The methanogens found in anaerobic digesters are actually not bacteria but from the kingdom archaea. Archaea are very different from bacteria and are their own unique life forms found in a variety of environments. In anaerobic systems, it is the methanogens that produce methane and maintaining a stable population of methanogens is key to successful digester operation. But, methanogens also have multiple metabolic pathways that impact digester performance. 

Archaea Methanogenic Pathways:
Acetoclastic                   CH3COOH  CH4 + CO2
Hydrogenotrophic       CO2 + 4H2  CH4 + 2H2O
Methylotrophic             4CH3OH  3CH4 + CO2 + 2H2O
 
Bacterial Pathways in Digesters:
            Fermentation              Produces organic acids from complex biological polymers
            Acetogenesis               Continued fermentation to CH3COOH
            Homoacetogeneis       4H2 + 2CO2  CH3COOH + 2H2O
                                                   Metabolic rate at 3 to 10x faster than HM Archaea
             SAO                             CH3COOH  4H2 + 2CO2
                                                  Syntrophic with Hydrogenotrophic methanogens
 
Causes of increases CO2 (lower methane %) in produced gas

• Acetoclastic and methylotrophic methanogens both produce CO2 along with methane. With the acetoclastic making equal parts CO2 and methane.  In digesters dominated by acetoclastic organisms (especially Methanothrix from Aster Bio's data), we see lower rates of methane production.
• Overgrowth by homoacetogenic bacteria - these organisms convert H2 and CO2 back to acetate which favors acetoclastic methanogen growth. These organisms deprive desirable hydrogenotrophic methanogens of needed inputs for growth.
• Need for SAO bacterial populations that work directly with hydrogenotrophic methanogens in a syntrophic relationship where the bacteria provide the archaea with H2 and CO2 - the result is improved methane yields. 

Good articles on the SAO & Homoacetogenesis bacteria in digester performance.

• Deep insights into the network of acetate metabolism in anaerobic digestion: focusing on syntrophic acetate oxidation and homoacetogenesis​ - ​www.sciencedirect.com/science/article/abs/pii/S0043135420313075
• Syntrophy mechanism, microbial population, and process optimization for volatile fatty acids metabolism in anaerobic digestion - www.sciencedirect.com/science/article/abs/pii/S1385894722046162
• Novel syntrophic bacteria in full-scale anaerobic digesters revealed by genome-centric metatranscriptomics   www.nature.com/articles/s41396-019-0571-0/

Odor problems are an ongoing nuisance in wastewater treatment. And, there are so many options to help control odors related to collection systems, headworks, and even aeration basins. When evaluating various control options, it may be beneficial to look at the chemistry of wastewater odors.

If you smell something it is evaporating. In effect, something in the water is moving into the surrounding air and our noses are able to detect smells. Factors that impact evaporation rate include:

• Temperature
• Surface area
• pH & speciation
• Solubility
• Henry's Law (dissolved gas in a liquid is related to partial pressure)
  
  

Our most common nuisance odors in wastewater arise from anaerobic biochemistry. And here is a short list of offensive compounds and how they smell:

• Sulfides (H2S) - classic rotten egg odor
• Organic acids - sharp rancid odors
• Mercaptans - RSH - putrid smell (detectable at extremely low concentrations)
• Dimethyl sulfide - cooked cabbage like odors
• Ammonia/urea - ammoniacal 

​Biochemistry Section
In wastewater, our odor causing problems are usually related to anaerobic biochemical reactions. We also need to consider speciation (or what form is present) of the compounds which is related to pH & ORP (redox). For example, H2S gas is the volatile form of sulfides in wastewater. As you decrease pH, you have increased % as H2S in the air versus S= (which is in the water). 

In a typical collection system pH, bacteria thrive. Organics are present for oxidation. Bacteria then search for the most available, highest energy yielding reaction that acts as an ELECTRON ACCEPTOR. Once you deplete oxygen, you enter anoxic/anaerobic metabolism. For odors, the key is what electron acceptor the bacteria are using; as this will help determine redox, pH, and resulting odor causing compounds.

Control Options & How They Work

• Masking agents (fragrances) - fast control of odors but an after the fact solution.
• Sulfide binding chemistry - works on H2S but ignores other odor problems
• Oxidants - can directly oxidize odorous compounds or increase redox potential (provide a higher energy electron acceptor)
• Nitrate - provides a convenient, low-cost alternative electron acceptor to increase redox potentials.
• pH adjustment - increasing pH can reduce volatilization of sulfides and organic acids. However it can increase problems with ammonia odors.
• Biological treatment - this includes biological scrubbers and cultures added to lines. However, they do require oxygen or other higher energy electron acceptor to reduce odors.
  
  

Putting it all together
No single odor control option fits every situation. If you have a consistent odor issue, you should investigate to determine what is causing the odors and work on the biochemistry of how and where these compounds are forming in the system. Once surveyed, you can take steps to prevent the formation of the underlying compounds by utilizing knowledge of biochemistry and speciation of volatile compounds. I have found success in using a combination of redox potentials (ORP), temperature, and pH to be highly effective in routine monitoring. 

It is often taught that bacteria can double every 20 minutes. This 20 minute number comes from common food spoilage organisms such as E. coli. In wastewater with diverse microbial populations doubling time is different for each species but we can use trends in growth rates to see how doubling time relates to F/M, MCRT, and problems with washout. First, let's step back and look at some of the general principals of microbial ecology as it relates to wastewater.

Two General Strategists

• r-strategists - which are faster growing organisms
  • Log phase in WWTP
  • Seen in high F/M
  • Changing influent concentrations or makeup
  • Lag & Log phase growth biomass
• K-strategists -  are the slower growing organisms vital for treatment
  • Tend to be slower growing
  • Exploit a niche
    1. Substrates for growth
    2. pH extremes
    3. Temperatures extremes
       
       

The Microbial Growth Curve
If you have read this site before, you know that I always like to relate microbial populations and biological treatment unit performance to your location on the Microbial Growth Curve. This gives us a convenient visualization to help explain they why bacteria are behaving as they do.

• During startup or near a lagoon influent you have a “bloom” of favoring fast growing organisms that thrive in high F/M environments. This tends to be your r-strategist microbes.
• As the easily used soluble biodegradable organics (BOD5) decreases, you see overall growth rates slow and organisms that exploit niche substrates emerge. This is where we use the terms "mature biomass", "steady-state", and "decline phase growth". At this point log growth stops and you have low F/M.
  
  

Examples in WW

1. Fast growing in WW
   1. Many common genera – well known since the also grow fast in the lab!
   2. Doubling rate is usually in the hours for WWTP
2. Niche organisms
   1. AOB & NOB - nitrifiers
   2. Phosphate Accumulating Organisms (PAO)
   3. Sulfur Oxidizing
   4. Nocardia forms (foaming bacteria)
   5. Some filamentous bacteria (low F/M)
      
3. Environments promoting niche organisms
   1. Longer sludge age – must be long enough to avoid wash out
   2. Presence of substrate at levels promoting their growth
   3. Environmental factors that favor their growth - examples include organic acids, low D.O., high temperature, low N or P, etc.
   4. F/M conditions that favor development of these niche organisms. In the case of nitrifiers, the rule of thumb is 80% of COD must be removed before you start to see substantial growth in the MLSS.

Over the past year, I have seen an increasing number of systems having zero to low populations of known nitrifiers yet they are still removing total inorganic nitrogen. Driven by data generated from molecular testing, we are learning a lot about the various pathways for nitrogen transformations in wastewater. And now we are seeing the role the Phosphate Accumulating Organisms (PAO) can also play in the process.

Nitrification Pathways

• AOB/NOB - this process is the well-known chemoautotrophic bacteria. This is the familiar oxidation of ammonia to nitrite by Nitrosomonas. And the most common NOB is actually Nitrospira. 
  
  
• COMAMMOX - Complete Ammonia Oxidation - A subset of Nitrospira can oxidize ammonia to nitrate. This reduces the number of single step AOB seen in the system.
  
  
• ANAMMOX - Anaerobic Ammonia Oxidation - organisms from the phylum Planctomycetes utilize NO2 + NH4 --> N2 + H2O. This requires AOB for nitrite, but the short cut to N2 gas results in substantial energy savings. The challenge here is ANAMMOX cultures have an extremely slow growth rate and require anaerobic conditions to have a metabolic advantage.
  
  
• Heterotrophic Nitrification - some heterotrophic organisms can oxidize ammonia and nitrite. These organisms grow various organic substrates but also have the ability to oxidize ammonia and nitrite.
  
  
• Simultaneous Nitrification Denitrification - building on heterotrophic nitrification with organisms such as Paracoccus denitrificans, Pseudomonas sp, and Alcaligenes sp having pathways for ammonia oxidation and denitrification even under aerobic conditions.  Factors impacting SND pathways include carbon source, C/N ratio, pH, temperature, and Dissolved Oxygen.
  
  
• Simultaneous Removal of Nitrogen and Phosphorus - SNDPR - this is often seen in low COD systems and aquaculture, but some PAO also have the ability to take a role in removal of inorganic nitrogen in wastewater. 

Challenge of using the above pathways in wastewater treatment

1. Just because an organism has metabolic capabilities or genes, it does not mean that they will be turned on or used. For example, Paracoccus denitrificans can oxidize ammonia and grow on organic acids. If you have sufficient organics (COD), it will obtain more energy using standard heterotrophic pathways and you may see little ammonia oxidation. This is why C/N ratios can become very important for heterotrophic nitrification - in WW, this means lower F/M may trigger other energy yielding pathways.
   
   
2. Do you need separate zones with various D.O. levels, or can we have multiple zones inside biofilms or granules. This is the mechanism seen in newer granular sludge systems where the individual granule has aerobic/anoxic/anaerobic zones. This also happens to a lesser extent in conventional floc/biofilm.
   
   
3. Perhaps a mix of all of the above will be more robust and less susceptible to upset from changing conditions. We actually see this in working systems where organisms that can exploit a ecological niche will grow and populations do change in response to loading and environmental/operational conditions.

Good articles on heterotrophic nitrification, SND, and SNDPR

Pertti J. Martikainen, “Heterotrophic nitrification – An eternal mystery in the nitrogen cycle” Soil Biology and Biochemistry Volume 168, 2022. https://www.sciencedirect.com/science/article/pii/S0038071722000682?via%3Dihub
 
Shivani Shukla, et al. “Simultaneous Nitrification–Denitrification by Phosphate Accumulating Microorganisms” World Journal of Microbiology and Biotechnology (2020) 36:151
 
Zhao W, Bi X, Peng Y, Bai M. Research advances of the phosphorus-accumulating organisms of Candidatus Accumulibacter, Dechloromonas and Tetrasphaera: Metabolic mechanisms, applications and influencing factors. Chemosphere. 2022 Nov;307(Pt 1):135675. doi: 10.1016/j.chemosphere.2022.135675. Epub 2022 Jul 13. PMID: 35842039

I have seen recent social media posts showing people pouring concentrated hydrogen peroxide into manholes, lift-stations, biological treatment plants, and even lakes. The claim is that hydrogen peroxide is not harmful to anything and solves all problems from FOG to hazardous algae blooms. In the videos, people are pouring 5 gallon pails of peroxide into all of the above. I know people want to sell their products and talk about the benefits of their technology. However, every product has strengths and weaknesses and may or may not fit the application. 

When it comes to oxidant chemistry in collection systems, you have many choices. You need to find the one that best matches site specific challenges and keeps costs under control. This often means that a multiple lift station collection system benefits from using multiple technologies. To choose among technologies it helps to:

• Know the strengths and weaknesses of each approach
• You need to know the system - map out ORP, discover anoxic/anaerobic zones, find out where FOG is accumulating and why - there is a lot to mapping out the problems, but it is a vital step in optimizing treatment.
• Implement solutions and monitor/adjust programs.

Now for most used oxidant technology options.

Oxygen Uptake Rates (OUR) - also called Dissolved Oxygen Uptake Rates (DOUR) are one of the common wastewater tests that give valuable information on microbial activity levels. A fast and simple test, OUR measures oxygen consumption by biomass over time. Results are expressed as mg O2/L/hr. It is similar to more complex respirometery testing; it is just faster and requires no special equipment.

Theoretical Background

• Aerobic Bacteria (microbes) use oxygen when growing
• More "food" = faster growth and higher OUR 
• Mature or older biomass = slower growth and lower OUR 
• More biological solids mean more endogenous respiration, so we introduce SOUR to account for different MLSS or MLVSS concentrations

Standardized Oxygen Uptake Rates (SOUR) - account for changes in biological solids
Important for activated sludge – divide OUR by MLVSS in g – to adjust for background respiration by solids. Note if you use MLSS instead of MLVSS as solids. MLSS > MLVSS. Whatever you use just be consistent!

Normal OUR
While there are "rules of thumb" for OUR, that does not replace routine testing of your specific system. Operations and sample points will make a difference in OUR. The test takes less than 10 minutes so take a few minutes and run the test as frequently as feasible.

Interpretation of Results
Remember we are using respiration rates to estimate where we are on the growth curve – works with microscopic exam, SV30/SVI, and other tests

Low OUR (< baseline average)

• Lower F/M (less BOD) or “older” biomass
• For industrial – can also indicate toxic shock where you actually killed viable microbes and have yet to enter log growth.
• Low OUR can indicate moving into endogenous phase of the growth curve

High OUR (> baseline averages)

• Increased soluble BOD5 - microbes are growing faster which is also higher F/M and "young" biomass
• Very high OUR can indicate log phase of the growth curve