The Design and Application of a Molecular Profiling Strategy to Identify Polymicrobial Acute Sepsis Infections
Abstract
Sepsis is a term used to describe an array of clinical presentations ranging from mild body dysfunction to multiple organ failure. These clinical signs result from a systemic inflammatory response to microbes or microbial products present in sterile sites such as blood. Current clinical diagnostics rely on culture techniques to identify systemic infections. However, culture lacks sensitivity and a positive result is only obtained in 40% of cases thereby limiting our knowledge of sepsis microbiology.
This doctoral study described the development of methods for direct detection of bacteria or bacterial products in blood. A method of lysing host cells and a bacterial DNA extraction protocol was developed and evaluated on mock bacterial communities spiked into whole blood. The results indicated that viable bacteria could be recovered down to 10 CFU/ml using this method. Paired-end Illumina sequencing of the 16S rRNA gene also indicated that the bacterial DNA extraction method enabled recovery of bacterial DNA from spiked blood.
This method demonstrated improved detection of systemic bacterial infections involving bacteria as well as their products in three cohorts of clinically septic patients. Application of the paired-end Illumina 16S rRNA sequencing to saponin treated blood from intensive care unit (ICU) and emergency department (ED) patients indicated there were bacterial DNA profiles present in whole blood. These patterns were examined alongside the patient’s clinical data and indicated common molecular profiling patterns were associated with the primary source of infection. Polymicrobial DNA was present in the blood samples with the taxonomic profiles suggesting commensal microbiota were implicated in addition to a principal pathogen. Bacterial DNA from Streptococcus and Staphylococcus were abundant in patients that died in the ICU.
Overall this study identified common bacterial DNA patterns in the blood of septic patients which were associated with the patients’ primary source of infection, implicated the commensal microbiota in systemic infection and suggested that sepsis infection may not always involve persistent bacterial bloodstream infections. Rather, this study concluded that bacterial products or viable organisms are likely cyclically present and cleared from the bloodstream resulting in a robust inflammatory response.
Description
Keywords
Microbiology
Citation
Faria Crowder, M. (2015). The Design and Application of a Molecular Profiling Strategy to Identify Polymicrobial Acute Sepsis Infections (Doctoral thesis, University of Calgary, Calgary, Canada). Retrieved from https://prism.ucalgary.ca. doi:10.11575/PRISM/28047