Those of us working with industrial wastewater treatment often complain about textbooks and training courses focusing on municipal wastewater. Rick Marshall & Steve Leach - both highly experienced wastewater professionals - are offering a petrochemical focused wastewater course in December 2020. This course will be online using Zoom but have the same content as in-person training. I highly recommend this course for those of you working in petrochemical or refining wastewater treatment. 

Microbial metabolism generates energy by moving electrons along metabolic pathways to a terminal electron acceptor. If this sounds complex or brings back images of the biology class electron transport chains, I apologize but it is good to know the general metabolic options in wastewater and not get bogged down by the individual biochemistry steps.

We often use the term aerobic, facultative anaerobe, or anaerobe when talking about wastewater bacteria. These terms are great, but the lines are not as clean as you may think. Let's start with the general options.

It all starts with where the electron ends
Going back to the electron tower (pictured above), microbes generate energy for growth by moving the electron to the highest energy acceptor. If the terminal electron acceptor is oxygen, you have an extremely high energy yield. As you move to other electron acceptors, less energy is produced. 

Terms that we use in wastewater

• Obligate aerobe
  Only grows when oxygen is present. Often we say NItrosomonas (AOB) are obligate aerobes, but even Nitrosomonas has limited growth using alternative anoxic pathways. 
• Facultative anaerobe
  In wastewater, a facultative anaerobe is any culture having the ability to grow when dissolved oxygen is not present. They can use various alternative electron acceptors. If they do not have an appropriate electron acceptor they do not grow. These are the most common cultures in wastewater treatment systems - example genera include: Pseudomonas, Bacillus, Zooglea, Thauera, & Paracoccus.
• Obligate anaerobe
  Here we have organisms that are poisoned by free oxygen. They only grow in reducing (low redox) environments. Here you have organisms found in sludge layers, inside dense granular floc, or anaerobic digesters. Common examples include Clostridium (an acid forming bacteria) and Methanogens (archaeal cultures that produce methane from short-chain organic acids and hydrogen).
• Fermentative 
  This group includes some facultative anaerobes and obligate anaerobes! Fermentative respiration requires the terminal electron acceptor to be organic compounds. This is the acid forming step in anaerobic digesters. Fermentative respiration yields less energy than growth using oxygen, nitrate, nitrite, or sulfate as electron acceptors.
• Microaerophilic
  This includes cultures that thrive at low oxygen levels. They use oxygen as an electron acceptor but exploit a relatively low D.O. ecological niche.

Foam can tell you a lot about the biological state of your aeration basin. Some foams including the normal white foam and the greasy thick stable foams are produced from the byproducts of microbial growth. Here are the most common foams that you see on aeration basins:

Normal Biological Foam
As bacteria grow and divide, they produce extracellular materials including polysaccharides, proteins (enzymes), and even DNA/RNA. Also important for biofilm or floc adhesion, these materials trap air bubbles in a light color foam that is easily broken by waters spray if it even builds up more than 1 - 2" on the water surface. When the system is in log phase growth, you may notice both a lighter color and more foam than when the system is mature in decline phase growth.

Nocardia/Microthrix Parvicella (filaments)
Nocardia (often actually a Gordonia sp) is promoted by long chain fatty acids (FOG) and Microthrix is triggered often by low F/M, long chain fatty acids and lower temperatures. Both genera can exist in wastewater and not cause foaming, but under appropriate conditions, they start to produce hydrophobic extracellular materials. The hydrophobic biological polymers trap air and create the thick, stable, greasy foam. The best way to prevent foaming is to reduce grease and keep appropriate MCRT or F/M ratios in the system.

Surfactant Foams
Surfactants are used in many products not just cleaning agents. Firefighting foams, food product emulsion stabilizers, even many personal care products contain surfactant chemistries. Under normal concentrations, bacteria readily remove surfactants or they are diluted beyond their ability to reduce surface tension (creating foam). Periodically, you may see higher loadings hit the aeration basin. With surfactants the foam is usually stable and has a light color. While their presence is temporary and foam levels should drop, you can use commercial antifoam products to prevent excess foam during peak events.

Plant soaps, Starches, & Proteins
The above chemicals are natural polymers found in wastewater. Foaming occurrs when normal biological foam beomes stabilized by the natural polymers. In this case, antifoams and water sprays are often effective. We have also effectively used pretreatment with enzymes to pre-digest the polymers prior to the aeration basin - this can be very valuable in food processing wastewater treatment.

For years, I have always been told that Nitrosomonas sp are obligate aerobic bacteria. We all accepted that Nitrosomonas  used oxygen to convert ammonia into nitrite. That was the extent of their metabolic capabilities. However, Nitrosomonas have now been found to have a few other metabolic tricks available to survive in anoxic and anaerobic environments. How well does Nitrosomonas grow under D.O. limited environments, surprisingly well. In fact, Aster Bio's MCA tests on multiple anaerobic digesters has found significant Nitrosomonas populations. Here is what Nitrosomonas does in aerobic, anoxic, and anaerobic environments:

• Aerobic - this is where Nitrosomonas exhibits most rapid growth and is the best-known metabolic process. Using Ammonia + Oxygen to produce Nitrite.
  
  
• Anaerobic - Nitrosomonas can function as an ANAMMOX culture. In this pathway, Ammonium and Nitrite are the inputs with the final products being Nitrogen gas and H2O.  
  
  
• Anoxic - Nitrosomonas uses nitrite as the terminal electron acceptor and can use various cell intermediates and a few organic compounds to provide energy for growth. Technically, the anaerobic pathways listed above can be included with the anoxic systems as both require nitrite.

What does this mean for wastewater treatment? Well you may have noticed much emphasis on ANAMMOX technology where ammonia oxidation and denitrification happen in the same step. This saves on energy for aeration and makes denitrification more efficient. You do have to account for the slower growth under anoxic/anaerobic conditions, but Nitrosomonas may turn out to be one of the most important ANAMMOX cultures in wastewater systems as it also provides the NO2 used in ANAMOX during aerobic growth.

Much of our animal based foods come from CAFO systems where animals are housed at high density. WIth the high density comes problems with manure and urine. The number and size of these operations means that CAFO can be the largest source of wastewater in many areas. The waste is high in BOD5 and ammonia which is often treated in lagoon based systems with land application via irrigation systems. Odors and treatment inefficiencies in ponds can cause problems with operations, so there was room for improving biological treatment processes.

As this is an ongoing R&D project, I thought this would be a way to introduce you to how biological products are developed for new applications. We started with a manure degrading formulation developed in the 1990s but knew that advances in our technologies could improve the results. Working with several groups on introducing biological pretreatment in the animal houses, we are using extensive analytical testing in both pilot scale and working houses. Testing includes standard water chemistry tests and DNA based testing for changes in the biomass composition. The goals for adding biological products being:

• Reduce difficulty in pumping manure from collection pits to wastetreatment lagoons
• Lower noxious gases inside buildings - this includes H2S, ammonia, and odor causing VOCs
• Control dense crust formation which intensifies files and other nuisance insects
• Start biological degradation upstream of the waste treatment lagoon thereby reducing treatment time required in the lagoon system

We noticed that many biolgoical additives were being marketed for CAFO and manure treatment operations and I had worked on manure treatment products years before without much scientific testing. So we started at the begining and looked at the differences in feeds, bedding (if used), and operation protocols. By learning where and why bottlenecks occur, we were able to see opportunities to modify the biomass and enzymes present. In the past, we assumed that Bacillus strains associated with plant based waste degradation were sufficient and used well known waste degrading Bacillus spores. They worked, but there was definitely room for improvement. We also did not limit ourselves to bioaugmentation technologies. The research aslo considred the use of micronutrients, enzymes, or other supportive chemistries also enhance pre-treatment. Here is what we have discovered and are continuing to evaluate:

• Pretreatment in animal housing is a cost-effective option
• Systems with straw bedding benefitted from adding a newly isolated high cellulase producing organism.
• The inclusion of biosurfactant and high cellulase strains improved manure liquefication which was the primary bottleneck in the pre-treatment system.
• Adding enzymes did improved the results
• The use of dry cultures immobilized on biosupportive carrier allowed for the inclusion of vegetative organisms for performance in lower temperatures and improved removal of H2S and other odor causing compounds
• The appropriate solution depends upon animals housed and other local conditons so there was no universal product

In many areas, wastewater and stormwater go through facultative or moderately aerated lagoon systems prior to discharge. These lagoons remove residual oragnics, nutrients, and solids that can pass through the upstream treatment. In the case of stormwater, the lagoons remove nutrients from fertilizer runoff and hydrocarbons from parking lots/roads. Unlike modern wastewater treatment sytems, the lagoons have low levels of biomass (MLSS) that can range from 30 - 100 mg/L volatile solids. The solids component consists of:

• Residual TSS from wastewater treatment - both living organisms and non-living components
• Facultative bacteria that thrive in the low F/M environment (we actually see AOB thriving in ponds with high sludge levels during summer months)
• Algae - both eukaryotic and cyanobacteria. Often excess phosphorus can trigger a cyanobacteria bloom causing problems with TSS, odors, and pH.

The reason for ammonia appearing and AOB growing during summer months is a phenomenon related to sludge degradation on the pond bottom. While the sludge layer degrades all year, the rate of anaerobic lysis increases with summer temperatures. The pond bottom remains anaerobic/anoxic producing organic acids, methane, and nutrients (N, P). The upper layers have oxygen present from algae photosynthesis, natural diffusion from the surface, and action of mixers/aerators if present. In ponds with more sludge, the problems related to ammonia release and algae blooms tend to cause more problems. So what can be done to control this other than a big mechanical dredging project?

• Add mixers for low velocity, high volume movement. The mixer circulates water from anerobic zone to the upper aerobic regions, allowing for more effective sludge stabilization/nutrient removal. We are also seeing benefits from adding new micro & nanobubble aeration technologies here too.
• If mixers are not enough to control the problem, you can then use algae control methods inclusing sonic systems, copper sulfate, and biological algae control methods.
• If sludge layer is visible at the surface and has produced "islands" - then it is time to evaluate dredging or various biodgredging technologies. If the sluge has a high organic fraction, then biodredging may prove less costly and effective.