Disinfection of aquarium effluents by chlorination and UV treatment

The study was conducted to investigate the efficacy of chlorine and UV irradiation in disinfecting aquarium effluent. A non-agglutinating, a virulent strain of Aeromonas salmonicida (NCIMB 11 02) was used as the test organism. Effluents from a fish tank were inoculated with a suspension of test organisms and subsequently treated with different concentrations of hypochlorite and UV irradiation separately and simultaneously. When used alone, 1.0 ppm hypochlorite reduced the viable cell count from 6.5 log to 3.0 log within 20 minutes of contact period. On the other hand, when used in combination with UV irradiation only 0.5 ppm hypochlorite exerted the same bactericidal effect within the same contact period as was observed with 1.0 ppm hypochlorite alone. This result indicated that required dose of disinfectant for the disinfection of aquarium effluents can be considerably reduced when it is used in combination with UV irradiation.

Formation and modeling of disinfection by-products in drinking water of six cities in China

Funding and support for this project was provided by NSFC (Grant No. 40771015), and Key International Science and Technology Cooperation Projects (Grant No. 22007DFC20180). The authors also gratefully acknowledge the support of Key Projects in the National Science & Technology Pillar Program in the Eleventh Five-year Plan Period (Grant No. 2006BAD01B06-02). The authors thank the CDCs of Daqing, Beijing, Tianjin, Zhengzhou, Changsha and Shenzhen cities for field and laboratory technical support.

Antisense Gene Silencing and Bacteriophages as Novel Disinfection Processes for Engineered Systems

The growth and proliferation of invasive bacteria in engineered systems is an ongoing problem. While there are a variety of physical and chemical processes to remove and inactivate bacterial pathogens, there are many situations in which these tools are no longer effective or appropriate for the treatment of a microbial target. For example, certain strains of bacteria are becoming resistant to commonly used disinfectants, such as chlorine and UV. Additionally, the overuse of antibiotics has contributed to the spread of antibiotic resistance, and there is concern that wastewater treatment processes are contributing to the spread of antibiotic resistant bacteria.

Due to the continually evolving nature of bacteria, it is difficult to develop methods for universal bacterial control in a wide range of engineered systems, as many of our treatment processes are static in nature. Still, invasive bacteria are present in many natural and engineered systems, where the application of broad acting disinfectants is impractical, because their use may inhibit the original desired bioprocesses. Therefore, to better control the growth of treatment resistant bacteria and to address limitations with the current disinfection processes, novel tools that are both specific and adaptable need to be developed and characterized.

In this dissertation, two possible biological disinfection processes were investigated for use in controlling invasive bacteria in engineered systems. First, antisense gene silencing, which is the specific use of oligonucleotides to silence gene expression, was investigated. This work was followed by the investigation of bacteriophages (phages), which are viruses that are specific to bacteria, in engineered systems.


For the antisense gene silencing work, a computational approach was used to quantify the number of off-targets and to determine the effects of off-targets in prokaryotic organisms. For the organisms of Escherichia coli K-12 MG1655 and Mycobacterium tuberculosis H37Rv the mean number of off-targets was found to be 15.0 + 13.2 and 38.2 + 61.4, respectively, which results in a reduction of greater than 90% of the effective oligonucleotide concentration. It was also demonstrated that there was a high variability in the number of off-targets over the length of a gene, but that on average, there was no general gene location that could be targeted to reduce off-targets. Therefore, this analysis needs to be performed for each gene in question. It was also demonstrated that the thermodynamic binding energy between the oligonucleotide and the mRNA accounted for 83% of the variation in the silencing efficiency, compared to the number of off-targets, which explained 43% of the variance of the silencing efficiency. This suggests that optimizing thermodynamic parameters must be prioritized over minimizing the number of off-targets. In conclusion for the antisense work, these results suggest that off-target hybrids can account for a greater than 90% reduction in the concentration of the silencing oligonucleotides, and that the effective concentration can be increased through the rational design of silencing targets by minimizing off-target hybrids.

Regarding the work with phages, the disinfection rates of bacteria in the presence of phages was determined. The disinfection rates of E. coli K12 MG1655 in the presence of coliphage Ec2 ranged up to 2 h-1, and were dependent on both the initial phage and bacterial concentrations. Increasing initial phage concentrations resulted in increasing disinfection rates, and generally, increasing initial bacterial concentrations resulted in increasing disinfection rates. However, disinfection rates were found to plateau at higher bacterial and phage concentrations. A multiple linear regression model was used to predict the disinfection rates as a function of the initial phage and bacterial concentrations, and this model was able to explain 93% of the variance in the disinfection rates. The disinfection rates were also modeled with a particle aggregation model. The results from these model simulations suggested that at lower phage and bacterial concentrations there are not enough collisions to support active disinfection rates, which therefore, limits the conditions and systems where phage based bacterial disinfection is possible. Additionally, the particle aggregation model over predicted the disinfection rates at higher phage and bacterial concentrations of 108 PFU/mL and 108 CFU/mL, suggesting other interactions were occurring at these higher concentrations. Overall, this work highlights the need for the development of alternative models to more accurately describe the dynamics of this system at a variety of phage and bacterial concentrations. Finally, the minimum required hydraulic residence time was calculated for a continuous stirred-tank reactor and a plug flow reactor (PFR) as a function of both the initial phage and bacterial concentrations, which suggested that phage treatment in a PFR is theoretically possible.

In addition to determining disinfection rates, the long-term bacterial growth inhibition potential was determined for a variety of phages with both Gram-negative and Gram-positive bacteria. It was determined, that on average, phages can be used to inhibit bacterial growth for up to 24 h, and that this effect was concentration dependent for various phages at specific time points. Additionally, it was found that a phage cocktail was no more effective at inhibiting bacterial growth over the long-term than the best performing phage in isolation.

Finally, for an industrial application, the use of phages to inhibit invasive Lactobacilli in ethanol fermentations was investigated. It was demonstrated that phage 8014-B2 can achieve a greater than 3-log inactivation of Lactobacillus plantarum during a 48 h fermentation. Additionally, it was shown that phages can be used to protect final product yields and maintain yeast viability. Through modeling the fermentation system with differential equations it was determined that there was a 10 h window in the beginning of the fermentation run, where the addition of phages can be used to protect final product yields, and after 20 h no additional benefit of the phage addition was observed.

In conclusion, this dissertation improved the current methods for designing antisense gene silencing targets for prokaryotic organisms, and characterized phages from an engineering perspective. First, the current design strategy for antisense targets in prokaryotic organisms was improved through the development of an algorithm that minimized the number of off-targets. For the phage work, a framework was developed to predict the disinfection rates in terms of the initial phage and bacterial concentrations. In addition, the long-term bacterial growth inhibition potential of multiple phages was determined for several bacteria. In regard to the phage application, phages were shown to protect both final product yields and yeast concentrations during fermentation. Taken together, this work suggests that the rational design of phage treatment is possible and further work is needed to expand on this foundation.

A comparison between ultraviolet disinfection and copper alginate beads within a vortex bioreactor for the deactivation of bacteria in simulated waste streams with high levels of colour, humic acid and suspended solids

We show in this study that the combination of a swirl flow reactor and an antimicrobial agent (in this case copper alginate beads) is a promising technique for the remediation of contaminated water in waste streams recalcitrant to UV-C treatment. This is demonstrated by comparing the viability of both common and UV-C resistant organisms in operating conditions where UV-C proves ineffective - notably high levels of solids and compounds which deflect UV-C. The swirl flow reactor is easy to construct from commonly available plumbing parts and may prove a versatile and powerful tool in waste water treatment in developing countries.

Haematite in Lateritic Soils Aids Groundwater Disinfection

Microbiologically contaminated water severely impacts public health in low-income countries, where treated water supplies are often inaccessible to much of the population. Groundwater represents a water source that commonly has better microbiological quality than surface water. A 2-month intensive flow and quality monitoring programme of a spring in a densely settled, unsewered parish of Kampala, Uganda, revealed the persistent presence of high chloride and nitrate concentrations that reflect intense loading of sewage in the spring’s catchment. Conversely, thermotolerant coliform bacteria counts in spring water samples remained very low outside of periods of intense rainfall. Laboratory investigations of mechanisms responsible for this behavior, achieved by injecting a pulse of H40/1 bacteriophage tracer into a column packed with locally derived granular laterite, resulted in near-total tracer adsorption. X-ray diffraction (XRD) analysis showed the laterite to consist predominantly of quartz and kaolinite, with minor amounts (<5%) of haematite. Batch studies comparing laterite adsorption capacity with a soil having comparable mineralogy, but with amorphous iron oxide rather than haematite, showed the laterite to have a significantly greater capacity to adsorb bacteriophage. Batch study results using pure haematite confirmed that its occurrence in laterite contributes substantially to micro-organism attenuation observed and serves to protect underlying groundwater.

Disinfection of Burkholderia cepacia complex from non-touch taps in a neonatal nursery

Burkholderia cepacia complex (Bcc) comprises nine closely related species or genomovars. It is an important causative agent of opportunistic infections and waterborne nosocomial infections. B. cepacia (formerly genomovar I) was identified from the blood culture of a baby in our neonatal unit (NU) in March 2005. B. cepacia was isolated four times from clinical specimens since the introduction of non-touch taps in the NU from 2000 to 2005 and only once from 1994 to 2000. Environmental samples were collected from the NU, including tap water from non-touch taps. Clinical and environmental isolates of Bcc were characterized using molecular identification and strain typing. A literature review was undertaken to delineate a method for eradication of Bcc. Several variations for hot water eradication of the organism from the taps were attempted. Genotyping and molecular analysis revealed that tap water isolates were B. cenocepacia which was a different species from the B. cepacia isolated from blood cultures of the neonate. However, B. cenocepacia has been known to cause nosocomial outbreaks and it was eventually eradicated from the NU by using repeated thermal shock (hot water at 65 degrees C for 10 min), changing taps and decolonizing sinks with hypochlorite. Molecular typing is useful in assisting the investigation of Bcc nosocomial infections.

The application of TiO2 photocatalysis for disinfection of water contaminated with pathogenic micro-organisms: a review

The use of semiconductor photocatalysis for treatment of water and air has been the topic of intense research activity over the past 20 years. This powerful process has also been extended to the disinfection of environments contaminated with pathogenic micro-organisms. This review summarizes recent developments concerned with the photocatalytic treatment of water contaminated with pathogenic micro-organisms presenting a potential hazard to animals and human beings.

Mechanistic studies of the photocatalytic oxidation of microcystin-LR: An investigation of byproducts of the decomposition process

Microcystins (cyclic heptapeptides) are produced by a number of freshwater cyanobacteria and cause concern in potable water supplies due to their acute and chronic toxicity. The present study reports the structural characterization of the degradation products of the photocatalytic oxidation of microcystin-LR, so aiding the mechanistic understanding of this process. TiO₂ photocatalysis is a promising technology for removal of these toxins from drinking water. However, before it can be adopted in any practical application it is necessary to have a sufficient knowledge of degradation byproducts and their potential toxicity. Liquid chromatography-mass spectrometry analysis demonstrated that the major destruction pathway of microcystin appears to be initiated via three mechanisms: UV irradiation, hydroxyl radical attack, and oxidation. UV irradiation caused geometrical isomerization of microcystin converting the (4E), (6E) of the Adda configuration to (4E), 6(Z) or 4(Z), 6(E). Hydroxyl radical attack on the conjugated diene structure of Adda moiety produced dihyroxylated products. Further oxidation cleaved the hydroxylated 4-5 and/or 6-7 bond of Adda to form aldehyde or ketone peptide residues, which then were oxidized into the corresponding carboxylic acids. Photocatalysis also hydrolyzed the peptide bond on the ring structure of microcystin to form linear structures although this appeared to be a minor pathway.

UV dosimetry for solar water disinfection (SODIS) carried out in different plastic bottles and bags

Solar water disinfection (SODIS) is a well-established inexpensive means of water disinfection in developing countries, but lacks an indicator to illustrate its end-point. A study of the solar UV dosage required for SODIS, in order to achieve a bacteria concentration below the detection limit for: Escherichia coli, Enterococcus spp. and Clostridium perfringens, in water in PET bottles, PE and PE/EVA bags showed disinfection to be most efficient in PE bags, with a solar UV (290–385 nm) dose of 389 kJ m−2 required. In parallel to the disinfection experiments, a range of polyoxometalate, semiconductor photocatalysis and photodegradable dye-based solar UV dosimeter indicators were tested under the same solar UV irradiation conditions. All three types of dosimeter produced indicators that largely and significantly change colour upon exposure to 389 kJ m−2 solar UV; further indicators are reported which change colour at higher doses and hence would be suitable for the less efficient SODIS containers tested. All indicators tested were robust, easy to use and inexpensive so as not to add significantly to the attractive low cost of SODIS. Furthermore, whilst semiconductor photocatalyst and photodegradable dye based indicators are disposable, one-use systems, the polyoxometalate based indicators recover colour in the dark overnight, allowing them to be reused, and hence further decreasing the cost of using indicators during the implementation of the SODIS method.

Comparative effectiveness of solar disinfection using small-scale batch reactors with reflective, absorptive and transmissive rear surfaces

This study investigated the enhancement of solar disinfection using custom-made batch reactors with reflective (foil-backed) or absorptive (black-backed) rear surfaces, under a range of weather conditions in India. Plate counts of Escherichia coli ATCC11775 were made under aerobic conditions and under conditions where reactive oxygen species (ROS) were neutralised, i.e. in growth medium supplemented with 0.05% w/v sodium pyruvate plus incubation under anaerobic conditions. While the addition of either an absorptive or a reflective backing enhanced reactor performance under strong sunlight, the reflective reactor was the only system to show consistent enhancement under low sunlight, where the process was slowest. Counts performed under ROS-neutralised conditions were slightly higher than those in air, indicating that a fraction of the cells become sub-lethally injured during exposure to sunlight to the extent that they were unable to grow aerobically. However, the influence of this phenomenon on the dynamics of inactivation was relatively small