User:Conkle.30/sandbox

Topic: What factors influenced the evolution of Streptococcus mutans? Specifically, how has evolution selected for the traits that increase the bacteria's cariogenicity.

Annotated Bibliography
'''Banas, J.A., J.D. Miller, M.E. Fuschino, et al. 2006. Evidence that Accumulation of Mutants in a Biofilm Reflects Natural Selection Rather than Stress-Induced Adaptive Mutation. Applied and Environmental Microbiology 73: 357-361.'''

This paper addressed the question as to whether adaptive mutation or natural selection was responsible for the increase in mutants within a biofilm. S. mutans was the bacteria that was studied. Specifically, the authors focused on the protein gtfA, which is responsible for successful biofilm architecture. Several strains and combinations of S. mutans were used including a wild type, a knockout variety for gtfA, and a mutant. Through genetic recombination of two homologous genes (gtfB and gtfC), the mutant was able to increase the success of its biofilms, compensating for its lack of gtfA. Rats were inoculated with these strains (wild type, or a varying ratio of knockouts to mutants) and mutant numbers were calculated after 30 days. The percentage of mutants increased significantly in rats inoculated with a mixture of low mutants to high knockouts. Furthermore, the mutant colonies collect at the end of the study were determined to be from the original inoculant and not from spontaneous or adaptive mutations. Therefore, the authors concluded that, under stressful situations (i.e. an unsuccessful biofilm environment) natural selection was responsible for the increase in mutants. While this paper does not focus directly on the evolution of S. mutans, it is important to cite because it provides insight into the evolutionary forces at play in a biofilm environment.

'''Cornejo, O.E., T. Lefébure, P.D. Pavinski Bitar, et al. 2012. Evolutionary and Population Genomics of the Cavity Causing Bacteria Streptococcus mutans. Molecular Biology and Evolution 30(4):881-893.'''

The study by Cornejo et al. analyzed the genome of Streptococcus mutans along with several other closely related bacterial species in order to look for sites of variation that increased S. mutans’ cariogenic ability. The authors used several next generation technologies to sequence the genome. They were specifically interested in single nucleotide polymorphisms. It was found that recombination was more significant to genetic changes than random mutation. The study also suggested that the population of S. mutans began expanding exponentially about 10,00 years ago, a time frame that coincides with the birth of agriculture. Additionally, the paper mentioned several adaptations acquired by S. mutans, such as improved carbohydrate metabolism and acid tolerance, that enable it to successfully colonize and adhere to the oral cavity of humans. The authors estimated that there are 14 genes under selection that contribute to these adaptations. This paper fails to include details of selection pressures that lead to the noted adaptations. Yet, it is important to cite this paper because it provides a genetic basis for the evolution of S. mutans.

'''Hoshino, T., T. Fujiwara, and S. Kawabata. 2012. Evolution of Cariogenic Character in Streptococcus mutans: Horizontal Transmission of Glycosyl Hydrolase Family 70 Genes. Scientific Reports 2:518-525.'''

The anthropological basis for the spread of dental caries has been well studied, but the factors that lead to Streptococcus mutans’ increase in cariogenicity have been generally ignored. The authors proposed a mechanism for acquisition of genes that enable S. mutans to adhere to hard surfaces, thus increasing their ability to cause dental caries. Multiple phylogenetic trees were constructed to trace the gene of interest, gtf, through several bacterial species, including Lactobacillus and Leuconostoc. The study suggested that when ancient peoples began to farm and consume fermented foods, such as yogurt, the bacteria found in these foods interacted with the bacteria of the oral cavity. The authors concluded that horizontal gene transfer was responsible S. mutans’ acquisition of gtf. This study will contribute to my research paper because it outlines another genetic mechanism contributing to the evolution of S. mutans.

'''Simon, L. 2007. The Role of Streptococcus mutans And Oral Ecology in the Formation of Dental Caries. Journal of Young Investigators.'''

This article addressed the role bacteria play in the formation of dental caries and why S. mutans is the primary etiological agent for this disease. The author did not use any novel methods for investigation, but instead, reviewed many other articles to provide a thorough description of the factors contributing to dental caries. These factors include initial acquisition of the bacteria, characteristics leading to virulence, and potential treatment methods. The primary characteristics of S. mutans that lead to virulence are its ability to adhere to hard surfaces, its production of lactic acid, and acidophilicity. The author concluded that the diet of industrialized societies plays a great role in selecting for the most pathogenic bacteria. Since my paper is about the evolution of S. mutans, specifically regarding the traits that increase its cariogenicity, it is important to include a description of dental caries formation, cause and effect, and treatments.

'''Simón, M., R. Montiel, A. Smerling, et al. 2014. Molecular Analysis of Ancient Caries. Proceedings of The Royal Society 281: 20140586.'''

The authors sought to discover sites of variation in the genome of S. mutans that were under selection or neutral evolution. They used S. mutans DNA samples from dental caries found in the remains of humans dating back to the Bronze Age and compared them to modern day samples. Specifically, they studied a gene that codes for dextranase, an enzyme that contributes to the bacteria’s virulence. They discovered no changes for this gene as a whole when comparing ancient samples to modern day samples. However, there was one location along the gene that shows evidence for positive selection and another location that suggests a shift away from neutral evolution. The results were promising, yet there is still much work to be done considering the evolution of S. mutans and its connection to human development. Areas of improvement include larger sample sizes of ancient DNA and the study of a larger genomic sequence. Additionally, this paper fails to provide an explanation of or suggestion for the selection pressures influencing the genome. Even though this gene did not show signs of strong evolutionary changes, this paper is nevertheless important to cite because it analyzed the evolution of a virulence factor of S. mutans.

October 1 Assignment Link and Content
Wiki Page Dental_caries This is where I added my suggestions to the talk page. Another Wiki user moved the sentence I added to this page to the page that's specifically for the bacteria. My sentence contribution can be found at: Streptococcus mutans

Talk Suggestions Vertical Transmission and Evolutionary Factors of Streptococcus mutans that increased its cariogenicity

I think providing information on the evolutionary forces that lead to S. mutans’s ability to cause dental caries can enhance this article.

1. One of the characteristics that make S. mutans the primary etiological agents for caries is its ability to adhere to the hard surfaces of teeth through formation of biofilms. It is thought that the bacteria acquired the gene to do this through horizontal gene transfer with other lactic acid producing bacteria, such as Lactobacillus. The gene under study is glucosyltranferase, or gtf.

2. Throughout evolutionary history, S. mutans has also acquired adaptations that have increased its fitness in the oral cavity. These traits include improved carbohydrate metabolism and greater acid tolerance. It is estimated that there are currently 14 genes under selection that contribute to these adaptations.

3. Lastly, I'd like to suggest that more detail be provided regarding vertical transmission of S. mutans, which is most often from mother (or caretaker) to child. This is listed in the article, but I would like to provide research findings that support this form of transmission. For example, it was previous thought that S. mutans isn't found in the oral cavity until young children's teeth erupt. However, studies have shown S. mutans to be present in the grooves of the tongue on pre-dentate infants. This suggests support for vertical transmission from mother to child shortly after birth.

Sentence added to main page It is believed that Streptococcus mutans acquired the gene that enables it to produce biofilms though horizontal gene transfer with other lactic acid bacterial species, such as Lactobacillus.

Full Length Paper (Final Draft)
Dental caries, more commonly known as cavities, is one of the most pervasive and non-discriminating infectious diseases on the planet (Simon 2007). It is caused primarily by the bacteria Streptococcus mutans, and evidence of the disease can be found as far back as 1.5 million years ago in ancient remains of human ancestors (Simón et al. 2007). Over time, the bacteria have evolved traits to survive in the dynamic environment of the oral cavity. However, these traits, while beneficial to the bacteria, amplify its virulence and the prevalence of dental caries increased significantly in human populations. With each evolutionary event, a change occurred in the genotype of the bacteria, which resulted in a change in its phenotype. These phenotypic changes, which were most often at the level of protein production, provided the material necessary for natural selection to act on. Thus, S. mutans evolved into the bacteria it is today.

It is thought that S. mutans is originally introduced into the oral cavity of humans at a very young age, even before primary teeth erupt. Acquisition of the virulent bacteria is most often due to vertical transmission from mother to child, most likely shortly after birth (Simon 2007). Evidence of geographic, racial and familial genetic differences between human samples of S. mutans strains supports the hypothesis of maternal transmission (Cornejo et al. 2012; Caufield et al. 2006). Furthermore, S. mutans strains among Caucasian populations are very genetically similar. This lack of genetic diversity could have been caused by a genetic drift event, which would have reduced the amount of genetic variation, at random, within the bacterial strains. Caufield et al. (2006) specifically suggests that a bottleneck event may have occurred. However, it was noted that more bacterial samples must be taken from additional Caucasian populations to further support the bottleneck hypothesis (Caufield et al. 2006).

Despite these genetic nuances between populations, there are three key traits that have evolved in all strains of S. mutans that help to increase its virulence, consequently making it the primary etiological agent for dental caries. These traits are increased organic acid production, the ability to survive and thrive in a low pH environment, and the capacity to form biofilms on the hard surfaces of teeth, the later of these traits being the most important (Banas et al. 2007). These virulence factors are significant because they can be used to study the evolutionary changes that occurred between the bacteria and its human host (Simón et al. 2014).

During its evolution, S. mutans acquired the ability to increase the amount of carbohydrates it could metabolize, and consequently more organic acid was produced as a byproduct (Cornejo et al. 2012). This is significant in the formation of dental caries because increased acidity in the oral cavity amplifies the rate of demineralization of the tooth, which leads to carious lesions (Takahashi and Nyvad 2010). The studies of Cornejo et al. (2012) propose that this trait evolved in S. mutans via lateral gene transfer with another bacterial species present in the oral cavity. Possible mechanisms for this genetic exchange include bacterial transformation and/or conjugation. There are several genes, SMU.438 and SMU.1561, involved in carbohydrate metabolism that are up-regulated in S. mutans. These genes possibly originated from Lactococcus lactis and S. gallolyticus, respectively (Cornejo et al. 2012). Acquiring this trait increased the fitness of S. mutans because it allowed the bacteria to utilize more resources. These bacteria would be able to outcompete others and thus experience positive selection. A second virulence trait that evolved in S. mutans is its ability to not only survive, but also thrive in acidic conditions (Takahashi and Nyvad 2010). One of the challenges for bacteria residing in the oral cavity are the dynamic pH conditions that vary throughout the day as food is consumed. In particular, carbohydrates significantly lower the pH of the oral cavity. Therefore, when ancient humans began to consume a more carbohydrate rich diet, S. mutans had to adapt to the changing environment (Cornejo et al. 2012). The ability to remain viable in low pH conditions gives S. mutans a selective advantage over other members of the oral microbiota. As a result, S. mutans could once again outcompete other species, and occupy additional regions of the mouth, such as advanced dental plaques, which can be as acidic as pH 4.0 (Takahashi and Nyvad 2010).

While increased acid production and acid tolerance are key characteristics of the cariogenic ability of S. mutans, no other acquired trait contributes to virulence as much as the ability to form biofilms. This is because the bacteria must be able to adhere to the surface of the tooth long enough for a cavity to form. One experiment that supports the significance of biofilms was conducted in 1971 by de Stoppelaar et al. The experiment used rodents and genetically manipulated strains of S. mutans that were unable to form biofilms. It was concluded that the rodents inoculated with wild-type S. mutans developed more carious lesions than the rodents that were inoculated with the genetically modified strain.

S. mutans acquired the trait for biofilm formation in the same way that it acquired the ability to increase its metabolism of carbohydrates, i.e. through horizontal gene transfer. By incorporating new genes into its genome, S. mutans would have gained additional proteins that could serve novel functions. In the case of biofilm production, S. mutans specifically acquired the glucosyltransferase (GTF) gene that allows the bacteria to produce polysaccharides from sucrose. These sticky polysaccharides are responsible for the bacteria’s ability to aggregate with one another and adhere to tooth enamel (Hoshino et al. 2012).

The GTF genes found in S. mutans most likely are derived from other anaerobic bacteria found in the oral cavity, such as Lactobacillus or Leuconostoc. These bacteria are commonly found in fermented foods and would have been introduced into the oral cavity when humans began consuming products such as yogurt, wine, and cheese. Moreover, the GTF genes in S. mutans display homology with similar genes found in Lactobacillus and Leuconostoc. The common ancestral gene is believed to have been used for hydrolysis and linkage of carbohydrates. Many function of this gene were preserved, but novel features arose in response to new conditions specific to each bacterial species (Hoshino et al. 2012).

Further evidence for the acquisition of the GTF gene can be found in the comparison of modern and historic caries samples. Ancient samples lack any indication of the present of biofilms or the GFT gene, which suggests that the GTF gene must have been acquired at some point in time. Additionally, it is important to note that strains of non-oral streptococci bacteria do not contain the GTF gene. Thus, this evolutionary event was unique to S. mutans present in the oral cavity (Hoshino et al. 2012).

The work by Banas et al. (2007) not only further supports the role of GTF genes in biofilm production, but also provides evidence of additional evolutionary forces at work within biofilms themselves. They observed biofilm formation in three separate groups of S. mutans: a wild-type group, an engineered knockout group (deletion of the GTF gene), and a knockout group, which included mutants that were able to partially restore the GFT gene. This mutation is significant since an inability to form successful biofilms increases the amount of stress the colony experiences. The research group was specifically interested in what was driving the increase in mutants in the third group. They concluded that it was natural selection, not adaptive mutation that drove selection. Since the mutation was beneficial and increased the fitness of the colony, those bacteria that could restore partial GTF function were selected for and consequently increased in number. There was no increase in the rate of mutation itself.

In discussing the evolution of S. mutans, it is imperative to include the role humans have played and the co-evolution that has occurred between the two species. As humans evolved anthropologically, the bacteria evolved biologically. It is widely accepted that the advent of agriculture in early human populations provided the conditions S. mutans needed to evolve into the virulent bacteria it is today. Agriculture introduced fermented foods, as well as more carbohydrate rich foods, into the diets of historic human populations. As a result, the environment of the oral cavity began to change (Cornejo et al. 2012).

With these environmental changes, the microflora that previously resided under relatively neutral and static pH conditions experienced novel selection pressure caused by the new, more acidic and dynamic pH state. This resulted in the selection of S. mutans strains that were more fit and possessed the adaptations needed to survive and reproduce in the harsher habitat. However, these traits, while beneficial for the bacteria, cause an increase in dental caries. Consequently, the relationship between humans and S. mutans evolved from one of mutualism to one of parasitism (Takahashi and Nyvad 2010). Now that humans and S. mutans have taken on novel roles, it is possible that this new association is driving additional evolutionary changes within both species, as described by the Red Queen hypothesis. The interplay between the two species was eloquently summarized by Takahashi and Nyvad (2010) as being amphibiotic, i.e. “the dynamic adaptation that occurs  in response to changing environmental conditions between two dissimilar organisms living  together.”

Lastly, another significant change to the oral environment occurred during the Industrial Revolution. More efficient manufacturing of foodstuffs increased the availability and amount of sucrose consumed by humans. This provided S. mutans with more energy resources, and thus exacerbated an already rising rate of dental caries (Hoshino et al. 2012).

It is important to note that there is alternative research, which suggests that human’s involvement in the evolution of S. mutans began many years before the onset of agriculture. Humphrey et al. (2013) cites the adoption of a hunter-gatherer lifestyle as the anthropological change that prompted the cariogenic traits of S. mutans. Remains of Pleistocene hunter-gatherers from Northern Africa provide evidence of an increased rate of dental caries. This could be contributed to their reliance on cariogenic plant foods, like acorns that are rich in carbohydrates, and increase wear and tear of teeth, which would have reduced protective tooth enamel (Humphrey et al. 2013).

Throughout the history of S. mutans, there have been three significant evolutionary events that have lead to an increase in the fitness of the bacteria. At the same time, these adaptations have increased the bacteria’s virulence, and consequently, the prevalence of dental caries has risen dramatically (Simon 2007). These events were largely induced by the anthropological changes of humans (Cornejo et al. 2012). Since the two species continue to share a parasitic relationship, it is possible that the two groups will continue to influence the evolution of one another (Takahashi and Nyvad 2010). Additionally, since dental caries is such a prevalent disease, it is important to continue researching the evolution of S. mutans (Simon 2007). Possible topics include the effect of modern dental practices on the evolution of S. mutans and additional prevention methods targeted at the three specific cariogenic adaptations.

References:

Banas, J. A., J. D. Miller, M. E. Fuschino, K. R. O. Hazlett, W. Toyofuku, K. A. Porter, S. B. 	Reutzel, M. A. Florczyk, K. A. McDonough, and S. M. Michalek. 2007. Evidence that accumulation of mutants in a biofilm reflects natural selection rather than stress-induced adaptive mutation. Applied and Environmental Microbiology 73(1): 357–361.

Caufield, P. W., D. Saxena, D. Fitch, and Y. Li. 2006. Population structure of plasmid-containing strains of Streptococcus mutans, a member of the human indigenous biota. Journal of Bacteriology 189(4): 1238–1243.

Cornejo, O. E., T. Lefébure, P.D. Pavinski Bitar, P. Lang, V. P. Richards, K. Eilertson, T. Do, D. Beighton, L. Zeng, S. J. Ahn, R. A. Burne, A. Siepel, C. D. Bustamante, and M. J. 	Stanhope. 2012. Evolutionary and population genomics of the cavity causing bacteria Streptococcus mutans. Mol. Biol. Evol. 30(4): 881–893.

De Stoppelaar, J.D., K. G. König, A. J. M. Plasschaer, and J. S. Van Der Hoeven. (1971). Decreased cariogenicity of a mutant of Streptococcus mutans. Archs Oral Biol. 16: 971-	975.

Hoshino, T., T. Fujiwara, and S. Kawabata. 2012. Evolution of cariogenic character in Streptococcus mutans: Horizontal transmission of glycosyl hydrolase family 70 genes. Scientific Reports 2: 518-524.

Humphrey, L. T., I. De Groote, J. Morales, N. Barton, S. Collcutt, C. B. Ramsey, and A. Bouzouggar. 2013. Earliest evidence for caries and exploitation of starchy plant foods in Pleistocene hunter-gatherers from Morocco. PNAS 111(3): 954–959.

Simon, L. 2007. The role of Streptococcus mutans and oral ecology in the formation of dental caries. Journal of Young Investigators.

Simón, M., R. Montiel, A. Smerling, E. Solórzano, N. Díaz, B. A. Àlvarez-Sandoval, A. R. Jiménez-Marin, and A. Malgosa. 2014. Molecular analysis of ancient caries. Proc. R. Soc. B 281: 20140586.

Takahashi, N., and B. Nyvad. 2010. The role of bacteria in the caries process: Ecological 	perspectives. Journal of Dental Research 90(3): 294-303.

Edits to "Live" Wiki Page Streptococcus mutans
I added the entire evolution section and added a sentence to the children section, which I highlighted in bold here.

Evolution

There are three key traits that have evolved in S. mutans that help increase its adaptability to the oral cavity, but at the same time, these traits have increased its virulence. These characteristics are increased organic acid production, the capacity to form biofilms on the hard surfaces of teeth, and the ability to survive and thrive in a low pH environment.

During its evolution, S. mutans acquired the ability to increase the amount of carbohydrates it could metabolize, and consequently more organic acid was produced as a byproduct. This is significant in the formation of dental caries because increased acidity in the oral cavity amplifies the rate of demineralization of the tooth, which leads to carious lesions. It is thought that the trait evolved in S. mutans via lateral gene transfer with another bacterial species present in the oral cavity. There are several genes, SMU.438 and SMU.1561, involved in carbohydrate metabolism that are up-regulated in S. mutans. These genes possibly originated from Lactococcus lactis and S. gallolyticus, respectively.

Another instance of lateral gene transfer is responsible for S. mutans’ acquisition of the glucosyltransferase (GTF) gene, which allows the bacteria to produce polysaccharides from sucrose. These sticky polysaccharides are responsible for the bacteria’s ability to aggregate with one another and adhere to tooth enamel, i.e. to form biofilms. The GTF genes found in S. mutans most likely are derived from other anaerobic bacteria found in the oral cavity, such as Lactobacillus or Leuconostoc. Additionally, the GTF genes in S. mutans display homology with similar genes found in Lactobacillus and Leuconostoc. The common ancestral gene is believed to have been used for hydrolysis and linkage of carbohydrates.

The third trait that evolved in S. mutans is its ability to not only survive, but also thrive in acidic conditions. This trait gives S. mutans a selective advantage over other members of the oral microbiota. As a result, S. mutans could outcompete other species, and occupy additional regions of the mouth, such as advanced dental plaques, which can be as acidic as pH 4.0. Natural selection is most likely the primary evolutionary mechanisms responsible for this trait.

In discussing the evolution of S. mutans, it is imperative to include the role humans have played and the co-evolution that has occurred between the two species. As humans evolved anthropologically, the bacteria evolved biologically. It is widely accepted that the advent of agriculture in early human populations provided the conditions S. mutans needed to evolve into the virulent bacteria it is today. Agriculture introduced fermented foods, as well as more carbohydrate rich foods, into the diets of historic human populations. These new foods would have introduced new bacteria into the oral cavity, as well as created new environmental conditions. For example, Lactobacillus or Leuconostoc are typically found in foods such as yogurt and wine. Also, consuming more carbohydrates would have increase the amount of sugars available to S. mutans for metabolism as well as lowered the pH of the oral cavity. This newly acidic habitat would select for those bacteria that could survive and reproduce at a lower pH.

Another significant change to the oral environment occurred during the Industrial Revolution. More efficient manufacturing of foodstuffs increased the availability and amount of sucrose consumed by humans. This provided S. mutans with more energy resources, and thus exacerbated an already rising rate of dental caries.

Children In general, S. mutans is acquired in the oral cavity at the moment of tooth eruption. But S. mutans has been detected in the oral cavity of predentate children. This suggests that the eruption of teeth is not a necessary prerequisite. Thus, this species may not be confined to dental plaque. Additionally, acquisition of the virulent bacteria is most often due to vertical transmission from mother to child, most likely shortly after birth. The adhesion, invasion, and persistence within the oral cells are considered the virulence mechanism of S. mutans to colonize and survive in the oral cavity in the absence of a tooth surface.