Polymicrobial sepsis is associated with worse patient outcomes than monomicrobial sepsis. Routinely used culture-dependent microbiological diagnostic techniques have low sensitivity, often leading to missed identification of all causative organisms. To overcome these limitations, culture-independent methods incorporating advanced molecular technologies have recently been explored. However, contamination, assay inhibition and interference from host DNA are issues that must be addressed before these methods can be relied on for routine clinical use. While the host component of the complex sepsis host–pathogen interplay is well described, less is known about the pathogen’s role, including pathogen–pathogen interactions in polymicrobial sepsis. This review highlights the clinical significance of polymicrobial sepsis and addresses how promising alternative molecular microbiology methods can be improved to detect polymicrobial infections. It also discusses how the application of shotgun metagenomics can be used to uncover pathogen/pathogen interactions in polymicrobial sepsis cases and their potential role in the clinical course of this condition.
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