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Early emergence of Yersinia pestis as a severe respiratory pathogen.

Zimbler DL, Schroeder JA, Eddy JL, Lathem WW - Nat Commun (2015)

Bottom Line: Y. pestis recently evolved from the gastrointestinal pathogen Y. pseudotuberculosis; however, it is not known at what point Y. pestis gained the ability to induce a fulminant pneumonia.As Y. pestis further evolved, modern strains acquired a single amino-acid modification within Pla that optimizes protease activity.While this modification is unnecessary to cause pneumonic plague, the substitution is instead needed to efficiently induce the invasive infection associated with bubonic plague.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology-Immunology, Northwestern University Feinberg School of Medicine, Chicago, Illinois 60611, USA.

ABSTRACT
Yersinia pestis causes the fatal respiratory disease pneumonic plague. Y. pestis recently evolved from the gastrointestinal pathogen Y. pseudotuberculosis; however, it is not known at what point Y. pestis gained the ability to induce a fulminant pneumonia. Here we show that the acquisition of a single gene encoding the protease Pla was sufficient for the most ancestral, deeply rooted strains of Y. pestis to cause pneumonic plague, indicating that Y. pestis was primed to infect the lungs at a very early stage in its evolution. As Y. pestis further evolved, modern strains acquired a single amino-acid modification within Pla that optimizes protease activity. While this modification is unnecessary to cause pneumonic plague, the substitution is instead needed to efficiently induce the invasive infection associated with bubonic plague. These findings indicate that Y. pestis was capable of causing pneumonic plague before it evolved to optimally cause invasive infections in mammals.

No MeSH data available.


Related in: MedlinePlus

Pestoides F is competent to carry pPCP1 and produce active Pla.(a) Relative copy number of pPCP1 (represented by the pPCP1-encoded genes pla and pst) in the pPCP1-reintroduced Pestoides F and CO92 strains, compared with CO92 (set at 1). pspA was used as a control for a chromosomal gene. Relative copy number for each gene was measured by quantitative PCR from gDNA isolated from cultures grown overnight at 37 °C and normalized to gyrB. Data are combined from three independent biological replicates repeated twice; error bars represent the s.e.m. (b) Immunoblot analysis of whole-cell lysates of indicated Y. pestis strains grown at 37 °C with antibodies against Pla and RpoA (as a loading control). Panel is representative of three independent replicates. The presence or absence of pPCP1 in each strain is indicated. Full blots are shown in Supplementary Fig. 6. (c) The Plg-activating ability of the indicated Y. pestis CO92 or Pestoides F strains cultured at 37 °C is shown. Data are representative of three independent experiments performed in triplicate; error bars represent the s.e.m.
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f3: Pestoides F is competent to carry pPCP1 and produce active Pla.(a) Relative copy number of pPCP1 (represented by the pPCP1-encoded genes pla and pst) in the pPCP1-reintroduced Pestoides F and CO92 strains, compared with CO92 (set at 1). pspA was used as a control for a chromosomal gene. Relative copy number for each gene was measured by quantitative PCR from gDNA isolated from cultures grown overnight at 37 °C and normalized to gyrB. Data are combined from three independent biological replicates repeated twice; error bars represent the s.e.m. (b) Immunoblot analysis of whole-cell lysates of indicated Y. pestis strains grown at 37 °C with antibodies against Pla and RpoA (as a loading control). Panel is representative of three independent replicates. The presence or absence of pPCP1 in each strain is indicated. Full blots are shown in Supplementary Fig. 6. (c) The Plg-activating ability of the indicated Y. pestis CO92 or Pestoides F strains cultured at 37 °C is shown. Data are representative of three independent experiments performed in triplicate; error bars represent the s.e.m.

Mentions: On the basis of the similarities of the respiratory infection between Pestoides F and CO92 Δpla, we asked whether the acquisition of pPCP1 (and specifically Pla) was sufficient for this ancestral Y. pestis strain to cause primary pneumonic plague. However, it is not yet known whether Pestoides F is able to stably harbour pPCP1 and produce active Pla. To test this, we introduced a kanamycin-marked pPCP1 derived from CO92 (ref. 14) into the pCD1+ and pCD1− strains of Pestoides F. To ensure that this marked version of pPCP1 did not affect its encoded functions, we reintroduced the kanamycin-marked pPCP1 plasmid back into the pCD1+ and pCD1− strains of CO92 lacking pPCP1; in all cases, the kanamycin resistance marker was subsequently excised by Flp-based recombination. To test the ability of Pestoides F to carry pPCP1, we measured the copy number of the pla and pst genes (both encoded on pPCP1) in Pestoides F relative to that in CO92 and Δpla CO92 carrying the reintroduced pPCP1. The copy number of pPCP1 carried by wild-type CO92 and the unmarked, reintroduced pPCP1 in CO92 are similar, indicating that the scar left by removal of the antibiotic cassette has no effect on plasmid replication (Fig. 3a). In addition, we found that Pestoides F naturally maintains pPCP1 without antibiotic selection (albeit at a higher relative copy number compared with CO92; Fig. 3a). Furthermore, neither Angola nor Pestoides A maintain pPCP1 at a significantly different copy number compared with wild-type CO92 (Supplementary Fig. 3a). To determine the conservation of pla regulation between ancestral and modern strains, a Ppla-gfp reporter containing the CO92 pla promoter cloned upstream of the coding sequence (CDS) for the green fluorescent protein (GFP) was integrated onto the chromosomes of CO92, Angola, Pestoides A and Pestoides F at the attTn7 site. Bacteria were cultured at 37 °C and the fluorescence of each strain was measured and normalized to the optical densities of the cultures. No significant change in fluorescence was observed between any of the ancestral strains compared with that of CO92 (Supplementary Fig. 3b), suggesting no differences in the regulation of pla transcription within these strains. Finally, immunoblot analysis of these strains cultured under the same conditions demonstrates that Pestoides F is able to synthesize Pla from pPCP1 at a similar level to that of CO92 (Fig. 3b).


Early emergence of Yersinia pestis as a severe respiratory pathogen.

Zimbler DL, Schroeder JA, Eddy JL, Lathem WW - Nat Commun (2015)

Pestoides F is competent to carry pPCP1 and produce active Pla.(a) Relative copy number of pPCP1 (represented by the pPCP1-encoded genes pla and pst) in the pPCP1-reintroduced Pestoides F and CO92 strains, compared with CO92 (set at 1). pspA was used as a control for a chromosomal gene. Relative copy number for each gene was measured by quantitative PCR from gDNA isolated from cultures grown overnight at 37 °C and normalized to gyrB. Data are combined from three independent biological replicates repeated twice; error bars represent the s.e.m. (b) Immunoblot analysis of whole-cell lysates of indicated Y. pestis strains grown at 37 °C with antibodies against Pla and RpoA (as a loading control). Panel is representative of three independent replicates. The presence or absence of pPCP1 in each strain is indicated. Full blots are shown in Supplementary Fig. 6. (c) The Plg-activating ability of the indicated Y. pestis CO92 or Pestoides F strains cultured at 37 °C is shown. Data are representative of three independent experiments performed in triplicate; error bars represent the s.e.m.
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f3: Pestoides F is competent to carry pPCP1 and produce active Pla.(a) Relative copy number of pPCP1 (represented by the pPCP1-encoded genes pla and pst) in the pPCP1-reintroduced Pestoides F and CO92 strains, compared with CO92 (set at 1). pspA was used as a control for a chromosomal gene. Relative copy number for each gene was measured by quantitative PCR from gDNA isolated from cultures grown overnight at 37 °C and normalized to gyrB. Data are combined from three independent biological replicates repeated twice; error bars represent the s.e.m. (b) Immunoblot analysis of whole-cell lysates of indicated Y. pestis strains grown at 37 °C with antibodies against Pla and RpoA (as a loading control). Panel is representative of three independent replicates. The presence or absence of pPCP1 in each strain is indicated. Full blots are shown in Supplementary Fig. 6. (c) The Plg-activating ability of the indicated Y. pestis CO92 or Pestoides F strains cultured at 37 °C is shown. Data are representative of three independent experiments performed in triplicate; error bars represent the s.e.m.
Mentions: On the basis of the similarities of the respiratory infection between Pestoides F and CO92 Δpla, we asked whether the acquisition of pPCP1 (and specifically Pla) was sufficient for this ancestral Y. pestis strain to cause primary pneumonic plague. However, it is not yet known whether Pestoides F is able to stably harbour pPCP1 and produce active Pla. To test this, we introduced a kanamycin-marked pPCP1 derived from CO92 (ref. 14) into the pCD1+ and pCD1− strains of Pestoides F. To ensure that this marked version of pPCP1 did not affect its encoded functions, we reintroduced the kanamycin-marked pPCP1 plasmid back into the pCD1+ and pCD1− strains of CO92 lacking pPCP1; in all cases, the kanamycin resistance marker was subsequently excised by Flp-based recombination. To test the ability of Pestoides F to carry pPCP1, we measured the copy number of the pla and pst genes (both encoded on pPCP1) in Pestoides F relative to that in CO92 and Δpla CO92 carrying the reintroduced pPCP1. The copy number of pPCP1 carried by wild-type CO92 and the unmarked, reintroduced pPCP1 in CO92 are similar, indicating that the scar left by removal of the antibiotic cassette has no effect on plasmid replication (Fig. 3a). In addition, we found that Pestoides F naturally maintains pPCP1 without antibiotic selection (albeit at a higher relative copy number compared with CO92; Fig. 3a). Furthermore, neither Angola nor Pestoides A maintain pPCP1 at a significantly different copy number compared with wild-type CO92 (Supplementary Fig. 3a). To determine the conservation of pla regulation between ancestral and modern strains, a Ppla-gfp reporter containing the CO92 pla promoter cloned upstream of the coding sequence (CDS) for the green fluorescent protein (GFP) was integrated onto the chromosomes of CO92, Angola, Pestoides A and Pestoides F at the attTn7 site. Bacteria were cultured at 37 °C and the fluorescence of each strain was measured and normalized to the optical densities of the cultures. No significant change in fluorescence was observed between any of the ancestral strains compared with that of CO92 (Supplementary Fig. 3b), suggesting no differences in the regulation of pla transcription within these strains. Finally, immunoblot analysis of these strains cultured under the same conditions demonstrates that Pestoides F is able to synthesize Pla from pPCP1 at a similar level to that of CO92 (Fig. 3b).

Bottom Line: Y. pestis recently evolved from the gastrointestinal pathogen Y. pseudotuberculosis; however, it is not known at what point Y. pestis gained the ability to induce a fulminant pneumonia.As Y. pestis further evolved, modern strains acquired a single amino-acid modification within Pla that optimizes protease activity.While this modification is unnecessary to cause pneumonic plague, the substitution is instead needed to efficiently induce the invasive infection associated with bubonic plague.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology-Immunology, Northwestern University Feinberg School of Medicine, Chicago, Illinois 60611, USA.

ABSTRACT
Yersinia pestis causes the fatal respiratory disease pneumonic plague. Y. pestis recently evolved from the gastrointestinal pathogen Y. pseudotuberculosis; however, it is not known at what point Y. pestis gained the ability to induce a fulminant pneumonia. Here we show that the acquisition of a single gene encoding the protease Pla was sufficient for the most ancestral, deeply rooted strains of Y. pestis to cause pneumonic plague, indicating that Y. pestis was primed to infect the lungs at a very early stage in its evolution. As Y. pestis further evolved, modern strains acquired a single amino-acid modification within Pla that optimizes protease activity. While this modification is unnecessary to cause pneumonic plague, the substitution is instead needed to efficiently induce the invasive infection associated with bubonic plague. These findings indicate that Y. pestis was capable of causing pneumonic plague before it evolved to optimally cause invasive infections in mammals.

No MeSH data available.


Related in: MedlinePlus