Cultivation of unculturable soil bacteria

Despite the abundance of bacterial species in soil, more than 99% of these species cannot be cultured by traditional techniques. In addition, the less than 1% of bacteria that can be cultured are not representative of the total phylogenetic diversity. Hence, identifying novel species and their new functions is still an important task for all microbiologists. Cultivating techniques have played an important role in identifying new species but are still low-throughput processes. This review discusses the issues surrounding cultivation, including achievements, limitations, challenges, and future directions. Despite the abundance of bacterial species in soil, more than 99% of these species cannot be cultured by traditional techniques. In addition, the less than 1% of bacteria that can be cultured are not representative of the total phylogenetic diversity. Hence, identifying novel species and their new functions is still an important task for all microbiologists. Cultivating techniques have played an important role in identifying new species but are still low-throughput processes. This review discusses the issues surrounding cultivation, including achievements, limitations, challenges, and future directions. the 16S rRNA gene is used for phylogenetic studies due to highly conserved sequences between different species of bacteria and archaea, and also hypervariable regions as species-specific signature sequences. There are two types of methodologies, 16S rRNA gene sequencing and ribotyping. 16S rRNA gene sequencing has become prevalent in microbiology because it provides a rapid accurate alternative to phenotypic methods of bacterial identification. Ribotyping is the fingerprinting of genomic DNA restriction fragments of the 16S rRNA genes. a FISH method that enhances the signal through conjugation of an enzyme peroxidase to the specific nucleic acid probe instead of a fluorescent dye. It is useful in phylogenetic studies of prokaryotes that may be small, slow growing, or starving bacteria. a method to analyze a clone library; a collection of DNA fragments that is stored and propagated in a population of microorganisms through the process of molecular cloning. a plot of the number of visible colonies on a plate against incubation time. a technique to detect differences in the melting behavior of small DNA fragments (200–700 base pairs) that differ by as little as a single base substitution. The separated bands on the gel are located according to the concentration gradient of denaturants. a cytogenetic technique that is used to detect and localize the presence or absence of specific DNA sequences on chromosomes. a strategy to allow growth of targeted microbes through highly selective media based on two selective agents, a carbon source and an antimicrobial. a method used for sequencing long DNA strands. Because DNA sequencing can only be used for relatively short strands (100–1000 base pairs), longer sequences must be subdivided into smaller fragments, and subsequently re-assembled to give the overall sequence. a method based on the incorporation of 13C-labeled substrate into cellular biomarkers such as nucleic acids, separation of labeled from unlabeled nucleic acids by density gradient centrifugation, and molecular identification of active populations carrying labeled nucleic acid.