DNA extraction from bovine faeces: current status and future trends

Rapp, Delphine

doi:10.1111/j.1365-2672.2009.04606.x

Cited by 23 publications

(20 citation statements)

References 59 publications

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“…Addressing this knowledge gap now appears feasible, as real-time quantitative PCR (qPCR) can be used as an alternative to culture-based methods for quantifying environmental pathogens (7,23,29). Improvements in sample preparation and nucleic acid cleanup methods have largely overcome problems associated with the molecular biology-based analysis of fecal matter (22). Further, qPCR can detect stressed, damaged, and otherwise nonculturable cells persisting in a state of dormancy or indeed dead (15,17,29).…”

mentioning

confidence: 99%

Diversity and Abundance of Zoonotic Pathogens and Indicators in Manures of Feedlot Cattle in Australia

Klein

Brown

Tucker

et al. 2010

Appl Environ Microbiol

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mentioning

confidence: 99%

Diversity and Abundance of Zoonotic Pathogens and Indicators in Manures of Feedlot Cattle in Australia

Klein

Brown

Tucker

et al. 2010

Appl Environ Microbiol

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“…A number of studies have highlighted the need for careful consideration of DNA extraction methods from different animal species (Goldberg et al, 2016;Hart et al, 2015;Rapp, 2010). For example, DNA was successfully extracted from fish faecal samples using the isopropanol method, while the MoBio Power Fecal DNA kit (Qiagen Pty Ltd, Chadstone, Victoria, Australia) was needed to extract quality DNA from equine faecal samples (Hart et al, 2015).…”

Section: (B) (A)mentioning

confidence: 99%

“…However, the complex matrix of faecal samples make it a challenging job to choose the most suitable extraction protocol as some common methods, such as the cell lysis by boiling method, are incapable of removing faecal inhibitors (Rapp, 2010;Wilson, 1997).…”

Section: (B) (A)mentioning

confidence: 99%

“…There are several methods (physical, mechanical and chemical), as well as specifically designed commercial kits (McOrist et al, 2002;Yu and Morrison, 2004) used to extract DNA from faeces. However, the complex matrix of faecal samples make it a challenging job to choose the most suitable extraction protocol as some methods, such as the cell lysis by boiling method, are incapable of removing faecal inhibitors (Rapp, 2010;Wilson, 1997).…”

Section: Introductionmentioning

confidence: 99%

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Studies on avian pathogenic Escherichia coli in commercial broiler chicken in South East Queensland

Awawdeh¹,

Turni²,

Henning³

et al.

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Avian pathogenic Escherichia coli (APEC) is the causative agent of avian colibacillosis, a localised or systemic infection resulting in clinical diseases such as colisepticemia, chronic respiratory disease and swollen-head syndrome. Globally, avian colibacillosis is the leading cause of morbidity and mortality in poultry, and it has been associated with massive economic losses and welfare problems.This organism is of public health significance as APEC is communicable to humans. The diagnosis of avian colibacillosis relies on clinical signs, typical pathological lesions and culture of E. coli from affected tissue(s). Antimicrobial therapy is often used both for treatment and control. Previous overseas studies have characterised APEC and identified virulence genes (VGs) that can be used as molecular markers for the identification of APEC. Little is known about APEC in broiler chickens in Australia.The aim of this thesis was to gain a better understanding of the epidemiology of APEC in Australian broiler flocks and how factors including presence of VGs and phylogenetic group can improve the identification of this pathotype in Australia. Firstly, three faecal DNA extraction methods were evaluated. Faeces were collected from healthy chickens and chickens with colibacillosis from commercial broiler farms in South East Queensland (SEQ). The extracted DNA was screened by a pentaplex-PCR for five APEC-associated VGs (iroN, iutA, iss, hlyF and ompT). DNA extracted from E. coli isolates cultured from the cloaca and organs of the birds were screened using the same PCR.Repeated bead beating plus column elution was the preferred DNA extraction method, as it yielded good PCR quality and adequate quantity DNA. However, identifying APEC by direct detection of the five VGs from the faecal material was not feasible as all of these genes were also detected in all of the birds. However, the VGs were more commonly detected in E. coli isolates cultured from birds with colibacillosis.A cross-sectional study was performed to estimate APEC farm-level prevalence in healthy broiler chickens in SEQ and to identify potential risk factors associated with the carriage of APEC. At the farm-level, all of the 40 farms sampled were positive for APEC, that is at least one bird per farm carried APEC, while the within-farm prevalence was 63% (95% Confidence Interval: 55.8, 70.2).Higher APEC within-farm bird-level prevalence was significantly associated with the usage of well water as a source of drinking water, failure to disinfect the water line after each flock, farm visitors not showering before entering the shed, distances greater than 20 metres between the car park and the poultry shed and the presence of wild birds within 50 metres of the shed. Chlorinating the drinking water combined with automatic water filtration reduced within-farm bird-level APEC prevalence. Page | 3Therefore, based on the results concluded from the multivariable model, improving biosecurity and water treatments might reduce APEC prevalence, decrease the risk of co...

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“…However, real-time PCR is not commonly used for the direct detection of foodborne pathogenic bacteria in feces because fecal components can inhibit real-time PCR (4). To obtain useable PCR templates from crude human feces, column purification methods that require numerous time-consuming technical steps are often used to remove inhibitors (5). We wanted to develop a very rapid, simple method within 5 steps for the extraction and purification of bacterial DNA from human feces, because actual outbreaks of foodborne illnesses require rapid pathogen detection.…”

Section: Introductionmentioning

confidence: 99%

A Rapid and Simple Real-Time PCR Assay for Detecting Foodborne Pathogenic Bacteria in Human Feces

Hanabara

Ueda

2016

Jpn J Infect Dis

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SUMMARY: A rapid, simple method for detecting foodborne pathogenic bacteria in human feces is greatly needed. Here, we examined the efficacy of a method that employs a combination of a commercial PCR master mix, which is insensitive to PCR inhibitors, and a DNA extraction method which used sodium dodecyl benzene sulfonate (SDBS), and Tween 20 to counteract the inhibitory effects of SDBS on the PCR assay. This method could detect the target genes (stx1 and stx2 of enterohemorrhagic Escherichia coli, invA of Salmonella Enteritidis, tdh of Vibrio parahaemolyticus, gyrA of Campylobacter jejuni, ceuE of Campylobacter coli, SEA of Staphylococcus aureus, ces of Bacillus cereus, and cpe of Clostridium perfringens) in a fecal suspension containing 1.0 × 10 1 to 1.0 × 10 3 CFU/ml. Furthermore, the assay was neither inhibited nor influenced by individual differences among the fecal samples of 10 subjects or fecal concentration (40-160 mg/ml in the fecal suspension). When we attempted to detect the genes of pathogenic bacteria in 4 actual clinical cases, we found that this method was more sensitive than standard culture method. These results showed that this assay is a rapid, simple detection method for foodborne pathogenic bacteria in human feces.

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DNA extraction from bovine faeces: current status and future trends

Cited by 23 publications

References 59 publications

Diversity and Abundance of Zoonotic Pathogens and Indicators in Manures of Feedlot Cattle in Australia

Diversity and Abundance of Zoonotic Pathogens and Indicators in Manures of Feedlot Cattle in Australia

Studies on avian pathogenic Escherichia coli in commercial broiler chicken in South East Queensland

A Rapid and Simple Real-Time PCR Assay for Detecting Foodborne Pathogenic Bacteria in Human Feces

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