scholarly journals Single tube confirmatory tests forEscherichia coli

1980 ◽  
Vol 85 (1) ◽  
pp. 51-57 ◽  
Author(s):  
Keyword(s):  
Escherichia Coli ◽  
False Positive ◽  
False Negative ◽  
Gas Production ◽  
Single Tube ◽  
E Coli ◽  
Peptone Water ◽  
Confirmatory Tests ◽  
Laboratory Trial ◽  
The Difference

SummaryIn a multi-laboratory trial, lauryl tryptose mannitol broth (LTMB) and minerals-modified glutamate medium with added tryptophan (MMGM + T) were compared as single tube tests for the confirmation ofEscherichia coliin water; the confirmed results were also compared with the production of gas from minerals-modified glutamate medium without added tryptophan (MMGM). LTMB and MMGM + T gave similar gas and indole results with about 90% of the water samples in most of the laboratories. When compared with the ‘correct’ results as judged by acid and gas production from lactose peptone water and indole from tryptone water, the difference in the rate of false positive reactions between LTMB and MMGM + T was insignificant; but LTMB gave a significantly lower rate of false negative reactions than MMGM + T. Gas production from MMGM without added tryptophan gave significantly higher rates of both false positive and false negative reactions. Lauryl sulphate is therefore a suitable inhibitory surfactant for use in single tubemedia for the confirmation ofE. coli, which can be recommended.

scholarly journals Membrane filtration media for the enumeration of coliform organisms and Escherichia coli in water: comparison of Tergitol 7 and lauryl sulphate with Teepol 610

1980 ◽  
Vol 85 (2) ◽  
pp. 181-191 ◽  
Author(s):  
Keyword(s):  
Escherichia Coli ◽  
Water Samples ◽  
False Positive ◽  
Membrane Filtration ◽  
Sodium Lauryl Sulphate ◽  
Standard Medium ◽  
E Coli ◽  
Laboratory Trial ◽  
Positive Results ◽  
Filtration Technique

SUMMARYIn a multi-laboratory trial with the mombrane filtration technique, three surfactants – Teepol 610 (T610), Tergitol 7 (T7) and sodium lauryl sulphate (LS) – were compared in media for the enumeration of coliform organisms and Escherichia coli in water. A total of 170 samples of water (87 raw and 92 marginally chlorinated) were examined for colony counts of coliform organisms, and 185 water samples (94 raw and 91 marginally chlorinated) for E. coli. Slight differences in the confirmed colony counts between the three media were noted, but few of these were observed consistently in every laboratory. In most laboratories, T7 gave slightly higher counts of E. coli than LS with chlorinated waters; a higher incidence of false-positive results for E. coli at 44 °C was also noted with T7. As there were no outstanding differences in the trial, sodium lauryl sulphate, which is chemically defined, cheap and readily available, is therefore recommended for use at a concentration of 0·1% instead of Teepol 610 in the standard medium for the enumeration of coliform organisms and E. coli in water by the membrane filtration technique.

Comparison of Enrichment Conditions for Rapid Detection of Low Numbers of Sublethally Injured Escherichia coli O157in Food

2009 ◽  
Vol 72 (9) ◽  
pp. 1862-1868 ◽  
Author(s):  
VICKY JASSON ◽  
ANDREJA RAJKOVIC ◽  
LEEN BAERT ◽  
JOHAN DEBEVERE ◽  
MIEKE UYTTENDAELE
Keyword(s):  
Escherichia Coli ◽  
Bile Salts ◽  
False Negative ◽  
Raw Milk ◽  
Detection Methods ◽  
Fermented Sausage ◽  
E Coli ◽  
Negative Results ◽  
Peptone Water ◽  
Sprouted Seeds

A comparative study of lag phases and growth rates of healthy, stressed, and sublethally injured Escherichia coli O157 cells in 10 enrichment broths was performed. The evaluation of enrichment protocols was validated by different end point detection methods (two PCR and two combined capture-plate methods). Tryptic soy broth b [TSB (b)] provided the fastest growth (μmax = 1.00 ± 0.06 h−1) but failed to recover oxidative-stressed E. coli O157. TSB (a), TSB–yeast extract medium, TSB supplemented with 8 mg/liter novobiocin plus 16 mg/liter vancomycin (TSB+), buffered peptone water (BPW), and BPW supplemented with 8 mg/liter vancomycin (BPW+V) enabled resuscitation of E. coli O157 cells independent from precultural conditions. Modified TSB plus 10 mg/liter novobiocin (mTSB+N), EC medium, EC reduced bile salts medium (ECred), TSB (b), and TSB supplemented with 8 mg/liter novobiocin plus 16 mg/liter vancomycin plus 2 mg/liter rifampin plus 1 mg/liter K-Telluriet plus 1.5 g/liter bile salts no. 3 (TSB++) all failed to recover E. coli O157 cells for at least one type of stress. The use of TSB (a), TSB+, BPW, and BPW+V was compared with that of mTSB+N (International Organization for Standardization reference broth) for reliable detection of low numbers of healthy, stressed, and sublethally injured E. coli O157 (approximately 10 CFU/10 g) from foods (sprouted seeds, fermented sausage, raw milk, and raw ground beef). When low numbers of healthy cells were inoculated, BPW, BPW+V, TSB, TSB+, and mTSB+N enabled growth until detectable numbers within 6 h of enrichment at 41.5°C. Results showed that mTSB+N failed to recover to detectable numbers E. coli O157 cells sublethally injured by freeze and food stresses, in contrast to what was obtained with BPW and BPW+V. This study highlights that using mTSB+N for recovery of E. coli O157 from foods may yield false-negative results.

Evaluation of Culture- and PCR-Based Detection Methods for Escherichia coli O157:H7 in Inoculated Ground Beef†

2005 ◽  
Vol 68 (8) ◽  
pp. 1566-1574 ◽  
Author(s):  
TERRANCE M. ARTHUR ◽  
JOSEPH M. BOSILEVAC ◽  
XIANGWU NOU ◽  
MOHAMMAD KOOHMARAIE
Keyword(s):  
Escherichia Coli ◽  
False Positive ◽  
False Negative ◽  
Ground Beef ◽  
Immunomagnetic Separation ◽  
Detection Methods ◽  
Escherichia Coli O157 ◽  
E Coli ◽  
Perishable Product ◽  
Specificity And Sensitivity

Currently, several beef processors employ test-and-hold systems for increased quality control of ground beef. In such programs, each lot of product must be tested and found negative for Escherichia coli O157:H7 prior to release of the product into commerce. Optimization of three testing attributes (detection time, specificity, and sensitivity) is critical to the success of such strategies. Because ground beef is a highly perishable product, the testing methodology used must be as rapid as possible. The test also must have a low false-positive result rate so product is not needlessly discarded. False-negative results cannot be tolerated because they would allow contaminated product to be released and potentially cause disease. In this study, two culture-based and three PCR-based methods for detecting E. coli O157:H7 in ground beef were compared for their abilities to meet the above criteria. Ground beef samples were individually spiked with five genetically distinct strains of E. coli O157: H7 at concentrations of 17 and 1.7 CFU/65 g and then subjected to the various testing methodologies. There was no difference (P > 0.05) in the abilities of the PCR-based methods to detect E. coli O157:H7 inoculated in ground beef at 1.7 CFU/65 g. The culture-based systems detected more positive samples than did the PCR-based systems, but the detection times (21 to 48 h) were at least 9 h longer than those for the PCR-based methods (7.5 to 12 h). Ground beef samples were also spiked with potentially cross-reactive strains. The PCR-based systems that employed an immunomagnetic separation step prior to detection produced fewer false-positive results.

scholarly journals Development of Molecular Rapid Detection for Vibrio cholerae and Escherichia coli

2020 ◽  
Author(s):  
Diana Elizabeth Waturangi ◽  
JASON PETRUS ◽  
RICO KOSASIH ◽  
GLORIA RAISSA
Keyword(s):  
Escherichia Coli ◽  
Vibrio Cholerae ◽  
False Positive ◽  
Rapid Detection ◽  
Pathogenic Bacteria ◽  
Molecular Detection ◽  
Detection Method ◽  
False Negative ◽  
E Coli ◽  
Drinking Water Samples

Abstract Background: Vibrio cholerae and Escherichia coli were main causative agent foodborne diseases, especially in many developing countries, such as Indonesia. Thereby, rapid detection of these pathogenic bacteria is necessary to quickly detect infection that occurred so it can be treated immediately. In this case, multiplex PCR allows multiple genes amplification in one reaction thereby enable to perform rapid detection of these pathogenic bacteria. The objective of this study is to develop rapid molecular detection of V. Cholerae and E. coli and analyze the sensitivity and specificity of this assay.Result: In this study, we used various virulence genes in each pathogenic bacteria as marker to develop rapid molecular detection. Based on this research, optimum results of V. cholerae and E. coli rapid detection were obtained with a primer concentration of 16 µM for ctxA and ompU, 30 µM for ace, and 50 µM for zot, and toxR; 2 µM for elt and 5 µM for stx, respectively. Finally, based on the method standardization by ISO/TS 20836 these assays had 0% false positive, 0% false negative, 100% specificity, and 100% sensitivity; 0% false positive, 4% false negative, 100% specificity, and 96% sensitivity for V. cholerae and E. coli respectively. Conclusion: The optimized method was qualified to be used as a detection method for V. cholerae and E. coli detection according to ISO/TS 20836 (2017) and EHEC and ETEC contamination in drinking water samples.

scholarly journals A comparison of confirmatory media for coliform organisms and Escherichia coli in water

1981 ◽  
Vol 87 (3) ◽  
pp. 369-375 ◽  
Keyword(s):  
Escherichia Coli ◽  
Brilliant Green ◽  
False Negative ◽  
Gas Production ◽  
Single Tube ◽  
Negative Results ◽  
False Negative Results ◽  
Indole Production

SummaryGas production by coliform organisms and Escherichia coli from lauryl tryptose lactose broth (LTLB) was compared with that from brilliant green (lactose) bile broth (BGB). These media were compared with lauryl tryptose mannitol broth (LTMB) with and without added tryptophan for both gas and indole production. At 37 °C, LTLB and BGB were both satisfactory for gas production, but at 44 °C, LTLB gave fewer false-negative results and was thus significantly less inhibitory than BGB. However when LTLB and LTMB were compared as single-tube confirmatory media, LTLB give a high proportion of false-negative reactions in the indole test at 44 °C. The substitution of mannitol for lactose and the addition of tryptophan yielded a satisfactory medium for both confirmation of gas production and the demonstration of indole at 44 °C.

scholarly journals OPTIMIZATION OF MOLECULAR DETECTION FOR Vibrio cholerae and PATHOGENIC Escherichia coli USING MULTIPLEX PCR

2021 ◽  
pp. e299
Author(s):  
Diana Elizabeth Waturangi ◽  
Jason Petrus ◽  
Rico Kosasih ◽  
Felicia Roseline
Keyword(s):  
Escherichia Coli ◽  
Multiplex Pcr ◽  
Vibrio Cholerae ◽  
False Positive ◽  
Rapid Detection ◽  
Pathogenic Bacteria ◽  
Molecular Detection ◽  
False Negative ◽  
E Coli ◽  
Pathogenic Escherichia Coli

Vibrio cholerae and pathogenic Escherichia coli were considered as main causative agent foodborne diseases especially in many developing countries, such as Indonesia. Thereby, rapid detection of these pathogenic bacteria is necessary to treat food-borne related diseases causing by these bacteria. In this case, multiplex PCR allows multiple genes amplification in one reaction thereby enable to perform rapid detection of these pathogenic bacteria. The objective of this study is to optimize uniplex and multiples PCR of V. cholerae and pathogenic E. coli detection and determine the sensitivity and specificity of this assays. We used various virulence genes for each pathogenic bacterium as markers for uniplex and multiplex PCR detection. Based on this research, the optimum results of V. cholerae and pathogenic E. coli were obtained with a primer concentration of 16 µM for ctxA and ompU, 30 µM for ace, and 50 µM for zot, and toxR; 2 µM for elt and 5 µM for stx, respectively. Finally, based on the standardization method by ISO/TS 20836 these assays had 0% false positive, 0% false negative, 100% specificity, and 100% sensitivity; 0% false positive, 4% false negative, 100% specificity, and 96% sensitivity for V. cholerae and pathogenic E. coli respectively. The optimized method was qualified to be used as a molecular detection for V. cholerae as well as EHEC and ETEC detection according to ISO/TS 20836 (2017)  from drinking water samples.

Contamination of Intact Apples after Immersion in an Aqueous Environment Containing Escherichia coli O157:H7†

1999 ◽  
Vol 62 (5) ◽  
pp. 444-450 ◽  
Author(s):  
R. L. BUCHANAN ◽  
S. G. EDELSON ◽  
R. L. MILLER ◽  
G. M. SAPERS
Keyword(s):  
Escherichia Coli ◽  
Outer Core ◽  
Core Region ◽  
Effect Of Temperature ◽  
Aqueous Environment ◽  
Escherichia Coli O157 ◽  
Dye Uptake ◽  
E Coli ◽  
Golden Delicious ◽  
Peptone Water

The extent and location of Escherichia coli O157:H7 contamination after intact apples were immersed in cold (2°C) 1% peptone water containing approximately 3 × 107 CFU/ml was assessed using four apple varieties, Golden Delicious, McIntosh, Red Delicious, and Braeburn. Room temperature and refrigerated apples were used to determine the effect of temperature differential on E. coli infiltration. The highest levels of E. coli were associated with the outer core region of the apple, followed by the skin. Apples were subsequently treated by immersing them for 1 min in 2,000 mg/liter sodium hypochlorite, followed by a 1-min tapwater rinse. This treatment reduced pathogen levels by 1- to 3-log cycles but did not eliminate the microorganism, particularly from the outer core region. While E. coli was not detected in the inner core of most apples, warm fruit immersed in cold peptone water occasionally internalized the pathogen. The frequency and extent of internalization of the pathogen was less when cold apples were immersed in cold peptone water. Subsequent dye uptake studies with Golden Delicious apples indicated that approximately 6% of warm apples immersed into a cold dye solution accumulated dye via open channels leading from the blossom end into the core region. However, dye uptake did not occur when the dye solution was warmer than the apple.

Simultaneous Enrichment of Shiga Toxin–Producing Escherichia coli O157 and O26 and Salmonella in Food Samples Using Universal Preenrichment Broth

2009 ◽  
Vol 72 (10) ◽  
pp. 2065-2070 ◽  
Author(s):  
MASASHI KANKI ◽  
KAZUKO SETO ◽  
JUNKO SAKATA ◽  
TETSUYA HARADA ◽  
YUKO KUMEDA
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Food Samples ◽  
Escherichia Coli O157 ◽  
E Coli ◽  
Significant Difference ◽  
Peptone Water ◽  
Radish Sprouts ◽  
Pork Samples ◽  
Better Than

Universal preenrichment broth (UPB) was compared with modified Escherichia coli broth with novobiocin (mEC+n) for enrichment of Shiga toxin–producing E. coli O157 and O26, and with buffered peptone water (BPW) for preenrichment of Salmonella enterica. Ten strains each of the three pathogens were inoculated into beef and radish sprouts following thermal, freezing, or no treatment. With regard to O157 and O26, UPB incubated at 42°C recovered significantly more cells from inoculated beef than UPB at 35°C and from radish sprout samples than UPB at 35°C and mEC+n. With regard to Salmonella, UPB incubated at 42°C was as effective as UPB at 35°C and BPW at recovering cells from beef and radish sprout samples. No significant difference was noted between the effectiveness of UPB at 42°C and UPB at 35°C or BPW in the recovery of Salmonella from 205 naturally contaminated poultry samples. By using UPB at 42°C, one O157:H7 strain was isolated from the mixed offal of 53 beef samples, 6 cattle offal samples, and 50 pork samples all contaminated naturally, with no pathogen inoculation. The present study found that UPB incubated at 42°C was as effective as, or better than, mEC+n for enrichment of O157 and O26 and comparable to BPW for preenrichment of Salmonella. These findings suggest that a great deal of labor, time, samples, and space may be saved if O157, O26, and Salmonella are enriched simultaneously with UPB at 42°C.

scholarly journals Isolation and Enumeration of Escherichia coli from Soil and Water

2020 ◽  
Vol 36 (2) ◽  
pp. 75-77
Author(s):  
Anindita Bhowmik ◽  
Sunjukta Ahsan
Keyword(s):  
Escherichia Coli ◽  
Water Samples ◽  
Gas Production ◽  
Soil Samples ◽  
Most Probable Number ◽  
Biochemical Tests ◽  
Probable Number ◽  
E Coli ◽  
Dilution Series

Majority of the population of Bangladesh depend on tap or surface water as their source of water supply. This study was carried out to examine the microbial quality of both water and soil collected from different places using the multiple tube fermentation technique to determine coliform count by the most probable number (MPN) method in brilliant green lactose broth (BGLB) media.Inoculum from positive tubes of the presumptive test were further transferred on eosinemethylene blue (EMB) and MacConkey agar.The organisms isolated were further characterized using biochemical tests. Out of 93 water samples, 30 (32.26%) indicated the presence of lactose fermenter and gas producer in all 3 tubes of dilution series using inoculum quantities of 1.0, 0.1 and 0.01 ml, whereas out of 85 soil samples, 45 (52.94%) showed acid and gas production in all 3 tubes of dilution series.Among 85 soil samples, 40 samples that contained at least one positive in each dilution series and among 93 water samples, 31 samples that contained at least one positive in each dilution series were further re-identified with biochemical tests.This study showed 30.59% soil isolates and 26.88% water isolates were Escherichia coli which highlighted the fact that both water and soil act as a major reservoir of E.coli, which indicates possible fecal contamination as well as presence of potentially pathogenic E. coli. Bangladesh J Microbiol, Volume 36 Number 2 December 2019, pp 75-77

scholarly journals Persistence of Escherichia coli O157:H12 and Escherichia coli K12 as Non-pathogenic Surrogates for O157:H7 on Lettuce Cultivars Irrigated With Secondary-Treated Wastewater and Roof-Collected Rain Water in the Field

2020 ◽  
Vol 4 ◽  
Author(s):  
Hsin-Bai Yin ◽  
Nidhi Gupta ◽  
Chi-Hung Chen ◽  
Ashley Boomer ◽  
Abani Pradhan ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Pathogenic Bacteria ◽  
Water Source ◽  
Water Sources ◽  
Rain Water ◽  
Treated Wastewater ◽  
E Coli ◽  
Water Characteristics ◽  
The Difference ◽  
Alternative Water

Treated wastewater (TW) and roof-collected rain water (RW) that meet the required microbial quality as per Food Safety Modernization Act (FSMA) regulation may serve as alternative irrigation water sources to decrease the pressure on the current water scarcity. Alternative water sources may have different water characteristics that influence the survival and transfer of microorganisms to the irrigated produce. Further, these water sources may contain pathogenic bacteria such as Shiga-toxigenic Escherichia coli. To evaluate the risk associated with TW and RW irrigation on the fresh produce safety, the effect of TW and RW irrigation on the transfer of two non-pathogenic E. coli strains as surrogates for E. coli O157:H7 to different lettuce cultivars grown in the field was investigated. Lettuce cultivars “Annapolis,” “Celinet,” and “Coastline” were grown in the field at the Fulton farm (Chambersburg, PA). Approximately 10 days before harvest, lettuce plants were spray-irrigated with groundwater (GW), TW, or RW containing 6 log CFU ml−1 of a mixture of nalidixic acid-resistant E. coli O157:H12 and chloramphenicol-resistant E. coli K12 in fecal slurry as non-pathogenic surrogates for E. coli O157:H7. On 0, 1, 3, 7, and 10 days post-irrigation, four replicate lettuce leaf samples (30 g per sample) from each group were collected and pummeled in 120 ml of buffered peptone water for 2 min, followed by spiral plating on MacConkey agars with antibiotics. Results showed that the recovery of E. coli O157:H12 was significantly greater than the populations of E. coli K12 recovered from the irrigated lettuce regardless of the water sources and the lettuce cultivars. The TW irrigation resulted in the lowest recovery of the E. coli surrogates on the lettuce compared to the populations of these bacteria recovered from the lettuce with RW and GW irrigation on day 0. The difference in leaf characteristics of lettuce cultivars significantly influenced the recovery of these surrogates on lettuce leaves. Populations of E. coli O157:H12 recovered from the RW-irrigated “Annapolis” lettuce were significantly lower than the recovery of this bacterium from the “Celinet” and “Coastline” lettuce (P < 0.05). Overall, the recovery of specific E. coli surrogates from the RW and TW irrigated lettuce was comparable to the lettuce with the GW irrigation, where GW served as a baseline water source. E. coli O157:H12 could be a more suitable surrogate compared to E. coli K12 because it is an environmental watershed isolate. The findings of this study provide critical information in risk assessment evaluation of RW and TW irrigation on lettuce in Mid-Atlantic area.

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