Characteristics of Bacillus cereus isolates from legume-based Indian fermented foods

An antibiogram of 48 strains of Bacillus cereus isolated from 6 different kinds of legume-based Indian fermented foods (amriti, dhokla, dosa, idli, papad and wadi) was generated against 18 different antibiotics that are commonly used against foodborne diseases, mainly gastroenteritis. Each of the isolates was found to be resistant against at least nine different antibiotics. Production of extracellular enzymes, namely protease, lipase and amylase by 33%, 27% and 46%, respectively, of the isolates indicates their potentiality for food spoilage. In brain–heart infusion broth supplemented with glucose, the D_100 °C-values for the tested 12 strains ranged from 3.0 to 9.2 min. In nutrient broth, the minimum and maximum pHs permitting growth of B. cereus were 5.3 and 11.6, respectively. The minimum inhibitory concentrations of sodium chloride, benzoic acid and sorbic acid for the growth of the isolates were 65–85 mg ml⁻¹, 400–700 μg ml⁻¹ (pH 5.0–4.2) and 500–600 μg ml⁻¹ (pH 5.0–4.8), respectively. Of the tested 10 strains, eight were resistant to 300 μg nisin ml⁻¹ (pH 5.0). While studying the combined effect of selected hurdles on the growth of an isolate, the judicious combination considered was 20 mg sodium chloride, 300 μg benzoic acid and 25 μg nisin ml⁻¹ at pH 5.6. The whole-cell protein fingerprinting (WCPF) analysis using SDS–PAGE revealed a high level of diversity among the isolates. At 60% similarity level, the WCPF profiles could be grouped into four major clusters which were divided into 34 subclusters. Most of the subclusters were source-wise homogeneous.     

Occurrence of pathogenic bacteria in raw milk, cultured pasteurised milk and naturally soured milk produced at small-scale dairies in Zimbabwe

The presence of selected pathogenic bacteria in raw milk (RM), cultured pasteurised milk (CPM), and naturally soured raw milk (NSRM) sold at three small-scale dairies in Zimbabwe was determined. Mean levels of Escherichia coli in RM, CPM and NSRM were 4.5, 7.1 and 7.8 log₁₀ CFU ml⁻¹, respectively. Enterotoxigenic E. coli (ETEC), producing heat-stable enterotoxin (ST1) was found in 16 of the samples. Presumptive Staphylococcus aureus was identified in 7 of 12 RM, 15 of 27 CPM and 20 of 21 NSRM samples at mean levels of 5.2, 7.3 and 7.8 log₁₀ CFU ml⁻¹, respectively. Klebsiella pneumonia spp., Enterobacter cloacae, Proteus mirabilis, Hafnia alvei, Citrobacter spp., Serratia marcescens and Aeromonas hydrophilia were found in 27 samples. The high number of S. aureus, E. coli, identification of ETEC-ST1 and the other pathogenic microorganisms found in pasteurised and unpasteurised milk products represent a health hazard to the consumers and emphasises the need for improved hygiene practice at all levels in the dairy.     

Survival of Campylobacter jejuni on vacuum or carbon dioxide packaged primal beef cuts stored at −1.5 °C

The ability of two Campylobacter jejuni strains (MOO3, a cattle isolate, and MOO8, a sheep isolate) to survive on chilled, preservatively packaged primal beef cuts was examined. Each of the strains was inoculated separately (10⁵ cfu g⁻¹) onto 500 g beef steaks, packaged under vacuum or 100% carbon dioxide, and stored, with uninoculated controls, for 41 days at −1.5 °C. Bacterial numbers were determined at 0, 1, 3, 6, 13, 20, 27, 34 and 41 days by dilution, plating on both Columbia Sheep Blood Agar and Campylobacter Blood-Free Selective Agar and incubation at 42 °C for 48 h under microaerophilic conditions. In addition, an aerobic count on all samples was determined by dilution, plating on Plate Count Agar and incubation at 25 °C for 72 h. No significant changes in numbers of the C. jejuni strains or aerobic bacteria occurred during storage at −1.5 °C on steaks packaged under either vacuum or carbon dioxide. The ability of these pathogens to survive standard preservative packaging conditions is different from that reported in the literature and therefore a cause for concern. Strict hygienic practice or the implementation of decontamination technologies are recommended as mechanisms to assure safety of meat with respect to this pathogen.     

Simple methods for rapid determination of sulfite in food products

Two simple methods for sulfite determination in food products were proposed. The modified para-rosaniline (PRA)-formaldehyde method showed a much broader linear range (0.05–5.0 mg l⁻¹ as SO₂) than the commonly used procedure for the sulfur dioxide detection in the atmosphere (0.05–1.0 mg l⁻¹ as SO₂). By using a standard reference color card, this method only needed 5 min to complete a test. The 5,5′-dithiobis(2-nitrobenzoic acid) (DTNB) method was another good alternative for rapid determination of sulfite since it only needed one main reagent and the method was robust and easy to operate with the linear range of 0.10–4.3 mg l⁻¹ as SO₂. When applying to real food samples, the DTNB method had good recoveries for all the tested samples and the results agreed well with those obtained by the official iodometric titration. The modified PRA-formaldehyde method worked well with all the tested dried food products, though matrix effect of sulfite binding was observed for the beer samples. Thus, the modified PRA-formaldehyde method has advantages of sensitivity and rapidity, but the DTNB method offers a wider range of applications. Both methods provided practical ways for in situ determination of sulfite by non-professional operators with the modified PRA-formaldehyde method suited for food samples with less sulfite binding problems.     

Microbiological quality assessment of garri with reference to thermonuclease activity

The microbiological quality of garri obtained from open markets and traditional processing industry in Benin City with reference to staphylococcal contamination was investigated. Enumeration of the total heterotrophic bacteria, total fungal propagules and Staphylococcus aureus in food samples was carried out using appropriate media. The staphylococcal thermonuclease activity determination was used as an indicator of previous (substantial) growth of S. aureus and possible presence of staphylococcal enterotoxins. The total heterotrophic bacterial count of the open markets’ samples was 7.75±2.87×10³ cfu/g with corresponding staphylococcal count of 1.65±0.82×10³ and fungal propagule count of 2.50±1.14×10² cfu/g. Samples obtained immediately after processing from the traditional garri industry revealed total heterotrophic bacterial count of 5.00×10¹ cfu/g; staphylococcal count of 5.00×10⁰ cfu/g and no fungal count. The zone diameter of thermonuclease activity was 8.13±0.52 mm for open markets’ samples while traditional processing industry samples have 7.83±0.54 mm. From the study, it was found that there was contamination and growth of S. aureus and possibly the presence of staphylococcal enterotoxin in the product. The public health significance of the microorganisms with the production of thermonuclease and enterotoxins by S. aureus are discussed.     

Microbiological profile of helva sold at retail markets in Izmir city and the survival of Staphylococcus aureus in this product

The objective of this research was to determine the microbiological profile of tahin helva, a low moisture confectionery. A total of 63 tahin helva samples were collected from the retail markets in Izmir, Turkey. These samples were examined by standard procedures for aerobic plate count (APC) and counts of moulds and yeasts, Enterobacteriaceae, Staphylococcus aureus and Salmonella spp. APC of samples ranged from <10 to 1.6 × 10⁵ cfu/g. The counts for moulds, yeasts and Enterobacteriaceae ranged from <10 to 1.8 × 10³, <10 to 7 × 10², <10 to 8.5 × 10² cfu/g, respectively. None of the samples contained S. aureus or Salmonella. The potential for survival of S. aureus in the product stored at refrigeration (4 °C) and room (20 °C) temperatures was evaluated by artificial contamination. S. aureus cells were still recovered after 9 months of storage at both temperatures. The results of this investigation indicate that S. aureus contamination of tahin helva, (a_w of 0.172) may constitute a potential public health hazard, depending on the extent of contamination and mishandled exercise at the sale area and the kitchen.     

The dissipation rates of cyprodinil, fludioxonil, procymidone and vinclozoline during storage of grape juice

The disappearance of the fungicides cyprodinil, fludioxonil, procymidone and vinclozoline, which are widely used to control grey mold in vineyards, from commercially sterilized white grape juice was studied. Fungicide losses from white grape juice stored at 40 °C were monitored for about 2 months in order to simulate in a fast way the shelf-life of the product at room temperature (about 1 year). The fungicides were determined by using a simple method based on a liquid–liquid extraction (LLE) and gas chromatographic separation, followed by mass spectrometric detection (GC–MSD). Fortified white grape juice subsamples were taken three times a week from the oven and the degradation of the fungicides monitored for 56 days. The half-lives at 40 °C for vinclozoline and procymidone were found to be 11 and 20 days, respectively. Those for the more recently introduced fungicides fludioxonil and cyprodinil were somewhat longer (33 and 44 days, respectively). Based on the first-order rate constants obtained, the fungicides dissipated in the following sequence: vinclozoline (0.062 d⁻¹) > procymidone (0.035 d⁻¹) > fludioxonil (0.021 d⁻¹) > cyprodinil (0.016 d⁻¹).     

Computer modelling for the control of particulate sterilization under dynamic flow conditions

A finite difference model was used for predicting the centre temperature of particulates during ultra-high temperature processing under continuous flow conditions. This model compared favourably with the analytical solutions. A sensitivity analysis was carried out on the input parameters to the numerical model, which indicated that particulate size and thermal diffusivity were the most critical parameters influencing process lethality and cook value. For example, a spherical particulate (thermal properties defined) of 20 mm diameter required a holding time greater than 5 min at 135°C to achieve a minimum safe cook (F₀ of 3.0 min for Clostridium botulinum), compared to a 10 mm spherical particulate which would have received an F₀ value of 68 min after 5 min processing at 135°C. The centre temperatures predicted using the model were sensitive to changes in the convective surface heat transfer coefficient in the range 100–300 W m⁻² K⁻¹, but became less sensitive as the values increased up to 500 W m⁻² K⁻¹. An initial product temperature change from 20 to 60°C resulted in an 8 min increase in process lethality for a 15 mm spherical particulate processed for 5 min at 135°C. The determination of experimental results in particulates of less than 20 mm (the critical dimension) was found to be significantly affected by conduction errors through the thermocouple wires, and therefore the model presented will be a useful prediction tool in situations of experimental uncertainty.     

Rapid gas-chromatographic method for the determination of diacetyl in milk, fermented milk and butter

In this work, a simple and fast method for the determination of diacetyl by gas-chromatographic technique coupled with flame ionisation detector (GLC-FID) was developed. Diacetyl is the typical butter flavour, but it is also commonly present in others fermented dairy products. Recently, diacetyl determination has also attracted interest because it is one of the parameters on which lactic acid bacteria (L.A.B.) are characterized and valued. Only acetone and 2,3-pentanedione were used as chemicals. After centrifugation of acetone–milk mixture, supernatant was filtered and directly injected into gas-chromatographic apparatus, without a further purification procedure step.

This method was accurate and precise; diacetyl recovery on milk was 97% and the detection limit was 1 mg L⁻¹. Finally, by using this method, diacetyl was easily determined in fresh and high-temperature treated milk, commercial butter, yoghurt and also in a series of L.A.B. performance tests.     

Effect of nisin on the storage of sous vide processed Korean seasoned beef

Seasoned beef called Jangzorim in Korea is produced by boiling in soy sauce, and is a popular food in Korea. The aim of this study is to evaluate the microbial safety and physical qualities of sous vide processed seasoned beef, and the effect of nisin during storage. Sous vide processed packages with or without nisin (100 IU or 500 IU) were stored at 4 °C or 25 °C for 60 days, and samples measured for quality at regular intervals throughout this storage period. In the case of 25 °C storage, the number of mesophilic microorganisms in seasoned beef packages without nisin increased markedly, but with nisin there was no observed increase. Psychrotrophic microorganisms, anaerobic microorganisms, and B. cereus cells showed similar trends, although C. perfringens was not detected in all samples. At 25 °C storage, changes in the cutting force of packages containing nisin showed no significant change, packages without nisin decreased markedly. The colour of packages without nisin showed a drastic decrease in lightness (‘L’) while no changes were observed in packages with nisin.     

An assessment of the microbial quality of the air within a pork processing plant

The aim of this study was to assess microbial quality of the air in a pork processing plant and from visual inspection to identify factors which may contribute to increase/decrease air-borne contamination. Total viable count (TVC), total coliform count (TCC) and the presence of Staphylococcus, Listeria and Salmonella species were used to evaluate the microbial quality of the air at five locations in the plant, at five time points during production. A visual inspection was used to identify risk factors which may increase the micro flora of the air in the plant. The highest TVC (133 cfu m⁻³), coliform counts (8 cfu m⁻³) and Staphylococcus aureus counts (8 cfu m⁻³) were found in the cooking area, blast chill and raw areas respectively. The higher levels of TVC, coliforms and Staphylococcus could most likely be attributed to people in the area, condensation and mechanically generated aerosol respectively although factors such as psychrometric properties of air and building ventilation factors which were outside of the scope of the current investigation would also need to be considered before a definite conclusion could be made in this regard. Listeria and Salmonella were not detected in the air in the plant. However, the inspection suggested the air in the raw area to be most at risk of microbial contamination, due to higher levels of microbial sources. Overall, microbial levels in the air were low, which suggests that the current control measures such as the positive air pressures from high-risk to low-risk, internal separators and closed drains were effective in reducing the microbial contamination in the air of the plant.     

Incidence of aflatoxins, ochratoxin A and zearalenone in tunisian foods

A total of 209 samples of different groups of foods widely consumed by the Tunisian population were collected during 2004–2005 years. Samples were analyzed for contamination with aflatoxins, ochratoxin A and zearalenone, using competitive enzyme-linked immunosorbent assay (ELISA). The predominant mycotoxin was ochratoxin A with a mean level of 3.5 ± 5.3 ng g⁻¹ in 59.8% of studied samples. Furthermore, Aflatoxins were detected in all analyzed commodities with a contamination frequency of 50.5%. In addition, aflatoxin B1 was found in 37% of the samples. The zearalenone was detected around 15% with a mean level of 10.4 ± 11.8 ng g⁻¹. Species, dried fruits and sorghum were the most contaminated samples by aflatoxin and ochratoxin mycotoxins, whereas Rice was the least contaminated commodity. The most frequent mycotoxins co-occurrence included aflatoxins and ochratoxin A, which have been detected in 33.8% of analyzed samples. Furthermore, the simultaneous contamination by aflatoxins, ochratoxin A and zearalenone was observed in 7.2% of studied samples.     

Determination of heavy metals in milk by potentiometric stripping analysis using a home-made flow cell

A method is described for sensitive and reliable determination of trace concentrations of cadmium, lead and copper in real samples by stripping potentiometry with a home-made flow cell. The selection of the experimental conditions was made using experimental design methodology. The optimum conditions for the method include an acetic acid–acetate buffer mixture (pH 3.4) as supporting electrolyte, an electrolysis potential of −1.1 V and a flow rate of 3 ml min⁻¹. The detection limits were 6.51 × 10⁻¹⁰ mol l⁻¹ for cadmium, 4.60 × 10⁻¹⁰ mol l⁻¹ for lead and 8.59 × 10⁻¹⁰ mol l⁻¹ for copper, with an electrolysis time of 900 s. The relative standard deviations at this concentration level were 0.038, 0.086 and 0.095, respectively. In order to check the analytical accuracy, standard solutions have been used and recoveries close to 100% were obtained. The method was successfully applied to the determination of cadmium, lead and copper in powdered milk, using the method of standard additions.     

Antibacterial activities of the surface microflora of Kefalograviera cheese

The microflora developed on the surface of Kefalograviera, a hard cheese, during ripening in the warm room was studied. The microbial total counts and yeasts decreased (P < 0.05) and the same was observed with the pH on ripening for 15 days. Bacilli dominated at day 4 while at 15 days Micrococcaceae and enterococci occurred frequently. A great number (38 out of 56) of the isolates exhibited narrow spectrum of antibacterial activities against Gram-positive indicator strains, while ten of the Bacillus isolates (Bacillus pumilus) inhibited the growth of Listeria monocytogenes. The extracellular antilisterial substances were sensitive to proteinase K and heat labile (at 100 °C).     

An evaluation of the effect of storage and processing temperatures on the microbiological status of sous vide extended shelf-life products

The microbiological and organoleptic quality of selected sous vide products was monitored over a storage period of up to five weeks at 3°C and 8°C, respectively. Sous vide products stored at the recommended temperature of 3°C showed negligible microbial growth and were found to be organoleptically acceptable throughout the storage period. Total plate counts at the end of the fourth week of storage were in the range <10–7×10³ CFU/g for all 19 products under study. Listeria monocytogenes, Salmonella, Clostridium perfringens, Bacillus cereus and Enterobacteriaceae were not detected in any of the processed samples. At 8°C, under temperature abuse conditions, while some products had acceptable microbial levels of 10²–10⁴ CFU/g after three weeks, others such as chicken chasseur had counts above 10⁶ CFU/g by the second week of storage. Lactic acid bacteria and Pseudomonas species were dominant in the microbial flora of spoilt samples and B. cereus (>3×10⁴ CFU/g) was isolated from spoilt chicken chasseur samples in the fourth week of storage at 8°C. Critical factors affecting sous vide product safety are discussed.     

Microbiological quality of fruit juices sold in Tripoli–Libya

Fruit juices are becoming an important part of the modern diet in many communities. Data regarding the microbiological quality of foods, particularly fruit juices, sold in Libya and the neighboring North African countries are few if not lacking. Using standard microbiological procedures, 146 fruit juice samples sold in Tripoli were examined. Total bacterial counts (TBC) of samples examined ranged between <1 and 3 × 10⁵ CFU/ml (mean = 5 × 10⁴). Almond juice showed the highest TBC ranging between 1.8 × 10³ and 3 × 10⁵ (mean = 1.7 × 10⁵). Staphylococcus aureus was detected in 8 (5.5%) samples, Streptococcus spp. in 4 (2.7%), coliforms in 33 (22.6%), Escherichia coli (none were of serogroup O157) in 3 (2.1%), Klebsiella pneumonia in 17 (11.6%), Aeromonas spp. in 3 (2.1%), Pseudomonas aeruginosa in 6 (4.1%), Candida albicans in 18 (12.3%), Candida spp. in 109 (74.7%) and other yeasts in 85 (58.2%). Many of these organisms can cause disease in humans, which indicates that a large number of juices tested did not fulfill the Libyan guidelines for the microbiological quality of juices. Application of the HACCP system should be introduced into the food industry sector to improve the quality of fruit juices and other foods manufactured in Libya.     

Microbiological characteristics of cake prepared from rice flour and sticky rice flour

Microbiological, chemical and sensory changes in MiGao (a traditional Chinese steamed cake) were studied during five days of storage at 25 °C. Microorganisms examined for were: Enterobactericeae, lactic acid bacteria, Gram-positive, catalase-positive cocci, Staphylococcus aureus, Bacillus strains, yeasts and moulds, and pH and moisture content were also analyzed. Total plate counts in the products under study in the third day of storage were in the range <10³ CFU/g and the shelf-life of the product was estimated to be only two or three days by the panelists. During the first two days, Gram-positive bacteria were dominant, mainly represented by Staphylococcus epidermidis. Bacillus strains occurred by the third day, reaching a maximum level of 1 × 10⁶ CFU/g after five days of storage. No Enterobactericeae or Lactic acid bacteria were detected in the processed products throughout the storage period. All the isolated strains from the mannitol salt agar belonged to the genus Staphylococcus. The predominant species were S. epidermidis and S. aureus. Most Bacillus strains, isolated from the tryptone soya agar, belonged to the species Bacillus brevis. The count of yeasts and moulds increased slowly but remained low throughout the storage period.     

The in vitro antimicrobial and antioxidant activities of the essential oils and methanol extracts of endemic Thymus spathulifolius

The present study was conducted to evaluate the in vitro antimicrobial and antioxidant properties of essential oil and methanol extracts from a unique and endemic plant, Thymus spathulifolius (Hausskn. and Velen.). The antimicrobial test results showed that the essential oil of T. spathulifolius strongly inhibited the growth of test microorganisms studied, except for 4 fungi species while polar and non-polar subfractions of the methanol extract had moderate antibacterial, but not antifungal and anticandidal activity. The antioxidative potential of the samples was evaluated using two separate methods, inhibition of free radical 2,2-diphenyl-1-picrylhydrazyl (DPPH) and β-carotene–linoleic acid systems. The polar subfraction of the methanol extract was able to reduce the stable free radical 2,2-diphenyl-1-picrylhydrazyl (DPPH) with an IC₅₀ of 16.15 ± 0.5 μg/ml, which was lower than that of synthetic antioxidant, BHT, (19.8 ± 0.5 μg/ml). Inhibition values of linoleic acid oxidation were calculated as 92% and 89% for the oil and the polar subfraction, respectively. Gallic acid equivalent total phenolic constituent of the polar subfraction was 141.00 ± 0.90 μg/mg (14.1%, w/w). The chemical composition of a hydrodistilled essential oil of T. spathulifolius was analyzed by a GC and GC/MS system. A total of 28 constituents representing 99.2% of the oil were identified; thymol (36.5%), carvacrol (29.8%), p-cymene (10.0%) and γ-terpinene (6.3%) were the main components comprising 82.6% of the oil. Results presented here may suggest that the essential oil and extracts of T. spathulifolius possess antimicrobial and antioxidant properties, and therefore, they can be used as a natural preservative ingredient in food and/or pharmaceutical industry.     

Application of electrolyzed oxidizing water on the reduction of bacterial contamination for seafood

For reducing bacterial contamination, electrolyzed oxidizing water (EO water) has been used to reduce microbial population on seafood and platform of fish retailer. The specimens of tilapia were inoculated with Escherichia coli and Vibrio parahaemolyticus, and then soaked into EO water for up to 10 min. EO water achieved additional 0.7 log CFU/cm² reduction than tap water on E. coli after 1 min treatment and additional treatment time did not achieved additional reduction. EO water treatment also reduced V. parahaemolyticus, by 1.5 log CFU/cm² after 5 min treatment and achieved 2.6 log CFU/cm² reduction after 10 min. The pathogenic bacteria were not detected in EO water after soaking treatment. In addition, EO water could effectively disinfect the platform of fish retailer in traditional markets and fish markets.     

Determination of cadmium and lead in fruit juices by stripping chronopotentiometry and comparison of two sample pretreatment procedures

A method for the determination of cadmium and lead in apple and pear fruit juices by stripping chronopotentiometry (SCP) is described. The results obtained after previous digestion of the sample with concentrated sulphuric acid and dry-ashing (sample pretreatment procedure “A”) and after a treatment with concentrated hydrochloric acid followed by heating in hot water bath (sample pretreatment procedure “B”) were compared. The sample pretreatment procedures were tested on real fruit juice samples and all data confirmed that the efficiency of procedure A was greater than procedure B. In particular, for samples treated with procedure A, good linearity was obtained in the range of examined concentration as is shown by the determination coefficients that were 0.998 (n = 4) for cadmium and 0.996 (n = 4) for lead. Recoveries of 86–104% for cadmium and of 87–102% for lead were obtained from a sample spiked at different levels. The accuracy was also evaluated by means of a matching reference sample of spiked skim milk powder (BCR 150—Community Bureau of Reference) to prove the reliability of the method. The detection limits were 2.0 ng g⁻¹ for cadmium and 4.8 ng g⁻¹ for lead. The relative standard deviations (mean of nine determinations), evaluated on a real sample, were 7.8% and 6.5% respectively. The average content was in the range not detectable–3.0 ng g⁻¹ for cadmium and 8.2–21.3 ng g⁻¹ for lead.