In composting, heterotrophic microbial activity and growth lead to the degradation of organic material. This degradation occurs via the formation of complex microbial communities that work in a delicate balance to drive changes in the temperature and pH of the compost pile. Although composting is an ancient method, it has evolved as a useful method for the reduction of municipal solid wastes and for the destruction of potentially hazardous pathogenic organisms. Composting of the agricultural wastes produces a biologically stable humic substance that may be used as a soil additive, thereby efficiently reducing the amount of waste generated.
Composting mixture of Switchgrass and dog food. Compost appears to be covered with fungi.
The complexity of the interactions among the microbial populations present in a compost pile makes the process a perfect habitat for studying the succession and dynamics of a microbial ecosystem. These interactions provide significant insight into the selection, adaptation and exchange of materials that facilitate the dominance of certain taxonomic groups at different stages in the process. Advances in molecular ecology methods such as probe hybridizations and genetic fingerprinting methods provide the tools necessary for monitoring the dynamics of microbial communities.
Probe hybridization is a molecular biological method that does not require the use of PCR, and is available for providing information about the microbial community in a compost sample. In probe hybridizations, a labeled fragment of a known DNA sequence is hybridized to its complimentary strand in an environmental sample. The labeling may be done via radioactivity or chemiluminescence to detect and visualize the presence of specific species in the microbial community. According to Spiegelman et al. (5), there are 3 main reasons for using oligonucleotide probes: (i) to investigate the presence of various taxonomic groups in the community, (ii) to measure the relative abundance of specific taxa, and (iii) to determine the spatial distribution of species or groups present. Probe hybridizations may be used (i) with clone libraries in the identification of groups of interest (4), (ii) along with other fingerprinting techniques (4) and (iii) directly in order to investigate the presence of taxonomic groups.
Capillary electrophoresis (CE) is a method used to separate a variety of molecules including organic acids, inorganic ions, amino acids, proteins and nucleic acids based on their charge and frictional forces (2). CE has also found applications in protein/drug and protein/DNA interactions and other physiochemical studies (2). In molecular biology, CE has emerged as a technique to meet the demand for high-throughput analysis of DNA, and consequently, of microbial communities (1). This technique may be coupled with many of the traditional molecular biology tools used to analyze microbial communities. One of the most common methods used is temperature gradient capillary electrophoresis (TGCE) which is the capillary version of temperature gradient gel electrophoresis (TGGE). TGGE is used to separate rDNA in a polyacrylamide gel containing a temperature gradient based on differences in G-C content of the sequences (3). As the DNA moves along the gel, the increased thermal gradient forces it to become single stranded, but it does not become completely denatured because of the presence of a GC clamp that is incorporated into one of the primers for the PCR amplification. Compost research completed using TGGE has produced microbial diversity data and identification when coupled with 16S rDNA sequencing.
Another genetic fingerprinting method that may be coupled with capillary electrophoresis is the ARISA (Automated Ribosomal Intergenic Spacer Analysis) method which creates a community profile of the PCR-amplified genetic region between 16S rRNA and 23S rRNA on the basis of species-specific length polymorphisms in the region (5). Coupling ARISA with capillary electrophoresis increases resolution and analytic power.
Separation spectra of compost sample using ARISA method with capillary electrophoresis.
(1) Hong, H., A. Pruden, and K. F. Reardon. 2007. Comparison of CE-SSCP and DGGE for monitoring a complex microbial community remediating mine drainage. Journal of Microbiological Methods 69:52-64.
(2) Issaq, H. J. 2002. Thirty-five years of capillary electrophoresis: Advances and perspectives. Journal of Liquid Chromatography & Related Technologies 25:1153-1170.
(3) Muyzer, G. 1999. Presented at the International Symposium on Microbial Ecology, Halifax, Canada.
(4) Schloss, P. D. 2002. Quantitative Molecular Analysis of Microbial Succession in Compost and its Ramifications on Process Reproducibility. Dissertation. Cornell University, Ithaca.
(5) Spiegelman, D., G. Whissell, and C. W. Greer. 2005. A survey of the methods for the characterization of microbial consortia and communities. Canadian Journal of Microbiology 51:355-386.