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Joseph A. Sorg - One of the best experts on this subject based on the ideXlab platform.
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The requirement for co-germinants during Clostridium difficile Spore Germination is influenced by mutations in yabG and cspA
2019Co-Authors: Ritu Shrestha, Alicia M. Cochran, Joseph A. SorgAbstract:Clostridium difficile Spore Germination is critical for the transmission of disease. C. difficile Spores germinate in response to cholic acid derivatives, such as taurocholate (TA), and amino acids, such as glycine or alanine. Although the receptor with which bile acids are recognized (germinant receptor) is known, the amino acid co-germinant receptor has remained elusive. Here, we used EMS mutagenesis to generate mutants with altered requirements for the amino acid co-germinant, similar to the strategy we used previously to identify the bile acid germinant receptor, CspC. Surprisingly, we identified strains that do not require co-germinants, and the mutant Spores germinated in response to TA alone. Upon sequencing these mutants, we identified different mutations in yabG. In C. difficile, yabG expression is required for the processing of key Germination components to their mature forms (e.g., CspBA to CspB and CspA). A defined yabG mutant exacerbated the EMS mutant phenotype. Building upon this work, we found that small deletions in cspA resulted in Spores that germinated in the presence of TA alone without the requirement of a co-germinant. cspA encodes a pseudoprotease that was previously shown to be important for incorporation of the CspC germinant receptor. Herein, our study builds upon the role of CspA during C. difficile Spore Germination by providing evidence that CspA is important for recognition of co-germinants during C. difficile Spore Germination. Our work suggests that two pseudoproteases (CspC and CspA) likely function as the C. difficile germinant receptors.
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Model for C. difficile Spore Germination.
2019Co-Authors: Ritu Shrestha, Alicia M. Cochran, Joseph A. SorgAbstract:(A) In our working model for wildtype C. difficile Spore Germination, CspB protease activity is inhibited by two pseudoproteases, CspC (the bile acid germinant receptor; [38]) and CspA (implicated as the co-germinant receptor; this study). Interaction of CspC with cholic acid derivatives [31] and the interaction of CspA with co-germinants [32, 68] results in these two proteins disassociating from CspB. Subsequently, CspB cleaves the inhibitory pro-domain from proSleC thereby activating SleC’s cortex degrading activity. (B) In the C. difficile yabG::ermB mutant, CspBA is not processed into the two proteins important for C. difficile Spore Germination. In this scenario, the CspA portion is not positioned correctly to inhibit CspB activity until the co-germinant signal is received. This results in CspB activity being inhibited only by CspC. In response to TA, CspC dissociates from CspBA and CspB (of CspBA) processes preproSleC to the active, SleC, form.
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a clostridium difficile alanine racemase affects Spore Germination and accommodates serine as a substrate
Journal of Biological Chemistry, 2017Co-Authors: Ritu Shrestha, Steve W Lockless, Joseph A. SorgAbstract:Clostridium difficile has become one of the most common bacterial pathogens in hospital-acquired infections in the United States. Although C. difficile is strictly anaerobic, it survives in aerobic environments and transmits between hosts via Spores. C. difficile Spore Germination is triggered in response to certain bile acids and glycine. Although glycine is the most effective co-germinant, other amino acids can substitute with varying efficiencies. Of these, l-alanine is an effective co-germinant and is also a germinant for most bacterial Spores. Many endoSpore-forming bacteria embed alanine racemases into their Spore coats, and these enzymes are thought to convert the l-alanine germinant into d-alanine, a Spore Germination inhibitor. Although the C. difficile Alr2 racemase is the sixth most highly expressed gene during C. difficile Spore formation, a previous study reported that Alr2 has little to no role in Germination of C. difficile Spores in rich medium. Here, we hypothesized that Alr2 could affect C. difficile l-alanine-induced Spore Germination in a defined medium. We found that alr2 mutant Spores more readily germinate in response to l-alanine as a co-germinant. Surprisingly, d-alanine also functioned as a co-germinant. Moreover, we found that Alr2 could interconvert l- and d-serine and that Alr2 bound to l- and d-serine with ∼2-fold weaker affinity to that of l- and d-alanine. Finally, we demonstrate that l- and d-serine are also co-germinants for C. difficile Spores. These results suggest that C. difficile Spores can respond to a diverse set of amino acid co-germinants and reveal that Alr2 can accommodate serine as a substrate.
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a clostridium difficile alanine racemase affects Spore Germination and accommodates serine as a substrate
Journal of Biological Chemistry, 2017Co-Authors: Ritu Shrestha, Steve W Lockless, Joseph A. SorgAbstract:Abstract Clostridium difficile has become one of the most common bacterial pathogens in hospital-acquired infections in the United States. Although C. difficile is strictly anaerobic, it survives in aerobic environments and transmits between hosts via Spores. C. difficile Spore Germination is triggered in response to certain bile acids and glycine. Although glycine is the most effective co-germinant, other amino acids can substitute with varying efficiencies. Of these, L-alanine is an effective co-germinant and is also a germinant for most bacterial Spores. Many endoSpore-forming bacteria embed alanine racemases into their Spore coats and these enzymes are thought to convert the L-alanine germinant into D-alanine, a Spore Germination inhibitor. Although, the C. difficile Alr2 racemase is the sixth most highly-expressed gene during C. difficile Spore formation, a previous study reported that Alr2 has little to no role in Germination of C. difficile Spores in rich medium. Here, we hypothesized that Alr2 could affect C. difficile L-alanine-induced Spore Germination, in a defined medium. We found that alr2 mutant Spores more readily germinate in response to L-alanine as a co-germinant. Surprisingly, D-alanine also functioned as a co-germinant. Moreover, we found that Alr2 could interconvert L- and D-serine and that Alr2 bound to L- and D-serine with approximately 2-fold weaker affinity to that of L- and D-alanine. Finally, we demonstrate that L- and D-serine are also co-germinants for C. difficile Spores. These results suggest that C. difficile Spores can respond to a diverse set of amino acid co-germinants and reveal that Alr2 can accommodate serine as a substrate.
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detecting cortex fragments during bacterial Spore Germination
Journal of Visualized Experiments, 2016Co-Authors: Michael B Francis, Joseph A. SorgAbstract:The process of endoSpore Germination in Clostridium difficile, and other Clostridia, increasingly is being found to differ from the model Spore-forming bacterium, Bacillus subtilis. Germination is triggered by small molecule germinants and occurs without the need for macromolecular synthesis. Though differences exist between the mechanisms of Spore Germination in species of Bacillus and Clostridium, a common requirement is the hydrolysis of the peptidoglycan-like cortex which allows the Spore core to swell and rehydrate. After rehydration, metabolism can begin and this, eventually, leads to outgrowth of a vegetative cell. The detection of hydrolyzed cortex fragments during Spore Germination can be difficult and the modifications to the previously described assays can be confusing or difficult to reproduce. Thus, based on our recent report using this assay, we detail a step-by-step protocol for the colorimetric detection of cortex fragments during bacterial Spore Germination.
Michael B Francis - One of the best experts on this subject based on the ideXlab platform.
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detecting cortex fragments during bacterial Spore Germination
Journal of Visualized Experiments, 2016Co-Authors: Michael B Francis, Joseph A. SorgAbstract:The process of endoSpore Germination in Clostridium difficile, and other Clostridia, increasingly is being found to differ from the model Spore-forming bacterium, Bacillus subtilis. Germination is triggered by small molecule germinants and occurs without the need for macromolecular synthesis. Though differences exist between the mechanisms of Spore Germination in species of Bacillus and Clostridium, a common requirement is the hydrolysis of the peptidoglycan-like cortex which allows the Spore core to swell and rehydrate. After rehydration, metabolism can begin and this, eventually, leads to outgrowth of a vegetative cell. The detection of hydrolyzed cortex fragments during Spore Germination can be difficult and the modifications to the previously described assays can be confusing or difficult to reproduce. Thus, based on our recent report using this assay, we detail a step-by-step protocol for the colorimetric detection of cortex fragments during bacterial Spore Germination.
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Spore cortex hydrolysis precedes dipicolinic acid release during clostridium difficile Spore Germination
Journal of Bacteriology, 2015Co-Authors: Michael B Francis, Charlotte A Allen, Joseph A. SorgAbstract:ABSTRACT Bacterial Spore Germination is a process whereby a dormant Spore returns to active, vegetative growth, and this process has largely been studied in the model organism Bacillus subtilis. In B. subtilis, the initiation of germinant receptor-mediated Spore Germination is divided into two genetically separable stages. Stage I is characterized by the release of dipicolinic acid (DPA) from the Spore core. Stage II is characterized by cortex degradation, and stage II is activated by the DPA released during stage I. Thus, DPA release precedes cortex hydrolysis during B. subtilis Spore Germination. Here, we investigated the timing of DPA release and cortex hydrolysis during Clostridium difficile Spore Germination and found that cortex hydrolysis precedes DPA release. Inactivation of either the bile acid germinant receptor, cspC, or the cortex hydrolase, sleC, prevented both cortex hydrolysis and DPA release. Because both cortex hydrolysis and DPA release during C. difficile Spore Germination are dependent on the presence of the germinant receptor and the cortex hydrolase, the release of DPA from the core may rely on the osmotic swelling of the core upon cortex hydrolysis. These results have implications for the hypothesized glycine receptor and suggest that the initiation of germinant receptor-mediated C. difficile Spore Germination proceeds through a novel Germination pathway. IMPORTANCEClostridium difficile infects antibiotic-treated hosts and spreads between hosts as a dormant Spore. In a host, Spores germinate to the vegetative form that produces the toxins necessary for disease. C. difficile Spore Germination is stimulated by certain bile acids and glycine. We recently identified the bile acid germinant receptor as the Germination-specific, protease-like CspC. CspC is likely cortex localized, where it can transmit the bile acid signal to the cortex hydrolase, SleC. Due to the differences in location of CspC compared to the Bacillus subtilis germinant receptors, we hypothesized that there are fundamental differences in the Germination processes between the model organism and C. difficile. We found that C. difficile Spore Germination proceeds through a novel pathway.
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Spore cortex hydrolysis precedes dipicolinic acid release during clostridium difficile Spore Germination
Journal of Bacteriology, 2015Co-Authors: Michael B Francis, Charlotte A Allen, Joseph A. SorgAbstract:ABSTRACT Bacterial Spore Germination is a process whereby a dormant Spore returns to active, vegetative growth, and this process has largely been studied in the model organism Bacillus subtilis. In B. subtilis, the initiation of germinant receptor-mediated Spore Germination is divided into two genetically separable stages. Stage I is characterized by the release of dipicolinic acid (DPA) from the Spore core. Stage II is characterized by cortex degradation, and stage II is activated by the DPA released during stage I. Thus, DPA release precedes cortex hydrolysis during B. subtilis Spore Germination. Here, we investigated the timing of DPA release and cortex hydrolysis during Clostridium difficile Spore Germination and found that cortex hydrolysis precedes DPA release. Inactivation of either the bile acid germinant receptor, cspC, or the cortex hydrolase, sleC, prevented both cortex hydrolysis and DPA release. Because both cortex hydrolysis and DPA release during C. difficile Spore Germination are dependent on the presence of the germinant receptor and the cortex hydrolase, the release of DPA from the core may rely on the osmotic swelling of the core upon cortex hydrolysis. These results have implications for the hypothesized glycine receptor and suggest that the initiation of germinant receptor-mediated C. difficile Spore Germination proceeds through a novel Germination pathway. IMPORTANCEClostridium difficile infects antibiotic-treated hosts and spreads between hosts as a dormant Spore. In a host, Spores germinate to the vegetative form that produces the toxins necessary for disease. C. difficile Spore Germination is stimulated by certain bile acids and glycine. We recently identified the bile acid germinant receptor as the Germination-specific, protease-like CspC. CspC is likely cortex localized, where it can transmit the bile acid signal to the cortex hydrolase, SleC. Due to the differences in location of CspC compared to the Bacillus subtilis germinant receptors, we hypothesized that there are fundamental differences in the Germination processes between the model organism and C. difficile. We found that C. difficile Spore Germination proceeds through a novel pathway.
Ritu Shrestha - One of the best experts on this subject based on the ideXlab platform.
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Model for C. difficile Spore Germination.
2019Co-Authors: Ritu Shrestha, Alicia M. Cochran, Joseph A. SorgAbstract:(A) In our working model for wildtype C. difficile Spore Germination, CspB protease activity is inhibited by two pseudoproteases, CspC (the bile acid germinant receptor; [38]) and CspA (implicated as the co-germinant receptor; this study). Interaction of CspC with cholic acid derivatives [31] and the interaction of CspA with co-germinants [32, 68] results in these two proteins disassociating from CspB. Subsequently, CspB cleaves the inhibitory pro-domain from proSleC thereby activating SleC’s cortex degrading activity. (B) In the C. difficile yabG::ermB mutant, CspBA is not processed into the two proteins important for C. difficile Spore Germination. In this scenario, the CspA portion is not positioned correctly to inhibit CspB activity until the co-germinant signal is received. This results in CspB activity being inhibited only by CspC. In response to TA, CspC dissociates from CspBA and CspB (of CspBA) processes preproSleC to the active, SleC, form.
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The requirement for co-germinants during Clostridium difficile Spore Germination is influenced by mutations in yabG and cspA
2019Co-Authors: Ritu Shrestha, Alicia M. Cochran, Joseph A. SorgAbstract:Clostridium difficile Spore Germination is critical for the transmission of disease. C. difficile Spores germinate in response to cholic acid derivatives, such as taurocholate (TA), and amino acids, such as glycine or alanine. Although the receptor with which bile acids are recognized (germinant receptor) is known, the amino acid co-germinant receptor has remained elusive. Here, we used EMS mutagenesis to generate mutants with altered requirements for the amino acid co-germinant, similar to the strategy we used previously to identify the bile acid germinant receptor, CspC. Surprisingly, we identified strains that do not require co-germinants, and the mutant Spores germinated in response to TA alone. Upon sequencing these mutants, we identified different mutations in yabG. In C. difficile, yabG expression is required for the processing of key Germination components to their mature forms (e.g., CspBA to CspB and CspA). A defined yabG mutant exacerbated the EMS mutant phenotype. Building upon this work, we found that small deletions in cspA resulted in Spores that germinated in the presence of TA alone without the requirement of a co-germinant. cspA encodes a pseudoprotease that was previously shown to be important for incorporation of the CspC germinant receptor. Herein, our study builds upon the role of CspA during C. difficile Spore Germination by providing evidence that CspA is important for recognition of co-germinants during C. difficile Spore Germination. Our work suggests that two pseudoproteases (CspC and CspA) likely function as the C. difficile germinant receptors.
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a clostridium difficile alanine racemase affects Spore Germination and accommodates serine as a substrate
Journal of Biological Chemistry, 2017Co-Authors: Ritu Shrestha, Steve W Lockless, Joseph A. SorgAbstract:Clostridium difficile has become one of the most common bacterial pathogens in hospital-acquired infections in the United States. Although C. difficile is strictly anaerobic, it survives in aerobic environments and transmits between hosts via Spores. C. difficile Spore Germination is triggered in response to certain bile acids and glycine. Although glycine is the most effective co-germinant, other amino acids can substitute with varying efficiencies. Of these, l-alanine is an effective co-germinant and is also a germinant for most bacterial Spores. Many endoSpore-forming bacteria embed alanine racemases into their Spore coats, and these enzymes are thought to convert the l-alanine germinant into d-alanine, a Spore Germination inhibitor. Although the C. difficile Alr2 racemase is the sixth most highly expressed gene during C. difficile Spore formation, a previous study reported that Alr2 has little to no role in Germination of C. difficile Spores in rich medium. Here, we hypothesized that Alr2 could affect C. difficile l-alanine-induced Spore Germination in a defined medium. We found that alr2 mutant Spores more readily germinate in response to l-alanine as a co-germinant. Surprisingly, d-alanine also functioned as a co-germinant. Moreover, we found that Alr2 could interconvert l- and d-serine and that Alr2 bound to l- and d-serine with ∼2-fold weaker affinity to that of l- and d-alanine. Finally, we demonstrate that l- and d-serine are also co-germinants for C. difficile Spores. These results suggest that C. difficile Spores can respond to a diverse set of amino acid co-germinants and reveal that Alr2 can accommodate serine as a substrate.
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a clostridium difficile alanine racemase affects Spore Germination and accommodates serine as a substrate
Journal of Biological Chemistry, 2017Co-Authors: Ritu Shrestha, Steve W Lockless, Joseph A. SorgAbstract:Abstract Clostridium difficile has become one of the most common bacterial pathogens in hospital-acquired infections in the United States. Although C. difficile is strictly anaerobic, it survives in aerobic environments and transmits between hosts via Spores. C. difficile Spore Germination is triggered in response to certain bile acids and glycine. Although glycine is the most effective co-germinant, other amino acids can substitute with varying efficiencies. Of these, L-alanine is an effective co-germinant and is also a germinant for most bacterial Spores. Many endoSpore-forming bacteria embed alanine racemases into their Spore coats and these enzymes are thought to convert the L-alanine germinant into D-alanine, a Spore Germination inhibitor. Although, the C. difficile Alr2 racemase is the sixth most highly-expressed gene during C. difficile Spore formation, a previous study reported that Alr2 has little to no role in Germination of C. difficile Spores in rich medium. Here, we hypothesized that Alr2 could affect C. difficile L-alanine-induced Spore Germination, in a defined medium. We found that alr2 mutant Spores more readily germinate in response to L-alanine as a co-germinant. Surprisingly, D-alanine also functioned as a co-germinant. Moreover, we found that Alr2 could interconvert L- and D-serine and that Alr2 bound to L- and D-serine with approximately 2-fold weaker affinity to that of L- and D-alanine. Finally, we demonstrate that L- and D-serine are also co-germinants for C. difficile Spores. These results suggest that C. difficile Spores can respond to a diverse set of amino acid co-germinants and reveal that Alr2 can accommodate serine as a substrate.
Charlotte A Allen - One of the best experts on this subject based on the ideXlab platform.
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Spore cortex hydrolysis precedes dipicolinic acid release during clostridium difficile Spore Germination
Journal of Bacteriology, 2015Co-Authors: Michael B Francis, Charlotte A Allen, Joseph A. SorgAbstract:ABSTRACT Bacterial Spore Germination is a process whereby a dormant Spore returns to active, vegetative growth, and this process has largely been studied in the model organism Bacillus subtilis. In B. subtilis, the initiation of germinant receptor-mediated Spore Germination is divided into two genetically separable stages. Stage I is characterized by the release of dipicolinic acid (DPA) from the Spore core. Stage II is characterized by cortex degradation, and stage II is activated by the DPA released during stage I. Thus, DPA release precedes cortex hydrolysis during B. subtilis Spore Germination. Here, we investigated the timing of DPA release and cortex hydrolysis during Clostridium difficile Spore Germination and found that cortex hydrolysis precedes DPA release. Inactivation of either the bile acid germinant receptor, cspC, or the cortex hydrolase, sleC, prevented both cortex hydrolysis and DPA release. Because both cortex hydrolysis and DPA release during C. difficile Spore Germination are dependent on the presence of the germinant receptor and the cortex hydrolase, the release of DPA from the core may rely on the osmotic swelling of the core upon cortex hydrolysis. These results have implications for the hypothesized glycine receptor and suggest that the initiation of germinant receptor-mediated C. difficile Spore Germination proceeds through a novel Germination pathway. IMPORTANCEClostridium difficile infects antibiotic-treated hosts and spreads between hosts as a dormant Spore. In a host, Spores germinate to the vegetative form that produces the toxins necessary for disease. C. difficile Spore Germination is stimulated by certain bile acids and glycine. We recently identified the bile acid germinant receptor as the Germination-specific, protease-like CspC. CspC is likely cortex localized, where it can transmit the bile acid signal to the cortex hydrolase, SleC. Due to the differences in location of CspC compared to the Bacillus subtilis germinant receptors, we hypothesized that there are fundamental differences in the Germination processes between the model organism and C. difficile. We found that C. difficile Spore Germination proceeds through a novel pathway.
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Spore cortex hydrolysis precedes dipicolinic acid release during clostridium difficile Spore Germination
Journal of Bacteriology, 2015Co-Authors: Michael B Francis, Charlotte A Allen, Joseph A. SorgAbstract:ABSTRACT Bacterial Spore Germination is a process whereby a dormant Spore returns to active, vegetative growth, and this process has largely been studied in the model organism Bacillus subtilis. In B. subtilis, the initiation of germinant receptor-mediated Spore Germination is divided into two genetically separable stages. Stage I is characterized by the release of dipicolinic acid (DPA) from the Spore core. Stage II is characterized by cortex degradation, and stage II is activated by the DPA released during stage I. Thus, DPA release precedes cortex hydrolysis during B. subtilis Spore Germination. Here, we investigated the timing of DPA release and cortex hydrolysis during Clostridium difficile Spore Germination and found that cortex hydrolysis precedes DPA release. Inactivation of either the bile acid germinant receptor, cspC, or the cortex hydrolase, sleC, prevented both cortex hydrolysis and DPA release. Because both cortex hydrolysis and DPA release during C. difficile Spore Germination are dependent on the presence of the germinant receptor and the cortex hydrolase, the release of DPA from the core may rely on the osmotic swelling of the core upon cortex hydrolysis. These results have implications for the hypothesized glycine receptor and suggest that the initiation of germinant receptor-mediated C. difficile Spore Germination proceeds through a novel Germination pathway. IMPORTANCEClostridium difficile infects antibiotic-treated hosts and spreads between hosts as a dormant Spore. In a host, Spores germinate to the vegetative form that produces the toxins necessary for disease. C. difficile Spore Germination is stimulated by certain bile acids and glycine. We recently identified the bile acid germinant receptor as the Germination-specific, protease-like CspC. CspC is likely cortex localized, where it can transmit the bile acid signal to the cortex hydrolase, SleC. Due to the differences in location of CspC compared to the Bacillus subtilis germinant receptors, we hypothesized that there are fundamental differences in the Germination processes between the model organism and C. difficile. We found that C. difficile Spore Germination proceeds through a novel pathway.
Peter Setlow - One of the best experts on this subject based on the ideXlab platform.
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intracellular membranes of bacterial endoSpores are reservoirs for Spore core membrane expansion during Spore Germination
Scientific Reports, 2018Co-Authors: Michael Laue, Christin Dittmann, Peter SetlowAbstract:Bacterial endoSpores are formed by certain bacteria, such as Bacillus subtilis or the pathogenic Bacillus anthracis and Clostridioides difficile, to allow survival in environmental conditions which are lethal to vegetative bacteria. The Spores possess a particular architecture and molecular inventory which endow them with a remarkable resistance against desiccation, heat and radiation. Another remarkable Spore feature is their rapid return to vegetative growth during Spore Germination and outgrowth. The underlying processes of this latter physiological and morphological transformation involve a number of different events, some of which are mechanistically not entirely understood. One of these events is the expansion of the central Spore core, which contains the DNA, RNA and most Spore enzymes. To date, it has been unclear how the ~1.3- to 1.6-fold expansion of the core membrane surface area that accompanies core expansion takes place, since this occurs in the absence of significant if any ATP synthesis. In the current work, we demonstrate the presence of intracellular membrane structures in Spores located just below the core membrane. During Spore Germination these internal core membranes disappear when the core size increases, suggesting that they are integrated into the core membrane to allow core expansion. These intracellular membranes are most probably present as more or less compressed vesicles or tubules within the dormant Spore core. Investigations of Spores from different species suggest that these intracellular membrane structures below the core membrane are a general feature of endoSpore forming bacteria.
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A Quasi-chemical Model for Bacterial Spore Germination Kinetics by High Pressure
Food Engineering Reviews, 2017Co-Authors: Christopher J. Doona, X. Philip Ye, Kenneth Kustin, Florence E. Feeherry, Runze Huang, Haiqing Chen, Peter SetlowAbstract:High pressure processing (HPP) is an emerging non-thermal technology that is growing exponentially in use worldwide for the pasteurization of commercial foodstuffs. At combinations of elevated pressures and temperatures, HPP inactivates bacterial Spores, but HPP has not yet been implemented commercially for food sterilization. Studies of the mechanisms of bacterial Spore inactivation by HPP using primarily Spores of Bacillus species have shown that Spore Germination precedes inactivation, with the release of dipicolinic acid from the Spore core as the rate-determining step. Investigations probing Spore resistance to and Germination by HPP using Bacillus subtilis , a number of selected B. subtilis mutants, Bacillus amyloliquefaciens , and Clostridium difficile Spores have compiled a wealth of detailed mechanistic information, while also accumulating abundant Germination kinetics data that has not previously been analyzed by predictive models. Presently, we devise a “quasi-chemical” model for bacterial Spore Germination dynamics by HPP. This quasi-chemical Germination model (QCGM) hypothesizes a three-step mechanism and derives a set of ordinary differential equations to model the observed Germination dynamics. The results with this model are viewed in the context of historical studies of Spore activation, Germination, and inactivation, with an eye toward potentially integrating differential equation models for Germination and inactivation into a single, comprehensive model for Spore dynamics by HPP. With the increasing use of high hydrostatic pressure to investigate mechanisms of bacterial Spore resistance and physiology, the QCGM results help promote the efficient control of bacterial Spores, whether for the inactivation of Clostridium botulinum Spores in low-acid foods or aerosolized Bacillus anthracis Spores on textiles used in protective clothing, tents, or shelters.
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characterization of bacterial Spore Germination using phase contrast and fluorescence microscopy raman spectroscopy and optical tweezers
Nature Protocols, 2011Co-Authors: Lingbo Kong, Peter Setlow, Pengfei Zhang, Guiwen Wang, Jing Yu, Yongqing LiAbstract:Characterization of bacterial Spore Germination using phase-contrast and fluorescence microscopy, Raman spectroscopy and optical tweezers
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workshop report modeling the molecular mechanism of bacterial Spore Germination and elucidating reasons for Germination heterogeneity
Journal of Food Science, 2009Co-Authors: Karl J Indest, Wallace G Buchholz, James R Faeder, Peter SetlowAbstract:Over the course of 2 days, top researchers in the fields of bacterial Spore biology and computational biology discussed approaches to determine the cause of Spore Germination heterogeneity. Biological and mathematical data gaps were identified, and experimental approaches and computational strategies for modeling Spore Germination were presented and evaluated. As a result of these interactions, future research directions were defined, the outcome of which should result in a robust model to help define the molecular mechanism(s) of Spore Germination. Mechanistic understanding of Germination will be instrumental for developing novel sterilization, treatment, and decontamination strategies to mitigate threats posed by Spores.
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clostridium perfringens Spore Germination characterization of germinants and their receptors
Journal of Bacteriology, 2008Co-Authors: Daniel Paredessabja, Peter Setlow, Antonio J Torres, Mahfuzur R SarkerAbstract:Clostridium perfringens food poisoning is caused by type A isolates carrying a chromosomal enterotoxin (cpe) gene (C-cpe), while C. perfringens-associated non-food-borne gastrointestinal (GI) diseases are caused by isolates carrying a plasmid-borne cpe gene (P-cpe). C. perfringens Spores are thought to be the important infectious cell morphotype, and after inoculation into a suitable host, these Spores must germinate and return to active growth to cause GI disease. We have found differences in the Germination of Spores of C-cpe and P-cpe isolates in that (i) while a mixture of l-asparagine and KCl was a good germinant for Spores of C-cpe and P-cpe isolates, KCl and, to a lesser extent, l-asparagine triggered Spore Germination in C-cpe isolates only; and (ii) l-alanine or l-valine induced significant Germination of Spores of P-cpe but not C-cpe isolates. Spores of a gerK mutant of a C-cpe isolate in which two of the proteins of a Spore nutrient germinant receptor were absent germinated slower than wild-type Spores with KCl, did not germinate with l-asparagine, and germinated poorly compared to wild-type Spores with the nonnutrient germinants dodecylamine and a 1:1 chelate of Ca2+ and dipicolinic acid. In contrast, Spores of a gerAA mutant of a C-cpe isolate that lacked another component of a nutrient germinant receptor germinated at the same rate as that of wild-type Spores with high concentrations of KCl, although they germinated slightly slower with a lower KCl concentration, suggesting an auxiliary role for GerAA in C. perfringens Spore Germination. In sum, this study identified nutrient germinants for Spores of both C-cpe and P-cpe isolates of C. perfringens and provided evidence that proteins encoded by the gerK operon are required for both nutrient-induced and non-nutrient-induced Spore Germination.