Detection and differentiation of Salmonella serotypes using surface enhanced Raman scattering (SERS) technique

This research was conducted to prove that developed silver biopolymer nanoparticle substrate for surface enhanced Raman scattering (SERS) technique could detect and differentiate three different serotypes of Salmonella. Nanoparticle was prepared by adding 100 mg of silver nitrate to a 2 % polyvinyl alcohol solution, then adding 1 % trisodium citrate to reduce silver nitrate and produce silver encapsulated biopolymer nanoparticles. Then, nanoparticle was deposited on a stainless steel plate and used as SERS substrate. Fresh cultures of Salmonella typhimurium, Salmonella enteritidis and Salmonella infantis were washed and suspended in 10 mL of sterile deionized water. Approximately 5 μl of the bacterial suspensions were placed on the substrate individually and exposed to 785 nm laser excitation. SERS spectral data were recorded between 400 and 1,800 cm^?1. SERS signals were collected from 15 different spots on the substrate for each sample. PCA model was developed to classify Salmonella serotypes. PC1 identified 92 % of the variation between the Salmonella serotypes, and PC2 identified 6 % and in total 98 % between the serotypes. Soft independent modeling of class analogies of validation set gave an average correct classification of 92 %. Comparison of the SERS spectra of Salmonella serotypes indicated that both isolates have similar cell walls and cell membrane structures which were identified by spectral regions between 520 and 1,050 cm^?1. However, major differences were detected in cellular genetic material and proteins between 1,200 and 1,700 cm^?1. SERS with silver biopolymer nanoparticle substrate could be a promising tool in pathogen detection and it would potentially be used to classify them.     

Use of Fourier transform infrared spectroscopy for rapid comparative analysis of Bacillus and Micrococcus isolates

The identification of foodborne microorganisms and their endospores in food products are important for food safety. The present work compares Bacillus (Bacillus licheniformis, Bacillus circulans and Bacillus subtilis) and Micrococcus (Micrococcus luteus) species with Fourier transform infrared (FTIR) spectroscopy. Our results show that there are several characteristic peaks belonging to both the Micrococcus and Bacillus species which can be used for the identification of these foodborne bacteria and their endospores. For Micrococcus species, a new band was observed at 1338 cm⁻¹ which may be due to acetate oxidation via the carboxylic acid cycle. The bands at 1313 cm⁻¹ and 1256 cm⁻¹ can be explained by an exopolymer formation and the other bands at 1074 cm⁻¹ and 550 cm⁻¹, may be due to the glycogen-like storage material in Micrococcus spp. There are also characteristic peaks at 993 cm⁻¹ and 801 cm⁻¹ for these bacterial species. Different Bacillus species also showed characteristic peaks at 1000–500 cm⁻¹ region. Dipicolinic acid (DPA) bands at ∼728 cm⁻¹ and ∼703 cm⁻¹ seen only in B. circulans were the marker of an endospore formation.     

Bacterial inactivation in fruit juices using a continuous flow Pulsed Light (PL) system

In this work, the susceptibility to pulsed light (PL) treatments of both a Gram-positive (L. innocua 11288) and a Gram-negative (E. coli DH5-??) bacteria inoculated in apple (pH = 3.49, absorption coefficient 13.9 cm^− 1) and orange juices (pH = 3.78, absorption coefficient 52.4 cm^− 1) was investigated in a range of energy dosages from 1.8 to 5.5 J/cm². A laboratory scale continuous flow PL system was set up for the experiments, using a xenon flash-lamp emitting high intensity light in the range of 100-1100 nm. The flashes lasted 360 ??s at a constant frequency of 3 Hz.The results highlighted how the lethal effect of pulsed light depended on the energy dose supplied, the absorption properties of liquid food as well as the bacterial strain examined. The higher the quantity of the energy delivered to the juice stream, the greater the inactivation level. However, the absorbance of the inoculated juice strongly influenced the dose deliver and, therefore, the efficiency of the PL treatment. Among the bacteria tested, E. coli cells showed a greater susceptibility to the PL treatment than L. innocua cells in both apple and orange juices. Following treatment at 4 J/cm², microbial reductions in apple and orange juices were, respectively, 4.00 and 2.90 Log-cycles for E. coli and 2.98 and 0.93 Log-cycles for L. innocua.Sublethally injured cells were also detected for both bacterial strains, thus confirming that membrane damage is an important event in bacterial inactivation by PL.     

Antioxidant and antimicrobial activities of Laetiporus sulphureus (Bull.) Murrill

Antioxidant capacity and antimicrobial activities of Laetiporus sulphureus (Bull.) Murrill. extracts obtained with ethanol were investigated in this study. The study was aimed at determining the antioxidant activity (DPPH free radical-scavenging, β-carotene/linoleic acid systems), total phenolic content and total flavonoid concentration of L. sulphureus. Inhibition values both of L. sulphureus ethanol and the standards increased parallel with the elevation of concentration in the linoleic acid system. Inhibition values of L. sulphureus (LS) extract, BHA and α-tocopherol standards were found to be 82.2%, 96.4% and 98.6%, respectively, at a concentration of 160 μg/ml. DPPH free radical-scavenging activity was found to exhibit 14%, 26%, 55% and 86% inhibition, respectively, at concentrations of 100, 200, 400 and 800 μg/ml. Total flavanoids were 14.2 ± 0.12 μg mg⁻¹ (quercetin equivalent) while the phenolics were 63.8 ± 0.25 μg mg⁻¹ (pyrocatechol equivalent) in the extract. Positive correlations were found between total phenolic content in the mushroom extracts and their antioxidant activities. Edible mushrooms may have potential as natural antioxidants. L. sulphureus showed narrow antibacterial activity against Gram-negative bacteria and strongly inhibited the growth of the Gram-positive bacteria tested. The crude extract exhibited high anticandidal activity on Candida albicans. Therefore, the extracts could be suitable as antimicrobial and antioxidative agents in the food industry.     

Comparison of free amino acid,carbohydrates concentrations in Korean edible and medicinal mushrooms

Ten popular species of both edible and medicinal Korean mushrooms were analysed for their free amino acids and disaccharides. The average total free amino acid concentration was 120.79 mg g⁻¹ in edible mushrooms and 61.47 mg g⁻¹ in medicinal mushrooms, respectively. The average total of free amino acids for all mushrooms, edible mushrooms and medicinal mushrooms was 91.13 mg g⁻¹. Agaricus blazei (227.00 mg g⁻¹) showed the highest concentration of total free amino acids; on the other hand, Inonotus obliquus (2.00 mg g⁻¹) showed the lowest concentration among the 10 species of mushrooms. The average total carbohydrates concentration was 46.67 mg g⁻¹ in the 10 species of mushrooms, where the edible mushrooms contained 66.68 mg g⁻¹ and the medicinal mushrooms contained 26.65 mg g⁻¹. The carbohydrates constituents of the 10 mushroom species were mainly mannose (36.23%), glucose (34.70%), and xylose (16.83%).     

Chemical characterization of a traditional honey-based Sardinian product: Abbamele

The first chemical characterization of abbamele, a traditional honey decoction from Sardinia (Italy) is hereby reported. Water content (from 17.7% to 27.7%), electrical conductivity (from 0.19 to 0.81 mS cm⁻¹), pH (from 3.21 to 3.92), free acidity (from 26.1 to 87.6 meq kg⁻¹), invertase activity (from 0 to 1.02 U kg⁻¹), 5-(hydroxymethyl)-2-furaldehyde, HMF (from 881 to 4776 mg kg⁻¹), total polyphenols (from 188 to 984 mg kg⁻¹) and free amino acid contents of thirteen abbamele samples, from industrial and traditional producers, were obtained in an attempt to compare this traditional product with honey and to study the relationship between its main features and the production procedures. The long thermal treatment involved in the production of abbamele has been identified as the main cause of very low (or absent) invertase activity and free amino acid content as well as the very high content of HMF.     

Assessment of the microbiological safety of dried spices and herbs from production and retail premises in the United Kingdom

A study of dried spices and herbs from retail and production premises to determine the microbiological status of such products was undertaken in the UK during 2004. According to EC Recommendation 2004/24/EC and European Spice Association specifications, 96% of 2833 retail samples and 92% of 132 production batches were of satisfactory/acceptable quality. Salmonella spp. were detected in 1.5% and 1.1% of dried spices and herbs sampled at production and retail, respectively. Overall, 3.0% of herbs and spices contained high counts of Bacillus cereus (1%, ≥10⁵ cfu g⁻¹), Clostridium perfringens (0.4%, ≥10³ cfu g⁻¹) and/or Escherichia coli (2.1%, ≥10² cfu g⁻¹). Ninety percent of samples examined were recorded as being ‘ready-to-use’, 96% of which were of satisfactory/acceptable quality. The potential public health risk of using spices and herbs as an addition to ready-to-eat foods that potentially undergo no further processing is therefore highlighted in this study. Prevention of microbial contamination in dried herbs and spices lies in the application of good hygiene practices during growing, harvesting and processing from farm to fork, and effective decontamination. In addition, the importance of correct food handling practices and usage of herbs and spices by end users cannot be overemphasised.     

Acaricidal activity of fennel seed oils and their main components against Tyrophagus putrescentiae, a stored-food mite

The acaricidal activities of components derived from Foeniculum vulgare (fennel) seed oils against Tyrophagus putrescentiae adults were examined using direct contact application and compared with those of the compounds benzyl benzoate, dibutyl phthalate and N,N-diethyl-m-toluamide. The biologically active constituent of the F. vulgare seeds was characterized as (+)-carvone by spectroscopic analyses. On the basis of LD₅₀ values, the compound most toxic to T. putrescentiae was naphthalene (4.28 μg/cm²) followed by dihydrocarvone (4.32 μg/cm²), (+)-carvone (4.62 μg/cm²), (−)-carvone (5.23 μg/cm²), eugenol (10.62 μg/cm²), benzyl benzoate (11.24 μg/cm²), thymol (11.42 μg/cm²), dibutyl phthalate (13.11 μg/cm²), N,N-diethyl-m-toluamide (13.53 μg/cm²), methyl eugenol (39.52 μg/cm²), myrcene (39.88 μg/cm²) and acetyleugenol (72.24 μg/cm²). These results indicate that acaricidal activity of the F. vulgare seed oil could be caused by carvone and naphthalene of which the former is likely to be more important because it is 74.7 times more abundant than naphthalene. Carvone and naphthalene merit further study as potential stored-food mite control agents or as lead compounds.     

Production of trehalose by intramolecular transglucosylation of maltose catalysed by a new enzyme from Thermus thermophilus HB-8

Thermus thermophilus HB-8 is a source of trehalose synthase (GTase), which catalyses conversion of maltose into trehalose. Specific activity of maltose transglucosylation by cell-free extracts of the bacteria was about 0.1 U mg⁻¹ protein and precipitation at 28% saturation of ammonium sulphate caused 3.5-fold enzyme purification. The optimum temperature for conversion of maltose into trehalose was 65 °C with about 27% of maximum activity at 85 °C. The highest GTase productivity was achieved at cultivation temperature over 60 °C and at NaCl concentration range of 0.1–0.5% (w/v). However, larger concentrations of sodium chloride in the growth media caused a remarkable decrease of GTase synthesis. The results, of ammonium sulphate fractionation and activity towards maltotriose (0.028 U mg⁻¹), maltotetraose (0.16 U mg⁻¹) and GlcαpNp (0.27 U mg⁻¹), show that trehalose synthase and α-glucosidase activities reside in separate protein fractions of cell-free extracts from T. thermophilus cells.     

Efficacy of neutral electrolyzed water for sanitization of cutting boards used in the preparation of foods

The effectiveness of neutral electrolyzed water (NEW) to sanitize cutting boards used for food preparation was investigated. Cutting boards made of hardwood and bamboo were inoculated with Escherichia coli K12 and Listeria innocua, dried for 1 h, washed, rinsed and sanitized with NEW, sodium hypochlorite (NaClO) solution, or tap water (control). After each washing protocol, surviving bacterial populations were determined. Results showed that both NEW and NaClO sanitizing solutions produced similar levels of bacterial reductions. In manual washing, the population reductions by NEW and NaClO were 3.4 and 3.6 log₁₀ CFU/100 cm² for E. coli, and 4.1 and 3.9 log₁₀ CFU/100 cm² for L. innocua, respectively. In the automatic washing, the reductions by NEW and NaClO were 4.0 and 4.0 log₁₀ CFU/100 cm² for E. coli, and 4.2 and 3.6 log₁₀ CFU/100 cm² for L. innocua, respectively. No significant differences (P > 0.05) were observed in surviving bacteria counts when comparing board material types.     

Interference of Origanum vulgare L. essential oil on the growth and some physiological characteristics of Staphylococcus aureus strains isolated from foods

The aim of this study was to investigate the chemical composition of Origanum vulgare L. essential oil, the inhibitory effect of the oil on the cell viability of Staphylococcus aureus strains isolated from foods, and the influence of sub-inhibitory concentrations of the oil on some physiological attributes of these strains. GC-MS analysis showed that carvacrol (57.71%) was the most prevalent compound in the oil, followed by p-cymene (10.91%), γ-terpinene (7.18%), terpinen-4-ol (6.68%) and thymol (3.83%). The results showed that O. vulgare essential oil at 0.03, 0.6 and 0.12 μL mL⁻¹ inhibited the cell viability of Staph. aureus. At 0.12 μL mL⁻¹ the oil caused cidal effect with decrease ≥3 log cycles of the initial inoculum after 15 min of exposure. Sub-inhibitory concentrations (0.03 and 0.015 μL mL⁻¹) of the oil suppressed some physiological attributes of the Staph. aureus strains such as coagulase, lipase and salt tolerance. The oil interfered on the microbial metabolic activity in a dose-dependent manner. O. vulgare essential oil could be a novel antimicrobial with capability to suppress some physiological characteristics, in addition to inhibit the growth and survival of pathogen bacteria in foods, particularly Staph. aureus.     

Combined effect of enterocin AS-48 and high hydrostatic pressure to control food-borne pathogens inoculated in low acid fermented sausages

The single and combined effects of enterocin AS-48 and high hydrostatic pressure (HHP) on Listeria monocytogenes, Salmonellaenterica, and Staphylococcus aureus was investigated in fuet (a low acid fermented sausage) during ripening and storage at 7 °C or at room temperature. AS-48 (148 AU g⁻¹) caused a drastic 5.5 log cfu g⁻¹ decrease in L. monocytogenes (P < 0.001) and a significant (P < 0.01) inhibition (1.79 logs) for Salmonella at the end of ripening (10 d). After pressurization (400 MPa) and storage Listeria counts remained below 5 cfu g⁻¹ in all fuets containing AS-48 (pressurized or not). HHP alone had no anti-Listeria effect. HHP treatment significantly reduced Salmonella counts, with lowest levels in pressurized fuets with AS-48. S. aureus showed similar growth for all treatments and storage conditions. These results indicate that AS-48 can be applied alone to control L. monocytogenes and combined with HHP treatment to control Salmonella in fuets.     

Salmonella surveillance and control at post-harvest in the Belgian pork meat chain

Salmonella remains the primary cause of reported bacterial food borne disease outbreaks in Belgium. Pork and pork products are recognized as one of the major sources of human salmonellosis. In contrast with the primary production and slaughterhouse phases of the pork meat production chain, only a few studies have focussed on the post-harvest stages. The goal of this study was to evaluate Salmonella and Escherichia coli contamination at the Belgian post-harvest stages. E. coli counts were estimated in order to evaluate the levels of faecal contamination. The results of bacteriological analysis from seven cutting plants, four meat-mincing plants and the four largest Belgian retailers were collected from official and self-monitoring controls. The prevalence of Salmonella in the cutting plants and meat-mincing plants ranged from 0% to 50%. The most frequently isolated serotype was Salmonella typhimurium. The prevalence in minced meat at retail level ranged from 0.3% to 4.3%. The levels of Salmonella contamination estimated from semi-quantitative analysis of data relating to carcasses, cuts of meat and minced meat were equal to −3.40 ± 2.04 log CFU/cm², −2.64 ± 1.76 log CFU/g and −2.35 ± 1.09 log CFU/g, respectively. The E. coli results in meat cuts and minced meat ranged from 0.21 ± 0.50 to 1.23 ± 0.89 log CFU/g and from 1.33 ± 0.58 to 2.78 ± 0.43 log CFU/g, respectively. The results showed that faecal contamination still needs to be reduced, especially in specific individual plants.     

Efficacy of diflubenzuron plus methoprene against Sitophilus oryzae and Rhyzopertha dominica in stored sorghum

The efficacy of diflubenzuron (1 mg kg⁻¹)+methoprene (1 mg kg⁻¹) against Sitophilus oryzae (L.) and Rhyzopertha dominica (F.) in sorghum was evaluated in a silo-scale trial in southeast Queensland, Australia. Sorghum is normally protected from a wide range of insects by mixtures of grain protectants. The chitin synthesis inhibitor diflubenzuron was evaluated as a potential new protectant for S. oryzae in combination with the juvenile hormone analogue methoprene, which is already registered for control of R. dominica. Sorghum (ca 200 t) was treated after harvest in 2000 and assessed for treatment efficacy and residue decline during 6.5 months storage. The reproductive capacity of S. oryzae and R. dominica was greatly reduced in bioassays of treated sorghum throughout the trial, and efficacy remained relatively stable during the trial. An initial exposure of S. oryzae adults to treated sorghum for 2 weeks reduced F₁ progeny production of all strains by 80.8-98.8%, but a second exposure of 4 weeks reduced F₁ progeny production by 98.5-100%. In addition, the reproductive capacity of any S. oryzae progeny produced was greatly reduced. Exposure of R. dominica adults to treated sorghum for 2 weeks reduced F₁ progeny production of all strains by 99.6-100%, including a methoprene-resistant strain. The results indicate that S. oryzae or R. dominica adults invading sorghum treated with diflubenzuron (1 mg kg⁻¹)+methoprene (1 mg kg⁻¹) would be incapable of producing sustainable populations.     

Toxicity of ethyl formate to adult Sitophilus oryzae (L.), Tribolium castaneum (herbst) and Rhyzopertha dominica (F.)

Fumigations were conducted using a continuous flow-through laboratory process to maintain constant concentrations of ethyl formate and low levels (<0.8%) of respiratory carbon dioxide. The procedure minimised the effects of sorption by exposing test insects without media and minimised the effect of carbon dioxide by use of continuous flow. The concentration×time (Ct) products of ethyl formate for adult Sitophilus oryzae, Tribolium castaneum and Rhyzopertha dominica at 25 °C and 70% relative humidity for the 6 h exposure were, respectively: (1) LD₅₀ 107.8, 108.8 and 72.8 mg h L⁻¹ and (2) LD_99.5 207.4, 167.1 and 122.2 mg h L⁻¹. Endpoint mortality was reached within 24 h of initial exposure.     

Smearing of soft cheese with Enterococcus faecium WHE 81, a multi-bacteriocin producer,against Listeria monocytogenes

Enterococcus faecium WHE 81, a multi-bacteriocin producer, was tested for its antimicrobial activity on Listeria monocytogenes in Munster cheese, a red smear soft cheese. The naturally delayed and superficial contamination of this type of cheese allowed the use of E. faecium WHE 81 at the beginning of the ripening as a surface culture. A brine solution inoculated at 10⁵ CFU of E. faecium WHE 81 per mL was sprayed on the cheese surface during the first smearing operation. On day 7, smearing of cheese samples with a brine solution at 10² CFU of L. monocytogenes per mL yielded initial cell counts of approximately 50 CFU g⁻¹ of the pathogen on the cheese surface. Although, in some instances, L. monocytogenes could survive (<50 CFU g⁻¹) in the presence of E. faecium WHE 81, it was unable to initiate growth. In control samples however, L. monocytogenes counts often exceeded 10⁴ CFU g⁻¹. In other respects, E. faecium WHE 81, which naturally existed in Munster cheese, did not adversely impact on the ripening process.     

Spoilage characteristics of Brochothrix thermosphacta and campestris in chilled vacuum packaged lamb,and their detection and identification by real time PCR

The spoilage potential of Brochothrix campestris and Brochothrix thermosphacta was investigated in vacuum-packed lamb. Striploins (n = 338) were inoculated and stored for twelve weeks at temperatures − 1.5, 0, 2 and 7 °C. Growth around 5–6 log₁₀ CFU/cm² was recorded after six weeks at 0, 2 and 7 °C, and ~ 3 log₁₀ CFU/cm² after nine weeks at − 1.5 °C. B. campestris was shown to cause spoilage by nine weeks at temperatures above 0 °C by the presence of green drip and unacceptable odours. Molecular based assays for the detection and differentiation of B. thermosphacta and B. campestris were developed and validated. A TaqMan assay was designed to target a unique single-nucleotide polymorphism in the Brochothrix 16 s rRNA gene with a sensitivity of < 7 CFU per reaction. Secondly a specific PCR was designed for B. campestris targeting the structural genes, brcA and brcB. These testing regimes offer a rapid and cost effective method for the detection and screening of Brochothrix species in meat products and processing environments.     

Sensory,microbiological and chemical assessment of the freshness of red mullet (Mullus barbatus) and goldband goatfish (Upeneus moluccensis) during storage in ice

The shelf life of red mullet and goldband goatfish during ice storage were studied in terms of sensory, microbiological and chemical changes. The sensory acceptability limit was 8 days for goldband goatfish (Upeneus moluccensis) and 11 days for red mullet (Mullus barbatus) stored in ice. The TVC level was correlated with sensory assessment. The TVC exceeded 7 log cfu g⁻¹ after 8 days for goldband goatfish, and 11 days for red mullet. At the end of storage period, pH, TVB-N, TBA, FFA and PV for red mullet were 7.84, 47.19 mg/100 g, 0.69 mg MA kg⁻¹, 1.17% oleic acid and 1.58 meq O₂/kg and for goldband goatfish they were 7.53, 43.97 mg/100 g, 0.74 mg MA kg⁻¹, 1.62% oleic acid and 1.68 meq O₂/kg, respectively. In red mullet, agmatine, serotonin, histamine and dopamine became the dominant amines, reaching 7.30, 5.97, 2.52 and 2.31 mg/100 g, respectively. Also the dominant amines for goldband goatfish were 4.37, 3.88, 3.38 and 2.00 mg/100 g for histamine, agmatine, dopamine and putrescine, respectively.     

Insecticidal activity of asarones identified in Acorus gramineus rhizome against three coleopteran stored-product insects

The insecticidal activities of Acorus gramineus rhizome-derived materials against adults of Sitophilus oryzae (L.), Callosobruchus chinensis (L.) and Lasioderma serricorne (F.) were examined using direct contact application and fumigation methods. The biologically active constituents of the Acorus rhizome were characterized as the phenylpropenes (Z)- and (E)-asarones by spectroscopic analysis. Responses varied with insect species, compound and exposure time rather than dose. In a filter paper diffusion test, (Z)-asarone caused 70% and 90% mortality against S. oryzae adults at 0.064 and 0.255 mg/cm² at 4 days after treatment, respectively, with 100% mortality at 7 days after treatment. (E)-Asarone at 0.255 mg/cm² was almost ineffective against S. oryzae adults at 7 days after treatment. Against C. chinensis adults at 0.064 mg/cm², (Z)- and (E)-asarones gave 100% mortality at 3 and 7 days after treatment, respectively. Against L. serricorne adults, (Z)-asarone gave 90% and 83% mortality at 0.255 and 0.064 mg/cm² at 7 days after treatment, respectively, whereas (E)-asarone at 0.255 mg/cm² was almost ineffective at 7 days after treatment. These results indicate that the toxicity of asarones might be due to the cis configuration rather than to the position of the double bond. In a fumigation test, (Z)-asarone at 0.577 mg/cm² was much more effective against adults of all three insect species in sealed containers than in open ones, indicating that the insecticidal activity of the compound was largely attributable to fumigant action.     

Flow injection spectrophotometry using natural reagent from Morinda citrifolia root for determination of aluminium in tea

A flow injection (FI) spectrophotometric method with using natural reagent extracted from Morinda citrifolia root has been developed for determination of aluminium. The extract contained anthraquinone compounds which could react with Al³⁺ to form reddish complexes which had maximum absorption wavelength at 499.0 nm. The extract could be used as a reagent in FI system without further purification to obtain pure compound. A sensitive method for determination of aluminium in concentration range of 0.1-1.0 mg L⁻¹, with detection limit of 0.05 mg L⁻¹ was achieved. Relative standard deviations of 1.2% and 1.7% were obtained for the determination of 0.1 and 0.6 mg L⁻¹ Al³⁺ (n = 11). Sample throughput of 35 h⁻¹ was achieved with the consumption of 3 mL each of carrier and reagent solutions per injection. The developed method was successfully applied to tea samples, validated by the FAAS standard method. The method is simple, fast, economical and could be classified as a greener analytical method.