The microbial ecology of novel horizontal flow biofilm reactors (HFBRs) used to treat methane, hydrogen sulphide and ammonia contaminated airstreams at 10°C
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Ammonia (NH3), hydrogen sulphide (H2S) and methane (CH4) are three problematic gaseous emissions that are regularly encountered in agricultural, industrial and municipal waste sectors. Methane is an important greenhouse gas, while hydrogen sulphide and ammonia are both highly toxic and odorous. It is therefore essential that these three gases are treated before being released into the atmosphere to prevent environmental damage. During this body of work the three gases were biologically treated using a novel Horizontal Flow Biofilm Reactor (HFBR) design at 10°C. The physicochemical parameters and the microbial ecology of the HFBRs were investigated to determine gas removal processes and to link microbial community structure to reactor performance. Four separate studies were conducted over the course of this work; one ammonia, one hydrogen sulphide and two methane trials. In the ammonia trial, three HFBRs were operated to treat an ammonia-contaminated airstream at 10°C for 90 d. Average removal efficiencies of 99.7% were achieved at loading rates of 4.8 g NH3 m3 h-1. Biological nitrification of the ammonia to nitrite (NO2-) and nitrate (NO3-) was performed by nitrifying bacterial and archaeal biofilm communities. Ammonia Oxidising Bacteria (AOB) were more abundant than Ammonia Oxidising Archaea (AOA) throughout the depths of the HFBRs. AOB from the Nitrosomonas and Nitrosospira genera were the dominant bacterial clones in the HFBRs, while an uncultured archaeal clone dominated the AOA community. The only Nitrite Oxidising Bacteria (NOB) species identified in the HFBRs was closely related to Candidatus nitrotoga arctica. The overall bacterial community structure between the HBFRs was highly conserved, although variations in community structure occurred between zones in the HFBRs. This study demonstrated that HFBRs are a suitable biotechnology for the treatment of ammonia contaminated airstreams at low temperatures and identified the key nitrifying microorganisms driving the removal process. During the hydrogen sulphide trial, three HFBRs were tested for the removal of H2S gas from air streams over a 180-day trial at 10°C. Removal rates of up to 15.1 g H2S m-3 h-1 were achieved during the trial. Bio-oxidation of H2S in the reactors led to the production of H+ and sulphate (SO42-) ions resulting in acidification of the liquid phase. Reduced removal efficiency was observed when a loading rate of 15.1 g H2S m-3 h-1 was applied to the HFBRs. The provision of additional NaHCO3 in the liquid nutrient feed (LNF) during maximum H2S loading rates, led to decreased liquid phase acidity and improved H2S removal. The bacterial diversity within the HFBRs was low and the community was dominated by two species from the genus Acidithiobacillus and Thiobacillus. The harsh environmental conditions present in the HFBRs were likely responsible for the lack of bacterial diversity. Depth-resolved genetic fingerprinting of the bacterial communities in the HFBRs using Temperature Gradient Gel Electrophoresis (TGGE), revealed differences in the community structure between zones in the reactors. The variation in bacterial community composition between zones was influenced by alkalinity, pH and SO4 concentrations. In spite of the low operating temperature, the results of this study indicate that HFBRs have excellent potential to biologically treat H2S contaminated airstreams. Two separate methane trials were carried out to treat methane contaminated airstreams using HFBRs at 10°C. In the first methane trial three HFBRs were operated to treat methane contaminated airstreams at low concentrations (1.12%) for 233 d. Removal rates of up to 7.1 g CH4 m-3 h-1 were achieved during the trial, demonstrating that HFBRs are a suitable technology for the treatment of methane contaminated airstreams at low temperatures. Methane removal rates were influenced by temperature, with reduced removal rates observed during a cold period when temperature fluctuated between 1-10°C (Q10 2.49 ±0.45). The composition of the LNF applied to the HFBRs impacted on methane removal rates. Increased removal was observed when organic carbon was omitted from the LNF, while the impact of nitrogen source and concentration was unclear. Terminal Restriction Fragment Length Polymorphism (TRFLP) fingerprinting of the bacterial communities in the HFBRs identified a diverse and dynamic bacterial population which varied with depth in the reactors and over time. Fluorescent in-situ Hybridisation (FISH) of the methanotrophic 16S rRNA genes indicated that Type II methanotrophs appeared to be more abundant in the HFBR biofilm than Type I methanotrophs, although the microbial community was dominated by other prokaryotes. Results from this study showed that HFBRs were capable of treating methane contaminated airstreams and that the process was facilitated by a diverse and dynamic bacterial population. In the second methane trial three HFBRs were operated at 10°C for 341 days treating methane-contaminated airstreams at low concentrations (1.6%, v/v). Removal rates of up to 8.2 g CH4 m-3 h-1, with removal efficiencies of 62.1%, were achieved at a loading rate of 13.2 g CH4 m-3 h-1. Maximum removal rates were observed following the addition of silicone oil and Brij 35 to the LNF. Silicone oil addition improved removal efficiencies by an average of 69% across the three HFBRs. The further addition of Brij35 to R1 and R2 resulted in increased methane removal rates of 33%, when compared to operation with silicone oil alone. The methane oxidation potential (MOP) of the HFBR biofilm and seed biomass was assessed in batch incubations at various temperatures. The highest MOP rate (19.0 mg CH4 g[VSS] h-1) for the HFBR biofilm was measured at 23°C, whereas highest oxidation rates in the seed biomass were measured at 37°C (33.6 mg CH4 g[VSS] h-1). Methanotrophs were present in abundance at all depths in the HFBRs based on the quantification of the functional pmoA gene using qPCR. TGGE fingerprinting and sequencing indicated that overall methanotroph diversity in the HFBRs was low with Type I Methylobacter and Methylomonas species, and Type II Methylocystis species, detected. The methanotroph communities between the three HBFRs was similar but changed over the duration of the trial. Results from this study showed that silicone oil and Brij35 improved methane removal in the HFBRs and that methane oxidation in the reactors was performed by a small group of methanotrophs who were abundant at all depths in the reactors. Results from across the four trials showed that HFBRs are a suitable biotechnology for the treatment of ammonia, hydrogen sulphide and methane contaminated airstreams at low temperatures. Microbial community structure and function in the HFBRs varied with depth in the HFBRs and was impacted by a number of different environmental parameters. In the future, manipulation of the physical and chemical environment in HFBRs to support the development of desired microbial communities can be implemented to improve reactor performance and operation.