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Prospects for Pathogen Reductions in Livestock Wastewaters: A ReviewVincent R. Hilla

a Division of Parasitic Diseases, National Center for Infectious Diseases, Centers for Disease Controland Prevention, 4770 Buford Hwy, MS/F-36, Atlanta, GA 30341-3724.

To cite this Article Hill, Vincent R.(2003) 'Prospects for Pathogen Reductions in Livestock Wastewaters: A Review', CriticalReviews in Environmental Science and Technology, 33: 2, 187 — 235To link to this Article: DOI: 10.1080/10643380390814532URL: http://dx.doi.org/10.1080/10643380390814532

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Critical Reviews in Environmental Science and Technology, 30(2):187–235 (2003)

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Prospects for Pathogen Reductions inLivestock Wastewaters: A ReviewVincent R. HillDivision of Parasitic Diseases, National Center for Infectious Diseases, Centers forDisease Control and Prevention, 4770 Buford Hwy, MS/F-36, Atlanta, GA 30341-3724

ABSTRACT: Swine, cattle, poultry, and other commercial livestock can be infected by numerouspathogenic enteric microbes that are also infectious for humans. Livestock waste generated byconcentrated animal feeding operations (CAFOs) therefore may contain human pathogens that caninfect humans exposed to them during direct contact with livestock waste or media contaminated withthe waste (e.g., surface water, ground water, food crops). This article reviews published literaturerelated to the detection and inactivation of human pathogens in flushed livestock waste(i.e., “wastewater”). Physical, biological, chemical, and energetic treatment technologies are dis-cussed, including research results demonstrating or indicating the potential efficacy of the treatmentapproaches for pathogen reductions in livestock wastewaters.

KEY WORDS: CAFO, animal waste, treatment, disinfection, constructed wetlands, lagoons.

I. INTRODUCTION

Since 1995, hurricanes and other substantial rain events have contributed to atleast six spills of greater than 1 million gallons from waste lagoons located onconcentrated animal feeding operations (CAFOs) in North Carolina, USA, as wellas the flooding of over 50 other animal waste lagoons. Water quality investigationsconducted following some of these events associated the lagoon failures with fishkills, increased enteric microbial concentrations in receiving waters, and shellfishbed closings.44,189 In addition to waste releases during lagoon failures and flooding,concerns have been raised regarding potential public health and environmentaleffects of the lagoon-sprayfield waste management systems typically used bycommercial livestock farms in North Carolina,322 including potential health effectsassociated with odors emanating from swine farms,259,260 ground water contamina-tion from lagoon liquid seepage and infiltration from sprayfields,114,282,315 surfacewater contamination from sprayfields,282 and eutrophication of nearby water bod-ies due to ammonia volatilization from lagoons.4 These environmental and publichealth concerns are often raised in reference to swine farm operations and lagoons,but waste storage ponds are also used frequently by dairy operations and cattlefeeding lots, and sometimes used by poultry and sheep farms.295

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Nonpoint source (NPS) pollution of surface water has been identified as themost important source of water pollution in the U.S.,301 leading the USEPA toproposed regulations for reducing NPS pollution from CAFOs. These proposedregulations, in addition to specifying actions to improve nutrient management byCAFOs, also include pathogen control as a waste management goal.296 In responseto concerns regarding the environmental and public health impacts of the lagoon-sprayfield system, the North Carolina State Legislature enacted a moratorium in1997 (N.C. House Bill 515) on the construction of new animal waste lagoons untilalternative waste management and treatment techniques can be identified. Numer-ous other U.S. states have taken legislative action to increase regulations on wastemanagement by CAFOs, although few identify specific technological standards foranimal waste treatment.297 A similar situation exists internationally, where mostregulations of commercial livestock production operations are focused on issuessuch as waste management plans, buffer zones surrounding swine farm operations,and appropriate waste loading rates to farm land. Some countries, such as Malaysiaand Taiwan, have established treatment technology requirements or effluent stan-dards, but these standards do not specify standards for the microbiological qualityof animal wastes.122 Although not directly addressing animal wastewaters, theWorld Health Organization (WHO) has established guidelines319 for the microbio-logical quality of wastewater used in “agriculture,” which has been defined asincluding pasture land and land used to grow crops for human or animal consump-tion. In the U.S., “sprayfields” used for land application of animal wastewaters aretypically used to grow fodder crops for animal consumption, but are also used togrow food for human consumption. The WHO guidelines recommend that waste-waters used to irrigate agricultural land contain one intestinal nematode egg(Ascaris and Trichuris spp. and hookworms) per liter of wastewater,319 whether thecrop produced is for humans or animals (Table 1). A fecal coliform concentrationof 1000 per 100 mL is recommended for wastewater used to grow crops likely tobe eaten uncooked, but no fecal coliform standard is recommended in the 1989WHO guidelines for irrigation of pasture or fodder-crop land. Research subse-quently published in the Bulletin of the World Health Organization25 suggests thata fecal coliform standard of 105 organisms per 100 mL would be protective of thehealth of agricultural workers and nearby communities exposed to spray-appliedwastewater. This research did not consider wastewater standards for risks posed tohuman health and the environment through NPS transport of pathogens from landapplication sites to water resources. A range of U.S. State and Federal microbio-logical standards have been established for the reuse of municipal wastewater andbiosolids, but have not been promulgated for the land application of animalmanures and wastewaters (Table 1).

Historically, livestock waste has been managed to minimize the transport ofnutrients and metals from land application fields by establishing and monitoringagronomic (nutrient) loading rates of animal waste onto land. Thus, little data areavailable regarding the presence and concentrations of pathogens or other enteric

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TABLE 1. Microbiological Quality Guidelines and Standards forApplication of Wastewater to Land

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microbes in livestock waste or waste management systems. These data are impor-tant because pathogens in animal wastes may survive waste storage, treatment, andland application and be transported from farms to water resources and nearbycommunities where they represent a risk of infection for people exposed to themeither through direct ingestion of environmental water or through contamination ofenvironmental waters that are used for drinking water or fishing. Between 1986and 1998, there were 136 outbreaks (and over 447,000 cases) of gastrointestinal(GI) illness in the U.S. attributed to the ingestion of drinking water contaminatedwith enteric microbes that can be found in both human and animalwastes11,135,176,180,181,213 (Table 2). In addition, 39 outbreaks and over 3000 cases ofGI illness in the U.S. were attributed to exposure to environmental surface water(e.g., rivers, lakes), which may have been contaminated by waste from animals(domestic or feral) or humans. It is likely that these data do not account for the vastmajority of GI illness cases associated with waterborne microbes, based on epide-miological research indicating that 1 to 10% of cases of GI illness are presented formedical attention and subsequently reported to public health agencies for account-ing.208,237,255

A. Prevalence and Human Health Risks of Zoonotic Pathogens

Commercial livestock can be infected with numerous enteric pathogens thatare also infectious to humans64 (Table 3). Cattle can be reservoirs for a wide rangeof zoonotic bacterial and protozoan pathogens. Campylobacter prevalence rates

TABLE 2. Waterborne outbreaks in the U.S. (1986-1998)

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have been reported to be 5 to 53% in dairy cattle.314 In a study of Northwestern U.S.cattle feedlots and dairy farms, E. coli O157:H7 was detected on all 12 farmsinvestigated and in 1 to 6% of the animals studied.125 In a nationwide study ofcommercial U.S. dairy operations, C. parvum was detected in 22% of calves.108

Other studies of cattle farms in North America have found herd prevalence ratesof 20% for C. parvum and 10 to 88% for G. lamblia.223,327 Of all the commerciallypopular livestock raised in the U.S., swine are the most physiologically similar tohumans, so much so that pigs have been proposed as potential “mixing vessels” forpotentially pandemic avian and human influenza viruses153 and are one of theleading candidates for xenotransplants (animal-to-human organ, tissue or celltransplants).81 Consequently, many enteric pathogens of swine are also infectiousto humans, including numerous bacteria (e.g., Salmonella, Yersinia, Campylobacter),protozoan parasites (e.g., Cryptosporidium parvum, Giardia lamblia), parasiticworms (e.g., Ascaris), and some emerging viruses (hepatitis E virus, Norwalk-likecaliciviruses).64,302 A study of Salmonella prevalence in swine herds in NorthCarolina found Salmonella spp. in at least one fecal sample at 24 of 29 swine farmsstudied (83%), and in 25% of fecal samples collected.71 C. parvum and G. lambliahave been found on U.S. and Canadian swine farms with a population prevalenceof 5 to 11% and 1 to 12%, respectively.223,256,327,328 A study of parasite prevalencein commercial swine herds throughout the U.S. found Ascaris at 70% of the farmsand in 13 to 68% of the pigs, depending on type and age.169

In addition to the research indicating the prevalence of zoonotic patho-gens on CAFOs, epidemiologic research indicates that these pathogens canbe transmitted to humans and cause illness. Cases of human Ascaris infectionoccurring in North America have been attributed to transmission from in-

TABLE 3. Enteric pathogens potentially present in livestock wastemanagement systems

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fected pigs.3 Research also indicates that swine farm workers and residentsare at an increased risk of infection with the protozoan parasite Toxoplasmagondii,312 which has been reported to infect 5 to 11% of swine in Iowa.335

While research suggests that humans on or in the near vicinity of swine farmsare at an increased risk of Toxoplasma infection, humans can only be infectedby oocysts shed by cats or present in undercooked meat, so the exposure of off-farm human populations to Toxoplasma occytes is not likely. Workers on swinefarms were also shown to be at increased risk of developing leptospirosis in anoutbreak in Missouri associated with exposure to infected pigs.52 A study inOntario, Canada, indicated that residents living in areas having high cattle densityhad an increased risk of infection with E. coli O157:H7,208 and a case of antibiotic-resistant salmonellosis has also been attributed to contact with cattle, on a ranch inNebraska.94

Limited information is available regarding off-farm risks to human health dueto exposure to waterborne pathogens from agriculture-related sources. The con-tamination of water resources with enteric microbes from livestock waste has beendocumented. The negative impacts of swine and poultry waste lagoon failures onthe microbiological quality of receiving stream water has been well documented,44,189

but these events can be considered the result of poor waste management byindividual farms under extreme weather conditions. However, other studies sug-gest that enteric microbes can be transported from CAFO waste managementsystems to on- and off-farm water resources under normal operations and moderateweather conditions. A study by the U.S. Centers for Disease Control and Preven-tion58 of swine farms in Iowa reported measuring C. parvum oocysts at concentra-tions of 9 to 15 oocysts/L in on-farm groundwater monitoring wells, as well as atconcentrations of 1200 to 2200 oocysts/L in farm lagoons. A study of dairy farmsin the northeastern U.S. reported that 9% of farm-associated stream samplescontained Cryptosporidium oocysts.271 Cattle waste has been implicated in a caseof E. coli O157:H7 diarrhoeal illness associated with well water.155 Dairy farmsusing different waste management techniques were associated with significantincreases in E. coli concentrations in adjacent streams in South Carolina.157 Runofffrom swine farm holding pens has been implicated in the contamination of riverwater with Yersinia enterocolitica.304 In laboratory and field investigations, surfaceand subsurface runoff from manure-treated soils has been shown to be capable oftransporting enteric microbes, including C. parvum oocysts and E. coli O157:H7to ground water and surface water.46,66,104,202 Thus, it is important for animal wastetreatment systems to reduce pathogen concentrations such that environmentalmedia and public health are not negatively impacted when the waste is applied toland. Few studies of livestock waste treatment systems have investigated theefficacy of the systems for reducing concentrations of frank pathogens(e.g., Salmonella, C. parvum). The majority of research on enteric microbe reduc-tions in animal waste treatment systems has relied on a few traditional indicatororganisms such as fecal coliforms and fecal streptococci, but there are numerous

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other candidate microbes that can provide additional insight into system treatmentefficacy and that may be more appropriate than traditional bacterial indicators forevaluating treatment efficacy for nonbacterial pathogens. An understanding of theconcepts underlying the use of microbial indicators and the array of proposedmicrobial indicators is important for comprehensive interpretation of treatmentsystem microbiological data.

B. Microbial Indicators of Pathogen Dynamics in Environmentaland Treatment Systems

Livestock can be infected with numerous bacterial, viral, protozoan, andhelminthic pathogens that are also infectious to humans. Ensuring that waste-water is sufficiently treated, or that environmental media are not contaminated,by analyzing samples for all potentially present pathogens is not feasible andwould be a time-consuming, technically challenging, and expensive effort ifeven just a few pathogens were assayed. Thus, it is desirable to analyzesamples for easily quantifiable microbes that can serve as models for the fateof pathogens having similar characteristics and dynamics in treatment andenvironmental systems. Many investigations focus on the fate and survival of“microbial indicators,” organisms that are indigenous to the gastrointestinal(GI) systems of warm-blooded animals and not considered to be frank patho-gens. Because they are natural inhabitants of the GI tract, their presence infecal waste is consistent and, depending on the microbe, they are present athigh concentrations. Enteric pathogens, in contrast, are only present in excretawhen an infected host animal is shedding the organism. In sewage and othercomposite fecal waste streams, the concentrations of enteric pathogens can beorders of magnitude lower than fecal indicator organisms due to dilution of theinfectious organisms in waste and wastewater from noninfected individuals. Inenvironmental waters and drinking water, the concentrations of enteric patho-gens can be expected to be much lower than in waste treatment systems. Thus,fecal indicator organisms are useful tools for characterizing the level of fecalcontamination of environmental media. They are also of interest in treatmentsystems as models for the presence, movement, and survival of enteric patho-gens, which can be difficult, time consuming, and expensive to detect andquantify. According to Bitton,21 the criteria for an ideal indicator organism are(1) it should be indigenous to the intestinal tract of warm-blooded animals,(2) it should be present when pathogens are present, and absent in uncontami-nated samples, (3) it should be present in greater numbers than the pathogen ofinterest, (4) it should not die-off or be removed at a greater rate than thepathogen in the treatment system, (5) it should not multiply in the environment,(6) it should be detectable using easy, rapid, and inexpensive methods, and(7) it should be nonpathogenic.

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Bacterial indicator organisms include total coliforms, fecal coliforms, E. coli,fecal streptococci, enterococci, C. perfringens, Bifidobacteria, Bacteroides fragilis,and Rhodococcus coprophilus. Total coliforms and fecal coliforms (a subset oftotal coliforms) are functionally derived groups of Enterobacteriaceae that fermentlactose either at 35°C (total coliforms) or 44.5°C (fecal coliforms). Both groupshave been widely used as indicators of enteric bacterial pathogen dynamics intreatment and environmental systems, although the total coliform group is notconsidered to be as specific an indicator of fecal contamination as fecal coliforms.85,186

Even so, U.S. National Primary Drinking Water Regulations still rely on totalcoliform detection for primary indication of fecal contamination of drinking water(40 CFR §141.63). However, for drinking water samples determined to be positivefor the presence of total coliforms, the federal drinking water regulations requireconfirmatory analysis of either fecal coliforms or E. coli. E. coli, a member of thefecal coliform group, is considered a more specific bacterial indicator of fecalcontamination than fecal coliforms because even the fecal coliform test detectssome nonfecal thermotolerant bacterial species (and E. coli are not generallythought to have non-animal environmental sources).83 E. coli concentrations infresh recreational water have been shown to be significantly correlated with anincreased incidence of swimming-associated GI illness;84,241 thus, they are used asambient water quality criteria for fecal pollution of fresh water.299 The Gram-positive fecal streptococci have been of interest for many years as indicators offecal contamination due to their high persistence in environmental systems relativeto coliforms and many bacterial pathogens, as well as their potential utility fordistinguishing between human and animal fecal pollution.62 The fecal coliform/fecal streptococci (FC/FS) ratio is reported to be less than 0.7 in animal feces andgreater than 4.0 in human feces, although it has been shown that the FC/FS ratiois affected by differential die-off rates of the bacterial groups and may only beuseful during the first 24 h following a fecal pollution event.107 Enterococci, asubset of the fecal streptococci that includes Enterococcus faecalis and Enterococ-cus faecium, are also of interest as environmentally persistent indicators of bacte-rial pathogens. Their concentrations in marine and fresh waters have been corre-lated with increased incidence of swimming-associated GI symptoms;47,49,50,241

thus, they are used as indicators for fecal pollution of marine and fresh waters.299

However, research on enterococci relationships with plants suggest that theseorganisms may exist naturally and reproduce on some plant species, therebymaking them inappropriate microbial indicators for certain systems.62,216,217

C. perfringens is a Gram-positive spore-forming anaerobic bacterium that hasbeen suggested as a possible indicator for the presence and removal of environ-mentally stable viruses and protozoan parasite cysts and oocysts (e.g., C. parvumand Giardia lamblia) in environmental waters and treatment systems.20,100,234

C. perfringens spore removals during sedimentation and filtration appear to corre-late well with removals of C. parvum oocysts and G. lamblia cysts in somestudies,233 but not in others.234 The usefulness of C. perfringens is limited for

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certain applications due to the high level of resistance of C. perfringens spores toenvironmental stress and disinfection.48

Bacterial indicators that have been suggested as being specific indicators ofhuman fecal pollution include sorbitol-forming bifidobacteria, a group of Gram-positive nonsporeforming anaerobes,156,193 and Bacteroides fragilis, an anaerobicbacterium that is a predominant species in human feces,143 but may die-off in theenvironment at a higher rate than enteric bacterial pathogens.95 However, one ofthe animals shown to harbor bifidobacteria are pigs.194,250 For the detection of fecalpollution from animals, it has been suggested that the bacterium, Rhodococcuscoprophilus, is animal specific (i.e., not typically present in human feces) and agood indicator for differentiating between human and animal fecal sources insurface water.156,193,225 A major drawback to the use of R. coprophilus is the lengthof the culture assay (16 to 21 days),225 although molecular methods for the detec-tion of Rhodococcus species are being developed that may increase the utility ofthis microbe as an indicator of fecal pollution.16

Viruses that have been studied as indicators of fecal pollution and treatmentefficacy include somatic coliphages, male-specific (F-specific) coliphages, and thephages of Bacteroides fragilis. Somatic and F-specific coliphages are viral patho-gens of coliform bacteria. They are differentiated by how the viruses infect theirhost bacteria, either by attaching to the bacterial cell wall (somatic coliphages) orto fimbrae (“male-specific,” F+ pili) protruding from the bacterial cell wall. Somebacterial hosts are susceptible to infection by either somatic or F-specific coliph-ages, and studies using these hosts typically refer to “coliphages” in the generalsense. Coliphages have been suggested to be good indicators for the presence ofenteroviruses in fresh water.270 Somatic coliphages are present in the feces ofhumans and animals at high concentrations.118,129 Somatic coliphages can be foundat higher concentrations in surface water and sediment than other viral indicatormicrobes6 and have been shown to be good indicators for the reduction of humanenteric viruses in municipal water treatment plants.234 The results of fresh waterstudies indicate that somatic coliphage concentrations show a general correlationwith degree of sewage pollution.152 However, it has been suggested that somaticcoliphages can infect environmental bacteria related to E. coli and multiply undercertain environmental conditons, thereby diminishing their usefulness as an indi-cator in these instances.118,263 F-specific coliphages are not likely to reproduce inthe environment because F+ pili are produced only at temperatures above 30°C, atemperature not often reached in sewage and ambient waters.118 F-specific coliph-ages have been detected in the feces of humans and numerous other animals,51,101,227

generally at lower concentrations than somatic coliphages.118,129 Although F-spe-cific coliphages can be excreted by humans and animals, there is evidence that F-specific RNA (FRNA) coliphages can be used to discriminate between human andanimal fecal pollution by classifying them into four groups based on their biologi-cal and physicochemical properties (e.g., serology). Phages in Groups II and IIIhave been predominantly isolated from human feces and domestic sewage.102

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Phages in Groups I and IV have been predominantly isolated from non-humananimals.227 However, the animal and human serogroups are not strictly specific;human waste has been shown to contain Groups I and IV FRNA coliphages, andsome animal wastes have been shown to contain “human-specific” serogroups(e.g., swine have been shown to harbor Group II FRNA coliphages).130,149 Molecu-lar hybridization and RT-PCR techniques are being developed that may increasethe utility of FRNA coliphages for fecal source tracking investigations.120,149,258

Whereas somatic and F-specific coliphages can be found in both human andanimal wastes, research indicates that the phages of Bacteroides fragilis may bespecific to humans.118,289 B. fragilis phages can be detected in sewage, fresh water,and marine water at appreciable concentrations288 and have been suggested to beas good a viral indicator of fecal pollution as somatic coliphages (and better thanF-specific coliphages) in marine water and shellfish.185 Although their concentra-tions in environmental waters are often very low, research suggests that theirdetection is evidence of fecal pollution from humans and not from animals.289

Other research indicates that B. fragilis phages are not as reliable an indicator offecal pollution as somatic and F-specific coliphages.7,156 Molecular methods arebeing developed that may improve the utility of B. fragilis phages for environmen-tal and treatment system investigations.240

II. ENTERIC MICROBE REDUCTIONS IN LAGOON SYSTEMS

CAFOs often manage animal waste by flushing waste from the productionfacilities using water, which is then collected in earthen basins typically referredto as lagoons (Figure 1). Lagoon storage has become a common managementtechnique for animal wastewaters because of the cost-effectiveness of lagoontechnology.97 Wastewater in these lagoons is not discharged directly to receivingwaters, but is periodically discharged to nearby farm land, typically by sprayingthe liquid (hence the term, “sprayfield”). Under normal conditions, nonaeratedanimal waste lagoons built and operated according to NRCS Code 359 (WasteTreatment Lagoon) are anaerobic and have hydraulic residence times (HRTs) of3 to 6 months or more. CAFOs typically use a single lagoon for waste storageand treatment, but lagoon systems at some farms may have two or more lagoonsin series.

Lagoons, often referred to as stabilization ponds, have been used for municipalwastewater treatment for decades.209 However, as opposed to animal waste lagoonsystems, municipal stabilization pond systems in the U.S. are typically multicell (inseries) flow-through systems having hydraulic residence times on the order of 60to 120 days.207 Lagoon systems consisting of a number of cells (e.g., three or more)in series have been shown to be more effective in reducing nutrients and pathogensthan single cell systems having the same volume and surface area.192 Multicelllagoon systems better approximate plug-flow reactor dynamics as additional cells

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are added in series,311 thereby allowing distinct biological ecosystems to developin each individual cell, which can more efficiently metabolize waste nutrients andcontribute to enteric microbe die-off.198

The treatment effectiveness of lagoons is influenced by numerous physical,chemical, and biological conditions, including sedimentation, pH, temperature,sunlight (irradiance), dissolved oxygen, and microbial metabolism and predation.Sedimentation is an effective process for gross removal of nutrients associated withlarge solid particles (e.g., manure pellets and animal feed), but is not likely to bea significant factor in the removal of most enteric microbes unless large residencetimes are provided.92,222 The exception to this general view would be sedimentationof helminth ova, which are large enough (e.g., 50 to 80 µm) to settle effectively inlagoon systems.9,195 The occurrence of elevated pH levels (above 8.5) in facultativeand aerobic lagoons, typically due to the biological activity of algae, has beenshown to increase the die-off of E. coli,230,235 Salmonella typhimurium,Campylobacter jejuni,235 and the Serratia marcescens bacteriophage,98 but may notbe solely responsible for the inactivation of enteric microbes such as enterococcior male-specific (F-specific) DNA coliphages.73 Neutrophilic bacteria, which aremost likely to be active in wastewater ponds, maintain their cytoplasm at pH values

FIGURE 1. Livestock waste-handling options.295

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between 7.5 to 8.0 under normal external pH conditions (i.e., circumneutral, pH 6to 8). An increase in pH above the optimal cytoplasmic pH range causes a decreasein proton differential across the cell membrane and thus a decrease in the energy-generating proton motive force. The production of acidic fermentation productsand the uptake of potassium ions (K+) in exchange for export of H+ are twomechanisms that bacteria have for acidifying their external environment andincreasing internal pH.30 However, enteric bacteria may be more sensitive toperturbations in internal pH, especially in the alkaline range.333,334

Increases in temperature are associated with increases in microbial metabolicactivity when they are within the tolerance ranges of the lagoon microflora(e.g., as occurs in mesophilic lagoons). Improved nutrient removal is typicallyobserved in lagoon systems at warmer temperatures,192,247 as is improved reductionof enteric pathogenic bacteria, enteric viruses, and coliforms.12,56 Laboratory stud-ies designed to investigate the role that temperature plays in increasing themicrobiocidal effects of other environmental conditions have shown that the die-off of fecal coliforms is more rapid at higher temperatures when cultures aremaintained at pH 9.0 instead of circumneutral pH levels.235 However, for lagoonsystems in tropical areas where seasonal temperature variations are small and highambient temperatures are sustained through much of the year, wastewater lagoonscan experience substantial stratification, leading to reduced performance and patho-gen inactivation.198 The effects of summer stratification are not likely to be asimportant in anaerobic lagoons (e.g., swine waste lagoons) relative to aerobiclagoons, where stratification leads to the creation of larger anaerobic zones andassociated lower levels of metabolic activity and biological treatment performance.

For lagoon systems open to the atmosphere, sunlight irradiance can be animportant factor in lagoon ecosystem dynamics and pathogen reduction. It is wellknown that UV radiation is absorbed by nucleic acids and can cause damage tothese molecules. Sunlight contains a wide spectrum of radiation wavelengths,including a portion of the UV spectrum, visible wavelengths, and a portion of theinfrared spectrum. Research indicates that UVB (λ = 280 to 315 nm) may be theprimary light wavelength range responsible for the inactivation of E. coli andF-specific DNA coliphages in stabilization pond wastewater, likely due to directabsorption and endogenous damage to DNA.72 Sunlight (radiant energy) canpenetrate water, but the depth of penetration is limited by absorption by watermolecules, dissolved organic matter, microorganisms, and inorganic particulates.70

The depth that radiant energy can effectively penetrate wastewater lagoons isdependent on the concentration of dissolved and particulate matter in the lagoonwastewater. The depth that UV radiation can penetrate into aerobic lagoons maybe 10 cm or less. In anaerobic lagoons, UV penetration is on the order of only afew centimeters.68 However, less energetic wavelengths (400 to 700 nm) can havebactericidal effects in the presence of dissolved oxygen and humics through anexogenous process referred to as “photooxidation”.68 Humic substances, which areubiquitous in wastewater, may act as sensitizers in stabilization ponds by creating

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toxic forms of oxygen through the absorption and transfer of light energy todissolved oxygen molecules. The bactericidal effects of this mechanism have beenfound to be more pronounced with elevated pH, likely due to the increasedabsorbance of radiant energy by humic substances at higher pH values. It appearsthat toxic oxygen species can inactivate fecal coliforms by damaging the cellmembrane of the microorganisms, thereby disrupting membrane transport pro-cesses and allowing toxic substances to enter the cells. At a pH of 7.5, increasedabsorbance of sunlight was found to have a greater effect on enterococci andF-specific RNA coliphages in unfiltered lagoon liquid than in filtered lagoon liquidand control buffer, but did not have a differential effect for F-specific DNAcoliphages or E. coli.73 At pH values less than 8.0, wavelengths longer than 440 nm(which penetrate farther into water) do not appear to be able to damage fecalcoliforms, but have been shown to inactivate enterococci.69,72 Thus, the bactericidaleffect of radiant energy would be expected to be greatest when pond water is clearand has elevated dissolved oxygen (DO) and pH values, conditions that are morelikely to be present in aerobic lagoons than in anaerobic lagoons (such as can beexpected to be present on CAFOs).68,69 In ponds where algal concentrations arehigh, the depth of radiant energy penetration will be reduced due to absorbance byalgal cells, but the bactericidal effects should still be present due to elevated DOand pH conditions. This view is supported by other research showing reductionsin total and fecal coliforms due to the interaction of radiant energy, pH, andDO.211,235,292 The effect of radiant energy on the inactivation of enteric viruses inlagoons has not been fully investigated, but it appears that sunlight irradiance isalso associated with increased rates of viral inactivation in lagoons.98

Oxygen is introduced into lagoon water through diffusion from ambient air andthe activity of photosynthetic organisms. When available, oxygen is preferentiallyused by microorganisms for energy generation because they can generate moreenergy through aerobic respiration (using O2 as an electron acceptor) than fermen-tation or anaerobic respiration (which rely on the use of alternate, less oxidativeelectron acceptors). Because oxygen diffusion into water is slow, the presence ofdissolved oxygen in wastewater lagoons is typically due to the photosyntheticactivity of algae. Toxic oxygen species can be produced, both chemically andbiologically, when dissolved oxygen is present. These toxic species include singletoxygen, superoxide (O2

–), peroxide (O22–), hydrogen peroxide (H2O2), and hy-

droxyl radical (OH•). Although many of these species are short-lived, they arehighly reactive and can cause oxidative damage to important cellular and viralcomponents. Data from lagoons indicate that the survival of fecal coliforms isenhanced under anaerobic conditions.172,198 However, other data indicate that in-creasing DO from 0% to saturation is not associated with increased die-off of fecalcoliforms in stabilization pond wastewater203 or in buffer solution at pH 9.0.235

Therefore, dissolved oxygen may not exert a biocidal effect on its own but, as otherresearch demonstrates, likely contributes to the increased inactivation of entericbacteria and viruses in conjunction with other environmental conditions

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(e.g., sunlight and pH). Thus, the inactivation rates of enteric bacteria in the upperlayers of stabilization ponds may be influenced by the creation of toxic oxygenspecies due to irradiance and increase with increasing DO concentration.68,73 Thedependence of radiant energy inactivation effects on DO concentrations in waste-water ponds appears to also hold for the inactivation of F-specific RNA coliphages,but not F-specific DNA coliphages.73

Wastewater lagoons are biological reactors, and as such the biodegradation oforganic materials and the reduction of pathogens is intricately connected to theinteractions of the various organisms comprising the biological community of thelagoon ecosystem. Biological factors influencing the survival of enteric bacterialin aerobic and anaerobic lagoons include natural die-off due to starvation andcompetition with other microorganisms, predation by microinvertebrates, and themicrobiocidal effects of toxins, antibiotics, and enzymes produced by lagoonorganisms. Microbial products excreted into the lagoon environment may alsocontribute to the inactivation of enteric viruses. In the animal host, enteric bacteriaare surrounded by high concentrations of the nutrients necessary for cell mainte-nance and growth. In a way, the water environment within an animal host can beconsidered to be highly eutrophic. Raw wastewater is also nutrient rich, but moredilute than the environment in the intestinal tracts of animals. As additionaltreatment is provided in lagoons, the concentrations of nutrients in wastewaterdecrease. When viewed through Monod growth kinetics, it can be reasoned thatenteric microorganisms have high nutrient requirements (ks values) because theirmetabolic systems are acclimated to the high substrate environment of the gas-trointestinal tract. When outside the host, for example, in an anaerobic swine wastelagoon, enteric microbes are exposed to a lower substrate environment in whichthey are less efficient in utilizing available substrates than other microorganismsthat most likely have lower nutrient requirements (and lower ks values). Thesituation is even more pronounced in aerobic lagoons because they tend to haveeven lower ambient nutrient concentrations. Thus, enteric microbes do not usuallyincrease in concentration in wastewater lagoons, but instead die-off due to lowcompetitiveness with other microorganisms. In addition, the weakened state thatenteric microorganisms may be in due to their inability to obtain essential nutrientsmay contribute to their injury and die-off when under stress from other environ-mental factors. Research on experimental ponds has shown that at the same reactorHRT, coliform and Salmonella die-off decreased as the organic loading to thereactors increased (i.e., as the nutrient level of the reactors increased).54 However,some researchers suggest that prior starvation can protect enteric bacteria from theeffects of hydrogen peroxide and other toxic oxygen species by activating cellulardefense mechanisms associated with oxidative stress.69,159

The predation of enteric microbes by invertebrates, such as the Cladocera,Moina dubia, and Daphnia magna, can also occur in wastewater lagoons,182

although the rate of predation is likely to be far less than the rate of die-off due toother causes. Predation may actually improve enteric bacterial survival in some

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aerobic lagoons if zooplankton (e.g., Daphnia) are present in sufficient numbersthat they can feed on and decrease the population of algae in the pond such thatbactericidal pH and DO levels are reduced. Numerous other studies comparingenteric bacterial reductions in sterilized vs. nonsterilized water suggest that theincreased relative die-off of enteric bacteria in nonsterilized water may be due topredation or competition.205 However, while it may be true in certain situations thatpredation and competition are responsible for enteric bacterial die-off in nonsterilizedwater, it may be that the differences observed in sterilized vs. nonsterilized waterare due to the increased and/or consistent presence of microbiocidal compounds(e.g., antibiotics, toxins, enzymes) produced by microorganisms in the nonsterilizedwater. Antibiotics are chemical substances produced by microorganisms(e.g., bacteria and filamentous fungi) during fermentation, which can damage themetabolic activity of bacteria, including enteric bacterial pathogens and coliforms.The Gram-positive bacteria Actinomyces have been shown to produce antibioticsantagonistic to Salmonella.39 Toxins are proteins typically produced by bacteriaduring invasion and colonization of animal hosts, but can be found in environmen-tal samples (e.g., the various clostridia neurotoxins, and cholera enterotoxin). Thedegradation of organic wastes in lagoons depends on the release of enzymes(e.g., protease, catalase, phosphatase, urease) into the lagoon liquid.146,294 Proteaseactivity, which has been associated with the inactivation of enteric viruses,63 has beenshown to be optimal at pH 8.0, but also reduced in deep ponds.103,145 Anaerobiclagoons tend to be deep (10 to 15 ft) and are likely to have pH values in the rangeof 8.0.137,139 It has been shown that algae produce antibacterial substances,74,154 so theeffects of bactericidal algal products are more likely to be observed in aerobiclagoons than in anaerobic lagoons (where algae are not present or are present at lowlevels). Other research has shown that algae and other microorganisms in stabiliza-tion pond liquid can significantly reduce poliovirus concentrations.275 Algal activitymay be more of a factor in lagoons treating flushed swine waste than those treatingcattle waste based on research indicating that swine waste is a better culture mediumfor algae than are dairy or beef cattle wastes.137 Investigations into the inactivationof G. lamblia and hepatitis A virus in septic tank effluent mixed with swine wasteindicate that biological activity in the swine waste was responsible for the increaseddie-off of the microbes in the mixed waste compared to septic tank effluent alone.77,78

While the same general physical, chemical, and biological processes occur inswine waste lagoons and municipal waste lagoons, swine lagoons are designed toachieve wastewater quality sufficient for subsequent application to crop land.Municipal lagoon systems typically discharge to surface water and have far morestringent treatment requirements. Because animal waste lagoon design and landapplication standards are based on solids, metals, and nutrient reduction, there iscurrently little data indicating the levels of enteric pathogens and other microbesin CAFO lagoon liquid. In municipal waste lagoon systems, it has been proposedby Marais198 that fecal bacteria die off in a series of n identical, completely mixedponds according to the first-order equation:

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NN

k teT

n=+

0

1( )

where Ne and No are effluent and influent concentrations, respectively; t is the HRTin days; and kT, the first-order decay constant for fecal coliforms removal. Maraisproposed that kT for fecal coliform die-off can be estimated according to theArrhenius equation, with the rate constant at 20°C, k20, equal to 2.6 and theArrhenius constant, θ, equal to 1.19, within a temperature range of 2 to 21°C:

kTT= −2 6 1 19 20. ( . )

Other values of k20 and 2 have been proposed203,210 that are lower than thevalues of 2.6 and 1.19, respectively, proposed by Marais.198 According to theMarais model, a 90% reduction of fecal coliforms can be expected in each lagooncell (at an ambient temperature of 20EC) having an approximate HRT of 3.5 days.These equations have been supported by some research,197 but other models havebeen proposed that do not incorporate temperature,8 or that incorporate additionalparameters such as solar intensity, algal concentration, pH, pond depth, reactordispersion number, and influent chemical oxygen demand (COD).203,238 In addi-tion, the use of baffles in stabilization ponds has been shown to increase removalrates for solids, nutrients, and enteric bacteria.160,218 For the removal of helminthparasite eggs in stabilization ponds, an empirical first-order equation dependentonly on HRT has been proposed.9 In municipal waste systems, fecal coliformreductions of 92 to 99% have been reported in stabilization ponds having HRTs of5 to 20 days (at average ambient temperatures of 26EC).196 Somatic coliphageshave been reported to be reduced by 80 to 90% in stabilization ponds having HRTsof 8 to 20 days and average temperatures of 30°C.224 In CAFO lagoon systems,indigenous enteric microbe and pathogens have not been sufficiently quantified ortheir inactivation kinetics characterized to make comprehensive evaluations of theefficacy of lagoon treatment for reducing pathogens. Research has been conductedat a swine nursery in North Carolina that indicates that concentrations of fecalcoliforms E. coli, enterococci, somatic coliphages, and F-specific coliphages influshed swine waste can be reduced by 90 to 99% in a single-stage anaerobiclagoon.142

III. ALTERNATIVES TO LAGOON TREATMENT

Alternatives to lagoons can employ biological, chemical, physical, or energeticprocesses. Traditionally, physical and biological treatment techniques have beenfavored by CAFOs because they are relatively inexpensive and can achieve suffi-

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cient nutrient reduction for application of the treated waste to farm land at agro-nomic rates. Chemical treatment may be used for the removal of solids or nutrients(e.g., through chemical coagulation) or for disinfection (e.g., chlorination, ozonation).Energetic treatment techniques have been used primarily for the inactivation ofpathogens in wastewater (e.g., UV disinfection, gamma irradiation, electrolysis).

A. Physical Treatment Techniques

Physical treatment techniques include sedimentation, screening, and filtrationto separate larger solids particles from bulk liquid. Filtration (e.g., sand filtration)is not a common physical separation technique for animal waste managementsystems. The primary purpose of sedimentation and screening in animal wastemanagement systems is to remove solids from the bulk liquid, thereby reducing theorganic load to subsequent treatment systems (to improve their potential perfor-mance) and minimizing operation and maintenance problems. Increasingly, solidsseparation is of interest for the recovery of biosolids as commercial fertilizer.Pathogens can be expected to be only slightly reduced in swine wastewaterfollowing sedimentation or screening. In a sedimentation column experiment withswine waste slurry, reductions of Salmonella typhimurium, E. coli, Streptococcusfaecalis were 68, 80, and 0% after 48 h of settling (and also reflecting reductiondue to die-off)222 (Table 4). Little additional data are available to estimate potentialenteric microbe reductions in solids separation units for animal waste. In municipalwaste systems, many studies have been performed investigating pathogen remov-als in primary sedimentation tanks. In general, removals of viruses, bacteria, andprotozoa can all be expected to be less than 90%, with slightly higher removals ofhelminth ova possible.92,109,316

B. Biological Treatment Techniques

Alternative biological treatment techniques utilize the same fundamental pro-cesses described for lagoons and stabilization ponds. Biological treatment tech-niques that have been investigated for swine and other animal wastes include:constructed wetlands, overland flow, anaerobic digestion, aeration/aerobic diges-tion, and activated sludge systems (Table 5).

1. Constructed Wetlands

Constructed wetlands have been receiving substantial interest for the treatmentof swine, dairy, and poultry waste based on a well-established record of goodtreatment performance for municipal waste163 and anticipated cost-effectiveness

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for animal waste treatment.53,65,87,124 Wetlands are land areas where the surface soilis saturated during all or part of the year. Because of the presence of both aerobicand anaerobic biological activity during seasonal flooding and drainage, naturalwetlands have been shown to be possibly the most biologically productive ecosys-tems on Earth.147 Constructed “treatment” wetlands are engineered wetland sys-tems for the improvement of water or wastewater quality. Designs of constructedwetlands are increasing as our knowledge improves of how these systems function,but they essentially consist of surface flow (SF— e.g., “typical” free water surfacewetlands and pond-wetland combination designs) and subsurface flow (SSF—e.g., horizontal flow, vertical flow, “root-zone method,” and reciprocating) hy-draulic designs. Numerous biological processes occur in constructed wetlands thataffect the removal of nutrients and pathogens in wastewater. As with stabilizationponds, biological activity in constructed wetlands consists of both aerobic metabo-lism and a wide spectrum of anaerobic processes under different oxidation-reduc-tion conditions. Surface flow wetlands are typically designed to have a small “freewater” zone (about 30 cm) overlying the subsurface soil in which the wetlandvegetation is growing, thereby creating a large specific surface area (surface area/volume water) that takes advantage of the tendency for microbial communities tobe more dense and metabolically active on solid surfaces where nutrients accumu-late.2,170,239 Subsurface flow wetlands take advantage of this biological principle byeliminating the bulk surface water zone and maximizing the contact betweenwastewater and biofilms covering the subsurface reactor media.13 Lagoons havecomparatively far less specific surface areas than SF or SSF constructed wetlands.Constructed wetlands also utilize the additional biological activity of plants, whichhave been shown to improve the performance of the treatment wetland systems byimproving reactor hydraulics through their physical presence36,123 and increasingmicrobiological activity by providing additional surface area for microbial growth.121

In addition, the oxygen released into the subsurface by wetland plants during theaeration of their root systems is used for subsurface aerobic metabolism andnitrification/denitrification.246 SSF wetlands are designed to maximize the expo-sure to wastewater to the biological activity associated with wetland vegetationroot systems, referred to for some designs as the “root-zone method”.33

As discussed for stabilization ponds, the presence of different physical,chemical, and biological conditions in constructed wetlands will affect the sur-vival and removal of enteric pathogens in these treatment systems. Physicalremoval of solid particles (including microbes) occurs in constructed wetlandsthrough sedimentation and filtration, a physical process not present in lagoonsystems.162,257 Filtration is especially important in SSF systems, which are de-signed to maximize the contact between wastewater and subsurface gravel me-dia, because the subsurface media and biofilms can act as a filter for wastewatersolids and microorganisms.163,321 Sometimes the filtration process works toowell, because clogging of SSF systems can be a problem.23,178 The efficiency offiltration for removing wastewater particles or microbes can be expected to be

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dependent on the size of the particles and microbes,183 as well temperature204 andhydraulic loading rate.257 Because the flooding of soils tends to cause pH conver-gence (i.e., acidic soils increasing in pH and alkaline soils decreasing in pH),treatment wetland pH values tend to be circumneutral (pH 6 to 8) when influentsare not strongly acidic or basic.91,163 In general, the circumneutral pH values ofhuman and animal wastewaters suggest that pH is not a major parameter forpathogen reductions in wetland systems treating these wastes. As with pH,temperature and DO also tend to be buffered in constructed wetland treatmentsystems.163 Increased temperature can improve the performance of constructedwetland systems in the same general way that stabilization ponds are affected, byincreasing microbial activity.220,245,324 However, the relationship is not alwaysapparent.138,188,277 In a study of a SF constructed wetland system treating dairywaste, researchers reported that fecal coliform reductions were significantlylower in winter than during summer conditions.220 Other studies support thepositive relationship between increased ambient temperatures and enteric mi-crobe removal, but the effect is not always apparent.248 Sunlight irradiance islikely to not be as important in constructed wetland systems as for lagoonsbecause of light attenuation caused by the canopy of emergent wetland shoots.35

Thus, algal growth and the microbiocidal effects of sunlight are diminished intypical wetland systems, unless they incorporate open-water pond zones.

Emergent wetland vegetation also has the effect of diminishing wind velocityover the wetland surface, thereby decreasing oxygenation of the water surface.35,36

However, research suggests that wetland vegetation is responsible for the activeaeration of the wetland subsurface through oxygen leakage from roots into therhizosphere,214 as well as passive aeration when wind blowing across dead andbroken shoots forces atmospheric air through the shoots into the underground rootsystem.34 Research differs as to whether the presence of wetlands vegetationcontributes to the improved performance of constructed wetlands. Most controlstudies of planted and unplanted wetland cells show that the presence of vegetationcan have a positive impact on the reduction of nitrogen compounds (e.g., totalnitrogen, total Kjeldahl nitrogen (TKN), ammonium) and biochemical oxygendemand (BOD) and/or COD.42,111,254,278,285,286,323,332 Oxygen release in the root zoneof wetland plants also appears to reduce methane emissions from constructedwetlands.284 Wetland vegetation activity may not, however, have an effect onsuspended solids removal.10,285 Research on the relative reductions of entericmicrobes in planted and unplanted wetlands indicates that the presence of wetlandvegetation can increase the removal of F-specific coliphages,113 E. coli,309 fecalcoliforms, fecal streptococci, and C. perfringens,278 but that the apparent effect onenteric microbes is not consistent.252,285 In addition to providing vegetation toincrease subsurface oxygenation of wetland soils, other constructed wetland de-signs have been investigated to take advantage of the treatment benefits of subsur-face oxygenation, including “upland-wetland” systems where unsaturated uplandmounds are included for wastewater treatment,148 and “recirculating” subsurface

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flow systems, where water levels are changed as the reactors are cyclically drainedand filled.15,168

In addition to the biological activity of wetlands vegetation, microbes withinwetland soil and gravel biofilms play a major role in chemical transformations andpathogen reductions. In vegetated wetlands, microbial populations of bacteria,actinomycetes, and fungi have been shown to be present at much higher concen-trations than in nonvegetated cells.128 As in stabilization ponds, these microbes canaffect pathogen survival in wetlands through predation and the excretion of anti-biotics, toxins, and enzymes. Enhanced development in the rhizosphere of popu-lations of bacteria having antibiotic activity has been documented.37 Wetlandvegetation has also been shown to exude antibiotics that can have antagonisticeffects on pathogens.264 Ciliated protozoa from vegetated treatment wetlands havebeen observed to consume E. coli at faster rates than protozoa from a nonvegetatedsoil bed, suggesting that ciliate grazing plays a role in E. coli removal in wetlandsystems.76 Ciliated protozoa common to wetlands have also been shown to becapable of ingesting C. parvum oocysts.283 However, protozoan predation activitycan be diminished in subsurface wetland media if pore sizes are small enough(≤6 µm) to physically protect the microbes from ingestion by the protozoa.325

Constructed wetland designs have typically assumed that expected treatmentperformance can be modeled using deterministic first-order models that incorpo-rate reaction rate constants for the various processes and parameters discussedpreviously. A commonly used form of these first-order models for pollutantreduction assumes plug flow conditions in the constructed wetland:

ln*

*

C C

C C

k

q

−−

= −0

where C0 is the influent concentration, C is the effluent concentration, C* is thebackground concentration (if known or applicable), k is the first-order areal rateconstant (cm/d) and q is the hydraulic loading rate (cm/d).163 Values for first-orderreaction rate constants for various parameters (e.g., BOD, TSS, ammonium, fecalcoliforms) in constructed wetlands are typically calculated as has been discussedfor fecal coliforms in stabilization ponds. Changes in loading rate and HRT havebeen shown to affect wetland treatment performance for reduction of BOD andCOD,279,285,287 total nitrogen,43,286,287 and fecal coliforms.119,171,285,287 The inclusionof dispersion into the first-order decay model has been suggested for improving theprediction of the fate of enteric viruses60 and fecal coliforms171 in constructedwetlands. An alternate approach to using first-order models is the use of regressionequations to account for the effects of multiple parameters on treatment perfor-mance.162 However, the usefulness of first-order rate models and regression modelsare limited for the prediction of treatment performance due to intersystem variabil-ity and the variability of rate constants and threshold/background concentrations in

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response to changes in hydraulic loading and influent concentrations.164 Nonlineardecay rates may also occur for pathogen removal if the compositional character ofthe pathogen population differs in system influent relative to system effluent, asmay be the case if there is a high percentage of injured microbes in the influent thatmay die-off at a faster rate than other members of the population.309

Published reaction rate constants indicate that at the same loading rates and/orHRTs the treatment performance of SSF wetlands can be expected to be better thanin SF wetlands.163 The higher treatment efficiencies reported for SSF wetlands arelikely due to increased filtration, higher specific surface area, and increased expo-sure of wastewater to rhizosphere biological activity. Reductions of fecal coliformsin SSF wetlands have been reported to be between 90 and 99.9% at HRTs of 2 to11 days.228,287 At hydraulic residence times of 5 days, seeded MS2 (an F-specificcoliphage) and human poliovirus type 1 were removed by approximately 99.9 and99.98%, respectively, in vegetated SSF gravel reactors treating municipal waste-water.112 Average removals of 99% of indigenous F-specific coliphages wereachieved in a SSF wetland having a nominal HRT of 5 days.113 In a study of parallelmunicipal waste treatment SF and SSF wetlands, the SSF wetlands (having anominal HRT of 5 days) reduced fecal coliforms, Giardia cysts, Cryptosporidiumoocysts, and coliphages by 98, 88, 69, and 95%, respectively. At a nominal HRTof 4 days, the SF wetlands reduced fecal coliforms, Giardia cysts andCryptosporidium oocysts by 93, 73, and 58%, respectively.110 Free-living amoebaewere removed by an average of 82% in different SSF wetland cell designs formunicipal sewage treatment.251 Helminth ova have been reported to be removed byapproximately 90% in SSF wetlands at short HRTs (1 to 4 h).190 These resultssuggest that protozoan parasites may not be substantially reduced (<90% reduc-tion) in SF or SSF treatment wetlands compared with viral and bacterial microbes.However, research on pilot-scale SSF wetland reactors indicates that the protozoanparasites Giardia and Cryptosporidium can be removed to a similar, or greater,extent (93 to 97%) as for fecal coliforms (91 to 95%), coliphages (86 to 91%), andenteric viruses (91 to 96%).242 Although research has shown that enteric microbesin wastewater can be substantially removed by constructed wetland treatmentsystems, it should be kept in mind that removal efficiency can be affected by theinfluent concentrations of these microbes,164 and that apparently negative removalefficiencies are possible if influent concentrations are below background/thresholdconcentrations.291,303

An analysis of fecal coliform reductions from over 30 constructed wetlandsystems summarized in the Livestock Wastewater Treatment Database (LWDB)estimated that the average reduction of fecal coliforms in constructed wetlandstreating livestock wastewaters was approximately 92 to 97%.59,173 In a study of theeffects of changes in hydraulic loading on treatment performance of a SSF wetlandtreating dairy wastewater, fecal coliforms were reduced by 95 to 99.6% at nominalHRTs of 2 to 7 days, respectively.285 A study of a SF wetland treating dairywastewater in Connecticut measured an overall fecal coliform reduction of 98% in

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the system at an average HRT of 41 days.220 In a study of a SF wetland systemtreating dairy wastewater in Arizona, fecal coliforms and Listeria monocytogeneswere reduced by 83 and 99%, respectively.165 In a later study at this site, coliphageswere reported to be reduced by 95% in the wetland system at a nominal HRT of4 days, but the effectiveness of the system for reducing fecal coliforms andL. monocytogenes was much lower (13 and 32%, respectively).166 A study of a SFconstructed wetland system treating effluent from a fish production pond thatreceived flushed swine waste reported reductions of fecal coliforms of 87 to 92%at HRTs of 2 to 5 days.187 In a study of a two-cell SF constructed wetland treatinglagoon liquid at a swine nursery, reductions of Salmonella, fecal coliforms, E. coli,and C. perfringens spores were 96, 98, 99, and 97%, respectively. Somatic andF-specific coliphage viral indicators were reduced by 99 and 98%, respectively, inthe constructed wetland system. Enterococci were reduced by only 87% and weredetermined to not be reliable indicators of the microbial reduction efficacy of thewetland system.140 Hill and Sobsey140 also reported significantly higher fecalcoliform, E. coli and Salmonella typhimurium reductions in a laboratory-scalevegetated SSF reactor treating swine lagoon liquid than in a similarly loaded SFwetland reactor and a nonvegetated SSF control reactor.

2. Overland Flow

Overland flow (OF) has been in use for over 20 years for the treatment ofmunicipal wastewater, primarily from small communities. By the 1980s, thetechnique had been established sufficiently that the U.S. EPA published designstandards for the construction and operation of overland flow systems.300 The OFprocess typically consists of the application of wastewater to the upper portion ofa sloping, grass-covered field of low permeability soil. Wastewater is allowed toflow as a sheet through the grass (i.e., no standing water) to runoff collectionditches at the bottom of the slope. The treatment mechanisms for OF systems aresimilar to those utilized in constructed wetlands systems: filtration and adsorptionin the subsurface matrix and emergent vegetation, nitrification primarily in theoxidized water film and denitrification in the subsurface soil, and the varioustreatment effects that result from the presence of a wide range of oxidation-reduction conditions in the surface and subsurface zones.150 Typical hydraulicloading rates are 2 to 10 cm/d,300 but with wastewater applications only occurringfor a period of time each day (e.g., 6 h/day). Treatment performance can be affectedby season (e.g., temperature, precipitation), as well as hydraulic loading rate(HLR) and system slope. At a HLR of 32 cm/2 h daily application and slope of 5%,reductions of COD, TKN, and total solids concentrations have been shown to beminimal: 40, 9, and 29%, respectively.131 At a lower HLR (1.25 cm/6 h) and slope(2%), treatment of effluent from a municipal facultative lagoon was better, withreductions of BOD, TKN, and TSS concentrations of 54, 80, and 60%, respec-

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tively.151 Hunt et al.151 found that background concentrations of fecal streptococciin a control OF cell were quite high (~30,000 CFU/100 mL), approximately equalto the effluent concentrations from the treatment OF cell and about 10 times higherthan in the municipal waste lagoon liquid entering the cell. Fecal coliforms werealso shown to be an unreliable indicator in the OF system due to substantialseasonal variations in their concentrations in OF system effluent, with effluentconcentrations during the summer being over an order of magnitude higher thaninfluent concentrations (100,000s CFU/100 mL vs. 10,000s CFU/100 mL). Astudy of a pilot-scale OF cell at a swine nursery in North Carolina reportedreductions of fecal coliforms, E. coli, enterococci, somatic coliphages, andF-specific coliphages to be 60 to 75% when the cell (sloped at a 2.5% grade) wasloaded with swine lagoon liquid at a rate of 3 cm/d.142

3. Anaerobic Digestion

Anaerobic digestion is a well-established treatment method for animal wastesand domestic wastewater sludge and is often used when a design goal is theproduction of methane gas for energy recovery. Sometimes the energy recoveredis used to heat anaerobic digesters to create consistent mesophilic (~ 35°C) orthermophilic (> 50°C) digestion conditions. In general, considering the energyinputs needed to maintain thermophilic reactor conditions, thermophilic digestioncan be expected to have greater operational costs than mesophilic digestion. Fixedmedia is also used in some designs (referred to as “fixed-media anaerobic digest-ers” or “anaerobic filters”) to increase the reactor surface area for biofilm growth,thereby improving reactor treatment performance, allowing for decreased reactorvolume, or allowing increased throughput flow rates. When considering funda-mental treatment processes, anaerobic digesters differ from typical CAFO wastelagoons primarily in that they are not open to the atmosphere and therefore do notallow for any aerobic biodegradation activity or enteric pathogen reduction throughmany of the environmental processes discussed previously (e.g., dissolved oxygentoxicity, sunlight irradiation, elevated pH due to algal activity). Depending onreactor design (e.g., presence/number of baffles, presence of fixed media), meso-philic anaerobic digestion can be expected to achieve reductions of COD and TKNof 60 to 80 and 40 to 60%, respectively, in swine waste at HRTs of 1 to 15days.28,29,329 Pathogens and enteric microbial indicators have been shown to besubstantially reduced during anaerobic digestion, with the magnitude of the reduc-tions affected primarily by reactor temperature, but also by pH, HRT, and volatilefatty acid (VFA) concentration.86,133,308 Mechanisms for pathogen inactivationduring anaerobic digestion include chemical toxicity, thermal inactivation, preda-tion, and microbial antagonism through the production of antibiotics, enzymes, andtoxins. During the anaerobic digestion of swine waste, VFAs can reach concentra-tions sufficient to exert biocidal effects on bacteria, although the effect is more

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pronounced at pH values (pH 4 to 5) not likely to be present under typicalanaerobic digestion conditions.133,136,177 Volatile fatty acids cause damage to bac-teria by inhibiting active transport of amino acids and other molecules.99 At higherdigester pH values (pH ≥ 8.0), ammonia is more likely to be present in itsuncharged form and has been shown to be toxic to enteric viruses306,307 and C.parvum oocysts.158 One seemingly protective factor for enteric bacteria duringanaerobic digestion is antibiotic resistance. Antibiotic-resistant bacteria have beenshown to have lower inactivation rates in anaerobic digesters treating swine waste.1

Fecal coliform reductions have been shown to increase in anaerobic digestersas temperatures increase, ranging from 0.3 to 2 to 3 to 4 to >4 log10 at temperaturesof 28, 35, 50, and 55°C, respectively.17,82,167,201,222 An anaerobic filter treating septictank effluent at a HRT of 5 days was found to reduce fecal coliform concentrationsby approximately 90% when reactor temperatures were 10 to 20°C.67 Culturableenteric viruses in domestic sludge have been found to be inactivated by 1.1 log10

when anaerobically digested for 20 days at a temperature of 35°C, and by 3.3 log10

when anaerobically digested for 20 days at 49°C.17 Mesophilic anaerobic digestionappears to be effective for the inactivation of protozoan cysts and oocysts (e.g.,3 log10 reduction after digestion for 24 h at 37°C), but thermophilic temperaturesmay be required for effective inactivation of parasitic ova like Ascaris.21 Otherresearch indicates that the viability of C. parvum oocysts may be reduced by only80% after 3 days of anaerobic digestion at 35°C.318 In an anaerobic digester fedcattle slurry at an operating temperature of 28°C and HRT of 24 days, S. typhimurium,Yersinia enterocolitica, L. monocytogenes, and Campylobacter jejuni reductionswere modest: 0.7, 1.4, 0.9 and 0.1 log10.167 A study of anaerobic digestion of pigslurry at 35°C and HRT of 1 to 4 days measured higher S. typhimurium reductions(0.8 to 1.3 log10).222 Another study of a swine waste anaerobic digester found thatthe F-specific coliphage f2 was reduced by 2 to 4 log10 at 37°C and a HRT of 10to 18 days.201 In an ambient-temperature in-ground “covered lagoon” anaerobicdigester having a design HRT of 65 days, E. coli concentrations in raw wastewaterinfluent were reduced by 99% in digester effluent at a volatile solids loading of2000 lb/day.61 Research by Hill139 on this system reported similar reductions forfecal coliforms and E. coli, as well as reductions of 96, 96, and 98% for Salmonella,C. perfringens spores, and coliphages, respectively. A pilot-scale upflow anaerobicdigester treating cattle and swine abattoir wastewater was reported to achieve fecalcoliform reductions of only 18%, and fecal streptococci reductions of 95%, whenthe reactor temperature was between 20 and 30°C and the HRT was approximately2 days.127

4. Aeration and Aerobic Digestion

Aeration and aerobic digestion of livestock waste is done primarily for odorcontrol, nitrogen management, and biodegradation of waste organics, but questions

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regarding pathogen reduction effectiveness are increasing.24,45 Aerated biologicaltreatment processes can be substantially affected by changes in temperature andHRT.200,229,261 Enteric pathogens and indicator microbes have been shown to havehigher inactivation rates under aerated and aerobic conditions in pig and cattlewaste slurry than under anaerobic conditions at the same temperatures.132,215 Inmunicipal sludge treated by aerobic and anaerobic digestion, inactivation rates ofenteroviruses during aerobic digestion were more than twice the rates measuredduring anaerobic digestion.261 The differential in treatment and inactivation ratesunder aerated and nonaerated conditions is also likely to increase as waste tempera-tures increase within the tolerance range of the microbial community.14,132 Themicrobiocidal effect of aeration appears to be microbially mediated, becausecomparative studies of porcine enterovirus type 2 in distilled water and autoclavedslurry showed no difference in inactivation rate when the media were aerated andnot aerated.79 In general, mesophilic aerobic digestion can be expected to reduceenteric bacteria and viruses by 90 to 99.9%.21,90,200 Mesophilic aerobic digestion isnot effective for inactivating helminthic ova.199,236,249 The provision of solid mediain aerobic digesters allows for the development of greater biomass for a givenreactor volume and can result in greater treatment efficiency. This approach, oftenreferred to as “aerobic biofiltration,” was studied by Hill et al.141 for the treatmentof flushed swine manure. Their study reported that an ambient-temperature aerobicbiofilter system having a HRT of 1 day removed Salmonella, fecal coliforms,E. coli, enterococci, and coliphages by 90 to 99% at temperatures above 10°C, butremoved C. perfringens spores to a lesser extent (80 to 87%).

As discussed for anaerobic digesters, the use of thermophilic temperatureswith aeration conditions can be expected to achieve greater inactivation of patho-gens than at lower temperatures, but will likely incur greater operational costsassociated with maintaining reactor temperatures of >50°C. Thermophilic aerobicdigestion has been shown to reduce enteric bacteria, viruses and parasites by 3 to8 log10, 4 to 8 log10, and 3 to 6 log10, respectively, in pig slurry and municipalsludge.26,41,161,293 Salmonella dublin has been reported to be reduced by over 6 log10

in less than 4 h during thermophilic aerobic digestion of pig slurry at 55°C.117

5. Activated Sludge

Traditional activated sludge systems consist of an aeration tank, where influentwaste is mixed with an aerated suspension of microorganisms, followed by asedimentation tank, where the waste is clarified and a portion of the settled solidsreturned to the aeration tank. The activated sludge process can also be performedin a single tank, known as a “sequencing batch reactor” (SBR). Sequencing batchreactors are operated in a “fill-and-draw” mode, where waste is treated in batchesthrough the cyclical filling of the SBR with waste, aeration of the batch, clarifica-tion, and withdrawal of the treated waste before beginning the cycle again with a

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new batch of waste. Activated sludge treatment of swine waste has been investi-gated in both continuous flow activated sludge systems18 and in SBRs.5,93,226

Pathogens are inactivated and/or removed in activated sludge systems duringthe aeration phase due to environmental factors (e.g., oxygenated compoundtoxicity, temperature) and biological factors (e.g., predation, microbial antago-nism). Pathogens are also removed during the sedimentation process when theybecome entrained in floc particles that settle during clarification. Research onmunicipal waste activated sludge suggests that enteric viruses are inactivated inactivated sludge systems due to microbial activity, including the production ofproteolytic enzymes.174,305 A typical activated sludge system can be expected toreduce bacteria, viruses, protozoan parasites, and helminths in wastewater by 80 to99+, 90 to 99, 80 to 99, and 0 to 90%, respectively.21,92,262 In an intermittent-aeration conventional activated sludge system treating swine waste, fecal coliforms,and fecal streptococci were reduced by 99.8 and 99.9%, respectively, at a systemHRT of 17.5 days.19 A SBR treating swine waste was shown to be capable ofreducing fecal coliforms by 99 to 99.99%.5

C. Chemical Treatment Techniques

Chemicals can be used for animal waste treatment to improve solids removal(i.e., coagulation/flocculation) or disinfection. Coagulants and flocculants thathave been used to treat CAFO wastewater include lime [Ca(OH)2], alum (Al2SO4),ferric chloride (FeCl3), and organic polymers.106,269 These coagulants remove mi-crobes from water by entraining them and other particles in precipitant floc, whichsubsequently can be removed through solids separation processes (e.g., sedimen-tation or filtration). In addition to their use in wastewater and water treatment,coagulants have also been used in microbial assays for concentrating microbesfrom water and wastewater samples.274 Coagulant addition can also raise or loweranimal waste slurry pH values to levels that inactivate pathogenic microbes. Acidic(pH < 6) and alkaline (pH > 10) conditions achieved using sulfuric acid and sodiumhydroxide, respectively, have been shown to inactivate Foot-and-Mouth Disease(FMD) virus by 4 log10 in cattle slurry within 24 h contact time.231 However, pHlevels and contact times required for significant disinfection can vary greatlydepending on microbe type and environmental factors such as temperature.134,273

Depending on the concentrations used, lime can act as a disinfectant as wellas a coagulant. Lime has been used for the stabilization of sewage sludges fordecades and has also been investigated for bulk wastewater treatment prior to landdisposal.105 At concentrations sufficient to raise wastewater pH to 8 to 10, lime wasshown to not produce appreciable bactericidal or viricidal effects when pH was≤ 10.5).276 However, research on swine slurry containing Aujeszky’s Disease Virus(ADV) found that ADV concentrations could be reduced by 3 log10 in slurry at pH8 to 9 after 48 h contact time with lime.175 Sustained exposure to lime at pH 10.5

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has been shown to produce inactivation of bacteria276 and C. parvum oocysts.253

When Robertson et al.253 corrected the alum solution to pH 6, however, no inac-tivation of C. parvum oocyst viability was observed, so it appears that the primarymicrobiocidal effect of lime is a toxic effect caused by elevated pH. At a pH of 11.5and contact time of 8.5 h, a lime treatment system removed 91% of total solids and99.99, 99.8, ~100, and 97.9% of fecal coliforms, Salmonella spp., helminth eggs,and protozoan cysts in raw municipal wastewater105 (Table 4). Lime coagulationand sedimentation of swine waste slurry at pH 11.5 has been shown to reduceporcine enterovirus and adenovirus concentrations by 3 to 4 log10 through physicalremoval and chemical inactivation.79

Alum has been used for coagulation in wastewater treatment systems, as wellas septic tanks,32 lagoons,96 and lakes.40 When pH is controlled above 6.0, greaterthan 99.9% of virus particles can be removed from water and nearly 99% of theseviruses subsequently recovered (i.e., not inactivated by alum contact).38 For thisreason, alum precipitation has been investigated as a method for virus recovery inwaste samples.221 At circumneutral pH values, alum coagulation has been reportedto remove 99.9% of fecal coliforms in a wastewater lagoon,96 but this pH rangedoes not appear to affect C. parvum oocyst viability253 or Coxsackie virus infec-tivity.179 When pH is not controlled, alum acidifies wastewater and has been shownto be toxic to C. parvum oocysts.253 Ferric chloride addition to wastewater canremove greater than 70% of total suspended solids without polymer or polyelec-trolyte additives.219 As with alum, ferric chloride acidifies wastewater and there-fore can reduce pathogen concentrations through physical removal and inactiva-tion when circumneutral pH conditions are not maintained. At the same dosage,ferric chloride coagulation has been reported to be more effective at removing MS2coliphage in wastewater at pH 5 (nearly 99% reduction) than at pH 7 (approxi-mately 93% reduction), a relatively small reduction in effectiveness that, whencompared with the relatively larger decrease in turbidity removal efficiency athigher pH, suggests that ferric chloride also reduced MS2 through inactivation.191

Similarly, ferric chloride has been shown to decrease C. parvum oocyst viabilitywhen pH was not corrected (and went as low as pH 1.5), but not at pH 6.253 Thelong-term implementation of chemical coagulants for CAFO wastewater treatmentwill likely depend on the developing balance between costs (e.g., chemical costs,solids management costs) and benefits associated with the increased recovery ofsolids (e.g., biosolids reuse). However, due to their ability to create extremelyacidic (e.g., with ferric chloride) or basic conditions (e.g., with lime) in wastewatersystems, chemical coagulants may show utility as a biosecurity technique for large-scale disinfection of waste management systems during serious outbreaks ofdisease (e.g., Foot-and-Mouth Disease).

Disinfectant chemicals, including chlorine and ozone, are often used to reducepathogen concentrations in wastewater. Chlorine and other chlorine-based chemi-cals (e.g., chloramines, chlorine dioxide) are well-established disinfectants inmunicipal wastewater systems, but are not widely used in animal waste manage-

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ment systems. In municipal systems, alternatives to chlorine (e.g., ozone) areincreasingly being used due to concerns regarding chlorination byproducts(e.g., trihalomethanes and chlorite).21,244 Ozonation has been investigated for ani-mal waste treatment primarily as a method for odor reduction for swine waste.310,326

Watkins et al.310 found that an ozone concentration of 2.0 g/L reduced E. coliconcentrations by 3 log10 in swine waste slurry. Wu et al.326 measured average E.coli and coliphage reductions of approximately 1.4 and 0.6 log10, respectively, inswine waste slurry treated at an ozone concentration of 1.0 g/L. In secondarymunicipal wastewater, ozonation has been shown to be capable of reducing seededS. typhimurium by more than 4 log10, E. coli by 3 log10, and poliovirus by 2 log10.88

At an ozone dose of 0.4 mg/L, Norwalk virus, poliovirus, and MS2 coliphage wereshown to be significantly, and rapidly, reduced in buffered saline water (approxi-mately 3 log10 reductions after 60 s of contact).266 Research on a novel disinfectionapproach indicates that a combination of carbonate and alkali can achieve greaterthan a 5 log10 reduction of E. coli in dairy cattle slurry maintained at pH 8.5 for5 days.80

D. Energetic Treatment Techniques

Energetic treatment techniques rely on electrical, magnetic, shock wave, orradiation energy to inactivate pathogens through physical damage and oxidation.Electrolytic treatment of cattle slurry using copper electrodes and an electricpotential of 1 to 5 V has been reported to reduce thermotolerant coliforms from anaverage of 350 organisms/g to below the detection limit of 200 organisms/g,272 butdue to the small difference in starting concentration and the method detection limitit is difficult to evaluate whether this treatment approach achieved substantialreductions of E. coli. Pulsed electrical fields, which achieve microbial inactivationby rupturing the microbial cell membrane in a process referred to as “nonthermalpasteurization”,57,330 are primarily being investigated for disinfecting pumpablefoods. This technique has been shown to be able to reduce E. coli and othervegetative bacteria by 3 to 9 log10 at an energy input of 2 to 200 kJ/L.206,331

Electrohydraulic discharge (EHD) of pulsed-plasma electricity has been investi-gated for the treatment of chemically and microbially contaminated water.115,320

During EHD, electrohydraulic cavitation and supercritical water oxidation pro-cesses damage cellular enzymes and other components sufficiently to inactivateE. coli by 4 to 6 log10.115,116 Sonic or shock waves are higher pressure pulses thatcan destroy E. coli and bacteriophages by disrupting their outer membranes andfragmenting their nucleic acid.290 Electrohydraulic shock waves have been shownto inactivate E. coli by 4 log10 in saline solution,184 indicating that this method couldbe effective for flushed animal wastes. Irradiation (e.g., gamma or acceleratedelectron) is another potential technique for inactivating pathogens in CAFO waste-water. Gamma irradiation has been shown to reduce coliform concentrations in

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swine waste slurry by 4 to 6 log10, depending on the dosage applied75 (Table 4).Gamma irradiation of cattle manure has also been shown to effectively reducebovine enterovirus concentrations.212 Gamma irradiation of municipal wastewatercan reduce Salmonella and total coliforms by 5 to 6 log10.31 Accelerated electronirradiation can also be effective for reducing enteric microbes in raw sewage, butmay be somewhat less effective than gamma irradiation for the inactivation ofcoliforms and coliphages.89 Relatively low doses of electron beam radiation havebeen shown to inactivate helminth ova in sewage sludge144 and pig slaughterhouseslurry.55 Microwave heating of a liquid bovine-swine manure mixture was capableof inactivating enteric viruses by 3 to 4 log10 at contact times of less than 1 sec(resulting in reactor temperatures of 55 to 70°C.27

Ultraviolet (UV) irradiation is the most commonly used energetic techniquefor the disinfection of water and wastewater. UV irradiation has most often beenused for the disinfection of relatively high-quality water and wastewater becausethe disinfection efficacy of UV radiation is lessened by the absorbance of UV lightby background water color (e.g., organic compounds such as humic acids) andsuspended solids.22,232 Water temperature is not considered to be an importantfactor in the disinfection efficacy of UV irradiation.265 Relatively little research hasfocused on UV disinfection of low-quality wastewaters similar to the quality ofwastewater that would be expected in a CAFO waste management system. Resultsof a bench-scale study using low pressure collimated UV irradiation of flushedswine waste found that culturable concentrations of fecal coliforms, E. coli, andsomatic coliphages could be reduced by 2 to 2.5 log10 in untreated and treatedflushed wastewater when incident UV doses were 60 mJ/cm2.141 For relatively low-quality secondary municipal wastewater effluent, UV disinfection has been re-ported to be capable of reducing fecal coliforms, coliphages and C. perfringensspores by 6 log10, 6 to 7 log10, and 0.7 to 1.8 log10, respectively.243,317 Interestingly,UV irradiation inactivation rates for C. parvum oocysts and G. lamblia cysts havebeen shown to be higher than for vegetative bacteria such as E. coli.267,268 Inves-tigations of bacterial and viral survival following UV irradiation have demon-strated that the “inactivated” microbes may still be viable if they are able to repairDNA damage through light-dependent (i.e., “light repair”or photoreactivation) orlight-independent (i.e., “dark repair”) mechanisms.126,313 In general, UV irradiationis an effective disinfection technique that may have utility for CAFO wastewatertreatment, especially if beneficial reuse of wastewater is a goal (e.g., production offood for human consumption). More research is needed, however, to evaluate thelong-term cost-effectiveness of UV disinfection for livestock wastewaters.

IV. CONCLUSIONS

Livestock can be reservoirs for numerous bacterial, viral, and parasitic patho-gens of human health significance. Historically, farm animal waste has been

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managed by land application according to its nutrient content. With the continuingconcentration of animal production into larger and larger operations, concerns areincreasing regarding the potential impacts of pathogens and other animal wasteconstituents on community health and water quality. Although relatively littleliterature is currently available to characterize the risks posed by pathogens inCAFO waste management systems, substantial research is being invested intodeveloping and evaluating alternative animal waste treatment techniques usingenteric microbial parameters as treatment criteria. Guidelines have been estab-lished in the U.S. for the microbiological quality of domestic sewage sludge andreclaimed wastewater applied to land (Table 1). U.S. federal and state standards aretypically not directly applicable for the land application of CAFO wastes, althoughresearch has shown that livestock wastes can contain many of the same types ofpathogens as can be found in domestic wastes. Recent research suggests that thereare human health risks associated with the application of wastewater to limited-access agricultural lands,25 but the microbiological quality guidelines suggested bythis research and other agencies are generally limited to bacterial and helminthparameters that may not be indicative of risks associated with pathogenic virusesand protozoan parasites (e.g., C. parvum).

Traditional lagoon-based techniques can be effective for reducing pathogenconcentrations in livestock wastewaters, but high concentrations of enteric mi-crobes will likely remain in lagoon liquid unless multiple lagoon cells are used in-series or other reactor modifications are made (e.g., baffling). Little data areavailable regarding enteric microbe concentrations in lagoon systems, but theavailable research58,61,139 indicates that wastewater from single-stage anaerobiclagoons can contain fecal coliform concentrations that are higher than the humanhealth guideline (105 fecal coliforms per 100 mL) recommended by Blumenthal etal.25 for wastewaters applied to low-access agricultural lands.

Alternative biological treatment techniques have been developed that have thepotential to achieve similar or greater pathogen reductions as can be achieved inlagoon systems. Constructed wetlands are one of the most promising of thesealternative biological treatment techniques due to the potential cost-effectivenessof these treatment systems. However, there are numerous design approaches fortreatment wetlands and care must be taken to consider important treatment param-eters such as reactor HRT and temperature effects on treatment efficacy. Inaddition, it is unclear whether constructed wetland technology can be effective asan alternative to lagoon treatment because high-strength flushed waste may betoxic to wetland vegetation. Other biological treatment techniques, such as anaero-bic digestion, aeration, and activated sludge, have been used on farms for livestockwaste treatment. Although research on the cost-effectiveness of candidate treat-ment technologies was not a focus of this literature review, it appears that anaero-bic digestion is one of the techniques having the greatest potential for use onCAFOs because of the technology’s ability to generate methane for energy recov-ery. However, anaerobic digesters can be a challenge to operate and are not likely

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to provide substantially high pathogen reductions unless long residence times areprovided or elevated temperatures maintained. Aeration techniques can be ex-pected to achieve higher levels of pathogen reductions compared with anaerobictechniques, but the costs associated with aeration may make these systems lessattractive for use on commercial livestock operations. Similar questions exist forthe various chemical and energetic disinfection techniques discussed in this re-view. Chemical addition, ozonation, and energetic treatment techniques showpromise for livestock wastewater disinfection, but questions of cost-effectivenesswill necessarily guide the implementation of these techniques for on-farm wastemanagement applications. These disinfection techniques may receive increasedattention as alternative waste utilization scenarios are investigated, such as humancrop production.

Whether related to animal waste utilization or waste disposal scenarios, ques-tions regarding pathogen presence and removal will continue to be raised untilsufficient research has been performed to characterize the human health andenvironmental risks posed by wastes generated by CAFOs. A review of theresearch literature suggests that substantial pathogen reductions can be achieved byanimal waste management systems using available and developing treatment ap-proaches. However, available data are not yet sufficiently robust to identify tech-nologies that combine cost-effectiveness with comprehensive efficacy for protect-ing human health and environmental resources.

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