Abstract
Antibiotics from few culturable microorganisms have saved millions of lives since the 20th century. But with resistance formation, these compounds become increasingly ineffective, while the majority of microbial and with that chemical compound diversity remains inaccessible for cultivation and exploration. Culturing recalcitrant bacteria is a stochastic process. But conventional methods are limited to low throughput. By increasing (i) throughput and (ii) sensitivity by miniaturization, we innovate microbiological cultivation to comply with biological stochasticity. Here, we introduce a droplet-based microscale cultivation system, which is directly coupled to a high-throughput screening for antimicrobial activity prior to strain isolation. We demonstrate that highly parallelized in-droplet cultivation starting from single cells results in the cultivation of yet uncultured species and a significantly higher bacterial diversity than standard agar plate cultivation. Strains able to inhibit intact reporter strains were isolated from the system. A variety of antimicrobial compounds were detected for a selected potent antibiotic producer.
Highlights
The microbial diversity in environmental habitats provides a rich resource for medically relevant substances [1] and it is far from being exhaustively mapped [2]
We were able to screen for antibiotic production, which demanded the addition of reporter strains to the cultivation compartments
One major advantage of our method compared to previous microscale cultivation systems lies in the space efficient incubation of droplets in bulk
Summary
The microbial diversity in environmental habitats provides a rich resource for medically relevant substances [1] and it is far from being exhaustively mapped [2]. Elucidation and application of appropriate growth conditions are required for cell replication of species that are recalcitrant to cultivation. To narrow the gap between the naturally occurring complexity of microbial communities and the limited diversity of culture collections, several new techniques have been developed. Other techniques mimic the conditions in situ in more detail by supplying extracts derived from the environment containing unspecified mixtures of macro- and micronutrients and other growth factors [4, 10,11,12]. The most advanced development in this direction is to provide direct contact with the environment [13,14,15,16], which proved highly productive in culturing species regarded as unculturable
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