8.6 Activated Sludge Microbiology, Aeration Basins & Secondary Clarifiers

8.6 Activated Sludge Microbiology, Aeration Basins & Secondary Clarifiers

1. Principles of Suspended-Growth Biological Treatment

Secondary treatment in modern municipal wastewater facilities relies predominantly on the activated sludge process, a suspended-growth biological treatment method first developed by Edward Ardern and W.T. Lockett in England in 1914. While preliminary and primary treatment rely on physical separation to remove coarse debris and settleable solids, the secondary biological process utilizes a living ecosystem of microorganisms to remove soluble, dissolved, and colloidal organic carbon compounds that cannot be captured by gravity alone.

The Living Biological Suspension

Inside the aeration basin, primary effluent (containing dissolved sugars, organic acids, proteins, and carbohydrates) is vigorously mixed with recycled biological solids. This mixed biological suspension is termed Mixed Liquor:

• Mixed Liquor Suspended Solids (MLSS): The total concentration of all suspended organic and inorganic solids in the aeration basin, expressed in milligrams per liter (mg/L). In conventional activated sludge facilities, MLSS typically ranges from 1,500 to 3,500 mg/L. In extended aeration systems and oxidation ditches, MLSS ranges from 3,000 to 6,000 mg/L.
• Mixed Liquor Volatile Suspended Solids (MLVSS): The volatile fraction of the MLSS, determined by igniting the dried TSS filter residue in a muffle furnace at 550°C. The volatile fraction burns off, leaving behind non-combustible fixed (ash/mineral) solids. MLVSS serves as the practical operational surrogate for active biological mass. In municipal systems, MLVSS normally constitutes 70% to 85% of total MLSS (i.e., Volatile Fraction = 0.70 to 0.85).

Biochemical Metabolism and Assimilation

Microorganisms in the activated sludge basin consist primarily of aerobic and facultative heterotrophic bacteria. They absorb dissolved organics through their semi-permeable cell membranes and oxidize them via enzyme-catalyzed biochemical pathways:

$\text{Organic Matter } (BOD_5) + O_2 + \text{Nutrients } (N, P) + \text{Cells} \xrightarrow{\text{Enzymes}} CO_2 + H_2O + \text{New Cells} + \text{Energy}$

1. Synthesis (Assimilation): A portion of the metabolized organic carbon is converted into new cellular tissue, resulting in net biomass growth.
2. Endogenous Respiration (Auto-Oxidation): When soluble food becomes depleted, bacteria metabolize their own internal cellular reserves and protoplasm to obtain maintenance energy:

$\text{Bacterial Cells } (C_5H_7NO_2) + 5 O_2 \longrightarrow 5 CO_2 + 2 H_2O + NH_3 + \text{Energy}$

Influent Primary Effluent
         │
         ▼
┌──────────────────────────────────────────┐
│         AERATION BASIN                   │
│  - Mixed Liquor (MLSS / MLVSS)           │
│  - Aerobic Microorganisms Metabolize BOD │ ◄── Compressed Air / Diffusers (DO: 1.5-2.5 mg/L)
│  - Floc Formation (EPS Exopolysaccharide)│
└──────────────────────────────────────────┘
         │
         ▼ Mixed Liquor Flow
┌──────────────────────────────────────────┐
│       SECONDARY CLARIFIER                │ ──► Clarified Effluent (Over Weirs to Disinfection)
│  1. Clarification (Clear Supernatant)    │
│  2. Thickening (Bottom Sludge Blanket)   │
└──────────────────────────────────────────┘
         │
         ├─────────────────────────────────────────┐
         ▼                                         ▼
Return Activated Sludge (RAS)             Waste Activated Sludge (WAS)
Recycled to Aeration Basin                Removed Daily to Solids Handling
(Maintains MLSS Inventory)               (Controls Sludge Age / MCRT)

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2. Activated Sludge Microbiology & Ecological Succession

Routine microscopic analysis of mixed liquor wet mounts (using brightfield or phase-contrast microscopy at 100× and 400× total magnification) is an essential diagnostic tool for North Carolina certified operators. Changes in microbial populations occur days before deterioration is visible in routine laboratory testing (e.g., effluent BOD and TSS). The biological community progresses through a well-defined ecological succession governed by sludge age (Mean Cell Residence Time) and organic loading rates.

   Relative Population
      ▲
 High │  [Bacteria]     [Amoebas & Flagellates]
      │   ───────       ───────────────────────
      │                                   [Free-Swimming Ciliates]
      │                                   ────────────────────────
 Med  │                                                    [Stalked Ciliates]
      │                                                    ─────────────────
      │                                                                     [Rotifers & Nematodes]
 Low  │                                                                     ─────────────────────
      └──────────────────────────────────────────────────────────────────────────────────────────►
        Young Sludge / High F/M / Low MCRT                     Old Sludge / Low F/M / High MCRT
        Rapid Growth Phase                                     Endogenous Respiration Phase

Primary Microorganisms and Succession Indicators

1. Bacteria

Bacteria represent 90% to 95% of the total microbial biomass in activated sludge. They are responsible for direct carbonaceous decomposition:

• Floc-Forming Bacteria: Bacteria such as Zoogloea ramigera secrete sticky, gelatinous extracellular polymeric substances (EPS), consisting of polysaccharides and proteins. EPS causes individual, dispersed bacteria to aggregate into dense, rapidly settling clusters known as flocs.
• Filamentous Bacteria: Long, thread-like cellular chains that grow within the floc. In moderate numbers, filaments form a structural mesh or 'backbone' that gives strength to biological flocs.

2. Protozoa

Protozoa are single-celled eukaryotic organisms that feed on dispersed single bacteria, suspended colloidal particles, and other protozoa. They clean the liquid phase, producing a clear, sparkling effluent:

• Amoebas (Sarcodina): Move via sluggish cytoplasmic extensions called pseudopods. They predominate in very young sludge, during facility start-up, or under severe organic shock loads (high F/M, low dissolved oxygen). They indicate poor treatment efficiency.
• Flagellates (Mastigophora): Characterized by one or more whip-like flagella that propel them rapidly through the liquid. Flagellates feed directly on dissolved and colloidal organics. Like amoebas, they predominate in young sludge where soluble organic food is abundant. Their presence in large numbers indicates high effluent turbidity and young sludge.
• Free-Swimming Ciliates: Oval or kidney-shaped cells covered in rhythmic, beating hair-like cilia. They swim freely through the mixed liquor feeding on dispersed bacteria. They indicate a transitioning sludge of moderate age and improving health.
• Stalked (Vorticellid) Ciliates: Anchor themselves directly to biological flocs with contractible stalks (e.g., Vorticella, Epistylis, Carchesium). A crown of cilia beats at their anterior cytostome (mouth), creating feeding vortexes that capture dispersed bacteria. Stalked ciliates indicate a mature, stable, healthy activated sludge system with excellent flocculation, low effluent turbidity, and high BOD removal.

3. Metazoa (Multicellular Animals)

• Rotifers: Multicellular organisms with a distinct head crowned by circular cilia rings (corona) that resemble spinning wheels, a muscular mastax (grinding jaw), and a telescoping foot. Rotifers consume free bacteria and small floc particles.
• Nematodes: Microscopic roundworms that thrash in a slow, S-shaped motion, browsing on floc interiors.
• Significance: Rotifers and nematodes appear only in older sludge with high Mean Cell Residence Times (long MCRTs, low F/M), complete nitrification, and advanced endogenous respiration.

Filamentous Bulking and Foam Dynamics

When environmental conditions favor filamentous bacteria over floc-formers, filaments grow out from the floc core into the bulk liquid, mechanically bridging flocs and preventing them from compacting in the secondary clarifier. This condition is known as filamentous sludge bulking, characterized by a very slow-settling sludge blanket and Sludge Volume Indexes ($SVI$) exceeding 150 to 200 mL/g.

Control Measures for Filamentous Bulking

• Targeted RAS Chlorination: Chlorine acts as a non-selective biocide. Because filamentous organisms extend outside the protective floc matrix, dosing chlorine into the Return Activated Sludge (RAS) stream exposes the filaments directly to the oxidant while the interior floc-formers remain protected. The standard operating dosage is 2 to 3 lbs of chlorine ($Cl_2$) per 1,000 lbs of MLVSS per day. Chlorination must occur at a point of high turbulence (such as the suction side of the RAS pump) to ensure instantaneous, uniform contact. Over-chlorination destroys the floc structure, producing pin floc and milky, turbid effluent.
• Biological Selectors: Introducing an un-aerated zone (anaerobic, anoxic, or high-rate aerobic contact zone) ahead of the main aeration basin. Floc-forming bacteria have high uptake rates for soluble substrate under dynamic conditions and store food internally, while filaments cannot absorb organics rapidly in the selector, effectively starving them out.

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3. Aeration Systems and Basin Hydraulics

Dissolved Oxygen Targets & Oxygen Transfer Dynamics

Microorganisms require dissolved molecular oxygen ($O_2$) as the terminal electron acceptor in aerobic respiration.

• Target Basin DO: 1.5 to 2.5 mg/L throughout all active aerobic zones.
• Low DO Hazards (< 1.0 mg/L): Impairs complete carbonaceous oxidation, prevents autotrophic nitrification, and selects for low-DO filamentous organisms (Type 1701, S. natans).
• High DO Hazards (> 3.5 to 4.0 mg/L): Wastes substantial electrical power (aeration blowers account for 50% to 70% of total treatment plant energy consumption) and can mechanically shear delicate biological flocs, generating pinpoint floc.

Aeration Technologies

1. Fine Bubble Diffused Aeration: Compressed air is pumped through porous membrane disks, tubes, or ceramic panels submerged along the basin floor. Fine bubbles (1 to 3 mm diameter) have a high surface-area-to-volume ratio and slow rise velocities, achieving a high Standard Oxygen Transfer Efficiency (SOTE) of 20% to 35% in clean water. However, fine bubble diffusers require clean air filtration and periodic chemical or acid cleaning to prevent biological and mineral fouling.
2. Coarse Bubble Diffusers: Generate large bubbles (6 to 12 mm diameter) that create vigorous hydraulic turbulence and mixing. They exhibit a lower oxygen transfer efficiency (6% to 12% SOTE) but are non-clogging, making them ideal for aerated grit chambers, channels, and aerobic digesters.
3. Mechanical Surface Aerators: Motor-driven vertical impellers or horizontal brush rotors (common in oxidation ditches) splash wastewater into the atmosphere to facilitate gas transfer. Surface aerators are mechanically robust but transfer less oxygen per kilowatt-hour in deep basins and can cause aerosol misting and thermal heat loss during freezing winter conditions.

Basin Hydraulic Flow Configurations

1. Complete Mix (CMAS):
   Influent + RAS ──► [       Uniform Mixed Liquor       ] ──► Clarifier
                      [ DO, F/M, and MLSS identical throughout ]

2. Plug Flow:
   Influent + RAS ──► [ Inlet ] ──► [ Middle ] ──► [ Outlet ] ──► Clarifier
                      High DO Demand              Low DO Demand
                      High F/M                   Low F/M

3. Step Feed:
   Influent ───────┬────────────┬────────────┐
                   ▼            ▼            ▼
   RAS ──► [ Stage 1 ] ──► [ Stage 2 ] ──► [ Stage 3 ] ──► Clarifier

• Complete Mix Activated Sludge (CMAS): Influent wastewater and RAS are distributed uniformly throughout the entire basin volume. This provides maximum dampening of toxic shocks, organic slug loads, and pH variations, but the uniform low substrate concentration can encourage low-F/M filaments.
• Plug Flow: Wastewater and RAS enter at the inlet of a long, narrow serpentine channel and flow longitudinally toward the discharge end without longitudinal mixing. At the head of the tank, food concentration and oxygen demand are intense; at the outlet, food is depleted. Diffuser density is often tapered (highest at the inlet, lowest at the outlet) to match this declining oxygen demand.
• Step Feed: Influent wastewater is introduced at multiple discrete locations along the length of the aeration basin, while 100% of the RAS enters at the very head of the tank. This configuration stores a high mass of solids in the first stage without subjecting the secondary clarifier to an excessive solids loading rate, providing excellent flexibility during peak wet-weather events.
• Contact Stabilization: Operates using two distinct tanks. In the Contact Basin (short hydraulic detention of 30 to 60 minutes), starved microorganisms rapidly absorb colloidal and soluble organics onto their surfaces. Mixed liquor is then settled in the clarifier, and concentrated sludge is pumped to a separate Stabilization Basin (4 to 6 hours detention) where the absorbed organics are slowly metabolized before returning to the contact zone.
• Sequencing Batch Reactors (SBRs): A fill-and-draw batch technology where equalization, biological oxidation, and secondary clarification all take place sequentially in a single tank through five distinct cycles: Fill, React, Settle, Decant, and Idle.
• Extended Aeration: Utilizes long hydraulic detention times (18 to 36 hours), high sludge ages (MCRT of 20 to 30+ days), and low organic loading. It produces very stable effluent and low net sludge yield, making it standard for package plants and oxidation ditches across North Carolina.

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4. Secondary Clarification: Clarification and Thickening

Dual Operational Functions

A secondary clarifier (final clarifier) is far more complex than a primary clarifier. While primary settling is strictly a physical gravity process, secondary clarification is an integral part of the living biological system. It must perform two simultaneous, non-negotiable functions:

1. Clarification: Producing an exceptionally clear effluent by allowing biological flocs to form a uniform sludge blanket that settles to the tank floor, leaving a clear supernatant with TSS typically under 10 to 15 mg/L to overflow the effluent weirs.
2. Thickening: Compacting the settled biological flocs along the clarifier floor into a concentrated sludge layer to produce Return Activated Sludge (RAS) at a concentration of 4,000 to 10,000 mg/L TSS, which is recycled to maintain the target biomass inventory in the aeration basin.

Return Activated Sludge (RAS) and Waste Activated Sludge (WAS)

• RAS Recycling: The return sludge flow rate ($Q_{RAS}$) is typically operated between 25% and 100% of influent plant flow ($Q$). If RAS pumping is too slow, solids accumulate in the clarifier, increasing the sludge blanket depth and risking washouts. If RAS pumping is too rapid, the sludge does not have time to thicken, recycling thin mixed liquor back to the aeration basin and scouring the clarifier blanket.
• WAS Withdrawal: Biological growth continuously produces new microbial cells as BOD is metabolized. To keep the microbial culture in balance and maintain process stability, an exact mass of excess sludge must be wasted daily via Waste Activated Sludge (WAS) pumps to solids handling facilities.

Secondary Clarifier Operational Problems & Troubleshooting

              SECONDARY CLARIFIER TROUBLESHOOTING

  [Rising Sludge / Sheets of Sludge] ──► DENITRIFICATION
  - Clumps of dark sludge with tiny gas bubbles
  - Nitrate (NO3-) converted to N2 gas in anoxic blanket
  - Solution: Increase RAS rate, lower sludge blanket depth

  [Pinpoint Floc / Ashing] ────────────► OLD SLUDGE / OVER-AERATION
  - Supernatant is clear, but fine pin flocs float over weir
  - High MCRT, low F/M, or high DO shearing floc
  - Solution: Increase WAS wasting, reduce aeration blower output

  [Billowing / Bulking Blanket] ────────► FILAMENTOUS BULKING / HYDRAULIC SURGE
  - Uniformly rising blanket, high SVI (> 150 mL/g)
  - Feathery flocs that refuse to compact
  - Solution: Chlorinate RAS (2-3 lbs Cl2/1,000 lbs MLVSS), check DO/nutrients

• Rising Sludge (Biological Denitrification):
  • Symptoms: Large, dark brown sheets or clumps of sludge rise to the clarifier surface, accompanied by fine gas bubbles breaking the surface. The underlying mixed liquor settles rapidly, and effluent pH may be slightly elevated.
  • Mechanism: Facilities that achieve complete nitrification convert ammonia ($NH_4^+$) to nitrate ($NO_3^-$) in the aeration basin. If the settled sludge blanket remains too long on the clarifier floor (anoxic conditions with zero DO), facultative heterotrophic bacteria use the nitrate oxygen, reducing nitrate into nitrogen gas ($N_2$). Insoluble nitrogen gas micro-bubbles become entrapped inside the biological floc matrix, buoying large clumps of sludge to the surface.
  • Corrective Action: Increase the RAS pumping rate to reduce sludge detention time in the clarifier; decrease sludge blanket depth; if nitrification is not permit-mandated, reduce aeration basin MCRT or lower DO to suppress nitrate formation.
• Ashing:
  • Symptoms: Tiny, gray-to-brown particle flakes floating on the water surface, forming a thin scum.
  • Causes: Early-stage denitrification or high grease concentrations combined with slow-growing Nocardia actinomycetes.
• Pin Floc (Pinpoint Floc):
  • Symptoms: The liquid between flocs is crystal clear, but tiny, spherical, non-settling floc particles (< 1 mm) pass over the effluent weirs.
  • Causes: Old sludge (excessively high MCRT, low F/M) undergoing advanced endogenous breakdown, or excessive mechanical shear from over-aeration ($DO > 4.0 \text{ mg/L}$).
  • Corrective Action: Increase WAS wasting to lower sludge age; reduce aeration blower output to lower DO into the 1.5 to 2.5 mg/L range.
• Billowing Sludge Blanket (Solids Washout):
  • Symptoms: The sludge blanket rises uniformly throughout the basin until it spills directly over the effluent weirs.
  • Causes: Severe filamentous bulking ($SVI > 150 \text{ to } 200 \text{ mL/g}$), peak wet-weather hydraulic surging, or gross under-pumping of RAS relative to solids loading.