Bacterial Community Structure During Yard Trimmings Composting

Abstract

A long-term objective of our group is to understand how various composting parameters affect microbial community structure in composts. In this study, we used terminal restriction fragment length polymorphisms (T-RFLP) of PCR-amplified 16S rRNA genes to analyze bacterial community structure during the composting of yard trimmings. Community DNA was isolated from samples collected on days 0, 8, 29, 64, and 136 from a compost windrow (consisting of leaves, grass, and brush in a 4:2:1 ratio) at a large-scale municipal facility. The DNA was PCR-amplified using fluorescently labeled primers targeted to bacterial domain 16S rRNA genes. The products were restriction-digested with Hhal, Mspl,and Rsal to give fingerprints of the bacterial communities. Terminal restriction fragment (TRF) sizes obtained with the three digestions were compared to the three fragments determined by computer-simulated amplification and restriction digestions of complete 16S rRNA gene sequences. T-RFLP patterns indicated extensive bacterial diversity in all of the composts. A large percentage of the observed TRFs corresponded to sizes predicted for bacteria by computer-simulated digestion. Comparison of fragment sizes from three digestions to those predicted by computer-simulated digestions indicated a substantial shift from a bacterial community containing primarily Gram-negative $α$, $β$, and $γ$ Proteobacteria (day 0) to communities containing many members of the Gram-positive Bacillus-Clostridium group (days 8, 29, and 64) and members of the CFB and Actinobacteria (days 29 and 64). Bacterial species identified on days 8, 29, and 64 included those previously isolated from thermophilic composts by cultivation such as Bacillus and Pseudomonas spp. as well as many not previously described in composts. Abundant TRFs corresponding to E. coli and other Gram-negative $γ$ Proteobacteria, decreased dramatically after the first 8 days of composting. The day-136 composts contained a diverse group of bacteria including many fragment sizes consistent with known Pantoea and Pseudomonas biocontrol agents as well as Xanthomonas and Bacillus species. The greatest diversity of bacteria was observed in the stabilized day 64 and 136 composts where 115 and 111 TRFs corresponding to members of 7 and 6 different phylogenetic groups, respectively, were observed.