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1、1101 Environmental Technology, Vol. 26. pp 1101-1113 Selper Ltd., 2005 MICROBIAL COMMUNITY DYNAMICS IN MANURE COMPOSTS BASED ON 16S AND 18S rDNA T-RFLP PROFILES S. M. TIQUIA 115F Science Building, Department of Natural Sciences, The University of Michigan, Dearborn, MI 48128, USA (Received 20 March.
2、; Accepted 28 April 2005) ABSTRACT Compost processing is assumed to be related to the microbial communities present. However, methods that will evaluate these relationships are not well understood. In this study, terminal restriction fragment length polymorphism (T-RFLP) analysis was used to evaluat
3、e the diversity of PCR-amplified bacterial 16S and fungal 18S rDNA communities from manure composts at different stages of composting (initial day 0, thermophilic day 24, and mature day 104). Results showed that the bacterial and fungal community profiles changed over the composting process, with ba
4、cterial communities showing a higher diversity compared with the fungal communities. During the thermophilic stage (day 24), the diversity of the bacterial communities increased, while the fungal communities decreased. As the compost reached maturity (day 104), a reverse pattern was observed between
5、 the diversity of bacterial and fungal communities. That is, the 18S rDNA T-RFLP- based diversity indices increased, while the 16S rDNA T-RFLP-based diversity decreased. Differences in temperature profiles at different stages of composting impacted the chemical properties and the diversity of the mi
6、crobial communities. The day 104 compost (mature) had lower water, organic matter and C contents and higher C and OM loss compared with the day 0 (initial) and day 24 (thermophilic) composts, which affected the diversity of the microbial communities. The results presented here demonstrated that dist
7、inctive community patterns from manure composts could be rapidly generated using T-RFLP analysis. The succession of peaks in combination of increasing and decreasing peak heights at different stage of composting indicates the high potential of T-RFLP technique to monitor the dynamics of microbial co
8、mmunities, and their variation qualitatively and quantitatively. Keywords: T-RFLP, genetic diversity, principal components analysis, manure composting, compost maturity, organic matter INTRODUCTION Characterizing and understanding of microbial community structure and diversity may provide informatio
9、n needed to improve and evaluate compost processing and quality. While changes in physico-chemical properties (temperature, bulk density, cation-exchange capacity, C:N ratio, pH and organic C) during the composting process have been extensively studied 17, information on microbial communities during
10、 composting is underexplored. Moreoever, there is a need for a rapid method of assessment of microbial diversity during composting. Many early studies on the microbiology of composting dealt with the isolation and description of various microbes in compost using classical culture method 8-14. Since
11、this method cannot detect non-culturable species, there is a possibility that organisms which are difficult to isolate are predominant. Recently, molecular tools have been used to detect compost microbes independent of culture 15-17. Blanc et al. 15 used the amplified ribosomal DNA restriction analy
12、sis (ARDRA) to characterize thermotolerant populations such as Thermus strains and Bacillus-related bacteria isolated from composts. Kowalchuk et al. 16 characterized ammonia-oxidising bacteria in compost using denaturing gradient gel electrophoresis (DGGE) from reverse transcriptase-PCR (RT-PCR) am
13、plified 16S rRNA genes. Peters et al. 17 analyzed microbial community succession during composting at a broader phylogenetic level using single-strand-conformation polymorphisms (SSCP). Much of these works, however, were based on the organisms that could be found and utilized to speed up decompositi
14、on. Attempts have not been made to assess the diversity of the microbial communities at different stages of composting. In this study, terminal restriction fragment length polymorphisms (T-RFLP) of bacterial 16S and fungal 18S rDNA genes were used to characterize microbial community structure and di
15、versity in manure composts at different stages of composting. This analysis is based on the restriction endonuclease digestion of fluorescently end-labelled PCR products. The digested product is mixed with a DNA size standard, itself labelled with a distinct fluorescent dye, and the fragments are th
16、en separated by gel electrophoresis using 1102 an automated sequencer 18. Upon analysis, only the terminal end-labelled restrictrion fragments are detected. An electropherogram is produced, which shows a profile of compost microbial community as a series of peaks of varying height. This technique has been effectively used in the exploration of complex microbial environments 19-22. Bruce 19 exploited this technique to analyse the diversity of mercury resistance genes
