Antibiotic Resistance and Virulence Factors among Enterococci Isolated from Chourico, a Traditional Portuguese Dry Fermented Sausage

Enterococci are ubiquitous microorganisms, found as part of the normal intestinal microbiota of many animals. They can be present in food products, for example, the Portuguese dry fermented sausage chourico. Twenty enterococci were isolated from chourico in two processing units; after identification and typification by conventional-molecular methods, the isolates were screened for virulence factors and antibiotic resistance. Identification allocated all enterococci to the species Enterococcus faecalis, and PCR fingerprinting demonstrated that each isolate was specific to the processing unit and chourico from which it was recovered. Regarding the screening for virulence factors, I strain produced cytolysin and 4 were gelatinase positive, but none produced lipase. The ace gene was detected in I enterococci, ebpABC and efaA(fs). in 16 isolates each, esp in 3, fsrB in 5, gelE in 7, and cylA in I. A multiresistant phenotype was observed in 8 isolates, 6 belonging to factory A. The antibiotic resistance gene ere(B) was detected in 9 enterococci, whereas the genes tet(M), aac(6')-le-aph(2 ''), and vanA were detected in 8 isolates each. As some of the E. faecalis chourico isolates present a multiresistant profile and harbor virulence and/or resistance genes, to assess further the safety of Portuguese dry sausages, a larger number of products and processing units must by analyzed

Diversity of Staphylococcus aureus Isolates in European Wildlife

Staphylococcus aureus is a well-known colonizer and cause of infection among animals and it has been described from numerous domestic and wild animal species. The aim of the present study was to investigate the molecular epidemiology of S. aureus in a convenience sample of European wildlife and to review what previously has been observed in the subject field. 124 S. aureus isolates were collected from wildlife in Germany, Austria and Sweden; they were characterized by DNA microarray hybridization and, for isolates with novel hybridization patterns, by multilocus sequence typing (MLST). The isolates were assigned to 29 clonal complexes and singleton sequence types (CC1, CC5, CC6, CC7, CC8, CC9, CC12, CC15, CC22, CC25, CC30, CC49, CC59, CC88, CC97, CC130, CC133, CC398, ST425, CC599, CC692, CC707, ST890, CC1956, ST2425, CC2671, ST2691, CC2767 and ST2963), some of which (ST2425, ST2691, ST2963) were not described previously. Resistance rates in wildlife strains were rather low and mecA-MRSA isolates were rare (n = 6). mecC-MRSA (n = 8) were identified from a fox, a fallow deer, hares and hedgehogs. The common cattle- associated lineages CC479 and CC705 were not detected in wildlife in the present study while, in contrast, a third common cattle lineage, CC97, was found to be common among cervids. No Staphylococcus argenteus or Staphylococcus schweitzeri-like isolates were found. Systematic studies are required to monitor the possible transmission of human- and livestock- associated S. aureus/MRSA to wildlife and vice versa as well as the possible transmission, by unprotected contact to animals. The prevalence of S. aureus/MRSA in wildlife as well as its population structures in different wildlife host species warrants further investigation

Measurement of Low-level radiofrequency electromagnetic fields in the human environment

In recent years there has been an increase in development of electromagnetic (EM) technology in the telecommunication industry, resulting in an increase in human non-ionizing exposure. This fact has initiated a number of scientific studies on possible health effects of EM fields on human organism. Totally four representative microenvironments were investigated for RF EM fields distribution, namely: city center, residential area, rural area, and extra-village area. Each microenvironment was measured 20 times in accordance with the International Commission for Non-Ionizing Radiation Protection (ICNIRP) guidelines. The extra-village measurements were taken as the base values that reflect the E-field intensities with the lowest amplitudes. The statistical analysis revealed notable statistical significance (p < 0.001) in almost all measured frequency bands except the Wi-Fi where the p-values were less than 0.05 for the city center and residential area but not significant for rural area. The highest total E-field intensity was measured in the residential area (approximately 1.85 V/m). All measured values were below the legal limits of the Slovak Republic and ICNIRP safety guidelines. However, the ICNIRP safety limits were written in 1998 considering only the thermal effects of RF radiation. They were updated in 2009 without any changes in the limits and still recommend 27.5 – 61 V/m (2 – 10 W/m2) for the RF frequency band of 400–2,000 MHz. The BioInitiative Report of 2012 established the scientific benchmark for possible health risks as 30–60 μW/m2 (approximately 0.1 – 0.15 V/m). Thus, all measured values were above the scientifically derived limits

Microbial ecosystem of traditional dry fermented sausages in Mediterranean countries and Slovakia.

This study highlighted the wide diversity of the processes for the manufacturing of traditional dry sausages in the Mediterranean countries (Spain, Italy, France, Greece and Portugal) and East-Central Europe (Slovakia). In particular, the temperature range of the “fermentation” step was broad, from low temperature 2°C to high 26°C together with variable length (1-8 days). Also a wide range both in temperature (2-25°C) and length (5-90 days) was noticed for the ripening step. Statistical analyses revealed that Gram-positive catalase-positive cocci (GCC+), yeasts/moulds and Enterobacteriaceae discriminated the ripened sausages according to the geographical origins while Lactic Acid Bacteria (LAB) constituted the dominant bacteria of the ripened products in all the countries. LAB counts, which increased during the fermentation step and aw which decreases during ripening, discriminated the three manufacturing steps (batter, fermented, ripened). In general, traditional dry fermented sausages did not present health risks in general, although the presence of some pathogens and spoilage microbiota in some sausages highlighted the importance of maintaining sound hygienic procedures

Traditional dry fermented sausages produced in small-scale processing units in Mediterranean countries and Slovakia. 1. Microbial ecosystems of processing environments

Microbial ecosystems were surveyed in 314 environmental samples from 54 Southern and Eastern European small-scale processing units (PUs) manufacturing traditional dry fermented sausages. The residual microflora contaminating the surfaces and the equipment were analysed after cleaning and disinfection procedures. All the PU environments were colonised at various levels by spoilage and technological microflora with excessive contamination levels in some of the PUs. Sporadic contamination by pathogenic microflora was recorded. Salmonella and Listeria monocytogenes were detected in 4.8% and 6.7% of the samples, respectively, and Staphylococcus aureus was enumerated in 6.1% of the samples. Several critical points were identified, such as the machines for S. aureus and the tables and the knives for L. monocytogenes; this knowledge is crucial for the improvement of hygiene control systems in small and traditional meat processing industries. The variability of the residual contamination emphasized the different cleaning, disinfecting and manufacturing practices routinely followed by these small-scale processing units

Distribution of aminogenic activity among potential autochthonous starter cultures for dry fermented sausages

Different strategies in the reduction of biogenic amine accumulation during the manufacture of five European traditional fermented sausages were studied concerning sausage formulation, the increase of sugar in the Italian salame abruzzese reduced the accumulation of cadaverine up to 43%. However, the addition of sugar in the French saucisson did not show a significant amine reduction. The inoculation of a decarboxylase-negative autochthonous starter culture reduced the biogenic amine accumulation in a different manner depending on the species and strain(s). The highest reduction was achieved by Lactobacillus sakei used in the Greek aeros thasou, resulting in a total putrescine reduction and a significant decrease in tyramine (62%) and histamine (71%). In Portuguese chouriços cadaverine reduction was only of 45% when a single strain of Staphylococcus equorum was inoculated, whereas a single strain of L. sakei or a mixture of S. equorum yielded a 75% and 89% of reduction, respectively. In Spanish fuet, a combination of L. sakei CTC6626 plus S. xylosus CTC6013 had only a very slight effect on tyramine reduction (19%) in Spanish fuet, whereas L. sakei CTC494 plus S. xylosus CTC6013 was capable to reduce tyraminogenesis by nearly 50%, suggesting that L. sakei CTC494 was the strain responsible for the additional tyramine reduction

Biogenic amines in traditional fermented sausages produced inselected European countries

Aminogenesis in traditional fermented sausages produced in Europe was studied during manufacturing process taking into account technological, physico-chemical and microbial factors. Tyramine was the major amine, followed by putrescine and cadaverine, although the occurrence of di-amines was much more variable. By principal component analysis, relationships between aminogenesis and the country of origin, physico-chemical parameters, processing conditions and microbial counts, were not found, probably due to the high dispersion observed in those variables. Therefore, biogenic amines occurred irrespectively of physico-chemical changes and technological conditions applied for sausage manufacture. By cluster analysis, five groups of fermented sausages were identified on the basis of their quantitative and qualitative profile of total biogenic amine content. Group A included products from very low to low total amine content (from not detected to 150 mg/kg); group B, products with moderate levels (from 150 to 350 mg/kg) tyramine being the major amine; group C, also with moderate amine contents but cadaverine being the major amine; and groups D and E, comprising products with high (from 350 to 550 mg/kg) and very high (higher than 550 mg/kg) amine content, respectively. Samples with moderate, high or very high levels of biogenic amines could be considered as products of less quality, and their consumption could be unhealthy for sensitive individuals or for those under classical monoamine oxidase inhibitor drug therapy

Repeated Stress Exaggerates Lipopolysaccharide-Induced Inflammatory Response in the Rat Spleen

Spleen is an immune organ innervated with sympathetic nerves which together with adrenomedullary system control splenic immune functions. However, the mechanism by which prior stress exposure modulates the immune response induced by immunogenic challenge is not sufficiently clarified. Thus, the aim of this study was to investigate the effect of a single (2 h) and repeated (2 h daily for 7 days) immobilization stress (IMO) on the innate immune response in the spleen induced by lipopolysaccharide (LPS, 100 microg/kg). LPS elevated splenic levels of norepinephrine and epinephrine, while prior IMO prevented this response. LPS did not alter de novo production of catecholamines, however, prior IMO attenuated phenylethanolamine N-methyltransferase gene expression. Particularly repeated IMO exacerbated LPS-induced down-regulation of alpha1B- and beta1-adrenergic receptors (ARs), while enhanced alpha2A- and beta2-AR mRNAs. Elevated expression of inflammatory mediators (iNOS2, IL-1beta, IL-6, TNF-alpha, IL-10) was observed following LPS and repeated IMO again potentiated this effect. These changes were associated with enhanced Ly6C gene expression, a monocyte marker, and elevated MCP-1, GM-CSF, and CXCL1 mRNAs suggesting an increased recruitment of monocytes and neutrophils into the spleen. Additionally, we observed increased Bax/Bcl-1 mRNA ratio together with reduced B cell numbers in rats exposed to repeated IMO and treated with LPS but not in acutely stressed rats. Altogether, these data indicate that repeated stress via changes in CA levels and specific alpha- and beta-AR subtypes exaggerates the inflammatory response likely by recruiting peripheral monocytes and neutrophils to the spleen, resulting in the induction of apoptosis within this tissue, particularly in B cells. These changes may alter the splenic immune functions with potentially pathological consequences