Comparative evaluation of DNA extraction methods for amplification by qPCR of superficial vs intracellular DNA from Bacillus spores

Brauge, Thomas; Faille, Christine; Inglebert, Gaelle; Dubois, Thomas; Morieux, Paul; Slomianny, Christian; Midelet, Graziella

doi:10.1016/j.ijfoodmicro.2017.12.012

Cited by 14 publications

(12 citation statements)

References 33 publications

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“…Our approach is based on the ability of C. botulinum GIII to survive the InstaGene extraction protocol, which seems to eliminate most of the other bacterial contaminants that usually disturb the isolation process. The InstaGene matrix has previously been shown to fail to extract intracellular DNA from Bacillus cereus spores [36]. Moreover, the same study demonstrated that the majority of B. cereus spores appeared little or unaffected by the InstaGene matrix extraction protocol.…”

Section: Discussionmentioning

confidence: 83%

Development of An Innovative and Quick Method for the Isolation of Clostridium botulinum Strains Involved in Avian Botulism Outbreaks

Gratiet

Poëzévara

Rouxel

et al. 2020

Toxins

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Avian botulism is a serious neuroparalytic disease mainly caused by a type C/D botulinum neurotoxin produced by Clostridium botulinum group III, one of the entwined bacterial species from the Clostridium novyi sensu lato genospecies. Its isolation is very challenging due to the absence of selective media and the instability of the phage carrying the gene encoding for the neurotoxin. The present study describes the development of an original method for isolating C. botulinum group III strains. Briefly, this method consists of streaking the InstaGene matrix extraction pellet on Egg Yolk Agar plates and then collecting the colonies with lipase and lecithinase activities. Using this approach, it was possible to isolate 21 C. novyi sensu lato strains from 22 enrichment broths of avian livers, including 14 toxic strains. This method was successfully used to re-isolate type C, D, C/D, and D/C strains from liver samples spiked with five spores per gram. This method is cheap, user-friendly, and reliable. It can be used to quickly isolate toxic strains involved in avian botulism with a 64% success rate and C. novyi sensu lato with a 95% rate. This opens up new perspectives for C. botulinum genomic research, which will shed light on the epidemiology of avian botulism.

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Section: Discussionmentioning

confidence: 83%

Development of An Innovative and Quick Method for the Isolation of Clostridium botulinum Strains Involved in Avian Botulism Outbreaks

Gratiet

Poëzévara

Rouxel

et al. 2020

Toxins

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“…For example, in the anthrax mailing attack of 2001 in the US, Bacillus anthracis was cultured from the victim's cerebral spinal fluid and blood, and subsequently genotyped to identify the source of the microorganism (Keim, Budowle, & Ravel, 2011). Chemical‐, mechanical‐, or heat‐based DNA extraction methods are generally effective for bacteria and viruses (Alessandrini et al, 2019; Brauge et al, 2018; Cho et al, 2014; Hennechart‐Collette, Niveau, Martin‐Latil, Fraisse, & Perelle, 2019; Liu et al, 2012; Thomas et al, 2013), with the exception of bacteria endospores.…”

Section: Dna Extraction From Human Remains and Non‐conventional Samplesmentioning

confidence: 99%

“…Bacteria endospores are encased in a tough protein coat, rendering some DNA extraction methods ineffective (Brauge et al, 2018; Rose et al, 2011; Thomas et al, 2013; Timbers et al, 2014). However, recent studies based on mechanical lysis using a French press and homogenizer (Brauge et al, 2018), as well as a novel chemical lysis method (sporeLYSE) have demonstrated effective DNA extraction from endospores (de Bruin, Chiefari, Wroblewski, Egan, & Kelly‐Cirino, 2019). Nevertheless, it remains to be seen how mechanical lysis can be used efficiently in forensic DNA extraction, since it is a cumbersome process.…”

Section: Dna Extraction From Human Remains and Non‐conventional Samplesmentioning

confidence: 99%

Recent trends and developments in forensic DNA extraction

Chong

Thong

Syn

2020

WIREs Forensic Science

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The extraction of DNA is a fairly standard procedure routinely undertaken in biological academic and research settings. In a forensic DNA context, DNA extraction is the crucial first step toward the generation of DNA profiles used to support convictions or exonerations in the criminal justice system. However, it is often complicated by the additional challenge of yielding sufficient quantities of clean DNA that is free from environmental inhibitors, from a wide spectrum of sample types which may contain limited quantities of biological material. While improvements in methods and technology over the decades have sped up and simplified the process of DNA extraction, several issues remain to be fully addressed—such as the incomplete separation of spermic DNA from sperm‐epithelial cell mixtures, increasing demands for ever‐faster DNA results, or even on‐scene DNA processing, and processing of difficult samples such as skeletal remains or samples contaminated by Chemical, Biological, or Radiological (CBR) agents. These challenges are often compounded by high work volumes and limited quantities of starting material. As such, this review focuses on recent trends and developments in four areas of forensic DNA extraction: (a) analysis of sexual assault evidence which often represents a significant portion of a forensic DNA laboratory's casework, (b) use of portable rapid DNA instruments which has gained much traction among law enforcement in recent times, (c) DNA extraction from difficult and CBR‐contaminated samples which, albeit rare, would be essential in a DNA laboratory's toolbox in the event of an incident, and (d) bypassing DNA extraction altogether. This article is categorized under: Forensic Biology > Forensic DNA Technologies

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“…Alongside, we also tested the QIAamp DNA Mini kit, as it is commonly used for DNA extraction in laboratories [32]. Of note, there is residual DNA of the spore mother cell remaining on the spore surface, which, at least in part, is accessible for PCR [33][34][35]. Nevertheless, retrieved DNA concentrations were not sufficient for further downstream applications, such as nanopore sequencing, since library preparation requires a minimum of 0.5-1 µg of DNA.…”

Section: Dna Extraction Methods Impacts the Quality Of Sequencing Resultsmentioning

confidence: 99%

Evaluation of a Highly Efficient DNA Extraction Method for Bacillus anthracis Endospores

Knüpfer¹,

Braun²,

Baumann³

et al. 2020

Microorganisms

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A variety of methods have been established in order to optimize the accessibility of DNA originating from Bacillus anthracis cells and endospores to facilitate highly sensitive molecular diagnostics. However, most endospore lysis techniques have not been evaluated in respect to their quantitative proficiencies. Here, we started by systematically assessing the efficiencies of 20 DNA extraction kits for vegetative B. anthracis cells. Of these, the Epicentre MasterPure kit gave the best DNA yields and quality suitable for further genomic analysis. Yet, none of the kits tested were able to extract reasonable quantities of DNA from cores of the endospores. Thus, we developed a mechanical endospore lysis protocol, facilitating the extraction of high-quality DNA. Transmission electron microscopy or the labelling of spores with the indicator dye propidium monoazide was utilized to assess lysis efficiency. Finally, the yield and quality of genomic spore DNA were quantified by PCR and they were found to be dependent on lysis matrix composition, instrumental parameters, and the method used for subsequent DNA purification. Our final standardized lysis and DNA extraction protocol allows for the quantitative detection of low levels (<50 CFU/mL) of B. anthracis endospores and it is suitable for direct quantification, even under resource-limited field conditions, where culturing is not an option.

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Comparative evaluation of DNA extraction methods for amplification by qPCR of superficial vs intracellular DNA from Bacillus spores

Cited by 14 publications

References 33 publications

Development of An Innovative and Quick Method for the Isolation of Clostridium botulinum Strains Involved in Avian Botulism Outbreaks

Development of An Innovative and Quick Method for the Isolation of Clostridium botulinum Strains Involved in Avian Botulism Outbreaks

Recent trends and developments in forensic DNA extraction

Evaluation of a Highly Efficient DNA Extraction Method for Bacillus anthracis Endospores

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