Detection of Listeria monocytogenes in ready-to-eat food by Step One real-time polymerase chain reaction

The aim of this study was to follow contamination of ready-to-eat food with Listeria monocytogenes by using the Step One real time polymerase chain reaction (PCR). We used the PrepSEQ Rapid Spin Sample Preparation Kit for isolation of DNA and MicroSEQ® Listeria monocytogenes Detection Kit for the real-time PCR performance. In 30 samples of ready-to-eat milk and meat products without incubation we detected strains of Listeria monocytogenes in five samples (swabs). Internal positive control (IPC) was positive in all samples. Our results indicated that the real-time PCR assay developed in this study could sensitively detect Listeria monocytogenes in ready-to-eat food without incubation.     

The StepOne real-time polymerase chain reaction detection of Salmonella sp., Salmonella enterica ser. typhimurium and enteritidis in milk and meat

The aim of this study was to follow contamination of ready to eat milk and meat products with Salmonella spp. by using the StepOne real-time polymerase chain reaction (PCR). Classical microbiological methods for detection of foodborne bacteria involve the use of pre-enrichment and/or specific enrichment, following isolation of bacteria in solid media and the final confirmation by biochemical and/or serological tests. We used the PrepSEQ Rapid Spin Sample Preparation Kit for isolation of DNA and MicroSEQ? Salmonella spp. Detection Kit for pursuance of the real-time PCR (Applied Biosystems). In samples without incubation we detected strain of Salmonella sp. in 5 out of 25 samples (swabs), as well as in the internal positive control (IPC), which was positive in all samples. This StepOne real-time PCR assay is extremely useful for any laboratory equipped by real-time PCR. It is a fast, reproducible, simple, specific and sensitive way to detect nucleic acids, which could be used in clinical diagnostic tests in the future. Our results indicated that real-time PCR assay developed in this study could sensitively detect Salmonella spp. in ready-to-eat food. This could prevent infection caused by Salmonella, and also could benefit food manufacturing companies by extending their product's shelf-life as well as saving the cost of warehousing their food products while awaiting pathogen testing results.     

The StepOne real-time polymerase chain reaction detection of Salmonella sp., Salmonella enterica ser. typhimurium and enteritidis in milk and meat

The aim of this study was to follow contamination of ready to eat milk and meat products with Salmonella spp. by using the StepOne real-time polymerase chain reaction (PCR). Classical microbiological methods for detection of foodborne bacteria involve the use of pre-enrichment and/or specific enrichment, following isolation of bacteria in solid media and the final confirmation by biochemical and/or serological tests. We used the PrepSEQ Rapid Spin Sample Preparation Kit for isolation of DNA and MicroSEQ® Salmonella spp. Detection Kit for pursuance of the real-time PCR (Applied Biosystems). In samples without incubation we detected strain of Salmonella sp. in 5 out of 25 samples (swabs), as well as in the internal positive control (IPC), which was positive in all samples. This StepOne real-time PCR assay is extremely useful for any laboratory equipped by real-time PCR. It is a fast, reproducible, simple, specific and sensitive way to detect nucleic acids, which could be used in clinical diagnostic tests in the future. Our results indicated that real-time PCR assay developed in this study could sensitively detect Salmonella spp. in ready-to-eat food. This could prevent infection caused by Salmonella, and also could benefit food manufacturing companies by extending their product's shelf-life as well as saving the cost of warehousing their food products while awaiting pathogen testing results.     

Bactericidal effect of TiO2 photocatalyst on selected food-borne pathogenic bacteria

Titanium dioxide (TiO(2)) photocatalysts have attracted great attention as a material for photocatalytic sterilization in the food and environmental industry. This research aimed to design a new photobioreactor and its application to sterilize selected food borne pathogenic bacteria, Salmonella choleraesuis subsp., Vibrio parahaemolyticus, and Listeria monocytogenes. The photocatalytic reaction was carried out with various TiO(2) concentrations and Ultraviolet (UV) illumination time. A feasible synergistic effect was found that the bactericidal effect of TiO(2) on all bacterial suspension after UV light irradiation was much higher than that of without TiO(2). As the concentration of TiO(2) increased to 1.0 mg/ml, bactericidal effect increased. However, the bactericidal effect was rapidly abbreviated at TiO(2) concentration higher than 1.25 mg/ml to all selected bacteria. UV illumination time affected drastically the viability of all bacteria with different death rate. Similar trends were obtained from S. choleraesuis subsp. and V. parahaemolyticus that their complete killing was achieved after 3 h of illumination. However, L. monocytogenes was more resistant and its death ratio was about 87% at that time.     

Epidemiological study on Listeria monocytogenes in Egyptian dairy cattle farms’ insights into genetic diversity of multi-antibiotic-resistant strains by ERIC-PCR

Environmental Science and Pollution Research - Listeria monocytogenes (L. monocytogenes) is frequently detected in ruminants, especially dairy cattle, and associated with the sporadic and epidemic...     

Studies on the drug resistance profile of Enterococcus faecium distributed from poultry retailers to hospitals

The multidrug resistant Enterococcus faecium (MEF) strains originating from farm animals are proliferating at a substantial pace to impact downstream food chains and could reach hospitals. This study was conducted to elucidate the drug susceptibility profile of MEF strains collected from poultry products in Ann Arbor, MI area and clinical settings from Michigan State Lab and Moffitt Cancer Center (MCC) in Florida. Presumptive positive Enterococcus isolates at species level were identified by Matrix Assisted Laser Desorption/Ionization Time-of-Flight (MALDI-TOF) analysis. The antibiotic susceptibility profile for both poultry and clinical strains was determined by the Thermo Scientific's Sensititre conform to the National Committee for Clinical Laboratory Standards (NCCLS) and validated via quantitative real-time PCR (qPCR) methods. Out of 50 poultry samples (Turkey: n = 30; Chicken: n = 20), 36 samples were positive for Enterococcus species from which 20.83% were identified as E. faecium. All the E. faecium isolates were multidrug resistant and displayed resistance to the last alternative drug, quinupristin/dalfopristin (QD) used to treat vancomycin resistant E. faecium (VRE) in hospitals. Results indicate the presence of MEF strains in food animals and clinical settings that are also resistant to QD.     

Industrial processing versus home cooking: an environmental comparison between three ways to prepare a meal

Today there is a strong trend in Sweden for industrially processed meals to replace homemade meals. In the public debate this is often claimed to increase the environmental impact from foods. In the study presented in this article, we used life-cycle assessment to quantify the environmental impact of three meals: homemade, semiprepared, and ready-to-eat. The differences in environmental impact between the meals were small; the ready-to-eat meal used the most energy, whereas the homemade meal had higher emissions causing eutrophication and global warming. The dominating contributor to the environmental impact was agriculture, accounting for 30%, of the impact related to energy and 95% of that related to eutrophication. Industry, packaging, and consumer home transport and food preparation also contributed significantly. Important factors were raw material use, energy efficiency in industry and households, packaging, and residue treatment. To decrease the overall environmental impact of food consumption, improvements in agriculture are very important, together with raw-material use within industry and households.     

Sequestration of maize crop straw C in different soils: role of oxyhydrates in chemical binding and stabilization as recalcitrance

While biophysical controls on the sequestration capacity of soils have been well addressed with physical protection, chemical binding and stabilization processes as well as microbial community changes, the role of chemical binding and stabilization has not yet well characterized for soil organic carbon (SOC) sequestration in rice paddies. In this study, a 6-month laboratory incubation with and without maize straw amendment (MSA) was conducted using topsoil samples from soils with different clay mineralogy and free oxy-hydrate contents collected across Southern China. The increase in SOC under MSA was found coincident with that in Fe- and Al-bound OC (Fe/Al-OC) after incubation for 30 d (R(2)=0.90, P=0.05), and with sodium dithionate-citrate-bicarbonate (DCB) extractable Fe after incubation for 180 d (R(2)=0.99, P<0.01). The increase in SOC under MSA was found higher in soils rich in DCB extractable Fe than those poor in DCB extractable Fe. The greater SOC sequestration in soils rich in DCB extractable Fe was further supported by the higher abundance of (13)C which was a natural signature of MSA. Moreover, a weak positive correlation of the increased SOC under MSA with the increased humin (R(2)=0.87, P=0.06) observed after incubation for 180 d may indicate a chemical stabilization of sequestered SOC as humin in the long run. These results improved our understanding of SOC sequestration in China's rice paddies that involves an initial chemical binding of amended C and a final stabilization as recalcitrant C of humin.     

Polychlorinated biphenyls (PCBs), polybrominated diphenyl ethers (PBDEs) and perfluorinated alkylated substances (PFASs) in traditional seafood items from western Greenland

In this study, contamination levels were determined for polychlorinated biphenyls (PCBs), polybrominated diphenyl ethers (PBDEs) and perfluorinated alkylated substances (PFASs) in traditional Greenland seafood items, such as raw and smoked fish fillet (salmon and halibut), whale and seal meat and narwhal mattak (skin and blubber). The daily intake of PCBs, PBDEs and PFASs through traditional seafood items in Greenland was assessed. Based on the presented levels of contaminants, in combination with earlier food intake studies, suggests that the daily exposure was below the tolerable daily intake threshold for all compounds. BDE-47 was the only PBDE-congener detected in all food items, except in smoked halibut. The levels of BDE-47 varied from < LOD in smoked halibut up to 18 ng/g lw in narwhal mattak and 21 ng/g lw in whale beef. ∑PCB were lowest in smoked halibut (37 ng/g lw) and highest in narwhal mattak with 1,146 ng/g lw. Perfluorooctane sulfonate (PFOS) was the most common of the PFASs. However, ΣPFASs were below detection limits in most fish fillet samples, and varied from 2.9 ng/g ww in whale beef to 13.5 ng/g ww in seal beef. The present study shows that the exclusion from the diet of local food items such as intestines and blubber have a strong positive effect for the reduction of POPs levels in food, without a reducing the health benefits of traditional food intake considerably.     

Identification and quantitative detection of Legionella spp. in various aquatic environments by real-time PCR assay

In this study, a SYBR green quantitative real-time PCR was developed to quantify and detect the Legionella spp. in various environmental water samples. The water samples were taken from watershed, water treatment plant, and thermal spring area in Taiwan. Legionella was detected in 13.6 % (24/176), and the detection rate for river water, raw drinking water, and thermal spring water was 10, 21.4, and 16.6 %, respectively. Using real-time PCR, concentration of Legionella spp. in detected samples ranged between 9.75?×?10⁴ and 3.47?×?10⁵?cells/L in river water, 6.92?×?10⁴ and 4.29?×?10⁵?cells/L in raw drinking water, and 5.71?×?10⁴ and 2.12?×?10⁶?cells/L for thermal spring water samples. The identified species included Legionella pneumophila (20.8 %), Legionella jordanis (4.2 %), Legionella nautarum (4.2 %), Legionella sp. (4.2 %), and uncultured Legionella sp. (66.6 %). The presence of L. pneumophila in aquatic environments suggested a potential public health threat that must be further examined.     

Listeria monocytogenes batch culture growth response to metabolic inhibitors

In certain environments nutrient and energy sources available to microorganisms can be limited. Foodborne pathogens must efficiently adapt in order to be successfully transmitted through the food chain to their hosts. For the intracellular foodborne pathogen Listeria monocytogenes, little is known regarding its response to nutrient/energy-limiting conditions. The alternative stress responsive sigma factor σ^B has been reported to contribute to survival under specific stresses. Therefore, the effects of several metabolic inhibitors on growth of L. monocytogenes wild-type and a ΔsigB mutant were examined. In the absence of inhibitors, both strains reached stationary phase after 18 h at 23°C and 10 h at 37°C. All of the metabolic inhibitors slowed growth of either strain, with few differences observed among the different inhibitors.     

Perfluoroalkyl substances (PFASs) in food and water from Faroe Islands

Diet and drinking water are suggested to be major exposure pathways for perfluoroalkyl substances (PFASs). In this study, food items and water from Faroe Islands sampled in 2011/2012 were analyzed for 11 perfluoroalkyl carboxylic acids (PFCAs) and 4 perfluoroalkane sulfonic acids (PFSAs). The food samples included milk, yoghurt, crème fraiche, potatoes, fish, and fish feed, and the water samples included surface water and purified drinking water. In total, nine PFCAs and four PFSAs were detected. Generally, the levels of PFAS were in the lower picogram per gram range. Perfluorobutanoic acid was a major contributor to the total PFASs concentration in water samples and had a mean concentration of 750 pg/L. Perfluoroundecanoic acid (PFUnDA) was predominating in milk and wild fish with mean concentrations of 170 pg/g. Perfluorooctane sulfonic acid (PFOS) was most frequently detected in food items followed by PFUnDA, perfluorononanoic acid, and perfluorooctanoic acid (PFOA). Levels of PFUnDA and PFOA exceeded those of PFOS in milk and fish samples. Prevalence of long-chain PFCAs in Faroese food items and water is confirming earlier observations of their increase in Arctic biota. Predominance of short-chain and long-chain homologues indicates exposure from PFOS and PFOA replacement compounds.     

Polychlorinated biphenyl (PCB) anaerobic degradation in marine sediments: microcosm study and role of autochthonous microbial communities

Polychlorobiphenyl (PCB) biodegradation was followed for 1 year in microcosms containing marine sediments collected from Mar Piccolo (Taranto, Italy) chronically contaminated by this class of hazardous compounds. The microcosms were performed under strictly anaerobic conditions with or without the addition of Dehalococcoides mccartyi, the main microorganism known to degrade PCBs through the anaerobic reductive dechlorination process. Thirty PCB congeners were monitored during the experiments revealing that the biodegradation occurred in all microcosms with a decrease in hepta-, hexa-, and penta-chlorobiphenyls (CBs) and a parallel increase in low chlorinated PCBs (tri-CBs and tetra-CBs). The concentrations of the most representative congeners detected in the original sediment, such as 245-245-CB and 2345-245-CB, and of the mixture 2356-34-CB+234-245-CB, decreased by 32.5, 23.8, and 46.7 %, respectively, after only 70 days of anaerobic incubation without any bioaugmentation treatment. Additionally, the structure and population dynamics of the microbial key players involved in the biodegradative process and of the entire mixed microbial community were accurately defined by Catalyzed Reporter Deposition Fluorescence In Situ Hybridization (CARD-FISH) in both the original sediment and during the operation of the microcosm. The reductive dehalogenase genes of D. mccartyi, specifically involved in PCB dechlorination, were also quantified using real-time PCR (qPCR). Our results demonstrated that the autochthonous microbial community living in the marine sediment, including D. mccartyi (6.32E+06 16S rRNA gene copy numbers g^?1 sediment), was able to efficiently sustain the biodegradation of PCBs when controlled anaerobic conditions were imposed.     

Detection of veterinary drug residues in surface waters collected nearby farming areas in Galicia,North of Spain

The occurrence of pharmaceuticals in the aquatic environment has become a matter of concern in the last decade due to potential risks posed to non-target organisms and the potential for unintended human exposure via food chain. This concern has been driven by a high detection frequency for drugs in environmental samples; these substances are produced in large quantities and are used in both veterinary and human medicine, leading to deposition and potential effects in the environment. However, few studies have focused on the presence of pharmaceuticals in rural areas associated with farming activities in comparison to urban areas. The aim of this study is to investigate the occurrence of pharmaceutically active compounds in surface waters collected from urban and rural areas in northwestern Spain. A monitoring study was conducted with 312 river water samples analysed by high-performance liquid chromatography coupled to tandem mass spectrometry. Positive detection of pharmaceuticals was made for 51 % of the samples. Decoquinate, sulfamethazine, sulfamethoxypyridazine and trimethoprim were the drugs most frequently detected, being present in more than 10 % of the samples. The sampling sites located downstream of the discharge points for wastewater treatment plants yielded the highest number of positive samples, 13 % of the positive samples were detected in these sites and 38 % of the samples collected near the collection point of a drinking water treatment plant were positive.     

Molecular probes for the detection and identification of ichthyotoxic marine microalgae of the genus Pseudochattonella (Dictyochophyceae, Ochrophyta)

Phytoflagellates of the genus Pseudochattonella (Dictyochophyceae, Ochrophyta) form blooms in marine coastal waters in northern Europe, Japan, and New Zealand that at times cause fish kills with severe losses for the aquaculture industry. The aim of this study was to develop molecular probes for the detection and identification of Pseudochattonella at the genus and species level. A variety of probes were developed and applied to either dot blot hybridization, (q)PCR, or microarray format. In the dot blot hybridization assay, five different oligonucleotide probes targeting the small subunit (SSU) rDNA were tested against DNA from 18 microalgal strains and shown to be specific to the genus Pseudochattonella. A genus-specific PCR assay was developed by identifying an appropriate primer pair in the SSU—internal transcribed spacer 1 (ITS1) rDNA region. Its specificity was tested by screening against both target and non-target strains, and the assay was used to confirm the presence or absence of Pseudochattonella species in environmental samples. In order to distinguish between the two species of the genus, two PCR primer pairs each biased towards one of the species were designed in the large subunit (LSU) rDNA D1 domain and used for quantitative real-time PCR. Five selected probes (three SSU and two LSU rDNA) were adapted for the use on microarrays and included on a prototype multi-species microarray for the detection of harmful algae (http://www.midtal.com). Finally, microarrays and qPCR were used for the monthly monitoring of a sampling site in outer Oslofjorden during a 1-year period. Members of Pseudochattonella are difficult to identify by light microscopy in Lugol’s preserved samples, and the two species Pseudochattonella verruculosa and Pseudochattonella farcimen can be morphologically distinguished only by transmission electron microscopy. The molecular probes designed in this study will be a valuable asset to microscopical detection methods in the monitoring of harmful algae and for biogeographical and ecological studies of this genus.     

Use of dimethyl sulfoxide as solubilization agent in the detection of 2,3,7,8-TCDD by radioimmunoassay

Immunoassays are potentially valuable tools for use in screening environmental samples for a broad range of contaminants, such as polychlorinated dibenzo-p-dioxins (PCDDs). The performance of the radioimmunoassay (RIA) for PCDDs was characterized using 4 solubilization systems: Cutscum, Triton, horse serum, and dimethyl sulfoxide (DMSO). The DMSO based assay appeared to perform best at low 2,3,7,8-TCDD levels. The effects of assay incubation time and hapten storage conditions on the DMSO based assay were assessed. The separation of bound from unbound radioactivity was accelerated without adversely affecting assay performance. Further assay development through the use of an alternative labelled hapten is considered.     

Novel method for determining pyrene biodegradation using synchronous fluorimetry

To study the biodegradation rate of pyrene dissolved in liquid medium supplemented with mineral salts, a synchronous fluorimetry (SF) method was established. The limit of detection for pyrene dissolved in mineral salts medium (MSM) was determined as 0.19 ng/ml with a relative standard deviation of less than 1.3% (n = 9). The pyrene degrading rates of four bacterial strains were investigated using this method under the same experimental conditions. The degradation rates of the three active strains ranged from 76% to 87% after a 14-h incubation. The results were confirmed by the gas chromatography with a flame ionized detector (GC/FID) method. This implies that pyrene degradation can be directly monitored by the SF method without the solvent extraction of samples. The advantages of SF are that it is less laborious, faster, and less expensive than the GC/FID determination method with solvent extraction. The SF method provides a new tool for studying the degradation of polynuclear aromatic hydrocarbons (PAHs) in the natural environment and under experimental conditions.     

Overview of Petroleum Environmental Research Forum (PERF) dense gas dispersion modeling project

This paper provides a background for and an overview of the results of a comprehensive study of transport and dispersion of dense gas plumes over rough surfaces typical of industrial sites. The Petroleum Environmental Research Forum (PERF) 93-16 project involved model development and evaluations using observations from three wind tunnels and from the Kit Fox field experiment. Detailed discussions of the results of the research are given in the other papers in this special issue. The wind tunnel experiments produced data showing that the resulting best-fit vertical entrainment formula was close to (i.e., within about 30%) the vertical entrainment formulas already in use by current models, which were derived primarily from observations over smooth surfaces. Observations from the Kit Fox field experiment demonstrated the validity of the entrainment curves derived from the wind tunnel data. The Kit Fox data were also used to evaluate algorithms for along-wind dispersion and cloud advection speeds for short-duration releases typical of an industrial site, and to evaluate the HEGADAS dense gas dispersion model.