Replication Dynamics for Six Gram-Negative Bacterial Species during Bloodstream Infection

Anderson, Mark T.; Brown, Aric N; Pirani, Ali; Smith, Sara N.; Photenhauer, Amanda; Sun, Yuang; Snitkin, Evan S.; Bachman, Michael A.; Mobley, Harry L. T.

doi:10.1128/mbio.01114-21

Cited by 14 publications

(18 citation statements)

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“…The copyright holder for this preprint this version posted February 24, 2023. ; https://doi.org/10.1101/2023.02.23.529827 doi: bioRxiv preprint Even within the phase of bloodstream survival, different fitness factors are important in different sites of infection. Despite active replication in both organs, K. pneumoniae abundance increases in the liver and decreases in the spleen during bacteremia (19,20). Tissue-resident cells determine differential host responses across sites, and K. pneumoniae site-specific fitness has been minimally investigated.…”

Section: Discussionmentioning

confidence: 99%

“…Immune responses can also vary across organs as tissue-resident cells can have differential interactions with bacteria, including K. pneumoniae (19). Varying replication rates of multiple Gram-negative species across tissues during bacteremia highlights that host-pathogen interactions are not uniform between sites (20). Thus, defining bacterial site-specific fitness mechanisms can illuminate host defense strategies.…”

Section: Introductionmentioning

confidence: 99%

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Klebsiella pneumoniaecauses bacteremia using factors that mediate tissue-specific fitness and resistance to oxidative stress

Holmes

Wilcox

Forsyth

et al. 2023

Preprint

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Gram-negative bacteremia is a major cause of global morbidity involving three phases of pathogenesis: initial site infection, dissemination, and survival in the blood and filtering organs. Klebsiella pneumoniae is a leading cause of bacteremia and pneumonia is often the initial infection. In the lung, K. pneumoniae relies on many factors like capsular polysaccharide and branched chain amino acid biosynthesis for virulence and fitness. However, mechanisms directly enabling bloodstream fitness are unclear. Here, we performed transposon insertion sequencing (TnSeq) in a tail-vein injection model of bacteremia and identified 58 K. pneumoniae bloodstream fitness genes. These factors are diverse and represent a variety of cellular processes. In vivo validation revealed tissue-specific mechanisms by which distinct factors support bacteremia. ArnD, involved in Lipid A modification, was required across blood filtering organs and supported resistance to soluble splenic factors. The purine biosynthesis enzyme PurD largely enhanced liver fitness and was required for replication in serum. PdxA, a member of the endogenous vitamin B6 biosynthesis pathway, optimized replication in serum and lung fitness. The stringent response regulator SspA was required for splenic fitness yet was dispensable in the liver. In a bacteremic pneumonia model that incorporates initial site infection and dissemination, splenic fitness defects were enhanced, and DsbA, SspA, and PdxA increased fitness across bacteremia phases. SspA and PdxA enhanced K. pnuemoniae resistance to oxidative stress. SspA specifically resists oxidative stress produced by NADPH oxidase Nox2 in the lung, spleen, and liver, as it was a fitness factor in wild-type but not Nox2-deficient (Cybb-/-) mice. These results identify site-specific fitness factors that act during the progression of Gram-negative bacteremia. Defining K. pneumoniae fitness strategies across bacteremia phases could illuminate therapeutic targets that prevent infection and sepsis.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Klebsiella pneumoniaecauses bacteremia using factors that mediate tissue-specific fitness and resistance to oxidative stress

Holmes

Wilcox

Forsyth

et al. 2023

Preprint

Self Cite

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“…GmhB may be crucial under conditions where rapid LPS production is necessary. During murine bacteremia, K. pneumoniae exhibits exponential replication in the spleen at 24 h ( 50 ). Rapid replication requires substantial LPS export and, in the absence of GmhB, lower abundance of normal LPS may be produced.…”

Section: Discussionmentioning

confidence: 99%

The ADP-Heptose Biosynthesis Enzyme GmhB is a Conserved Gram-Negative Bacteremia Fitness Factor

et al. 2022

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“…GmhB may be crucial under conditions where rapid LPS production is necessary. During murine bacteremia, K. pneumoniae exhibits exponential replication in the spleen at 24 hours (51). Rapid replication requires substantial LPS export and, in the absence of GmhB, lower abundance of normal LPS may be produced.…”

Section: Discussionmentioning

confidence: 99%

The ADP-heptose biosynthesis enzyme GmhB is a conserved Gram-negative bacteremia fitness factor

Holmes

Smith

Gurczynski

et al. 2022

Preprint

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Klebsiella pneumoniae is a leading cause of Gram-negative bacteremia, which is a major source of morbidity and mortality worldwide. Gram-negative bacteremia requires three major steps: primary site infection, dissemination to the blood, and bloodstream survival. Since K. pneumoniae is a leading cause of healthcare-associated pneumonia, the lung is a common primary infection site leading to secondary bacteremia. K. pneumoniae factors essential for lung fitness have been characterized, but those required for subsequent bloodstream infection are unclear. To identify K. pneumoniae genes associated with dissemination and bloodstream survival, we performed insertion site sequencing (InSeq) using a pool of >25,000 transposon mutants in a murine model of bacteremic pneumonia. This analysis revealed the gene gmhB as important for either dissemination from the lung or bloodstream survival. In Escherichia coli, GmhB is a partially redundant enzyme in the synthesis of ADP-heptose for the lipopolysaccharide (LPS) core. To characterize its function in K. pneumoniae, an isogenic knockout strain (ΔgmhB) and complemented mutant were generated. During pneumonia, GmhB did not contribute to lung fitness and did not alter normal immune responses. However, GmhB enhanced bloodstream survival in a manner independent of serum susceptibility, specifically conveying resistance to spleen-mediated killing. In a tail-vein injection of murine bacteremia, GmhB was also required by K. pneumoniae, E. coli and Citrobacter freundii for optimal bloodstream survival. Together, this study identifies GmhB as a conserved Gram-negative bacteremia fitness factor that acts through LPS-mediated mechanisms to enhance bloodstream survival.IMPORTANCEKlebsiella pneumoniae frequently causes healthcare-associated infections including pneumonia and bacteremia. This is particularly concerning due to emerging antimicrobial resistance and the propensity for bacteremia to initiate sepsis, which has high mortality and is the most expensive hospital-treated condition. Defining mechanisms of bloodstream survival is critical to understanding the pathology of bacteremia and identifying novel targets for future therapies. In this study, we identified the K. pneumoniae enzyme GmhB as a bloodstream-specific fitness factor that enables the bacteria to survive in the spleen but is dispensable in the lung. Furthermore, GmhB is also needed by the related bacterial pathogens Escherichia coli and Citrobacter freundii to cause bacteremia. Conserved bacteremia fitness factors such a GmhB could be the basis for future therapeutics that would alleviate significant disease caused by from multiple diverse pathogens.

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Replication Dynamics for Six Gram-Negative Bacterial Species during Bloodstream Infection

Abstract: Bloodstream infections are a global public health problem. The goal of this work was to determine the replication characteristics of Gram-negative bacterial species in the host following bloodstream infection.

Cited by 14 publications

References 50 publications

Klebsiella pneumoniaecauses bacteremia using factors that mediate tissue-specific fitness and resistance to oxidative stress

Klebsiella pneumoniaecauses bacteremia using factors that mediate tissue-specific fitness and resistance to oxidative stress

The ADP-Heptose Biosynthesis Enzyme GmhB is a Conserved Gram-Negative Bacteremia Fitness Factor

The ADP-heptose biosynthesis enzyme GmhB is a conserved Gram-negative bacteremia fitness factor

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