Smaller and regional Wastewater Treatment Plants (WWTPs) have the capacity to recycle wastewater for agricultural use, but the cost of obtaining regulatory approval or ‘accreditation’ is prohibitive. One reason for this is that each WWTP must demonstrate that its processes and operations consistently remove pathogens that cause infectious diseases in humans. Operating conditions include flow rate through the WWTP, and temperature in the activated sludge component of the WWTPs. Although pathogen ‘log removal values (LVR)’ were obtained for a WWTP at 19-20°C in Part I of this project (WQRA project 2001), these values cannot also be attributed to summer temperatures of 26°C. This research determined LRVs for ‘new’ WWTP operating conditions and combined the data with data from Phase I (Project 2001) for analysis. One of the conclusions from Part II was that faster flow rates associated with increased rainfall reduced pathogen LRVs.

Ultrafiltration membranes are used to remove viruses from treated wastewater. This makes it safe for release to the environment or for recycling, but it is important to monitor integrity of the membrane to ensure there are no damaged sections that viruses can break through. This research demonstrated that a silver nanoparticle is a valid, safer alternative to using non-infectious bacteriophage viruses that are currently used to test membrane integrity. The silver nanoparticle was tested and validated in a laboratory-scale ultrafiltration membrane unit. It was concluded that work should proceed to full-scale validation and integrity-testing of ultrafiltration membranes in recycled water applications.

Sewage is delivered to wastewater treatment plants (WWTPs) where benign microbial organisms within ‘activated sludge’ vessels contribute to the removal of harmful pathogens from the sewage. The activity and pathogen-removing ability of these helpful organisms is affected by many factors including temperature, numbers of fine particles, pH, ammonia, and the time available to remove the pathogens. Regulatory authorities require at least 90% (one log removal value, LRV) of the pathogens to be removed, but as WWTP operating conditions vary, the LRVs change. This problem led to recognition of the need to develop models capable of predicting relationships between plant operating parameters (such as temperature) and pathogen removal. This research reviewed published reports and datasets, then set up and ran an experimental activated sludge pilot plant to generate data about a range of operating conditions and pathogen removals. These datasets were used to develop models which had only a ‘poor’ predictive value for clostridia but were ‘good’ for giardia and ‘very good to excellent’ for the removal of other pathogens. These models need to be extended with more operating conditions but have the potential to be used to attribute LRVs and for future integration into online real-time monitoring.

‘Microbial source tracking’ (MST) is a technique that aims to identify the animal that excreted faeces and polluted water. There are a number of ways to do this, but the problem is that no one method accurately identifies the origins of faecal pollution in environmental water samples. This research found that faeces could be stored in a freezer or a laboratory -80°C cold-store for up to a month without changing the relative numbers of the different types of bacteria in the samples of faeces. Up to seven faeces samples from different animals were mixed together and examined using 17 techniques to identify the original animals. Three of the most accurate and reliable methods used mitochondrial DNA, the analysis of a bacterial enzyme sequence (beta-glucuronidase), and specific DNA sequences form bacteria known to come from humans, horses and cows. These three types of tests were selected for inclusion in a ‘Toolbox’ from which a combination of methods will allow accurate and reliable management of faecal contaminants in source waters.

Chlorine removes harmful pathogens from water but has the disadvantage of forming disinfection by-products (DBPs) by reacting with organic matter sometimes found in water. Chloramine also disinfects, is less likely to form DBPs and is more stable, so remains active in for longer in the pipelines which distribute drinking water from the plant to the tap. The problem is that it is difficult to predict exactly how much chloramine to add; it needs to be enough to maintain disinfecting activity in the pipeline distribution system, but not so much that customers find the smell of chlorine in tap water unpleasant. Traditionally, the chemical reaction rate has been used to predict the gradual ‘decay’ of chloramine in pipelines, but this is inaccurate. This research developed a computer software statistical programme that uses ‘artificial neural network’ concepts and operations to predict the longevity of chloramine residuals in water distribution systems. This is more accurate than traditional methods.

Pathogenic microscopic organisms in source waters pose a risk to public health if water treatment plants do not remove them. It was thought that sensitive PCR tests could be developed to inform decision-making about the most appropriate treatment processes, and to check the absence of pathogens from drinking water. This research focussed on four pathogen classes: cryptosporidium, microcystis, adenovirus and ammonia oxidising bacteria; and evaluated six DNA extraction kits. The Qiagen kit was most cost-effective for extracting DNA and Promega To-Taq polymerase was best for carrying out the PCR test on pathogens in real-world water samples. Other components of the PCR tests that were developed included test controls and DNA standards. A test for each class of pathogen was established and written as a ‘Standard Operating Protocol’ (SOP) which was then applied in different laboratories around Australia. Between-laboratory comparison of results showed the developed PCR tests to be highly reproducible and reliable. They can now be added to the existing suite of tools used to minimise risks to public health.