Research
Microorganisms possess an incredible variety of metabolisms. They are extremely versatile and capable of using many different kinds of “food” while being able to survive in extreme environments. Broadly speaking, my research is focused on understanding, characterizing and harnessing these metabolisms to solve environmental issues.
Current Research
Optimizing Phosphorus Removal from Wastewater
Enhanced Biological Phosphorus Removal (EBPR) is a process which utilizes the metabolic capability of PAOs to release P during anaerobic periods and uptake the P and store it as polyphosphate during aerobic periods to remove P from wastewater. Since PAOs are critical to the performance of this process, understanding their metabolism can help optimize the performance and robustness of the process. Most importantly, PAOs require a sufficient amount of polyhydroxyalkanoates (PHA) in the aerobic process to be able to remove P effectively. This requires a sufficient supply of carbon (in the form of volatile fatty acids (VFAs)) in the anaerobic period. A major issue with the typical EBPR process is that it can have unstable performance when the influent wastewater has low/unreliable carbon content. A lot of wastewater resource recovery facilities (WRRFs) face this issue.
So an alternate process configuration…
Side-Stream EBPR
Since influent carbon is an important factor, why not produce it internally in the process? Side-Stream EBPR (S2EBPR) incorporates an anaerobic fermentation reactor in the side-stream to ferment settled sludge from the secondary clarifier (also called return activated sludge (RAS)). In this side-stream reactor, VFAs are produced by fermenting microorganisms which is then simultaneously used by PAOs to form PHA and release P. We have found that the extended time that PAOs spend in the VFA-rich anaerobic side-stream reactor ensures an adequate supply of PHA in their cells for subsequent P uptake in the aerobic reactor. This enables influent carbon-independent phosphorus removal thus ensuring the performance and stability of the process. I am working on understanding the fundamental mechanisms that govern the performance of S2EBPR along with the microbial ecology and dynamics in this process.
Polyphosphate Accumulating Organisms (PAOs)
Since PAOs are critical to removing P from wastewater, understanding the type of PAOs, their metabolism and their activity is important to optimizing the process. So far, two types of PAOs have been implicated in the EBPR process - Candidatus (Ca.) Accumulibacter (hereafter referred to as Accumulibacter) and Tetrasphaera spp. Other organisms such as Ca. Obscuribacter and Ca. Accumulimonas have also been proposed as putative PAOs, even though their relevance to the EBPR process is still unknown. My current research looks at the ecology of PAOs in WRRFs performing both typical EBPR and S2EBPR to elucidate differences in microbial ecology. PAOs also rely on fermenting organisms to breakdown complex organic matter into VFAs, so understanding the inter-species dynamics between these functional groups is also important. I am using a combination of 16s rRNA gene sequencing and genome-resolved metagenomics to characterize the microbial ecology and statistical techniques to elucidate differences. I am also using comparative genomics to understand the evolution of the PAO metabolism which can lead to development of robust markers and identification of other putative PAOs.
Glycogen Accumulating Organisms (GAOs)
GAOs are microorganisms that are capable of using VFAs under anaerobic conditions to form PHA without being able to accumulate polyphosphate under aerobic conditions. So they compete for VFAs with PAOs and have a negative impact on P-removal in EBPR processes. It has been hypothesized that S2EBPR may lead to lower relative abundances of GAOs due to higher decay rates and lack of multiple internal polymer stores under anaerobic conditions compared to PAOs. I am currently performing research to evaluate this hypothesis and characterize the ecology of GAOs in S2EBPR facilities.
Simultaneous Nitrogen and Phosphorus Removal
WRRFs face stringent nitrogen (N) and phosphorus (P) limits. Typically processes for N and P removal have been implemented and handled independently due to inherent incompatibilities due to differing carbon requirements. But there are enormous advantages in being able to accomplish both these goals simultaneously. I am currently performing research to integrate EBPR with N-removal processes, understand fundamental mechanisms that influence its performance and the microbial ecology and dynamics of such an integrated process.
P-Cycling in Natural Environments
I am currently also performing research to characterize the microbial ecology of phosphorus (P) cycling in natural environments. Nutrient loading into natural waters causes a variety of issues such as algal blooms (which can include toxic cyanobacterial blooms) and eutrophication which can be detrimental to aquatic life. Understanding and minimizing P loading to the environment is one of the critical issues facing the environment today. Polyphosphate accumulating organisms (PAOs) accumulate phosphorus intracellularly as polyphosphate in excess of their cellular needs which makes them a critical player in P cycling. Understanding their prevalence and identity along with their activity is very important to assess their impact on P cycling. For this I am developing protocols for performing fluorescence-activated cell sorting (FACS) and activity tests which can be used in conjunction with 16S rRNA gene sequencing and ‘Omic techniques.
Previous Research
Microbial Fuel Cells (MFCs)
Microbial fuel fells (MFCs)are a well-studied potential technology for bioremediation and decentralized wastewater treatment. However, progress has been somewhat stalled at the bench-scale. In well controlled experiments, electron recovery is high. In natural environments, wastewaters are complex and anode-respiring bacteria can be outcompeted in the presence of competing microorganisms,leading to a loss in electron-recovery and power production. My research on MFCs focused on characterizing this competition and its impact on scale-up of MFCs for anaerobic wastewater treatment.
Effect of Nitrate on Anode-Respiring Communities in MFCs
A poorly understood phenomenon with a potentially significant impact on electron recovery in MFCs is the role of competition between anode-respiring bacteria and microorganisms that use other electron acceptors. Nitrogen species are a major constituent of wastewater and nitrate can act as a competing electron acceptor in the anode. Studies investigating the impact of competition on population dynamics in mixed communities in the anode are lacking. I investigated the impact of nitrate at different C/N ratios, 1.8, 3.7and 7.4 mg-C/mg-N, on the electrochemical performance and the biofilm community in mixed-culture chemostat MFCs. The electrochemical performance of the MFC was not affected under electron donor non-limiting conditions, 7.4 mg-C/mg-N. At lower C/N ratios, electron donor limiting, electron recovery was significantly lower. The electrochemical performance recovered upon removal of nitrate at 3.7 mg-C/mg-N. Microbial community analysis showed a decrease of Deltaproteobacteria accompanied by an increase in Betaproteobacteria in response to nitrate at low C/N ratios,and no significant changes at 7.4 mg-C/mg-N. Transcriptional analysis showed increased transcription of nirK and nirS genes during nitrate flux suggesting that denitrification to N2 (and not facultative nitrate reduction by Geobacter spp.) might be the primary response to perturbation with nitrate.
Further details can be obtained from the published paper.
Modelling Intracellular Competition in a Denitrifying Biocathode
One potential MFC configuration uses an organic-oxidizing anode biofilm and a denitrifying cathode biofilm. However nitrite, a denitrification intermediate with environmental and public health impacts, has been reported to accumulate. In this study, before complete denitrification was achieved in a bench-scale, batch denitrifying cathode, nitrite concentrations reached 66.4 % ± 7.5 % of the initial nitrogen. Common environmental inhibitors such as insufficient electron donor, dissolved oxygen, insufficient carbon source, and pH, were considered as a cause of the accumulation. Improvement in these conditions did not mitigate nitrite accumulation. I used an activated sludge model with an integration of the Nernst-Monod model and indirect coupling of electrons (ASM-NICE) which effectively simulated the observed batch data, including nitrite-accumulation by coupling biocathodic electron transfer to intracellular electron mediators. The simulated half-saturation constants for mediated intracellular transfer of electrons during nitrate and nitrite reduction suggested a greater affinity for nitrate reduction when electrons were not limiting. The results implied that longer hydraulic retention times (HRTs) may be necessary for a denitrifying biocathode to ensure complete denitrification. These findings could play a role in designing full-scale MFC wastewater treatment systems to maximize total nitrogen removal.
Further details can be obtained from the published paper.
Fate and Transport of Endocrine Disrupting Compounds (EDCs) and Pharmaceuticals and Personal Care Products (PPCPs)
Numerous studies have reported the presence of several endocrine disrupting compounds (EDC) and pharmaceuticals and personal care products (PPCP) in wastewater effluents and consequently in natural water sources which serve as source water for drinking water utilities. However it is still unclear as to which of these compounds are important and need to be monitored. I proposed a new approach to identify indicators or surrogates to monitor these compounds in a watershed based on intensive sampling, analytical methods and statistical analysis. The watershed-level protocol involves identifying common patterns of occurrence in these trace chemicals and proposes indicators based on results of statistical analyses such as correlation, clustering and principal component analysis. I also evaluated the application of the indicators to predict concentration levels of other compounds by developing regression models and determining significance of the models. For this research, I performed sampling in the Assabet River in Massachusetts and analyzed 26 trace compounds and 3 tracers using solid phase extraction (SPE), liquid chromatography (LC) coupled with tandem mass spectrometery (MS/MS) and inductively coupled plasma mass spectrometry (ICP-MS). My research showed that that gadolinium served as an indicator for 14 other compounds and could be used as a surrogate. The application of this protocol will help drinking water utilities and regulators alike to more effectively utilize their allocated resources.
Further details can be obtained from the published paper and my thesis.
