Development and use of 16S rRNA gene targeted PCR primers for the identification of
            <i>Escherichia coli</i>
            cells in water

Tsen, Hau‐Yang; Lin, Chi-Ying; Chi, Wan-Rong

doi:10.1046/j.1365-2672.1998.853535.x

Cited by 98 publications

(44 citation statements)

References 15 publications

(25 reference statements)

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“…A probe designed to detect E. coli, but not Shigella spp., based upon 16S rRNA sequence differences that do exist would then detect several other bacteria, in particular bacteria within the Enterobacteriaceae family. This lack of 100% species-specific rRNA target sequences of E. coli is well known (22). Although Shigella spp.…”

Section: Discussionmentioning

confidence: 99%

“…The concept of coliforms has been further refined by differentiating between environmental coliforms and fecal coliforms exhibiting thermotolerance, such as E. coli. The development of E. coli-specific tests based on either detection of ␤-D-glucoronidase activity using media containing 4-methylumbelliferyl-␤-D-glucuronide (MUG) (7) or molecular methods (2,22) now allows fecal contamination to be monitored by specific detection of E. coli. In this way, no further identification is required and positive reactions due to coliforms not associated with fecal contamination are eliminated.…”

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confidence: 99%

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Rapid Detection, Identification, and Enumeration of Escherichia coli Cells in Municipal Water by Chemiluminescent In Situ Hybridization

Stender¹,

Broomer²,

Oliveira³

et al. 2001

Appl Environ Microbiol

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A new chemiluminescent in situ hybridization (CISH) method provides simultaneous detection, identification, and enumeration of culturable Escherichia coli cells in 100 ml of municipal water within one working day. Following filtration and 5 h of growth on tryptic soy agar at 35°C, individual microcolonies of E. coli were detected directly on a 47-mm-diameter membrane filter using soybean peroxidase-labeled peptide nucleic acid (PNA) probes targeting a species-specific sequence in E. coli 16S rRNA. Within each microcolony, hybridized, peroxidase-labeled PNA probe and chemiluminescent substrate generated light which was subsequently captured on film. Thus, each spot of light represented one microcolony of E. coli. Following probe selection based on 16S ribosomal DNA (rDNA) sequence alignments and sample matrix interference, the sensitivity and specificity of the probe Eco16S07C were determined by dot hybridization to RNA of eight bacterial species. Only the rRNA of E. coli and Pseudomonas aeruginosa were detected by Eco16S07C with the latter mismatch hybridization being eliminated by a PNA blocker probe targeting P. aeruginosa 16S rRNA. The sensitivity and specificity for the detection of E. coli by PNA CISH were then determined using 8 E. coli strains and 17 other bacterial species, including closely related species. No bacterial strains other than E. coli and Shigella spp. were detected, which is in accordance with 16S rDNA sequence information. Furthermore, the enumeration of microcolonies of E. coli represented by spots of light correlated 92 to 95% with visible colonies following overnight incubation. PNA CISH employs traditional membrane filtration and culturing techniques while providing the added sensitivity and specificity of PNA probes in order to yield faster and more definitive results.

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

Rapid Detection, Identification, and Enumeration of Escherichia coli Cells in Municipal Water by Chemiluminescent In Situ Hybridization

Stender¹,

Broomer²,

Oliveira³

et al. 2001

Appl Environ Microbiol

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“…Other primer sets designed for two different regions have been proposed for the detection of E. coli, one of them coding for an outer-membrane protein (phoE gene) (Spierings et al, 1993) and the other coding DNA sequences for the V3 and V6 regions of the 16S rRNA genes of pathogenicand non-pathogenic strains of E. coli (Tsen et al, 1998). These primer sets permit the specific detection of E. coli, but also of Shigella species when the suggested sequences are amplified.…”

Section: Molecular Methodsmentioning

confidence: 99%

Detection of Coliforms in Drinking Water and its Effect on Human Health - A Review

Pal¹

2014

ILNS

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The coliform group has been used extensively as an indicator of water quality and has historically led to the public health protection concept. Total coliforms are a group of bacteria commonly found in the environment, for example in soil or vegetation, as well as the intestines of mammals, including humans. Total coliform bacteria are not likely to cause illness, but their presence indicates that the water supply may be vulnerable to contamination by more harmful microorganisms. Escherichia coli (E. coli) is the only member of the total coliform group of bacteria that is found only in the intestines of mammals, including humans. The presence of E. coli in water indicates recent fecal contamination and may indicate the possible presence of disease-causing pathogens, such as bacteria, viruses, and parasites. Although most strains of E. coli bacteria are harmless, certain strains, such as E.coli 0157:H7, may cause illness. About 80 % of communicable diseases in the world are waterborne. According to WHO estimate about 80 % of water pollution in developing country, like India is carried by domestic waste. In India 70 % of the water is seriously polluted and 75 % of illness and 80 % of the child mortality is attributed to water pollution. The improper management of water systems may cause serious problems in availability and quality of water. The major pathogenic bacteria responsible for water borne disease are spread by the faeco-oral route, in which water may play an intermediate role. The aim of this review is to examine methods currently in use for the detection of coliforms in drinking water and also to evaluate the possible health hazards associated with drinking water contaminated with coliforms.

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“…Thus, with the advancement of science, molecular techniques for instance PCR are applied and appeared to be more sensitive, quicker, multiple targets can be detected at once and enabled the detection of fastidious, health-significant microorganisms that replicate poorly or not at all on standard microbiological growth media. (Bej et al [8]; Tsen et al [9]; Venkateswaran et al [10]; Yamamoto and Harayama [11]). This cuts down time, labor, and overall cost of the detection process (Mukhopadhyay and Mukhopadhyay [12]).…”

Section: Introductionmentioning

confidence: 99%

The inevitability of molecular methods for drinking water analysis

et al. 2012

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Drinking water regulations focus on the absence of indicator microorganisms in drinking water supplied to the end consumers to avoid the spread of potential diseases to members of the community. However, low counts of total heterotrophic bacteria are permissible. In general, total heterotrophic plate counts (HPC) may be used to represent the general hygiene of water where high HPC could indicate the presence of pathogenic bacteria, e.g. coliforms. Almost all of the standard methods for testing drinking water samples are culture-dependent methods that only detect microorganisms capable of utilizing standard microbiological culture media. However, the presence of viable but not culturable (unculturable) microorganisms (VBNC) makes it extremely important to employ techniques capable of detecting this fraction of water-borne microorganisms. Thus, the aim of the current study was to compare the outcome of applying traditional and molecular techniques for drinking water analysis. Results demonstrated the presence of VBNC bacteria such as Salmonella enterica, Corynebacterium glutamicum, Brachybacterium faecium, Clostridium phytofermentans, Enterobacter sp., Beutenbergia cavernae, Saccharomonospora viridis, Bacillus subtilis, Fervidobacterium nodosum in some water samples, which were not detected by traditional techniques. This suggested that, molecular techniques are inevitable and more reliable than traditional methods.

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Development and use of 16S rRNA gene targeted PCR primers for the identification of Escherichia coli cells in water

Cited by 98 publications

References 15 publications

Rapid Detection, Identification, and Enumeration of Escherichia coli Cells in Municipal Water by Chemiluminescent In Situ Hybridization

Rapid Detection, Identification, and Enumeration of Escherichia coli Cells in Municipal Water by Chemiluminescent In Situ Hybridization

Detection of Coliforms in Drinking Water and its Effect on Human Health - A Review

The inevitability of molecular methods for drinking water analysis

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