Microbe observation and cultivation array (MOCA) for cultivating and analyzing environmental microbiota

Weimin Gao,Deirdre R Meldrum,Jared Naimark,Shih-Hui Chao,Dena Navarroli,Weiwen Zhang

doi:10.1186/2049-2618-1-4

Abstract

BackgroundThe use of culture-independent nucleic acid techniques, such as ribosomal RNA gene cloning library analysis, has unveiled the tremendous microbial diversity that exists in natural environments. In sharp contrast to this great achievement is the current difficulty in cultivating the majority of bacterial species or phylotypes revealed by molecular approaches. Although recent new technologies such as metagenomics and metatranscriptomics can provide more functionality information about the microbial communities, it is still important to develop the capacity to isolate and cultivate individual microbial species or strains in order to gain a better understanding of microbial physiology and to apply isolates for various biotechnological applications.ResultsWe have developed a new system to cultivate bacteria in an array of droplets. The key component of the system is the microbe observation and cultivation array (MOCA), which consists of a Petri dish that contains an array of droplets as cultivation chambers. MOCA exploits the dominance of surface tension in small amounts of liquid to spontaneously trap cells in well-defined droplets on hydrophilic patterns. During cultivation, the growth of the bacterial cells across the droplet array can be monitored using an automated microscope, which can produce a real-time record of the growth. When bacterial cells grow to a visible microcolony level in the system, they can be transferred using a micropipette for further cultivation or analysis.ConclusionsMOCA is a flexible system that is easy to set up, and provides the sensitivity to monitor growth of single bacterial cells. It is a cost-efficient technical platform for bioassay screening and for cultivation and isolation of bacteria from natural environments.

Highlights

The use of culture-independent nucleic acid techniques, such as ribosomal RNA gene cloning library analysis, has unveiled the tremendous microbial diversity that exists in natural environments
We have found that bacterial growth across the droplet array has a high level of uniformity when the initial cell density is more than 10 cells/μl, and microbe observation and cultivation array (MOCA) provides a novel platform for bioassay screening
Bacterial growth in droplets Because MOCA droplet settings create a different physical environment from that of bulk cultures, this may influence many of the growth factors required by the bacteria [27,28]

Summary

Introduction

The use of culture-independent nucleic acid techniques, such as ribosomal RNA gene cloning library analysis, has unveiled the tremendous microbial diversity that exists in natural environments. In sharp contrast to this great achievement is the current difficulty in cultivating the majority of bacterial species or phylotypes revealed by molecular approaches. During the past over two decades, the use of cultureindependent nucleic acid techniques, represented by ribosomal RNA gene cloning library analysis, has unveiled the tremendous microbial diversity that exists in natural environments [1]. Knowledge obtained through metatranscriptome analysis has been used in directed cultivation of bacteria [12] These examples shown that many microbial species can be cultured as long as the environments are optimized for growth

Methods

Results

Conclusion