In nature, microbial communities contain several species of bacteria. It is therefore difficult to isolate individual species of bacteria and cultivate them in the laboratory. Researchers have now developed new tools to help. These tools allow researchers to genetically manipulate distinct bacterial species within their communities. One of the tools, called ET-Seq, allows scientists to identify microbes that can be genetically modified directly within mixed communities. The other tool, dubbed DART, allows scientists to test the function of genes of specific species within their communities.
Most bacteria live together in complex communities. With most current tools, scientists must isolate individual species in order to study them. However, environmental microbes such as soil bacteria cannot be easily isolated and cultured in the laboratory. Moreover, the behavior of microbial communities results from the combined contributions of its members. This is very different from the activity of isolated species. Developments in ET-Seq and DART tools help scientists study microbial communities without the need to isolate different members. By combining the two technologies, researchers can also track genetic changes as the community grows and examine the function of genes in microorganisms that cannot be cultured in the lab.
Culture and genetic analysis have been the main means used to study the function of genes and the behavior of microbes. However, these classical approaches require the isolation and cultivation of microorganisms in the laboratory. This severely limits scientists’ knowledge of microbes that cannot yet be cultured in the lab. Also, the interactions that occur between microbes when they grow together in a community cannot be studied in isolated organisms. To address these challenges, this research has developed two technologies to test functions and interactions directly in laboratory microbiomes. Environmental transformation sequencing (ET-seq) delivers a mobile genetic element (transposon) into a microbial community. The transposon inserts itself randomly into the genes of certain bacterial species. By sequencing the genomes of all the microbes in the community, scientists can detect which members of the community are transformed by the transposon and how often. In this way, they identify genetically treatable species. These species can be specifically targeted for manipulation of selected genes using the all-in-one RNA-guided CRISPR-Cas transposase (DART). Researchers also combined the two techniques to demonstrate enrichment of targeted bacterial species, confer new metabolic traits, and measure the genetic fitness of bacteria in a community laboratory setting. These new capabilities will provide important new insights into the activities of uncultivated microbes and the functions of key genes, metabolites and proteins, for example, in the soil carbon cycle and mediating beneficial microbial interactions with plants for sustainable bioenergy. .
Funding was provided by the m-CAFE Microbial Community Analysis & Functional Evaluation in Soils Science Focus Area, which is led by Lawrence Berkeley National Laboratory and supported by the Department of Energy Office of Science, Office of Biological & Environmental Research. This research was also developed with funding from the Defense Advanced Research Projects Agency. This material is based on work supported by the National Science Foundation. Support was also provided by the Innovative Genomics Institute at the University of California, Berkeley.
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