Enumeration Of Enterobacteriaceae Research Articles

Sanitary Investigation on Microbial and Heavy Metals of Grilled Fish Sold in the Night Street Market of Daloa (Ivory Coast)

The quality of food sold in the street is a major public health concern. This research was conducted to study the sanitary quality of grilled fish sold in the streets of three districts of Daloa. Seasoned and unseasoned grilled fish were collected in 3 districts of Daloa, namely Commerce, Soleil and Tazibouo. A total of 150 unseasoned and 150 seasoned grilled fish were collected and analyzed for the detection and enumeration of Enterobacteriaceae, Staphylococcus aureus and fungal flora. We also tested for the presence of heavy metals such as lead (Pb), cadmium (Cd), mercury (Hg) and zinc (Zn). The antibiotic resistance of S. aureus was also checked. The results showed that Enterobacteriaceae represented the highest level of microbial load in the samples of seasoned grilled fish from the 3 districts. Total coliform levels ranged from (7.05 ± 2.1) x 103 colony-forming units (CFU)/g to (4.1 ± 1.9) x 105 CFU/g and from 2.8 ± 3.3 x102 to 3.4 ± 2.1x103 CFU/g for thermotolerant coliforms. The presence of heavy metals showed higher values in seasoned grilled fish than in unseasoned grilled fish. The highest values of heavy metals are Al (1.98±0.120 mg/kg), Pb (1.46±0.23 mg/kg), Cu (0.7±0.1 mg/kg), Hg (0.40±0.201 mg/kg) and Hg (0.9±0.201 mg/kg). The microbiological quality of both types of fish samples was unsatisfactory. The presence of S. aureus resistance was noticed with 80% resistance to vancomycin, lincomycin, gentamycin and erythromycin but all strains tested were sensitive to imipenem.

Impact of organic dairy cattle manure on environmental and egg microbiology of organic free-range laying hens

The popularity of free-range and organic poultry has increased in recent years due to consumer demands. A study was conducted to evaluate how the presence of organic dairy cattle manure could influence the environmental and egg microbiology of organic free-range laying flocks. A flock of brown egg layers was split and maintained in a rotational paddock grazing schedule of which was free of or exposed to organic dairy manure. Environmental and egg sampling occurred approximately every 8 wk between 20 and 44 wk of age for a presence of Listeria spp., Campylobacter spp., Salmonella spp., and enumeration of Enterobacteriaceae. There was no significant difference in the occurrence of Listeria spp., Campylobacter spp. and Salmonella spp. in environmental and egg samples between integrated and control grazing organic free-range flocks. A random sampling of Enterobacteriaceae colonies produced 211 viable isolates for biochemical identification. Seventeen genera, species, or serotypes were identified. There was a greater (P < 0.05) prevalence of total coliforms found on shell emulsion, egg contents, nest box straw, and forage samples from integrated organic free-range compared to control organic free-range flocks. Seasons of the year impacted microbial levels recovered from environmental and egg samples, with which summer having the highest level of all populations monitored. Additional studies are needed to fully understand the effect of mixed production rotational grazing on the prevalence of pathogens and Enterobacteriaceae on organic nest-run eggs and the grazing environment.

The use of alkaline phosphatase and possible alternative testing to verify pasteurisation of raw milk, colostrum, dairy and colostrum-based products.

Pasteurisation of raw milk, colostrum, dairy or colostrum‐based products must be achieved using at least 72°C for 15 s, at least 63°C for 30 min or any equivalent combination, such that the alkaline phosphatase (ALP) test immediately after such treatment gives a negative result. For cows’ milk, a negative result is when the measured activity is ≤ 350 milliunits of enzyme activity per litre (mU/L) using the ISO standard 11816‐1. The use and limitations of an ALP test and possible alternative methods for verifying pasteurisation of those products from other animal species (in particular sheep and goats) were evaluated. The current limitations of ALP testing of bovine products also apply. ALP activity in raw ovine milk appears to be about three times higher and in caprine milk about five times lower than in bovine milk and is highly variable between breeds. It is influenced by season, lactation stage and fat content. Assuming a similar pathogen inactivation rate to cows’ milk and based on the available data, there is 95–99% probability (extremely likely) that pasteurised goat milk and pasteurised sheep milk would have an ALP activity below a limit of 300 and 500 mU/L, respectively. The main alternative methods currently used are temperature monitoring using data loggers (which cannot detect other process failures such as cracked or leaking plates) and the enumeration of Enterobacteriaceae (which is not suitable for pasteurisation verification but is relevant for hygiene monitoring). The inactivation of certain enzymes other than ALP may be more suitable for the verification of pasteurisation but requires further study. Secondary products of heat treatment are not suitable as pasteurisation markers due to the high temperatures needed for their production. More research is needed to facilitate a definitive conclusion on the applicability of changes in native whey proteins as pasteurisation markers.

Microbiologique Microfilm™ EBEc - a Rapid Method for the Simultaneous Enumeration of Enterobacteriaceae and Escherichia coli.

The Enterobacteriaceae and generic Escherichia coli are routinely enumerated in foods as part of product release criteria, or in the case of swabs, for environmental monitoring. Microbiologique Microfilm™ EBEc is intended to provide a rapid and easy-to-use method for simultaneous enumeration of Enterobacteriaceae and E. coli on foods and environmental surfaces. Methods: This study evaluated the performance of Microfilm™ EBEc against ISO methods (ISO 21528-2:2017 for Enterobacteriaceae and ISO 16649-2: 2001 for E. coli) in 20 food matrixes and two environmental surfaces. Inclusivity, exclusivity, lot-to-lot reproducibility, ruggedness and stability studies were also performed on Microfilm™ EBEc. No significant differences and high correlation coefficients (R2) were observed between the Microfilm™ EBEc and the corresponding ISO methods in spiked food matrixes and environmental samples. Inclusivity studies showed expected results for all the E. coli and Enterobacteriaceae strains tested. In terms of exclusivity testing, all the strains tested failed to grow on Microfilm™ EBEc. A lot-to-lot study showed no significant differences in mean difference (log10) counts among the three lots of the Microfilm™ EBEc. Ruggedness studies showed no significant differences in mean difference (log10) counts at varying incubation temperatures and times. Stability studies on the Microfilm™ EBEc showed that it is stable at 2-25°C for 12 months, and at 45°C for 6 weeks. The results of this study indicate that the Microfilm™ EBEc is equivalent to the corresponding ISO methods for enumeration of Enterobacteriaceae and E. coli. Highlights: Microfilm™ EBEc offers a convenient and relatively fast test method for simultaneous enumeration of Enterobacteriaceae and E. coli in 24 h and has an advantage over the corresponding ISO methods that require two assays on the same sample for enumeration ofEnterobacteriaceae and E. coli Gram-negative indicator groups.

Validation by an interlaboratory collaborative trial of EN ISO 21528 - microbiology of the food chain - horizontal methods for the detection and enumeration of Enterobacteriaceae

The methods for the detection and enumeration of Enterobacteriaceae, described in EN ISO 21528, parts 1 and 2, were validated by order of the European Commission under the mandate M/381. Fourteen laboratories from seven European countries participated in the collaborative trials, organized by The Netherlands Food and Consumer Product Safety Authority (Wageningen/Utrecht, The Netherlands).Five different matrices from different food categories were selected to be tested in the collaborative trials, in order to validate the method horizontally, according to ISO 16140. The matrices included meat, tiramisu, infant formula, liquid egg, smoked salmon (detection method only) and animal feed (enumeration method only). The raw meat and liquid egg were naturally contaminated and the other matrices were artificially contaminated with a cocktail of four different Enterobacteriaceae strains.The samples used in the trial were tested for homogeneity and stability before distribution. The method for detection of Enterobacteriaceae showed a specificity and sensitivity above 95% for all matrices. The method for the enumeration had a repeatability limit r of 0.37 (expressed as a difference between log10-transformed test results) and a reproducibility limit R of 0.87 (expressed as a difference between log10-transformed test results).The validation data were incorporated in the newly published ISO standards EN ISO 21528:2017- Microbiology of the food chain - Horizontal methods for the detection and enumeration of Enterobacteriaceae - Part 1: Detection method, and Part 2: Colony-count technique.

Microbiological, chemical-physical and sensory characteristics of Fabriano salami from pigs fed Oregano vulgaris extract.

This study aimed at evaluating the effect of Oregano vulgaris extract supplementation of heavy pig diet on the microbiological, chemical-physical and sensory characteristics of Fabriano salami. Fabriano salamis were produced processing meat derived from pigs belonging to two dietary groups of 16 pigs each: CTRL group fed a commercial pelleted feed, and OR group fed the CTRL diet integrated with 0.2% of oregano extract. Ten salamis per group were collected at 0, 7, 20 and 45 days of ripening and analysed for total viable count, enumeration of Enterobacteriaceae, Enterococcus spp, coagulase negative and positive staphylococci, Lactobacillus spp., Lactococcus spp., presence of Salmonella spp. and Listeria monocytogens, pH, water activity (aw) and colour (CIE L*a*b* colour system). At the end of the ripening (day 45) chemical composition (AOAC, 1990), total antioxidant capacity (ORACFL), thiobarbituric reactive substances (TBARS), total phenolic content (TPC) (Folin-Ciocalteu method) and consumer tests were performed. The integration of oregano extract in pig diets did not interfere with the microbial evolution, chemical composition, pH and aw. After 20 days of ripening the OR salami were redder then CTRL samples, which were also yellower at the end of ripening. Furthermore, the diet has significantly improved the oxidative status, the polyphenolic content and antioxidant power in OR salami. The consumers gave a higher score to OR salami in informed condition.

Impact of oral and intramuscular administration amoxicillin on the selection of amoxicillin-resistant &lt;i&gt;Enterobacteriaceae&lt;/i&gt; in the digestive flora of piglets

Objective: The aim of this study was to evaluate the level of selection of amoxicillin-resistant Enterobacteriaceae in the digestive microbiota of piglets during oral and intramuscular administration of amoxicillin. Methodology and Results: Enumeration of Enterobacteriaceae was carried out on MacConkey agar with and without amoxicillin. E. coli isolates were identified and subjected to antimicrobial susceptibility test. The percentages of amoxicillin-resistant Enterobacteriaceae before treatment were between 10-13% for the three groups of piglets. After starting treatment of amoxicillin, from day 1 to day 4, the percentages of resistant Enterobacteriaceae were between 54 to 81% for the intramuscular treated groups and 58 to 87% for those treated orally. In the control group, percentage was 11%. During days of treatments, the percentage of amoxicillin-resistant E. coli strains to the associated antibiotics evolved during the treatments. In the control piglets, the percentages of E. coli resistant antibiotics did not increase. Conclusion and application of findings: Administration of oral amoxicillin resulted in a greater selection of resistant enterobacteria in the digestive microbiota of piglets than those induced by the intramuscular route. Escherichia coli , an important strain in animal and human pathology, was the target of detection in this selection of these resistant enterobacteria. They constitute a public health risk due to the transfer of resistance genes from pathogenic bacteria of porcine origin to the human digestive flora via the food chain. The results obtained from this study are part of a bacterial resistance monitoring policy that will prevent pig farmers from Cote d’Ivoire, the majority user of these antibiotics. Keywords: Piglets, Enterobacteriaceae , Amoxicillin, Resistance, Digestive microbiota

Development of quantitative real-time PCR for detection and enumeration of Enterobacteriaceae

The family Enterobacteriaceae, members of which are widely distributed in the environment, includes many important human pathogens. In this study, a rapid real-time PCR method targeting rplP, coding for L16 protein, a component of the ribosome large subunit, was developed for enumerating Enterobacteriaceae strains, and its efficiency was evaluated using naturally contaminated food products. The rplP-targeted real-time PCR amplified Enterobacteriaceae species with Ct values of 14.0–22.8, whereas the Ct values for non-Enterobacteriaceae species were >30, indicating the specificity of this method for the Enterobacteriaceae. Using a calibration curve of Ct=−3.025 (log CFU/g)+37.35, which was calculated from individual plots of the cell numbers in different concentrations of 5 Enterobacteriaceae species, the rplP-targeted real-time PCR was applied to 51 food samples. A <1log difference between the real-time PCR and culture methods was obtained in a majority of the food samples (81.8%), with good correlation (r2=0.8285). This study demonstrated that the rplP-targeted real-time PCR method could detect and enumerate Enterobacteriaceae species in foods rapidly and accurately, and therefore, it can be used for the microbiological risk analysis of foods.

Bacteriological Quality and Risk Factors for Contamination of Raw Mixed Vegetable Salads Served in Collective Catering in Abidjan (Ivory Coast)

The aim of this study was to determine the bacteriological quality of raw mixed vegetable salads served in collective catering in Abidjan and to assess the factors likely to favor contamination. An investigation was conducted to determine the practical risk of contamination. A total of 306 samples were collected and analyzed for the detection and enumeration of Enterobacteriaceae, E. coli and Salmonella according to standard microbiology methods. The most important risk factors were lack of training (96.7%), decontamination (84.0%), cross-contamination of vegetables in contact with fresh chicken or fish (76.7%) and salads that were uncovered and unprotected, thus exposed to the air and at ambient temperature for sale (97.7%). Theprevalence was 100%, 77.8% and 2.6% respectively for Enterobacteriaceae, E. coli and Salmonella. The average load of Enterobacteriaceae and E. coli was 4.93 log10 (CFU/g) and 1.89 log10 (CFU/g), respectively. Unsatisfactory microbiological quality samples were 52.3%. The results showed a failure of the sanitary quality in more than half of the products. Raising awareness in all of the workers in the food chain for vegetable salads is thus necessary to protect the health of the consumers.

Risk Assessment in the Recovery of Food for Social Solidarity Purposes: Preliminary Data.

The most recent study, conducted by Politecnico of Milan, on food surplus management in Italy shows that in the Italian food supply chain the food surplus is around 5.5 million tons/year, and the amount of food wasted is around 5.1 million tons/year. During 2015, the charitable organizations (COs) belonging to the Italian Food Bank Network, active in recovering and distributing food for social solidarity’s purposes, reused 381,345 tons of food from 2292 donors. The main supplying sources of the Banco Alimentare Network are: food industries, organized large-scale retail trade and collective catering service. The aim of this study was to analyze several aspects of the food surplus recovery thanks to the collaboration with the Banco Alimentare Foundation Onlus and Caritas Italiana. In particular, two main features were analyzed in the food recovery chain: the microbiological profiles of specific food categories recovered from catering service with the aim to evaluate their conformity in relation to food safety and process criteria. For this purpose 11 samples were analyzed in four different moments: T0, same day of the collection; T1, after four hours of storage at 4°C; T2, 24 hours from the collection (storage at 4°C); T3, after four days at frozen storage (-18°C). For all samples several microbiological parameters were investigated: enumeration of mesophilic aerobic bacteria (AFNOR 3M 01/1-09/89), enumeration of Enterobacteriaceae (AFNOR 3M 01/06-09/97), enumeration of E. coli (AFNOR 3M 01/08-06/01), enumeration of coagulase-positive Staphylococci AFNOR 3M 01/9-04/03 A), enumeration of Bacillus cereus (UNI EN ISO 7932:2005), research of Salmonella spp. [UNI EN ISO 6579 (2008b)], and research of Listeria monocytogenes [AFNOR BRD 07/4-09/98 (AFNOR, 2010a)]. Furthermore, the volunteer’s knowledge on the correct hygienic procedures during the recovery was evaluated by the 71 questionnaires with the aim to prevent foodborne diseases. The results show that the recovery of surplus from catering service and their reuse at COs should be planned with correct procedures, and the volunteer’s knowledge on the hygienic aspects appears to be a critical point. The recovery and the charitable activities require an appropriate assessment and careful risk analysis, in order to manage the complexity of no profit organization.

Evaluation of the Dry Sheet Medium (Compact Dry ETBR) for Enumerating Enterobacteriaceae in Meat Samples

Compact Dry ETB(R) (CD-ETB, ETB; Enterobacteriaceae) , a ready-to-use and self-diffusible dry medium sheet culture system for the detection and enumeration of Enterobacteriaceae, was evaluated. A total of 35 Enterobacteriaceae strains, which were studied for the inclusivity study, grew and formed red-purple colored colonies on the CD-ETB(R). When 17 gram-negative bacteria other than Enterobacteriaceae were inoculated for the exclusivity study, 2 strains grew and formed red-purple colonies, 9 strains formed colorless colonies, and 6 strains failed to grow. A total of 43 gram-positive bacteria and 3 yeasts failed to grow. The CD-ETB method was compared with the Violet Red Bile Glucose (VRBG) agar method according to ISO 21528-2 and the 3M Petrifilm(TM) EB (3M-EB(R)) method in 53 meat samples. The correlation coefficients between CD-ETB(R) and VRBG agar, CD-ETB(R) and 3M-EB(R), and 3M-EB(R) and VRBG agar were 0.962, 0.992 and 0.960, respectively.

Enterobacteriaceae and Related Organisms Isolated from Shell Eggs Collected During Commercial Processing

In the United States, commercial shell eggs are washed and graded before retail. Since passage of the Egg Products Inspection Act in 1971, processing guidelines have been set to ensure that external and internal characteristics are maintained. However, less is known about how commercial processing affects the safety of shell eggs. To identify enteric bacteria entering plants and persisting throughout processing, eggs were collected from 3 US commercial shell egg-processing plants on 3 separate visits. On each plant visit, 12 eggs were collected from each of 12 sites along the processing line: accumulator, prewash rinse, first washer, second washer, sanitizer rinse, dryer, oiler, check detection/scales, 2 egg grader/packer head lanes, rewash belt entrance, and rewash belt exit. Each egg was sampled by a rinse technique, and the rinsate was plated onto violet red bile glucose agar with overlay for the detection and enumeration of Enterobacteriaceae. From each plate, up to 5 colonies were randomly selected and isolated for identification to genus or species by using biochemical tests. Several genera and species were detected at each of the 3 plants. Sites from which the greatest numbers of isolates were identified were those collected from eggs during preprocessing (accumulator, prewash rinse) or from eggs judged as dirty (rewash belt entrance or exit). Sites yielding the smallest number of isolates were those during or at the end of processing. Escherichia coli and Enterobacter spp. were isolated from each of the 9 plant visits. Other genera isolated from at least 1 of the 3 plants included Cedecea, Citrobacter, Erwinia, Hafnia, Klebsiella, Kluyvera, Leclercia, Morganella, Proteus, Providencia, Rahnella, Salmonella, and Serratia. Non-Enterobacteriaceae isolated and identified included Aeromonas, Chryseomonas, Listonella, Pseudomonas, Sphingobacterium, Vibrio, and Xanthomonas. All of the genera and species were recovered less frequently from fully processed eggs than from unwashed eggs, indicating that shell eggs are less contaminated with bacteria as a result of commercial washing procedures.

Isolation of Enterobacter sakazakii and other Enterobacteriaceae from powdered infant formula milk and related products

The presence of Enterobacter sakazakii and other Enterobacteriaceae was surveyed in 82 powdered infant formula milk (IFM) and 404 other food products. The presence of Ent. sakazakii was detected using the conventional method (growth on violet red bile glucose agar plus yellow pigment production on TSA) and a new chromogenic medium (Druggan–Forsythe–Iversen agar, DFI) which enables results to be obtained 2 days earlier than the conventional method. Ent. sakazakii was isolated from 2/82 powdered IFM, 5/49 dried infant foods, 3/72 milk powder, 2/62 cheese products and various dry food ingredients, especially herbs and spices (40/122). Ent. sakazakii was isolated from 67 samples using the DFI medium, however only 19 of the samples were positive following the conventional method. The largest difference in isolation between the two methods was with dry food ingredients. Although Enterobacteriaceae were enumerated from one powdered IFM sample ( Klebsiella ozaenae, 200 cfu/g), 7/82 had detectable Enterobacteriaceae after enrichment in EE broth. Using the ISO 6579 2002 method and immuno-magnetic separation technique no Salmonella serovars were isolated from powdered IFM, dried infant foods or milk samples. Therefore hygienic production of powdered IFM and milk production as monitored by control of Salmonella and enumeration of Enterobacteriaceae did not control Ent. sakazakii.

5601998 Culture medium and device for detection and enumeration of Enterobacteriaceae : Mach Patrick A; Wickert Peter D; Adams Carl A St Paul, MN, United States

5601998 Culture medium and device for detection and enumeration of Enterobacteriaceae : Mach Patrick A; Wickert Peter D; Adams Carl A St Paul, MN, United States

Effect of Acidification with Encapsulated Lactate on Microbiological, Textural and Sensory Properties of Cooked, Comminuted Fish Loaves

A standard formulation for cooked, comminuted fish loaf was supplemented with encapsulated lactate (melting temperature =60oC) to assess the usefulness of this additive for shelf-life extension. Slices of standard (pH= 6.45) and acidified (pH =4.86) loaf were packaged in air and under vacuum, stored at 4oC and subjected to bacteriological analysis (total mesophilic and psychrotrophic plate counts, enumeration of Enterobacteriaceae and lactic acid bacteria) at weekly intervals. Texture profile analysis and sensory evaluation were also performed to determine the effect of acidification on the physico-chemical properties of the loaves. Use of encapsulated lactate in the formulation doubled the shelf-life from 2 to 4 weeks, due in part to strong inhibition of the Enterobacteriaceae. Texture profile analysis revealed that acidified fish loaf was softer and less rigid than the standard product. Cohesiveness scores determined by sensory evaluation confirmed this change in texture. Panelists also recorded undesir...

Acid injury of Escherichia coli.

Two strains of Escherichia coli sublethally treated in acidic environments showed an increased sensitivity to violet red bile agar. The death of cells, treated in tryptic soy broth which had been acidified with two inorganic and five organic acids, was followed on both nutrient agar and the selective medium violet red bile agar. The death rates determined on violet red bile agar were much faster than those established on nutrient agar. Therefore, it appeared that unreliable results may occur if selective media are used for the detection and enumeration of Enterobacteriaceae in acid foods.

Use of a modified MacConkey agar medium for the selective growth and enumeration of Enterobacteriaceae.

Use of a modified MacConkey agar medium for the selective growth and enumeration of Enterobacteriaceae.