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Metagenomic sequencing of fecal DNA can usefully characterise an individual's intestinal resistome but is limited by its inability to detect important pathogens that may be present at low abundance, such as carbapenemase or extended-spectrum beta-lactamase producing Enterobacteriaceae. Here we aimed to develop a hybrid protocol to improve detection of resistance genes in Enterobacteriaceae by using a short period of culture enrichment prior to sequencing of DNA extracted directly from the enriched sample. Volunteer feces were spiked with carbapenemase-producing Enterobacteriaceae and incubated in selective broth culture for 6 hours before sequencing. Different DNA extraction methods were compared, including a plasmid extraction protocol to increase the detection of plasmid-associated resistance genes. Although enrichment prior to sequencing increased the detection of carbapenemase genes, the differing growth characteristics of the spike organisms precluded accurate quantification of their concentration prior to culture. Plasmid extraction increased detection of resistance genes present on plasmids, but the effects were heterogeneous and dependent on plasmid size. Our results demonstrate methods of improving the limit of detection of selected resistance mechanisms in a fecal resistome assay, but they also highlight the difficulties in using these techniques for accurate quantification and should inform future efforts to achieve this goal.
This article was published in the following journal.
Name: PloS one
Subclinical colonization in piglets could be a potential source of this bacterium for community-acquired infection. The purposes of this study were to assess the effect of specimen type and processi...
This study compared the impact of different approaches, namely, nonenrichment, nonselective enrichment, and selective (antibiotic-containing) enrichment steps, for detecting extended-spectrum β-lacta...
Routine sequencing of MRSA could bring about significant improvements to outbreak detection and investigation. Sequencing is commonly performed using DNA extracted from a pure culture, but overcoming ...
We characterized clinical isolates of Campylobacter using whole-genome sequencing (WGS) for detection of virulence genes, antimicrobial resistance markers and phylogenetic analysis in order to increas...
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Sepsis remains a major challenge, even in modern intensive care medicine. The identification of the causative pathogen is crucial for an early optimization of the antimicrobial treatment r...
Patients with negative blood cultures represent 85-90 % from all patients with a blood culture taken during hospital admissions. This population usually includes an heterogeneous group of ...
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Pathogen identification is of paramount importance for bacterial meningitis. At present, the pathogen of bacterial meningitis is still mainly based on Gram stain and bacterial culture. How...
Currently, for a patient with intellectual disability without a recognizable syndrome (most cases), the way to diagnosis is often long, tedious and expensive because different approaches a...
An industry that creates products and procedures designed to enhance physical appearance and aesthetic appeal.
Detection of drugs that have been abused, overused, or misused, including legal and illegal drugs. Urine screening is the usual method of detection.
Techniques of nucleotide sequence analysis that increase the range, complexity, sensitivity, and accuracy of results by greatly increasing the scale of operations and thus the number of nucleotides, and the number of copies of each nucleotide sequenced. The sequencing may be done by analysis of the synthesis or ligation products, hybridization to preexisting sequences, etc.
A phenethylamine found in EPHEDRA SINICA. PSEUDOEPHEDRINE is an isomer. It is an alpha- and beta-adrenergic agonist that may also enhance release of norepinephrine. It has been used for asthma, heart failure, rhinitis, and urinary incontinence, and for its central nervous system stimulatory effects in the treatment of narcolepsy and depression. It has become less extensively used with the advent of more selective agonists.
Culture of an isolated organism free from any other associating or contaminating organisms.
DNA sequencing is the process of determining the precise order of nucleotides within a DNA molecule. During DNA sequencing, the bases of a small fragment of DNA are sequentially identified from signals emitted as each fragment is re-synthesized from a ...
Bioinformatics is the application of computer software and hardware to the management of biological data to create useful information. Computers are used to gather, store, analyze and integrate biological and genetic information which can then be applied...
DNA is extracted from human cells for a variety of reasons. With a pure sample of DNA you can test a newborn for a genetic disease, analyze forensic evidence, or study a gene involved in cancer. DNA isolation is a process of purification of DNA from...