hourThe human body is filled with trillions of microbial cells, most of which are bacteria, that make up the microbiome. Although small, some of these bacteria keep us healthy, while others promote disease. These differences often come down to the genes in each bacterial genome, but finding and sequencing rare strains can be difficult.
Gang Fang’s team developed mEnrich-seq, a sequencing technology to enrich bacteria of interest from samples.
Brian Schucha
GangbangA geneticist at the Icahn School of Medicine at Mount Sinai, he proposed a new solution: mEnrich-seq This is the result of 10 years of bacterial epigenomics research to distinguish DNA from various species for metagenomic research.One In an interview with ScientistFang describes his vision for how mEnrich-seq can help scientists answer difficult questions about humans’ companion bacteria.
What are some of the challenges associated with studying the human microbiome using metagenomics?
We have many technologies for understanding the microbiome in different ways, but there are common challenges. If a bacterial species is abundant in a sample, we can learn almost anything about it, but if the species’ abundance is very low, it is very difficult to study. Two or three lineages may coexist in the same species, and the important lineage may not be the relatively more numerous lineage. These different strains are very similar in terms of their genomes, making them very difficult to distinguish.
What motivated you to develop mEnrich-seq?
If the target is rare, most of the sequencing throughput is consumed by more abundant species. The moment we sequenced we had already lost this battle, so we needed a new strategy before we sequenced.
– Gang Fang, Mount Sinai Icahn School of Medicine
If the target is rare, most of the sequencing throughput is consumed by more abundant species. The moment we sequenced we had already lost this battle, so we needed a new strategy before we sequenced. The natural epigenetic barcoding of bacteria provides a unique way to address this problem. Although different species and strains have similar genomes, they often encode different DNA methyltransferases. DNA methylation patterns.2 Bacteria do this to distinguish between their own DNA and foreign DNA. We can use this to differentiate the genomes of species or strains based on their overall methylation patterns.
If you target a specific genome and know its methylation pattern, you can rationally select restriction enzymes that cut specific sequences, called methylation motifs. The enzyme digests most background DNA that lacks methylation to match it. mEnrich-seq allows you to enrich your bacteria of interest by more than 100-fold.
How does this protocol compare to standard metagenomic sequencing experiments?
We actually took this into consideration when designing it. We wanted it to be effective but also very easy to connect to existing pipelines. Ultimately, mEnrich-seq involves only two steps beyond standard library preparation. In the first step, the DNA is digested using rationally selected restriction enzymes after adapter ligation and before amplification. By digesting DNA with already ligated adapters, only the intact DNA will have adapters ligated at both ends and can be amplified over its entire length. The other DNA is much shorter, leading to step 2. After amplification, size selection is performed. Other steps such as quality control remain the same.
In what situations do you think this might be most useful?
One application is to combat antibiotic resistance, for example in urinary tract infections (UTIs). Ideally, clinicians would like to sensitively detect antibiotic resistance genes carried by a patient’s UTI strain, but urine samples also contain large amounts of host DNA and other bacteria. Currently, the best method for a UTI is to culture a urine sample, which takes three days to get results. This is not ideal. We want to build antibiotic resistance profiles quickly, within a day, so doctors can decide which antibiotics to give to their patients.
Another application is beneficial bacteria, or probiotics, such as: Bifidobacterium. Different strains can have very different health benefits. Discovering probiotics associated with human disease or drug responses requires performing some initial screening in stool samples to narrow down the species and recover more promising candidates.
Many people are interested in these applications, and we believe that mEnrich-seq provides a new, more sensitive, reliable, and cost-effective method to address these issues.
This interview has been condensed and edited for clarity.
References
- Cao L, et al. mEnrich-seq: Sequencing methylation-driven enrichment of bacterial taxa of interest in the microbiome.. Nat method. 2024;21(2):236-246.
- Beaulaurier J, et al. Metagenomic binning using DNA methylation and association of plasmids with bacterial host genomes. Nat Biotechnology. 2018;36(1):61-69.
