Assessment of the Sensitivity of Foodborne Cronobacter spp. Strains and Other Enterobacteria to Temperature Stresses and Chlorine-Containing Biocides.

We examined the effects of elevated temperatures and biocides on survivability of food isolates of Cronobacter spp. (C. sakazakii) and concomitant enterobacteriaceae obtained in microbiological control of infant nutrition products. Increased resistance of certain strains of Cronobacter, Enterobacter cloacae, and Pantoea spp. to thermal processing was revealed. Salmonella, Pantoea, and Cronobacter bacteria were least sensitive to antimicrobial action of chlorine-containing agents. The above properties varied in the strains of the same species. Specifically, only two of three examined isolates of Cronobacter spp. demonstrated lower sensitivity to heat in comparison with the enterobacterial test-cultures of other species.

Improvement of Methods for Safety Control of Microbial Producers and Food Produced Therewith.

The study substantiates the necessity to implement the algorithm of molecular-genetic assessment of biosafety of the genetically modified microorganisms (GMM) and to develop standardized methods to test the genetically modified strains producing enzymes, bioactive substances, and other products of microbial synthesis prior to their use in food industry. Analysis of microbial producers and related food products for the presence of GMM-associated DNA revealed high incidence of the marker genes amp and lacZ in enzyme preparations and in mycelium of industrial genetically modified producer of Aspergillus genus. The procedure of extraction of DNA from mycelium of mold fungi is optimized by including the stage of additional purification of the extracts, assessment of their purity by PCR with universal ITS primers, and determination of effective DNA concentration in the samples prior to conduction of the molecular genetic assay. For identification and genotyping of mold fungi (the biotechnological producers of enzyme preparations), the Sanger sequencing method was adapted. Using this modified method, we determined the species of five equivocally identified strains of Aspergillus genus. To identify the closely-related micromycetes of Ascomycota division, a genotyping algorithm was developed based on amplification of total DNA with expanded panel of primers and DNA sequencing by capillary electrophoresis.

[Assessment of the influence of an enzymal preparation - a complex of glucoamylase and xylanase from Aspergillus awamori Xyl T-15 on the intestinal microbiom and immunological indicators of rats].

The requirements for the safety of food products obtained by microbial synthesis are including as obligation for to conduct toxicological studies - the study of various biochemical and immunological markers of toxic effects. The necessity of these studies is explained by a possible change in the structure of food ingredients produced by a microbial cell and, consequently, a change in their biological properties, as well as the possible presence of living forms and/or DNA of producer strains or of their toxic metabolites in these ingredients. At the same time, it is well known that the nutrient composition of foods has a significant impact on the composition and properties of microorganisms that make up the gut microbiome, which, in turn, determines the immune status. The purpose of the research was to justify the analyses of gut microbiocenosis composition for inclusion in the protocol of safety investigation of foods obtained by microbial synthesis [on the example of an enzyme preparation (EP) - a complex of glucoamylase and xylanase from a genetically modified strain of Aspergillus awamori Xyl T-15]. Material and methods. In experimental studies carried out for 80 days, Wistar rats (males and females) were used. The study of the effect of EP (a complex of glucoamylase and xylanase from a genetically modified Aspergillus awamori Xyl T-15 strain) in dozes 10, 100 and 1000 mg/kg body mass on the cecum microbiome and the immune status (content of cytokines and chemokines: IL-1a, IL-4, IL-6, IL-10, IL-17A, INF-γ, TNF-α, MCP-1, MIP-1a and Regulated on Activation Normal T-cell Expressed and Secreted - RANTES) was carried out. Results. It has been shown that EP - a complex of glucoamylase and xylanase from A. awamori Xyl T-15 at doses of 100 mg/kg or more causes mild disturbances in the composition of gut microbiocenosis. At the same time, these disorders have a significant immunomodulat ory and immunotoxic effect on the body, which manifests itself in a dose-dependent change in the profile of pro-inflammatory cytokines and chemokines in blood and spleen. The adverse effect of EP on the body is probably due to the formation of metabolites that are not formed during usual digestive processes in the gastrointestinal tract. The minimum effective dose (LOAEL) of EP was 100 mg/kg body weight In accordance with established requirements, the activity of the EP should not appear in ready-to-use food. Subject to this requirement, amount of EP entering the body cannot exceed the established LOAEL level. Therefore, a complex of glucoamylase and xylanase can be used in food industry, subject to the establishment of regulations «for technological purposes¼ for A. awamori Xyl T-15 strain. Conclusion. The data obtained on the relationship between the state of the microbiome and the immune status upon the introduction of EP indicate the need to include indicators of the state of gut microbiocenosis in the test protocol of safety.

[Minor antibiotics residues in food: what are the risks for consumers].

The problem of contamination of foodstuffs with antibiotic residues does not lose its relevance everywhere, and the most widespread are the quantities of contaminants at the level of regulated values. This raises the concern of specialists in the field of production and processing of livestock products and initiates their appeal to scientific organizations of the hygienic profile for an explanation of the potential health risks associated with the consumption of low doses of antibiotics with food. Material and methods. Analysis and synthesis of data from scientific sources and official documents in the field of assessing health risks when consuming antibiotics with food, with an emphasis on the effects of minor amounts [at the level of sub-inhibitory values below minimum inhibitory concentrations has been carried out. Results and discussion. The issues of direct and indirect human exposure to antibiotics in low doses, including the formation of resistance of intestinal bacteria and the acceleration of the evolution of microbes, accumulation in the organism, the likelihood of allergic reactions, as well as preservation in foodstuffs during heat treatment, have been highlighted. The role of low doses of antibiotics as analogues of biologically active metabolites of bacteria is demonstrated, which, without exerting a toxic effect on the macroorganism, serve as triggers of changes in the microbial ecosystems of humans, animals and habitats through the mechanism of switching on regulation transcription in microbes and activation of horizontal transfer of genes encoding resistance and associated traits. The negativity of the disproportionately wide use of tetracyclines in agriculture, as the cause of the globalization of transferable resistance, has been emphasized, which was justified by the data on the ability of their sub-inhibitory doses to induce the expression of the largest number of mechanisms of its formation and most strongly provoke the horizontal transfer of linked genes between microbes. The need to preserve the current in the EAEU MRL for tetracyclines (0.01 mg/kg product), located in the concentration zone 0.05-0.1 minimum inhibitory concentrations for most sensitive bacteria, safe in terms of resistance induction, has been confirmed. Conclusion. Adequate rationing of antibiotics in food is recognized as a risk management measure for both direct and indirect negative consequences for human health, since the need to ensure MRLs requires manufacturers to strictly adhere to doses, duration of use and withdrawal periods of drugs. This reduce the likelihood of developing resistance in the gastrointestinal tract of animals, the load of the environment by resistant microbes and their transmission along the food chain. The underlying establishment of microbiological acceptable daily intake should be improved by including the selection of intestinal bacteria co-resistance under the influence of a regulated drug, as a marker of the induction of horizontal transfer of resistance genes, in the biological endpoints of determining.