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Staphylococcus aureus is the most common cause of surgical site infections and its treatment is challenging due to the emergence of multi-drug resistant strains such as methicillin-resistant S. aureus (MRSA). Natural berry-derived compounds have shown antimicrobial potential, e.g., ellagitannins such as sanguiin H-6 and lambertianin C, the main phenolic compounds in Rubus seeds, have shown antimicrobial activity. The aim of this study was to evaluate the effect of sanguiin H-6 and lambertianin C fractionated from cloudberry seeds, on the MRSA growth, and as treatment of a MRSA biofilm development in different growth media in vitro and in vivo by using a murine wound infection model where sanguiin H-6 and lambertianin C were used to prevent the MRSA infection. Sanguiin H-6 and lambertianin C inhibited the in vitro biofilm development and growth of MRSA. Furthermore, sanguiin H-6 showed significant anti-MRSA effect in the in vivo wound model. Our study shows the possible use of sanguiin H-6 as a preventive measure in surgical sites to avoid postoperative infections, whilst lambertianin C showed no anti-MRSA activity.

In this study, tall oil fatty acid ester of softwood kraft lignin (TOFA-L) was used to prepare TOFA lignin nanoparticles (TLNP) in water. The average diameters for two prepared TLNPs in 0.1 mg/ml concentration were 140 nm and 160 nm. TLNPs were attached covalently onto modified and unmodified cellulose fibres to form an antimicrobial composite material. The modified cellulose fibres contained reactive allylic double bonds with a degree of substitution of 0.05. The antimicrobial properties of both TLNPs and TLNP coated fibres (TLNP-C) were studied against Escherichia coli, Staphylococcus aureus and Pseudomonas aeruginosa using silver nanospheres (average size 10 nm) and Lignoboost lignin particles with 300 and 400 nm sizes as references. Antimicrobial activity of the samples was stronger against Gram-positive S. aureus cells than against Gram-negative target microbes E. coli and P. aeruginosa.

This study aimed to establish the differences between enzymatically extracted hydrolysates and pH shifted protein isolates from whole Baltic herring and roach in terms of polypeptide patterns, functionality, sensory properties, microbial quality, yield, and composition. Alkaline extraction resulted in the highest yields, whereas the hydrolysates showed the highest protein contents. The hydrolysates showed higher protein solubility (86.0%-88.5%) than the protein isolates (5.1-14.5%) as well as the higher foam capacity for Baltic herring. However, for roach, alkaline extracted protein isolates exhibited the highest foam capacity. All hydrolysates showed poor foam stability (0-13%) while the protein isolates showed notably higher stability (30-55%). The hydrolysates showed relatively low bitterness, whereas alkaline extracted roach proteins were perceived as bitter. This study demonstrated that it was possible to produce protein isolates and hydrolysates from whole fish with good microbial quality. However, both processes need to be optimised according to the food application and fish species.

In order to assist developing a natural, safe food-preservative, aqueous ethanolic extracts of leaves and berries of eight Finnish berry plants were fractionated with Sephadex LH-20 column chromatography. For each fraction, phenolic compounds were analyzed with NMR, UPLC-DAD-ESI-MS and HPLC-DAD. The antioxidant activities of the fractions were investigated using oxygen radical absorbance capacity (ORAC) assay, and the antibacterial activities were evaluated against foodborne pathogens Staphylococcus aureus and Escherichia coli. Antioxidant activities of the fractions correlated highly with both the total concentration and structural feature of phenolic compounds, including both flavonoids and non-flavonoid phenolics. ORAC value correlated strongly with the concentration of (+)-catechin, (−)-epicatecin, quercetin glycosides, and anthocyanins. Increase in size and number of sugar moieties may reduce the antioxidative activities of quercetin glycosides. Type of sugar moieties may have a significant role in influencing peroxyl-radicals scavenging ability of quercetin glycosides with monosaccharides as a single sugar moieties. Most of the fractions inhibited the target microbes. S. aureus strains expressed a higher sensitivity to phenolic compounds than E. coli strains.

Traditionally grass has been considered a suitable feed only for ruminants. Due to global challenges related to growing human population, sustainable feed materials for pigs are actively searched and grasses as a high yielding biomass have been seen as an interesting addition to pig feeds. The digestible nutrients in grasses are embedded in a fibre matrix not digestible to pigs, but suitable processing could alleviate this. The aim of the present study was to evaluate whether silage juice could provide a new sustainable diet ingredient for fattening pigs. Grass silage was fractionated with a twin-screw press to liquid and solid fractions. The silage juice was included into liquid feed of 112 fattening pigs from 62 kg live weight until slaughter (124 kg live weight) and compared to 96 pigs on a conventional control diet. Feed allowance was ad libitum until it reached 33.6 MJ net energy day−1 per pig which was kept as the maximum feed allowance until slaughter. The average daily gain of all the pigs was over 1000 g day−1 in the experimental period but no difference between silage juice fed and control group was found. The feed conversion ratio was, as expected, higher in barrows than gilts, but no difference was found between the two treatment groups. Variation in silage juice dry matter content during the experimental period may have led to a lower dry matter intake of silage juice fed pigs. Gilts had statistically significantly higher meat percentage than barrows in both treatments, but no difference was found between the treatment groups. Meat quality (pH, colour, drip loss and sensory analyses) results were similar in both treatment groups. For gut microbiota, no statistically significant differences were found in any of the bacterial families studied (Actinobacteriota, Bacteroidota, Firmicutes, Deferrivacterota, Spirochaetota, Minor Phyla). In conclusion, silage juice is a potential feed ingredient in liquid feed for pigs. Microbiological data indicates that silage juice does not cause gastrointestinal microbiota imbalance in pigs.

Phenolic compounds were extracted with food grade solvent of acidified aqueous ethanol from leaves, berries, berry press cakes, and branches of Finnish berry plants and analyzed with HPLC-DAD, UPLC-DAD-ESI-MS and NMR. In addition, press cakes from two berry species and branches from one species were also extracted and analyzed with the same methods. The antioxidant activities of the extracts were evaluated using Folin-Ciocalteau, oxygen radical absorbance capacity (ORAC), DPPH free radical scavenging, and total radical trapping antioxidant parameter (TRAP) assays. The antibacterial activities were investigated against various Gram-negative and Gram-positive foodborne pathogens. The leaf extracts showed higher antioxidative activities (3–20 fold in ORAC assay, 10–20 fold in TRAP) than the berry extracts, in association with the higher contents of phenolic compounds in the leaf extracts; Strongest anti-bacterial effects was observed in the leaf extracts of lingonberry (Vaccinium vitis-idaea), sea buckthorn (Hippophaë rhamnoides ssp. rhamnoides) and saskatoon (Amelanchier alnifolia) on Staphylococcus aureus, Listeria monocytogenes, and Bacillus cereus. However, the antibacterial efficacy varied with bacterial species and strains. The Folin-Ciocalteu, ORAC, and TRAP values was strongly correlated with the total content of flavonoids with less association shown with the content of total phenolics and flavonol glycosides. The results suggest a major contribution of pranthocyanidins and flavan‐3-ols to the antioxidative activities of the extracts. The growth inhibition on Staphylococcus aureus and Bacillus cereus was clearly associated with the content of total phenolics and ellagitannins.

The food industry, including the meat industry, is currently looking for natural preservatives to prevent the growth of harmful microbes in foods. The potential of plant-derived antimicrobial extracts to increase the shelf life and to delay the microbiological spoilage of marinated broiler chicken cuts in modified atmosphere packages during cold storage was investigated in this study. We evaluated the impact of aqueous ethanolic extracts of Finnish sea buckthorn berries and lingonberries and supercritical CO2-extracted herbal extracts from an antimicrobial blend and oregano leaves on the shelf life of broiler meat. The commercial antimicrobial blend extract and the oregano extract inhibited the growth of lactic acid bacteria (LAB) and Brochothrix thermosphacta in the marinated samples. The antimicrobial blend extract also reduced the growth of psychrotrophic aerobic bacteria, whereas the sea buckthorn and lingonberry extracts did not. Only minor antimicrobial activity against Enterobacteriaceae by all the extracts was observed. Plate count analysis, denaturing gradient gel electrophoresis, and quantitative real-time PCR indicated that LAB, which are the major spoilage group in marinated modified atmosphere–packaged poultry products, were not significantly affected by the berry extracts studied. During this shelf-life study, LAB isolates of Lactobacillus and Leuconostoc were identified in the marinated samples. Antimicrobial blends and oregano leaf extracts can act as antimicrobial agents in marinade blends, although tailoring of the dose is needed because of their strong taste. Further studies for exploiting synergistic effects of plant extracts could contribute to the development of potential and more effective antimicrobial blends. Studies are needed in meat matrices and in product applications to demonstrate the efficacy of these compounds.

Antimicrobial plant extracts used in food packaging provide a healthy packaging alternatives. They contain aromatic and phenolic compounds that are responsible for their antibacterial properties. In this study, we report the antibacterial effects of extracts obtained from sea buckthorn (Hippophaë rhamnoides L.) leaves, and inner bark of pine trees (Pinus silvestris) that were applied as coatings on paper suitable for packaging application. Extracts from sea buckthorn leaves exhibited antibacterial effect both as a solvent extract and as a coating on paper against Pseudomonas aeruginosa as test bacteria. However, coatings of pine bark extract did not exhibit antibacterial effect as coatings even though the solvent extracts exhibited antibacterial effects. Staphylococcus aureus demonstrated resistance towards both plant extracts after they had been applied as coatings on paper for packaging.

Cosmetics are moving towards pharma and cosmeceuticals are entering the market. Modern biotechnology offers many advantages to develop new type of ingredients for industrial applications in a sustainable way. Due to the arctic growth conditions Nordic wild berries are rich sources of natural biomolecules with multifunctional effects to the skin. We have shown that phenolic compounds in berries very efficiently inhibit the growth of many human pathogens including skin pathogens, such as Staphylococcus aureus, S. epidermis and Pseudomonas aeruginosa without effecting the growth of beneficial bacteria. Several mechanisms of action, including the weakening of the outer membrane of Gram- negative bacteria, are involved in the growth inhibition. More interestingly, various berry phenolics are shown to specifically block cell-to-cell signalling (quorum sensing) in a bacterial community. Thus these phytochemicals can act in cosmetic preparations in dual way, as natural preservatives to prevent spoilage of the product, and as stabilizers of healthy skin microbiota by inhibiting the growth of harmful skin microbes and by supporting the growth of beneficial microbes.Bioprocessing including enzyme treatment and fermentation combined with special dry fractionation technologies of the berry material has resulted in increased bioactivities. Using plant cell and tissue culture technology whole biosynthetic capacity of the plant can be advantaged and even totally new beneficial compounds for the skin can be produced. The unusual phenolic profile of the cultured berry cells, as well as their fatty acid composition with a high proportion of α-linolenic acid and high protein content makes them a unique and interesting alternative for cosmetic industry. Efficient utilization of the biomolecules requires controlled delivery systems into the skin, such as microcurrent patches developed at VTT, which are based on renewable enzyme catalysts and renewable materials. The possibilities using natural plant cell-derived ingredients combined with new delivery system will also be discussed.

Anti-adhesion and antimicrobial coatings were prepared and applied on commercial thin-film-composite (TFC) polyamide (PA) membrane to enhance anti-biofouling performance. Polyvinyl alcohol (PVA) coating was modified with cationic polyhexamethylene guanidine hydrochloride (PHMG) polymer to obtain antimicrobial performance. ATR-FTIR, SEM and AFM investigated the surface chemistry and morphology of the coated membranes. The contact angle measurement was used to determine hydrophilicity and surface energy. All coated membranes revealed more hydrophilic and lower surface roughness compared to uncoated membrane. Lower number of adhered Pseudomonas aeruginosa (P. aeruginosa) bacteria was detected on coated membranes, indicating anti-adhesion performance. The colony forming unit (CFU) and diffusion inhibition zone (DIZ) tests determined antimicrobial activity of the coated membranes against Escherichia coli (E. coli) and Bacillus subtilis (B. subtilis), showing the antimicrobial performance of PHMG. The results suggested that an optimal anti-fouling surface could be obtained applying a coating, which combines anti-adhesion and antimicrobial performance.

The potential of UV mutagenesis combined with a specific selection step to generate more acid-resistant Bifidobacterium animalis subsp. lactis Bb-12 strains with improved viability in low pH food matrices was investigated in this study. A total of 144 Bb-12 UV-mutants were initially characterised. After prolonged storage in apple juice (pH 3.5) and various pheno- and genotypic tests (acid and bile tolerance, substrate utilisation, antibiotic susceptibility, aerotolerance, RAPD) two mutants (2.20 and 2.56) were chosen for further studies including cell surface morphology, stability of various traits after repeated inoculations, and performance in a fermenter and during down-stream processing. 2.20 and 2.56 showed over two Log-values better viability in pH 3.5 juice compared to Bb-12. Alterations in cell surface structures of 2.20 and 2.56 were detected with AFM, whereas other studied traits remained unchanged.In conclusion, UV mutagenesis and subsequent incubation in acidic medium enabled improving the stability of B. animalis subsp. lactis in low pH juice. Acid tolerance testing (HCl, pH 2.5, 2 h) results did not predict long-term stability of the strains in acidic food matrix (apple juice pH 3.5).

Aims: To investigate the stability of Bifidobacterium animalis ssp. lactis VTT E‐012010 (=Bb‐12) during freeze‐drying, storage and acid and bile exposure. The effect of harvesting time and composition and pH of the cryoprotectant on the survival was evaluated. The procedure was performed by using a milk‐free culture medium and cryoprotectants to produce cells for nonmilk‐based applications. Methods and Results: Bifidobacterial cells were grown in fermenters in general edible medium for 15 or 22 h. The cell mass was freeze‐dried either as non‐neutralized or neutralized using sucrose, betaine or reconstituted skim milk (control) as cryoprotectants. For stability studies freeze‐dried powders were stored at 37, 5 and −20°C for 2–6 months. In addition, acid and bile tolerance of the powders was tested. Sucrose‐formulated B. animalis ssp. lactis preparations had an excellent stability during storage at refrigerated and frozen temperatures for 5–6 months. They also had a good survival during storage at 37°C for 2 months as well as during exposure to pH 3 and 1% bile acids. No difference was observed between 15 and 22 h grown cells or between non‐neutralized and neutralized cells. Betaine proved to be a poor cryoprotectant compared with sucrose. Conclusions: Fermentation time and neutralization of cell concentrate before freeze‐drying had no impact on the storage stability and bile and acid tolerance of freeze‐dried bifidobacterial cells. The nonmilk‐based production protocol using sucrose as a cryoprotectant yielded powdery preparations with excellent stability in adverse conditions (storage at elevated temperatures and during acid and bile exposure). Significance and Impact of the Study: The results indicate that it is feasible to develop nonmilk‐based production technologies for probiotic cultures. This provides new possibilities for the development of nondairy‐based probiotic products.

Berry fruits are rich sources of bioactive compounds, such as phenolics and organic acids, which have antimicrobial activities against human pathogens. Among different berries and berry phenolics, cranberry, cloudberry, raspberry, strawberry and bilberry especially possess clear antimicrobial effects against, e.g. Salmonella and Staphylococcus. Complex phenolic polymers, like ellagitannins, are strong antibacterial agents present in cloudberry and raspberry. Several mechanisms of action in the growth inhibition of bacteria are involved, such as destabilisation of cytoplasmic membrane, permeabilisation of plasma membrane, inhibition of extracellular microbial enzymes, direct actions on microbial metabolism and deprivation of the substrates required for microbial growth. Antimicrobial activity of berries may also be related to antiadherence of bacteria to epithelial cells, which is a prerequisite for colonisation and infection of many pathogens. Antimicrobial berry compounds may have important applications in the future as natural antimicrobial agents for food industry as well as for medicine. Some of the novel approaches are discussed.

The aim of the study was to characterize the heterotrophic microbiota in surface samples and in the air of the Roman catacombs of St. Callistus and St. Domitilla. The microbiotas inhabiting different environments and substrates, including plaster, marble and tufa in illuminated and dark sites, were studied. Microbial groups examined were aerobic microorganisms, anaerobic and facultatively anaerobic bacteria, proteolytic microorganisms, actinobacteria, yeasts and moulds. Filamentous actinobacteria, Streptomyces spp., were dominant in the biofilm samples. 16S rDNA sequence similarity analysis indicated that many of the isolates were novel species. Gram-negative bacteria were a minority among the isolated bacteria: few slime forming bacteria or types most closely related to them, like Pseudomonas spp., Stenotrophomonas nitritireducens, Sinorhizobium morelense and Bosea thiooxidans, were isolated. Fungi, mainly white Lecanicillium psalliotae, Torrubiella spp., Beauveria alba and L. aranearum, were isolated from both air and biofilm samples.

Aims: The Lactobacillus plantarum strains VTT E-78076 (E76) and VTT E-79098 (E98) were studied for their antifungal potential against Fusarium species. Methods and Results:In vitro screening with automated turbidometry as well as direct and indirect impedimetric methods clearly showed Lact. plantarum cell-free extracts to be effective against Fusarium species including Fusarium avenaceum, F. culmorum, F. graminearum and F.oxysporum. However, great variation in growth inhibition was observed between different Fusarium species and even between strains. The antifungal potential of Lact. plantarum E76 culture, including cells and spent medium, was also examined in laboratory-scale malting with naturally contaminated two-rowed barley from the crops of 1990–96. The growth of the indigenous Fusarium flora was restricted by the addition of Lact. plantarum E76 to the steeping water. However, the antifungal effect was greatly dependent on the contamination level and the fungal species/strains present on barley in different years. Conclusions:Lactobacillus plantarum strains E76 and E98 had a fungistatic effect against different plant pathogenic, toxigenic and gushing-active Fusarium fungi. Significance and Impact of the Study: The present study indicates that Lact. plantarum strains with known and selected characteristics could be used as a natural, food-grade biocontrol agent for management of problems caused by Fusarium fungi during germination of cereals.

This study investigates the bacterial anti-adhesion performance of uncoated and coated reverse osmosis (RO) membranes. All the membranes were commercially available fully-aromatic thin-film-composite (TFC) polyamide (PA) membranes. Two of the TFC PA membranes (SW30 and BW30) were coated using polyvinyl alcohol (PVA) coating, while the other three membranes (LE, XLE and NF90) were uncoated. Among the characterised TFC PA membranes, the PVA coated were more hydrophilic and their surface energy was higher in comparison to uncoated. In addition, the PVA coated membranes had lower surface roughness. AFM interaction force measurement demonstrated higher repellence performance for the more polar surface. Bacteria attachment test showed differences between the coated and the uncoated membranes. Indeed, the increase in hydrophilicity and surface polarity showed decrease in the attachment of Pseudomonas aeruginosa cells. Moreover, the results demonstrated that the surface polarity showed better correlation with the attachment of the bacteria. In addition, the type of the surface roughness may somehow contribute to the bacteria repellence.

Open circuit potentials of stainless steels increased when immersed in the Baltic Sea. The ennoblement potential was +200 mVsce in 40 to 50 days when sea water temperature was below 52°C and +300–400 mVsce within

The effect of lactic acid on the outer membrane permeability of Escherichia coli O157:H7, Pseudomonas aeruginosa , and Salmonella enterica serovar Typhimurium was studied utilizing a fluorescent-probe uptake assay and sensitization to bacteriolysis. For control purposes, similar assays were performed with EDTA (a permeabilizer acting by chelation) and with hydrochloric acid, the latter at pH values corresponding to those yielded by lactic acid, and also in the presence of KCN. Already 5 mM (pH 4.0) lactic acid caused prominent permeabilization in each species, the effect in the fluorescence assay being stronger than that of EDTA or HCl. Similar results were obtained in the presence of KCN, except for P. aeruginosa , for which an increase in the effect of HCl was observed in the presence of KCN. The permeabilization by lactic and hydrochloric acid was partly abolished by MgCl 2 . Lactic acid sensitized E. coli and serovar Typhimurium to the lytic action of sodium dodecyl sulfate (SDS) more efficiently than did HCl, whereas both acids sensitized P. aeruginosa to SDS and to Triton X-100. P. aeruginosa was effectively sensitized to lysozyme by lactic acid and by HCl. Considerable proportions of lipopolysaccharide were liberated from serovar Typhimurium by these acids; analysis of liberated material by electrophoresis and by fatty acid analysis showed that lactic acid was more active than EDTA or HCl in liberating lipopolysaccharide from the outer membrane. Thus, lactic acid, in addition to its antimicrobial property due to the lowering of the pH, also functions as a permeabilizer of the gram-negative bacterial outer membrane and may act as a potentiator of the effects of other antimicrobial substances.

The antagonistic potential of nonpathogenic Staphylococcus strains against Bacillus subtilis wild and type strains were studied under conditions simulating a paper- and board-machine environment. The antimicrobial activity was measured by growth inhibition in an automated turbidimeter. The antagonistic potential was compared with that of generally used chemical biocides in a paper- and board-machine environment. The siderophore-containing extracts of Staphylococcus strains significantly inhibited vegetative growth of B. subtilis and delayed the germination of spores both in synthetic and in white-water media. The mill strains were more resistant than type strain against Staphylococcus siderophores and against chemical biocides. The Staphylococcus siderophore-containing extracts did not interfere with the bacteriostatic effect of chemical biocides, but no synergy was detected. The results indicate the potential for application of Staphylococcus siderophore-containing extracts as biocontrol agents in paper- and boardmachine environment.

Aims: To investigate how cell physiological functions can predict the stability of freeze‐dried probiotics. In addition, the effect of the fermentation pH on the stability of probiotics was investigated. Methods and Results: Fermenter‐grown (pH 5·8 or 5·0) Lactobacillus rhamnosus cells were freeze‐dried and their survival was evaluated during storage at 37°C, in apple juice and during acid [hydrochloric acid (HCl) and malic acid] and bile exposure. Cells grown at pH 5·0 were generally coping better with acid‐stress than cells grown at pH 5·8. Cells were more sensitive to malic acid compared with HCl. Short‐term stability results of Lact. rhamnosus cells in malic acid correlated well with the long‐term stability results in apple juice, whereas the results of cell membrane integrity studies were in accordance with bile exposure results. Conclusions: Malic acid exposure can prove useful in evaluating the long‐term stability of probiotic preparations in apple juice. Fermentation at reduced pH may ensure a better performance of Lact. rhamnosus cells during the subsequent acid‐stress. Significance and Impact of the Study: The beneficial effect of lowered fermentation pH to Lact. rhamnosus stability during storage in apple juice and the usefulness of malic acid test in predicting the stability were shown.

Aim: To evaluate the influence of doxycycline therapy on the composition and antibiotic susceptibility of intestinal bifidobacteria. Methods and Results: Faecal samples were collected from nine subjects receiving doxycycline therapy and ten control subjects, and analysed for bifidobacteria by culturing and PCR–DGGE (denaturing gradient gel electrophoresis). A marked decrease in the diversity (average number of amplicons detected by PCR–DGGE 0·8 in the antibiotic vs 4·3 in the control group) of Bifidobacterium populations was observed during doxycycline therapy. The proportion of a tetracycline‐resistant bifidobacterial population was higher in the antibiotic group than in the control group (83%vs 26%). Based on the tet gene PCR, resistance could be associated with the presence of tet(W). In two subjects, strains representing highly similar genetic fingerprints but different tetracycline susceptibilities were detected. A mutation causing lack of functionality in the tet(W) was observed in one of the susceptible strains. Conclusions: Doxycycline therapy had a drastic effect on the diversity and tetracycline susceptibility of intestinal Bifidobacterium populations. Significance and Impact of the Study: The use of broad‐spectrum antibiotics increased the pool of tetracycline‐resistant commensal bacteria in the intestine. The detection of resistance genes alone is not sufficient for the evaluation of bacterial antibiotic resistance.

The effect of EDTA on Salmonella enterica serovar Typhimurium was studied in different growth phases with cells grown with or without Ca2+ and Mg2+ supplementation. EDTA affected the outer membrane much more strongly in the early exponential phase than in the mid- or late exponential phase, as indicated by uptake of 1-N-phenylnaphthylamine (a nonpolar hydrophobic probe, M r 219), and detergent (SDS) susceptibility. This effect was, however, not paralleled by LPS release (determined by measuring LPS-specific fatty acids or 14C-labelled LPS in cell-free supernatants, per a standardized cell density), which remained unchanged as a function of the growth curve. The conclusion from these results is that in the early exponential phase the effect of EDTA in S. enterica involves a component that is independent of LPS release.

The ability of selected lactic acid bacteria to inhibit the growth of rope-forming Bacillus strains in laboratory experiments and in wheat bread was investigated. Growth of Bacillus subtilis and Bacillus licheniformis was inhibited by Lactobacillus plantarum VTT E-78076 and Pediococcus pentosaceus VTT E-90390 in an automated turbidometry assay and in test bakings. Rope spoilage of wheat bread was inhibited by adding 20-30 g of sourdough/100 g of wheat dough if the sourdough was fermented with Lactobacillus plantarum VTT E-78076, Pediococcus pentosaceus VTT E-90390 or Lactobacillus brevis (commercial starter culture) and the pH values of sourdoughs were adjusted below 4.0 and the amount of total titratable acidity value was >12. Addition of lactic acid alone in concentrations comparable with those formed in sourdoughs did not prevent rope spoilage. Copyright 2002 Elsevier Science Ltd.

The effect of the polycation polyethyleneimine (PEI) on the permeability properties of the Gram-negative bacterial outer membrane was investigated using Escherichia coli, Pseudomonas aeruginosa and Salmonella typhimurium as target organisms. At concentrations of less than 20 µg ml−1, PEI increased the bacterial uptake of 1-N-phenylnaphthylamine, which is a hydrophobic probe whose quantum yield is greatly increased in a lipid environment, indicating increased hydrophobic permeation of the outer membrane by PEI. The effect of PEI was comparable to that brought about by the well-known permeabilizer EDTA. Permeabilization by PEI was retarded but not completely inhibited by millimolar concentrations of MgCl2. PEI also increased the susceptibility of the test species to the hydrophobic antibiotics clindamycin, erythromycin, fucidin, novobiocin and rifampicin, without being directly bactericidal. PEI sensitized the bacteria to the lytic action of the detergent SDS in assays where the bacteria were pretreated with PEI. In assays where PEI and SDS were simultaneously present, no sensitization was observed, indicating that PEI and SDS were inactivating each other. In addition, a sensitizing effect to the nonionic detergent Triton X-100 was observed for P. aeruginosa. In conclusion, PEI was shown to be a potent permeabilizer of the outer membrane of Gram-negative bacteria.

Gram-negative bacteria are harmful in various surroundings. In the food industy their metabolites are potential cause of spoilage and this group also includes many severe or potential pathogens, such as Salmonella. Due to their ability to produce biofilms Gram-negative bacteria also cause problems in many industrial processes as well as in clinical surroundings. Control of Gram-negative bacteria is hampered by the outer membrane (OM) in the outermost layer of the cells. This layer is an intrinsic barrier for many hydrophobic agents and macromolecules. Permeabilizers are compounds that weaken OM and can thus increase the activity of antimicrobials by facililating entry of hydrophobic compounds and macromolecules into the cell where they can reach their target sites and inhibit or destroy cellular functions. The work described in this thesis shows that lactic acid acts as a permeabilizer and destabilizes the OM of Gram-negative bacteria. In addition, organic acids present in berriers, i.e. malic, sorbic and benzoic acid, were shown to weaken the OM of Gram-negative bacteria. Organic acids can poteniate the antimicrobial activity of other compounds. Microbial colonic degradation products of plant-derived phenolic compounds (3,4-dihydroxyphenylacetic acid, 3-hydroxyphenylacetic acid, 3,4-dihydroxyphenylpropionic acid, 4-hydroxyphenylpropionic acid, 3-phenylpropionic acid and 3-hydroxyphenylpropionic acid) efficiently destabilized OM of Salmonella. The studies increase our understanding of the mechanism of action of the classical chelator, ethylenediaminetetra-acetic acid (EDTA). In addition, the results indicate that the biocidic activity of benzalkonium chloride against Pseudomonas can be increased by combined use with polyethylenimine (PEI). In addition to PEI, several other potential permeabilizers, such as succimer, were shown to destabilize the OM of Gram-negative bacteria. Furthermore, combination of the results obtained from various permeability assays (e.g. uptake of a hydrophobic probe, sensitization to hydrophobic antibiotics and detergents, release of lipopolysaccharide (LPS) and LPS-specific fatty acids) with atomic force microscopy (AFM) image results increases our knowledge of the action of permeabilizers. Gram-negatiivisten bakteerien ryhmään kuuluu laaja joukko haittamikrobeja, esimerkiksi potentiaalisia patogeeneja (kuten salmonella) ja elintarvikkeissa esiintyviä pilaajamikrobeja. Monille Gram-negatiivisille bakteereille on ominaista kyky kasvaa pinnoilla ja muodostaa biofilmiä, minkä vuoksi ne aiheuttavat ongelmia teollisuudessa sekä kliinisissä ympäristöissä. Gram-negatiivisille bakteereille ominainen ulkokalvorakenne heikentää monien antimikrobisten yhdisteiden kulkeutumista solun sisään, minkä vuoksi Gram-negatiivisten bakteereiden kasvun estäminen on hankalaa. Permeabilisaattorit ovat yhdisteitä, jotka kykenevät vaurioittamaan Gram-negatiivisten bakteereiden ulkokalvorakennetta ja mahdollistavat hydrofobisten yhdisteiden, kuten antibioottien ja desinfektioaineiden, kulkeutumisen solun sisään, jossa ne pääsevät reagoimaan vaikutuskohteidensa kanssa. Tässä työssä osoitettiin, että maitohappo ja lukuisat muut orgaaniset hapot (kuten bentsoe-, omena- ja sorbiinihappo, joita luontaisesti esiintyy marjoissa) heikensivät Gram-negatiivisten bakteerien ulkokalvoa. Orgaaniset hapot voivat tehostaa antimikrobisten yhdisteiden vaikutuksia. Suolistossa marjojen sisältämistä fenoliyhdisteistä muodostuvien mikrobien metaboliatuotteiden (fenyylipropioni- ja etikkahappojohdannaisia) osoitettiin heikentävän salmonellan ulkokalvoa ja herkistävän ne hydrofobisille antibiooteille. Lisäksi työssä selkiytettiin klassisen permeabilisaattorin, EDTA-kelaattorin, vaikutusmekanismeja salmonellaa kohtaan. Yhdistämällä useita permeabilisaattoriominaisuuksia mittaavia menetelmiä ja atomivoimamikroskopian antamat tulokset havaittiin, että polyetyleeni-imiini voimakkaasti vaurioitti Pseudomonas-bakteerien ulkokalvoa ja herkisti solut kvaternaariselle bentsalkoniumkloridi-biosidille. Yhdistämällä permeabilisaattoreiden käyttö antimikrobisiin yhdisteisiin voidaan laajentaa antimikrobisten yhdisteiden vaikutuksia Gram-negatiivisiin bakteereihin. Orgaanisten happojen salmonellan ulkokalvoa heikentävä vaikutus selittää osittain esimerkiksi maitohappobakteerien luontaisen kyvyn estää näiden haittamikrobien kasvua.