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Klebsiella pneumoniae is a common member of the intestinal flora of vertebrates. In addition to opportunistic representatives, hypervirulent (hvKp) and antibiotic-resistant K. pneumoniae (ABR-Kp) occur. While ABR-Kp isolates often cause difficult-to-treat diseases due to limited therapeutic options, hvKp is a pathotype that can infect healthy individuals often leading to recurrent infection. Here, we investigated the clinical K. pneumoniae isolate PBIO3459 obtained from a blood sample, which showed an unusual colony morphology. By combining whole-genome and RNA sequencing with multiple in vitro and in vivo virulence-associated assays, we aimed to define the respective Klebsiella subtype and explore the unusual phenotypic appearance. We demonstrate that PBIO3459 belongs to sequence type (ST)20 and carries no acquired resistance genes, consistent with phenotypic susceptibility tests. In addition, the isolate showed low-level virulence, both at genetic and phenotypic levels. We thus suggest that PBIO3459 is an opportunistic (commensal) K. pneumoniae isolate. Genomic comparison of PBIO3459 with closely related ABR-Kp ST20 isolates revealed that they differed only in resistance genes. Finally, the unusual colony morphology was mainly associated with carbohydrate and amino acid transport and metabolism. In conclusion, our study reveals the characteristics of a Klebsiella sepsis isolate and suggests that opportunistic representatives likely acquire and accumulate antibiotic resistances that subsequently enable their emergence as ABR-Kp pathogens.
Abstract
DNA extraction and preservation bias is a recurring topic in DNA sequencing‐based microbial ecology. The different methodologies can lead to distinct outcomes, which has been demonstrated especially in studies investigating prokaryotic community composition. Eukaryotic microbes are ubiquitous, diverse, and increasingly a subject of investigation in addition to bacteria and archaea. However, little is known about how the choice of DNA preservation and extraction methodology impacts perceived eukaryotic community composition. In this study, we compared the effect of two DNA preservation methods and six DNA extraction methods on the community profiles of both eukaryotes and prokaryotes in phototrophic biofilms on seagrass (Zostera marina) leaves from the Baltic Sea. We found that, whereas both DNA preservation and extraction method caused significant bias in perceived community composition for both eukaryotes and prokaryotes, extraction bias was more pronounced for eukaryotes than for prokaryotes. In particular, soft‐bodied and hard‐shelled eukaryotes like nematodes and diatoms, respectively, were differentially abundant depending on the extraction method. We conclude that careful consideration of DNA preservation and extraction methodology is crucial to achieving representative community profiles of eukaryotes in marine biofilms and likely all other habitats containing diverse eukaryotic microbial communities.