Marek Koutný

Formation, Degradation, and Detoxification of Putrescine by Foodborne Bacteria: A Review

Biogenic amines (BAs) represent a considerable toxicological risk in some food products. Putrescine is one of the most common BAs in food. Its increased occurrence in food may lead to alimentary poisoning, due to enhancement of the toxic effects of other BAs, and also to lower quality of food, this amine is potentially carcinogenic. Increased occurrence of putrescine in food is mainly due to the bacterial metabolism of the Gram-negative as well as Gram-positive bacteria present. The bacterial metabolism of putrescine is very specific due to its complexity (in comparison with the metabolism of other BAs). There are 3 distinct known pathways leading toward the formation of putrescine, in some splices involving up to 6 different enzymes. The existence of more metabolic pathways and the possibility of their simultaneous use by different bacteria complicate the specification of the best conditions for food production and storage, which could lead to a lower content of putrescine. This review provides a summary of the existing knowledge about putrescine production and detection (mainly detection of specific genes for different enzymes using polymerase chain reaction) in both starter and contaminating microorganisms. Thus, this comprehensive review gives a useful overview for further research.

Xanthan and gellan degradation by bacteria of activated sludge

Owing to increasing amounts of xanthan and gellan in food, cosmetics and pharmaceuticals, as well as in some technical spheres, studies were carried out on the xanthan and gellan degrading bacteria present in activated sludge. The activated sludge used in the study was able to degrade both carbohydrates over 7 days, with levels of xanthan and gellan utilizing microbes estimated at 10(5) cells/g of dry sludge weight. Isolating key degrading bacteria revealed the important role of genus Paenibacillus in xanthan degradation and prosthecate bacterium Verrucomicrobium sp. GD, which was capable of gellan utilization. Further tests performed with both strains showed they were able to degrade other types of carbohydrate polymers, and that Verrucomicrobium sp. GD did not possess extracellular free gellan depolymerase.

Novel touchdown-PCR method for the detection of putrescine producing gram-negative bacteria in food products.

Formation of biogenic amines may occur in food due to metabolic activities of contaminating Gram-negative bacteria. Putrescine is assumed to be the major biogenic amine associated with microbial food spoilage. Gram-negative bacteria can form putrescine by three metabolic pathways that can include eight different enzymes. The objective of this study was to design new sets of primers able to detect all important enzymes involved in the production of putrescine by Gram-negative bacteria. Seven new sets of consensual primers based on gene sequences of different bacteria were designed and used for detection of the speA, adiA, adi, speB, aguA, speC, and speF genes. A newly developed touchdown polymerase chain reaction (PCR) method using these primers was successfully applied on several putrescine-producers. Selected PCR products were sequenced and high similarity of their sequences (99-91%) with known sequences of the corresponding genes confirmed high specificity of the developed sets of primers. Furthermore, all the investigated bacteria produced both putrescine and agmatine, an intermediate of putrescine production, which was confirmed by chemical analysis. The developed new touchdown PCR method could easily be used to detect potential foodborne Gram-negative producers of putrescine. The newly developed sets of primers could also be useful in further research on putrescine metabolism in contaminating microbiota.

Antibacterial effect of phosphates and polyphosphates with different chain length

The aim of this study was to monitor the antibacterial effect of seven phosphate salts on selected strains of Gram-negative and Gram-positive bacteria, which could be considered responsible for food-borne diseases (Bacillus cereus, Bacillus subtilis, Enterococcus faecalis, Micrococcus luteus, Staphylococcus aureus, Citrobacter freundii, Escherichia coli, Proteus mirabilis, Salmonella enterica ser. Enteritidis and Pseudomonas aeruginosa). For these purposes, phosphates differing in chain length were used. The tested concentrations were in the range of 0.1–2.0% (wt v−1) applied at the model conditions. In the majority of cases the visible inhibitory effect on the growth of observed microorganisms could be seen. Due to the chemical structure of salts and their dissociation both the pH values of cultivation broth and similarly the growth characteristics of bacterial strains were affected. The inhibition of above mentioned bacteria was apparently supported by this dissociation. Phosphates obviously made the development of most Gram-positive bacteria impossible. Especially Micrococcus luteus was extremely sensitive to the presence of these substances. On the other hand, Gram-negative bacteria seemed to be resistant to the phosphate incidence. The exemption clause from the tested salts was represented by a high alkaline trisodium phosphate. It should be pointed out that generally the most significant antibacterial effects were shown by polyphosphates HEXA68 and HEXA70, trisodium phosphate undecahydrate, tetrasodium pyrophosphate and finally trisodium phosphate. By comparing the inhibitory effects of various phosphate salts can be concluded that the antibacterial activity was not determined only by the condensation degree but there was also proved the dependence on pH values.

Accelerated Biodegradation of Agriculture Film Based on Aromatic–Aliphatic Copolyester in Soil under Mesophilic Conditions

A study was conducted on the biodegradation of aromatic-aliphatic copolyester-based agricultural film in soil at 25 °C. The polymer is known to be biodegradable under composting conditions although rather recalcitrant under mesophilic conditions. The material investigated comprised of the copolyester filled with approximately 25% of starch containing biodegradable plasticizers, and its behavior was compared to the corresponding material without the filler. Mineralization followed by CO2 production merely reached the point of about 6% after 100 days of incubation in the pure copolyester film, whereas the value of around 53% was recorded for the filled copolyester film, which exceeded the readily biodegradable starch filler content in the material by more than 20% and could be accounted for biodegradation of the copolyester. It was suggested that the accelerated copolyester biodegradation in the starch-filled material was most likely explained by the increase in the active surface area of the material available for the microbial attack after biodegradation of the filler. The results were supported by changes in molecular weight distributions of the copolyester and observations made by several microscopic techniques. These findings encourage further development of biodegradable agricultural films based on this material.

Changes of physical properties of PLA-based blends during early stage of biodegradation in compost

Three biodegradable plastics materials, namely pure poly(l-lactide) (PLA), PLA with plasticizer triacetine (TAC) and the mixture PLA/polyhydroxybutyrate (PHB) and TAC were investigated concerning changes of physical properties due to biodegradation in compost at 58°C up to 16days. With rising time of degradation in compost, both number and weight molecular masses were decreasing progressively, but only marginal change of the polydispersity index was observed which indicates that biodegradation is not random process. FTIR spectroscopy revealed that in spite of the extensive decrease of molecular weight, no substantial change in chemical composition was found. The most significant modification of the spectra consisted in an appearing of the broad band in region 3100-3300cm-1, which was assigned to a formation of biofilm on the sample surfaces. This effect appeared for all three materials, however, it was much more pronounced for samples containing also triacetine. Measurement of changes in crystalline portion confirmed that amorphous phase degrades substantially faster compared to crystalline part. The plasticizer triacetine is disappearing also rather fast from the sample resulting besides other effect also in a temporary increase of Tg, which at the beginning grows almost to the value typical for PLA without plasticizer but later the Tg is decreasing due to substantial changes in molecular weight. Generally during composting, the samples keep shape for up to 8days, after that time the material disintegrates to rough powder.