Abstract:
:In the current study, a duplex PCR-ELISA method was developed targeting the specific genes, invA of Salmonella spp. and rfbE of Escherichia coli O157: H7, to detect one or both bacteria in food. In brief, PCR product amplified by PCR primer labeled with digoxin at the 5'-end and a probe labeled with biotin at the 3'-end can form dimer by nucleic acid hybridization which can be captured by binding of biotin to streptomycin coated in ELISA plate before using enzyme-labeled anti-digoxin antibody and substrate to develop color. Also, evaluation of the duplex PCR-ELISA method was conducted in different food samples including milk, juice, cabbage, shrimp, chicken, pork and beef. Results indicated that the duplex PCR-ELISA developed here was specific when using 25 non-target bacteria strains as controls and was sensitive with a limit of detection (LOD) of 1 CFU/mL, 1, 000 times higher than that of the duplex PCR method and was repeatable regardless of inter- and intra-batch variations. The duplex PCR-ELISA method established in the present study has proven to be highly specific, sensitive and repeatable. It has the potential to be applied in such fields as clinical diagnosis of food-borne diseases, food hygiene monitoring and pathogen detection in food.
journal_name
J Microbiol Methodsjournal_title
Journal of microbiological methodsauthors
Hu J,Huang R,Wang Y,Wei X,Wang Z,Geng Y,Jing J,Gao H,Sun X,Dong C,Jiang Cdoi
10.1016/j.mimet.2018.10.017subject
Has Abstractpub_date
2018-11-01 00:00:00pages
127-133eissn
0167-7012issn
1872-8359pii
S0167-7012(18)30844-3journal_volume
154pub_type
杂志文章abstract::The use of DNA microarrays for detection and identification of bacteria and genes of interest from various environments (e.g. soil, sediment, water column...) is a major challenge for microbiologists working on functional diversity. So far, most of the genomic methods that have been described rely on the use of taxono...
journal_title:Journal of microbiological methods
pub_type: 杂志文章,评审
doi:10.1016/j.mimet.2006.11.007
更新日期:2007-05-01 00:00:00
abstract::In wastewater, biological phosphate removal can fail because of the presence of glycogen accumulating organism (GAO), therefore measuring glycogen stored in microbial cultures provides information on the bacterial population type. Once glycogen is hydrolysed to glucose it was accurately measured using a human glucose ...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2014.03.002
更新日期:2014-05-01 00:00:00
abstract::Leishmaniasis, caused by protozoa belonging to the genus Leishmania, is an important public health problem found in >90 countries and with still limited options for treatment. Development of new anti-leishmanial drugs is an urgent need and the identification of new active compounds is a limiting factor that can be acc...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2019.105732
更新日期:2019-11-01 00:00:00
abstract::An effective microbial preservation technology for the long-term storage of viable prokaryotic cells is described. The method combines an almost instantaneous drying step with minimal stress to the cells during drying to maximize survival on rehydration. This is achieved by contact of a microlitre aliquot of a bacteri...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2005.02.014
更新日期:2005-10-01 00:00:00
abstract::Yeast cell wall fractions have been proposed to bind enteropathogenic bacteria. The aim of this study was to develop a quantitative assay by measuring the optical density as growth parameter of adhering bacteria. The exponential growth phase of adhering bacteria was determined by optical density reading and compared w...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2010.08.016
更新日期:2010-11-01 00:00:00
abstract::Using robotics and the MPN technique, a 96-microwell method was developed to compare two procedures for enumeration of viable chlorine-treated B. atrophaeus spores: broth-culture enrichment followed by real-time polymerase chain reaction analysis; and filter plating on agar. Recoveries of chlorine-treated spores were ...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2007.07.011
更新日期:2007-10-01 00:00:00
abstract::In this study, food-borne yeast isolates (n=96), comprising at least 33 species, were identified using MALDI-TOF MS and conventional methods (API ID 32 C and Phoenix Yeast ID). Discrepancies of both methods were resolved by sequencing the ITS1-5.8S-rRNA-ITS2 region. For ten isolates, mainly classified to Rhodotorula a...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2014.08.021
更新日期:2014-11-01 00:00:00
abstract::There is a paucity of knowledge on gene expression in dead bacterial cells. Why would this knowledge be useful? The cells are dead. However, the time duration of gene expression following cell death is often unknown, and possibly in the order of minutes. In addition, it is a challenge to determine if bacterial cells a...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2012.04.004
更新日期:2012-07-01 00:00:00
abstract::Several outbreaks of foodborne yersiniosis have been documented and this disease continues to be source of infections transmitted through foods. The selective agars most commonly used to isolate Yersinia enterocolitica in clinical, food and environmental samples, cefsulodin-irgasan-novobiocin (CIN) and MacConkey (MAC)...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2007.11.019
更新日期:2008-02-01 00:00:00
abstract::A rapid bead assay for detecting pathogenic bacteria with a simple microfluidic chip-based system was developed. Five oligonucleotide probes corresponding to the 16S rRNA of the targeted bacteria were coupled covalently to fluorescent beads. Four species of bacteria (Escherichia coli, Salmonella enterica subsp. enteri...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2006.03.014
更新日期:2006-11-01 00:00:00
abstract::This paper describes a method that facilitates the extraction of PCR-compatible DNA from different activated sludge samples. The approach involves a novel preprocessing step in DNA extraction, which removes potential PCR inhibitors. The sludge was washed with different ratios of acetone and petroleum ether after pretr...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/s0167-7012(02)00185-9
更新日期:2003-03-01 00:00:00
abstract::Fluorescence in situ hybridization has been used for the identification and analysis of populations of the dinoflagellate Symbiodinium that lives symbiotically in marine invertebrates. Conditions for in situ hybridization of Symbiodinium were optimized and used to identify the clade to which the isolate belongs using ...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/s0167-7012(02)00250-6
更新日期:2003-06-01 00:00:00
abstract::A 16S rRNA gene-based fingerprinting method was developed for the identification of Azotobacteraceae and tested onto 48 soil isolates and 28 reference strains belonging to the free-living nitrogen-fixing bacterial group and to the most common species found in soil samples. According to this method, the 16S rRNA gene w...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2004.01.006
更新日期:2004-05-01 00:00:00
abstract::Analysis of microbial community structure in complex environmental samples using nucleic acid techniques requires efficient unbiased DNA extraction procedures; however, humic acids and other contaminants complicate the isolation of PCR-amplifiable DNA from compost and other organic-rich samples. In this study, combina...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/s0167-7012(01)00377-3
更新日期:2002-05-01 00:00:00
abstract::The Carba NP test was developed to detect carbapenemase-producing Enterobacteriaceae, and uses imipenem as the reaction substrate. In Japan, IMP-6 metallo-β-lactamase (MBL) producers, which are usually resistant to meropenem but susceptible to imipenem, and IMP-1 MBL producers, which are usually resistant to both carb...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2016.12.015
更新日期:2017-02-01 00:00:00
abstract::Real-time TaqMan-PCR assays were developed for detection, differentiation and absolute quantification of the pathogenic subspecies of Clavibacter michiganensis (Cm) in one single PCR run. The designed primer pair, targeting intergenic sequences of the rRNA operon (ITS) common in all subspecies, was suitable for the am...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/s0167-7012(02)00152-5
更新日期:2003-01-01 00:00:00
abstract::The direct counts of bacteria inhabiting coral mucus were performed by flow cytometry testing four fluorescent dyes (SYBR®Green I, HCS, TOPRO®3, SYTO®62) with three different scleractinian species. Results obtained with SYTO62 were the most reliable based on the comparison with standardized epifluorescence counts and ...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2016.05.032
更新日期:2016-09-01 00:00:00
abstract::A digital image analysis method previously used to evaluate leaf color changes due to nutritional changes was modified to measure the severity of several foliar fungal diseases. Images captured with a flatbed scanner or digital camera were analyzed with a freely available software package, Scion Image, to measure chan...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2008.03.008
更新日期:2008-08-01 00:00:00
abstract::Centrifugation is a necessary concentrating step for the detection of Mycobacterium tuberculosis in a liquid culture. However, centrifugation is biologically hazardous and presents an obstacle in the development of an automated culture system. A bead-based bacterial concentration method, TRICORE, was recently develope...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2012.04.018
更新日期:2012-09-01 00:00:00
abstract::A fused protein composed of a carbohydrate-binding module and green fluorescence protein (GFP) was developed to measure the exopolysaccharides (EPShs) present in Escherichia coli microcolonies. The cleavage of the GFP part of this protein using a site-specific protease allowed for the non-invasive and quantitative eva...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2015.05.013
更新日期:2015-07-01 00:00:00
abstract::In this study an on-line electrochemical method was developed to examine the one-step growth cycle (OSGC) of the bacteriophage Bam35. The on-line conditions for monitoring the OSGC and the effect of aeration on the duration of the OSGC were defined. The data indicate that binding of phenyldicarbaundecaborane anions to...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2006.12.023
更新日期:2007-04-01 00:00:00
abstract::A simple, fast and cost-effective method for the taxon-specific separation of various 16S rRNAs was developed. This primer extension mediated separation of different RNA species was obtained on agarose gels. The method provides an easy way for the investigation of substrate mediated radioisotope incorporation into the...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2009.07.010
更新日期:2009-10-01 00:00:00
abstract::Quantification of the biosurfactants produced by a variety of microorganisms is a time taking and difficult task due to the lack of rapid, efficient and accurate methods. This work presents a simple turbidometric method for quantification of crude biosurfactants based on their property to become insoluble at low pH va...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2008.09.010
更新日期:2009-01-01 00:00:00
abstract::Central nervous system (CNS) infections have multiple potential causative agents for which simultaneous pathogen screening can provide a useful tool. This study evaluated a multiplexed microarray for the simultaneous detection of antibodies against CNS pathogens. The performance of selected microarray antigens for the...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2014.02.011
更新日期:2014-05-01 00:00:00
abstract::A strain-specific assay was developed for the detection of viable Lactobacillus on cattle feed. The DNA sequences of the 16S rRNA gene and four different 16S/23S rRNA intergenic spacer regions (ISR) from Lactobacillus sp. HOFG1 were determined. Based on these sequences, a strain-specific primer was designed for the am...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2004.12.002
更新日期:2005-05-01 00:00:00
abstract::Nosocomial infections caused by Stenotrophomonas maltophilia resistant to SXT are increasingly reported worldwide. In this study, a novel melting-curve based multiplex real-time PCR assay for the simultaneous detection of the ssrA and sul1, sul2 and sul3 genes was first established. The assays were performed on a Roch...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2018.12.002
更新日期:2019-01-01 00:00:00
abstract::Neisseria gonorrhoeae (NG) has developed resistance to most antibiotics, making it increasingly difficult to treat. Previous studies have predicted antimicrobial NG susceptibility based on the antimicrobial gene target DNA gyrase subunit A (gyrA) codon serine 91 and the penicillin-binding protein 2 (penA) using Roche ...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2019.105783
更新日期:2020-01-01 00:00:00
abstract::The European Union Reference Laboratory (EURL) for Listeria monocytogenes (Lm) collaborates with a network of 35 National Reference Laboratories (NRLs) throughout Europe. Most of these NRLs are in charge of detecting and typing Lm strains from food, environment and animals, which are isolated nationally. The past few ...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2014.06.001
更新日期:2014-09-01 00:00:00
abstract::Cryptosporidium is a zoonotic coccidian parasite associated with diarrhea, and the disinfectant-resistant oocysts are threats to public health even in industrialized countries. In order to make an accurate assessment of the risk to public health, a detection method that has a high recovery rate of oocysts in water is ...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/s0167-7012(03)00209-4
更新日期:2003-12-01 00:00:00
abstract::Ruthenium red is a polycationic stain used to visualize acid polysaccharides on the outer surface of cells. Ruthenium red staining followed by electron microscopic analysis was used to demonstrate the presence of an external glycoprotein layer surrounding the spore of both Bacillus anthracis and Bacillus subtilis. Thi...
journal_title:Journal of microbiological methods
pub_type: 杂志文章
doi:10.1016/j.mimet.2004.02.012
更新日期:2004-07-01 00:00:00