Electrochemical disinfection of Escherichia coli in the presence and absence of primary sludge particulates

Electrochemical (EC) residual disinfection of Escherichia coli (E. coli) in the presence and absence of primary sludge particulates (PSPs) was studied. The kinetics followed a first-order rate law. When PSPs were absent, the EC residual disinfection rate coefficient (k) increased linearly with EC pretreatment energy (E_C, 0–0.63 kWh/m³). However, with 143 mg PSPs/L, k first increased linearly with E_C (0–0.28 kWh/m³) and then decreased linearly with E_C (0.28–0.42 kWh/m³). H₂O₂ was detected during EC pretreatment in PSPs-free samples and the H₂O₂ concentration (C_H) increased with E_C (0–0.83 kWh/m³) linearly. Chloride was detected in PSPs aqueous samples (143 mg PSPs/L) and its concentration (C_C) changed during EC pretreatment: initially, a decrease of C_C was observed when E_C increased from 0 to 0.28 kWh/m³, followed by an increase of C_C when E_C increased 0.28–0.42 kWh/m³. In both cases, k correlated to the initial post-EC chloride concentration (C_CI) in an inverse linear relationship. This two-stage change of C_C and k was caused by a combination of two reactions: anodic oxidation of chloride and the reaction of chloramines with excess chlorine. This paper explains the mechanisms underlying EC residual disinfection in the presence and absence of PSPs, and proposes a feasible strategy for EC disinfection when PSPs are present, an approach that could be useful in the treatment of combined sewage overflow (CSO).     

Hydroxyapatite-supported Ag-TiO2 as Escherichia coli disinfection photocatalyst

A series of hydroxyapatite (HAP), 1wt% Ag-TiO(2) (AT1), 1wt% Ag-HAP and 5wt% AT1/HAP composite catalysts were prepared by incipient wetness and mechanical mixing methods. They were characterized by X-ray diffraction (XRD), FT-IR, SEM and ESCA analyses and their photocatalytic bactericidal activities were measured in suspension using Escherichia coli (E. coli), a water pollutant indicator. The surface analysis revealed that the Ag/Ti ratio is found to be ca. 0.0273 and also it indicated that the titania is present in the form of Ti(4+) and Ag is present as metallic silver. Both the XRD and ESCA analyses confirmed the phase of metallic Ag particles, which played a significant role on the bactericidal activity of the Ag doped TiO(2) catalysts. The FT-IR analysis of HAP revealed that the peak intensity is due to the absorbance of surface PO(4)(3-) group centered at wave number 1030cm(-1) and is drastically decreased upon exposure to UV for 1h. The HAP displayed high amount of bacteria adsorption, ca. 80% during the dark experiments compared to other catalytic systems tested. The cumulative photocatalytic properties of AT1/HAP catalytic system resulted in 100% E. coli bacteria reduction within 2min.     

UV inactivation and characteristics after photoreactivation of Escherichia coli with plasmid: health safety concern about UV disinfection

Occurrence and degree of photoreactivation after ultraviolet (UV) exposure have been widely studied. However, the characteristics of photoreactivated microorganisms were rarely investigated. Hence, in this study, Escherichia coli with plasmids of ampicillin (amp)-resistance or fluorescence was used as indicators to examine the UV inactivation efficiencies and variations of characteristics of E. coli after subsequent photoreactivation.The experimental results indicate that the amp-resistant bacteria and the fluorescent bacteria used in this study had similar trends of UV dose-response curves. 3.5-log₁₀ and 3-log₁₀ reductions were achieved with a UV dose of 5 mJ/cm² for the amp-resistant and fluorescent E. coli, respectively. There was no significant difference in the UV inactivation behavior, as compared with common strains of E. coli.For the amp-resistant E. coli and the fluorescent E. coli, after exposures with UV doses of 5, 15, 25, 40 and 80 mJ/cm², the corresponding percent photoreactivations after a 4 h exposure to photoreactivating light were 1% and 46% respectively for a UV dose of 5 mJ/cm², and essentially negligible for all other UV doses. Furthermore, the photoreactivated amp-resistant bacteria still have the ability of amp-resistance. And the revived fluorescent E. coli showed similar fluorescent behavior, compared with the untreated bacteria. The experimental results imply that after UV inactivation and subsequent photoreactivation, the bacteria retained the initial characteristics coded in the plasmid. This reveals a possibility that some characteristics of bacteria can retain or recover through photoreactivation, and a safety concern about pathogenicity revival might need to be considered with UV disinfection and photoreactivation.     

Pulsed ultra-violet inactivation spectrum of Escherichia coli

Inactivation of Escherichia coli is examined using ultra-violet (UV) radiation from a pulsed xenon flashlamp. The light from the discharge has a broadband emission spectrum extending from the UV to the infrared region with a rich UV content. The flashlamp provides high-energy UV output using a small number of short-duration pulses (30 micros). The flashlamp is used with a monochromator to investigate the wavelength sensitivity of E. coli to inactivation by the pulsed UV light. Using 8 nm wide pulses of UV radiation, the most efficient inactivation is found to occur at around 270 nm and no inactivation is observed above 300 nm. A pyroelectric detector allows the energy dose to be determined at each wavelength, and a peak value for E. coli population reduction of 0.43 log per mJ/cm(2) is measured at 270 nm. The results are compared with the published data available for continuous UV light sources.     

Bioaccumulation of nickel from aqueous solutions by genetically engineered Escherichia coli

This study constructed a genetically engineered Escherichia coli JM109 which simultaneously expressed nickel transport system and metallothionein to remove and recover Ni(2+) from aqueous solution. Bioaccumulation process was rapid and followed linearized Langmuir isotherm. A more than six-fold increase of Ni(2+) binding capacity was obtained by genetically engineered E. coli cells compared with original host E. coli cells. A pH assay showed genetically engineered E. coli cells accumulated Ni(2+) effectively over a broad range of pH (4-10). The presence of 1000 mg/L Na(+) and Ca(2+), or 50mg/L Cd(2+) or Pb(2+) did not have a significant effect on Ni(2+) bioaccumulation, while Mg(2+), Hg(2+) and Cu(2+) posed a severe adverse influence on Ni(2+) uptake by genetically engineered E. coli. Furthermore, genetically engineered E. coli cells did not require extra nutrients for Ni(2+) bioaccumulation.     

Recovery of Escherichia coli in fresh water fish, Jenynsia multidentata and Bryconamericus iheringi

Escherichia coli concentration was determined in digestive tract and muscle of Jenynsia multidentata and Bryconamericus iheringi through bioassays. Field experiments were also conducted with J. multidentata collected in the Suquía River, Córdoba, Argentina. E. coli was quantified by the most probable number, using lauryl sulphate tryptose broth with 4-methylumbelliferyl-beta-D-glucuronide. For bioassays, E. coli concentrations 10(2), 10(3), 10(4), 10(5), 10(6)CFU/ml were introduced in aquarium water. E. coli was recovered from the digestive tracts of J. multidentata and B. iheringi in all the concentrations assayed. Bacterial critical load in water for the recovery of bacteria from muscle, was 10(3)CFU/ml for both species. The regression analysis between E. coli loads in water and those found in digestive tract and muscle showed a positive linear relationship for J. multidentata and B. iheringi. The same relation was observed between the concentration of bacteria in digestive tract and muscle in both species. In field experiments, E. coli was recovered from digestive tract and muscle of J. multidentata. The presence of E. coli in the studied fish suggests that they can carry bacteria to non-polluted waters. However, further studies are necessary to evaluate its significance for public and environmental health.     

Relative survival of Escherichia coli and Salmonella typhimurium in a tropical estuary

Microcosm studies have been carried out to find out the relative survival of Escherichia coli and Salmonella typhimurium in a tropical estuary. Survival has been assessed in relation to the important self-purifying parameters such as biotic factors contained in the estuarine water, toxicity due to the dissolved organic and antibiotic substances in the water and the sunlight. The results revealed that sunlight is the most important inactivating factor on the survival of E. coli and S. typhimurium in the estuarine water. While the biological factors contained in the estuarine water such as protozoans and bacteriophages also exerted considerable inactivation of these organisms, the composition of the water with all its dissolved organic and inorganic substances was not damaging to the test organisms. Results also indicated better survival capacity of E. coli cells under all test conditions when compared to S. typhimurium.     

Escherichia coli virulence genes profile of surface waters as an indicator of water quality

We compared the presence of 58 known virulence genes (VGs) associated with Escherichia coli strains causing intestinal (InPEC) and extra-intestinal (ExPEC) infections in three estuarine, four brackish and 13 freshwater sites during the dry and wet seasons. The most common VGs observed in water samples during the dry season belonged to ExPEC (traT; 80% and ompA; 70%) whilst east1 (70%) gene was the most common among InPEC. More types of VGs were observed in water samples during wet season and included those found among InPEC (e.g. eaeA; 100%; fyuA, 90%; paa, 65%; cdt, 60%; and stx₂, 60%) and ExPEC (e.g. iroN_E.coli, 90%; iss, 90% and kpsMTII, 80%). Eight VGs were found exclusively in the wet season, of which four were found in all three water types indicating their association with storm-water run off. The number of VGs associated with ExPEC were significantly (P < 0.05) higher in only brackish and estuarine waters during the wet season compared to the dry season. There was no correlation between the number of E. coli and the presence of VGs in any of the water types in both seasons but we found similarities in VG profiles of sites with similar land uses.     

Escherichia coli, enterococci, and Bacteroides thetaiotaomicron qPCR signals through wastewater and septage treatment

Fecal indicators such as Escherichia coli and enterococci are used as regulatory tools to monitor water with 24 h cultivation techniques for possible input of sewage or feces and presence of potential enteric pathogens yet their source (human or animal) cannot be determined with routine methods. This critical uncertainty has furthered water pollution science toward new molecular approaches. Members of Bacteroides genus, such as Bacteroides thetaiotaomicron are found to have features that allow their use as alternative fecal indicators and for Microbial Source Tracking (MST). The overall aim of this study was to evaluate the concentration and fate of B. thetaiotaomicron, throughout a wastewater treatment facility and septage treatment facility. A large number of samples were collected and tested for E. coli and enterococci by both cultivation and qPCR assays. B. thetaiotaomicron qPCR equivalent cells (mean: 1.8 × 10⁷/100 mL) were present in significantly higher concentrations than E. coli or enterococci in raw sewage and at the same levels in raw septage. The removal of B. thetaiotaomicron target qPCR signals was similar to E. coli and enterococci DNA during the treatment of these wastes and ranged from 3 to 5 log₁₀ for wastewater and was 7 log₁₀ for the septage. A significant correlation was found between B. thetaiotaomicron marker and each of the conventional indicators throughout the waste treatment process for both raw sewage and septage. A greater variability was found with enterococci when compared to E. coli, and CFU and equivalent cells could be contrasted by various treatment processes to examine removal and inactivation via septage and wastewater treatment. These results are compared and contrasted with other qPCR studies and other targets in wastewater samples providing a view of DNA targets in such environments.     

Microbial toxicity of metal oxide nanoparticles (CuO, NiO, ZnO, and Sb2O3) to Escherichia coli, Bacillus subtilis, and Streptococcus aureus

In this study, the microbial toxicities of metal oxide nanoparticles were evaluated for Escherichia coli, Bacillus subtilis, and Streptococcus aureus in laboratory experiments. The nanoparticles tested were CuO, NiO, ZnO, and Sb₂O₃. The metal oxide nanoparticles were dispersed thoroughly in a culture medium, and the microorganisms were cultivated on Luria-Bertani agar plates containing different concentrations of metal oxide nanoparticles. The bacteria were counted in terms of colony forming units (CFU). The CFU was reduced in a culture medium containing metal oxide NP, and the dose-response relationship was characterized. CuO nanoparticles were found to be the most toxic among the tested nanoparticles, followed by ZnO (except S. aureus), NiO, and Sb₂O₃ nanoparticles. We determined that the intrinsic toxic properties of heavy metals are also associated with the toxicity of metal oxide nanoparticles. Ion toxicity was also evaluated to determine the effects of metal ions dissolved from metal oxide NPs, and the toxicity induced from the dissolved ions was determined to be negligible herein. To the best of our knowledge, this is the first study of the toxicity of NiO and Sb₂O₃ NPs on microorganisms. We also discuss the implications of our findings regarding the effects of the intrinsic toxic properties of heavy metals, and concluded that the apparent toxicities of metal oxide NPs can largely be understood as a matter of particle toxicity.     

Influence of tetracycline resistance on the transport of manure-derived Escherichia coli in saturated porous media

In this research, tetracycline resistant (tet^R) and tetracycline susceptible (tet^S) Escherichia coli isolates were retrieved from dairy manure and the influence of tetracycline resistance on the transport of E. coli in saturated porous media was investigated through laboratory column transport experiments. Experimental results showed that tet^RE. coli strains had higher mobility than the tet^S strains in saturated porous media. Measurements of cell surface properties suggested that tet^RE. coli strains exhibited lower zeta potentials than the tet^S strains. Because the surface of clean quartz sands is negatively charged, the repulsive electrostatic double layer (EDL) interaction between the tet^R cells and the surface of sands was stronger and thus facilitated the transport of the tet^R cells. Although no difference was observed in surface acidity, cell size, lipopolysaccharides (LPS) sugar content and cell-bound protein levels between the tet^R and tet^S strains, they displayed distinct outer membrane protein (OMP) profiles. It was likely that the difference in OMPs, some potentially related to drug efflux pumps, between the tet^R and tet^S strains led to alteration in cell surface properties which in turn affected cell transport in saturated porous media. Findings from this research suggested that manure-derived tet^RE. coli could spread more widely in the groundwater system and pose serious public health risks.     

Optimization of DGGE community fingerprinting for characterizing Escherichia coli communities associated with fecal pollution

We evaluated the use of DGGE fingerprinting to differentiate communities of Escherichia coli from animal and geographic sources. An initial screening of 15 gene candidates revealed the ability of three target genes (mdh, phoE and uidA-4) to effectively differentiate E. coli communities originating in horses, pigs, geese and goats. Cluster and jackknife analyses performed on the communities from a more extensive number of hosts (n = 150) including humans (via raw sewage), horses, pigs, geese and cows revealed that the internal accuracy of classification of E. coli community fingerprints to their origin was similar for each of the three genes (85-86%). Each of the three genes were tested for their ability to associate E. coli source- and sink communities in two settings featuring contaminated water; (i) a stream receiving municipal wastewater effluent and (ii) a pond inhabited by geese. For each gene, DGGE fingerprints effectively matched effluent- and downstream E. coli communities (98-100% similarity) and excluded upstream communities, while communities from goose fecal material were 77-79% similar to communities in pond water, indicating fecal inputs from geese. Furthermore, each gene discriminated against E. coli communities from hosts non-indigenous to either setting. DGGE analysis of E. coli communities appears to be a promising tool to augment existing efforts aiming to address the dynamics of bacteria pollution in complex, natural environments.     

Effects of sludge dewatering and digestion processes on Escherichia coli enumeration

Centrifuge dewatering digested sewage sludge has been reported to significantly increase numbers of Escherichia coli, potentially exceeding the UK microbiological standards of 10³ or 10⁵ E. coli/g dry solids (DS) for enhanced or conventional treatment for agricultural use, respectively. Here, we report an investigation of the effects of different types of dewatering process on E. coli enumeration in conventionally treated, anaerobically digested sludge (primary and secondary liquid digestion), as well as raw sludge at eight wastewater treatment works in the United Kingdom. The dewatering methods evaluated included: centrifuge conditioning and filter-belt and filter-plate pressing. The results demonstrated that conventional treatment by primary and secondary liquid digestion and dewatering conditioning produces biosolids compliant with UK maximum microbiological limits for agricultural application.     

DNA extraction and Escherichia coli quantification of anaerobically digested biosolids using the competitive touchdown PCR method

Accurate enumeration of indicator organisms such as Escherichia coli is important for assessing the safety of water and wastewater samples. Recent research has shown that E. coli can enter a viable but non-culturable state; therefore, traditional cultivation methods could potentially underestimate the quantities of the organisms. The goals of the research were to develop and verify a DNA extraction protocol and a quantitative polymerase chained reaction (PCR) method for E. coli enumeration in digested biosolids. A solvent-based DNA extraction protocol with extensive cell lysis recovered approximately 78-84% of spiked DNA. In comparison, a commercial kit only recovered 28-42% of DNA, likely from inefficient cell lysis. The developed competitive touchdown PCR (cPCR) method for E. coli enumeration was comparable to both real-time PCR (rt-PCR) and cultivation methods with sensitivity of approximately 50,000-500,000 E. coli per gram dry solids (DS), which is suitable for Class B biosolids monitoring in the US and "conventional" biosolids in the European Union. The cPCR protocol provides a less expensive alternative than the rt-PCR as a culturing independent method for enumerating E. coli.     

Detection of virulence genes in Escherichia coli of an existing metabolic fingerprint database to predict the sources of pathogenic E. coli in surface waters

A collection of 366 Escherichia coli strains from 10 host groups and surface waters were tested for the presence of 15 virulence genes associated with strains causing intestinal and extra-intestinal infections. The virulence genes included eaeA, VT1, 2 and 2e, LT1, ST1 and 2, Einv gene, EAgg gene, CNF1 and 2, papC, O111 and O157 side chain LPS. Of the 262 strains obtained from nine different hosts, 39 (15%) carried one or more of these virulence genes. These included six strains from humans, two from horses, eight from dogs, two from ducks, five from cattle, seven from chickens, four from pigs, two from sheep and three from deer. Of the remaining 104 strains obtained from water samples, 10 (10%) also carried one or more of the tested virulence genes. Of these, six had identical biochemical phenotypes (BPTs) to strains isolated from humans (two strains), dogs (two strains), chickens (one strain) and sheep (one strain) with 4 BPTs also carrying same virulence genes. Our results indicate that the sources of clinically important E. coli strains found in surface waters due to faecal contamination can be predicted by using a combination of biochemical fingerprinting method and the detection of virulence genes. From the public health point of view this information will be of great importance for evaluating the risk associated with public use of the catchment.     

Deer diet affects ribotype diversity of Escherichia coli for bacterial source tracking

Ribotyping is one of a number of genotypic methods for bacterial source tracking. This method requires a host origin database of one bacterial species be established in order to identify environmental isolates. Researchers establishing these databases have observed considerable ribotype diversity within a specific bacterial species. One source of this diversity may be diet. We determined the effect of diet on ribotype diversity for Escherichia coli in penned and wild deer (Odocoileus virginianus) in a 13-ha forested watershed. A total of 298 E. coli isolates was obtained, 100 from penned deer, 100 from wild deer, and 98 from the stream in the watershed to which all deer had access. The wild deer had significantly more ribotypes (35) than the penned deer (11 ribotypes, p = 0.05). This result suggests that diet affected ribotype diversity, and that a host origin database for bacterial source tracking should contain bacterial isolates from wild rather than from captive animals. Also, 42 of 98 (42.9%) environmental isolates matched penned and wild deer ribotypes. If bacterial source tracking determines that fecal contamination is predominantly from wildlife, then it may be unnecessary to monitor these watersheds because control over wildlife is difficult.     

Transport of Escherichia coli in saturated porous media: dual mode deposition and intra-population heterogeneity

Because of heterogeneity among members of a bacteria population, deposition rates of bacteria may decrease upon the distance bacteria are transported in an aquifer. Such deposition rate decreases may result in retained bacteria concentrations, which decrease hyper-exponentially as a function of transport distance, and may therefore significantly affect the transport of colloids in aquifers. We investigated the occurrence of hyper-exponential deposition of Escherichia coli, an important indicator for fecal contamination, and the causes for such behavior. In a series of column experiments with glass beads of various sizes, we found that attachment of E. coli decreased hyper-exponentially, or, on logarithmic scale in a bimodal way, as a function of the transported distance from the column inlet. From data fitting of the retained bacteria concentration profiles, the sticking efficiency of 40% of the E. coli population was high (alpha=1), while the sticking efficiency of 60% was low (alpha=0.01). From the E. coli total population, an E. coli subpopulation consisting of slow attachers could be isolated by means of column passage. In subsequent column experiments this subpopulation attached less than the E. coli total population, consisting of both slow and fast attachers. We concluded that the main driver for the observed dual mode deposition was heterogeneity among members of the bacteria population. Intra-population may result in some microbes traveling surprisingly high distances in the subsurface. Extending the colloid filtration theory with intra-population variability may provide a valuable framework for assessing the transport of bacteria in aquifers.     

Genetic diversity of Escherichia coli isolates in irrigation water and associated sediments: implications for source tracking

Identifying the sources of fecal contaminants in surface water bodies such as rivers and lakes is of significant importance for environmental quality, food safety and regulatory purposes. Current DNA library-based source tracking approaches rely on the comparison of the genetic relatedness among the fecal contaminants. The objective of this study was to determine the genetic relatedness of Escherichia coli isolated from irrigation water and associated sediments using pulse field gel electrophoresis (PFGE) and to evaluate the genetic stability of the E. coli PFGE patterns. The isolates were obtained over a 4-month period from specific locations within irrigation canals and sediments associated with the Rio Grande River along the Texas-Mexico border. Fifty E. coli isolates were genotyped using PFGE. Different E. coli genotypes were identified among samples collected in 11 different locations. Some isolates obtained over successive months showed similar genotypic patterns. In the laboratory experiment, the PFGE pattern of one E. coli strain changed during survival in irrigation water. The genetic relatedness of this strain changed from >95% to <83% over 8-week survival. These results imply that PFGE is of such extreme resolution that it may be a challenging task to rely solely on a PFGE-based source tracking DNA fingerprint library for large watersheds.     

Impact of sonication at 20 kHz on Microcystis aeruginosa, Anabaena circinalis and Chlorella sp

Blooms of toxic cyanobacteria such as Microcystis aeruginosa periodically occur within wastewater treatment lagoons in the warmer months, and may consequently cause contamination of downstream water and outages of the supply of recycled wastewater. Lab-scale sonication (20 kHz) was conducted on suspensions of M. aeruginosa isolated from a wastewater treatment lagoon, and two other algal strains, Anabaena circinalis and Chlorella sp., to investigate cell reduction, growth inhibition, release of microcystin and sonication efficiency in controlling the growth of the M. aeruginosa. For M. aeruginosa, for all sonication intensities and exposure times trialled, sonication led to an immediate reduction in the population, the highest reduction rate occurring within the initial 5 min. Sonication for 5 min at 0.32 W/mL, or for a longer exposure time (>10 min) at a lower power intensity (0.043 W/mL), led to an immediate increase in microcystin level in the treated suspensions. However, prolonged exposure (>10 min) to sonication at higher power intensities reduced the microcystin concentration significantly. Under the same sonication conditions, the order of decreasing growth inhibition of the three algal species was: A. circinalis > M. aeruginosa > Chlorella sp., demonstrating sonication has the potential to selectively remove/deactivate harmful cyanobacteria from the algal communities in wastewater treatment lagoons.     

The application of bioluminescence assay with culturing for evaluating quantitative disinfection performance

Various methods, including bioluminescence assay, were investigated regarding their suitability for quantitatively evaluating the disinfection performance. Although bioluminescence assay itself has been widely reported as a rapid, easy and suitable method for analyzing live microorganisms, the limited sensitivity of its measurement (approximately 10(3)-10(4)cells/assay vial), which is insufficient for disinfection study, requires further study. Among three methods (amplifying by enzymatic method, membrane filtration, and amplification by culturing) examined for increasing the detection sensitivity, amplification by culturing showed the best performance as Escherichia coli was employed as an indicating microorganism. Even with a short culturing time of 4h, the detection limit of E. coli measurement was successfully improved 200-fold, and the analytical results were not dependent upon the state of E. coli growth (stationary state with E. coli stock suspension vs. growth state with E. coli). In addition, the analytical integrity of bioluminescence assay with culturing was further demonstrated in comparison with spread plate method as free chlorine and UV irradiation were employed in the disinfection study.