1Microbiology of WastewaterTreatmentLecture 25Other Sources:http://www.splammo.net/JLbactsite.htmlBlack, Microbiology Principles and Exploration 7th EdMadigan, Brock Biology of Microorganisms, 11th Ed.Lecture Topics Why treat wastewater The wastewatertreatment plant Important microbialprocesses Microbial monitoringWhat is wastewater? Industrial sources:– Petrochemical, dairy, food, pharmaceutical,metallurgical, etc. Domestic sources:– Form households and non-industrial businesses Domestic sewage:– Sinks, toilets, and showers– US domestic sewage varies little from community tocommunity across the country– Our waste is not unique. We even flush our toilets atthe same time.What’s in domesticwastewater? Also called sewage: it looks likespent dishwater The chemists only care aboutthe organic content,specifically carbohydrates,fats, and proteins. Sewage is 99.9% water and0.02-0.04% solids Example: Washington DC, 200tons of solids per day isproduced– 40-50% is proteins, 40-50%carbohydrates, 5-10% fats What would happen if this wasreleased into the surroundingenvironment?http://nsm1.nsm.iup.edu/tsimmons2Environmentalconsequences of nottreating sewage: People can get sick frompathogen contaminatedwater. Big problem indeveloping nations. Releasing wastewaterdirectly into a streamleads to oxygen depletion.– This is caused by aerobicrespiration linked to highorganic carbon loads inthe waste.– Low oxygen in water cancause fish kills. The degree of oxygenconsumption in wastewatercan be quantified.– This is called theBiological Oxygen Demand(BOD)http://ian.umces.eduHow tomeasure BOD Five day bioassay for oxygenconsumption, BOD5 300 ml bioassay in special bottles. Uses an oxygen meter to measuredissolved oxygen (DO)consumption in a five day period The BOD5 is calculated by: BOD5 (mg/L) = D1-D2/P– D1= initial DO (mg/L) of thesample– D2= sample DO (mg/L) after 5days– P= decimal volumetric fraction ofsample used.• Domestic sewage: 150-200 mg/L• Milk processing/cannery waste: 5000-6000• Pulping operations: 10,000-15,000 mg/LWhy the different numbers?The need for wastewater treatment plants Goal of wastewater treatment:– Protect health– Preserve natural resources– Prevent ecological damage How to accomplishes thesegoals:– Use wastewater treatment plants(WWTP)– The WWTP removes energy-richorganic matter before dischargeinto the environment.– And uses technology toprevent/lower the occurrence ofwater borne diseases.3Sewage TreatmentPrimary treatment: Non-biological treatment Removes solids Waste has high nutrientload (eg C, N, S, and P)Secondary treatment: Decreases dissolvedorganic carbon (DOC) Uses biological treatment Aerobic and anaerobicsecondary treatmentOverview of sewage treatment plantMicrobiology of anoxic secondary treatment This is used for breaking downsolid waste. Done in an anoxic sludgedigester. Solids are complex polymerse.g. cellulose and fiber.– Microbes secret lipases,proteases, amylases, etc. Fermentation is the majormetabolism in this treatment Lots of methane is producedby methanogenic archaea. The methane is collected andused to generate electricity.Anoxic secondary wastetreatmentTo the landfill4Aerobic secondarywaste treatment:trickle filter Trickling filter is a bed ofcrushed rocks—the 1º-treated sewage is trickledover it. Lots of surfaces formicroorganisms to attach to. Complete mineralization ofwaste to CO2, ammonia,nitrate, sulfate, and phosphate Same process occurring in afish aquarium.www.unclestu.comAerobic secondary wastetreatments: activated sludge Air bubbled through wastewater. Bacteria form large flocs.–Zoogloea ramigera is oneof the key species thatforms a slime and is thebase of the floc. After the flocs form theyare allowed to settle out Filamentous bacteria cancause sludge bulkingproblems--sludge thickensAerobic Activated sludge• Much of the organic material binds to the floc and iseventually taken up by the microbial biofilm.• 95% of the BOD is reduced during this stage.Important microbes in thesewage treatment plant Nitrifying bacteria– Aerobes– Convert nitrogenouswaste into nitrate Denitrifying bacteria– Anaerobes– Convert nitrate to N2 Methanogens– Generate methane fromacetate– Or use H2 and CO2 tomake methane– Mostly archaea5Nitrifying bacteria Ammonia is converted into nitrate Ammonia has a high BOD because NH3 oxidationrequires oxygen. Two groups of microbes are involved:– Ammonia oxidizing bacteria (AOB)– Nitrite oxidizing bacteria (NOB) AOB oxidize NH3 to NO2- in two steps:– Ammonia monooxygenase (AMO)– Hydroxylamine oxidoreductase (HAO) NOB oxidize NO2- to NO3-– Uses the Nor enzyme complex Both AOB and NOB respire oxygenNitrification EnzymesAMO= converts ammonia to hydroxylamine (toxic)HOA = converts toxic hydroxylamine to nitriteNOR = nitrite oxidoreductaseNO3-NH3NH2OH NO2-NO2-1/2O2H2OH2O1/2O2H2OPMFAMO HOA NOR∆G˚’ = -275 kJ/mol ∆G˚’ = -76 kJ/mole-e-H+H+Ammonia Oxidizing Bacteria (AOB):(A,B) Nitrsomonas; (C,D) NitrosolobusNitrite Oxidizing Bacteria (NOB):(E, F) Nitrospira; (G, H) Ntirococcus6Sewage treatment andenvironmental monitoring Monitoring effluents and the surroundingenvironment is important (Why?):– Asses the efficacy of the treatment process– C, N, P, metals, microbes, effluent toxicity It is often too difficult to directlymonitor a specific pathogen or virus/phage(Why?). Instead, monitoring is usually done forindicator organisms.What is an indicator organism? An organism that can be readily cultured thatindicates the presence of a pathogenicmicroorganism or correlates to a health problem. Five criteria for an indicator organism:– Consistently present in feces and at higher concentrations thanpathogens.– Should not multiply outside the human intestinal tract.– Should be as resistant or more resistant than the pathogen toenvironmental conditions and to disinfection.– Easy to assay (culture and quantify) and differentiate from otherorganisms.– Environmental concentrations should correlate with pathogens ormeasurable health hazards.Common indicator bacteria Coliforms:– Facultatively aerobic, gram-negative, nonspore-forming, rod-shaped bacteria; ferment lactosewith gas formation at 35˚C within 48 hrs.– Usually enteric bacterial group (E. coli,Klebsiella,