Identification of shiga toxin-producing bacteria by a new immuno-capture toxin gene PCR

Luo, Weiyi

doi:10.1016/s0378-1097(02)01005-4

Cited by 3 publications

(2 citation statements)

References 12 publications

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“…2002). PCR‐based diagnostic methods are used in a qualitative, not quantitative format (Luo et al. 2002).…”

Section: Introductionmentioning

confidence: 99%

“…Multiplex polymerase chain reaction (PCR) analysis has been used to detect the genes specific for E. coli O157:H7 . PCR-based diagnostic methods are used in a qualitative, not quantitative format (Luo et al 2002). Because as low as 10 E. coli O157:H7 cells have been found to sicken people (Doyle et al 1997), it is imperative to be able to detect this extremely low dosage of E. coli O157:H7 in foods.…”

Section: Introductionmentioning

confidence: 99%

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REAL‐TIME REVERSE TRANSCRIPTION PCR DETECTION OF VIABLE SHIGA TOXIN‐PRODUCING ESCHERICHIA COLI O157:H7 IN FOOD

Di¹,

Tumer

2010

Journal of Food Safety

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Shiga toxin (Stx)‐producing Escherichia coli (STEC), particularly E. coli O157:H7, have been associated with food‐related outbreaks and have become a global health concern. Detection of low numbers of viable STEC in food is undoubtedly challenging. The present study demonstrated that 7 × 102 to 7 × 103 cfu E. coli O157:H7 cells in 50 mL of inoculated food samples including apple juice, lettuce and ground beef could be concentrated through a two‐step filtration technique. Bacterial RNA was effectively isolated from the concentrated E. coli O157:H7 cells with the combination of Trizol Max Bacterial RNA Isolation Enhancement Reagent and TriReagent®. Real‐time reverse transcription polymerase chain reaction with seven sets of primers targeting virulence and serotype genes of E. coli O157:H7 specifically detected the bacterial RNA representing 1 × 102 to 1 × 103 cfu viable bactersial cells. The total detection time from sampling to measurement was approximately 4 h. This study provides a rapid and specific detection method for viable STEC in food. PRACTICAL APPLICATIONS Rapid detection of viable STEC including Escherichia coli O157:H7 cells in food has been difficult, especially when present in low numbers. This study has shown that 7 × 102 to 7 × 103 cfu E. coli O157:H7 in 50 mL of inoculated food samples, i.e., 14–140 cfu/mL bacterial cells, could be readily concentrated and the bacterial RNA could be detected by real‐time reverse transcription polymerase chain reaction (PCR) in a relatively short time. Use of seven primer sets makes this method very specific for real‐time PCR detection of E. coli O157:H7.

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“…2002). PCR‐based diagnostic methods are used in a qualitative, not quantitative format (Luo et al. 2002).…”

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

REAL‐TIME REVERSE TRANSCRIPTION PCR DETECTION OF VIABLE SHIGA TOXIN‐PRODUCING ESCHERICHIA COLI O157:H7 IN FOOD

Di¹,

Tumer

2010

Journal of Food Safety

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Analysis of Hepatitis B Virus‐Immunoglobulin Isotype Complexes by a Novel Immuno‐Capture Polymerase Chain Reaction Method

et al. 2003

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Hepatitis B virus (HBV) can be present in the circulating blood either as free virus or as a virion-immunoglobulin (Ig) complex. Presently, it remains unclear what specific role each Ig plays in the clearance of HBV. In this study, a novel method that combined immuno-capture and polymerase chain reaction (PCR) amplification was used for detecting and distinguishing different HBV-Ig complexes. Three isotypes of Ig (IgM, IgG and IgA) bound to HBV were detected in the four clinically defined stages of HBV infection in 108 patients. The results showed that all the three isotypes of Ig could bind to HBV, and the patterns of HBV-Ig complexes varied according to disease categories. Interestingly, the frequency of HBV DNA-Ig complexes in hepatitis B e antigen (HBeAg)-positive patients was significantly lower than that in HBeAg-negative patients. All the data suggest that the three isotypes of HBV DNA-Ig circulating immune complex (CIC) may have different biological meanings. In summary, HBV bound to an antibody is a common feature of hepatitis B, and immuno-capture PCR is a valuable method for the analysis of the composition of the immune complexes. The detection of HBV-Ig complexes may provide new and valuable insights into HBV pathogenesis.

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A Pentaplex PCR Assay for the Detection and Differentiation ofShigellaSpecies

Ojha

Yean

Ismail

et al. 2013

BioMed Research International

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The magnitude of shigellosis in developing countries is largely unknown because an affordable detection method is not available. Current laboratory diagnosis of Shigella spp. is laborious and time consuming and has low sensitivity. Hence, in the present study, a molecular-based diagnostic assay which amplifies simultaneously four specific genes to identify invC for Shigella genus, rfc for S. flexneri, wbgZ for S. sonnei, and rfpB for S. dysenteriae, as well as one internal control (ompA) gene, was developed in a single reaction to detect and differentiate Shigella spp. Validation with 120 Shigella strains and 37 non-Shigella strains yielded 100% specificity. The sensitivity of the PCR was 100 pg of genomic DNA, 5.4 × 104 CFU/ml, or approximately 120 CFU per reaction mixture of bacteria. The sensitivity of the pentaplex PCR assay was further improved following preincubation of the stool samples in Gram-negative broth. A preliminary study with 30 diarrhoeal specimens resulted in no cross-reaction with other non-Shigella strains tested. We conclude that the developed pentaplex PCR assay is robust and can provide information about the four target genes that are essential for the identification of the Shigella genus and the three Shigella species responsible for the majority of shigellosis cases.

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Identification of shiga toxin-producing bacteria by a new immuno-capture toxin gene PCR

Cited by 3 publications

References 12 publications

REAL‐TIME REVERSE TRANSCRIPTION PCR DETECTION OF VIABLE SHIGA TOXIN‐PRODUCING ESCHERICHIA COLI O157:H7 IN FOOD

REAL‐TIME REVERSE TRANSCRIPTION PCR DETECTION OF VIABLE SHIGA TOXIN‐PRODUCING ESCHERICHIA COLI O157:H7 IN FOOD

Analysis of Hepatitis B Virus‐Immunoglobulin Isotype Complexes by a Novel Immuno‐Capture Polymerase Chain Reaction Method

A Pentaplex PCR Assay for the Detection and Differentiation ofShigellaSpecies

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