Raw milk and allergy prevention – a possible feature for organic milk?

This review aims to give some update on the role of raw milk and/or its constituents in allergy prevention, also considering possible advantages of organic milk in such a context. In addition, challenges and strategies will be discussed to control and maintain safety of a raw milk-based formula product

Comparison of Selective Media for the Enumeration of Probiotic Enterococci from Animal Feed

The project »Methods for the Official Control of Probiotics Used as Feed Additives« has been undertaken to develop and validate methods for the selective enumeration and strain identification of six probiotic microorganism genera (enterococci, lactobacilli, bifidobacteria, pediococci, bacilli and yeast). A diversity of media has been used for the detection, isolation and enumeration of enterococci. Aiming at the selective enumeration of enterococci (mainly Enterococcus faecium) present in probiotic animal feeds, either as a single component or in combination with other microorganisms, an extensive screening of published methods for culturing and enumerating enterococci was carried out. A collection of enterococcal strains used as probiotics in animal feeds and of isolates as well as reference strains from culture collections was established. Moreover, selected strains of lactobacilli, pediococci and streptococci were included for reference purposes. Based on a multitude of publications, twelve commercially available media were selected for testing and then compared with regard to their usefulness and selectivity. Bile esculin azide (BEA) agar showed good selectivity and pronounced growth of most enterococcal strains. Good reproducibility and electivity (esculin hydrolysis) as well as no influence of the feed matrix on the colony counts and a simple preparation procedure formed the basis for the proposed enumeration protocol. This work formed the basis for the enumeration protocol that was adapted to ISO format and validated in a collaborative study involving twenty laboratories from twelve European countries

Comparison of Selective Media for the Enumeration of Probiotic Enterococci from Animal Feed

The project »Methods for the Official Control of Probiotics Used as Feed Additives« has been undertaken to develop and validate methods for the selective enumeration and strain identification of six probiotic microorganism genera (enterococci, lactobacilli, bifidobacteria, pediococci, bacilli and yeast). A diversity of media has been used for the detection, isolation and enumeration of enterococci. Aiming at the selective enumeration of enterococci (mainly Enterococcus faecium) present in probiotic animal feeds, either as a single component or in combination with other microorganisms, an extensive screening of published methods for culturing and enumerating enterococci was carried out. A collection of enterococcal strains used as probiotics in animal feeds and of isolates as well as reference strains from culture collections was established. Moreover, selected strains of lactobacilli, pediococci and streptococci were included for reference purposes. Based on a multitude of publications, twelve commercially available media were selected for testing and then compared with regard to their usefulness and selectivity. Bile esculin azide (BEA) agar showed good selectivity and pronounced growth of most enterococcal strains. Good reproducibility and electivity (esculin hydrolysis) as well as no influence of the feed matrix on the colony counts and a simple preparation procedure formed the basis for the proposed enumeration protocol. This work formed the basis for the enumeration protocol that was adapted to ISO format and validated in a collaborative study involving twenty laboratories from twelve European countries

Susceptibility of Bifidobacteria of Animal Origin to Selected Antimicrobial Agents

Strains of the genus Bifidobacterium are frequently used as probiotics, for which the absence of acquired antimicrobial resistance has become an important safety criterion. This clarifies the need for antibiotic susceptibility data for bifidobacteria. Based on a recently published standard for antimicrobial susceptibility testing of bifidobacteria with broth microdilution method, the range of susceptibility to selected antibiotics in 117 animal bifidobacterial strains was examined. Narrow unimodal MIC distributions either situated at the low-end (chloramphenicol, linezolid, and quinupristin/dalfopristin) or high-end (kanamycin, neomycin) concentration range could be detected. In contrast, the MIC distribution of trimethoprim was multimodal. Data derived from this study can be used as a basis for reviewing or verifying present microbiological breakpoints suggested by regulatory agencies to assess the safety of these micro-organisms intended for the use in probiotics

Temporal expression of the staphylococcal enterotoxin D gene under NaCl stress conditions

Staphylococcus aureus is one of the most osmotolerant food-borne pathogens. While its growth is repressed by competing bacteria, the organism exhibits a growth advantage at increased salt concentrations. Staphylococcal enterotoxin D leads to vomiting and diarrhea upon ingestion. To date, the effect of NaCl on both sed expression and its regulatory control are unclear. We determined the impact of NaCl stress on sed expression and the influence of agr, sarA and sigB on sed expression under NaCl stress. The temporal expression of sed in LB and LB with 4.5% NaCl was compared, as well as sed expression of wild-type (wt) strains and isogenic Δagr, ΔsarA and ΔsigB mutants. In general, NaCl stress led to decreased sed expression. However, one strain exhibited a trend towards increased sed expression under NaCl stress. No significant effect of agr on sed expression was detected and only one ΔsigB mutant showed a significant decrease in sed expression in the early stationary phase under NaCl stress. One ΔsarA mutant showed decreased sed expression in the early stationary and another increased sed expression in the stationary growth phase under NaCl stress. These findings suggest high strain-specific variation in sed expression and its regulation under NaCl stres

Molecular Methods Used for the Identification of Potentially Probiotic Lactobacillus reuteri Strains

Forty potentially probiotic Lactobacillus strains as well as reference strains of different genera were grown under standardised conditions. Cell masses were harvested and DNA was isolated. For identification, all strains were subjected to genus-specific polymerase chain reaction (PCR), and the affiliation with the genus Lactobacillus was confirmed for all isolates. Using two species-specific primer-pairs for Lactobacillus reuteri, specific amplicons were observed for eight of the forty investigated strains. For differentiation, these eight strains as well as the reference strains of the species L. reuteri and closely related species were subjected to randomly amplified polymorphic DNA (RAPD)-PCR using fourteen arbitrary primers. Two selected strains as well as probiotic and common reference strains were further investigated applying pulsed field gel electrophoresis (PFGE). With the latter two methods, individual profiles were found for most strains, but no difference between probiotic and common strains could be made out

Molecular Methods Used for the Identification of Potentially Probiotic Lactobacillus reuteri Strains

Forty potentially probiotic Lactobacillus strains as well as reference strains of different genera were grown under standardised conditions. Cell masses were harvested and DNA was isolated. For identification, all strains were subjected to genus-specific polymerase chain reaction (PCR), and the affiliation with the genus Lactobacillus was confirmed for all isolates. Using two species-specific primer-pairs for Lactobacillus reuteri, specific amplicons were observed for eight of the forty investigated strains. For differentiation, these eight strains as well as the reference strains of the species L. reuteri and closely related species were subjected to randomly amplified polymorphic DNA (RAPD)-PCR using fourteen arbitrary primers. Two selected strains as well as probiotic and common reference strains were further investigated applying pulsed field gel electrophoresis (PFGE). With the latter two methods, individual profiles were found for most strains, but no difference between probiotic and common strains could be made out

Safety of Novel Microbes for Human Consumption : Practical Examples of Assessment in the European Union

Novel microbes are either newly isolated genera and species from natural sources or bacterial strains derived from existing bacteria. Novel microbes are gaining increasing attention for the general aims to preserve and modify foods and to modulate gut microbiota. The use of novel microbes to improve health outcomes is of particular interest because growing evidence points to the importance of gut microbiota in human health. As well, some recently isolated microorganisms have promise for use as probiotics, although in-depth assessment of their safety is necessary. Recent examples of microorganisms calling for more detailed evaluation include Bacteroides xylanisolvens, Akkermansia muciniphila, fructophilic lactic acid bacteria (FLAB), and Faecalibacterium prausnitzii. This paper discusses each candidate's safety evaluation for novel food or novel food ingredient approval according to European Union (EU) regulations. The factors evaluated include their beneficial properties, antibiotic resistance profiling, history of safe use (if available), publication of the genomic sequence, toxicological studies in agreement with novel food regulations, and the qualified presumptions of safety. Sufficient evidences have made possible to support and authorize the use of heat-inactivated B. xylanisolvens in the European Union. In the case of A. muciniphila, the discussion focuses on earlier safety studies and the strain's suitability. FLAB are also subjected to standard safety assessments, which, along with their proximity to lactic acid bacteria generally considered to be safe, may lead to novel food authorization in the future. Further research with F. prausnitzii will increase knowledge about its safety and probiotic properties and may lead to its future use as novel food. Upcoming changes in EUU Regulation 2015/2283 on novel food will facilitate the authorization of future novel products and might increase the presence of novel microbes in the food market.Peer reviewe