Deletion of Braun Lipoprotein and Plasminogen-Activating Protease-Encoding Genes Attenuates Yersinia pestis in Mouse Models of Bubonic and Pneumonic Plague

Lier, Christina J. van; Sha, Jian; Kirtley, Michelle L.; Cao, Anthony; Tiner, Bethany L.; Erova, Tatiana E.; Cong, Yingzi; Kozlova, Elena; Popov, Vsevolod L.; Baze, Wallace B.; Chopra, Ashok K.

doi:10.1128/iai.01595-13

Cited by 22 publications

(66 citation statements)

References 52 publications

(118 reference statements)

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“…The Pla (plasminogen activator) surface protease of Y. pestis is a multifunctional protein that contributes to bacterial adherence to host cells, intracellular survival, complement resistance, and bacterial dissemination by virtue of possessing fibrinolytic and coagulase activities (48,(57)(58)(59)(60)(61)(62)(63). To examine whether the deletion of three membrane protein-encoding genes (lpp, msbB, and ail) from WT CO92 altered Pla levels, we performed Western blot analysis.…”

Section: Methodsmentioning

confidence: 99%

“…Consequently, WT CO92 and ⌬ail single, ⌬lpp ⌬msbB double, and ⌬lpp ⌬msbB ⌬ail triple mutant cultures grown overnight were diluted 1:20 and grown in either HIB or calcium-depleted modified M9 medium (42 mM Na 2 HPO 4 , 22 mM KH 2 PO 4 , 8.6 mM NaCl, 18.6 mM NH 4 Cl, 0.001 mg/ml FeSO 4 , 0.0001% thiamine, 1 mM MgSO 4 , 0.4% dextrose, and 1% Casamino Acids) at 28°C with shaking (180 rpm) for 3 h and then at 37°C for 2 h. When the bacteria were grown in HIB, 5 mM EGTA (Sigma-Aldrich, St. Louis, MO) was added to trigger the low-calcium response 5 min before harvesting of the cultures. Aliquots of the cultures grown in either medium (representing similar CFU) were removed, and 1 ml of the supernatants was precipitated with 55 l of 100% trichloroacetic acid (TCA) on ice for 2 h. The TCA precipitates were dissolved in SDS-PAGE buffer and analyzed by immunoblotting with antibodies to YopE, LcrV (Santa Cruz Biotechnology, Santa Cruz, CA), and YopH (Agrisera, Stockholm, Sweden) (49,55,57). The anti-DnaK monoclonal antibody (Enzo) was employed to probe bacterial pellets to ensure that the bacterial supernatants were obtained from similar numbers of bacteria across the tested strains.…”

Section: Methodsmentioning

confidence: 99%

“…For visualization of membrane alterations, bacterial strains were grown to exponential phase at 28°C (optical density at 600 nm [OD 600 ] of 0.6). The cells were washed, pelleted, fixed, and subjected to transmission electron microscopy (57).…”

Section: Methodsmentioning

confidence: 99%

“…The samples were incubated at 37°C for 2 h with shaking at 500 rpm. The number of surviving bacteria (CFU) in each sample was determined by serial dilutions and plating onto SBA plates (49,57). Percent bacterial survival was calculated by dividing the average number of CFU in samples incubated in normal serum by the average number of CFU in samples incubated in heat-inactivated serum and multiplying by 100.…”

Section: Vt)mentioning

confidence: 99%

“…Six-to eightweek-old female Swiss Webster mice (17 to 20 g) were purchased from Taconic Laboratories (Germantown, NY). All of the animal studies were [57]) of WT CO92 or the ⌬ail single, ⌬lpp ⌬msbB or ⌬lpp ⌬ail double, or ⌬lpp ⌬msbB ⌬ail triple mutant strain. Also, the animals were challenged by the i.n.…”

Section: Vt)mentioning

confidence: 99%

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Combinational Deletion of Three Membrane Protein-Encoding Genes Highly Attenuates Yersinia pestis while Retaining Immunogenicity in a Mouse Model of Pneumonic Plague

et al. 2015

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f Previously, we showed that deletion of genes encoding Braun lipoprotein (Lpp) and MsbB attenuated Yersinia pestis CO92 in mouse and rat models of bubonic and pneumonic plague. While Lpp activates Toll-like receptor 2, the MsbB acyltransferase modifies lipopolysaccharide. Here, we deleted the ail gene (encoding the attachment-invasion locus) from wild-type (WT) strain CO92 or its lpp single and ⌬lpp ⌬msbB double mutants. While the ⌬ail single mutant was minimally attenuated compared to the WT bacterium in a mouse model of pneumonic plague, the ⌬lpp ⌬ail double mutant and the ⌬lpp ⌬msbB ⌬ail triple mutant were increasingly attenuated, with the latter being unable to kill mice at a 50% lethal dose (LD 50 ) equivalent to 6,800 LD 50 s of WT CO92. The mutant-infected animals developed balanced T H 1-and T H 2-based immune responses based on antibody isotyping. The triple mutant was cleared from mouse organs rapidly, with concurrent decreases in the production of various cytokines and histopathological lesions. When surviving animals infected with increasing doses of the triple mutant were subsequently challenged on day 24 with the bioluminescent WT CO92 strain (20 to 28 LD 50 s), 40 to 70% of the mice survived, with efficient clearing of the invading pathogen, as visualized in real time by in vivo imaging. The rapid clearance of the triple mutant, compared to that of WT CO92, from animals was related to the decreased adherence and invasion of human-derived HeLa and A549 alveolar epithelial cells and to its inability to survive intracellularly in these cells as well as in MH-S murine alveolar and primary human macrophages. An early burst of cytokine production in macrophages elicited by the triple mutant compared to WT CO92 and the mutant's sensitivity to the bactericidal effect of human serum would further augment bacterial clearance. Together, deletion of the ail gene from the ⌬lpp ⌬msbB double mutant severely attenuated Y. pestis CO92 to evoke pneumonic plague in a mouse model while retaining the required immunogenicity needed for subsequent protection against infection. P athogenic yersiniae lead to two types of diseases: yersiniosis (typified by gastroenteritis caused by Yersinia enterocolitica and Y. pseudotuberculosis) (1) and plague (evoked by Y. pestis) (2, 3). Y. pestis has evolved from Y. pseudotuberculosis within the last 20,000 years by acquiring additional plasmids and pathogenicity islands as well as by deactivating some genes (4-6). This evolutionary adaptation allowed the plague bacterium to maintain a dual life-style in fleas and rodents/mammals and conferred the ability to survive in the blood instead of the intestine (3). Plague manifests itself in three forms: bubonic (acquired from an infected rodent through a flea bite), pneumonic (acquired either directly by aerosol transmission from an infected host's lungs through respiratory droplets or secondarily from bubonic plague), and septicemic (severe bacteremia either directly due to a flea bite or subsequent to bubonic or pneumonic plague) (2). T...

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Vt)mentioning

confidence: 99%

Section: Vt)mentioning

confidence: 99%

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Combinational Deletion of Three Membrane Protein-Encoding Genes Highly Attenuates Yersinia pestis while Retaining Immunogenicity in a Mouse Model of Pneumonic Plague

et al. 2015

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Plague vaccines: new developments in an ongoing search

Rosenzweig

Hendrix

Chopra

2021

Appl Microbiol Biotechnol

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As the reality of pandemic threats challenges humanity, exemplified during the ongoing SARS-CoV-2 infections, the development of vaccines targeting these etiological agents of disease has become increasingly critical. Of paramount concern are novel and reemerging pathogens that could trigger such events, including the plague bacterium Yersinia pestis . Y. pestis is responsible for more human deaths than any other known pathogen and exists globally in endemic regions of the world, including the four corners region and Northern California in the USA. Recent cases have been scattered throughout the world, including China and the USA, with serious outbreaks in Madagascar during 2008, 2013–2014, and, most recently, 2017–2018. This review will focus on recent advances in plague vaccine development, a seemingly necessary endeavor, as there is no Food and Drug Administration–licensed vaccine available for human distribution in western nations, and that antibiotic-resistant strains are recovered clinically or intentionally developed. Progress and recent development involving subunit, live-attenuated, and nucleic acid–based plague vaccine candidates will be discussed in this review. Key points • Plague vaccine development remains elusive yet critical. • DNA, animal, and live-attenuated vaccine candidates gain traction.

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