Abstract
Most bacteria live in biofilms in their natural habitat rather than the planktonic cell stage that dominates during
traditional laboratory cultivation and enrichment schemes. The present study describes the establishment of a fow-based enrichment method based on multispecies biofilm communities for directing bioflm functionality using an
environmental inoculum. By controlling fow conditions and physio-chemical properties, the set-up aims to simulate
natural conditions ex situ for bioflm formation. The functionality of the method was demonstrated by enrichment of
bioflm microbiomes using consortia from a warm compost pile and industrial waste materials as growth substrate,
and further exploring the metagenomes by biotechnological tools. The 16S rRNA gene sequencing results revealed a
diference in consortium composition and especially in genus abundance, in fow experiments compared to tradi‑
tional liquid-shake experiments after enrichment, indicating good bioflm development and increased abundance of
bioflm-forming taxa. The shotgun sequence mining demonstrated that diferent enzymes classes can be targeted by
enriching bioflms on diferent substrates such as oat husk, pine saw dust, and lignin. The fow-based bioflm method
is efective in reducing bacterial consortia complexity and in selecting biofilm-forming bacteria, and it is possible to
enrich the biofilm community in various directions based on the choice of sample material, environmental conditions,
and nutritional preferences, targeting enzymes or enzyme classes of industrial interest.
traditional laboratory cultivation and enrichment schemes. The present study describes the establishment of a fow-based enrichment method based on multispecies biofilm communities for directing bioflm functionality using an
environmental inoculum. By controlling fow conditions and physio-chemical properties, the set-up aims to simulate
natural conditions ex situ for bioflm formation. The functionality of the method was demonstrated by enrichment of
bioflm microbiomes using consortia from a warm compost pile and industrial waste materials as growth substrate,
and further exploring the metagenomes by biotechnological tools. The 16S rRNA gene sequencing results revealed a
diference in consortium composition and especially in genus abundance, in fow experiments compared to tradi‑
tional liquid-shake experiments after enrichment, indicating good bioflm development and increased abundance of
bioflm-forming taxa. The shotgun sequence mining demonstrated that diferent enzymes classes can be targeted by
enriching bioflms on diferent substrates such as oat husk, pine saw dust, and lignin. The fow-based bioflm method
is efective in reducing bacterial consortia complexity and in selecting biofilm-forming bacteria, and it is possible to
enrich the biofilm community in various directions based on the choice of sample material, environmental conditions,
and nutritional preferences, targeting enzymes or enzyme classes of industrial interest.