Saccharibacteria deploy two distinct Type IV pili, driving episymbiosis, host competition, and twitching motility

All cultivated Patescibacteria, or CPR, exist as obligate episymbionts on other microbes. Despite being ubiquitous in mammals and environmentally, molecular mechanisms of host identification and binding amongst ultrasmall bacterial episymbionts are largely unknown. Type 4 pili (T4P) are well conserved in this group and predicted to facilitate symbiotic interactions. To test this, we targeted T4P pilin genes in Saccharibacteria Nanosynbacter lyticus strain TM7x to assess their essentiality and roles in symbiosis. Our results revealed that N. lyticus assembles two distinct T4P, a non-essential thin pili that has the smallest diameter of any T4P and contributes to host-binding, episymbiont growth, and competitive fitness relative to other Saccharibacteria, and an essential thick pili whose functions include twitching motility. Identification of lectin-like minor pilins and modification of host cell walls suggest glycan binding mechanisms. Collectively our findings demonstrate that Saccharibacteria encode unique extracellular pili that are vital mediators of their underexplored episymbiotic lifestyle.