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Metagenomics Study Uncovers Rapid Shifts in Mouth Microbe Immune Sequences

By Andrea Anderson

NEW YORK (GenomeWeb News) – The anti-viral immune sequences used by some mouth microbes changes rapidly and varies dramatically from one individual to the next, according to a metagenomic study appearing online last night in Genome Research.

A California research team collected saliva samples from four healthy individuals at various time points over a year to a year-and-a-half and used 16S ribosomal RNA sequencing to characterize the individuals' mouth microbiomes at each sampling time. They then incorporated information on the clustered regularly spaced interspersed short palindromic repeats, or CRISPR, sequences that streptococcal bacteria in these samples were using to protect themselves against potentially harmful viruses at each sampling time.

While mouth bacterial communities tended to remain fairly stable, the researchers reported, the Streptococcus CRISPR sequences changed quickly and markedly, even within the same individual.

"The CRISPRs are constantly changing, which is giving us pretty good evidence that these bacteria are encountering and becoming resistant to these viruses on what's likely a daily basis," lead author David Pride, a pathology researcher at the University of California at San Diego, told GenomeWeb Daily News.

Because viruses can influence the composition of bacterial communities, Pride explained, understanding the nature of bacteriophage infections and the strategies used to combat them is relevant to understanding microbial communities, including those in and on the human body.

"Viruses really are sort of a neglected part of the microbiome in humans," he said, explaining that most microbiome studies to date have focused on bacterial and/or archaeal members of these microbial communities.

"By killing off bacteria [viruses] can ultimately determine what bacteria are present and not present," Pride added. "We also think that viruses are the main tools by which new genes come in and out of the community."

Pride and his co-workers did 16S rRNA sequencing with the Roche 454 GS FLX Titanium platform to assess bacterial communities in saliva samples collected from four individuals over the course of 11 to 17 months.

They also used Sanger sequencing to determine what sorts of viral sequences were present in Streptococcus CRISPR sequences to get clues about how often these bacteria encounter viruses and the effects of these interactions.

The team focused on bacteria in this genera because Streptococcus is known to be an important bacteria for the human oral cavity, Pride noted.

CRISPR sequences in general are a means by which bacteria protect themselves against viral interlopers. By incorporating bits of viral DNA into these CRISPR sequences, bacteria are able to mount a defense against subsequent introgressions by the same sorts of viruses.

"As bacteria see new viruses, the bacteria actually cut out pieces of the viral genome, put them into their own DNA, and then use that through a mechanism that's called nucleic acid interference," Pride explained, blocking viral reproduction.

When they began analyzing their sequence data — which included sequence representing 427,917 16S rRNA genes and 6,859 CRISPR sequences — the researchers found that the overall microbial communities appeared to be fairly stable within each individual.

In contrast, just seven to 22 percent of the viral sequences in streptococcal CRISPR regions remained present in each person throughout the course of the study.

And, they explained, roughly a third of the sequences shifted within the same individual each time they were tested, suggesting Streptococcus CRISPRs in the mouth microbiome shift daily or perhaps even more frequently.

In addition, there was little to no overlap in the streptococcal CRISPR spacer sequences found in different individuals sampled. For instance, Pride noted, even though two of the four study participants came from the same household, these individuals only shared about two percent of their CRISPR sequences.

Based on their findings so far, those involved in the study speculate that it may ultimately be possible to incorporate information on an individual's mouth microbiome immune patterns in tracking health and treating disease.

"The individual-specific and traceable character of CRISPR spacer complements could potentially open the way for expansion of the domain of personalized medicine to the oral microbiome," they wrote, "where lineages may be tracked as a function of health and other factors."

In the future, the researchers plan to continue looking at viral and bacterial interactions in the mouth microbiome by studying the same and additional individuals, Pride said. They also hope to begin exploring how such interactions influence oral health and disease.

"The next couple of stages are to try to understand how these things, particularly the CRISPRs and the viruses and the bacteria, differ between oral health and folks who have periodontal disease or gingivitis," he said.

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