Sandra M. Osés

Prevalence and quantification of Shiga-toxin producing Escherichia coli along the lamb food chain by quantitative PCR.

Shiga-toxin producing Escherichia coli (STEC), which is responsible for numerous food-borne disease outbreaks, is the most important human pathogen found in ruminants. In this study, conventional microbiology and quantitative PCR (qPCR) were used to detect and quantify STEC along the lamb food chain, from slaughterhouses to butcheries, in both meat and environmental samples. Microbial Assessment Scheme (MAS) was used to select Critical Sampling Locations (CSLs) in each establishment. The rpoB gene was used to enumerate total E. coli by qPCR, whereas the genes stx(1), stx(2) and eae were directly amplified for quantification of E. coli virulent populations. The results obtained show that E. coli carrying all three virulence genes were the most prevalent in slaughterhouses (69%), whereas E. coli with the eae gene alone were found more frequently in the processing plant (32%), and stx(1)- and stx(2)-positive E. coli were predominant in butcheries (9-10%). E. coli virulent populations were not common in butcheries. Samples determined to be positive for E. coli virulent populations after enrichment were quantified by qPCR and compared with conventional microbiology counts using validated methods. The results showed a higher number of positive CSLs for E. coli virulent populations, and higher counts were obtained when qPCR was used than when using conventional methods.

Bioactive properties of honey with propolis.

Nowadays, propolis is used as an innovative preservative and as a bioactive food supplement. Due to its bitter and astringent flavour, propolis is hardly accepted by consumers. The aim of this study was to obtain a likeable food product made with honey and propolis, whose antimicrobial, antioxidant and anti-inflammatory properties were enhanced in comparison with those of the base honeys used. 0.1%, 0.3% and 0.5% soft propolis extracts were added to honeys and the products that most appealed to the users were subjected to further research. Total phenolics, flavonoids, ABTS free radical and hydroxyl radicals scavenging and anti-inflammatory activities increased in all mixtures. Antimicrobial activity of the combined products showed synergic effects, resulting in higher results than those of the base honeys and propolis extracts. Therefore, honeys enriched with small amounts of propolis extracts are promising functional foods.

Characterization by culture-dependent and culture-independent methods of the bacterial population of suckling-lamb packaged in different atmospheres.

This study offers insight into the dynamics of bacterial populations in fresh cuts of suckling lamb under four different atmospheric conditions: air (A), and three Modified Atmosphere Packaging (MAP) environments, 15%O2/30%CO2/55%N2 (C, commercial), 70%O2/30%CO2 (O), and 15%O2/85%CO2 (H) for 18 days. Microbial analyses by both conventional methods and PCR-DGGE were performed. Controversial and surprising results emerged from comparing both methods in relation to the genus Pseudomonas. Thus, conventional methods detected the presence of high numbers of Pseudomonas colonies, although PCR-DGGE only detected this genus in air-packaged samples. PCR-DGGE detected higher microbial diversity in the control samples (A) than in the modified atmospheres (C, O, H), having atmosphere H the fewest number of species. Brochothrix thermosphacta, LAB (Carnobacterium divergens and Lactobacillus sakei), and Escherichia spp. were detected in all the atmospheres throughout storage. Moreover, previously undescribed bacteria from lamb meat such as Enterobacter hormaechei, Staphylococcus equorum and Jeotgalicoccus spp. were also isolated in this study by DGGE. Additionally, qPCR analysis was used to detect and characterize strains of Escherichia coli. Virulence genes (stx1, stx2 and eae) were detected throughout storage in 97% of the samples. A high CO2 atmosphere was the most effective packaging combination doubling storage time in comparison with commercial atmosphere.

Control of Escherichia coli and Listeria monocytogenes in suckling-lamb meat evaluated using microbial challenge tests

Escherichia coli and Listeria monocytogenes microbial challenge tests were performed on fresh suckling-lamb meat. Hind leg slices were chilly stored under two modified atmosphere packaging (MAP) environments (A: 15%O2/60%CO2/25%N2, B: 15%O2/30%CO2/55%N2) and vacuum packaging (V). Only E. coli was reduced between 0.72-1.25 log cfu/g from day 1 to day 4 by the combined use of MAP/V, chilling storage and the growth of native lactic acid bacteria. However, L. monocytogenes was not inhibited by the application of V or MAP. Even do, in inoculated samples, this pathogen increased between 1.2-2.7 log cfu/g throughout the study. Consequently, a second experiment that combined the effects of MAP/V and a protective culture (Leuconostoc pseudomesenteroides PCK 18) against L. monocytogenes was designed. Two different levels of protective cultures were assayed (4 and 6 log cfu/g). Lc. pseudomesenteroides PCK 18 was able to control the growth of L. monocytogenes when the differences between them are higher than 2 log cfu/g. Moreover, when high level of protective culture was used a significant reduction of L. monocytogenes counts were noticed in samples packaged in 60% of CO2 along the storage period, although sensory properties were also affected.

Methods of analysis of honey

A thorough updated review of both standardized and the most used and novel analytical methods for the analysis of honey is presented. The methodologies applied to honey in the analysis of the physical parameters (electrical conductivity, rheological properties, specific rotation, color and water activity), the analysis of the properties and the most important components of honey (moisture, sugars, enzymes, HMF, types of acidity and pH, formol index, insoluble solids, organic acids, proteins, amino acids, vitamins, minerals, volatile and semi-volatile compounds and polyphenols), and the antioxidant and antimicrobial activities are described. Finally, the most applied methods for multicomponent analysis and/or for honey authenticity verification (both the botanical and/or geographical origin honey classification and the detection of honey adulteration) are provided.

Analysis of Polyphenols in Honey: Extraction, Separation and Quantification Procedures

Polyphenols are secondary plant metabolites playing a major role as potentially functional components. They can also be used for honey authentication. This review gathers the recent literature references about honey extraction procedures, as well as instrumental analysis of phenolic compounds found in honey. Liquid-Liquid extraction is widely used for both extraction and purification purposes, with adequate recovery percentages. However, the use of high solvent volumes is a major disadvantage. More environmentally friendly methods include accelerated solvent extraction, and dispersive and inverse dispersive liquid-liquid microextraction. Solid phase extraction is the most common method for honey polyphenols’ isolation. Polyphenol isolation by a combination of liquid-liquid and solid phase extraction allows good recoveries for a variety of different compounds. High-performance liquid chromatography with ultraviolet or mass spectrometry detectors is by far, the most commonly employed instrumental procedure to separate and quantify polyphenols in honey although capillary electrophoresis has been also successfully used for these purposes. The use of new sorbents, the optimization of current procedures and the development of other simple and rapid analytical techniques are challenges for future analysis of polyphenols found in honey.

Design of a food product composed of honey and propolis

Propolis is a natural bee hive product with potential functional activities. Nowadays, many propolis products are available on the market, including cosmetics and food commodities. However, it is still unknown if edible products with propolis are fully accepted by consumers and are thus commercially viable, due to the bitter and resinous flavor and taste of propolis. This work is focused on the design of a “honey with propolis” foodstuff that can be welcomed by consumers, and, if possible, provide added benefits compared to the honey itself. Propolis tinctures containing three different concentrations of ethanol and times of maceration were prepared, giving the best results with 90% ethanol and two days of maceration. Then, in order to study the maximum amount of propolis in honey that could be accepted by a representative group of consumers, a forced choice paired comparison preference test with 69 people was carried out, giving the best results with a honey product containing 0.3-0.5% propolis extract. Phenolic and flavonoid contents, as well as TEAC antioxidant and antimicrobial activities of propolis, honey, and the product “honey with propolis” were then analyzed. As expected, propolis showed the highest phenolics and flavonoids contents, as well as the highest antioxidant and antimicrobial activities. The product “honey with propolis” showed a significantly higher antimicrobial activity, however, when compared to honey alone. In conclusion, the commercialization of a food product of “honey and up to 0.5% propolis” is promising, interesting and viable, as such a product is greatly appreciated by consumers, as well as being potentially healthy.