Survival of Escherichia coli in two sewage treatment plants using UV irradiation and chlorination for disinfection

We investigated the survival of Escherichia coli in two STPs utilising UV irradiation (STP-A) or chlorination (STP-B) for disinfection. In all, 370 E. coli strains isolated from raw influent sewage (IS), secondary treated effluent (STE) and effluent after the disinfection processes of both STPs were typed using a high resolution biochemical fingerprinting method and were grouped into common (C-) and single (S-) biochemical phenotypes (BPTs). In STP-A, 83 BPTs comprising 123 isolates were found in IS and STE, of which 7 BPTs survived UV irradiation. Isolates tested from the same sites of STP-B (n = 220) comprised 122 BPTs, however, only two BPTs were found post-chlorination. A representative isolate from each BPT from both STPs was tested for the presence of 11 virulence genes (VGs) associated with uropathogenic (UPEC) or intestinal pathogenic (IPEC) E. coli strains. Strains surviving UV irradiation were distributed among seven phylogenetic groups with five BPTs carrying VGs associated with either UPEC (4 BPTs) or IPEC (1 BPT). In contrast, E. coli strains found in STP-B carried no VGs. Strains from both STPs were resistant to up to 12 out of the 21 antibiotics tested but there was no significant difference between the numbers of antibiotics to which surviving strains were resistant to in these STPs. Our data suggests that some E. coli strains have a better ability to survive STPs utilising chlorination and UV irradiation for disinfection. However, strains that survive UV irradiation are more diverse and may carry more VGs than those surviving SPTs using chlorination.     

Comparative disinfection efficiency of pulsed and continuous-wave UV irradiation technologies

Pulsed UV (PUV) is a novel UV irradiation system that is a non-mercury lamp-based alternative to currently used continuous-wave systems for water disinfection. PUV polychromatic irradiation disinfection efficiency was compared to that from continuous-wave monochromatic low-pressure (LP) and polychromatic medium-pressure (MP) UV systems, using two types of actinometry (ferrioxalate and iodide-iodate) and an absolute spectral emission method for fluence measurement. All three methods were in good agreement. Once accurate and reliable methods for fluence measurement were established, the inactivation of Escherichia coli and pathogen surrogates phage T4 and T7 were investigated under each technology. Inactivation was significantly faster using PUV irradiation compared to LP or MP UV lamps at equivalent fluence levels. A significant fraction of the enhanced PUV inactivation efficiency was due to wavelengths greater than 295 nm.     

Photocatalytic inactivation of E. coli in surface water using immobilised nanoparticle TiO2 films

Photocatalysis is a promising method for the disinfection of potable water in developing countries where solar irradiation can be employed, thus reducing the cost of treatment. In addition to microbial contamination, water normally contains suspended solids, dissolved inorganic ions and organic compounds (mainly humic substances) which may affect the efficacy of solar photocatalysis. In this work the photocatalytic and photolytic inactivation rates of Escherichia coli using immobilised nanoparticle TiO₂ films were found to be significantly lower in surface water samples in comparison to distilled water. The presence of nitrate and sulphate anions spiked into distilled water resulted in a decrease in the rate of photocatalytic disinfection. The presence of humic acid, at the concentration found in the surface water, was found to have a more pronounced affect, significantly decreasing the rate of disinfection. Adjusting the initial pH of the water did not markedly affect the photocatalytic disinfection rate, within the narrow range studied.     

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.     

Comparison of immunomagnetic separation/adenosine triphosphate rapid method to traditional culture-based method for E. coli and enterococci enumeration in wastewater

Untreated wastewater samples from California, North Carolina, and Ohio were analyzed by the immunomagnetic separation/adenosine triphosphate (IMS/ATP) method and the traditional culture-based method for E. coli and enterococci concentrations. The IMS/ATP method concentrates target bacteria by immunomagnetic separation and then quantifies captured bacteria by measuring bioluminescence induced by release of ATP from the bacterial cells. Results from this method are available within 1 h from the start of sample processing. Significant linear correlations were found between the IMS/ATP results and results from traditional culture-based methods for E. coli and enterococci enumeration for one location in California, two locations in North Carolina, and one location in Ohio (r values ranged from 0.87 to 0.97). No significant linear relation was found for a second location in California that treats a complex mixture of residential and industrial wastewater. With the exception of one location, IMS/ATP showed promise as a rapid method for the quantification of faecal-indicator organisms in wastewater.     

Assessing the ecotoxicity of vinyl chloride using green alga P. subcapitata, nematode C. elegans, and the SOS chromotest in a closed system without headspace

The ecotoxicity of vinyl chloride (VC) was evaluated using green alga, nematode, and the SOS chromotest. The green alga and nematode tested were Pseudokirchneriella subcapitata, and Caenorhabditis elegans, respectively. Because of the tendency of VC to escape from an aqueous exposure medium to the air phase, all tests in the present study were performed in a closed system without headspace to minimize the losses of VC. Previous studies on VC toxicity were performed in an open system or closed system with headspace. VC inhibits the growth of P. subcapitata. The 48-h IC50 value for P. subcapitata exposed to VC was calculated to be 5.15 mg/L. The survival of C. elegans was not influenced at concentrations of up to 60 mg/L; however, VC has an adverse effect on the reproduction of C. elegans. In a stress-related gene expression test using C. elegans, a significant and concentration-dependent expression of heat shock protein 16.2 was observed, indicating that VC induces the stress of C. elegans at the genetic level. The results of the SOS chromotest using Escherichia coli PQ37 showed an IF_max value of 1.11, indicating that VC is not genotoxic. The present study demonstrated that VC has an adverse effect on the algal growth and reproductive and genetic levels of C. elegans. A closed system without headspace is an effective method of testing the aquatic toxicity of volatile organic compounds such as VC.     

Effect of hybrid UV-thermal energy stimuli on inactivation of S. epidermidis andB. subtilis bacterial bioaerosols

Bioaerosols have become an increasingly important issue due to their harmful effects on human health. As the concern over airborne microorganisms grows, so does the need to develop and study efficient methods of controlling them. In this study, we designed a hybrid system involving ultraviolet (UV) irradiation and thermal energy and investigated its effects on bacterial bioaerosols, followed by a comparison with thermal energy alone and UV irradiation alone. The results show that the hybrid effect caused no variation in the shape of the normalized particle size distributions of S. epidermidis and B. subtilis bioaerosols. However, a physical transport loss of bacterial bioaerosols developed as the temperature inside the glass quartz tube increased. When bacterial bioaerosols were simultaneously exposed to UV irradiation and thermal energy for less than 1.05 s, more than 99% of S. epidermidis bioaerosols were inactivated at 120 °C with exposure to one UV lamp and at 80 °C with exposure to two UV lamps; and 93.5% and 98.5% of B. subtilis bioaerosols were inactivated at 280 °C with exposure to one and two UV lamps, respectively. Moreover, the hybrid UV-thermal stimuli significantly reduced the concentration of ozone, which is a secondary UV-induced pollutant. Our results show that to obtain the same inactivation efficiency, the hybrid UV-thermal stimuli were more efficient than thermal energy alone in terms of energy consumption and produced significantly less ozone than UV irradiation alone. The hybrid stimuli also had higher inactivation efficiency than UV alone. Therefore, these results provide valuable information for the development of new methods for controlling bioaerosols.     

Mechanisms of Escherichia coli inactivation by several disinfectants

The objective of this study was to elucidate dominant mechanisms of inactivation, i.e. surface attack versus intracellular attack, during application of common water disinfectants such as ozone, chlorine dioxide, free chlorine and UV irradiation. Escherichia coli was used as a representative microorganism. During cell inactivation, protein release, lipid peroxidation, cell permeability change, damage in intracellular enzyme and morphological change were comparatively examined. For the same level of cell inactivation by chemical disinfectants, cell surface damage was more pronounced with strong oxidant such as ozone while damage in inner cell components was more apparent with weaker oxidant such as free chlorine. Chlorine dioxide showed the inactivation mechanism between these two disinfectants. The results suggest that the mechanism of cell inactivation is primarily related to the reactivity of chemical disinfectant. In contrast to chemical disinfectants, cell inactivation by UV occurred without any changes measurable with the methods employed. Understanding the differences in inactivation mechanisms presented herein is critical to identify rate-limiting steps involved in the inactivation process as well as to develop more effective disinfection strategies.     

Survival of manure-borne E. coli in streambed sediment: Effects of temperature and sediment properties

Escherichia coli bacteria are commonly used as indicator organisms to designate of impaired surface waters and to guide the design of management practices to prevent fecal contamination of water. Stream sediments are known to serve as a reservoir and potential source of fecal bacteria (E. coli) for stream water. In agricultural watersheds, substantial numbers of E. coli may reach surface waters, and subsequently be deposited into sediments, along with fecal material in runoff from land-applied manures, grazing lands, or wildlife excreta. The objectives of this work were (a) to test the hypothesis that E. coli survival in streambed sediment in the presence of manure material will be affected by sediment texture and organic carbon content and (b) to evaluate applicability of the exponential die-off equation to the E. coli survival data in the presence of manure material. Experiments were conducted at three temperatures (4 °C, 14 °C, and 24 °C) in flow-through chambers using sediment from three locations at the Beaverdam Creek Tributary in Beltsville, Maryland mixed with dairy manure slurry in the proportion of 1000:1. Indigenous E. coli populations in sediments ranged from ca. 10¹ to 10³ MPN g⁻¹ while approx 10³ manure-borne E. coli MPN g⁻¹ were added. E. coli survived in sediments much longer than in the overlaying water. The exponential inactivation model gave an excellent approximation of data after 6-16 days from the beginning of the experiment. Slower inactivation was observed with the increase in organic carbon content in sediments with identical granulometric composition. The increase in the content of fine particles and organic carbon in sediments led not only to the slower inactivation but also to lower sensitivity of the inactivation to temperature. Streambed sediment properties have to be documented to better evaluate the role of sediments as reservoirs of E. coli that can affect microbiological stream water quality during high flow events.     

Differentiation and identification of Shigella spp. and enteroinvasive Escherichia coli in environmental waters by a molecular method and biochemical test

Both Shigella spp. and enteroinvasive Escherichia coli (EIEC) are important human pathogens that are responsible for the majority of cases of endemic bacillary dysentery. However, they are difficult to identify and differentiate by biochemical tests or molecular methods alone. In this study, we developed a procedure to detect Shigella spp. and EIEC from environmental water samples using membrane filtration followed by nutrient broth enrichment, isolation using selective culture plates, and identification of the invasion plasmid antigen H (ipaH) gene by PCR amplification and DNA sequencing. Finally, we used a biochemical test and a serological assay to differentiate between Shigella and EIEC. Among the 93 water samples from nine reservoirs and one watershed, 76 (81.7%) water samples of culture plates had candidate colonies of Shigella and EIEC and 5 water samples were positive (5.4%) for a Shigella- and EIEC-specific polymerase chain reaction targeting the ipaH gene. Guided by the molecular method, the biochemical test, and the serological assay, 11 ipaH gene-positive isolates from 5 water samples were all identified as EIEC.     

Multi-scale temporal and spatial variation in genotypic composition of Cladophora-borne Escherichia coli populations in Lake Michigan

High concentrations of Escherichia coli in mats of Cladophora in the Great Lakes have raised concern over the continued use of this bacterium as an indicator of microbial water quality. Determining the impacts of these environmentally abundant E. coli, however, necessitates a better understanding of their ecology. In this study, the population structure of 4285 Cladophora-borne E. coli isolates, obtained over multiple three day periods from Lake Michigan Cladophora mats in 2007-2009, was examined by using DNA fingerprint analyses. In contrast to previous studies that have been done using isolates from attached Cladophora obtained over large time scales and distances, the extensive sampling done here on free-floating mats over successive days at multiple sites provided a large dataset that allowed for a detailed examination of changes in population structure over a wide range of spatial and temporal scales. While Cladophora-borne E. coli populations were highly diverse and consisted of many unique isolates, multiple clonal groups were also present and accounted for approximately 33% of all isolates examined. Patterns in population structure were also evident. At the broadest scales, E. coli populations showed some temporal clustering when examined by year, but did not show good spatial distinction among sites. E. coli population structure also showed significant patterns at much finer temporal scales. Populations were distinct on an individual mat basis at a given site, and on individual days within a single mat. Results of these studies indicate that Cladophora-borne E. coli populations consist of a mixture of stable, and possibly naturalized, strains that persist during the life of the mat, and more unique, transient strains that can change over rapid time scales. It is clear that further study of microbial processes at fine spatial and temporal scales is needed, and that caution must be taken when interpolating short term microbial dynamics from results obtained from weekly or monthly samples.     

Formulation of a mathematical model to predict solar water disinfection

A mathematical model was formulated that will facilitate the prediction of solar disinfection by analyzing the effect of sunlight exposure (x(1)) and the load of bacterial contamination (x(2)), as predictor variables, on the efficiency of solar disinfection (y). Aliquots of 0.1 ml containing average numbers of E. coli, ranging between 1 and 5 x 10(3)cells/ml raw water, were introduced into each of the 96 wells of polystyrene microtitre plates. Plates, with the lid on, were exposed to sunlight for varying exposures ranging between 1.04 x 10(3) and 8.40 x 10(3)kJ m(-2). Double strength nutrient broth was then added. After 48 h incubation wells containing visible contamination were considered as containing one cell or more that survived the exposure. Data showed that disinfection is dependent both on the load of bacterial contamination and sunlight exposure. This relationship is characterized by curves having shoulders followed by a steep decline and then tailing off in an asymptotic fashion. The shoulder size increased with the increase of the contamination load, however, the slope remains the same. Statistical analysis indicates a positive correlation among the variables (R(2) = 0.893); the mathematical model, y=1-(1-e(-kx(1)))(x(2)), represents the relationship, with k being the solar inactivation constant. The exposure required to produce a given decontamination level can be predicted using the equation: x(1)=-1/kln[1-(1-y)(-1/x(2))]e(-micro/rho.m/A), where micro is the linear attenuation coefficient (m(-1)), rho is the density, m is the mass and A is the area of the exposed part of the sample. The predictor variables (x(1), x(2)) strongly influence the efficiency of solar disinfection, which can be predicted using the suggested mathematical model. The present data provides a means to predict the efficiency of solar disinfection as an approach to improve the quality of drinking water mainly in developing countries with adequate sunshine all year-round.     

Investigating synergism during sequential inactivation of MS-2 phage and Bacillus subtilis spores with UV/H2O2 followed by free chlorine

A sequential application of UV as a primary disinfectant with and without H₂O₂ addition followed by free chlorine as secondary, residual disinfectant was performed to evaluate the synergistic inactivation of selected indicator microorganisms, MS-2 bacteriophage and Bacillus subtilis spores. No synergism was observed when the UV irradiation treatment was followed by free chlorine, i.e., the overall level of inactivation was the same as the sum of the inactivation levels achieved by each disinfection step. With the addition of H₂O₂ in the primary UV disinfection step, however, enhanced microbial inactivation was observed. The synergism was observed in two folds manners: (1) additional inactivation achieved by hydroxyl radicals generated from the photolysis of H₂O₂ in the primary UV disinfection step, and (2) damage to microorganisms in the primary step which facilitated the subsequent chlorine inactivation. Addition of H₂O₂ in the primary disinfection step was also found to be beneficial for the degradation of selected model organic pollutants including bisphenol-A (endocrine disruptor), geosmin (taste and odor causing compound) and 2,4-D (herbicide). The results suggest that the efficiency of UV/free chlorine sequential disinfection processes, which are widely employed in drinking water treatment, could be significantly enhanced by adding H₂O₂ in the primary step and hence converting the UV process to an advanced oxidation process.     

Adsorption, sedimentation, and inactivation of E. coli within wastewater treatment wetlands

Bacteria fate and transport within constructed wetlands must be understood if engineered wetlands are to become a reliable form of wastewater treatment. This study investigated the relative importance of microbial treatment mechanisms in constructed wetlands treating both domestic and agricultural wastewater. Escherichia coli (E. coli) inactivation, adsorption, and settling rates were measured in the lab within two types of wastewater (dairy wastewater lagoon effluent and domestic septic tank effluent). In situ E. coli inactivation was also measured within a domestic wastewater treatment wetland and the adsorption of E. coli was also measured within the wetland effluent.Inactivation of E. coli appears to be the most significant contributor to E. coli removal within the wastewaters and wetland environments examined in this study. E. coli survived longer within the dairy wastewater (DW) compared to the domestic wastewater treatment wetland water (WW). First order rate constants for E. coli inactivation within the WW in the lab ranged from 0.09 day⁻¹ (d⁻¹) at 7.6 °C to 0.18 d⁻¹ at 22.8 °C. The average in situ rate constant observed within the domestic wetland ranged from 0.02 d⁻¹ to 0.03 d⁻¹ at an average water temperature of 17 °C. First order rate constants for E. coli inactivation within the DW ranged from 0.01 d⁻¹ at 7.7 °C to 0.04 d⁻¹ at 24.6 °C. Calculated distribution coefficients (K_d) were 19,000 mL g⁻¹, 324,000 mL g⁻¹, and 293 mL g⁻¹ for E. coli with domestic septic tank effluent (STE), treated wetland effluent (WLE), and DW, respectively. Approximately 50%, 20%, and 90% of E. coli were “free floating” or associated with particles <5 μm in size within the STE, WLE, and DW respectively. Although 10-50% of E. coli were found to associate with particles >5 μm within both the STE and DW, settling did not appear to contribute to E. coli removal within sedimentation experiments, indicating that the particles the bacteria were associated with had very small settling velocities.The results of this study highlight the importance of wastewater characterization when designing a treatment wetland system for bacterial removal. This study illustrated the level of variability in E. coli removal processes that can be observed within different wastewater, and wetland environments.     

The impact of selected water quality parameters on the inactivation of Bacillus subtilis spores by monochloramine and ozone

Selected water quality parameters-pH, dissolved organic carbon, turbidity (NTU), and temperature-were tested for their potential effects on ozone and monochloramine inactivation of Bacillus subtilis spores. In oxidant demand-free phosphate-buffer, temperature had the strongest influence on inactivation kinetics when using ozone, pH had a smaller but significant impact on B. subtilis spore inactivation with both monochloramine and ozone. Where monochloramine was applied, modeling and experimental measurements confirmed that dichloramine levels were too low to produce significant inactivation effects under these experimental conditions. It was demonstrated that oxidant demand-free phosphate buffer may not be an adequate environmental analogue for inactivation responses in natural waters.     

Large scale analysis of virulence genes in Escherichia coli strains isolated from Avalon Bay, CA

Contamination of recreational waters with Escherichia coli and Enterococcus sp. is a widespread problem resulting in beach closures and loss of recreational activity. While E. coli is frequently used as an indicator of fecal contamination, and has been extensively measured in waterways, few studies have examined the presence of potentially pathogenic E. coli strains in beach waters. In this study, a combination of high-throughput, robot-assisted colony hybridization and PCR-based analyses were used to determine the genomic composition and frequency of virulence genes present in E. coli isolated from beach water in Avalon Bay, Santa Catalina Island, CA. A total of 24,493 E. coli isolates were collected from two sites at a popular swimming beach between August through September 2007 and from July through August 2008. All isolates were examined for the presence of shiga-like toxins (stx1/stx2), intimin (eaeA), and enterotoxins (ST/LT). Of the 24,493 isolates examined, 3.6% contained the eaeA gene, indicating that these isolates were potential EPEC strains. On five dates, however, greater than 10% of the strains were potential EPEC, suggesting that incidence of virulence genes at this beach has a strong temporal component. No STEC or ETEC isolates were detected, and only eight (<1.0%) of the potential EPEC isolates were found to carry the EAF plasmid. The potential EPEC isolates mainly belonged to E. coli phylogenetic groups B1 or B2, and carried the β intimin subtype. DNA fingerprint analyses of the potential EPEC strains indicated that the isolates belonged to several genetically diverse groups, although clonal isolates were frequently detected. While the presence of virulence genes alone cannot be used to determine the pathogenicity of strains, results from this study show that potential EPEC strains can be found in marine beach water and their presence needs to be considered as one of the factors used in decisions concerning beach closures.     

A cross comparison of QPCR to agar-based or defined substrate test methods for the determination of Escherichia coli and enterococci in municipal water quality monitoring programs

Molecular methods such as quantitative, real-time polymerase chain reaction (QPCR) are intended to shorten the period between sampling and publicly available results. Cross comparison studies in Racine, WI, USA evaluated QPCR against agar-based (US EPA Method 1600) and defined substrate (IDEXX Colilert-18^®) methods for the detection and quantification of Escherichia coli and enterococci in a variety of aqueous environments (wastewater, stormwater, and surface water). Regulatory outcomes were also compared based on choice of indicator and method. Positive correlation was seen between QPCR cell equivalents and viable cells through the wastewater treatment process and in all surface water samples (river or freshwater bathing beach) but not in direct stormwater discharge. For surface water samples, correlation improved with the application of a site-specific corrective factor, with regulatory action correctly predicted 98% of the time at bathing beaches. This study suggests the potential utility of QPCR for certain water quality monitoring applications.     

Molecular characterization of antimicrobial resistance in enterococci and Escherichia coli isolates from European wild rabbit (Oryctolagus cuniculus)

A total of 44 Escherichia coli and 64 enterococci recovered from 77 intestinal samples of wild European rabbits in Portugal were analyzed for resistance to antimicrobial agents. Resistance in E. coli isolates was observed for ampicillin, tetracycline, sulfamethoxazole/trimethoprim, streptomycin, gentamicin, tobramycin, nalidixic acid, ciprofloxacin and chloramphenicol. None of the E. coli isolates produced extended-spectrum beta-lactamases (ESBLs). The bla_TEM, aadA, aac(3)-II, tet(A) and/or tet(B), and the catA genes were demonstrated in all ampicillin, streptomycin, gentamicin, tetracycline, and chloramphenicol-resistant isolates respectively, and the sul1 and/or sul2 and/or sul3 genes in 4 of 5 sulfamethoxazole/trimethoprim resistant isolates. Of the enterococcal isolates, Enterococcus faecalis was the most prevalent detected species (39 isolates), followed by E. faecium (21 isolates) and E. hirae (4 isolates). More than one-fourth (29.7%) of the isolates were resistant to tetracycline; 20.3% were resistant to erythromycin, 14.1% were resistant to ciprofloxacin and 10.9% were resistant to high-level-kanamycin. Lower level of resistance (< 10%) was detected for ampicillin, quinupristin/dalfopristin and high-level-gentamicin, -streptomycin. No vancomycin-resistance was detected in the enterococci isolates. Resistance genes detected included aac(6′)-aph(2″), ant(6)-Ia, tet(M) and/or tet(L) in all gentamicin, streptomycin and tetracycline-resistant isolates respectively. The aph(3′)-IIIa gene was detected in 6 of 7 kanamycin-resistant isolates, the erm(B) gene in 11 of 13 erythromycin-resistant isolates and the vat(D) gene in the quinupristin/dalfopristin-resistant E. faecium isolate. This survey showed that faecal bacteria such as E. coli and enterococci of wild rabbits could be a reservoir of antimicrobial resistance genes.     

Toxicity of three phenolic compounds and their mixtures on the gram-positive bacteria Bacillus subtilis in the aquatic environment

Although phenolic compounds are intensively studied for their toxic effects on the environment, the toxicity of catechol, resorcinol and hydroquinone mixtures are still not well understood because most previous bioassays are conducted solely using single compound based on acute tests. In this work, the adverse effect of individual phenolic compounds (catechol, resorcinol and hydroquinone) and the interactive effect of the binary and tertiary mixtures on Bacillus subtilis (B. subtilis) using microcalorimetric method were examined. The toxicity of individual phenolic compounds follows the order catechol > resorcinol > hydroquinone with their respective half inhibitory concentration as 437, 728 and 934 µg mL⁻¹. The power-time curve of B. subtilis growth obtained by microcalorimetry is in complete agreement with the change in turbidity of B. subtilis against time, demonstrating that microcalorimetric method agrees well with the routine microbiological method. The toxicity data obtained from phenolic compound mixtures show that catechol and hydroquinone mixture possess synergistic effect while the other mixtures display additive joint actions. Furthermore, the concentration addition (CA) and independent action (IA) models were employed to predict the toxicities of the phenolic compounds. The experimental results of microcalorimetry show no significant difference on the toxicity of the phenolic compound mixtures from that predicted by CA. However, IA prediction underestimated the mixture effects in all the experiments.     

Inactivation of Bacillus subtilis spores with ozone and monochloramine

The inactivation kinetics of Bacillus subtilis spores with ozone and monochloramine was characterized by a lag phase followed by a pseudo-first-order rate of inactivation. The lag phase decreased and the post-lag phase rate constant increased with increasing temperature within the range investigated (1-30 degrees C for ozone, 1-20 degrees C for monochloramine). The corresponding activation energies were 46820 J/mol for ozone and 79640 J/mol for monochloramine. The CT concept was found to be valid within the concentration range investigated of 0.44-4.8 mg/l for ozone, and 3.8-7.7 mg/l as Cl(2) for monochloramine. The inactivation kinetics of B. subtilis spores with both ozone and monochloramine varied with pH within the range of pH 6-10 investigated. The fastest ozone and monochloramine inactivation rates were observed at pH 10 and 6, respectively. Different stocks of the same strain of B. subtilis spores had different resistance to ozone and monochloramine mainly because of discrepancies in the extent of the lag phase. B. subtilis spores might not be conservative surrogates for C. parvum oocysts for ozone disinfection at relatively low temperature mainly due to the spores having a lower activation energy compared to that for the oocysts. In contrast, the activation energy for monochloramine was comparable for both microorganisms but differences in the extent of the lag phase might result in the spores being overly conservative surrogates for the oocysts at relatively low temperature.