Next-Generation Therapy Analyzes Intestinal Phages, Propelling Future of C. difficile Treatment


Using metagenomic information, the technology isolates phages whose enzymes can specifically target the bacteria that causes C. difficile infection.

Next-generation sequencing is improving genome analysis technology to make phage therapy more efficacious against Clostridioides difficile (C. difficile), according to findings published in the journal Frontiers. With advancing technology, phage therapy could become more effective than fecal microbiota transplantation (FMT), the most effective therapy to defend against C. difficile infection (CDI) thus far.

Image credit: luchschenF |

Image credit: luchschenF |

“Scientific research on bacteriophages has become a hot topic because of the impending problem of multidrug-resistant bacteria,” the study authors wrote in the paper. “Given the versatility of phage therapy for CDI, there is a need for a high-throughput system that can identify phages that infect C. difficile.”

C. difficile is a Gram-positive, spore-forming anaerobic and pathogenic bacterium that colonizes the gut and causes mild to severe nosocomial diarrhea. Patients who contract C. difficile are initially given antibiotic treatment.

However, due to the overuse of antibiotics and the nature of the disease, many strains of C. difficile have become multidrug-resistant and disinfectant-resistant, resulting in a growing number of patients having relapsed CDI. This also makes the disease harder to treat, more able to spread, and even more dangerous.

This review looks at recent findings on phage therapy for CDI. With many types of phages having been identified, it’s necessary to understand which ones could be beneficial for the treatment of CDI. This means having the ability to isolate phages that can control pathobionts, which contribute to the pathogenesis of CDI.

In some parts of the world, phage (bacteriophage) therapy was developed as an alternative treatment to antibiotics, which are becoming less effective against resistant disease. With traditional phage therapy, host bacteria are collected from healthy human fecal samples. A plaque assay is used to isolate phages for therapy.

But next-generation phage therapy takes it a step further—the host bacteria can actively target pathobionts. Using next-generation sequencing, researchers can also collect intestinal bacterial and viral metagenomic information from these human fecal samples. Doing so can help identify the phage-derived antibacterial enzymes that target specific host bacteria.

Phages encode the enzyme endolysin, which can infect Gram-positive C. difficile, degrading the envelope surrounding the bacteria (peptidoglycans) to destroy its biofilm. There are 4 N-terminal enzyme active domains (EAD): amidases, acetylmuramidases, endopeptidases, and glucosaminidases, with amidases and muramidases most commonly able to target these peptidoglycans.

“To isolate phages used in phage therapy, it has been necessary to isolate and culture target bacteria,” the study authors wrote.

And because the phages are isolated from culture bacteria in the intestine, an organ that contains a vast and diverse microbiota, investigators anticipate that this therapeutic technique can be used for other targeted bacteria.

“In the near future, phage science will be developed further by integration with a wide range of fields including medicine, microbiology, bioinformatics, and synthetic biology,” the study authors concluded.


Fujimoto K, Uematsu S. Phage therapy for Clostridioides difficile infection. Front. Immunol. Vol 13;2022. DOI: 10.3389/fimmu.2022.1057892

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