Hygiene auditing in mass catering: a 4-year study in a university canteen.

OBJECTIVES: The outcomes of hygiene audits carried out two times per year were used to determine the correct execution of the procedures foreseen by the Hazard Analysis and Critical Control Points (HACCP) plan over 4 years (2013-2016) in a university canteen producing about 1200 meals a day. STUDY DESIGN: Critical analysis of hygiene audits. METHODS: Hygiene audits were carried out on the basis of a checklist divided into seven main items and subitems that covered all the production areas of the canteen. For each audit subitem, total percentage of inadequacy was calculated as the total number of negative answers (N) divided by the total number of answers (n = 8) collected in the period 2013-2016. RESULTS: The results showed a discontinuous trend among years. In more detail, the highest percentage of inadequacy was seen for food maintaining temperatures, thus highlighting management issues mainly related to time taken for food preparation. A relatively high level of inadequacy was also recorded for staff clothing and hygiene. CONCLUSIONS: The critical analysis of data emerged from the audits was useful to obtain an overview of improvements and emerging criticalities.

Salmonellosis associated with mass catering: a survey of European Union cases over a 15-year period.

Salmonella spp. is the causative agent of a foodborne disease called salmonellosis, which is the second most commonly reported gastrointestinal infection in the European Union (EU). Although over the years the annual number of cases of foodborne salmonellosis within the EU has decreased markedly, in 2014, a total of 88 715 confirmed cases were still reported by 28 EU Member States. The European Food Safety Authority reported that, after the household environment, the most frequent settings for the transmission of infection were catering services. As evidenced by the reviewed literature, which was published over the last 15 years (2000-2014), the most frequently reported causative agents were Salmonella Enteritidis and Salmonella Typhimurium serovars. These studies on outbreaks indicated the involvement of various facilities, including hospital restaurants, takeaways, ethnic restaurants, hotels, in-flight catering, one fast-food outlet and the restaurant of an amusement park. The most commonly reported sources of infection were eggs and/or egg-containing foods, followed by meat- and vegetable-based preparations. Epidemiological and microbiological studies allowed common risk factors to be identified, including the occurrence of cross-contamination between heat-treated foods and raw materials or improperly cleaned food-contact surfaces.

In vivo chronic nicotine exposure differentially and reversibly affects upregulation and stoichiometry of α4ß2 nicotinic receptors in cortex and thalamus.

Studies with heterologous expression systems have shown that the α4ß2 nicotinic acetylcholine receptor (nAChR) subtype can exist in two stoichiometries (with two [(α4)2(ß2)3] or three [(α4)3(ß2)2] copies of the α subunit in the receptor pentamer) which have different pharmacological and functional properties and are differently regulated by chronic nicotine treatment. However, the effects of nicotine treatment in vivo on native α4ß2 nAChR stoichiometry are not well known. We investigated in C57BL/6 mice the in vivo effect of 14-day chronic nicotine treatment and subsequent withdrawal, on the subunit expression and ß2/α4 subunit ratio of (3)H-epibatidine labeled α4ß2*-nAChR in total homogenates of cortex and thalamus. We found that in basal conditions the ratio of the ß2/α4 subunit in the cortex and thalamus is different indicating a higher proportion in receptors with (α4)2(ß2)3 subunit stoichiometry in the thalamus. For cortex exposure to chronic nicotine elicited an increase in receptor density measured by (3)H-epibatidine binding, an increase in the α4 and ß2 protein levels, and an increase in ß2/α4 subunit ratio, that indicates an increased proportion of receptors with the (α4)2(ß2)3 stoichiometry. For thalamus we did not find a significant increase in receptor density, α4 and ß2 protein levels, or changes in ß2/α4 subunit ratio. All the changes elicited by chronic nicotine in cortex were transient and returned to basal levels with an average half-life of 2.8 days following nicotine withdrawal. These data suggest that chronic nicotine exposure in vivo favors increased assembly of α4ß2 nAChR containing three ß2 subunits. A greater change in stoichiometry was observed for cortex (which has relatively low basal expression of (α4)2(ß2)3 nAChR) than in thalamus (which has a relatively high basal expression of (α4)2(ß2)3 nAChR).

Different physiological and behavioural effects of e-cigarette vapour and cigarette smoke in mice.

Nicotine is the primary addictive substance in tobacco smoke and electronic cigarette (e-cig) vapour. Methodological limitations have made it difficult to compare the role of the nicotine and non-nicotine constituents of tobacco smoke. The aim of this study was to compare the effects of traditional cigarette smoke and e-cig vapour containing the same amount of nicotine in male BALB/c mice exposed to the smoke of 21 cigarettes or e-cig vapour containing 16.8 mg of nicotine delivered by means of a mechanical ventilator for three 30-min sessions/day for seven weeks. One hour after the last session, half of the animals were sacrificed for neurochemical analysis, and the others underwent mecamylamine-precipitated or spontaneous withdrawal for the purposes of behavioural analysis. Chronic intermittent non-contingent, second-hand exposure to cigarette smoke or e-cig vapour led to similar brain cotinine and nicotine levels, similar urine cotinine levels and the similar up-regulation of α4ß2 nicotinic acetylcholine receptors in different brain areas, but had different effects on body weight, food intake, and the signs of mecamylamine-precipitated and spontaneous withdrawal episodic memory and emotional responses. The findings of this study demonstrate for the first time that e-cig vapour induces addiction-related neurochemical, physiological and behavioural alterations. The fact that inhaled cigarette smoke and e-cig vapour have partially different dependence-related effects indicates that compounds other than nicotine contribute to tobacco dependence.

Microbiological monitoring of air quality in a university canteen: an 11-year report.

Over the past decade, an increased tendency to consume meals at dining facilities outside the home has been highlighted; moreover, meals supplied in food businesses have been involved in many foodborne disease outbreaks. Therefore, microbial air contamination in food processing facilities could be a concern and an increase of microbial loads could represent a risk factor, especially for the potential contamination of foods due to undesirable spoiling and pathogenic bacteria. In this paper, the results of an 11-year microbiological monitoring of air quality in a university canteen are reported. The study, which started in the year 2000, was performed within a hazard analysis and critical control point (HACCP) plan implementation of a canteen that produces about 1,000 meals a day in order to verify the effectiveness of corrective actions on the indoor air quality. The primary food preparation room, the kitchen, and three cold rooms underwent air sampling by using a calibrated impaction sampler. Our investigation detected a general and progressive improvement in the air quality of the canteen since the beginning of the study, thus suggesting the appropriateness of the corrective action undertaken during the HACCP implementation program.

Bacterial dynamics in a raw cow's milk Caciotta cheese manufactured with aqueous extract of Cynara cardunculus dried flowers.

AIMS: To investigate the bacterial dynamics of a Caciotta cheese traditionally manufactured in the Montefeltro area (Central Italy) with raw cow's milk and an aqueous extract of dried flowers from Cynara cardunculus as a coagulating agent. METHODS AND RESULTS: Conventional methods and a combined PCR-DGGE approach, relying on culture-dependent and -independent analyses, were used to investigate the cheese bacterial community, with a special focus on lactic acid bacteria. A heterogeneous population, including enterococci, lactococci, lactobacilli, food spoilage and other banal micro-organisms, was found. SIGNIFICANCE AND IMPACT OF THE STUDY: The study contributed to highlighting the influence of different technological parameters on bacterial dynamics of a raw milk Caciotta cheese coagulated with vegetable rennet. CONCLUSIONS: None of the species found in the vegetable rennet became dominant during the cheese-making and a prevailing role of the adventitions microbita coming from the raw milk and the dairy environment was highlighted.

Characterisation of Lactobacillus helveticus strains producing antihypertensive peptides by RAPD and inverse-PCR of IS elements.

Lactobacillus helveticus is used for the manufacture of cheeses and milk-based products. Although it is not considered a probiotic microorganism, some strains demonstrated beneficial effects through the production of antihypertensive peptides from the hydrolysis of casein during milk fermentation. Strain-specificity of bioactive peptide production by L. helveticus makes the availability of reliable typing methods essential for both legal and good manufacturing processes. Accordingly, RAPD and inverse-PCR of five insertion sequence elements were comparatively evaluated for the molecular characterisation of four L. helveticus dairy cultures producing antihypertensive peptides and fourteen reference strains. Calculation of discriminatory indices and cluster analysis of the DNA fingerprints confirmed the suitability of both approaches for acceptable strain differentiation. Although RAPD was more discriminating, for a few test strains a neat discrimination was only achieved through multiplex inverse-PCR, thus suggesting the suitability of a combined analytical approach for a finer strain discrimination.

Paclitaxel eluting balloon: from bench to bedside.

Despite the impressive progress of percutaneous treatment modalities, restenosis remains the major Achilles heel of interventional cardiology. Approximately 25% of the general population treated for coronary diseases with a bare-metal stent and about 10% of patients treated with a drug-eluting stent develop an overgrowth of vascular tissue and renarrowing inside the stent, or in-stent restenosis. These rates are even greater in diabetics and patients at higher risk of restenosis both for clinical presentation (patients in dialysis, low ejection fraction) or anatomical characteristics (ostial, bifurcation, long lesions). Non-stent based local drug delivery and particularly the use of paclitaxel eluting balloon (PEB) could be one promising strategy to reduce restenosis. This review will briefly explore the different characteristics of PEB devices currently present in the market and summarize the results obtained both in animal models and clinical practice, giving an indication of the potential field of application of this new technology.

PCR-DGGE analysis of lactic acid bacteria and yeast dynamics during the production processes of three varieties of Panettone.

AIMS: To study lactic acid bacteria (LAB) and yeast dynamics during the production processes of sweet-leavened goods manufactured with type I sourdoughs. METHODS AND RESULTS: Fourteen sourdough and dough samples were taken from a baking company in central Italy during the production lines of three varieties of Panettone. The samples underwent pH measurements and plating analysis on three solid media. The microbial DNA was extracted from both the (sour)doughs and the viable LAB and yeast cells collected in bulk, and subjected to PCR-denaturing gradient gel electrophoresis (DGGE) analysis. The molecular fingerprinting of the cultivable plus noncultivable microbial populations provide evidence of the dominance of Lactobacillus sanfranciscensis, Lactobacillus brevis and Candida humilis in the three fermentation processes. The DGGE profiles of the cultivable communities reveal a bacterial shift in the final stages of two of the production processes, suggesting an effect of technological parameters on the selection of the dough microflora. CONCLUSIONS: Our findings confirm the importance of using a combined analytical approach to explore microbial communities that develop during the leavening process of sweet-leavened goods. SIGNIFICANCE AND IMPACT OF THE STUDY: In-depth studies of sourdough biodiversity and population dynamics occurring during sourdough fermentation are fundamental for the control of the leavening process and the manufacture of standardized, high-quality products.

Phenotypic, genotypic and technological characterization of predominant lactic acid bacteria in Pecorino cheese from central Italy.

AIMS: Identification and biotyping of lactic acid bacteria (LAB) isolated from raw-milk Pecorino cheese manufactured in the Marche region (central Italy) for selection of suitable starter cultures or adjuncts. METHODS AND RESULTS: Preliminary characterization with morphological and biochemical assays were undertaken for 112 Gram-positive and catalase-negative isolates. Unequivocal identification of the isolates was obtained through restriction analysis of the amplified 16S rRNA gene and sequencing of 360-380 bp amplicons. Fifty-nine isolates belonging to LAB species generally recognized as safe and potentially utilized as starters or flavour-producing adjuncts were preselected and tested for their acidifying, proteolitic and autolytic activities. Fifty-five of these isolates were also subject to RAPD (randomly amplified polymorphic DNA) fingerprinting and unweighted pair-group method with arithmetic averages (UPGMA) cluster analysis for the estimation of genotypic intra-species variation. As a result, in Pecorino cheese, a heterogeneous lactic acid bacteria population, which includes strains with metabolic characteristics of technological interest, was characterized. CONCLUSIONS: The polyphasic approach proposed allows the bacterial ecology of Pecorino cheese to be investigated and allows to assess the potential role of autochthonous LAB strains for the dairy industry. SIGNIFICANCE AND IMPACT OF THE STUDY: The great economic importance of Pecorino cheese encouraged a deeper knowledge of its microbiota, which is known to influence the peculiar sensory properties of this cheese, also in view of its exploitation.

Resident lactic acid bacteria in raw milk Canestrato Pugliese cheese.

AIMS: Investigation of the autochthonous lactic acid bacteria (LAB) population of the raw milk protected designation of origin Canestrato Pugliese cheese using phenotypic and genotypic methodologies. METHODS AND RESULTS: Thirty phenotypic assays and three molecular techniques (restriction fragment length polymorphism, partial sequencing of the 16S rRNA gene and recA multiplex PCR assay) were applied to the identification of 304 isolates from raw milk Canestrato Pugliese cheese. As a result, 168 of 207 isolates identified were ascribed to genus Enterococcus, 25 to Lactobacillus, 13 to Lactococcus and one to Leuconostoc. More in details among the lactobacilli, the species Lactobacillus brevis and Lactobacillus plantarum were predominant, including 13 and 10 isolates respectively, whereas among the lactococci, Lactococcus lactis subsp.cremoris [corrected] was the species more frequently detected (seven isolates). CONCLUSIONS: Except for the enterococci, phenotypic tests were not reliable enough for the identification of the isolates, if not combined to the genotype-based molecular techniques. The polyphasic approach utilized allowed 10 different LAB species to be detected; thus suggesting the appreciable LAB diversity of the autochthonous microbial population of the Canestrato Pugliese cheese. SIGNIFICANCE AND IMPACT OF THE STUDY: A comprehensive study of the resident raw milk Canestrato Pugliese cheese microbial population has been undertaken.

Brain neuronal nicotinic receptors as new targets for drug discovery.

Neuronal nicotinic receptors (nAChRs) are a heterogeneous family of ion channels differently expressed in the nervous system where, by responding to the endogenous neurotransmitter acetylcholine, they contribute to a wide range of brain activities and influence a number of physiological functions. Over recent years, the application of newly developed molecular and cellular biological techniques has made it possible to correlate the subunit composition of nAChRs with specific nicotine-elicited behaviours, and refine some of the in vivo physiological functions of nAChR subtypes. The major new findings are the widespread expression of nAChRs, outside the nervous system, their specific and complex organisation, and their relevance to normal brain function. Moreover, the combination of clinical and basic research has better defined the involvement of nAChRs in a growing number of nervous pathologies other than degenerative diseases. However, there are still only a limited number of nicotinic-specific drugs and, although some nicotinic agonists have an interesting pharmacology, their clinical use is limited by undesirable side effects. Some selective nicotinic ligands have recently been developed and used to explore the complexity of nAChR subtype structure and function in the expectation that they will become rational therapeutic alternatives in a number of neurodegenerative, neuropsychiatric and neurological disorders. In this review, we will discuss the molecular basis of brain nAChR structural and functional diversity mainly in pharmacological and biochemical terms, and summarise current knowledge concerning the newly discovered drugs used to classify the numerous receptor subtypes and treat the brain diseases in which nAChRs are involved.

Effect of the implementation of HACCP on the microbiological quality of meals at a university restaurant.

A study was conducted to evaluate the microbiological quality, including total mesophilic counts and markers of bacteriological hygiene, as indicator of food safety of three categories of the most consumed meals in a university restaurant, before and after implementation of the HACCP system and personnel training. Cold gastronomy products, cooked warm-served products, and cooked cold-served products were tested for bacterial contamination. Throughout the experiment, 894 samples were examined for total counts of aerobic bacteria, counts of indicator organisms (coliform organisms and Escherichia coli) and pathogens (Staphylococcus aureus, Bacillus cereus, Salmonella spp., and Listeria monocytogenes). Implementation of the HACCP system, together with training in personnel hygiene, good manufacturing practices, and cleaning and sanitation procedures, resulted in lower aerobic plate counts and a lower incidence of S. aureus, coliform organisms, E. coli, and B. cereus, whereas Salmonella spp. and L. monocytogenes were not found in all samples studied. The microbial results of this study demonstrate that personnel training together with HACCP application contributed to improve the food safety of meals served in the restaurant studied.

Interactions between Saccharomyces cerevisiae and malolactic bacteria: preliminary characterization of a yeast proteinaceous compound(s) active against Oenococcus oeni.

AIMS: To investigate the occurrence and extent of Saccharomyces cerevisiae and Oenococcus oeni interactions. METHODS AND RESULTS: Interactions between S. cerevisiae and O. oeni were investigated by double-layer and well-plate assays showing the occurrence of specific interactions for each yeast-malolactic bacteria (MLB) coupling. Heat and protease treatments of synthetic grape juice fermented by the S. cerevisiae strain F63 indicated that the inhibitory activity exerted by this yeast on O. oeni is due to a proteinaceous factor(s) which exerts either bacteriostatic or bactericidal effect depending on concentration and affects malolactic fermentation in natural grape juice and wine. CONCLUSIONS: A proteinaceous factor(s) produced by a S. cerevisiae wine strain able to inhibit O. oeni growth and malic acid fermentation was characterized. SIGNIFICANCE AND IMPACT OF THE STUDY: The individuation, characterization and exploitation of yeast proteinaceous factor(s) exerting inhibitory activity on MLB may offer new opportunities for the management of malolactic fermentation.

Cholinergic nicotinic receptor involvement in movement disorders associated with Lewy body diseases. An autoradiography study using [(125)I]alpha-conotoxinMII in the striatum and thalamus.

The presence of alpha6 subunit containing nicotinic acetylcholine receptors on nigrostriatal dopaminergic neurons has been demonstrated in rodents and monkeys. [(125)I]alpha-conotoxinMII is a radioligand that binds to alpha6, and also alpha3 subunits of nicotinic acetylcholine receptors (nAChRs). In the present study, we have compared the distribution of [(125)I]alpha-conotoxinMII binding in post mortem human tissue from four groups of patients: individuals with dementia with Lewy bodies displaying extra-pyramidal features (DLB + EPF), DLB without extra-pyramidal features (DLB - EPF) Parkinson's disease without dementia (PD) and age-matched controls. Reduced binding was observed in the putamen and caudate in PD and both DLB groups. In DLB patients, the decline was greater in DLB + EPF compared to DLB - EPF group. The declines in nicotinic receptor binding in the striatum were in part paralleled by reductions in the striatal dopamine transporter. In the thalamus, [(125)I]alpha-conotoxinMII binding was significantly reduced in the centromedian nucleus in both DLB groups, and also in the parafascicular nucleus in the DLB - EPF group. In DLB + EPF and PD patients, there was decreased binding in the ventral lateral nucleus. This study demonstrates alterations of alpha6 and/or alpha3 nAChRs binding in DLB and PD, which are likely to relate to extra-pyramidal symptoms.

Neuronal nicotinic receptors: from structure to pathology.

Neuronal nicotinic receptors (NAChRs) form a heterogeneous family of ion channels that are differently expressed in many regions of the central nervous system (CNS) and peripheral nervous system. These different receptor subtypes, which have characteristic pharmacological and biophysical properties, have a pentameric structure consisting of the homomeric or heteromeric combination of 12 different subunits (alpha2-alpha10, beta2-beta4). By responding to the endogenous neurotransmitter acetylcholine, NAChRs contribute to a wide range of brain activities and influence a number of physiological functions. Furthermore, it is becoming evident that the perturbation of cholinergic nicotinic neurotransmission can lead to various diseases involving nAChR dysfunction during development, adulthood and ageing. In recent years, it has been discovered that NAChRs are present in a number of non-neuronal cells where they play a significant functional role and are the pathogenetic targets in several diseases. NAChRs are also the target of natural ligands and toxins including nicotine (Nic), the most widespread drug of abuse. This review will attempt to survey the major achievements reached in the study of the structure and function of NAChRs by examining their regional and cellular localisation and the molecular basis of their functional diversity mainly in pharmacological and biochemical terms. The recent availability of mice with the genetic ablation of single or double nicotinic subunits or point mutations have shed light on the role of nAChRs in major physiological functions, and we will here discuss recent data relating to their behavioural phenotypes. Finally, the role of NAChRs in disease will be considered in some details.

Developmental expression of heteromeric nicotinic receptor subtypes in chick retina.

Acting through nicotinic acetylcholine receptors (nAChRs), acetylcholine plays an important role in retinal development and the formation of retinal connections to target tissues, but very little is known about the nAChR subtypes expressed in vertebrate retina during neuronal development. We used immunoprecipitation and [3H]epibatidine binding to study the expression of chick retina alpha-bungarotoxin-insensitive heteromeric nAChRs during development and adulthood, and found that it is strictly developmentally regulated, reaching a peak on postnatal day 1. The increase in [3H]epibatidine receptors is caused mainly by an increase in the receptors containing the alpha2, alpha6, beta3, and beta4 subunits. The contribution of beta subunits to [3H]epibatidine receptors significantly changes during development: the beta2 subunit is contained in the majority (84%) of receptors on embryonic day (E) 7 but in only 32% on postnatal day (P) 1, whereas the beta4-containing receptors increase from 22% to 78% during the same period. Using a sequential immunodepletion procedure, we purified the beta2- and beta4-containing subtypes and found that they coassemble with alpha4 and/or alpha3 on E11, and also with the alpha2, alpha6, and beta3 on P1. After the immunodepletion of alpha6-containing receptors, the beta2- and beta4-containing receptors have a very similar pharmacological profile on P1. Parallel immunoprecipitation experiments in other brain areas showed that the developmentally regulated receptors in optic lobe are those containing the alpha2, alpha5, and beta2 subunits and those containing the alpha4 and beta2 subunits, whereas the receptors in forebrain-cerebellum contain the alpha4 and beta2 subunits with or without the alpha5 subunit. These results indicate that there is an increase in receptor heterogeneity and complexity in chick retina during development that is also maintained in adulthood.

Sp proteins and Phox2b regulate the expression of the human Phox2a gene.

Phox2a is a vertebrate homeodomain transcription factor that is involved in the specification of the autonomic nervous system. We have isolated the 5' regulatory region of the human Phox2a gene and studied the transcriptional mechanisms underlying its expression. We first identified the minimal gene promoter by means of molecular and functional criteria and demonstrated that its activity relies on a degenerate TATA box and a canonical Sp1 site. We then concentrated on the region immediately upstream of the promoter and found that it stimulates transcription in a neurospecific manner because its deletion caused a substantial decline in reporter gene expression only in neuronal cells. This DNA region contains a putative binding site for homeodomain transcription factors, and its mutation severely affects the transcriptional activity of the entire 5' regulatory region, thus indicating that this site is necessary for the expression of Phox2a in this cellular context. The use of the electrophoretic mobility shift assay showed that Phox2b/PMX2b is capable of specifically interacting with this site, and cotransfection experiments demonstrated that it is capable of transactivating the human Phox2a promoter. Many data obtained from knock-out mice support the hypothesis that Phox2a acts downstream of Phox2b during the development of most of the autonomic nervous system. We have provided the first molecular evidence that Phox2b can regulate the expression of Phox2a by directly binding to its 5' regulatory region.