The toxic mode of action of cyclic lipodepsipeptide fusaricidins, produced by Paenibacillus polymyxa, toward mammalian cells.

AIMS: Toxigenic strains of Paenibacillus polymyxa were isolated from buildings connected with the symptoms of ill health. Our aim was to identify the toxic compounds of Paenibacillus polymyxa and to describe their toxic actions. METHODS AND RESULTS: The toxins of Paenibacillus polymyxa were purified and analysed by HPLC and mass spectrometry. Toxic fusaricidins A and B, and LI-F05a with mass ions at m/z 883·7, 897·6 and 897·6, respectively, were found. The cytotoxicity of purified fusaricidins A and B was measured using boar sperm, porcine tubular kidney epithelial cells and murine fibroblasts. The ion channel forming properties of fusaricidins were studied using the black lipid membrane (BLM) technique. Fusaricidins A and B depolarized the mitochondria of boar sperm, porcine tubular kidney epithelial cells and murine fibroblasts at concentrations of 0·5-1 µg ml-1 and caused nuclear fragmentation and induced apoptosis at concentrations of 2·5-5 µg ml-1 . Furthermore, fusaricidins A and B induced K+ permeating single channels. CONCLUSIONS: It was concluded that fusaricidins were toxic to mitochondria and induced apoptosis in mammalian cells. It was proposed that the observed toxicity of fusaricidins is due their ion channel forming properties. SIGNIFICANCE AND IMPACT OF THE STUDY: This paper revealed, for the first time, the mode of action of Paenibacillus polymyxa fusaricidins toxins towards mammalian cells. Fusaricidins, due to their potassium ionophoricity and mitochondria depolarizing impacts, may have contributed to the health damage observed at sites where the producer strains were isolated at high density.

Proposed minimal standards for describing new taxa of aerobic, endospore-forming bacteria.

Minimal standards for describing new taxa within the aerobic endospore-forming bacteria are proposed, following Recommendation 30b of the Bacteriological Code (1990 Revision). These minimal standards are recommended as guidelines to assist authors in the preparation of descriptions for novel taxa. They encourage broad polyphasic characterization and the construction of descriptions that are practically useful in routine diagnostic laboratories. The proposals have been endorsed by the Subcommittee on the Taxonomy of the Genus Bacillus and Related Organisms of the International Committee on Systematics of Prokaryotes.

Acrebol, a novel toxic peptaibol produced by an Acremonium exuviarum indoor isolate.

AIMS: To identify a toxin and its producer isolated from woody material in a building where the occupants experienced serious ill health symptoms. METHODS AND RESULTS: Hyphal extracts of an indoor fungus, identified as the cycloheximide-tolerant species Acremonium exuviarum, inhibited motility of boar spermatozoa (EC(50) 5 +/- 2 microg of crude solids ml(-1)) and caused cytolysis of murine neuroblastoma cells (MNA) and feline fetal lung cells (FL). The responsible substances were purified and identified as two structurally similar, heat-stable, novel, toxic peptaibols, 1726 Da and 1740 Da, respectively, with amino acid sequences of Acetyl-Phe-Iva/Val-Gln-Aib-Ile-Thr-Leu-Aib-Pro-Aib-Gln-Pro-Aib-(X-X-X)-SerOH and Acetyl-Phe-Iva/Val-Gln-Aib-Ile-Thr-Leu-Val-Pro-Aib-Gln-Pro-Aib-(X-X-X)-SerOH. Purified acrebol inhibited motility of boar sperm, depleted ATP half-content in 1 day (EC(50) of 0.1 microg ml(-1), 60 nmol l(-1)) depolarised the mitochondria after 2 days, but did not affect the cellular content in NADH. This indicates mitochondrial toxicity. Plate-grown biomass of A. exuviarum BMB4 contained 0.1-1% (w/w) of acrebol, depending on the culture medium. CONCLUSIONS: Acrebol paralysed the energy generation of mammalian cells suggesting that mitochondria were its target of action. SIGNIFICANCE AND IMPACT OF THE STUDY: Acremonium exuviarum, as an indoor fungus, is potentially hazardous to health because of the toxic peptaibols that it produces.

Proliferation of mycobacteria in a piggery environment revealed by mycobacterium-specific real-time quantitative PCR and 16S rRNA sandwich hybridization.

Pig mycobacteriosis is the most common animal mycobacterial disease in Finland with a long-term average prevalence of 0.34% and temporary peaks as high as 0.85%. In the current study Mycobacterium-specific real-time qPCR and 16S rRNA sandwich hybridization were utilized for culture-independent detection and measurement of potentially infectious mycobacteria in selected piggeries. Participating herds (n=5) were selected according to prevalence of tuberculous lesions (>4%) in slaughtered carcasses. When DNA extracted from piggery bedding materials was analyzed by Mycobacterium-targeted qPCR using the SYBR green I dye for detection of amplification products, 10(5) to 10(7) cell equivalents of mycobacterial DNA were detected in unused bedding materials and 10(8) to 10(10)g(-1) dry weight in used bedding materials. When Mycobacterium-specific hybridization probes were used for detection of amplification products, 10(5) to 10(7) cell equivalents of mycobacterial DNA g(-1) dry weight were detected in unused bedding materials in four out of the five piggeries studied and up to 10(8) cell equivalents in used bedding material. The results were confirmed by the Mycobacterium-specific 16S rRNA sandwich hybridization assay. The present results show, that mycobacteria occur in organic materials commonly used on pig farms, and may proliferate in bedding materials during use. We also show that DNA- and RNA-based methods may be utilized for detection of environmental reservoirs of mycobacteria causing porcine and human infection.

Characterization of adhesion threads of Deinococcus geothermalis as type IV pili.

Deinococcus geothermalis E50051 forms tenuous biofilms on paper machine surfaces. Field emission electron microscopy analysis revealed peritrichous appendages which mediated cell-to-surface and cell-to-cell interactions but were absent in planktonically grown cells. The major protein component of the extracellular extract of D. geothermalis had an N-terminal sequence similar to the fimbrial protein pilin annotated in the D. geothermalis DSM 11300 draft sequence. It also showed similarity to the type IV pilin sequence of D. radiodurans and several gram-negative pathogenic bacteria. Other proteins in the extract had N-terminal sequences identical to D. geothermalis proteins with conservative motifs for serine proteases, metallophosphoesterases, and proteins whose function is unknown. Periodic acid-Schiff staining for carbohydrates indicated that these extracellular proteins may be glycosylated. A further confirmation for the presence of glycoconjugates on the cell surface was obtained by confocal laser scanning imaging of living D. geothermalis cells stained with Amaranthus caudatus lectin, which specifically binds to galactose residues. The results indicate that the thread-like appendages of D. geothermalis E50051 are glycosylated type IV pili, bacterial attachment organelles which have thus far not been described for the genus Deinococcus.

Atmospheric oxygen and other conditions affecting the production of cereulide by Bacillus cereus in food.

Factors influencing the production of cereulide, the emetic toxin of Bacillus cereus in food and laboratory media were investigated, using liquid chromatography-ion trap mass spectrometry and sperm motility inhibition bioassay for detection and quantitation. Oxygen was essential for production of the emetic toxin by B. cereus. When beans, rice or tryptic soy broth were inoculated with cereulide producing strains B203, B116 (recent food isolates) or the strain F-4810/72, high amounts (2 to 7 microg ml(-1) or g(-1) wet wt) of cereulide accumulated during 4-day storage at room temperature. In parallel cultures and foods, stored under nitrogen atmosphere (> 99.5% N2), less than 0.05 microg of cereulide ml(-1) or g(-1) wet wt accumulated. The outcome of the bioassay matched that of the chemical assay, with no indication of interference by substances in the rice or beans. Boiling for 20 to 30 min did not inactivate cereulide or cereulide producing strains in rice or the beans. Adding l-leucine and l-valine (0.3 g l(-1)) stimulated cereulide production 10- to 20-fold in R2A and in rice water agar. When the B. cereus strains were grown on agar media under permissive conditions (air, room temperature), cereulide was produced overnight with little or no increase when the incubation was extended to 4 days. In broth culture, the production of cereulide started later than 16-24 h. Anoxic storage prevented cereulide production also when the amino acids had been supplied. Packaging with modified atmosphere low in oxygen may thus be used to reduce the risk of cereulide formation during storage of food.

In vitro assay for human toxicity of cereulide, the emetic mitochondrial toxin produced by food poisoning Bacillus cereus.

The in vitro boar spermatozoon test was compared with the LC ion trap MS analysis for measuring the cereulide content of a pasta dish, implemented in serious emetic food poisoning caused by Bacillus cereus. Both assays showed that the poisonous food contained approximately 1.6 microg of cereulide g(-1) implying the toxic dose in human as < or =8 microg kg(-1) body weight. The threshold concentration of cereulide provoking visible mitochondrial damage in boar sperm exposed in vitro was 2 ng of cereulide ml(-1) of extended boar sperm. The same threshold value was found for cereulide extracted from the food and from the cultured bacteria. This shows that other constituents of the food did not enhance or mask the effects of cereulide. Exposure of four human cell lines (HeLa, Caco-2, Calu-3 and Paju) to cereulide showed that the threshold concentration for the loss of mitochondrial membrane potential in human cells was similar to that observed in boar sperm. Human cells and boar sperm were equally sensitive to cereulide. The results show that boar spermatozoan assay is useful for detecting cereulide concentrations toxic to humans. Spermatozoa in commercially available extended fresh boar and cryopreserved bull semen were compared, boar sperms were 100 times more sensitive to cereulide than bull sperms.

A new method for in vitro detection of microbially produced mitochondrial toxins.

Sperm motility inhibition assay, earlier shown valuable for the detection of food poisoning non-protein toxins of Bacillus species was developed into an assay useful for specific detection of mitochondria damaging toxins. This was done by assessing the dissipation of the mitochondrial inner membrane transmembrane potential, Deltapsim under conditions where the plasma membrane permeability barrier remained intact. The Deltapsim was estimated as the intensity of orange JC-1 fluorescence in the mitochondrial sheath of the exposed spermatozoa. The plasma membrane integrity of the same cells was assessed by observing the exclusion of propidium iodide from the cytoplasm. Three types of mitochondrial toxic responses to microbially made bioactive substances were recognised. Mitochondrial toxicity by gramicidin (A, B, C, D), nigericin, salinomycin, narasin, monensin, calcimycin and antimycin A was characterised by gradual fading of the JC-1 fluorescence in the mitochondria. Dissipation of the Deltapsim by cereulide, valinomycin and enniatin (A, A1, B, B1) was visible as spotwise quenching of the mitochondrial JC-1 fluorescence. In addition these substances caused hyperpolarisation of the plasma membrane. Oligomycin (A, B, C), ionomycin and staurosporine inhibited the spermatozoan motility, but Deltapsim was fully preserved. Surfactin and lichenysin A caused mitochondrial damage at concentrations where the plasma membrane was also damaged.

Colored moderately thermophilic bacteria in paper-machine biofilms.

Biofilms cause several problems in papermaking. This report describes a microbiological survey of colored biofilms in six paper and board machines, including two case studies of outbreaks of colored slimes in which the causative bacteria were found. A total of 95 pink-, red-, orange- or yellow-pigmented strains were isolated. Nearly all (99%) of the strains grew at 52 degrees C, 72% grew at 56 degrees C, but only 30% grew at 28 degrees C, indicating that most of the strains were moderately thermophilic. Biofilm formation potential and biocide susceptibility of the strains were analyzed with a microtiter plate assay. In the presence of 5 ppm of methylene bisthiocyanate or 2,2-dibromo-3-nitrilopropionamide in paper-machine water, 55 strains formed biofims. Moreover, 39 strains increased biofilm production by 5-753% in the presence of biocide, suggesting that biocide concentrations inhibitory to planktonic but not to surface-attached cells may actually promote biofouling. The cells may have inactivated a portion of the biocides, as the cell density in this assay was high, corresponding to the highest cell densities occurring in the circulating waters. Four groups of colored bacteria that were isolated from several mills were identified. Pink-pigmented Deinococcus geothermalis and red-pigmented Meiothermus silvanus occurred as common primary biofilm-formers in paper machines. This report is the first description of the involvement of Meiothermus species in red-slime formation in the paper industry. The third group of bacteria (putative new species related to Roseomonas) contained strains that were not biofilm formers, but which were commonly found in slimes of neutral or alkaline machines. The fourth group contained red-pigmented biofilm-forming strains representing a novel genus of alpha- Proteobacteria related to Rhodobacter. Many colored paper-machine bacteria are species previously known from microbial mats of hot springs. Some characteristics of the bacterial groups are described here in order to facilitate their recognition in future cases of colored-slime outbreaks in the paper industry.

Impaired semen quality of AI bulls fed with moldy hay: a case report.

The daily quality control of semen at a Finnish artificial insemination (AI) bull station is based on subjective motility and sperm morphology of young bulls entering the semen collection program. Semen quality dropped suddenly in autumn 1998. During 5 consecutive months, the number of rejected ejaculates and discarded frozen semen batches due to poor motility increased, and the number of all forms of abnormal spermatozoa increased. However, for the accepted ejaculates, a 60 day nonretum rate was normal. The summer of 1998 in Finland was rainy, and the hay used in the AI station was visibly moldy. Immunoassay and gas chromatography-mass spectrometry (GC-MS) detected Fusarium mycotoxins HT-2 and T-2, but no zearalenone in the hay. Occurrence of mycotoxins such as T-2 and HT-2 in the moldy hay coincided with, and may have been responsible for the impaired semen quality in AI bulls. This case report will draw the attention to the possible hazards when feeding moldy hay.

Inhibition of human natural killer cell activity by cereulide, an emetic toxin from Bacillus cereus.

The lipophilic toxin, cereulide, emitted by emetic food poisoning causing strains of Bacillus cereus, is a powerful mitochondria toxin. It is highly lipophilic and rapidly absorbed from the gut into the bloodstream. We tested how this toxin influences natural killer (NK) cells, which are important effectors in defence against infections and malignancy. Cereulide inhibited cytotoxicity and cytokine production of natural killer cells, caused swelling of natural killer cell mitochondria, and eventually induced natural killer cell apoptosis. The suppressive effect on cytotoxicity was fast and toxic concentration low, 20-30 microg/l. As the emesis causing concentration of cereulide is around 10 microg/kg of total body mass, our results suggest that emesis causing or even lower doses of cereulide may also have a systemic natural killer cell suppressive effect.

Structure and on-site formation of biofilms in paper machine water flow.

Paper machine biofilms formed in situ on stainless steel surfaces were studied. A robust flow cell was fitted to side stream (1.8 m s(-1)) of the spray water circuit of a paper machine. This on-site tool allowed for assessing the efficacy of antifoulants and the adequacy of steel polishing under mill conditions. A rapid fluorescence-based assay was developed to quantify the biomass of shallow biofilms on machine steel. The fluorescence matched the ATP content measured for the same biofilms. Electrolytic polishing reduced the tendency of biofouling of 500 grit surface steel. Biofilm grew under machine conditions as clusters on the steels, showing uniformly coccoid, filaments or short rods; only one cell type in each cluster. The biofilm clusters excluded latex beads of 0.02 microm with hydrophilic or with hydrophobic surfaces from penetrating more than three to four layers of cells. Under the high hydraulic flow at the machine (1.8 m s(-1)), the biofilm grew in 7 days 6-10 microm thick. The high flow rate guided the shape of the biofilm clusters emerging after the primary attachment of cells. Adhered individual bacteria were the platform on steel to which solids such as paper machine fines then accumulated.

Thermomonas haemolytica gen. nov., sp. nov., a gamma-proteobacterium from kaolin slurry.

Four aerobic, gram-negative bacterial strains isolated from kaolin slurry used in the production of paper were subjected to a polyphasic analysis and characterization to determine their taxonomic position. Analysis of the 16S rDNA sequences of the four strains revealed that they represent a new lineage within the gamma-Proteobacteria, related to the genera Xanthomonas, Pseudoxanthomonas, Stenotrophomonas, Luteimonas, Xylella and Rhodanobacter. Analysis of the quinone system, the polyamines, the fatty acids and the polar lipids revealed a combination of characteristics that is unique and not described for the phylogenetic relatives. The four strains contain a ubiquinone Q-8, spermidine as the major polyamine, diphosphatidylglycerol, phosphatidylglycerol and phosphatidylethanolamine as the predominant polar lipids, and a fatty acid profile with predominantly iso-branched fatty acids. The G+C content of the genomic DNA was determined to be within the narrow range 67.1-68.7 mol%. Determination of DNA relatedness, as well as riboprint band patterns and amplified fragment length polymorphism profiles, clearly demonstrated that the four strains are members of a single species. Antibiotic-susceptibility patterns were identical for the four strains. Although showing a high degree of similarites in physiological and biochemical patterns, each of the four strains could be distinguished from the others on the basis of a few biochemical characteristics. On the basis of the estimates of phylogenetic relationships derived from the 16S rDNA sequence analyses, the observed chemotaxonomic characteristics and other phenotypic traits, a new genus, Thermomonas gen. nov., and species, Thermomonas haemolytica sp. nov., are proposed for the strains A50-7-3T (= DSM 13605T = LMG 19653T), B 50-7-1 (= DSM 13598 = LMG 19655), D50-7-1 (= DSM 13610 = LMG 19656) and B50-8-1 (= DSM 13599 = LMG 19654), with strain A50-7-3T as the type strain.

Firm but slippery attachment of Deinococcus geothermalis.

Bacterial biofilms impair the operation of many industrial processes. Deinococcus geothermalis is efficient primary biofilm former in paper machine water, functioning as an adhesion platform for secondary biofilm bacteria. It produces thick biofilms on various abiotic surfaces, but the mechanism of attachment is not known. High-resolution field-emission scanning electron microscopy and atomic force microscopy (AFM) showed peritrichous adhesion threads mediating the attachment of D. geothermalis E50051 to stainless steel and glass surfaces and cell-to-cell attachment, irrespective of the growth medium. Extensive slime matrix was absent from the D. geothermalis E50051 biofilms. AFM of the attached cells revealed regions on the cell surface with different topography, viscoelasticity, and adhesiveness, possibly representing different surface layers that were patchily exposed. We used oscillating probe techniques to keep the tip-biofilm interactions as small as possible. In spite of this, AFM imaging of living D. geothermalis E50051 biofilms in water resulted in repositioning but not in detachment of the surface-attached cells. The irreversibly attached cells did not detach when pushed with a glass capillary but escaped the mechanical force by sliding along the surface. Air drying eliminated the flexibility of attachment, but it resumed after reimmersion in water. Biofilms were evaluated for their strength of attachment. D. geothermalis E50051 persisted 1 h of washing with 0.2% NaOH or 0.5% sodium dodecyl sulfate, in contrast to biofilms of Burkholderia cepacia F28L1 or the well-characterized biofilm former Staphylococcus epidermidis O-47. Deinococcus radiodurans strain DSM 20539(T) also formed tenacious biofilms. This paper shows that D. geothermalis has firm but laterally slippery attachment not reported before for a nonmotile species.

Isolation of toxigenic Nocardiopsis strains from indoor environments and description of two new Nocardiopsis Species, N. exhalans sp. nov. and N. umidischolae sp. nov.

Nocardiopsis strains were isolated from water-damaged indoor environments. Two strains (N. alba subsp. alba 704a and a strain representing a novel species, ES10.1) as well as strains of N. prasina, N. lucentensis, and N. tropica produced methanol-soluble toxins that paralyzed the motility of boar spermatozoa at <30 microg of crude extract (dry weight) x ml(-1). N. prasina, N. lucentensis, N. tropica, and strain ES10.1 caused cessation of motility by dissipating the mitochondrial membrane potential, Deltapsi, of the boar spermatozoa. Indoor strain 704a produced a substance that destroyed cell membrane barrier function and depleted the sperm cells of ATP. Indoor strain 64/93 was antagonistic towards Corynebacterium renale. Two indoor Nocardiopsis strains were xerotolerant, and all five utilized a wide range of substrates. This combined with the production of toxic substances suggests good survival and potential hazard to human health in water-damaged indoor environments. Two new species, Nocardiopsis exhalans sp. nov. (ES10.1T) and Nocardiopsis umidischolae sp. nov. (66/93T), are proposed based on morphology, chemotaxonomic and physiological characters, phylogenetic analysis, and DNA-DNA reassociations.

Sugar composition of biofilms produced by paper mill bacteria.

Biofilms of paper mill bacteria were cultivated in paper mill white water-simulating conditions on glass slides or stainless steel coupons in a laboratory culture system. The sugar content and composition of the biofilms were analysed and compared with the sugar composition of paper mill slimes. Acid methanolysis followed by gas chromatography revealed that Burkholderia was the major biofilm producer in pure culture, producing up to 50 microg of biofilm sugar cm(-2) in 5 days in rich medium and 10 microg in paper mill simulating medium. A mixture of simulated paper mill water with a culture medium yielded more biofilm (100 microg cm(-2)) than either of the media alone, so the biofilm accumulation was not proportional to the available substrate. More biofilm accumulated on stainless steel coupons than on glass slides, and the steel-coupon biofilms contained slightly more uronic acids. The biofilm sugars contained mainly galactose, glucose, mannose, and rhamnose. In paper mill medium, the Burkholderia biofilm contained more galactose and glucose, and less rhamnose, than in rich laboratory medium. The sugar composition of paper mill slimes was quite similar to those of steel-cultured Burkholderia cepacia biofilms. This suggests that Burkholderia cepacia is responsible for much of the slime in the paper mill.

Toxic-metabolite-producing bacteria and fungus in an indoor environment.

Toxic-metabolite-emitting microbes were isolated from the indoor environment of a building where the occupant was suffering serious building-related ill-health symptoms. Toxic substances soluble in methanol and inhibitory to spermatozoa at <10 microg (dry weight) ml(-1) were found from six bacterial isolates and one fungus. The substances from isolates of Bacillus simplex and from isolates belonging to the actinobacterial genera Streptomyces and Nocardiopsis were mitochondriotoxic. These substances dissipated the mitochondrial membrane potential (Deltapsi) of boar spermatozoa. The substances from the Streptomyces isolates also swelled the mitochondria. The substances from isolates of Trichoderma harzianum Rifai and Bacillus pumilus damaged the cell membrane barrier function of sperm cells.

Inhibition of bacilli in industrial starches by nisin.

The properties of Bacillus coagulans and of other bacilli that contaminate paper and paperboard manufacturing processes were investigated under simulated industrial conditions. Nisin (0.05 to 0.125 microg ml(-1) blocked growth of indigenous bacilli that contaminate sizing starches. B. coagulans starch isolates, B. licheniformis, B. amyloliquefaciens, and B. stearothermophilus grew at > or = 50 degrees C in industrial starch and produced alpha-glucosidase and cyclodextrins. The industrial isolates and reference strains of B. amyloliquefaciens, B. cereus, B. coagulans, B. flexus, B. licheniformis, B. pumilus, B. sporothermodurans, B. stearothermophilus and Alicyclobacillus acidoterrestris were inhibited by < or = 0.125 microg of nisin on agar. B. coagulans and B. stearothermophilus were similarly inhibited by < or = 0.025 microg of nisin ml(-1) and by 3 microg of the biocide DBNPA ml(-1) in industrial starch. B. licheniformis and B. amyloliquefaciens strains were less sensitive. About 40% of nisin added to starch was retained after cooking. Fifty percent of the nisin remained active after 11 h of storage at 60 degrees C. The results show that nisin has potential as a preservative for modified industrial starches.

In situ polychlorophenol bioremediation potential of the indigenous bacterial community of boreal groundwater.

The composition and chlorophenol-degrading potential of groundwater bacterial community in a permanently cold, oxygen-deficient chlorophenol contaminated aquifer at Kärkölä, Finland was studied with the aim of evaluating in situ bioremediation potential. The groundwater contained from 10(4) to 10(7) microscopically counted cells/ml and up to 10(5) CFU/ml heterotrophic bacteria cultivable at 8 and 20 degrees C. Of the 102 pure cultures, of which 86% Gram-negative, from the plume area (10,000 microg of chlorophenols/l), 57% degraded 2, 3, 4, 6-tetrachlorophenol (TeCP), the main component of the wood preservative which was the source of contamination: 17% also degraded pentachlorophenol (PCP). The degraders were scattered among 16 different clusters of Gram-negatives mainly proteobacteria and members of Cytophaga/Flexibacter/Bacteroides phylum judged by the composition of whole-cell fatty acids. Only one Gram-positive degrading cluster was found containing seven actinobacteria closest to Nocardioides. Of the 88 pure cultures isolated from outside the plume (< 10 microg of chlorophenols/l) 67% were Gram-negative. Seven percent of the isolates degraded 2, 3. 4, 6-TeCP and/or PCP. Five of the Gram-positive isolates from outside the plume were Mycobacterium/Rhodococcus-related actinobacteria and O-methylated 2, 3, 4, 6-TeCP and PCP. The results show that chlorophenol degrading bacterial flora had been enriched as a result of contamination of the aquifer. This suggests significant in situ bioremediation potential of the site.

Degradation of paper mill water components in laboratory tests with pure cultures of bacteria.

The degradation of dissolved and colloidal substances from thermomechanical pulp (TMP) by bacteria isolated from a paper mill was studied in a laboratory slide culture system. Burkholderia cepacia strains hydrolysed triglycerides to free fatty acids, and the liberated unsaturated fatty acids were then degraded to some extent. Saturated fatty acids were not notably degraded. However, the branched anteiso-heptadecanoic fatty acid was degraded almost like the unsaturated fatty acids. About 30% of the steryl esters were degraded during 11 days, increasing the concentrations of free sterols. Approximately 25% of the dehydroabietic, and 45% of the abietic and isopimaric resin acids were degraded during 11 days. The degree of unsaturation seemed to be of greater importance for the degradation of fatty acids than the molar mass. No degradation of dissolved hemicelluloses could be observed with any of the nine bacterial strains studied. Burkholderia cepacia strains and one Bacillus coagulans strain degraded monomeric fructose and glucose in winter TMP water, but in summer TMP water, with much lower sugar concentrations, also other Bacillus strains degraded monomeric sugars.