300747 Advanced Topics and Research Skills

Assessment 1: PROJECT PROPOSAL

PROJECT TITLE
Identifying risk factors for potentially-toxic cyanobacterial occurrence in stored recycled water

SUMMARY OF PROJECT PROPOSAL

Hawkesbury Water Recycling Scheme (HWRS) is a recycling scheme at the agricultural campus of
UWS which aims to treat effluent for water irrigation. Sydney Water Corporation supplies treated
effluent from stabilisation ponds at Richmond STP (Sewage Treatment Plant) to UWS for
agricultural and irrigational uses. The Hawkesbury area contains sport fields, horse paddocks and
five dams which are all used for water activities such as dairy pasture irrigation etc. This project
proposal describes the potential risk factors for toxic cyanobacterial blooms which can
unexpectedly occur in recycled water inside the dams. Immediate action is required to avoid the
harmful risks to humans, animals, livestock etc.

BACKGROUND
Hawkesbury Agricultural College which is currently known as University of Western Sydney and
Sydney Water has fostered a mutually-beneficial agreement thirty years ago. This partnership has
led to the emergence of the Hawkesbury Water Reuse Scheme, which aims to treat effluent for
water irrigation at the Hawkesbury area. This provides an opportunity for the partners to initiate
development and to raise awareness in the community about the benefits of the recycled water
(Booth, et al., 2003). Hawkesbury University campus consists of valuable sources such as vegetable
beds, lawns, fruit orchards, sport fields, deer, sheep, pasture for horses and irrigation of dairy cattle.
(Derry, et al., 2005)
Sydney Water is Australias largest water utility company, which is owned by the New South
Wales Government. The company operated on a range of different areas which include Sydney, the
Illawara and Blue Mountains districts. This water utility has a sewage treatment plant in Richmond,
which is situated at the foot of the Blue Mountains (Sydney Water, n.d.). The dry-weather flow
from this structure is received by Hawkesbury Agricultural College and then utilised for irrigation
of experimental crops and pastures (Booth, et al., 2003).
Growth of cyanobecteria has been a regular occurrence in recycled water due to a combination of
chemical, physical and biological factors. The continuous growth rate of the cyanobecteria results in
the formation of blooms or scums in water. Factors for growth are dependent on the temperature of
the water, climatic conditions, availability of nutrients, degree of stratification and the water
columns hydrodynamic stability. In many cases, phosphorus is a key element impacted in the
growth of cyanobacterial blooms due to the direct relationship between the photosynthetic pigment
chlorophyll-a and the concentration of total phosphorus (Global Water Research Coalition, 2009).
The decision of diverting the effluent away from the Hawkesbury Campus to the HawkesburyNepean (H-N) River was not a practical solution due to the continuous draught present in the area.
This restricted the use of portable water for irrigation and high costs are associated with the
replaced portable water. Due to the high presence of phosphates and nitrates in the effluent, this can

cause the eutrophication system to be aggravated and can worsen the conditions of the H-N River.
Sydneys most valuable water source is the H-N River. This is due to its many different uses such
as agriculture, aquaculture, and recreation. (Derry, et al., 2005). In 1998, Sydney Water suggested
the urban residents to drink boiled water or buy bottled water due to the presence of
microorganisms present in the water from the of Hawkesbury-Nepean River. Consumption of
contaminated water or seafood such as shellfish, crustaceans and fish can pose a risk to human
health due to the presence of alkaloid toxins (Derry, et al., 2005).
Recently in 2009, a South Australian River known as the Murray River which is located
downstream of Hume Dam was reported with 800km long growth of Algal Blooms. The factors
responsible for increasing blooms included high nutrients, high temperatures, low storage level and
often poor water quality. Due to the formation of algal blooms and the high associate risks, public
and environmental warnings were issued around the Murray River area. Based on the data, this
algal bloom became such an issue that it was reported to the local Minister for immediate action to
resolve the issue. Regular media warnings and posters were also released to provide information for
public safety especially for local campers and the users of rivers (Water Research Australia, 2009).
The word algae refers to large group of aquatic and photosynthetic plant-like organisms which have
simple retroactive systems and not vascular tissues. Algae does not have real stems, leaves and
roots. Algae is different from the phylogenetic groups and it is categorised into divisions of
taxonomic. Algae belongs to the kingdom of protiste. Algae can be mostly found in different parts
of the world which includes freshwater, seawater and in moist conditions. Generally the size of the
algae is microscopic, however they can also exist in the form of seaweed where their length can
range up to 50 meters. Algae uses photosynthesis to produce their own food for survival because
algae contains chlorophyll. Algae is considered to be eukaryotice as it contains chloroplasts. The
main role of chloroplasts is to use the bound structure of the membrane to perform photosynthesis.
Comparatively, cyanobacteria can perform photosynthesis without the requirement of specialized
organelles (Lenntech, n.d.).
Common cyanotoxins are microcystin, anatoxin-a, saxitoxin and cylinderospermopsin. Microcystin
(MCs) belong to cyanobacterias first genera, which are the most widespread hepatotoxins. MCs are
life threatening to living organisms as they are water-soluble and immediately accumulates inside
the liver. In 1996, 60 people were poisoned in Brazil due to MCs (Sylvain, et al., 2013). Similarly,
anatoxin-a (ANTX-a), which belongs to the third genera of cyanobecteria and it is extremely watersoluble. However, ANTX-a becomes unstable when it is at a pH of N10. The toxin must be exposed
to sunlight in order to render it non-toxic. This can have many negative impact on humans and
animals if this toxin is inhaled and can cause vomiting, respiratory arrest, convulsion and death
(Sylvain, et al., 2013).
Saxitoxin (STXs) are water-soluble and are starble in freshwater for more than 90 days. These can
be degraded or changed through the alteration of high temperatures. STXs prevent sodium ion
channels going into the nerve axon membrane, and can cause paralysis, numbness and dysfunction
of nerves. This can also lead to loss of life due to respiratory system failure. Due to the lack of
evidence for the presence of STXs in water, no guidelines have been developed for drinking water.
Similarly, cylinderospermopsin (CYL) is known as a tropical toxin where its half-life in highly
pure water is greater than 10 days. This toxin has a huge impact on the liver and can cause cells to
die. One study reported an incident where over 100 children had been admitted in hospital for
treatment following consumption of this present in drinking water (Sylvain, et al., 2013).
There are different type of toxins that can have a huge negative impact on ecosystems, agriculture
and wildlife. Ecosystem can be negatively affected due to high presence of cyanotoxins and they
can easily contaminate finfish, prawns and selfish. Furthermore, additional exposure to

contamination can happen through gill surfaces. Similarly Cyanobecteria can also affect wildlife
such as mammals, invertebrates and birds through drinking water which contains poison and
cyanotoxins. There is also risk for the crops to become contaminated with cyanotoxins because
water is used for spray-irrigation. One of the crop facility in United Kingdom stored water in a pond
for spray-irrigation and the water was highly contaminated with microcystins (MCs). Through
spray-irrigation, the cyanotoxins slowly spread into lettuce and other crops. The research revealed
that due to high concentration, this possessed health risks to humans. (Foundation of Water
Research, 2014).
Formation of microcystins scums can make the treatment of drinking water more difficult as it adds
additional stress. Study reveals that when cyanobecteria cells are processed through the water
treatment process, they regrow in recycled water tanks. Similarly the process is also harder as
difficulty is experienced in the process of filtration and coagulation (Global Water Research
Coalition, 2009 ).
For the past few years, UWSs Hawkesbury campus had an agreement with Sydney Water
Corporation for them to use tertiary-treated effluent for irrigation from the STP (Sewage treatment
plant) due to UWSs strong code for environmental and agriculture. In 2002, laboratory reports
from STP showed that the bacterial quality was worsened due to the supply of effluent to the
campus. After this, Sydney Water Corporation replaced STP with Intermittently Decanted Aerated
Lagoon (IDAL) in 2005 due to the supply of low water quality from STP. The replaced IDAL
provided supply of high quality of water, however study on the Schemes dam showed increase in
FC and stable effluent. One of the possible sources of effluent is group of birds which bring faecal
matter to the dam. The IDAL upgrade had a positive impact on the Hawkesbury Nepean Rivers
irrigation runoff which improved local eutrophication. Research shows that when phosphates are
leached from old sediments and extraction of faecal matter leads to algal growth.
In one of the dam, Algal blooms unexpectedly started to occur and complaints were made due to
strong odor. Investigation was carried out as one of the contractors was heavily exposed to the
contaminated water in the dam and then the risk assessment was further improved for the safety of
staff, volunteers and students around the campus (Derry, et al., 2005). For risk assessment,
qualitative and quantitative data were recorded through the use of different types of indicators.
These indicators include Thermotolerant coliform (TC), Biochemical Oxygen Demand (BOD),
Dissolve Oxygen(DO), pH ,Temperature (Temp), Conductivity (Cond.) and Suspended solids (SS).
To achieve high water quality for food-crop irrigation, monitoring point was placed closed to
supply lines control point which goes to the horticulture dam. (Aiken, et al., 2010)
Data samples for pH, dissolved oxygen and temperature were recorded in the field using relevant
Hach test kits. Results show that increase in biochemical oxygen demand (BOD) leads to
releasing of nutrients such as carbon, nitrogen and phosphorus. The process of releasing nutrients
was observed in Horticultural Dam and supports the initial interest for risk assessment. Similarly,
pathogens started to rise due to the failing STP in the Turkey Nest dam and immediate action was
required to defend against pathogens. Ecological activities must be improved for purification,
however further research is required. (Derry, et al., 2005)
My motivation for this study is that it will allow the implementation of enhanced water recycling
processes and management of recycled water. This means that the quality of water will be improved
when cyanobecteria growth is controlled and reduced. Reduction of El Nino variations climate
change and ocean flow can improve draught conditions in Hawkesbury-Nepean (H-N) River and
also reduce negative impact on marine and terrestrial ecosystems (Vecchi & Wittenberg, 2010).

Aim of this research study is to determine aquatic and environmental risks factors for cyanobecteria
blooms in stored recycled water. HWRS located at UWSs Hawkesbury campus uses treated
effluent from stabilization ponds for irrigation. This is received from Sydney Water Corporations
STP (Sewage Treatment Plant). Water flows from Richmond STP into the first storage dam (Turkey
Nest Dam) and then flows into the second storage dam (Horticulture Dam). Water is then pumped
to vegetable crops, pasture (sheep, horses, cattle, etc.), sports fields and lawns. Control of
cyanobecteria is achieved by reduction of total Nitrogen and total phosphate in water at STP as they
are the offending chemicals. Control points are used to determine the use of recycled water and
direct the flow of water to less demanding uses such as diluting nutrients with storm water,
silviculture etc. It is vital that high quality water must be used drinking and food-crop irrigation.
Research shows that agriculturalists prefer water with high nutrients for irrigation while
environmentalists prefer low lower of nutrients. (Aiken, et al., 2010)
UWS Ethic Committee approval is not required due to no participation of humans or animals in this
research project.

HYPOTHESIS
My prediction for this study is that cyanobacteria blooms grow due to high level of nutrients present
in recycled water.

SIGNIFICANCE
This research study is important as cyanobacteria or blue-green algae can cause high level of risks
to humans, environment, aquatic animals and livestock. If the growth of cyanobecteria blooms is
not controlled, this can develop and increase the chances of life threatening diseases. El Nino causes
variation in ocean flow and climate patterns which subsequently affects the equilibrium of marine
and terrestrial ecosystems (Vecchi & Wittenberg, 2010). As a result, this increased the rate of
occurrence and magnitude of cyanobecteria blooms. As a consequence, this lead to excessive
drought conditions and potential cyanobacteria blooms in the Hawkesbury-Nepean (H-N) River.
Immediate action must be taken to resolve this problem for the safety of staff, students, visitors,
Hawkesbury residents, agriculture land, farm animals etc.

EXPERIMENTAL DESIGN AND METHOD

The purpose of this research is to monitor risk factor for cyanobacteria in recycled water to enhance
water quality as cyanobacteria possess many health related problems associated with it.
Experimental Design and method for recycled water sample consists of field and laboratory
analysis. Laboratory analysis will be used to analyse water samples for different water quality
parameters. The Hawkesbury Water Reuse Scheme (HWRS) illustrates the current extensively used
water recycling schemes which makes this case study ideally suitable for this research project.
1. Weekly samples of tertiary treated sewage effluent will be collected from two of the Hawkesbury
Water Recycling Scheme (HWRS) impoundments, Turkey Nest Dam, and Horticulture (Hort) Dam.
These will include the following three dependent variables:
Phycocyanin, as proxy for cyanobacterial concentration in water
Chlorophyll-a, as proxy for algal growth in water
Chlorophyll, as indicator of all chlorophyll in water.

Analysis for these indicators using an AquaFluor 8000 field test kit and Hach laboratory
fluorometer will be carried out with regression analysis to determine relationships between the three
indicators. This will establish the validity of field monitoring as opposed to laboratory monitoring
for indicators of algal and cyanobacterial growth, in the establishment of a cost-effective
monitoring subset of indicators for the dependent variables.
2. Indication of organisms is accessible through the use of growth curves, however laboratory work
can be performed in order to record and compare changes in field as oppose to the treated effluent
with E. coli and Enterococci.
3. A photographic record of representative algal genera will be carried out using, confocal
microscopy to add a qualitative dimension to the quantitative proxy record. Identification of these
illustrative examples will be carried out to the level of genus. There are some genus of
cyanobacteria such as: the genera anabaena, schizothrix, umezakia, lyngabya and
cylindrospermopsis. (Cyanosite, n.d.)
4. Weekly assessment of the following climatic risk factors as independent variables will be
temperature (maximum and minimum), sunlight, rainfall, time of day (morning, noon, afternoon,
evening), cloud cover.
5. Weekly samples for the following risk factor for algal growth as correlated dependent variable
will be carried out:
pH- will be measured electrochemically using a combination electrode and is calibrated
against two or three commercially available buffer solution. pH meter reflects if water is
acidity (pH < 7) ,neutral (pH=7)and alkaline (pH > 7)
Temperature- can be measured by using a thermometer with a range of 0-50C or a suitable
electronic thermometer. The thermometer is placed in water to measure temperature. The
reading of the temperature remains stable for few minutes on the thermometer.
Turbidity- is caused by suspended matter in water such as clay, slit, divided inorganic and
organic matter, plankton and other microscopic organism. To achieve more accurate results
for turbidity test, laboratory analysis should be performed because probes in field give
inaccurate value and especially in very shallow water (<0.5).
Total Suspended Solids (TSS) - describe as the portion of total solids in a water sample
retained by a glass finer (GF/C) filter of probe size >2m.Water sample will be send to ALS
Lab for analysis.
Dissolved Oxygen (DO)-The amount of dissolved oxygen in water is widely dependent
upon the water temperature. Colder water can take more dissolve oxygen in comparison to
warmer water when atmosphere is balance water will be 100% saturation of dissolve oxygen
at maximum concentration. The unit of measurement is mg/L (dissolved oxygen
concentration) or % (saturation).
Electrical Conductivity (EC) - Electrical conductivity determines supply of direct
measurement of dissolve ionic matter in the water and are valuable in aquatic studies. The
Unit of measurement (mS/cm).
To achieve valid results, the results will be transferred to other organisations such as local
government, and for use of results for modification of existing water quality guidelines, chemical
analyses have to be carried out at a NATA accredited lab, so samples will be sent to ALS (277-289
Woodpark Rd, Smithfield NSW 2164) in terms of Standard Methods for the Examination of Water
and Wastewater (AS/NZS 5667.1:1998 - water quality sampling) (Department of Water, 2009).
CW&F will be paid for laboratory analysis water samples by prior agreement relating to an overarching water quality research project.

6. Field Horiba Unit (Water quality analyser) will be used to analyse water samples.
7. Following nutrients present in recycled water will be sent to ALS lab for analysis. Total
phosphorus (TP), Nitrogen oxides (
), Total nitrogen (TN), Total Kjeldahl nitrogen (TKN),
Biochemical oxygen demand (
), Nitrate-Nitrogen (
-,
-), Ammonia-Nitrogen
(
-N), and Nitrite-Nitrogen (
-N).
8. Data for all parameters will be saved using Excel as an extension of the HWRS data base.
9. My opinion will be provided for any change in total coliform and Enterococci in HWRS water
samples. This will be determined by using regression analysis. Data analysis will be carried out
using a program combination of Excel, MINITAB, and SPSS (Statistical Package for the Social
Sciences) (IBM SPSS Statistics is the most recent and current version). After determination of
normality, kurtosis and skewness, data will be subjected to bivariate linear regression analysis
and multiple regression analysis to identify simple linear relationships between dependent and
independent variables and more complex interrelationships within a model standardised for
interactive effects of co-variables.
10. Data will be presented in terms of quantitative regression results indicating the strength and
nature of relationships between the main risk factors for cyanobacterial growth, with qualitative
illustrative material to outline some genera which might flourish under relevant environmental
conditions.
11. A descriptive literature review is ongoing for two years, targeting on risk factor for
cyanobacteria in recycled water in Hawkesbury Campus (UWS).

Hawkesbury Water Recycling Scheme Health Risk Assessment Budget

#
Item
1 Salary
2 Performance
Indicators

2 Nutrients

3 Algal
Indicator

Description
Hidden Cost: staff indirectly
involved with the project
pH
Temperature
Total Dissolved Solids
Dissolved Oxygen
Total Suspended Solids
NT-8
Total Phosphorus (TP)
Nitrogen Oxides (
)
- Total Nitrogen
- Total Kjeldahl Nitrogen
- Ammonia-nitrogen
- Nitrate-nitrogen
Chlorophyll-a (Chl-a)
Chlorophyll
Phycocyanin

4 Administration Computer, Electricity etc.

5 Maintenance
Sample Storage
6 Travel and
Sending sample to laboratory
Transportation for analysis from UWS
(Handling
(Hawkesbury Campus)
Sample)

Cost ($)
1 Staff * $30p/h * 10 hours per week
* 30 weeks = $9000
$0
$0
$0
$0
$11
$60 Per Sample * 32 Samples = $1920
$28 Per Sample * 32 Samples = $896
$0
$0

$28 Per Sample * 32 Samples = $896

$0
$0
Negligible
Negligible
Travel/Week (km): 54.2km
Cost/km: 65c/km
Total Weeks: 30 Weeks
Travel/Week * Cost/km * 30 Weeks
= 54.2*0.65*30 = $1056.9

7 Analysis
Analysis of collected data
TOTAL BUDGET

Negligible
$ 13,789.90

TIMELINE
2015
M A M

2016

J A S O

N M A

Literature Review

Develop Project Proposal

Project Proposal Submission

Oral Presentation

14

14

M J

Formulate Presentation

Final Oral Presentation

HWRS Data Collection

Data Analysis
Prepare Final Report
Final Report Submission

J A S O

REFERENCES
Aiken, J. T., Derry, C. & Attwater, R., 2010. Impact of Improved Recycled water quality on a
sydney irrigation scheme. Agriculture Use, pp. 89-90.
Booth, C. A., Attwater, R., Derry, C. & Simmons, B., 2003. Water Reuse: The Hawkesbury Water
Reuse Scheme. [Online]
Available at: http://www.uws.edu.au/__data/assets/pdf_file/0020/134903/Water_2003.pdf
[Accessed 25 September 2014].
Cyanosite, n.d. The Toxic Cyanobacteria. [Online]
Available at: http://www-cyanosite.bio.purdue.edu/cyanotox/toxiccyanos.html
[Accessed 29 October 2014].
Department of Water, 2009. Surface water sampling methods and analysis technical appendices.
[Online]
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[Accessed 29 October 2014].
Derry, C., Attwater, R. & Booth, S., 2005. Rapid health-risk assesment of effluent irrgation on an
Austalian usinversity campus. International Journal of Hygene and Environmental Health, Issue
209, pp. 159-171.
Foundation of Water Research, 2014. Cyanobacterial Toxins (Cyanotoxins) in Water. [Online]
Available at: http://www.fwr.org/cyanotox.pdf
[Accessed 29 October 2014].

Global Water Research Coalition, 2009 . WQRA: Chapter 1. [Online]

Available at: http://www.waterra.com.au/cyanobacteria-manual/Chapter1.htm
[Accessed 15 November 2014].
Global Water Research Coalition, 2009. WQRA: Chapter 2. [Online]
Available at: http://www.waterra.com.au/cyanobacteria-manual/Chapter2.htm
[Accessed 27 September 2014].
Lawton, L., Marsalek, B., Padisk,, J. & Chorus, I., 1999. In: I. C. a. J. Bartram, ed. Toxic
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[Accessed 25 September 2014].
Sylvain, M. et al., 2013. State of knowledge and concerns on cyanobeterial blooms. Science Direct:
Environment International, Volume 59, pp. 303-327.
Vecchi, G. A. & Wittenberg, A. T., 2010. El Nio and our future climate: where do we stand?.
WIREs Climate Change, 1(1), pp. 260-270.
Water Research Australia, 2009. Cyanobacterial Bloom Management: Current and Future Options.
[Online]
Available at: http://www.waterra.com.au/publications/document-search/?download=158,
http://www.google.com.au/url?sa=t&rct=j&q=&esrc=s&source=web&cd=5&ved=0CEIQFjAE&ur
l=http%3A%2F%2Fwww.waterra.com.au%2Fpublications%2Fdocumentsearch%2F%3Fdownload%3D158&ei=3d9VVLPT
[Accessed 12 October 2014].