Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2021 Dec;12(1):415-429.
doi: 10.1080/21505594.2020.1870336.

The lipoprotein NlpD in Cronobacter sakazakii responds to acid stress and regulates macrophage resistance and virulence by maintaining membrane integrity

Xuemeng Ji  1 , Ping Lu  2 , Juan Xue  3 , Ning Zhao  1 , Yan Zhang  1 , Lu Dong  1 , Xuejiao Zhang  1 , Ping Li  4 , Yaozhong Hu  1 , Jin Wang  1 , Bowei Zhang  1 , Jingmin Liu  1 , Huan Lv  1 , Shuo Wang  1
Affiliations

The lipoprotein NlpD in Cronobacter sakazakii responds to acid stress and regulates macrophage resistance and virulence by maintaining membrane integrity

Xuemeng Ji et al. Virulence. 2021 Dec.

Abstract

Cronobacter sakazakii, an emerging opportunistic pathogen, is implicated in severe foodborne outbreak infections in premature and full-term infants. Generally, acid tolerance is vital for the pathogenesis of foodborne pathogens; however, its role in C. sakazakii virulence remains largely unknown. To screen out acid-tolerance determinants from transposon mutants, anovel counterselection method using gentamicin and acid was developed. Using the counterselection method and growth assay, we screened several acid-sensitive mutants and found that nlpD encodes an acid-resistance factor in C. sakazakii. Compared to the wild-type strain, the nlpD mutant exhibited attenuated virulence in a rat model. Using macrophage THP-1 cells and a pH probe, we verified that nlpD enables bacteria to resist macrophages by resisting acidification. Finally, we confirmed that nlpD maintains C. sakazakii membrane integrity in acid using propidium iodide permeabilization assays via flow cytometry. Our results confirm that nlpD is a novel virulence factor that permits C. sakazakii to survive under acid stress conditions. Considering that NlpD is a conserved lipoprotein located in the bacterial outer membrane, NlpD could be used as a target for drug development.

Keywords: Cronobacter sakazakii; acid resistance; lipoprotein; membrane integrity.

PubMed Disclaimer

Conflict of interest statement

The authors have no competing interests to disclose.

Figures

Figure 1.
Figure 1.
Phenotype verification of five acid determinants using markerless deletion mutants
Figure 2.
Figure 2.
The acid determinant nlpD is involved in the pathogenicity of Cronobacter sakazakii.
Figure 3.
Figure 3.
nlpD is not involved in adherence
Figure 4.
Figure 4.
The ΔnlpD mutant shows attenuated survival within activated macrophages
Figure 5.
Figure 5.
nlpD deletion attenuates bacterial survival within IFN-γ-activated macrophages
Figure 6.
Figure 6.
Reduced nlpD expression in the ΔevnZ mutant
Figure 7.
Figure 7.
Membrane permeabilization of C. sakazakii by PI uptake assay
See this image and copyright information in PMC

References

    1. Elkhawaga AA, Hetta HF, Osman NS, et al. Emergence of Cronobacter sakazakii in cases of neonatal sepsis in Upper Egypt first report in North Africa. BMC Microbiol. 2020;11:215. .Epub 2019/12/29. 10.3389/fmicb.2020.00215. PubMed PMID: 32210926; PubMed Central PMCID: PMCPmc6935241 Pmc7075355. - PMC - PubMed
    1. Guo D, Wang S, Li J, et al. The antimicrobial activity of coenzyme Q(0) against planktonic and biofilm forms of Cronobacter sakazakii. Food Microbiol. 2020;86:103337. .Epub 2019/11/11. PubMed PMID: 31703870. - PubMed
    1. Chen Z, Zhang Y, Lin R, et al. Cronobacter sakazakii induces necrotizing enterocolitis by regulating NLRP3 inflammasome expression via TLR4. J Med Microbiol. 2020;69(5):748–758. .Epub 2020/03/27. PubMed PMID: 32209170. - PubMed
    1. Control CfD, Prevention . Enterobacter sakazakii infections associated with the use of powdered infant formula–tennessee, 2001. MMWR Morb Mortal Wkly Rep. 2002;51(14):297. - PubMed
    1. Wang M, Wang L, Wu P, et al. Genomics and experimental analysis reveal a novel factor contributing to the virulence of cronobacter sakazakii strains associated with neonate infection. J Infect Dis. 2019;220(2):306–315. .Epub 2019/03/06. PubMed PMID: 30835279. - PubMed

Publication types

MeSH terms