Staphylococcus aureus is a gram-positive bacterium that 20-30% of the general population carries in their nasal cavity. Most S. aureus infections are not serious as they are skin and soft tissue infections like abscesses and furuncles. However, S. aureus bloodstream, joint and bone infections, pneumonia and endocarditis are sometimes fatal. S. aureus is an adaptable bacterium and this has contributed to its success in building resistance to antibiotics such as methicillin. There are two main types of staph infections: methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-susceptible Staphylococcus aureus (MSSA) with MRSA being the more common. An individual who is an S. aureus carrier may take antibiotics which eliminate the non-resistant strains and leave the resistant strains behind and when the bacteria multiply, the infections they may cause are more serious and difficult to treat.
How is S. aureus so good at what it does? Thankfully, a group of scientists led by Frédéric Peyrusson has given us insight into the processes S. aureus undergoes to transform into a stronger, more resistant strain of bacteria that is marked by one trait - persistence.
How is S. aureus so good at what it does? Thankfully, a group of scientists led by Frédéric Peyrusson has given us insight into the processes S. aureus undergoes to transform into a stronger, more resistant strain of bacteria that is marked by one trait - persistence.
Staphylococcus aureus can form persister cells. Persisters are a subpopulation of bacterial cells that experience a non-growing state and a tolerance to high antibiotic concentrations. These persister cells exist in a transient state and therefore, when the external stresses such as antibiotics are removed, they are able to return to their usual state in which they multiply. The existence of persister cells is one of the reasons S. aureus has been successful at causing recurring infections.
The experimenters classified S. aureus cells as persisters by first treating a population of bacterial cells with three different antibiotics - oxacillin, clarithromycin and moxifloxacin. They found that after 24 hours of exposure to high concentrations of antibiotics there was a tolerant pool of S. aureus and they underwent biphasic killing. Biphasic killing is a feature of persister cells in which more susceptible bacteria are killed quickly followed by a slower bacterial cell death in persister subpopulations.
The existence of S. aureus persister cells is marked by the activation of the stringent response. Rapid synthesis of alarmones mediates the stringent response and leads to alterations in the S. aureus transcriptome, namely the activation of genes responsible for resisting stress and surviving starvation conditions as well a cessation of bacterial division.
There is differential gene expression between wild type S. aureus and S. aureus persister cells wherein some genes in the persister subpopulation are upregulated and others are downregulated. Upregulated genes correspond to galactose metabolism while downregulated genes correspond to proliferation and metabolic processes.
One striking discovery from the experiment was that S. aureus could develop tolerance to multiple antibiotic classes after exposure to a single antibiotic. For the experimenters, this was an indication that the bacterium undergoes many intracellular mechanisms. With this, they proposed a model of the factors that allow S. aureus to develop tolerance and persistence. They carried out a series of experiments using murine J774A.1 macrophages, human macrophages and S. aureus strain ‘SH1000’ in a series of cell cultures to observe the pathways shown in Figure 2.
The experimenters classified S. aureus cells as persisters by first treating a population of bacterial cells with three different antibiotics - oxacillin, clarithromycin and moxifloxacin. They found that after 24 hours of exposure to high concentrations of antibiotics there was a tolerant pool of S. aureus and they underwent biphasic killing. Biphasic killing is a feature of persister cells in which more susceptible bacteria are killed quickly followed by a slower bacterial cell death in persister subpopulations.
The existence of S. aureus persister cells is marked by the activation of the stringent response. Rapid synthesis of alarmones mediates the stringent response and leads to alterations in the S. aureus transcriptome, namely the activation of genes responsible for resisting stress and surviving starvation conditions as well a cessation of bacterial division.
There is differential gene expression between wild type S. aureus and S. aureus persister cells wherein some genes in the persister subpopulation are upregulated and others are downregulated. Upregulated genes correspond to galactose metabolism while downregulated genes correspond to proliferation and metabolic processes.
One striking discovery from the experiment was that S. aureus could develop tolerance to multiple antibiotic classes after exposure to a single antibiotic. For the experimenters, this was an indication that the bacterium undergoes many intracellular mechanisms. With this, they proposed a model of the factors that allow S. aureus to develop tolerance and persistence. They carried out a series of experiments using murine J774A.1 macrophages, human macrophages and S. aureus strain ‘SH1000’ in a series of cell cultures to observe the pathways shown in Figure 2.
Environmental factors like antibiotic pressure and a carbon source shift, in this case between glucose and lactose, exert stress onto the cell. These factors, in unison, cause changes to the intracellular environment as the cell carries out a number of responses. The heat shock, SOS and stringent response, cell wall and translation maintenance and changes in the proton motive force cause the cell to build multiple tolerances. Long term survival is dependent on these tolerances. Removal of the stressors can either allow S. aureus to revert to its replicative form and replenish its population or become a more resistant strain through the potentiation of higher mutation rates and substantial horizontal gene transfer.
RNA sequencing of wild-type S. aureus and S. aureus persister cells allowed the experimenters to get an overview of the bacterial persisters transcriptomic profile. Environmental stresses of different natures can collectively push the bacterial cell beyond a threshold and initiate a series of responses that ultimately lead to a persister phenotype.
The aforementioned stringent response inhibits proliferation and cellular processes which require high energy. This response is always changing so the researchers decided to explore the expression of its regulatory network during S. aureus infection. Soon after the bacteria are taken up by macrophages, quantitative RT-PCR indicated a rapid, temporary boost in the response regulators relQ, relP, codY and rsh.
The SOS response allows for DNA repair and therefore improves DNA integrity. Effectors of the SOS response network are transcribed along with genes encoding some antibiotic targets and this may be one of the reasons for tolerance.
The heat shock response decreases the number of misfolded proteins and helps reduce the chances of protein aggregation in the cell thereby controlling and maintaining translation.
Differential expression analysis showed that a majority of the genes for oxidative phosphorylation are repressed in bacterial persisters. This inhibits the proton motive force by limiting energy input.
The cell builds multiple tolerances as the external factors trigger the intracellular responses.
These multiple tolerances lead to long term survival of S. aureus persister cells. Maintenance of ATP also contributes to long term survival. Persister cells that have regulated their intracellular processes as a response to stressors can revert to their regular state and multiply thereby resulting in a population of bacteria after stressor removal. The bacterial persisters may also lead to a population of more resistant cells if the stressors are maintained. This higher resistance is attributed to higher rates of mutation and horizontal gene transfer.
The experimenters believe persistence is a potential critical trigger for the failure of therapeutic treatments. S. aureus owes some of its success to its ability to form bacterial persister cells. Treatment of staph infections with antibiotics may not eliminate the infection but rather, push these bacteria to quiescence. When antibiotic treatments stop and the S. aureus population is revived, so is the infection. This makes serious staph infections difficult to treat and sometimes fatal. We mentioned before that S. aureus could develop tolerance to multiple antibiotic classes after exposure to a single antibiotic. The experimenters noted the clinical meaning or significance of this finding remains to be established. Perhaps this could mean a change in the way S. aureus infections are treated in that multiple antibiotics could be administered at once, as opposed to a single antibiotic, and there may be less chances for the bacteria to develop this comprehensive tolerance.
The goal of the experiment was to provide evidence of S. aureus persister cells. An understanding of persistence in S. aureus including the mechanisms leading to a persistent phenotype and the alterations persistent cells make to survive harsh conditions is essential to improving therapies targeting the treatment of S. aureus infections.
RNA sequencing of wild-type S. aureus and S. aureus persister cells allowed the experimenters to get an overview of the bacterial persisters transcriptomic profile. Environmental stresses of different natures can collectively push the bacterial cell beyond a threshold and initiate a series of responses that ultimately lead to a persister phenotype.
The aforementioned stringent response inhibits proliferation and cellular processes which require high energy. This response is always changing so the researchers decided to explore the expression of its regulatory network during S. aureus infection. Soon after the bacteria are taken up by macrophages, quantitative RT-PCR indicated a rapid, temporary boost in the response regulators relQ, relP, codY and rsh.
The SOS response allows for DNA repair and therefore improves DNA integrity. Effectors of the SOS response network are transcribed along with genes encoding some antibiotic targets and this may be one of the reasons for tolerance.
The heat shock response decreases the number of misfolded proteins and helps reduce the chances of protein aggregation in the cell thereby controlling and maintaining translation.
Differential expression analysis showed that a majority of the genes for oxidative phosphorylation are repressed in bacterial persisters. This inhibits the proton motive force by limiting energy input.
The cell builds multiple tolerances as the external factors trigger the intracellular responses.
These multiple tolerances lead to long term survival of S. aureus persister cells. Maintenance of ATP also contributes to long term survival. Persister cells that have regulated their intracellular processes as a response to stressors can revert to their regular state and multiply thereby resulting in a population of bacteria after stressor removal. The bacterial persisters may also lead to a population of more resistant cells if the stressors are maintained. This higher resistance is attributed to higher rates of mutation and horizontal gene transfer.
The experimenters believe persistence is a potential critical trigger for the failure of therapeutic treatments. S. aureus owes some of its success to its ability to form bacterial persister cells. Treatment of staph infections with antibiotics may not eliminate the infection but rather, push these bacteria to quiescence. When antibiotic treatments stop and the S. aureus population is revived, so is the infection. This makes serious staph infections difficult to treat and sometimes fatal. We mentioned before that S. aureus could develop tolerance to multiple antibiotic classes after exposure to a single antibiotic. The experimenters noted the clinical meaning or significance of this finding remains to be established. Perhaps this could mean a change in the way S. aureus infections are treated in that multiple antibiotics could be administered at once, as opposed to a single antibiotic, and there may be less chances for the bacteria to develop this comprehensive tolerance.
The goal of the experiment was to provide evidence of S. aureus persister cells. An understanding of persistence in S. aureus including the mechanisms leading to a persistent phenotype and the alterations persistent cells make to survive harsh conditions is essential to improving therapies targeting the treatment of S. aureus infections.
About the authors:
Sonako Jacas ‘21 (left) and Clarke Feng ‘21 (right) met in both of their first Biology classes at Mount Holyoke. It is fitting that they are now taking their final Biology class together (and somehow in the same lab classroom as first year!). Sonako is a Biology major and Chemistry minor and strongly believes Biology is her one true love. She hopes to be involved in biomedical research after MHC. Clarke is a Biology major with a Religion minor, with a passion for Marine Biology and Rabbinic Judaism. They love sharks and teeth, and hopefully can find a job that lets them study both!
We want to extend a special thank you to Professor Amy Camp. Her genuine love for Microbiology and the compassion she has for her students made our last undergraduate Biology course one to remember!
We want to extend a special thank you to Professor Amy Camp. Her genuine love for Microbiology and the compassion she has for her students made our last undergraduate Biology course one to remember!
No comments:
Post a Comment