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Wastewater W t t Microbiology Mi bi l Microscopic review and Fil Filaments t Steve Leach Novozymes Biologicals Disturbing Water Facts 10 million people die every year from dirty drinking water 40% of the worlds population has no access to basic sanitation (toilets & running water) According d to USEPA 17% off treated d water is lost l in lleaky k pipes 40% of US rivers are identified as heavily polluted (75% in China) population p will live with Its estimated that half the worlds p chronic water shortages by 2050 Underground aquifers are being depleted at higher rates than natural nat al processes p ocesses can replenish eplenish them AWWA Journal May 2010 issue Bacterial Enzymatic y Activity y small, soluble compounds Extracellular enzymes Intracellular enzymes CO2 & H2O Membrane-bound Membraneenzymes P ti Protein G Grease large molecules, molecules particulate or colloidal material • Microbiology Bacteria do vast majority of organic degradation. Generation Time: replication in PURE culture: Bacillus sp. (BOD eating bacteria) 20-30 minutes Nit ifi Nitrifiers 22-48 22 48 hours h Methanogens 10-30 days Protozoa Their are useful as bacteria predators. number and sophistication tell us what the state of the bacterial population is is. A Aeration ti b basin i Home of active biomass Biomass Growth Pressures Operator SRT Inert material pH Temperature Toxins Dissolved Oxygen Competition Substrate/ concentration Bacteria formation Zoogloea g Filamentous bacteria Free motile rod-shaped bacteria Floc’d bacteria Spored bacteria Encapsulated bacteria in India Ink Stain Brightfield View 100X Phase contrast View 100X Daily Microexams (on site) Should be performed at least 3X per week Limited in scope Find Funnyy Hungry g y Bugs g Floc Size, shape, density Filaments Fil Abundance, type Higher life forms Presence, abundance, diversity Bulk water Clean,, containingg inerts or bacteria When should a detailed microexam be performed? When daily microexams show signs of change. When treatment performance is poor or worsens. If foaming problems occur. When the plant is undergoing operational changes. For plant confirmation and confidence When everything is running smoothly* Objective progression 10X darkfield 40X Phase contrast 10X Phase contrast 40X Phase contrast 40X Phase contrast 100X Phase contrast w/oil immersion (Microlife / Indicator Organisms) Amoeba Flagellates Swimming ciliates Crawling ciliates Stalked ciliates Suctoria Rotifers Nematodes A change in the microlife can indicate a change in the wastestream and/or plant conditions. P t Protozoa Facts F t Single-celled organisms Feed heterotrophically heterotrophically* Various forms of motility Eukaryotes Approx. 65,000 currently identified* g or in colonies Occur single Some are parasitic Oldest identified- over 500 million years (Cambrian) (Cambrian)* Found almost everywhere P t Protozoa Facts F t cont.; t Reproduce asexually asexually* Some species are the cause of human diseases Many M species i form f cysts t International Society of Protistologists (Protozoologists) Single Cell- Amazing! Motility Reproduction Respiration Feeding Defense(encystment) Adaption Protozoa Amoebas Flagellates Free-swimming F i i ciliates ili t Crawling ciliates Carnivore ciliates Stalked Stalked ciliates Suctorians M t Metazoa F t Facts (regards to wastewater) Multicellular Slower growing Worms, rotifers, gastrotrich, etc… Evolved l dd during the h Precambrian b age ((545 million ll years ago)* )* Worms and higher have approx. 55 specialized cells Typically larger than protozoa Sexual and asexual reproduction Heterotrophic All are motile* Microexam Sample Fil Filament t Facts F t Filaments are chains of individual cells Filaments aid in Floc formation Filaments remove BOD Filaments grow in adverse conditions Filaments have no Boundaries Filaments grow in all types of WWT systems Filaments can create poor settling Filaments cost “Money” Fil Filament t environment i t Low Dissolved Oxygen yg (D.O.) ( ) Low Nutrients (N and P) Low Food to Mass ratio ((F/M) / ) or High g F/M / Septicity/Organic acids Readily degradable substrate Manual on the Causes and Control of Activated Sludge Bulking, Foaming, and Other Solids Separation Problems 3rd edition Jenkins, Richard, Daigger Lewis Publishers Filaments and associated causes Manual on the Causes and Control of Activated Sludge Bulking, Foaming, and Other Solids Separation Problems 3rd edition Jenkins, Richard, Daigger Lewis Publishers Low Dissolved oxygen Low F/M Septicity Oil and Grease Sphaerotilus natans, Type 1701, Haliscomenobacter hydrosis Type 0041, Type 0675, Type 1851, Type 0803 Type 021N, Thiothrix I&II, Type 0914,, Type yp 0411,, Type yp 0961,, Type yp 0581, Type 0092, Nostocoida limicola I, II, and III Nocardia spp., Microthrix parvicella, Type 1863 Nutrient Deficiency Type 021N 021N, Thi Thiothrix th i I and d II II, S. S T natans, N. limicola III, H. hydrosis Low pH Fungi Morphological Features Attached Growth: False Branching: True Branching: Cell Shape: Square q Spherical Disc-shaped Disc shaped Rectangular Rod Morphological Features Constrictions: Crosswalls: Sheath: Shape: Straight Curved Coiled N Nocardia di Oil and Grease Gram stained slide 1000X/ Bright field w/oil Nostocoida Limicola sp. Nutrient deficiency Organic acids F t Factors affecting ff ti “Chlorination” “Chl i ti ” Chlorine, Peroxide, or Chlorine Dioxide Handling and cost influence this decision Exposure to Filaments ( RAS vs. Basin) Where is the best possible application point Frequency of Exposure A minimum of 1X per day (2-3X preferred) Concentration based on filament type Certain filaments require higher concentrations and longer exposure time Residual organic material Will the BOD/COD be oxidized by available chlorine Closely monitor microscopically Look k for f cell ll changes h and d possible bl d damage Ch Chemical i l facility f ilit Chlorination Chl i ti 1,299 , m3//day y Activated sludge (pure oxygen) yg ) Thiothrix II N. limicola II N Type 1851 Type T 0041 Before Chlorination Last day of Chlorination Last day of Chlorination B f Before/ / Aft After Dense floc Too many solids Weak floc and filaments Problem #1 Refinery y ASU Solids S lid fl floating ti on surface of secondary Eff. Eff TSS violation i l i No lab testing available 1000X Phase Solution Problem Denitrification Increase wasting ( d (reduce totall inventory) Monitor ORP in 2nd S Sweet t crude d Encapsulated floc S Sour crude d Low phosphate O Opportunity t it crude d Hyphomicrobium Problem #2 Chemical ASU- Pure oxygen Scum S on surface f off secondary Eff. Eff TSS violation i l i No lab testing available 1000X Phase monocolonies Solution Problem Extreme Low F/M Institute Step-feed Inf. COD monitoring Purchase Purchase microscope 100X Phase Problem #3 Paper p Mill ASB Loss L off COD reduction No N llab b testing i available 400X Phase Solution Problem Enzyme addition for Pitch control BICHEM BICHEM 1005 addition to compensate for increased COD 400X Phase High ferric addition Unknown dump Chlorination of filaments Sulphide presence Nocardia Inter-floc bridging