Isolation and Characterization of anti-<em>Streptococcus Mutans</em> Phage as a Possible Treatment Agent for Caries

Abstract

Streptococcus mutans is a key bacterium in dental caries- one of the most prevalent chronic infectious diseases. Conventional treatment both fails to specifically target the pathogenic bacteria and at-tempts to eradicate commensal bacteria as well. Thus, caries remains one of most common and challenging diseases. The use of bacterial viruses as anti-bacterial agents, is gaining interest worldwide. Hardly any phages were described against S. mutans. The objective of this study was to isolate anti-S. mutans phages and to characterize their antimi-crobial properties. Human saliva samples were filtered and screened for potential phages. Standard double-layered agar method was used for isolation. Whole genome sequence analysis and morphology visualization by TEM, were used for anti-S. mutans phage identification. Antibacterial properties were evaluated using clinical strains and ATCC strains of S. mutans in various states. Antibacterial effect was also tested on human cariogenic dentin. One phage against S. mutans was isolated and termed SMHBZ8. This phage showed effective lytic activity in vitro against both planktonic and biofilm S. mutans cultures. Moreover, the phage showed antibacterial effect when used on cariogenic dentin. The isolation and characterization of SMHBZ8 may be the first step in developing a potential phage therapy for dental caries.

Investigating CRISPR spacer targets and their impact on genomic diversification of Streptococcus mutans

“…1D) which may indicate that on average foreign DNA elements have higher GC content than the average S. mutans genome (36.8%). Phages that infect S. mutans , M102AD, M102, and SMHBZ8 have GC content of 39.6%, 39.21% and 38.8% respectively [31, 34, 46]. The GC content of S. mutans genomes was consistent for strains with 1-20 (36.80%), 20-40 (36.82%) and 40-60 (36.79%) CRISPR spacers (Figure 6).…”

“…Isolation of anti- S. mutans phage is rare. For example, □APCM01 was the only S. mutans phage isolated from 85 saliva samples [33] and smHBZ8 was the single S. mutans phage isolated from 254 samples (diverse sources; saliva, dental sewage, cariogenic dentin, extracted teeth, and dental plaque) [31]. To understand how important CRISPR-Cas is in explaining the low frequency of infective phage isolation we calculated the frequency of phage immunity spacers across our genome database.…”

“…We anticipate that rather than infecting S. mutans, these phage genes are present either as prophage elements or as orphan genes in S. mutans genomes. The four S. mutans infecting phage that S. mutans has acquired spacer immunity against are known as M102, M102AD, APCM01 and smHBZ8 [31][32][33][34]. These are the only S. mutans phage that have been genome sequenced and all belong to the Siphoviridae family of double-stranded DNA viruses.…”

Investigating CRISPR spacer targets and their impact on genomic diversification of Streptococcus mutans

“…The four S. mutans infecting phage that S. mutans has acquired spacer immunity against are known as M102, M102AD, ɸAPCM01 and smHBZ8 (Delisle and Rostkowski, 1993;Delisle et al, 2012;Dalmasso et al, 2015;Zaken et al, 2021). These are the only S. mutans phage that have been genome sequenced and all belong to the Siphoviridae family of doublestranded DNA viruses; they are lytic phage.…”

“…Overall, it appears that immunity to the variable region is less commonly acquired, and this was most evident for M102 and M102AD. Isolation of S. mutans phage is rare, For example, ɸAPCM01 was the only S. mutans phage isolated from 85 saliva samples (Dalmasso et al, 2015) and smHBZ8 was the single S. mutans phage isolated from 254 samples (diverse sources; saliva, dental sewage, cariogenic dentin, extracted teeth, and dental plaque) (Zaken et al, 2021). To understand how important CRISPR-Cas is in explaining the low frequency of infective phage isolation we calculated the frequency of phage immunity spacers across our genome database.…”