Microbes self-organize in microcolonies at solid surfaces while transitioning to a sessile form within a protective biofilm matrix. Microbes also have complex community dynamics at fluid interfaces. While the biological implications of surface-attached and interfacial biofilms for the environment, health, and industry are widely appreciated, the earlier developmental stage of microbes as microcolonies has received scant attention. This presentation elucidates two new approaches to investigate microbial dynamics in spatially and interfacially confined microsystems. First, a new approach to studying microcolony formation and community dynamics is described. Using microfluidics-enabled fabrication, a nanoliter-scale sessile culture system (the nanoculture) is designed to grow synthetic microbial communities. Each nanoculture begins as a several nanoliter droplet of suspended cells, encapsulated by a polydimethylsiloxane (PDMS) membrane. The physicochemical properties of the encapsulation materials allow the diffusion of functional probes to interrogate cell physiology under chemical insults, allowing microbial interactions to be probed within or across the confining vessel. We use this versatile platform to investigate bacterial-fungal (inter-kingdom) dynamics that play a central role in early childhood dental caries and many infections. Second, microbial response to confinement at fluid-fluid interfaces are studied both in terms of physico-chemical effects and metabolic implications. We study two strains of P. aeruginosa, PAO1 and PA14. The PAO1 cells remodel the hexadecane-water interface to form highly elastic Films of Bacteria at Interfaces (FBI), i.e. elastic, solid films of bacteria and excreted polysaccharides, whereas the PA14 cells form active FBI that feature interface-associated microbes that remain highly motile. Transcriptional profiles of the interfacially confined strains suggest that the elastic FBI provides protection, in a manner akin to biofilms, enabling cells to cope with the detrimental effects of the interfacial environment. Together, these studies provide a basis for new strategies to minimize the deleterious impacts and to optimize the beneficial effects of microbial communities relevant to the environment and health. The nanoculture system and FBI-encapsulated droplets can also be exploited in upstream bioprocessing technologies, with uses ranging from the encapsulation of beneficial microbial communities to high-throughput screening of bioactive molecules.