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This study focused on the capacity of finished compost, often used as packing material in biofiltration units, to support microbial biodegradation of trichloroethylene (TCE). Finished compost was enriched with methane or propane (10% head space) to stimulate cometabolic biodegradation of gaseous TCE. Successful hydrocarbon enrichment, as indicated by rapid depletion of hydrocarbon gas and measurable growth of hydrocarbon-utilizing micro-organisms, occurred within a week. Within batch reactor flasks, approximately 75% of head space TCE (1–40 ppmv) was rapidly sorbed onto compost material. Up to 99% of the remaining head space TCE was removed via biodegradation in compost enriched with either hydrocarbon. Hydrocarbon enrichment with methane or propane corresponded to 10-fold increases in methanotrophic or propanotrophic populations, respectively. Based on growth assessment under different nutritional regimes, there appeared to be complex metabolic interactions within the microbial community in enriched compost. Five separate bacterial cultures were derived from the hydrocarbon-enriched compost and assayed for the ability to degrade TCE.
During the past decade, composting technology has received considerable attention in two distinct areas: microbial ecology of the composting process and potential uses of the end product compost. The composting process is ideally a controlled microbial digestion utilizing indigenous bacteria, actinomycetes and fungi to decompose an organic substrate typically under aerobic conditions. During the composting process, thermophilic temperatures are achieved at various stages because metabolic heat production exceeds losses (Miller 1993). Observed patterns of microbial succession are influenced by the relative degree of decomposition of the organic matter as well as the changing temperatures within the system. Physical and biochemical characteristics of finished compost vary depending on the starting material and the specific operational parameters maintained during the composting process. However, certain generalizations can be made. Finished compost typically consists of porous organic material, with an increased density and a decreased C/N ratio, which is rich in diverse microflora and suitable for a wide variety of applications.
Finished compost has historically been utilized as organic fertilizer and to modify texture and water retention capacity of agricultural soils (Finstein & Morris 1975; Miller 1993). Compost has also been successfully used as an economically feasible substrate for mushroom cultivation and as packing material in odour-removing biofilters for treatment of wastes from food and other industries (Carlson & Leiser 1966; Bohn 1975; Leson & Winer 1991; Segall 1995). More recently, gaseous organic contaminants have also been successfully treated using compost-packed biofiltration units (Leson & Winer 1991).
Although strict methanotrophy appears to be limited to a few specific bacterial genera, there have been reports of apparent facultative methanotrophy, in which isolates can grow on methane or on other organic carbon sources, such as glucose (Zhao & Hanson 1984). Propanotrophy is considered to be more widespread in the microbial world than true methanotrophy, and many reports of facultative propanotrophy have been cited for bacterial and actinomycetes species (Perry 1980). Methane and propane consumption in mixed cultures from subsurface and other environmental samples has been reported, even in cases where pure cultures exhibiting methanotrophy or propanotrophy were not isolated (Fliermans et al. 1988; Phelps et al. 1990). As the starting material for compost typically consists of organic debris, which has often had close and prolonged contact with the soil environment, conditions supporting mixed methanotrophic or propanotrophic populations would not be unexpected in finished compost.
The current research was conducted to test the efficacy of stimulating TCE biodegradation in compost material with methane or propane enrichment, and to characterize the effects of key parameters, such as TCE concentration and hydrocarbon concentration, on the biodegradation process. Additionally, effects of hydrocarbon enrichment on the numbers and types of compost micro-organisms in compost were investigated, and bacterial cultures derived from enriched compost were assayed individually for TCE degradation capacity.
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- Materials and methods
Considerable research has focused on the stimulation of TCE biodegradation in soil by methane or propane. Wilson & Wilson (1985) first established the feasibility of methane enrichment for stimulating TCE removal in soils. Phelps et al. (1990) successfully enriched subsurface sediments with methane and propane under pulse feeding and continuous feeding conditions, and observed up to 90% TCE removal (20 ppm) within 5 d. These researchers found propane to be superior to methane as an enrichment gas, and considered this to be due to the higher energy value of propane vs methane. Conversely, Fliermans et al. (1988) enriched contaminated aquifer sediments with a variety of carbon sources, including methane and propane. They reported substantial TCE degradation (91% of 50 ppm within 3 weeks) in sediments spiked with 50 ppm TCE and enriched with methane, but negligible degradation with propane enrichment. Likewise, Lanzarone & McCarty 1990) were unable to enrich subsurface sediments with propane; propane was never consumed in their column reactor, and TCE was not removed as it was in methane-enriched reactors. They concluded that propanotrophic populations were non-existent in their sediment samples. In the current study, compost was successfully enriched with either propane or methane, but previous work demonstrated that certain compost materials are more amenable to this purpose than others (Sukesan & Watwood 1997). Enrichment with hydrocarbon gas is an excellent approach to consider for TCE remediation needs, but assessment of each system is obviously necessary to ensure suitability.
With respect to finished compost as a biofiltration matrix, both absorption and biological degradation contribute to initial TCE removal. It is likely that sorption is actually the initial step in the biodegradation process, as it increases contact between cells and chemical (Barrio-Lage et al. 1987). However, even an extremely sorptive matrix such as compost has a finite sorptive capacity which limits the amount of TCE which can be removed in this manner from a gaseous waste stream. Beyond this point biological degradation of the contaminant must proceed in order for removal to continue. It has been demonstrated that this process can be stimulated by enriching compost material with hydrocarbon gas (Watwood & Sukesan 1995; Sukesan & Watwood 1997). The current results confirm that exposure of compost material to propane or methane gas results in elevated populations of propanotrophs and methanotrophs, respectively. It is therefore considered that the stimulation of TCE removal is due to the cometabolic activity of propane monooxygenase and methane monooxygenase, respectively.
For such a complex matrix as finished compost, however, the situation may not be this straightforward. In this study, there was apparent overlap between methane, propane and complex carbon utilization in compost microbial populations. In separate TCE degradation experiments where the compost was first enriched with methane then switched to propane enrichment, there was a 6 d lag period, during which propane consumption and TCE removal were considerably lower than in compost continuously exposed to propane (data not shown). However, even during this lag period, low but measurable levels of propane and TCE depletion did occur. These observations indicate that methane enrichment may impact propanotrophic activity within the compost. Replica plating experiments revealed that significant numbers of propane-utilizing colonies could grow successfully on Higgins-methane medium, and vice versa. Many colonies originally grown on Higgins-methane or L-salts-propane medium could also grow on NA, although very few colonies could be successfully transferred from NA to Higgins-methane or L-salts-propane medium. This type of apparent metabolic overlap has been described by other researchers. For example, Brockman et al. (1994) found that 1% methane injections into subsurface sediments resulted in an elevated population of propanotrophs able to degrade TCE.
One likely explanation for the apparent metabolic overlap observed here and by other researchers is the existence of complex microbial interactions. In the current study, two of five cultures, derived from enriched compost and exhibiting apparent metabolic overlap, proved to be mixed. The tentative genera assigned to the other three cultures, based on fatty acid analysis, have never been cited as methanotrophic. This is difficult to reconcile with the fact that these cultures grew consistently on Higgins-methane medium. There have been several well publicized claims of new facultative methanotrophs (Patt et al. 1974; Lynch et al. 1980), which later proved difficult to verify and in certain cases, were recanted (Lidstrom-O’Connor et al. 1983). Dalton & Leak (1985) therefore recommended caution when asserting new claims of facultative methanotrophy, and no such claim is made here. Rather, based on the unusual metabolic patterns exhibited by these cultures and the apparent metabolic discrepancies in identification, it appears that each of the five compost-derived cultures was mixed, although the methods employed in this study could not separate the components. It is likely that complex interactions between members in the mixed cultures prevented their separation.
This type of complexity has frustrated other researchers in this field. Fliermans et al. (1988) observed TCE degradation in methane and propane enrichment cultures from subsurface sediments, but they were unable to establish stable methanotrophic enrichment cultures and could not isolate methanotrophic TCE degraders in pure culture. These researchers theorized that methane enrichment might serve to enrich the general microbial community slowly, promoting diverse populations capable of tolerating and degrading TCE.
It is also important to note in the current study that the separate cultures derived from enriched compost exhibited substantially lower levels of TCE biodegradation than those exhibited by the consortium present in the compost material. When taken together, these observations suggest that microbial populations present in enriched compost participate in intricate, and perhaps obligatory, metabolic interactions. In this system, the complexity of the microbial community may contribute significantly to cometabolic TCE removal; syntrophic, commensalic or symbiotic associations may result in conversions that are not feasible for separate cultures.
Another issue of interest is the optimum hydrocarbon concentration to target in the treatment system concomitant to contaminant introduction. In theory, some minimal hydrocarbon concentration must be maintained, even after successful enrichment, to ensure continuous induction of the appropriate cometabolic enzymes. However, high hydrocarbon concentrations could theoretically result in substrate competition which could lead to decreased TCE degradation. An additional factor to consider for actual treatment systems is the expense associated with the hydrocarbon supply. In the current study, hydrocarbon enrichment was routinely performed with 10% hydrocarbon, based on previous results, but different hydrocarbon concentrations were assessed during TCE degradation experiments. The results indicate that for methane and propane, TCE degradation in previously enriched compost was independent of hydrocarbon concentration within the range of 0·2 to 2·0%. This result contrasts with a report by Strandberg et al. (1989). This group studied TCE removal in a fixed film bioreactor infused with aqueous growth medium and methane and inoculated with an enrichment culture from contaminated subsurface material. While they found no evidence of substrate competition between methane and TCE with atmospheric methane concentrations between 4 and 20%, when methane was reduced to 2%, TCE biodegradation efficiency declined, presumably due to a lack of sustained induction of methane monooxygenase. These divergent results may not be totally unexpected for two such different systems, again highlighting the difficulty inherent in extrapolation of results between systems.
More research is needed to address specific aspects of biofiltration microbiology. While it is not surprising that such a complex process will vary between systems and materials, it is extremely important that researchers and practitioners continue to report observations and results with compost packed systems so that this promising remediation technology can be further improved and refined. Critical to advancing this type of technology is a more thorough understanding of the microbial ecology and metabolic constraints within the biofiltration matrix.