed that triclosan impaired oxidative phosphorylation, downshifted ATP synthesis, uncoupled respiration and provoked excessive oxygen uptake. These exposure concentrations are 100-1000 fold lower that those permitted in consumer goods. The mitochondriotoxic mechanism of triclosan differs from that of valinomycin, cereulide and the enniatins by not involving potassium ionophoric activity.

Ophiobolin A from Bipolaris oryzae perturbs motility and membrane integrities of porcine sperm and induces cell death on mammalian somatic cell lines.

Bipolaris oryzae is a phytopathogenic fungus causing a brown spot disease in rice, and produces substance that strongly perturbs motility and membrane integrities of boar spermatozoa. The substance was isolated from the liquid culture of the fungal strain using extraction and a multi-step semi-preparative HPLC procedures. Based on the results of mass spectrometric and 2D NMR techniques, the bioactive molecule was identified as ophiobolin A, a previously described sesterterpene-type compound. The purified ophiobolin A exhibited strong motility inhibition and viability reduction on boar spermatozoa. Furthermore, it damaged the sperm mitochondria significantly at sublethal concentration by the dissipation of transmembrane potential in the mitochondrial inner membrane, while the plasma membrane permeability barrier remained intact. The study demonstrated that the cytotoxicity of ophiobolin A toward somatic cell lines is higher by 1-2 orders of magnitude compared to other mitochondriotoxic mycotoxins, and towards sperm cells unique by replacing the progressive motility by shivering tail beating at low exposure concentration.

Potato crop as a source of emetic Bacillus cereus and cereulide-induced mammalian cell toxicity.

Bacillus cereus, aseptically isolated from potato tubers, were screened for cereulide production and for toxicity on human and other mammalian cells. The cereulide-producing isolates grew slowly, the colonies remained small (~1 mm), tested negative for starch hydrolysis, and varied in productivity from 1 to 100 ng of cereulide mg (wet weight)(-1) (~0.01 to 1 ng per 10(5) CFU). By DNA-fingerprint analysis, the isolates matched B. cereus F5881/94, connected to human food-borne illness, but were distinct from cereulide-producing endophytes of spruce tree (Picea abies). Exposure to cell extracts (1 to 10 µg of bacterial biomass ml(-1)) and to purified cereulide (0.4 to 7 ng ml(-1)) from the potato isolates caused mitochondrial depolarization (loss of ΔΨm) in human peripheral blood mononuclear cells (PBMC) and keratinocytes (HaCaT), porcine spermatozoa and kidney tubular epithelial cells (PK-15), murine fibroblasts (L-929), and pancreatic insulin-producing cells (MIN-6). Cereulide (10 to 20 ng ml(-1)) exposed pancreatic islets (MIN-6) disintegrated into small pyknotic cells, followed by necrotic death. Necrotic death in other test cells was observed only after a 2-log-higher exposure. Exposure to 30 to 60 ng of cereulide ml(-1) induced K(+) translocation in intact, live PBMC, keratinocytes, and sperm cells within seconds of exposure, depleting 2 to 10% of the cellular K(+) stores within 10 min. The ability of cereulide to transfer K(+) ions across biological membranes may benefit the producer bacterium in K(+)-deficient environments such as extracellular spaces inside plant tissue but is a pathogenic trait when in contact with mammalian cells.

20-Residue and 11-residue peptaibols from the fungus Trichoderma longibrachiatum are synergistic in forming Na+/K+ -permeable channels and adverse action towards mammalian cells.

Certain species of the filamentous fungal genus Trichoderma (e.g. Trichoderma longibrachiatum and Trichoderma citrinoviride) are among the emerging clinical pathogens and also the most common species in the indoor space of mould-damaged buildings. The molecules involved in its pathology are not known. In the present study, we report that 0.5-2.6 wt% of the T. longibrachiatum mycelial biomass consisted of thermostable secondary metabolites mitochondriotoxic to mammalian cells. These were identified by LC/MS as one 11-residue and eight 20-residue peptaibols, AcAib-Asn-Leu/Ile-Leu/Ile-Aib-Pro-Leu/Ile-Leu/Ile-Aib-Pro-Leuol/Ileol (1175 Da) and AcAib-Ala-Aib-Ala-Aib-Ala/Aib-Gln-Aib-Val/Iva-Aib-Gly-Leu/Ile-Aib-Pro-Val/Iva-Aib-Val/Iva/Aib-Gln/Glu-Gln-Pheol(1936-1965 Da) (Aib, α-aminoisobutyric acid; Ac, acetyl; Ileol, isoleucinol; Iva, isovaline; Leuol, leucinol; Pheol, phenylalaninol). The toxic effects on boar sperm cells depended on these peptaibols, named trilongins. The trilongins formed voltage dependent, Na(+)/K(+) permeable channels in biomembranes. The permeability ratios for Na(+) ions, relative to K(+), of the 11-residue trilongin channel (0.95 : 1) and the 20-residue trilongin channel (0.8 : 1) were higher than those of alamethicin. The combined 11-residue and 20-residue trilongins generated channels that remained in an open state for a longer time than those formed by either one of the peptaibols alone. Corresponding synergy was observed in toxicokinetics. With 11-residue and 20-residue trilongins combined 1 : 2 w/w, an effective median concentration (EC(50) ) of 0.6 µg·mL(-1) was reached within 30 min, and the EC(50) shifted down to 0.2 µg·mL(-1) upon extended exposure. By contrast, with 11-residue or 20-residue trilonging separately in 30 min of exposure, the EC(50) values were 15 and 3 µg·mL(-1) , respectively, and shifted down to 1.5 and 0.4 µg·mL(-1) upon extended exposure. This is the first report on ion-channel forming peptaibols with synergistic toxicity from T. longibrachiatum strains isolated from clinical samples.

BC4707 is a major facilitator superfamily multidrug resistance transport protein from Bacillus cereus implicated in fluoroquinolone tolerance.

Transcriptional profiling highlighted a subset of genes encoding putative multidrug transporters in the pathogen Bacillus cereus that were up-regulated during stress produced by bile salts. One of these multidrug transporters (BC4707) was selected for investigation. Functional characterization of the BC4707 protein in Escherichia coli revealed a role in the energized efflux of xenobiotics. Phenotypic analyses after inactivation of the gene bc4707 in Bacillus cereus ATCC14579 suggested a more specific, but modest role in the efflux of norfloxacin. In addition to this, transcriptional analyses showed that BC4707 is also expressed during growth of B. cereus under non-stressful conditions where it may have a role in the normal physiology of the bacteria. Altogether, the results indicate that bc4707, which is part of the core genome of the B. cereus group of bacteria, encodes a multidrug resistance efflux protein that is likely involved in maintaining intracellular homeostasis during growth of the bacteria.

Psychrotolerant Paenibacillus tundrae isolates from barley grains produce new cereulide-like depsipeptides (paenilide and homopaenilide) that are highly toxic to mammalian cells.

Paenilide is a novel, heat-stable peptide toxin from Paenibacillus tundrae, which colonizes barley. P. tundrae produced 20 to 50 ng of the toxin mg(-1) of cells (wet weight) throughout a range of growth temperatures from +5°C to +28°C. Paenilide consisted of two substances of 1,152 Da and 1,166 Da, with masses and tandem mass spectra identical to those of cereulide and a cereulide homolog, respectively, produced by Bacillus cereus NS-58. The two components of paenilide were separated from those of cereulide by high-performance liquid chromatography (HPLC), showing a structural difference suggesting the replacement of O-Leu (cereulide) by O-Ile (paenilide). The exposure of porcine spermatozoa and kidney tubular epithelial (PK-15) cells to subnanomolar concentrations of paenilide resulted in inhibited motility, the depolarization of mitochondria, excessive glucose consumption, and metabolic acidosis. Paenilide was similar to cereulide in eight different toxicity endpoints with porcine and murine cells. In isolated rat liver mitochondria, nanomolar concentrations of paenilide collapsed respiratory control, zeroed the mitochondrial membrane potential, and induced swelling. The toxic effect of paenilide depended on its high lipophilicity and activity as a high-affinity potassium ion carrier. Similar to cereulide, paenilide formed lipocations, i.e., lipophilic cationic compounds, with K(+) ions already at 4 mM [K(+)], rendering lipid membranes electroconductive. Paenilide-producing P. tundrae was negative in a PCR assay with primers specific for the cesB gene, indicating that paenilide was not a product of plasmid pCER270, encoding the biosynthesis of cereulide in B. cereus. Paenilide represents the first potassium ionophoric compound described for Paenibacillus. The findings in this paper indicate that paenilide from P. tundrae is a potential food-poisoning agent.

Physiology of resistant Deinococcus geothermalis bacterium aerobically cultivated in low-manganese medium.

This dynamic proteome study describes the physiology of growth and survival of Deinococcus geothermalis, in conditions simulating paper machine waters being aerobic, warm, and low in carbon and manganese. The industrial environment of this species differs from its natural habitats, geothermal springs and deep ocean subsurfaces, by being highly exposed to oxygen. Quantitative proteome analysis using two-dimensional gel electrophoresis and bioinformatic tools showed expression change for 165 proteins, from which 47 were assigned to a function. We propose that D. geothermalis grew and survived in aerobic conditions by channeling central carbon metabolism to pathways where mainly NADPH rather than NADH was retrieved from the carbon source. A major part of the carbon substrate was converted into succinate, which was not a fermentation product but likely served combating reactive oxygen species (ROS). Transition from growth to nongrowth resulted in downregulation of the oxidative phosphorylation observed as reduced expression of V-type ATPase responsible for ATP synthesis in D. geothermalis. The battle against oxidative stress was seen as upregulation of superoxide dismutase (Mn dependent) and catalase, as well as several protein repair enzymes, including FeS cluster assembly proteins of the iron-sulfur cluster assembly protein system, peptidylprolyl isomerase, and chaperones. Addition of soluble Mn reinitiated respiration and proliferation with concomitant acidification, indicating that aerobic metabolism was restricted by access to manganese. We conclude that D. geothermalis prefers to combat ROS using manganese-dependent enzymes, but when manganese is not available central carbon metabolism is used to produce ROS neutralizing metabolites at the expense of high utilization of carbon substrate.

Cereulide produced by Bacillus cereus increases the fitness of the producer organism in low-potassium environments.

Cereulide, produced by certain Bacillus cereus strains, is a lipophilic cyclic peptide of 1152 Da that binds K(+) ions with high specificity and affinity. It is toxic to humans, but its role for the producer organism is not known. We report here that cereulide operates for B. cereus to scavenge potassium when the environment is growth limiting for this ion. Cereulide-producing B. cereus showed higher maximal growth rates (µ(max)) than cereulide non-producing B. cereus in K(+)-deficient medium (K(+) concentration ~1 mM). The cereulide-producing strains grew faster in K(+)-deficient than in K(+)-rich medium with or without added cereulide. Cereulide non-producing B. cereus neither increased µ(max) in K(+)-deficient medium compared with K(+)-rich medium, nor benefited from added cereulide. Cereulide-producing strains outcompeted GFP-labelled Bacillus thuringiensis in potassium-deficient (K(+) concentration ~1 mM) but not in potassium-rich (K(+) concentration ~30 mM) medium. Exposure to 2 µM cereulide in potassium-free medium lacking an energy source caused, within seconds, a major efflux of cellular K(+) from B. cereus not producing cereulide as well as from Bacillus subtilis. Cereulide depleted the cereulide non-producing B. cereus and B. subtilis cells of a major part of their K(+) stores, but did not affect cereulide-producing B. cereus strains. Externally added 6-10 µM cereulide triggered the generation of biofilms and pellicles by B. cereus. The results indicate that both endogenous and externally accessible cereulide supports the fitness of cereulide-producing B. cereus in environments where the potassium concentration is low.

Pseudoxanthomonas bacteria that drive deposit formation of wood extractives can be flocculated by cationic polyelectrolytes.

Runnability problems caused by suspended bacteria in water using industries, have, in contrast to biofilms, received little attention. We describe here that Pseudoxanthomonas taiwanensis, a wide-spread and abundant bacterium in paper machine water circuits, aggregated dispersions of wood extractives ("pitch") and resin acid, under conditions prevailing in machine water circuits (10(9) cfu ml(-1), pH 8, 45°C). The aggregates were large enough (up to 50 µm) so that they could be expected to clog wires and felts and to reduce dewatering of the fiber web. The Pseudoxanthomonas bacteria were negatively charged over a pH range of 3.2-10. Cationic polyelectrolytes of the types used as retention aids or fixatives to flocculate "anionic trash" in paper machines were effective in flocculating the Pseudoxanthomonas bacteria. The polyelectrolyte most effective for this purpose was of high molecular weight (7-8 × 10(6) g mol(-1)) and low charge density (1 meq g(-1)), whereas polyelectrolytes that effectively zeroed the electrophoretic mobility (i.e., neutralized the negative charge) of the bacterium were less effective in flocculating the bacteria. Based on the results, we concluded that the polyelectrolytes functioning by bridging mechanism, rather than by neutralization of the negative charge, may be useful as tools for reducing harmful deposits resulting from interaction of bacteria with wood extractives in warm water industry.

Chemical and microbiological hazards associated with recycling of anaerobic digested residue intended for agricultural use.

In the present study, three full-scale biogas plants (BGP) were investigated for the concentration of heavy metals, organic pollutants, pesticides and the pathogenic bacteria Bacillus cereus and Escherichia coli in the anaerobically digested residues (ADR). The BGPs mainly utilize source-separated organic wastes and industrial food waste as energy sources and separate the ADR into an ADR-liquid and an ADR-solid fraction by centrifugation at the BGP. According to the Norwegian standard for organic fertilizers, the ADR were classified as quality 1 mainly because of high zinc (132-422 mg kg(-1) DM) and copper (23-93 mg kg(-1) DM) concentrations, but also because of high cadmium (0.21-0.60 mg kg(-1) DM) concentrations in the liquid-ADR. In the screening of organic pollutants, only DEHP (9.7-62.1 mg kg(-1)) and ∑ PAH 16 (0.2-1.98 mg kg(-1) DM) were detected in high concentrations according to international regulations. Of the 250 pesticides analyzed, 11 were detected, but only imazalil (<0.30-5.77 mg kg(-1) DM) and thiabendazol (<0.14-0.73 mg kg(-1) DM) were frequently detected in the ADR-fiber. Concentrations of imazalil and thiabendazol were highest during the winter months, due to a high consumption of citrus fruits in Norway in this period. Ten percent of the ADR-liquid samples contained cereulide-producing B. cereus, whereas no verotoxigenic E. coli was detected. The authors conclude that the risk of chemical and bacterial contamination of the food chain or the environment from agricultural use of ADR seems low.

Effects of polarization in the presence and absence of biocides on biofilms in a simulated paper machine water.

The antifouling potential of electric polarization combined and not combined with biocides was studied in nonsaline warm water with high organic content. Deinococcus geothermalis is a bacterium known for forming colored biofilms in paper machines and for its persistence against cleaning and chemical treatments. When D. geothermalis biofilms grown for 24 h in simulated paper machine water were exposed to cathodic or cathodically weighted pulsed polarization at least 60% (P < 0.05) of the biofilms were removed from stainless steel (AISI 316L). Biofilm removal by 25 ppm (effective substances 5-25 ppm) of oxidizing biocides (bromochloro-5,5-dimethylhydantoin, 2,2-dibromo-2-cyanoacetamide, peracetic acid) increased to 70% when combined with cathodically weighted pulsed polarization. Using a novel instrument that allows real-time detection of reactive oxygen species (ROS) we showed that the polarization program effective in antifouling generated ROS in a pulsed manner on the steel surface. We thus suggest that the observed added value of oxidative biocides combined with polarization depended on ROS. This suggestion was supported by the finding that a reductive biocide, methylene bisthiocyanate, counteracted the antifouling effect of polarization.

Adsorption of bacteria and polycations on model surfaces of cellulose, hemicellulose and wood extractives studied by QCM-D.

Quartz crystal microbalance with dissipation monitoring (QCM-D), atomic force microscopy (AFM) and field emission scanning electron microscopy (FESEM) were used as the tools to study the adsorption of bacteria onto surfaces of silica and polystyrene coated with materials related to papermaking. Cationic polyelectrolytes used as fixatives and retention aids in paper industry were found to promote irreversible adsorption of the ubiquitous white water bacterium, Pseudoxanthomonas taiwanensis, onto model surfaces of cellulose (pH 8). The high charged low molecular weight polyelectrolyte, poly(diallyldimethyl) ammonium chloride (pDADMAC) adsorbed to silica surface as a flat and rigid layer, whereas the low charged cationic polyacryl amide (C-PAM) of high molecular weight adsorbed as a thick and loose layer. AFM images showed that the polyelectrolytes accumulated as layers around each bacterial cell. In the presence of wood hemicellulose (O-acetyl-galactoglucomannan) the bacteria adsorbed massively, as large, tightly packed rafts (up to 0.05mm in size) onto the polystyrene crystal surface coated with wood extractives (pH 4.7). AFM and FESEM micrographs also showed large naked areas (with no bacteria) in between the bacterial rafts on the crystal surface. In this case, QCM-D only incompletely responded to the massiveness of the bacterial adsorption. The results indicate that cationic polymers can be used to increase the retention of bacteria from the process water onto the fibre web and that, depending on the balance between hemicelluloses and wood extractives and pH of the process waters, bacteria can be drawn from process waters onto surfaces.

Burkholderia fungorum DBT1: a promising bacterial strain for bioremediation of PAHs-contaminated soils.

An extensive taxonomic analysis of the bacterial strain Burkholderia sp. DBT1, previously isolated from an oil refinery wastewater drainage, is discussed here. This strain is capable of transforming dibenzothiophene through the 'destructive' oxidative pathway referred to as the Kodama pathway. Burkholderia DBT1 has also been proved to use fluorene, naphthalene and phenanthrene as carbon and energy sources, although growth on the first two compounds requires a preinduction step. This evidence suggests that the strain DBT1 exerts a versatile metabolism towards polycyclic aromatic hydrocarbons other than condensed thiophenes. Phylogenetic characterization using a polyphasic approach was carried out to clarify the actual taxonomic position of this strain, potentially exploitable in bioremediation. In particular, investigations were focused on the possible exclusion of Burkholderia sp. DBT1 from the Burkholderia cepacia complex. Analysis of the sequences of 16S, recA and gyrB genes along with the DNA-DNA hybridization procedure indicated that the strain DBT1 belongs to the species Burkholderia fungorum, suggesting the proposal of the taxonomic denomination B. fungorum DBT1.

Biofilm-forming bacteria with varying tolerance to peracetic acid from a paper machine.

Biofilms cause runnability problems in paper machines and are therefore controlled with biocides. Peracetic acid is usually effective in preventing bulky biofilms. This study investigated the microbiological status of a paper machine where low concentrations (≤ 15 ppm active ingredient) of peracetic acid had been used for several years. The paper machine contained a low amount of biofilms. Biofilm-forming bacteria from this environment were isolated and characterized by 16S rRNA gene sequencing, whole-cell fatty acid analysis, biochemical tests, and DNA fingerprinting. Seventy-five percent of the isolates were identified as members of the subclades Sphingomonas trueperi and S. aquatilis, and the others as species of the genera Burkholderia (B. cepacia complex), Methylobacterium, and Rhizobium. Although the isolation media were suitable for the common paper machine biofoulers Deinococcus, Meiothermus, and Pseudoxanthomonas, none of these were found, indicating that peracetic acid had prevented their growth. Spontaneous, irreversible loss of the ability to form biofilm was observed during subculturing of certain isolates of the subclade S. trueperi. The Sphingomonas isolates formed monoculture biofilms that tolerated peracetic acid at concentrations (10 ppm active ingredient) used for antifouling in paper machines. High pH and low conductivity of the process waters favored the peracetic acid tolerance of Sphingomonas sp. biofilms. This appears to be the first report on sphingomonads as biofilm formers in warm water using industries.

Deinobacterium chartae gen. nov., sp. nov., an extremely radiation-resistant, biofilm-forming bacterium isolated from a Finnish paper mill.

A rod-shaped, non-spore-forming, non-motile, aerobic, oxidase and catalase-positive and radiation-resistant bacterium (designated strain K4.1(T)) was isolated from biofilm collected from a Finnish paper mill. The bacterium grew as pale pink colonies on oligotrophic medium at 12 to 50 °C (optimum 37 to 45 °C) and at pH 6 to 10.3. The DNA G+C content of the strain was 66.8 l%. According to 16S rRNA gene sequence analysis, strain K4.1(T) was distantly related to the genus Deinococcus, sharing highest similarity with Deinococcus pimensis (90.0  %). In the phylogenetic tree, strain K4.1(T) formed a separate branch in the vicinity of the genus Deinococcus. The peptidoglycan type was A3ß with L-Orn-Gly-Gly and the quinone system was determined to be MK-8. The polar lipid profile of strain K4.1(T) differed markedly from that of the genus Deinococcus. The predominant lipid of strain K4.1(T) was an unknown aminophospholipid and it did not contain the unknown phosphoglycolipid predominant in the polar lipid profiles of deinococci analysed to date. Two of the predominant fatty acids of the strain, 15 : 0 anteiso and 17 : 0 anteiso, were lacking or present in small amounts in species of the genus Deinococcus. Phylogenetic distinctness and significant differences in the polar lipid and fatty acid profiles suggest classification of strain K4.1(T) as a novel genus and species in the family Deinococcaceae, for which we propose the name Deinobacterium chartae gen. nov., sp. nov. The type strain is K4.1(T) (=DSM 21458(T) =HAMBI 2721(T)).