H7 In Samples Research Articles

Sustained-release modeling of clove essential oil in brine to improve the shelf life of Iranian white cheese by bioactive electrospun zein

In this study, the sustained-release of clove essential oil (CEO) loaded into the structure of electrospun zein was used as a biopreservative to extend the shelf life of Iranian white cheese. CEO was loaded at levels of 5, 7.5, and 10% w/w in the structure of electrospun nanofibers. In this study, a concentration of 35% w/v zein was used to produce electrospun fibers, and in the field emission scanning electron microscope (FESEM) it was observed that by increasing the loading of CEO from 5 to 10% w/w in the fiber structure, their diameter decreased from 517.96 ± 41.57 nm to 457.88 ± 32.45 nm. Although increasing the level of CEO reduced the diameter of the electrospun nanofibers, Young's modulus, tensile strength, and a higher level of CEO increased elongation at break of the films. The results of mechanical properties showed that by increasing the amount of CEO application in the structure of electrospun zein nanofibers from 5 to 10% w/w tensile strength from 8.18 ± 0.62 to 4.43 ± 0.86 MPa, and Young's modulus from 38.25 ± 2.81 to 27.25 ± 3.48 MPa decreased. Successful encapsulation of CEO in designed structures and the absence of adverse bonds between the encapsulant material (zein) and the core (CEO) were confirmed by the Fourier-transform infrared spectroscopy (FTIR) test. The in vitro sustained-release of the CEO in 8% w/v brine during 45 days of storage at 4 °C was modeled. The Fickian diffusion was the dominant release mechanism of the CEO and the Peppas-Sahlin model was the best model describing the essential oil release behavior. The electrospun films containing CEO were well able to suppress the growth of Listeria monocytogenes and Escherichia coli O157: H7 in samples of Iranian white cheese kept in 8% brine for 45 days at 4 °C. The samples treated with the electrospun film containing 7.5% w/w of CEO had the highest acceptability among different treatments.

Outbreak of Shiga toxin-producing Escherichia coli O157:H7 linked to raw drinking milk resolved by rapid application of advanced pathogen characterisation methods, England, August to October 2017.

An outbreak of Shiga toxin-producing Escherichia coli (STEC) O157:H7 occurred on the Isle of Wight between August and October 2017. Of the seven cases linked to the outbreak, five were identified through the statutory notification process and two were identified through national surveillance of whole genome sequencing data. Enhanced surveillance questionnaires established a common link to a farm, and link to the likely food vehicle, raw drinking milk (RDM). Microbiological investigations, including PCR, identified the presence of STEC O157:H7 in samples of RDM. Analysis of core genome single nucleotide polymorphism (SNP) data of STEC O157:H7 from human stool specimens, animal faecal samples and RDM demonstrated a one SNP difference between isolates, and therefore close genetic relatedness. Control measures that were put in place included suspension of sales and recall of RDM, as well as restrictions on public access to parts of the farm. Successful integration of traditional epidemiological surveillance and advanced laboratory methods for the detection and characterisation of STEC O157:H7 from human, animal and environmental samples enabled prompt identification of the outbreak vehicle and provided evidence to support the outbreak control team’s decision-making, leading to implementation of effective control measures in a timely manner.

Electrochemical Detection of E. coli O157:H7 in Water after Electrocatalytic and Ultraviolet Treatments Using a Polyguanine-Labeled Secondary Bead Sensor.

The availability of clean drinking water is a significant problem worldwide. Many technologies exist for purifying drinking water, however, many of these methods require chemicals or use simple methods, such as boiling and filtering, which may or may not be effective in removing waterborne pathogens. Present methods for detecting pathogens in point-of-use (POU) sterilized water are typically time prohibitive or have limited ability differentiating between active and inactive cells. This work describes a rapid electrochemical sensor to differentially detect the presence of active Escherichia coli (E. coli) O157:H7 in samples that have been partially or completely sterilized using a new POU electrocatalytic water purification technology based on superradicals generated by defect laden titania (TiO2) nanotubes. The sensor was also used to detect pathogens sterilized by UV-C radiation for a comparison of different modes of cell death. The sensor utilizes immunomagnetic bead separation to isolate active bacteria by forming a sandwich assay comprised of antibody functionalized secondary magnetic beads, E. coli O157:H7, and polyguanine (polyG) oligonucleotide functionalized secondary polystyrene beads as an electrochemical tag. The assay is formed by the attachment of antibodies to active receptors on the membrane of E. coli, allowing the sensor to differentially detect viable cells. Ultravioloet (UV)-C radiation and an electrocatalytic reactor (ER) with integrated defect-laden titania nanotubes were used to examine the sensors’ performance in detecting sterilized cells under different modes of cell death. Plate counts and flow cytometry were used to quantify disinfection efficacy and cell damage. It was found that the ER treatments shredded the bacteria into multiple fragments, while UV-C treatments inactivated the bacteria but left the cell membrane mostly intact.

Dual FITC lateral flow immunoassay for sensitive detection of Escherichia coli O157:H7 in food samples

A pattern of signal amplification lateral flow immunoassay (LFIA) for pathogen detection, which used fluorescein isothiocyanate (FITC) labeled antigen and antibody for dual FITC-LFIA was developed. Escherichia coli O157:H7 (E.coli O157:H7) was selected as the model analyte. In the signal amplification LFIA method, FITC was mixed with sample culture medium, with the presence of E.coli O157:H7 in the samples, the bacteria could emit a yellow-green fluorescence after incubation, creating a fluorescent antigen probe. This antigen probe was added to LFIA, which already contained E.coli O157:H7 monoclonal antibodies-FITC (McAb-E.coli O157:H7-FITC) dispersed in the conjugate pad. Another E.coli O157:H7 McAb was the test line, and goat anti-mouse IgG antibody was the control line in nitrocellulose (NC) membrane. The visual limit of detection (LOD) of the strip for qualitative detection was 105 CFU/mL while the LOD for semi-quantitative detection could down to 104 CFU/mL by using scanning reader. Signal amplification LFIA was perfectly applied to the detection of food samples with E.coli O157:H7. The LOD was substantially improved to 1 CFU/mL of the original bacterial content after pre-incubation of the bread, milk and jelly samples in broth for 10, 8 and 8h respectively. The results of this method was more sensitive by 10-fold than the conventional colloidal gold (CG) based strips and comparable to the traditional ELISA. This simple, low-cost and easy to be popularized method served as a significant step towards the development of monitoring food-borne pathogens in food-safety testing.

Simple sensitive rapid detection of Escherichia coli O157:H7 in food samples by label-free immunofluorescence strip sensor

A simple, one-step, rapid method to detect Escherichia coli O157: H7 (E. coli O157: H7) using a label-free immunofluorescence strip sensor is presented. Fluorescein isothiocyanate (FITC) was added to the sample culture medium to prepare the fluorescent probe for the label-free strip sensor. With the presence of E. coli O157: H7 in the samples, the bacteria could emit a yellow-green fluorescence after incubation and maintain good affinity to the monoclonal antibodies (McAb) against E. coli O157: H7. The direct-type immunofluorescence strip sensor was based on the binding between fluorescent bacteria and the unlabeled McAb immobilized at the test line in nitrocellulose membrane (NC membrane) reaction zone. The visual limit of detection (LOD) of the strip for qualitative detection was 106cells/mL while the LOD for semi-quantitative detection could go down to 105cells/mL by using scanning reader. The LOD was substantially improved to 1cells/mL of the original bacterial content after pre-incubation of the bread, milk and jelly samples in broth for 10, 10 and 8h respectively, which was competitive to some current rapid E. coli O157: H7 detection methods. Besides the obvious advantages, including reduced detection time and operation procedures, the results of this method meet the various detection requirements for E. coli O157: H7 and are comparable to the traditional enzyme-linked immunosorbent assay (ELISA) and double antibody sandwich gold-labeled strips. This is the first report of semi-quantitative immunofluorescence strip for directly detecting foodborne pathogen using only one unlabeled antibody. All detections could be achieved in less than 5min. In addition, this simple, low-cost and easy to be popularized method served as a significant step towards the development of monitoring foodborne pathogens in food-safety testing.

Chemiluminescent aptasensor capable of rapidly quantifying Escherichia Coli O157:H7

Cost-effective and easy-to-use biosensor was developed for the rapid quantification and monitoring of Escherichia (E.) Coli O157:H7 in sample using E. Coli O157:H7 aptamer, graphene oxide (GO)/iron nanocomposites, and guanine chemiluminescence detection. E. Coli O157:H7 aptamer-conjugated 6-carboxyfluorescein (6-FAM) with excellent specificity captured E. Coli O157:H7 in a sample when the mixture was incubated for 1h at 37°C. Free E. Coli O157:H7 aptamers remaining in sample after the incubation were removed with GO/iron nanocomposites based on the principle of π–π stacking interaction between free aptamer and GO/iron nanocomposites. Then, E. Coli O157:H7 bound with aptamer-conjugated 6-FAM in sample emitted strong light when guanine chemiluminescent reagents (e.g., 3,4,5-trimethoxylphenylglyoxal hydrate, Tetra-n-propylammonium hydroxide) were added in the sample. The strength of light emitted in guanine chemiluminescence reaction was proportionally enhanced with the increase of E. Coli O157:H7 concentration. The limit of detection (LOD) of biosensor capable of quantifying E. Coli O157:H7 with good accuracy, precision, and reproducibility was as low as 4.5×103cfu/ml. We expect that the rapid analytical system can be applied in the field of food safety as well as public health.

Detection of rfbO157 and fliCH7 Genes in Escherichia coli Isolated from Human and Sheep in Basrah Province

This study aimed to detect Escherichia coli O157:H7 in samples from patients with diarrhea, sheep milk, and sheep drinking water by growth on selective media, biochemical tests, latex agglutination test, and PCR technique. During the period from November 2008 to May 2009, a total of 340 samples were collected; stools diarrhoeic patients (125), healthy sheep feces (125), milk (45), and sheep drinking water samples (45). Samples were screened for the presence of non-sorbitol fermenting colonies (NSF) on sorbitol MacConkey agar supplemented with cefixime and potassium tellurite (CT-SMAC). Out of the 340 samples, 58 (46.4%) stool from diarrhea, 98 (78.4%) from sheep feces, 19 (42.2%) milk, and 17 (37.7%) drinking water were nonsorbitol fermenting. E. coli isolates were serotyped as E. coli O157:H7 by latex agglutination test, 3 (5.7%) isolates of stool from diarrhea patients, 7 (7.3%) isolates of sheep feces, 2 (11.1%) isolates of milk, and none of drinking water samples were positive. All latex agglutination positive isolates were positive to PCR test except one isolate from sheep feces. PCR positive isolates were produced enterohemolysin on sheep blood agar, and were found resistant to at least 3 antibiotics. The highest rate of E. coli O157:H7 isolation was observed in April (7.6%) followed by December (4.7%) , while those of sheep feces showed the highest rate in March (10%) followed by April (8.6%).

Inoculation of beef with low concentrations of Escherichia coli O157:H7 and examination of factors that interfere with its detection by culture isolation and rapid methods.

Currently used industry testing programs require the ability to detect Escherichia coli O157:H7 in samples of beef trim or ground beef at levels as low as 1 CFU/375 g. We present a reliable protocol for generating a control inoculum for verification testing at this low concentration and evaluate its use. Results show that half of all samples received no cells when 1 CFU was the target concentration and that targets greater than 3 CFU were much more reliable. Detection by culture isolation and two commercial assays, Qualicon BAX-MP and BioControl GDS, detected 94% ± 11%, 92% ± 10%, and 92% ± 7% of samples inoculated with 5.4 CFU (range 1 to 9 CFU), respectively. We also examined the effect of background aerobic plate count (APC) bacteria and fat content effects on the detection of E. coli O157:H7. At APC concentrations below 6 log CFU/g, the rapid methods detected all beef trim samples inoculated with 26 CFU of E. coli O157:H7 per 65 g. At an APC of 6.7 log CFU/g, culture, BAX-MP, and GDS detected 100, 75, and 13%, respectively, of inoculated samples. Neither commercial method detected E. coli O157:H7 in the samples when APC was 7.7 log CFU/g, whereas culture was able to detect 63% of E. coli O157:H7 in the samples when APC was at this concentration. Increased fat content correlated with decreasing recovery of immunomagnetic separation beads, but this was not observed to interfere with detection of E. coli O157:H7.

Validation of low-volume enrichment protocols for detection of Escherichia coli O157 in raw ground beef components, using commercial kits.

Testing of beef destined for use in ground beef products for the presence of Escherichia coli O157:H7 has become an important cornerstone of control and verification activities within many meat supply chains. Validation of the ability of methods to detect low levels of E. coli O157:H7 is critical to confidence in test systems. Many rapid methods have been validated against standard cultural methods for 25-g samples. In this study, a number of previously validated enrichment broths and commercially available test kits were validated for the detection of low numbers of E. coli O157:H7 in 375-g samples of raw ground beef component matrices using 1 liter of enrichment broth (large-sample:low-volume enrichment protocol). Standard AOAC International methods for 25-g samples in 225 ml of enrichment broth, using the same media, incubation conditions, and test kits, were used as reference methods. No significant differences were detected in the ability of any of the tests to detect low levels of E. coli O157:H7 in samples of raw ground beef components when enriched according to standard or large-sample:low-volume enrichment protocols. The use of large-sample:low-volume enrichment protocols provides cost savings for media and logistical benefits when handling and incubating large numbers of samples.

Effect of chemical sanitizer combined with modified atmosphere packaging on inhibiting Escherichia coli O157:H7 in commercial spinach

Escherichia coli O157:H7 contaminated spinach has recently caused several outbreaks of human illness in the USA and Canada. However, to date, there has been no study demonstrating an effective way to eliminate E. coli O157:H7 in spinach. Therefore, this study was conducted to investigate the effect of chemical sanitizers alone or in combination with packaging methods such as vacuum and modified atmosphere packaging (MAP) on inactivating E. coli O157:H7 in spinach during storage time. Spinach inoculated with E. coli O157:H7 was packaged in four different methods (air, vacuum, N 2 gas, and CO 2 gas packaging) following treatment with water, 100 ppm chlorine dioxide, or 100 ppm sodium hypochlorite for 5 min at room temperature and stored at 7±2 °C. Treatment with water did not significantly reduce levels of E. coli O157:H7 in spinach. However, treatment with chlorine dioxide and sodium hypochlorite significantly decreased levels of E. coli O157:H7 by 2.6 and 1.1 log 10 CFU/g, respectively. Levels of E. coli O157:H7 in samples packaged in air following treatments grew during storage time, whereas levels were maintained in samples packaged in other packaging methods (vacuum, N 2 gas, and CO 2 gas packaging). Therefore there were significant differences (about 3–4 log) of E. coli O157:H7 populations between samples packed in air and other packaging methods following treatment with chemical sanitizers after 7 days storage. These results suggest that the combination of treatment with chlorine dioxide and packaging methods such as vacuum and MAP may be useful for improving the microbial safety of spinach against E. coli O157:H7 during storage.

Rapid Detection of Escherichia coli O157:H7 Inoculated in Ground Beef, Chicken Carcass, and Lettuce Samples with an Immunomagnetic Chemiluminescence Fiber-Optic Biosensor

A biosensor was evaluated with regard to its usefulness in the rapid detection of Escherichia coli O157:H7 inoculated in ground beef, chicken carcass, and romaine lettuce samples. The biosensor consisted of a chemiluminescence reaction cell, a fiber-optic light guide, and a luminometer linked to a personal computer in conjunction with immunomagnetic separation. The samples inoculated with E. coli O157:H7 were first centrifuged and suspended in buffered peptone water and then incubated with anti–E. coli O157 antibody–coated magnetic beads and horseradish peroxidase(HRP)–labeled anti–E. coli O157 antibodies to form antibody-coated bead–bacterium–HRP-labeled antibody sandwich complexes. Finally, the sandwich complexes were separated from the samples in a magnetic field and reacted with luminol in the reaction cell. The number of E. coli O157:H7 cells was determined by collecting the HRP-catalyzed chemiluminescence signal from the bead surface through a fiber-optic light guide and measuring the signal with a luminometer. The chemiluminescence biosensor was specific for E. coli O157:H7 in samples containing other bacteria, including Salmonella Typhimurium, Campylobacter jejuni, and Listeria monocytogenes. The chemiluminescence signal was linear on a log scale from 102 to 105 CFU of E. coli O157:H7 per ml in samples. Detection could be completed within 1.5 h without any enrichment. The detection limits for ground beef, chicken carcass, and lettuce samples were 3.2 × 102, 4.4 × 102, and 5.5 × 102 CFU of E. coli O157:H7 per ml, respectively.

IMMUNOMAGNETIC‐ELECTROCHEMILUMINESCENT DETECTION OF E. COLI O157:H7 IN GROUND BEEF1

An immunomagnetic electrochemiluminescence approach using a commercially available instrument was developed to detect the presence of E. coli O157:H7 in samples of ground beef. The approach can detect E. coli O157:H7 that have been incurred with 0.05 CFU of the bacterium per gram of beef after an enrichment for 18 h at 37C. Results of comparison tests indicate that the approach is at least 100 times more sensitive than the Reveal dipstick method with a probable false positive rate less than 4–5%. To enhance the adaptability of the approach, a heat treatment was introduced to eliminate the possibility of generating aerosols of live pathogens during the sample loading process. To minimize the carryover of contaminants associated with the capture of E. coli O157:H7 by immuno magnetic beads (IMB), an immuno magnetic separation procedure was developed. The procedure involved the use of a magnetic field to first retain the IMB together with captured bacteria on a paramagnetic column. After washing away the contaminants, the target bacteria were then released from the column by removing the magnetic field. With the proven sensitivity and improved sample treatment procedures, the ECL approach could be considered as an attractive alternative for screening the presence of E. coli O157:H7 in foods.