Limits...
Endopeptidase-mediated beta lactam tolerance.

Dörr T, Davis BM, Waldor MK - PLoS Pathog. (2015)

Bottom Line: In response to a wide variety of cell wall--acting antibiotics, this pathogen loses its rod shape, indicative of cell wall degradation, and becomes spherical.Other autolysins proved dispensable for this process.Our findings suggest the enzymes that mediate cell wall degradation are critical for determining bacterial cell fate--sphere formation vs. lysis--after treatment with antibiotics that target cell wall synthesis.

View Article: PubMed Central - PubMed

Affiliation: Division of Infectious Diseases, Brigham and Women's Hospital and Howard Hughes Medical Institute, Boston, Massachusetts, United States of America; Department of Microbiology and Immunobiology, Harvard Medical School, Boston, Massachusetts, United States of America.

ABSTRACT
In many bacteria, inhibition of cell wall synthesis leads to cell death and lysis. The pathways and enzymes that mediate cell lysis after exposure to cell wall-acting antibiotics (e.g. beta lactams) are incompletely understood, but the activities of enzymes that degrade the cell wall ('autolysins') are thought to be critical. Here, we report that Vibrio cholerae, the cholera pathogen, is tolerant to antibiotics targeting cell wall synthesis. In response to a wide variety of cell wall--acting antibiotics, this pathogen loses its rod shape, indicative of cell wall degradation, and becomes spherical. Genetic analyses revealed that paradoxically, V. cholerae survival via sphere formation required the activity of D,D endopeptidases, enzymes that cleave the cell wall. Other autolysins proved dispensable for this process. Our findings suggest the enzymes that mediate cell wall degradation are critical for determining bacterial cell fate--sphere formation vs. lysis--after treatment with antibiotics that target cell wall synthesis.

No MeSH data available.


Related in: MedlinePlus

V. cholerae amiB is not required for sphere formation in response to inhibition of cell wall synthesis.(A-C) A V. cholerae ΔamiB mutant was treated as described in Fig 1 A-C. Scale bar = 5 μm. No AB = no antibiotic added.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4401780&req=5

ppat.1004850.g002: V. cholerae amiB is not required for sphere formation in response to inhibition of cell wall synthesis.(A-C) A V. cholerae ΔamiB mutant was treated as described in Fig 1 A-C. Scale bar = 5 μm. No AB = no antibiotic added.

Mentions: In E. coli, lysis following exposure to cell-wall acting antibiotics appears to be partially dependent on the redundant PG amidases AmiA, AmiB, and AmiC, whose cleavage of septal PG enables daughter-cell separation [12]. The V. cholerae genome encodes a single PG amidase (AmiB), and an amiB deletion mutant forms long chains of unseparated daughter cells comparable to those of amidase-deficient E. coli [28]. Unexpectedly, V. cholerae amidase-deficient cells were more, rather than less, susceptible than wild type cells to killing by penicillin, phosphomycin and D-cycloserine (Fig 2A). However, this susceptibility was not associated with the bacterial lysis seen in drug-treated E. coli. We observed no significant decrease in culture density (OD600) of the V. cholerae amiB mutant in response to cell-wall acting antibiotics (S3 Fig), and bacterial lysis was rarely observed using light microscopy. Instead, antibiotic treatment ultimately resulted in formation of spherical cells that were similar to those formed by wild type bacteria (Fig 2B and 2C). We speculate that the ultimate decline in viability of the ΔamiB mutant under these conditions may reflect its previously noted compromised cell envelope [28]. It is important to note that interpretation of cfu and OD600 data is complicated by the fact that the mutant’s multi-cell chains disintegrate into single spheres upon beta lactam treatment (Fig 2B and 2C; discussed below). This is likely the cause of the observed increase in cfu directly after addition of antibiotic in Fig 2A, which may somewhat obscure a loss of viability. The mutant’s increased lag phase compared to the wild type further complicated direct comparisons based on culture density and cfu. Therefore, to directly assess AmiB’s role in the sphere formation process, we turned to single cell analysis.


Endopeptidase-mediated beta lactam tolerance.

Dörr T, Davis BM, Waldor MK - PLoS Pathog. (2015)

V. cholerae amiB is not required for sphere formation in response to inhibition of cell wall synthesis.(A-C) A V. cholerae ΔamiB mutant was treated as described in Fig 1 A-C. Scale bar = 5 μm. No AB = no antibiotic added.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4401780&req=5

ppat.1004850.g002: V. cholerae amiB is not required for sphere formation in response to inhibition of cell wall synthesis.(A-C) A V. cholerae ΔamiB mutant was treated as described in Fig 1 A-C. Scale bar = 5 μm. No AB = no antibiotic added.
Mentions: In E. coli, lysis following exposure to cell-wall acting antibiotics appears to be partially dependent on the redundant PG amidases AmiA, AmiB, and AmiC, whose cleavage of septal PG enables daughter-cell separation [12]. The V. cholerae genome encodes a single PG amidase (AmiB), and an amiB deletion mutant forms long chains of unseparated daughter cells comparable to those of amidase-deficient E. coli [28]. Unexpectedly, V. cholerae amidase-deficient cells were more, rather than less, susceptible than wild type cells to killing by penicillin, phosphomycin and D-cycloserine (Fig 2A). However, this susceptibility was not associated with the bacterial lysis seen in drug-treated E. coli. We observed no significant decrease in culture density (OD600) of the V. cholerae amiB mutant in response to cell-wall acting antibiotics (S3 Fig), and bacterial lysis was rarely observed using light microscopy. Instead, antibiotic treatment ultimately resulted in formation of spherical cells that were similar to those formed by wild type bacteria (Fig 2B and 2C). We speculate that the ultimate decline in viability of the ΔamiB mutant under these conditions may reflect its previously noted compromised cell envelope [28]. It is important to note that interpretation of cfu and OD600 data is complicated by the fact that the mutant’s multi-cell chains disintegrate into single spheres upon beta lactam treatment (Fig 2B and 2C; discussed below). This is likely the cause of the observed increase in cfu directly after addition of antibiotic in Fig 2A, which may somewhat obscure a loss of viability. The mutant’s increased lag phase compared to the wild type further complicated direct comparisons based on culture density and cfu. Therefore, to directly assess AmiB’s role in the sphere formation process, we turned to single cell analysis.

Bottom Line: In response to a wide variety of cell wall--acting antibiotics, this pathogen loses its rod shape, indicative of cell wall degradation, and becomes spherical.Other autolysins proved dispensable for this process.Our findings suggest the enzymes that mediate cell wall degradation are critical for determining bacterial cell fate--sphere formation vs. lysis--after treatment with antibiotics that target cell wall synthesis.

View Article: PubMed Central - PubMed

Affiliation: Division of Infectious Diseases, Brigham and Women's Hospital and Howard Hughes Medical Institute, Boston, Massachusetts, United States of America; Department of Microbiology and Immunobiology, Harvard Medical School, Boston, Massachusetts, United States of America.

ABSTRACT
In many bacteria, inhibition of cell wall synthesis leads to cell death and lysis. The pathways and enzymes that mediate cell lysis after exposure to cell wall-acting antibiotics (e.g. beta lactams) are incompletely understood, but the activities of enzymes that degrade the cell wall ('autolysins') are thought to be critical. Here, we report that Vibrio cholerae, the cholera pathogen, is tolerant to antibiotics targeting cell wall synthesis. In response to a wide variety of cell wall--acting antibiotics, this pathogen loses its rod shape, indicative of cell wall degradation, and becomes spherical. Genetic analyses revealed that paradoxically, V. cholerae survival via sphere formation required the activity of D,D endopeptidases, enzymes that cleave the cell wall. Other autolysins proved dispensable for this process. Our findings suggest the enzymes that mediate cell wall degradation are critical for determining bacterial cell fate--sphere formation vs. lysis--after treatment with antibiotics that target cell wall synthesis.

No MeSH data available.


Related in: MedlinePlus