Turntable Paper-Based Device to Detect Escherichia coli

Escherichia coli has been known to cause a variety of infectious diseases. The conventional enzyme-linked immunosorbent assay (ELISA) is a well-known method widely used to diagnose a variety of infectious diseases. This method is expensive and requires considerable time and effort to conduct and complete multiple integral steps. We previously proposed the use of paper-based ELISA to rapidly detect the presence of E. coli. This approach has demonstrated utility for point-of-care (POC) urinary tract infection diagnoses. Paper-based ELISA, while advantageous, still requires the execution of several procedural steps. Here, we discuss the design and experimental implementation of a turntable paper-based device to simplify the paper-based ELISA protocols for the detection of E. coli. In this process, antibodies or reagents are preloaded onto zones of a paper-based device and allowed to dry before use. We successfully used this device to detect E. coli with a detection limit of 105 colony-forming units (colony-forming unit [CFU])/mL.

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A PCR-Based Method of Detection and Differentiation of K88+ Adhesive Escherichia Coli

Journal of Veterinary Diagnostic Investigation ◽

10.1177/104063879600800410 ◽

1996 ◽

Vol 8 (4) ◽

pp. 460-463 ◽

Cited By ~ 14

Author(s):

Mark A. Franklin ◽

David H. Francis ◽

Diane Baker ◽

Alan G. Mathew

Keyword(s):

Escherichia Coli ◽

Enzyme Linked Immunosorbent Assay ◽

Antigenic Variant ◽

Variable Regions ◽

E Coli ◽

Clinical Assay ◽

Pcr Product ◽

Structural Subunit ◽

Pcr Products ◽

Polymerase Chain

The objective of this study was to develop a polymerase chain reaction (PCR)-based method to detect and differentiate among Escherichia coli strains containing genes for the expression of 3 antigenic variants of the fimbrial adhesin K88 (K88ab, K88ac, and K88ad). Five primers were designed that allowed detection of K88+ E. coli, regardless of antigenic variant, and the separate detection of the ab, ac, and ad variants. Primers AM005 and AM006 are 21 base pair (bp) oligomers that correspond to a region of the K88 operon that is common to all 3 antigenic variants. Primers MF007, MF008, and MF009 are 24-bp oligomers that matched variable regions specific to ab, ac, and ad, respectively. Using primers AM005 and AM006, a PCR product was obtained that corresponds to a 764-bp region within the large structural subunit of the K88 operon common to all 3 antigenic variants. Primer AM005 used with MF007, MF008, or MF009 produced PCR products approximately 500-bp in length from within the large structural subunit of the K88 operon of the 3 respective antigenic variants. Fragments were identified by rates of migration on a 1% agarose gel relative to each other as well as to BstEII-digested lambda fragments. This PCR-based method was comparable to the enzyme-linked immunosorbent assay and western blot test in the ability to differentiate between the antigenic variants. K88+ E. coli were differentiated from among laboratory strains and detected in ileal samples taken from cannulated pigs challenged with a known K88+ variant. K88+ E. coli were also detected from fecal swabs taken from newly weaned pigs, thus confirming that this PCR-based test could provide a convenient clinical assay for the detection of K88+ E. coli. Detection and differentiation of K88+ E. coli using general and specific primers was successful. PCR methods of detection should permit identification of K88+ antigenic variants regardless of the level of expression of the antigen.

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The combination of Elephantopus scaber and Sauropus androgynus promotes erythroid lineages and modulates follicle-stimulating hormone and luteinizing hormone levels in pregnant mice infected with Escherichia coli

Veterinary World ◽

10.14202/vetworld.2021.1398-1404 ◽

2021 ◽

pp. 1398-1404

Author(s):

Muhammad Sasmito Djati ◽

Yuyun Ika Christina ◽

Muhaimin Rifa'i

Keyword(s):

Escherichia Coli ◽

Luteinizing Hormone ◽

Follicle Stimulating Hormone ◽

Relative Number ◽

Enzyme Linked Immunosorbent Assay ◽

Hormone Levels ◽

E Coli ◽

Pregnant Mice ◽

Elephantopus Scaber ◽

Stimulating Hormone

Background and Aim: Escherichia coli infection produces an adverse effect on the erythrocyte lineage and hormone levels during pregnancy. This study aimed to evaluate the effects of Elephantopus scaber (ES) and Sauropus androgynus (SA) in combination on circulating follicle-stimulating hormone (FSH) and luteinizing hormone (LH) levels and erythropoiesis changes in E. coli-infected pregnant mice. Materials and Methods: Female Balb/c mice were mated with normal male mice and pregnancies were identified by the formation of vaginal plugs. Twenty-eight pregnant mice were divided randomly into seven groups: A control group (N), E. coli-infected pregnant mice (K+), and infected pregnant mice received the following five treatments: (1) Only ES; (2) ESSA1 (75:25); (3) ESSA2 (50:50); (4) ESSA3 (25:75); and (5) only SA, beginning from the 1st to the 16th day of pregnancy. Pregnant mice were infected with 107 CFU/mL of E. coli on day 4. Blood serum was collected on days 8, 12, and 16 of pregnancy and LH and FSH levels were measured by enzyme-linked immunosorbent assay. Bone marrow was isolated to determine the relative number of TER-119+VLA4+ and TER-119+CD34+ using flow cytometry. Results: The ESSA1 and SA groups exhibited a marked increase in LH levels. The combination of ES and SA administered at a 25:75 ratio (ESSA3) altered FSH levels and the relative number of TER-119+VLA4+ in infected pregnant mice. Combined with SA at an equal ratio (50:50), ESSA2 group exhibited a significant increase in the expression of TER119+CD34+ compared with the other treatment groups. Conclusion: ES and SA combined at a ratio of 25:75 exhibited optimal results in altering hormonal and erythropoiesis in infected pregnant mice.

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Comparison of Rapid Enzyme-Linked Immunosorbent Assay and Immunomagnetic Separation Methods for Detection of Escherichia coli O157 in Fecal, Hide, Carcass, and Ground Beef Samples

Journal of Food Protection ◽

10.4315/0362-028x-70.10.2230 ◽

2007 ◽

Vol 70 (10) ◽

pp. 2230-2234 ◽

Cited By ~ 4

Author(s):

T. W. THOMPSON ◽

T. P. STEPHENS ◽

G. H. LONERAGAN ◽

M. F. MILLER ◽

M. M. BRASHEARS

Keyword(s):

Escherichia Coli ◽

Ground Beef ◽

Immunomagnetic Separation ◽

Separation Method ◽

Enzyme Linked Immunosorbent Assay ◽

Escherichia Coli O157 ◽

Perfect Agreement ◽

Fecal Samples ◽

E Coli ◽

Beef Products

Rapid enzyme-linked immunosorbent assays (ELISAs) are approved for detection of Escherichia coli O157 in beef products. However, these kits have also been used in the industry to detect this pathogen on hides or in feces of cattle, although this use has not been validated. The objective of this study was to compare commercially available ELISAs (E. coli Now, Reveal, and VIP) with immunomagnetic separation along with selective media to detect E. coli O157 on hides, in feces, and in medium- and low-level-inoculated ground beef and carcasses (simulated by using briskets) samples. Naturally infected hide and fecal samples were subjected to both the immunomagnetic separation method and ELISAs for the detection of E. coli O157. Additionally, E. coli O157 inoculated and noninoculated ground beef and beef briskets were used to simulate meat and carcass samples. When comparing the detection results from the ELISAs (E. coli Now, Reveal, and VIP) to the immunomagnetic separation method, poor agreement was observed for fecal samples (kappa = 0.10, 0.02, and 0.03 for E. coli Now, Reveal, and VIP, respectively), and fair-to-moderate agreement was observed for hide samples (kappa = 0.30, 0.51, and 0.29 for E. coli Now, Reveal, and VIP, respectively). However, there was near-perfect agreement between the immunomagnetic separation method and ELISAs for ground beef (kappa = 1, 1, and 0.80 for E. coli Now, Reveal, and VIP, respectively) and brisket (kappa = 1, 1, and 1 for E. coli Now, Reveal, and VIP, respectively) samples. Assuming immunomagnetic separation is the best available method, these data suggest that the ELISAs are not useful in detecting E. coli O157 from hide or fecal samples. However, when ELISAs are used on ground beef and beef brisket samples they can be used with a high degree of confidence.

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Development of an Electrochemical Biosensor for Rapid and Effective Detection of Pathogenic Escherichia coli in Licorice Extract

Applied Sciences ◽

10.3390/app9020295 ◽

2019 ◽

Vol 9 (2) ◽

pp. 295 ◽

Cited By ~ 3

Author(s):

Haixia Wang ◽

Yuwen Zhao ◽

Songtao Bie ◽

Tongchuan Suo ◽

Guangcheng Jia ◽

...

Keyword(s):

Escherichia Coli ◽

Electrochemical Biosensor ◽

Linear Trend ◽

Differential Pulse ◽

E Coli ◽

Pathogenic Escherichia Coli ◽

Colony Forming Units ◽

Relative Standard ◽

Licorice Extract ◽

Pulse Voltammetry

An aptamer-based electrochemical biosensor was successfully developed and applied in the rapid detection of pathogenic Escherichia coli (E. coli) in licorice extract. The thiolated capture probes were firstly immobilized on a gold electrode, and then the biotinylated aptamer probes for E. coli were introduced by hybridization with the capture probes. Due to the stronger interaction between the aptamer and the E. coli, a part of the biotinylated aptamers will dissociate from the capture probes in the presence of E. coli. The residual biotinylated aptamer probes can quantitatively bind with streptavidin-alkaline phosphatase. Subsequently, α-naphthyl phosphate substrate was catalytically hydrolyzed to generate electrochemical response, which could be recorded by a differential pulse voltammetry. The dependence of the peak current on the logarithm of E. coli concentration in the range from 5.0 × 102 colony forming units (CFU)/mL to 5.0 × 107 CFU/mL exhibited a linear trend with a detection limit of 80 CFU/mL. The relative standard deviation of 5 successive scans was 5.3%, 4.5% and 1.1% for 5.0 × 102, 5.0 × 105 and 5.0 × 107 CFU/mL E. coli, respectively. In the detection of the licorice extract samples, the results obtained from the proposed strategy and traditional culture counting method were close to each other, but the time consumption was only ~1/30 compared with the traditional method. These results demonstrate that the designed biosensor can be potentially utilized for rapid microbial examination in traditional Chinese medicine and relevant fields.

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Use of a Shiga Toxin (Stx)-Enzyme-Linked Immunosorbent Assay and Immunoblot for Detection and Isolation of Stx-Producing Escherichia coli from Naturally Contaminated Beef

Journal of Food Protection ◽

10.4315/0362-028x-63.9.1167 ◽

2000 ◽

Vol 63 (9) ◽

pp. 1167-1172 ◽

Cited By ~ 22

Author(s):

HEBA NASHED ATALLA ◽

ROGER JOHNSON ◽

SCOTT MCEWEN ◽

R. W. USBORNE ◽

C. L. GYLES

Keyword(s):

Escherichia Coli ◽

Vero Cell ◽

Immunosorbent Assay ◽

Shiga Toxin ◽

Total Coliform ◽

Complete Agreement ◽

Enzyme Linked Immunosorbent Assay ◽

Spearman Correlation ◽

Southern Ontario ◽

E Coli

The purpose of this study was to evaluate an enzyme-linked immunosorbent assay (ELISA) and an immunoblot procedure for detection and isolation of Shiga toxin-producing Escherichia coli (STEC) from beef, and to correlate the presence of STEC in beef with E. coli and total coliform counts. A total of 120 samples of boneless beef supplied to a meat processor in southern Ontario were tested for the presence of STEC, E. coli, and total coliforms. Following enrichment in modified tryptic soy broth, samples were screened for Shiga toxin (Stx) by a Stx-ELISA and a Vero cell assay (VCA). Samples that were positive in the Stx-ELISA were subjected to the Stx-immunoblot for STEC isolation. Overall, 33.3% of samples were positive in the VCA, and 34.2% were positive in the Stx-ELISA. There was almost complete agreement between the Stx-ELISA and the VCA results (kappa = 0.98). The sensitivity and specificity of the Stx-ELISA with respect to the VCA were 100% and 98.75%, respectively. STEC were isolated by the Stx-immunoblot from 87.8% of the samples that were positive in the Stx-ELISA. The STEC isolates belonged to 19 serotypes, with serotype O113:H21 accounting for 10 of 41 isolates. No STEC of serotype O157:H7 were isolated. There was a significant correlation between E. coli counts and total coliform counts (Spearman correlation coefficient = 0.68, P < 0.01). The E. coli count was positively correlated with detection of STEC by both the Stx-ELISA and the VCA (P < 0.01).

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Evaluation of Three Commercially Available Enzyme-Linked Immunosorbent Assay Kits for Detection of Shiga Toxin

Journal of Food Protection ◽

10.4315/0362-028x-72.4.741 ◽

2009 ◽

Vol 72 (4) ◽

pp. 741-747 ◽

Cited By ~ 34

Author(s):

JOHN WILLFORD ◽

KENNETH MILLS ◽

LAWRENCE D. GOODRIDGE

Keyword(s):

Escherichia Coli ◽

Immunosorbent Assay ◽

Shiga Toxin ◽

Screening Method ◽

Enzyme Linked Immunosorbent Assay ◽

Microplate Assay ◽

E Coli ◽

Elisa Kit ◽

Pure Cultures ◽

Order Of Magnitude

Three commercially available Shiga toxin (Stx) enzyme-linked immunosorbent assay (ELISA) kits were evaluated for their ability to detect Stx in pure cultures of Stx-producing Escherichia coli (specificity). The detection limits (sensitivity) of each ELISA kit were also evaluated. Seventy-eight Stx-producing E. coli (STEC) isolates that produced Stx1, Stx2, or Stx1 and Stx2 variants were examined in this study. The specificities of the tests were comparable, and the sensitivities of two of the tests (Premier EHEC and rBiopharm Ridascreen Verotoxin Enzyme Immunoassay) were within the same order of magnitude. The ProSpecT Shiga Toxin E. coli Microplate Assay was approximately 10-fold less sensitive. The inability of all three tests to detect the Stx2d and Stx2e variants indicated that some STEC strains may not be detected by Stx ELISA. The ability of the Premier EHEC ELISA to detect toxin in artificially inoculated bovine fecal samples (following enrichment) indicated that this kit may be used to screen cattle for the presence of Stx as an indicator of the presence of STEC. In particular, such a screening method could be useful during the summer, when the number of STEC-positive animals and the number of STEC that they shed increase.

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AVIAN INFLUENZA ANTIBODY RESPONSE IN PIGLETS THATAREGIVEN ESCHERICHIA COLI–AVIAN INFLUENZA VACCINES

Indonesia Medicus Veterinus ◽

10.19087/imv.2021.10.1.61 ◽

2021 ◽

Vol 10 (1) ◽

pp. 61-70

Author(s):

Audrey Febiannya Putri Bhaskara ◽

I Gusti Ngurah Kade Mahardika ◽

I Nyoman Suartha

Keyword(s):

Escherichia Coli ◽

Avian Influenza ◽

Optical Density ◽

Antibody Test ◽

Influenza Vaccines ◽

Enzyme Linked Immunosorbent Assay ◽

Virus Antibody ◽

E Coli ◽

Test Kit ◽

Virus Influenza

Babi berperan penting dalam ekologi virus influenza, karena babi dapat berperan sebagai wahana untuk reasorsi virus influenza dari unggas dan mamalia. Vaksinasi dengan antigen influenza universal, yaitu nukleoprotein, dapat menurunkan peluang babi dalam memunculkan virus influenza baru. Penelitian ini bertujuan untuk mengentahui respons antibodi dari vaksinasi dengan nukleoprotein rekombinan - Escherichia coli. Sebanyak 12 anak babi landrace dari tiga induk yang berbeda dipilih secara acak. Enam ekor divaksinasi dengan vaksin nucleoprotein-E. coli pada umur tujuh hari dan diulang pada umur 21 hari. Enam ekor tidak divaksinasi. Serum diambil pada umur 35 hari. Nilai optical density (OD) antibodi terhadap nukleoprotein diuji dengan teknik Enzyme-Linked Immunosorbent Assay (ELISA) dengan menggunakan Kit ELISA komersial, Avian Influenza Virus Antibody Test Kit. Hasil penelitian menunjukkan bahwa nilai Optical Density rata-rata babi yang divaksinasi (0,367) secara statistika nyata lebih tinggi dibandingkan dengan yang tidak divaksinasi (0,054). Vaksin rekombinan nucleoprotein-E. coli yang dicobakan mampu meningkatan antibodi terhadap virus avian influenza pada anak babi.

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Role of endotoxin in the response to experimentally induced bacteremia in chronically prepared rats

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.1997.272.5.r1562 ◽

1997 ◽

Vol 272 (5) ◽

pp. R1562-R1570 ◽

Cited By ~ 2

Author(s):

D. E. Carlson ◽

J. K. Babus ◽

N. Nguyuza ◽

H. Melhem-Stancofski ◽

B. J. Eastridge

Keyword(s):

Escherichia Coli ◽

Total Activity ◽

E Coli ◽

Plasma Endotoxin ◽

Endotoxin Activity ◽

Colony Forming Units ◽

Related Increase ◽

Experimentally Induced

Chronically prepared rats were injected intravenously with live Escherichia coli in doses from approximately 10(5) to approximately 10(9) colony-forming units (CFU). Significant dose-related increase in plasma adrenocorticotropin and corticosterone occurred after approximately 10(7) CFU. Fever occurred after approximately 10(7) CFU but not after approximately 10(9) CFU. These responses changed significantly but were not blocked completely when > 94% of the viable E. coli was removed from the inoculates. The remaining endotoxin activity in the inoculates resembled lipopolysaccharide (LPS) extracted from the same strain of E. coli on electrophoretic gels. Plasma endotoxin increased for > or = 240 min to 5.1 +/- 0.9 endotoxin units (EU)/ml after approximately 10(7) CFU and to 440 +/- 59 EU/ml after approximately 10(9) CFU. Endotoxin at approximately 10(9) CFU caused death within 24 h that was not predicted by the total activity of endotoxin that was injected. In contrast, extracted LPS with its strain and total activity matched to approximately 10(7) CFU mimicked the responses to this nonfatal dose. The total endotoxin activity of the injected bacteria appears to account for the effects of nonfatal doses of E. coli but not for the effects of fatal doses.

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Prevalence and characterization of Escherichia coli in the Kelantan River and its adjacent coastal waters

Water Science & Technology Water Supply ◽

10.2166/ws.2020.018 ◽

2020 ◽

Vol 20 (3) ◽

pp. 930-942

Author(s):

Chui Wei Bong ◽

Siong Kiat Chai ◽

Lay Ching Chai ◽

Ai Jun Wang ◽

Choon Weng Lee

Keyword(s):

Escherichia Coli ◽

Coastal Waters ◽

Membrane Filtration ◽

Sea Water ◽

Phylogenetic Group ◽

Animal Origin ◽

Phylogenetic Groups ◽

E Coli ◽

Colony Forming Units ◽

Phylogenetic Grouping

Abstract The presence of Escherichia coli in river and sea water may cause different levels of infections and constitutes a risk to public health. In this study, water samples were collected from 15 sites along the Kelantan River, estuaries and its adjacent coastal waters to investigate the prevalence and diversity of E. coli. A membrane filtration technique was used to enumerate E. coli and phylogenetic grouping was performed using triplex polymerase chain reaction. E. coli abundance ranged from 3.1 × 10 to 1.6 × 105 colony forming units 100 mL−1, and total suspended solids correlated significantly with E. coli abundance (r2 = 0.165, p < 0.001) and rainfall (r2 = 0.342, p < 0.001). Phylogenetic group B1 and A (59.4%) were the most prevalent, whereas groups B2 and D were least abundant. The higher abundance of phylogenetic group D at upstream sites of the Kelantan River suggested fecal contamination mainly of animal origin. Canonical-correlation analysis showed phylogenetic group B2, and phylogenetic groups A and D were greater in waters with higher inorganic nutrients (e.g. NH4, NO2 and NO3), whereas phylogenetic group B1 appeared to have better salinity tolerance between phylogenetic groups.

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An endosomal LAPF is required for macrophage endocytosis and elimination of bacteria

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1903896116 ◽

2019 ◽

Vol 116 (26) ◽

pp. 12958-12963 ◽

Cited By ~ 4

Author(s):

Tianliang Li ◽

Kewei Qin ◽

Nan Li ◽

Chaofeng Han ◽

Xuetao Cao

Keyword(s):

Escherichia Coli ◽

Immune Response ◽

Complex Formation ◽

Infectious Diseases ◽

Proinflammatory Cytokines ◽

Inflammatory Responses ◽

E Coli ◽

Caveolin 1 ◽

Early Endosomes ◽

Deficient Mice

Macrophages can internalize the invading pathogens by raft/caveolae and/or clathrin-dependent endocytosis and elicit an immune response against infection. However, the molecular mechanism for macrophage endocytosis remains elusive. Here we report that LAPF (lysosome-associated and apoptosis-inducing protein containing PH and FYVE domains) is required for caveolae-mediated endocytosis.Lapf-deficient macrophages have impaired capacity to endocytose and eliminate bacteria. Macrophage-specificLapf-deficient mice are more susceptible toEscherichia coli(E. coli) infection with higher bacterial loads. Moreover,Lapfdeficiency impairs TLR4 endocytosis, resulting in attenuated production of TLR-triggered proinflammatory cytokines. LAPF is localized to early endosomes and interacts with caveolin-1. Phosphorylation of LAPF by the tyrosine kinase Src is required for LAPF-Src-Caveolin complex formation and endocytosis and elimination of bacteria. Collectively, our work demonstrates that LAPF is critical for endocytosis of bacteria and induction of inflammatory responses, suggesting that LAPF and Src could be potential targets for the control of infectious diseases.

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