Occurrence of mycotoxins in talkan: a cereal-based food traditional for Turkic population.

The consumption of cereal contaminated with mycotoxins poses a serious health risk for humans and animals. The present work aims to evaluate the presence of mycotoxins in talkan, a cereal-based food commonly consumed by the Turkic population. The presence of mycotoxins was investigated in a total of 50 samples obtained from Kazakhstan. After a preliminary screening using various ELISA kits, mycotoxins were confirmed and quantified by HPLC-MS/MS method. More than 28% of the samples were positive for at least one mycotoxin. The calculated probably daily intake for adults and children was 20% above the tolerable daily intake for aflatoxin B1 and deoxynivalenol, while it was above 100% for zearalenone, indicating a high risk for the Kazakh population. A total of 12 samples exhibited concentrations above the European maximum level for ochratoxin A, zearalenone and deoxynivalenol, however, these values were within the limits established by the Russia-Kazakhstan-Belarus Customs Union (TR CU 015/2011).

Sleep disorders in Latin-American children with asthma and/or allergic rhinitis and normal controls.

BACKGROUND: Asthma and/or allergic rhinitis have been associated with sleep disorders. The aim of this study was to evaluate sleep disorders in Latin-American children (4-10 years) from nine countries, with persistent asthma (A) and/or allergic rhinitis (AR) and in normal controls (C). METHODS: Parents from 454 C children and 700 A and/or AR children followed up in allergy reference clinics completed the Children's Sleep Habits Questionnaire (CSHQ) which is a retrospective one-week questionnaire composed of 33 questions composed of seven subscales (bedtime resistance, sleep duration, sleep anxiety, night wakings, parasomnias, sleep-disordered breathing and daytime sleepiness). The total scale of CSHQ and the subscales were compared between groups C and A+AR, A (n=285) vs. AR (n=390), and between controlled A (CA, n=103) vs. partially controlled/uncontrolled A (UA, n=182). RESULTS: The comparison between C and A+AR showed no significant differences in age (6.7 years vs. 7.0 years, respectively), mean Body Mass Index and total scale of CSHQ (53.3 vs. 63.2, respectively) and the subscales were significantly higher in the A+AR group. Comparison between groups A and AR, except for sleep anxiety, showed significantly higher values for CSHQ total scale (66.9 vs. 61.0, respectively) and subscales for group A. The UA group showed significantly higher values for total CSHQ scale and subscales in comparison to CA (71.1 vs. 59.4, respectively). CONCLUSIONS: Latin-American children with asthma and/or allergic rhinitis showed sleep disorders identified by the CSHQ when compared to normal controls. Despite being treated, asthma causes sleep impairment, especially when uncontrolled.

Sleep-related disorders in Latin-American children with atopic dermatitis: A case control study.

BACKGROUND: Atopic dermatitis (AD) has been associated with impairment of sleep. The aim of this study was to evaluate sleep disorders in AD Latin-American children (4-10 years) from nine countries, and in normal controls (C). METHODS: Parents from 454 C and 340 AD children from referral clinics answered the Children Sleep Habits Questionnaire (CSHQ), a one-week retrospective 33 questions survey under seven items (bedtime resistance, sleep duration, sleep anxiety, night awakening, parasomnias, sleep-disordered breathing and daytime sleepiness). Total CSHQ score and items were analysed in both C and AD groups. Spearman's correlation coefficient between SCORAD (Scoring atopic dermatitis), all subscales and total CSHQ were also obtained. RESULTS: C and AD groups were similar regarding age, however, significantly higher values for total CSHQ (62.2±16.1 vs 53.3±12.7, respectively) and items were observed among AD children in comparison to C, and they were higher among those with moderate (54.8%) or severe (4.3%) AD. Except for sleep duration (r=-0.02, p=0.698), there was a significant Spearman's correlation index for bedtime resistance (0.24, p<0.0001), sleep anxiety (0.29, p<0.0001), night awakening (0.36, p<0.0001), parasomnias (0.54, p<0.0001), sleep-disordered breathing (0.42, p<0.0001), daytime sleepiness (0.26, p<0.0001) and total CSHQ (0.46, p<0.0001). AD patients had significantly higher elevated body mass index. CONCLUSION: Latin-American children with AD have sleep disorders despite treatment, and those with moderate to severe forms had marked changes in CSHQ.

Assessment of food safety using a new real-time PCR assay for detection and quantification of virulence factors of enterococci in food samples.

AIMS: Development of Taqman MGB real-time PCR (q-PCR) assays for the quantitative detection of virulence factor genes in pure culture and food samples with regard to food safety assessment. METHODS AND RESULTS: New Taqman primers and probes were designed for the ace, esp and gelE genes based on the determinants of virulence profiles of enterococcal strains from GenBank. The high specificity and accuracy of the Taqman probe assay was confirmed. The limit of detection for the different virulence genes was 102  CFU ml-1 or CFU g-1 for pure culture and meat samples, and 103  CFU g-1 for cheese samples. CONCLUSION: This method provides the specific and rapid detection and quantification of ace, esp and gelE genes compared to conventional PCR assays, thus allowing the rapid and direct safety assessment of Enterococcus genus in food samples. SIGNIFICANCE AND IMPACT OF THE STUDY: This study presents efficient methods that can be used directly on food products for the rapid quantification and tracing of virulence genes, regarding food safety assessment. Moreover, this is the first study to quantify these virulence factors using a specific Taqman q-PCR assay in food samples.

Development of a real-time PCR assay for direct detection and quantification of Bacillus sporothermodurans in ultra-high temperature milk.

In our study, a new and highly sensitive real-time PCR Taqman assay was developed for the direct and specific detection of Bacillus sporothermodurans in UHT milk. The target region was selected based on the 16S rRNA gene profiles of 11 B. sporothermodurans from GenBank. A standard curve was created using a reference strain of B. sporothermodurans, DSM 10599. A low limit of detection for B. sporothermodurans in UHT milk (10 cfu/mL) was obtained. Furthermore, a total of 110 UHT milk samples from several supermarkets were directly assessed to detect and quantify B. sporothermodurans using the real-time PCR Taqman developed. The B. sporothermodurans counts obtained were highly correlated with the microbial plate counts in the UHT milk samples. This is the first time that B. sporothermodurans has been quantified directly from UHT milk. This technique could be applicable as a routine tool for preventing the growth of these bacteria by allowing for the rapid screening of raw milk samples in dairy plants. As expected, the probability of bacterial growth in UHT milk packages increased with the B. sporothermodurans counts in the raw milk.

A study on toxic and essential elements in rice from the Republic of Kazakhstan: comparing the level of contamination in rice from the European Community.

Selected toxic elements (total As, Cd, Cr, Hg, Pb, Sr, U and V) and essential elements (Co, Cu, Fe, Mn and Zn) were analyzed using an inductively coupled plasma mass spectrometry (ICP-MS) in unpolished and milled rice collected from Kazakhstan and milled rice from Spain and Portugal to evaluate the potential health risk to the population. Arsenic species (arsenite, arsenate, arsenobetaine, dimethylarsinate and monomethilarsonate) were analyzed using HPLC-IC-MS. From 146 samples analyzed, none of them exceeded the maximum limit set by the European Legislation for Cd or Pb or values recommended by the Codex Alimentarius. Concentrations of Sr, U and V were below LOD and those of Hg, Pb, Co and Cr between

Characterisation and profiling of Bacillus subtilis, Bacillus cereus and Bacillus licheniformis by MALDI-TOF mass fingerprinting.

The Bacillus genus includes species such as Bacillus cereus, Bacillus licheniformis and Bacillus subtilis, some of which may be pathogenic or causative agents in the spoilage of food products. The main goal of this work was to apply matrix-assisted laser desorption ionisation-time of flight (MALDI-TOF) mass fingerprinting to the classification of these Bacillus species. Genetic analyses were also compared to phyloproteomic analyses. A collection of 57 Bacillus strains isolated from fresh and processed food and from culture collections were studied and their mass spectra compiled. The resulting mass fingerprints were compared and characteristic peaks at the strain and species levels were assigned. The results showed that MALDI-TOF was a good complementary approach to 16S rRNA sequencing and even a more powerful tool in the accurate classification of Bacillus species, especially for differentiating B. subtilis and B. cereus from Bacillus amyloliquefaciens and Bacillus thuringiensis, respectively. MALDI-TOF was also found to provide valuable information at both intra- and interspecies levels in the Bacillus species studied.

Detection and quantification of spoilage and pathogenic Bacillus cereus, Bacillus subtilis and Bacillus licheniformis by real-time PCR.

A new primer-probe set for the detection and quantification of Bacillus cereus, Bacillus licheniformis and Bacillus subtilis by real-time PCR (Rti-PCR) was developed. For it, forty-eight strains belonging to these species were considered. The DNA of these strains was isolated and a fragment of the 16S rRNA gene amplified. The amplicons were sequenced and the obtained sequences were aligned with reference sequences from the GenBank. For the development of the Real-Time PCR (RTi-PCR) methodology based on TaqMan probes, a primer pair and probe, specific for the studied Bacillus spp., were designed. To establish the quantification method, two RTi-PCR standard curves were constructed; one with DNA extracted from a serially-diluted B. cereus culture and a second curve with DNA extracted from a sterilised food product inoculated with serial dilutions of B. cereus. The curves exhibited R(2) values of 0.9969 and 0.9958 respectively. Linear correlations between the log(10) input DNA concentration and the threshold cycle (Ct) values were observed with a magnitude of linearity in the range of 1.65 × 10(1) CFU/mL to 1.65 × 10(6) CFU/mL for both standard curves. The specificity of the designed primers and probe was tested with DNA extracted from B. cereus, B. licheniformis and B. subtilis strains, which gave Ct values between 14 and 15, whereas non-specific amplifications of the DNA from other microbial species of food interest exhibited a Ct value above 28.5. To our knowledge, this method represents the first study about the quantification of spoilage and/or pathogenic B. cereus, B. licheniformis and B. subtilis in food products, with the aim to prevent the presence of these undesirable species in the food chain.

Differentiation of farmed and wild turbot (Psetta maxima): proximate chemical composition, fatty acid profile, trace minerals and antimicrobial resistance of contaminant bacteria.

The proximate, cholesterol, fatty acid and trace mineral compositions in the flesh of farmed and wild turbot (Psetta maxima) were evaluated. Additionally, the potential influence of the use of antimicrobial agents in the bacteria carried by farmed turbot was investigated. For this purpose, a total of 144 Pseudomonas spp. and 127 Aeromonas spp. were isolated and tested for their susceptibility to 12 antimicrobials by a disk diffusion method. Farmed turbot contained higher fat, cholesterol and calories as well as lower moisture content than its wild counterpart. The fatty acid profile of farmed turbot included higher levels of myristic, pentadecanoic, palmitoleic, gadoleic, cetoleic, linoleic, linolenic, stearidonic, eicosadienoic and eicosapentaenoic acids, and lower levels of stearic, arachidonic, docosapentaenoic and docosahexaenoic acids than its wild counterpart. The proportions of polyunsaturated fatty acids and n-3/n-6 ratios were higher in wild turbot than in farmed turbot. With respect to trace minerals, no toxic levels were found, and higher amounts of Cd, Co, Cu, Fe, Mn, Pb and Zn, as well as lower amounts of Cr, were found in farmed turbot relative to wild turbot. The antimicrobial resistance of Pseudomonas spp. and Aeromonas spp. were quite similar, with only the trimethoprim-sulfamethoxazole resistance of Aeromonas spp. isolated from farmed turbot being higher than those isolated from wild turbot. In the case of ampicillin, Pseudomonas spp. isolated from wild turbot showed higher resistance levels than those of their counterparts isolated from farmed turbot. In conclusion, the nutritional parameters of wild turbot are more adequate with respect to nutritional recommendations, while no differences were observed in food safety derived from trace mineral concentrations or the antimicrobial resistance of bacteria isolated from wild and farmed turbot.

Evolution of breast milk fatty acids in Spanish mothers after one year of uninterrupted lactation.

The WHO recommends exclusive breastfeeding of infants for the first sixth months of life and advises that it shall continue for up to two years of age or beyond in combination with complementary foods. However, the image of a woman breastfeeding a toddler or a preschooler is unusual in western societies. Exploring the nutritional properties of milk during prolonged lactation can help normalizing prolonged breastfeeding. Human milk fatty acid composition was determined in sixteen lactating mothers practicing prolonged lactation (≥12 months) and sixteen women on their first twelve months of lactation. Breast milk after one year is richer in saturated fatty acids, particularly lauric and myristic, showing a tendency towards lower levels of oleic acid, and higher of arachidonic, α-linolenic and docosahexaenoic acids, in comparison to early milk (< 1 year). The age and body condition of the mother, parity, sex of the baby, and diet influence also the fattyacidome of milk.

Influence of farming methods on microbiological contamination and prevalence of resistance to antimicrobial drugs in isolates from beef.

The presence of Escherichia coli, Staphylococcus aureus, Listeria monocytogenes and Salmonella spp. was determined in 75 samples of conventional beef and in 75 samples of organic beef. All samples came from cattle slaughtered and processed in the same slaughterhouse and quartering room. A total of 180 E. coli, 180 S. aureus and 98 L. monocytogenes strains were analyzed by an agar disk diffusion assay for their resistance to 11 antimicrobials, for the case of E. coli and S. aureus, or 9 antimicrobials, for the case of L. monocytogenes. Salmonella spp. were not isolated from any of the beef samples. No significant differences in prevalence were obtained for any of the bacterial species tested between organic and conventional beef. E. coli isolated from organic beef exhibited significant differences in antimicrobial resistance against 5 of the 11 antimicrobials tested as compared to isolates recovered from conventional beef. In the case of S. aureus, these differences were only found for 3 of the 11 antimicrobials tested and for L. monocytogenes, no differences were obtained between isolates obtained from organic or conventional beef. Although no significant differences were obtained in microbiological contamination, E. coli and S. aureus isolates from organically farmed beef samples showed significantly lower rates of antimicrobial resistance in E. coli and S. aureus isolates.

Comparison of antimicrobial resistance in Escherichia coli, Staphylococcus aureus, and Listeria monocytogenes strains isolated from organic and conventional poultry meat.

The presence of Escherichia coli, Staphylococcus aureus, and Listeria monocytogenes was determined in 55 samples of organic poultry meat and in 61 samples of conventional poultry meat. A total of 220 E. coli, 192 S. aureus, and 71 L. monocytogenes strains were analyzed by an agar disk diffusion assay for their resistance to ampicillin, cephalothin, chloramphenicol, ciprofloxacin, doxycycline, fosfomycin, gentamicin, nitrofurantoin, streptomycin, and sulfisoxazole (E. coli); chloramphenicol, ciprofloxacin, clindamycin, doxycycline, erythromycin, gentamicin, nitrofurantoin, oxacillin, and sulfisoxazole (S. aureus); and chloramphenicol, doxycycline, erythromycin, gentamicin, sulfisoxazole, and vancomycin (L. monocytogenes). The results indicated a significantly higher (P < 0.0001) prevalence of E. coli but not of S. aureus and L. monocytogenes in organic poultry meat as compared with conventional poultry meat. E. coli isolated from organic poultry meat exhibited lower levels of antimicrobial resistance against 7 of the 10 antimicrobials tested as compared with isolates recovered from conventional meat. In the case of S. aureus and L. monocytogenes isolated from conventional poultry, antimicrobial resistance was significantly higher only for doxycycline as compared with strains isolated from organic poultry. In the case of E. coli, the presence of multiresistant strains was significantly higher (P < 0.0001) in conventional poultry meat as compared with organic poultry meat. Organically farmed poultry samples showed significantly lower development of antimicrobial resistance in intestinal bacteria such as E. coli.

Evolution of resistance in poultry intestinal Escherichia coli during three commonly used antimicrobial therapeutic treatments in poultry.

The resistance rates of intestinal Escherichia coli populations from poultry were determined during treatment and withdrawal period with 3 antimicrobial agents commonly used as therapeutics in poultry medicine. A total of 108 chickens were considered: 18 were treated orally with enrofloxacin, 18 with doxycycline, and 18 with sulfonamides, whereas another 18 chickens were maintained as controls for each antimicrobial group. Fecal samples were taken during the treatment and after the withdrawal period, and E. coli were isolated through Fluorocult media plating. A total of 648 E. coli strains (216 per antimicrobial tested) were isolated and identified though biochemical methods. Minimal inhibitory concentrations to the antimicrobials used were also determined using a broth microdilution method. The resistance rates of intestinal E. coli to all of the antimicrobials tested significantly increased during the course of the therapeutic treatment. In addition, significant differences (P = 0.0136) in resistance rates persisted between the intestinal E. coli of the enrofloxacin-treated and control batches until the end of the withdrawal period, but this difference was not observed for the cases of doxycycline or sulfonamides treatments. Antimicrobial use in poultry medicine seems to select for antimicrobial-resistant strains of pathogenic bacterial species such as E. coli. In some cases, the higher frequencies of resistant strains may persist in the avian intestinal tract until the end of the withdrawal period, when it is legal to use these animals for human consumption.

Fatty acid composition of breast milk in Galicia (NW Spain): A cross-country comparison.

Based on the premise that the fatty acid composition of human milk can be substantially altered by diet, the current study investigated the fatty acid profile (fattyacidome) of breast milk in Galicia, a small region located in the north-west of Spain and characterized by the Southern European Atlantic Diet (SEAD). A cross-country comparison was also performed to assess worldwide variety and diet impact, reviewing the profiles reported various European, North and South American, Asian and African countries and Australia. Galician human milk appeared similar to the rest of Europe, with some particular features related to the SEAD (dairy, pork, beef and sunflower and olive oils), such as relatively high levels of linoleic acid and lower α-linolenic acid. The results also showed the existence of woman-specific profiles and significant changes over lactation in some fatty acids. Worldwide, the fatty acid profiles were similar, with the clear exception of Asiatic breast milk. The impact of fatty acids on infant health warrants further investigation.

In vitro evaluation of the prebiotic effect of red and white grape polyphenolic extracts.

Nowadays, the strong relationship between diet and health is well known. Although the primary role of diet is to provide nutrients to fulfill metabolic requirements, the use of foods to improve health and the state of well-being is an idea increasingly accepted by society in the last three decades. During the last years, an important number of scientific advances have been achieved in this field and, although in some situations, it is difficult to establish a distinction between "harmful" and "good" bacteria, experts agree in classifying the genera Bifidobacterium and Lactobacillus as beneficial bacteria. Thus, several strategies can be used to stimulate the proliferation of these beneficial intestinal bacteria, being one of them the consumption of prebiotics. The development of new prebiotics, with added functionality, is one of the most serious challenges shared not only by the scientific community but also by the food industry. The objective of this work was to evaluate the potential prebiotic effect of red and white grape residues, both obtained during the winemaking process. For such purpose, an in vitro study with pure cultures of Lactobacillus and Bifidobacterium was first conducted. Secondly, a study with mixed cultures using human fecal inocula was carried out in a simulator of the distal part of the colon. The obtained results showed an increase in the Lactobacillus and Bifidobacterium population, indicating that these ingredients are serious candidates to be considered as prebiotics.

Study of a room temperature phosphorescence phenomenon to allow the detection of aflatoxigenic strains in culture media.

A novel screening method based on room temperature phosphorescence (RTP) for the visual detection of aflatoxigenic strains from Aspergillus genus is described. Strains were cultured on media widely used in food mycology to which methyl-beta-cyclodextrin plus bile salts (0.6% sodium deoxycholate) were added. Aflatoxin production was readily detectable after 3 days of incubation at 28 degrees C by RTP emission from the mycelium of aflatoxigenic strains observed after exposure to UV light. The method was tested on thirty-two Aspergillus sp. strains. The phosphorescence phenomenon was reproduced in vitro by immobilizing aflatoxin B1 on ion exchange resin beads.

Liquid chromatography-electrospray ionization-mass spectrometry method in multiple reaction monitoring mode to determine 17alpha-ethynylestradiol residues in cattle hair without previous digestion.

A liquid chromatography-mass spectrometric (LC-MS/MS) method has been developed for determination of ethynylestradiol residues in cattle hair. Hair samples were pulverized with a cryogenic mill followed by a simple extraction with acetonitrile. A dansyl derivatization procedure to improve ethynylestradiol detection was used before the LC-MS/MS analysis in multiple reaction monitoring (MRM) mode using alpha-estradiol as an internal standard. The method was validated following the latest EU guidelines using blank hair samples spiked at 2 ng g(-1). The detection capability (CCbeta) was less than 2 ng g(-1), and the decision limit (CCalpha) was 1 ng g(-1). Incurred samples obtained 56 days after cow treatment with ethynylestradiol were analyzed, and the presence of ethynylestradiol in the hair was confirmed in all cases.

Antimicrobial resistance in Enterococcus spp. strains isolated from organic chicken, conventional chicken, and turkey meat: a comparative survey.

The mean counts of Enterococcus spp. were determined for 30 samples each of organic chicken meat, conventional chicken meat, and turkey meat, and differences for Enterococcus contamination in meat were determined. Two enterococci strains from each sample were isolated to obtain a total of 180 strains, and resistance to ampicillin, chloramphenicol, doxycycline, ciprofloxacin, erythromycin, gentamicin, nitrofurantoin, and vancomycin was determined by a disk diffusion method. Average counts obtained showed that Enterococcus mean counts from organic chicken meat (3.18 log CFU/g) were significantly higher than those obtained from conventional chicken meat (2.06 log CFU/g) or conventional turkey meat (1.23 log CFU/g). However, the resistance data obtained showed that isolates from organic chicken meat were less resistant than enterococci isolates from conventional chicken meat to ampicillin (P = 0.0067), chloramphenicol (P = 0.0154), doxycycline (P = 0.0277), ciprofloxacin (P = 0.0024), erythromycin (P = 0.0028), and vancomycin (P = 0.0241). In addition, isolates from organic chicken were less resistant than conventional turkey meat isolates to ciprofloxacin (P = 0.001) and erythromycin (P = 0.0137). Multidrug-resistant isolates were found in every group tested, but rates of multidrug-resistant strains were significantly higher in conventional chicken and turkey than those obtained from organic chicken meat. Enterococcus faecalis was the most common species isolated from organic chicken (36.67%), whereas Enterococcus durans was the most common species isolated from conventional chicken (58.33%) and turkey (56.67%). The rates obtained for antimicrobial resistance suggest that although organic chicken meat may have higher numbers of Enterococcus, these bacteria present a lower level of antimicrobial resistance.

Effects of bovine pregnancy on the fatty acid composition of milk: the significance for humans needs.

Milk from 40 Holstein dairy cows was collected from two different farms in Galicia (Spain). The differences in the fatty acid composition of two groups of cows, 20 pregnant and 20 non-pregnant, was studied to determine whether pregnancy status is a determinant factor that can alter the fatty acid profile of milk. Gas-chromatography (GC) coupled to flame ionisation detection (FID) was used for the determination of the fatty acids. Differences in the milk fatty acids between pregnant and non-pregnant cows were pronounced showing statistically significant differences for some fatty acids and the total saturated and monounsaturated fatty acids. Milk from non-pregnant cows was lower in saturated fatty acids and higher in monounsaturated fatty acids (unlike milk from pregnant cows). The effects of the consumption of bovine milk, particularly milk fat, on human health have been studied in depth and sometimes are associated with negative effects, but milk has also several beneficial characteristics linked to some fatty acids.