Studying the Symbiotic Bacterium in Individual, Living Nematodes Using Microfluidic Systems.

Animal-microbe symbioses are ubiquitous in nature and scientifically important in diverse areas, including ecology, medicine, and agriculture. nematodes and bacteria compose an established, successful model system for investigating microbial pathogenesis and mutualism. The bacterium is a species-specific mutualist of insect-infecting nematodes. The bacterium colonizes a specialized intestinal pocket within the infective stage of the nematode, which transports the bacteria between insects that are killed and consumed by the pair for reproduction. Current understanding of the interaction between the infective-stage nematode and its bacterial colonizers is based largely on population-level, snapshot time point studies on these organisms. This limitation arises because investigating temporal dynamics of the bacterium within the nematode is impeded by the difficulty of isolating and maintaining individual living nematodes and tracking colonizing bacterial cells over time. To overcome this challenge, we developed a microfluidic system that enables us to spatially isolate and microscopically observe individual, living nematodes and monitor the growth and development of the associated bacterial communities-starting from a single cell or a few cells-over weeks. Our data demonstrate, to our knowledge, the first direct, temporal, visual analysis of a symbiosis system and the application of this system to reveal continuous dynamics of the symbiont population in the living host animal. This paper describes an experimental system for directly investigating population dynamics of a symbiotic bacterium, , in its host-the infective stage of the entomopathogenic nematode . Tracking individual and groups of bacteria in individual host nematodes over days and weeks yielded insight into dynamic growth and topology changes of symbiotic bacterial populations within infective juvenile nematodes. Our approach for studying symbioses between bacteria and nematodes provides a system to investigate long-term host-microbe interactions in individual nematodes and extrapolate the lessons learned to other bacterium-animal interactions.