Escherichia coli O157:H7 is one of the major foodborne pathogenic bacterial that cause infectious diseases in humans.The previous found that a combination of kojic acid and tea polyphenols exhibited better activity ag...Escherichia coli O157:H7 is one of the major foodborne pathogenic bacterial that cause infectious diseases in humans.The previous found that a combination of kojic acid and tea polyphenols exhibited better activity against E.coli O157:H7 than using either alone.This study aimed to explore responses underlying the antibacterial mechanisms of kojic acid and tea polyphenols from the gene level.The functional enrichment analysis by comparing kojic acid and tea polyphenols individually or synergistically against E.coli O157:H7 found that acid resistance systems in kojic acid were activated,and the cell membrane and genomic DNA were destructed in the cells,resulting in“oxygen starvation”.The oxidative stress response triggered by tea polyphenols inhibited both sulfur uptake and the synthesis of ATP,which affected the bacteria's life metabolic process.Interestingly,we found that kojic acid combined with tea polyphenols hindered the uptake of iron that played an essential role in the synthesis of DNA,respiration,tricarboxylic acid cycle.The results suggested that the iron uptake pathways may represent a novel approach for kojic acid and tea polyphenols synergistically against E.coli O157:H7 and provided a theoretical basis for bacterial pathogen control in the food industry.展开更多
To prepare monoclonal antibodies （MAb） and antisera specific for Escherichia coli （E.coli） O157, and to develop a sandwich enzyme-linked immunosorbent assay （ELISA） to detect Eocoli O157 in foods. Methods Spleen...To prepare monoclonal antibodies （MAb） and antisera specific for Escherichia coli （E.coli） O157, and to develop a sandwich enzyme-linked immunosorbent assay （ELISA） to detect Eocoli O157 in foods. Methods Spleen cells from BALB/c mice immunized with the somatic antigen of E.coli O157：H7 were fused with routine Sp2/0 myeloma cells. The hybridoma cell line specific for E.coli O157 was established after having been subcloned. Antisera specific for E.coli O157 was prepared by intravenous injection into New Zealand rabbits with a stain of E.coli O157：H7. The sandwich ELISA was developed with the polyclonal antibody as the capture antibody and the MAb 3A5 as the detection antibody. The inoculated ground poultry meat and pasteurized miLk were tested to confirm efficiency of the method. Results MAb 3A5 specific for E.coli O157 and O 113：H21 belonged to subtype IgM. The ascetic titers of the antibody was 1：1× 10^6. No cross-reactivity of the MAb was observed with strains of Salmonella spp, Yersinia enterocolitica, Shigella dysenteriae, etc. The purified polyclonal antibody had a titer of 1： 1× 10^5 with E.coli O 157. The detection limit of this sandwich ELISA was 10^3- 10^4 cfu E.coli O157/mL in pure culture with a high specificity, which was characterized by every non-O157 strain with negative response. With 10h enrichment procedure, E.coli O157：H7 recovered well from inoculated ground poultry meat and pasteurized milk at levels of 0.1 cfu/g and 0.1 cfu/mL. Conclusion MAb 3A5 specific for E.coli O 157 and O 113：H21 can be produced by immunizing BALB/c mice with a strain of E.coli O157：H7. Then a sandwich ELISA can be developed with the polyclonal antibody as the capture antibody and the MAb 3A5 as the detection antibody. The method is proved to be a sensitive and specific technique to detect low number of E.coli O157 in food.展开更多
Objective:To survey the prevalence severe diarrhea arising from these bacteria in children under 5 years old in Marvdasht.Methods:In this study faecal sample from 615 children aged <5years old who were hospitalized...Objective:To survey the prevalence severe diarrhea arising from these bacteria in children under 5 years old in Marvdasht.Methods:In this study faecal sample from 615 children aged <5years old who were hospitalized lor gastroenteritis in Fars hospitals in Iran were collected and then enriched in Escherichia coli(E.coli) broth and modified tryplone soy broth with novobiocin media,fermentation of sorbitol,lactose and β— glucoronidase activity of isolated strains was examined by CT—SMAC,VRBA and chromogenic media respectively.Then isolation of E.coli O157:H7 have been confirmed with the use of specific antisera and with multiplex PCR method presence of virulence genes including:xtx_1.stx_2,eae.A.hly has been analyzed.Results:E.coli O157:H7 was detected in 7(1.14%) stool specimens.A significanl difference was seen between detection rale of isolated bacteria from age groups 18-23 months and other age groups(P=0.004).Out of considered virulence genes.only 1 of the isolated strains(0.16%)he stx,and eaeA genes were seen and also all isolated hacleria had resistance to penicillin,ampicillin and erythromycin antibiotics.Conclusions:We found thai children < 2 years of age were at highest risk of infection with E.coli O157:H7.Regarding severity of E.coli O157:H7 pathogenesis,low infectious dose and lack of routine assay for detection ol these bacleria in clinical laboratory,further and completed studies on diagnosis and genolyping of this E.coli O157:H7 strain has been recommended.展开更多
By means of the specific immuno-recognition and ultra-sensitive mass detection, a quartz crystal microbalance (QCM) biosensor for Escherichia coli O157:H7 detection was developed in this work. As a suitable surfactant...By means of the specific immuno-recognition and ultra-sensitive mass detection, a quartz crystal microbalance (QCM) biosensor for Escherichia coli O157:H7 detection was developed in this work. As a suitable surfactant, 16-mercaptohexadecanoic acid (MHDA) was introduced onto the Au surface of QCM, and then self-assembled with N-hydroxysuccinimide (NHS) raster as a reactive intermediate to provide an active interface for the specific antibody immobilization. The binding of target bacteria with the immobilized antibodies decreased the sensor’s resonant frequency, and the frequency shift was correlated to the bacterial concentration. The stepwise assembly of the immunosensor was characterized by means of the electrochemical techniques. Using the immersion-dry-immersion procedure, this QCM biosensor could detect 2.0×102 colony forming units (CFU)/ml E. coli O157:H7. In order to reduce the fabrication time, a polyelectrolyte layer-by-layer self-assembly (LBL-SA) method was adopted for fast construction. Finally, the reproducibility of this biosensor was discussed.展开更多
This study investigated the intervention effects of Lactobacillus casei LC2W in murine(SPF C57BL/c)challenge infection models induced by Escherichia coli O157:H7.Mice were fed streptomycin with water for 3 days prior ...This study investigated the intervention effects of Lactobacillus casei LC2W in murine(SPF C57BL/c)challenge infection models induced by Escherichia coli O157:H7.Mice were fed streptomycin with water for 3 days prior to intragastric gavage by E.coli O157:H7(Control)or L.casei LC2W together with E.coli O157:H7(Intervention)to explore the role of L.casei LC2W by biochemical indicators,histological evaluation,expression of colonic pro-inflammatory and intestinal barrier factors related to enteritis.Results showed that the administration of L.casei LC2W was able to alleviate the symptoms of colitis induced by E.coli O157:H7,exhibiting lower weight loss as well as more intact colon tissue.Furthermore,L.casei LC2W could down-regulate the expression of pro-inflammatory cytokines(TNF-α,IL-6,and IL-1β)and protect the intestinal barrier function by improving the level of MUC2,ZO-1 and E-cadherin 1 compared to the control group.These results demonstrate that L.casei LC2W can reduce the severity of E.coli O157:H7 infection,and suggest L.casei LC2W may maintain the immune balance and intestinal barrier to reduce colitis.In addition,we found the effect of intervention is similar to that of prevention,which is better than that of treatment.展开更多
The antagonistic activity of Lactobacillus acidophilus KLDS1.0901, KLDS1.0902, KLDSI.1003 and NCFM against Escherichia coli O157 ： H7 were investigated in this study. The culture supematants of all the L. acidophilus...The antagonistic activity of Lactobacillus acidophilus KLDS1.0901, KLDS1.0902, KLDSI.1003 and NCFM against Escherichia coli O157 ： H7 were investigated in this study. The culture supematants of all the L. acidophilus stains showed high bacteriostatic activities against E. coli O 157 ： H7 and the bacteriostatic substances of their Cell-Free Supernatants （CFS） were preliminarily determined from organic acids. The bacteriostatic activity from CFS or viable L. acidophilus against E. coli O157 ： H7 was also assessed by using co-incubation methods, CFS had high bactericidal activity against E. coli O157 ： H7, no viable E. coli O157 ： H7 was detected when 5×10^7 CfU ofE. coli O157 ： H7 was added to 5 mL of CFS and incubated at 37℃ for 2 h. However, L. acidophilus themselves had no bacteriostatic activity after directly contacted with E. coli O157 ： H7. The inhibition E. coli O157 ： H7 adhesion and colonization of L. acidophilus were also investigated based on competition, exclusion and displacement assays. L. acidophilus KLDS1.0901, KLDSI.1003 and NCFM strains were effective to displace E. coli O157 ： H7 from a Caco-2 cell layer in competition and exclusion assays. However, in displacement assay, all of the strains showed no significant antagonistic activities. Meanwhile, the probiotic potential of L. acidophilus strains was investigated based on adhesion assay to Caco-2 cells and anti- inflammatory effects by IL-8 produced in Caco-2 cells. The adhesion ability and anti-inflammatory effects of L. acidophilus strains showed a strain-dependent manner. In general, L. acidophilus KLDS 1.0901 and NCFM showed better probiotic potential than KLDS1.0902 and KLDSI.1003. Thus, the use ofL. acidophilus KLDS1.0901 and NCFM to prevent or treat of diseases associated induced E. coli O157 ： H7 in vivo was suggested.展开更多
AIM： To establish the rapid, specific, and sensitive method for detecting O157：H7 with DNA microchips. METHODS： Specific oligonucleotide probes （26-28 nt） of bacterial antigenic and virulent genes of E. coliO157...AIM： To establish the rapid, specific, and sensitive method for detecting O157：H7 with DNA microchips. METHODS： Specific oligonucleotide probes （26-28 nt） of bacterial antigenic and virulent genes of E. coliO157：H7 and other related pathogen genes were pre-synthesized and immobilized on a solid support to make microchips. The four genes encoding O157 somatic antigen （rfbE）, H7 flagellar antigen （fliC） and toxins （SLT1, SLT2） were monitored by multiplex PCR with four pairs of specific primers. Fluorescence-Cy3 labeled samples for hybridization were generated by PCR with Cy3-1abeled single prime. Hybridization was performed for 60 min at 45 ℃. Microchip images were taken using a confocal fluorescent scanner. RESULTS： Twelve different bacterial strains were detected with various combinations of four virulent genes. All the O157：H7 strains yielded positive results by multiplex PCR. The size of the PCR products generated with these primers varied from 210 to 678 bp. All the rfbE/fliC/SLT1/SLT2 probes specifically recognized Cy3-1abeled fluorescent samples from O157：H7 strains, or strains containing O157 and H7 genes. No cross hybridization of O157：H7 fluorescent samples occurred in other probes. Non-O157：H7 pathogens failed to yield any signal under comparable conditions. If the Cy3-1abeled fluorescent product of O157 single PCR was diluted 50-fold, no signal was found in agarose gel electrophoresis, but a positive signal was found in microarray hybridization. CONCLUSION： Microarray analysis of O157：H7 is a rapid, specific, and efficient method for identification and detection of bacterial pathogens.展开更多
The objective of this study was to determine the effect of sodium lactate on the survival of Listeria monocytogenes, Escherichia coli O157: H7, and Salmonella spp. in cooked ham during storage at refrigerated and abus...The objective of this study was to determine the effect of sodium lactate on the survival of Listeria monocytogenes, Escherichia coli O157: H7, and Salmonella spp. in cooked ham during storage at refrigerated and abuse temperatures. Cooked ham was added with 0% - 3% lactate, inoculated with a multiple-strain mixture of L. monocytogenes, E. coli O157: H7, or Salmonella spp. and stored at 4oC - 15oC for up to 35 day. The growth of the three pathogens was inhibited in ham containing 3% lactate, and no growth of E. coli O157: H7 and Salmonella spp. occurred at the lowest storage tem- peratures of 6 and 8oC, respectively. In ham containing no lactate, the average growth rates were 0.256 - 0.380 log CFU/day for L. monocytogenes at 4oC - 8oC, 0.242 - 0.315 log CFU/day for E. coli O157: H7 at 8oC - 15oC, and 0.249 - 0.328 log CFU/day for Salmonella spp. at 10oC - 15oC. The addition of 1% or 2% lactate significantly (P < 0.05) reduced the growth rates of the three pathogens, and the effect was more profound at lower temperatures. Salmonella spp. were more sensitive to the effect of lactate than L. monocytogenes and E. coli O157: H7. Polynomial models were developed to describe the growth rates of the three pathogens as affected by the lactate concentration and storage tem- perature. Results from this study demonstrate the effect of lactate on the growth of L. monocytogenes, E. coli O157: H7, and Salmonella spp. in cooked ham and indicate the effective lactate concentrations and storage temperatures that can be used to enhance the microbiological safety of ready-to-eat ham products.展开更多
In order to test if the intestinal bacteria play an important role in antibacterial ability of earthworm, we chose Escherichia coli O157:H7, an anthropozoonosis pathogen, as a biological stressor and studied the chang...In order to test if the intestinal bacteria play an important role in antibacterial ability of earthworm, we chose Escherichia coli O157:H7, an anthropozoonosis pathogen, as a biological stressor and studied the change of intestinal bacteria community of earthworm by PCR-DGGE analysis. Results showed that the pathogen merely existed 1 - 3 days, then almost disappeared after through the earthworm’s gut. In this period, the diversity and abundance index of intestinal bacteria increased first, then decreased, and finally kept stably after 7 days. The result demonstrated that the intestinal bacteria of earthworm had ability of adjust community structure to eliminate the pathogen E. coli O157:H7, and the amount of bacteria Bacillus increased significantly, which might be the positive antagonism to E. coli O157:H7.展开更多
Escherichia coli O157:H7 is a human pathogen that was first identified from a foodborne outbreak in 1982, and in the 25 years that followed, many new strains were identified and emerged in numerous outbreaks of human ...Escherichia coli O157:H7 is a human pathogen that was first identified from a foodborne outbreak in 1982, and in the 25 years that followed, many new strains were identified and emerged in numerous outbreaks of human disease. Extensive research has been conducted to identify virulence factor genes involved in the pathogenesis of E. coli O157:H7 and many genome sequences of E. coli O157:H7 strains have become available to the scientific community. Here, we provide a comprehensive overview of the research that has been conducted over the first 25 years to identify 394 known or putative virulence factor genes present in the genomes of E. coli O157:H7 strains. Finally, an examination of the conservation of these 394 virulence factor genes across additional genomes of E. coli O157:H7 is provided which summarizes the first 25 years and 13 genomes of this human pathogen.展开更多
Unpasteurized fruit juice has been implicated in outbreaks of Escherichia coli O157:H7 (E. coli O157:H7). Meanwhile, certain fruit, such as pomegranate, contains antimicrobial components. The objective of this study w...Unpasteurized fruit juice has been implicated in outbreaks of Escherichia coli O157:H7 (E. coli O157:H7). Meanwhile, certain fruit, such as pomegranate, contains antimicrobial components. The objective of this study was to investigate the survival and growth characteristics of E. coli O157:H7 on pomegranate juice in laboratory medium and in pomegranate-carrot and pomegranate-apple blend juices at different concentrations. Single strain of E. coli O157:H7 (E0019, H1730, and Cider) was inoculated into brain heart infusion (BHI) broth or a mixture of three strains was inoculated into the blended juices containing pomegranate juice. Our results showed that the addition of pomegranate juice inhibited the growth of tested E. coli O157:H7 in both laboratory medium and blended juices.The antimicrobial activity increased with increased concentrations of pomegranate juice (P < 0.001) and incubation times.The bacterial population was reduced by at least 2 log CFU/ml in BHI broth and juice blend samples in the presence of 20% and 40% Pomegranate juice respectively. Sensory evaluations performed using a 9 point hedonic scale showed significant satisfaction on using 40% pomegranate juice blend with carrot and apple juices. Our study suggests that pomegranate juice could be used as a natural antimicrobial in different food systems including juices to inhibit the growth of E. coli O157:H7.展开更多
Antimicrobial efficacy of three novel organic sanitizers, CHICO WashTM, C8C10 and CG100, was evaluated for the reduction of Escherichia coli O157:H7 during flume-washing of organic leafy greens. Organic formulations a...Antimicrobial efficacy of three novel organic sanitizers, CHICO WashTM, C8C10 and CG100, was evaluated for the reduction of Escherichia coli O157:H7 during flume-washing of organic leafy greens. Organic formulations at various concentrations: CHICO (C3H8I0C3O7) WashTM (1:20 ratio) and C8C10 and CG100 (0.2 and 0.4%), along with the controls: hydrogen-peroxide and water, were used for washing organic baby and mature spinach and romaine and iceberg lettuce. Leafy greens were inoculated with a 2-strain cocktail of E. coli O157:H7 (6 logs CFU/mL) and washed in each treatment for 1 or 2 minutes. The treated leafy greens were stored at 4°C and surviving pathogen populations determined on days 0, 1, and 3 of storage. Organic sanitizers, for both treatment times, significantly (P E. coli O157:H7 on all the leafy greens during storage. Highest reduction (3.4 logs CFU/g) was observed after treatment with CG100 (0.4%) in romaine lettuce, while CHICO WashTM showed greater than 2 logs CFU/g reduction on all the leafy greens, by day 3. This study demonstrates the potential application of organic sanitizers in flume-washing of organic leafy greens for the reduction of E. coli O157:H7.展开更多
Objective： To determine epidemiologic features of an Escherichia coli O157：H7 outbreak occurred in Xuzhou, Jiangsu Province, China in 1999, and assess the incidence of E. coli O157：H7 in diarrhea patients and host ...Objective： To determine epidemiologic features of an Escherichia coli O157：H7 outbreak occurred in Xuzhou, Jiangsu Province, China in 1999, and assess the incidence of E. coli O157：H7 in diarrhea patients and host animals and its relationship with disease onset, and provide a scientific basis for establishing prevention and control strategies. Methods： Epidemiological, microbiological, and molecular methods were performed to identify risk factors and describe the ecology of E. coli O157：H7 in the enviromnent. Results： From May to September, in 1999, 99 cases of E. coli O157：H7 infection were confirmed. Fifty-six patients were enrolled in the case-control study. Bad personal health habits and poor sanitary conditions in the kitchen were associated with increased risks of infection, whereas hand washing was protective. The household survey indicated that residents in the epidemic region during the outbreak had higher than expected rates of diarrhea. The total E. coli O157：H7 carrier rate in the livestock was 12.36%（22/178）, specifically 19.15% in cattle, 12.50% in goat, and 11.11% in swine. Numerical analysis of pulsed-field gel electrophoresis（PFGE） profiles divided strains into two clusters with 77.5% homology. One cluster contained 11 strains isolated from diarrheal patients, foods, and animals. The other cluster comprised 10 strains from patients and environment. Conclusion： In a large outbreak of E. coli O157：H7 infection among predominantly elderly residents in Xuzhou, high rates of carriage of E. coli O157：H7 among host animals most likely resulted in contamination of the environment, thereby leading to the outbreak. Effective and preventive control measures should be taken to avoid contamination, including environmental and family health improvement, good personal hygiene, and safe food handling practices.展开更多
Field effect transistors (FET) based on Single-Walled Carbon Nanotubes (SWNTs) become the hot topic in fields of nano-electronic, clinical diagnostics, environmental testing etc. in recent years. In this paper, we rep...Field effect transistors (FET) based on Single-Walled Carbon Nanotubes (SWNTs) become the hot topic in fields of nano-electronic, clinical diagnostics, environmental testing etc. in recent years. In this paper, we reported a simple, scalable way to enrich semiconducting SWNTs by using HNO3/H2SO4. Then carbon nanotube field-effect transistors (CNTFET) biosensor was fabricated with the enrichment SWNTs for Escherichia coli O157︰H7 detection. The response of each CNTFET was monitored in real time before and after introduction of the Escherichia coli O157︰H7 at various concentrations. The results show that CNT-FET biosensors we fabricated are sensitive to change of concentration of solution and response time is really short.展开更多
To construct and express the fusion protein Stx2B-IntiminC300 of EHEC O157 ： H7, and to further investigate its immunoprophylactic potential, the gene of Stx2B （stx2b） from EHEC O157：H7 chromosome was cloned into ...To construct and express the fusion protein Stx2B-IntiminC300 of EHEC O157 ： H7, and to further investigate its immunoprophylactic potential, the gene of Stx2B （stx2b） from EHEC O157：H7 chromosome was cloned into pMD18-T vector. Thereafter, the amplified gene was cloned into prokary- otic expression plasmid pET-28a （ ＋ ）-eaeC300, which was constructed previously. The recombinant pasmid pET-28a（ ＋ ）-stx2b-eaeC300 was transformed into E. coli BL21 （DE3）. After inducement, the protein Stx2B-IntiminC300 was successfully expressed and analyzed with sodium dodecyl sulfate polyacrylamide gel electrophoresis （SDS-PAGE）, Western blotting and N-terminal amino acid residual sequencing. To evaluate its immunoprophylactic potential, it was primarily purified by ion-exchange chromatography and injected into 30 BALB/c mice with AI（OH）3 in the subscapular region. Ten days after the last booster vaccination, 20 mice were attacked with EHEC O157：H7 lysate and the protective efficacy was observed. In the present study, the gene of Stx2B-intiminC300 was successfully cloned into pET-28a （ ＋ ） vector. The results of SDS-PAGE and Western blotting assay showed that the fusion protein was successfully expressed in the inclusion body form, accounting for 25 % of total expression products, and its molecular weight was about 43 kDa. The result of the N-terminal amino acid residual sequencing showed that it was identical to that of the molecular designed. The purity was about 75 % after primary purification. Animal tests revealed that the fusion protein Stx2B-intiminC300 has elicited high titer of protective antibody relatively. These results demonstrate that the fusion protein Stx2B-IntiminC300 is successfully expressed in prokaryotic expression system and shows certain immunoprophylactic potential.展开更多
基金supported by National Natural Science Foundation of China(31972021)R&D Projects in Key Areas of Guangdong Province(2019B020212003)+4 种基金the Science and Technology Program of Guangzhou,China(202206010177)Guangdong key research and development program(2021B0202060001)Foshan and agricultural academy cooperation projectGuangdong Modern Agriculture project(2022KJ117)Aquatic Products Center Project of GAAS。
文摘Escherichia coli O157:H7 is one of the major foodborne pathogenic bacterial that cause infectious diseases in humans.The previous found that a combination of kojic acid and tea polyphenols exhibited better activity against E.coli O157:H7 than using either alone.This study aimed to explore responses underlying the antibacterial mechanisms of kojic acid and tea polyphenols from the gene level.The functional enrichment analysis by comparing kojic acid and tea polyphenols individually or synergistically against E.coli O157:H7 found that acid resistance systems in kojic acid were activated,and the cell membrane and genomic DNA were destructed in the cells,resulting in“oxygen starvation”.The oxidative stress response triggered by tea polyphenols inhibited both sulfur uptake and the synthesis of ATP,which affected the bacteria's life metabolic process.Interestingly,we found that kojic acid combined with tea polyphenols hindered the uptake of iron that played an essential role in the synthesis of DNA,respiration,tricarboxylic acid cycle.The results suggested that the iron uptake pathways may represent a novel approach for kojic acid and tea polyphenols synergistically against E.coli O157:H7 and provided a theoretical basis for bacterial pathogen control in the food industry.
文摘To prepare monoclonal antibodies （MAb） and antisera specific for Escherichia coli （E.coli） O157, and to develop a sandwich enzyme-linked immunosorbent assay （ELISA） to detect Eocoli O157 in foods. Methods Spleen cells from BALB/c mice immunized with the somatic antigen of E.coli O157：H7 were fused with routine Sp2/0 myeloma cells. The hybridoma cell line specific for E.coli O157 was established after having been subcloned. Antisera specific for E.coli O157 was prepared by intravenous injection into New Zealand rabbits with a stain of E.coli O157：H7. The sandwich ELISA was developed with the polyclonal antibody as the capture antibody and the MAb 3A5 as the detection antibody. The inoculated ground poultry meat and pasteurized miLk were tested to confirm efficiency of the method. Results MAb 3A5 specific for E.coli O157 and O 113：H21 belonged to subtype IgM. The ascetic titers of the antibody was 1：1× 10^6. No cross-reactivity of the MAb was observed with strains of Salmonella spp, Yersinia enterocolitica, Shigella dysenteriae, etc. The purified polyclonal antibody had a titer of 1： 1× 10^5 with E.coli O 157. The detection limit of this sandwich ELISA was 10^3- 10^4 cfu E.coli O157/mL in pure culture with a high specificity, which was characterized by every non-O157 strain with negative response. With 10h enrichment procedure, E.coli O157：H7 recovered well from inoculated ground poultry meat and pasteurized milk at levels of 0.1 cfu/g and 0.1 cfu/mL. Conclusion MAb 3A5 specific for E.coli O 157 and O 113：H21 can be produced by immunizing BALB/c mice with a strain of E.coli O157：H7. Then a sandwich ELISA can be developed with the polyclonal antibody as the capture antibody and the MAb 3A5 as the detection antibody. The method is proved to be a sensitive and specific technique to detect low number of E.coli O157 in food.
基金the Islamic Azad University, Jahrom branch,for their executive support of this project
文摘Objective:To survey the prevalence severe diarrhea arising from these bacteria in children under 5 years old in Marvdasht.Methods:In this study faecal sample from 615 children aged <5years old who were hospitalized lor gastroenteritis in Fars hospitals in Iran were collected and then enriched in Escherichia coli(E.coli) broth and modified tryplone soy broth with novobiocin media,fermentation of sorbitol,lactose and β— glucoronidase activity of isolated strains was examined by CT—SMAC,VRBA and chromogenic media respectively.Then isolation of E.coli O157:H7 have been confirmed with the use of specific antisera and with multiplex PCR method presence of virulence genes including:xtx_1.stx_2,eae.A.hly has been analyzed.Results:E.coli O157:H7 was detected in 7(1.14%) stool specimens.A significanl difference was seen between detection rale of isolated bacteria from age groups 18-23 months and other age groups(P=0.004).Out of considered virulence genes.only 1 of the isolated strains(0.16%)he stx,and eaeA genes were seen and also all isolated hacleria had resistance to penicillin,ampicillin and erythromycin antibiotics.Conclusions:We found thai children < 2 years of age were at highest risk of infection with E.coli O157:H7.Regarding severity of E.coli O157:H7 pathogenesis,low infectious dose and lack of routine assay for detection ol these bacleria in clinical laboratory,further and completed studies on diagnosis and genolyping of this E.coli O157:H7 strain has been recommended.
基金Project supported by the Talent Foundation of Zhejiang Province (No. R205502)the Program of Education Department of Zhejiang Province (No. 20040197), China
文摘By means of the specific immuno-recognition and ultra-sensitive mass detection, a quartz crystal microbalance (QCM) biosensor for Escherichia coli O157:H7 detection was developed in this work. As a suitable surfactant, 16-mercaptohexadecanoic acid (MHDA) was introduced onto the Au surface of QCM, and then self-assembled with N-hydroxysuccinimide (NHS) raster as a reactive intermediate to provide an active interface for the specific antibody immobilization. The binding of target bacteria with the immobilized antibodies decreased the sensor’s resonant frequency, and the frequency shift was correlated to the bacterial concentration. The stepwise assembly of the immunosensor was characterized by means of the electrochemical techniques. Using the immersion-dry-immersion procedure, this QCM biosensor could detect 2.0×102 colony forming units (CFU)/ml E. coli O157:H7. In order to reduce the fabrication time, a polyelectrolyte layer-by-layer self-assembly (LBL-SA) method was adopted for fast construction. Finally, the reproducibility of this biosensor was discussed.
基金supported by the National Key Research and Development Program of China(No.2018YFD0501600)National Natural Science Foundation of China(No.31972056)Shanghai Agriculture Applied Technology Development Program,China(Grant No.2019-02-08-00-07-F01152).
文摘This study investigated the intervention effects of Lactobacillus casei LC2W in murine(SPF C57BL/c)challenge infection models induced by Escherichia coli O157:H7.Mice were fed streptomycin with water for 3 days prior to intragastric gavage by E.coli O157:H7(Control)or L.casei LC2W together with E.coli O157:H7(Intervention)to explore the role of L.casei LC2W by biochemical indicators,histological evaluation,expression of colonic pro-inflammatory and intestinal barrier factors related to enteritis.Results showed that the administration of L.casei LC2W was able to alleviate the symptoms of colitis induced by E.coli O157:H7,exhibiting lower weight loss as well as more intact colon tissue.Furthermore,L.casei LC2W could down-regulate the expression of pro-inflammatory cytokines(TNF-α,IL-6,and IL-1β)and protect the intestinal barrier function by improving the level of MUC2,ZO-1 and E-cadherin 1 compared to the control group.These results demonstrate that L.casei LC2W can reduce the severity of E.coli O157:H7 infection,and suggest L.casei LC2W may maintain the immune balance and intestinal barrier to reduce colitis.In addition,we found the effect of intervention is similar to that of prevention,which is better than that of treatment.
基金Supported by the Agro-scientific Research of China(201203009)the Ministry of Agriculture,China
文摘The antagonistic activity of Lactobacillus acidophilus KLDS1.0901, KLDS1.0902, KLDSI.1003 and NCFM against Escherichia coli O157 ： H7 were investigated in this study. The culture supematants of all the L. acidophilus stains showed high bacteriostatic activities against E. coli O 157 ： H7 and the bacteriostatic substances of their Cell-Free Supernatants （CFS） were preliminarily determined from organic acids. The bacteriostatic activity from CFS or viable L. acidophilus against E. coli O157 ： H7 was also assessed by using co-incubation methods, CFS had high bactericidal activity against E. coli O157 ： H7, no viable E. coli O157 ： H7 was detected when 5×10^7 CfU ofE. coli O157 ： H7 was added to 5 mL of CFS and incubated at 37℃ for 2 h. However, L. acidophilus themselves had no bacteriostatic activity after directly contacted with E. coli O157 ： H7. The inhibition E. coli O157 ： H7 adhesion and colonization of L. acidophilus were also investigated based on competition, exclusion and displacement assays. L. acidophilus KLDS1.0901, KLDSI.1003 and NCFM strains were effective to displace E. coli O157 ： H7 from a Caco-2 cell layer in competition and exclusion assays. However, in displacement assay, all of the strains showed no significant antagonistic activities. Meanwhile, the probiotic potential of L. acidophilus strains was investigated based on adhesion assay to Caco-2 cells and anti- inflammatory effects by IL-8 produced in Caco-2 cells. The adhesion ability and anti-inflammatory effects of L. acidophilus strains showed a strain-dependent manner. In general, L. acidophilus KLDS 1.0901 and NCFM showed better probiotic potential than KLDS1.0902 and KLDSI.1003. Thus, the use ofL. acidophilus KLDS1.0901 and NCFM to prevent or treat of diseases associated induced E. coli O157 ： H7 in vivo was suggested.
基金Supported by the Key Military Medical Science and Technique Program during the 10~(th) Five Year Plan Period, No. 01L006
文摘AIM： To establish the rapid, specific, and sensitive method for detecting O157：H7 with DNA microchips. METHODS： Specific oligonucleotide probes （26-28 nt） of bacterial antigenic and virulent genes of E. coliO157：H7 and other related pathogen genes were pre-synthesized and immobilized on a solid support to make microchips. The four genes encoding O157 somatic antigen （rfbE）, H7 flagellar antigen （fliC） and toxins （SLT1, SLT2） were monitored by multiplex PCR with four pairs of specific primers. Fluorescence-Cy3 labeled samples for hybridization were generated by PCR with Cy3-1abeled single prime. Hybridization was performed for 60 min at 45 ℃. Microchip images were taken using a confocal fluorescent scanner. RESULTS： Twelve different bacterial strains were detected with various combinations of four virulent genes. All the O157：H7 strains yielded positive results by multiplex PCR. The size of the PCR products generated with these primers varied from 210 to 678 bp. All the rfbE/fliC/SLT1/SLT2 probes specifically recognized Cy3-1abeled fluorescent samples from O157：H7 strains, or strains containing O157 and H7 genes. No cross hybridization of O157：H7 fluorescent samples occurred in other probes. Non-O157：H7 pathogens failed to yield any signal under comparable conditions. If the Cy3-1abeled fluorescent product of O157 single PCR was diluted 50-fold, no signal was found in agarose gel electrophoresis, but a positive signal was found in microarray hybridization. CONCLUSION： Microarray analysis of O157：H7 is a rapid, specific, and efficient method for identification and detection of bacterial pathogens.
文摘The objective of this study was to determine the effect of sodium lactate on the survival of Listeria monocytogenes, Escherichia coli O157: H7, and Salmonella spp. in cooked ham during storage at refrigerated and abuse temperatures. Cooked ham was added with 0% - 3% lactate, inoculated with a multiple-strain mixture of L. monocytogenes, E. coli O157: H7, or Salmonella spp. and stored at 4oC - 15oC for up to 35 day. The growth of the three pathogens was inhibited in ham containing 3% lactate, and no growth of E. coli O157: H7 and Salmonella spp. occurred at the lowest storage tem- peratures of 6 and 8oC, respectively. In ham containing no lactate, the average growth rates were 0.256 - 0.380 log CFU/day for L. monocytogenes at 4oC - 8oC, 0.242 - 0.315 log CFU/day for E. coli O157: H7 at 8oC - 15oC, and 0.249 - 0.328 log CFU/day for Salmonella spp. at 10oC - 15oC. The addition of 1% or 2% lactate significantly (P < 0.05) reduced the growth rates of the three pathogens, and the effect was more profound at lower temperatures. Salmonella spp. were more sensitive to the effect of lactate than L. monocytogenes and E. coli O157: H7. Polynomial models were developed to describe the growth rates of the three pathogens as affected by the lactate concentration and storage tem- perature. Results from this study demonstrate the effect of lactate on the growth of L. monocytogenes, E. coli O157: H7, and Salmonella spp. in cooked ham and indicate the effective lactate concentrations and storage temperatures that can be used to enhance the microbiological safety of ready-to-eat ham products.
文摘In order to test if the intestinal bacteria play an important role in antibacterial ability of earthworm, we chose Escherichia coli O157:H7, an anthropozoonosis pathogen, as a biological stressor and studied the change of intestinal bacteria community of earthworm by PCR-DGGE analysis. Results showed that the pathogen merely existed 1 - 3 days, then almost disappeared after through the earthworm’s gut. In this period, the diversity and abundance index of intestinal bacteria increased first, then decreased, and finally kept stably after 7 days. The result demonstrated that the intestinal bacteria of earthworm had ability of adjust community structure to eliminate the pathogen E. coli O157:H7, and the amount of bacteria Bacillus increased significantly, which might be the positive antagonism to E. coli O157:H7.
文摘Escherichia coli O157:H7 is a human pathogen that was first identified from a foodborne outbreak in 1982, and in the 25 years that followed, many new strains were identified and emerged in numerous outbreaks of human disease. Extensive research has been conducted to identify virulence factor genes involved in the pathogenesis of E. coli O157:H7 and many genome sequences of E. coli O157:H7 strains have become available to the scientific community. Here, we provide a comprehensive overview of the research that has been conducted over the first 25 years to identify 394 known or putative virulence factor genes present in the genomes of E. coli O157:H7 strains. Finally, an examination of the conservation of these 394 virulence factor genes across additional genomes of E. coli O157:H7 is provided which summarizes the first 25 years and 13 genomes of this human pathogen.
文摘Unpasteurized fruit juice has been implicated in outbreaks of Escherichia coli O157:H7 (E. coli O157:H7). Meanwhile, certain fruit, such as pomegranate, contains antimicrobial components. The objective of this study was to investigate the survival and growth characteristics of E. coli O157:H7 on pomegranate juice in laboratory medium and in pomegranate-carrot and pomegranate-apple blend juices at different concentrations. Single strain of E. coli O157:H7 (E0019, H1730, and Cider) was inoculated into brain heart infusion (BHI) broth or a mixture of three strains was inoculated into the blended juices containing pomegranate juice. Our results showed that the addition of pomegranate juice inhibited the growth of tested E. coli O157:H7 in both laboratory medium and blended juices.The antimicrobial activity increased with increased concentrations of pomegranate juice (P < 0.001) and incubation times.The bacterial population was reduced by at least 2 log CFU/ml in BHI broth and juice blend samples in the presence of 20% and 40% Pomegranate juice respectively. Sensory evaluations performed using a 9 point hedonic scale showed significant satisfaction on using 40% pomegranate juice blend with carrot and apple juices. Our study suggests that pomegranate juice could be used as a natural antimicrobial in different food systems including juices to inhibit the growth of E. coli O157:H7.
文摘Antimicrobial efficacy of three novel organic sanitizers, CHICO WashTM, C8C10 and CG100, was evaluated for the reduction of Escherichia coli O157:H7 during flume-washing of organic leafy greens. Organic formulations at various concentrations: CHICO (C3H8I0C3O7) WashTM (1:20 ratio) and C8C10 and CG100 (0.2 and 0.4%), along with the controls: hydrogen-peroxide and water, were used for washing organic baby and mature spinach and romaine and iceberg lettuce. Leafy greens were inoculated with a 2-strain cocktail of E. coli O157:H7 (6 logs CFU/mL) and washed in each treatment for 1 or 2 minutes. The treated leafy greens were stored at 4°C and surviving pathogen populations determined on days 0, 1, and 3 of storage. Organic sanitizers, for both treatment times, significantly (P E. coli O157:H7 on all the leafy greens during storage. Highest reduction (3.4 logs CFU/g) was observed after treatment with CG100 (0.4%) in romaine lettuce, while CHICO WashTM showed greater than 2 logs CFU/g reduction on all the leafy greens, by day 3. This study demonstrates the potential application of organic sanitizers in flume-washing of organic leafy greens for the reduction of E. coli O157:H7.
基金supported by National High-tech R&D Program (863 Program) (2007AA02Z409)
文摘Objective： To determine epidemiologic features of an Escherichia coli O157：H7 outbreak occurred in Xuzhou, Jiangsu Province, China in 1999, and assess the incidence of E. coli O157：H7 in diarrhea patients and host animals and its relationship with disease onset, and provide a scientific basis for establishing prevention and control strategies. Methods： Epidemiological, microbiological, and molecular methods were performed to identify risk factors and describe the ecology of E. coli O157：H7 in the enviromnent. Results： From May to September, in 1999, 99 cases of E. coli O157：H7 infection were confirmed. Fifty-six patients were enrolled in the case-control study. Bad personal health habits and poor sanitary conditions in the kitchen were associated with increased risks of infection, whereas hand washing was protective. The household survey indicated that residents in the epidemic region during the outbreak had higher than expected rates of diarrhea. The total E. coli O157：H7 carrier rate in the livestock was 12.36%（22/178）, specifically 19.15% in cattle, 12.50% in goat, and 11.11% in swine. Numerical analysis of pulsed-field gel electrophoresis（PFGE） profiles divided strains into two clusters with 77.5% homology. One cluster contained 11 strains isolated from diarrheal patients, foods, and animals. The other cluster comprised 10 strains from patients and environment. Conclusion： In a large outbreak of E. coli O157：H7 infection among predominantly elderly residents in Xuzhou, high rates of carriage of E. coli O157：H7 among host animals most likely resulted in contamination of the environment, thereby leading to the outbreak. Effective and preventive control measures should be taken to avoid contamination, including environmental and family health improvement, good personal hygiene, and safe food handling practices.
文摘Field effect transistors (FET) based on Single-Walled Carbon Nanotubes (SWNTs) become the hot topic in fields of nano-electronic, clinical diagnostics, environmental testing etc. in recent years. In this paper, we reported a simple, scalable way to enrich semiconducting SWNTs by using HNO3/H2SO4. Then carbon nanotube field-effect transistors (CNTFET) biosensor was fabricated with the enrichment SWNTs for Escherichia coli O157︰H7 detection. The response of each CNTFET was monitored in real time before and after introduction of the Escherichia coli O157︰H7 at various concentrations. The results show that CNT-FET biosensors we fabricated are sensitive to change of concentration of solution and response time is really short.
文摘To construct and express the fusion protein Stx2B-IntiminC300 of EHEC O157 ： H7, and to further investigate its immunoprophylactic potential, the gene of Stx2B （stx2b） from EHEC O157：H7 chromosome was cloned into pMD18-T vector. Thereafter, the amplified gene was cloned into prokary- otic expression plasmid pET-28a （ ＋ ）-eaeC300, which was constructed previously. The recombinant pasmid pET-28a（ ＋ ）-stx2b-eaeC300 was transformed into E. coli BL21 （DE3）. After inducement, the protein Stx2B-IntiminC300 was successfully expressed and analyzed with sodium dodecyl sulfate polyacrylamide gel electrophoresis （SDS-PAGE）, Western blotting and N-terminal amino acid residual sequencing. To evaluate its immunoprophylactic potential, it was primarily purified by ion-exchange chromatography and injected into 30 BALB/c mice with AI（OH）3 in the subscapular region. Ten days after the last booster vaccination, 20 mice were attacked with EHEC O157：H7 lysate and the protective efficacy was observed. In the present study, the gene of Stx2B-intiminC300 was successfully cloned into pET-28a （ ＋ ） vector. The results of SDS-PAGE and Western blotting assay showed that the fusion protein was successfully expressed in the inclusion body form, accounting for 25 % of total expression products, and its molecular weight was about 43 kDa. The result of the N-terminal amino acid residual sequencing showed that it was identical to that of the molecular designed. The purity was about 75 % after primary purification. Animal tests revealed that the fusion protein Stx2B-intiminC300 has elicited high titer of protective antibody relatively. These results demonstrate that the fusion protein Stx2B-IntiminC300 is successfully expressed in prokaryotic expression system and shows certain immunoprophylactic potential.
