Seaweed

Agronomy
Cropping in inland saline
groundwater evaporation
basins

A report for the Rural Industries Research

and Development Corporation

by Robert Cordover

April 2007

RIRDC Publication No 07/033

RIRDC Project No SWT-1A
© 2007 Rural Industries Research and Development Corporation.
All rights reserved.

ISBN 1 74151 432 0

ISSN 1440-6845

Seaweed Agronomy, cropping in inland saline water evaporation basins

Publication No. 07/033
Project No.SWT-1A

The information contained in this publication is intended for general use to assist public knowledge and discussion
and to help improve the development of sustainable regions. You must not rely on any information contained in
this publication without taking specialist advice relevant to your particular circumstances.

While reasonable care has been taken in preparing this publication to ensure that information is true and correct,
the Commonwealth of Australia gives no assurance as to the accuracy of any information in this publication.

The Commonwealth of Australia, the Rural Industries Research and Development Corporation (RIRDC), the
authors or contributors expressly disclaim, to the maximum extent permitted by law, all responsibility and liability
to any person, arising directly or indirectly from any act or omission, or for any consequences of any such act or
omission, made in reliance on the contents of this publication, whether or not caused by any negligence on the
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The Commonwealth of Australia does not necessarily endorse the views in this publication.

This publication is copyright. Apart from any use as permitted under the Copyright Act 1968, all other rights are
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should be addressed to the RIRDC Publications Manager on phone 02 6272 3186.

Researcher Contact Details

Robert Cordover
71 Mountain Road
Allens Rivulet
Tasmania 7150
Phone: 03 6239 6784
Fax: 03 6239 6784
Email: cordover@seaweedtrading.com

In submitting this report, the researcher has agreed to RIRDC publishing this material in its edited form.

RIRDC Contact Details

Rural Industries Research and Development Corporation
Level 2, 15 National Circuit
BARTON ACT 2600
PO Box 4776
KINGSTON ACT 2604

Phone: 02 6272 4819

Fax: 02 6272 5877
Email: rirdc@rirdc.gov.au.
Web : http://www.rirdc.gov.au

Published in April 2007

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ii
Foreword
The objective of this project is to develop an agronomy of Gracilaria seaweed. Mobilisation of
geologically stored salt and rising saline groundwater tables in parts of Australia have resulted in
thousands of hectares of salinised, degraded and waterlogged soils that can no longer support
traditional crops. The development of an agronomy for Gracilaria seaweed could enable the
profitable transformation of destructive saline groundwater into a useful resource that could provide
Australian farmers with a new export crop. Gracilaria seaweed is the raw material for the extraction
of agar gel used as a thickener in the food and cosmetics industries and as a base for tissue culture and
bacteriological media. A variety of Gracilaria chilensis, a red seaweed that is the source of most of
the world’s agar, is native to Australian waters, though not in commercially harvestable quantities.

This study is the first step toward an agronomy of Gracilaria seaweed in inland saline water basins
and provides the results of field studies using saline groundwater from several aquifers in Victoria to
test the hypothesis that these waters can support a commercial rate of growth for the seaweed. Since
nearly every aquifer has a unique chemistry, the results demonstrate the need for bio-chemical analysis
of each aquifer to test for seaweed viability. A review of the seaweed literature and the results of bio-
assays in four saline groundwater sources provide a baseline of information from which the agronomy
is being developed.

This project was funded from RIRDC core funds which are provided by the Australian Government.
Part of the strategic plan for the Environment and Farm Management R&D sub-program is to support
emerging agriculture technologies that will underpin Australia’s global competitiveness. The large
international demand for seaweed products and the potential to develop a resource from the currently
significant environmental problem of degraded saline lands meets the program objectives of
supporting sustainable agricultural production systems, addressing the threats and impediments to
sustainable agriculture and encouraging the emergence of new technologies.

This report, an addition to RIRDC’s diverse range of over 1600 research publications, forms part of
our Environment and Farm Management R&D program, which aims to support innovation in
agriculture and the use of frontier technology to meet market demands for accredited sustainable
production.

Most of our publications are available for viewing, downloading or purchasing online through our
website:

downloads at www.rirdc.gov.au/fullreports/index.html
purchases at www.rirdc.gov.au/eshop

Peter O’Brien
Managing Director
Rural Industries Research and Development Corporation

iii
Acknowledgments
This study benefited from the generous cooperation of Goulburn-Murray Water who provided free use
of the test ponds at their Girgarre evaporation basin for this work.

Thanks to George Wilson for his continued support and to Nica Cordover for her help with scientific
direction, her encouragement and her outstanding patience with the slow pace of agronomic
development of seaweed cultivation.

I am grateful for the encouragement and generous intellectual and financial support given to this study
by Phil Dyson, Rob Fisher, Susie Hedt, Ming-Long Liao and the CRC for Biopolymers, Dennis
McHugh, Warwick Noonan, Sergey Shabala, John West, and members of the Donald and District
Landcare Group.

Abbreviations
Measurements of the nutrients dissolved in water are made in mg/l (milligrams per litre) which is
about equivalent to ppm (parts per million). For seawater, salinity is often shown in ppt (parts per
thousand) or grams (of salt) per litre (of water).

For trace elements, i.e. small quantities of solutes, μg/l (micrograms per litre) is used. The
abbreviation M is occasionally used, following certain research papers. M stands for Molar, the mass
of an element divided by its atomic weight in a litre of water, for example 14 mg of nitrogen per litre
is one Mole (ar). Also mM (milli mole (ar), one thousandth Molar) and μΜ (micro-mole (ar), one
millionth Molar) are used.

For those not familiar with chemistry, the abbreviation pH (the hydrogen pressure) is a logarithmic
measure of the disassociation of water molecules (HOH) between 1 (very acid or high in H+ hydrogen
ions) and 14 (very basic or high in OH- hydroxyl ions), with distilled water at pH 7 being neutral.

Among the ions and elements found in seawater and saline groundwater, about twenty are relevant to
the agronomy of seaweed and they are often identified by an abbreviation, which is their symbol in all
international chemical literature. These are:

B - boron ( and its compound H3BO3, borate)

• C - carbon (and its compound HCO3 bicarbonate and the gas CO2 carbon dioxide)
• Ca - calcium
• Cl - chlorine
• Cu - copper
• Fe - iron
• K - potassium
• Mg - magnesium
• Mn - manganese
• Mo - molybdenum
• N - nitrogen
• Na - sodium
• O - oxygen (and the eponymous gas O2)
• P - phosphorous (and it’s compound PO4= phosphate)
• S - sulphur (and its compound SO4= sulphate)
• Zn - zi

iv
Contents
FOREWORD ...........................................................................................................................III
ACKNOWLEDGMENTS ........................................................................................................ IV
ABBREVIATIONS.................................................................................................................. IV
EXECUTIVE SUMMARY ....................................................................................................... VI
1. INTRODUCTION: BACKGROUND TO THE PROJECT ..................................................1
1.1. Salinised farmland....................................................................................................................... 1
1.2. Productive use of saline ground water......................................................................................... 2
1.3. Comparison of groundwater with seawater................................................................................. 2
1.5. Objectives of the research ........................................................................................................... 5
1.6. Markets for Gracilaria seaweed.................................................................................................. 5
1.6.1. Agar ..................................................................................................................................... 6
1.6.2. Commercialisation Strategy ................................................................................................ 7
1.7. Potential benefits of seaweed cultivation .................................................................................... 7
2. LITERATURE REVIEW ....................................................................................................9
2.1. Current Gracilaria farming production methods ........................................................................ 9
2.1.1. Constraints to Gracilaria Seaweed Farming ..................................................................... 11
2.1.2. Spores ................................................................................................................................ 12
2.2. Seaweed Physiology and Nutrition ........................................................................................... 13
2.2.1. Physiological limits in inland saline groundwaters ........................................................... 15
2.2.2. Growth rates of Gracilaria................................................................................................ 16
2.2.3. Nutrient media for algae.................................................................................................... 19
3. METHODOLOGY............................................................................................................20
3.1. Seaweed species selection ......................................................................................................... 20
3.1.1. Why Gracilaria? ............................................................................................................... 21
3.1.2. Taxonomy and local eco-types.......................................................................................... 22
3.2. Description of sites and basins .................................................................................................. 23
3.2.1. Girgarre ............................................................................................................................. 23
3.2.2. Jeparit ................................................................................................................................ 24
3.2.3. Lake Charm ....................................................................................................................... 24
3.2.4. Donald ............................................................................................................................... 25
3.3. Experimental methods and equipment ...................................................................................... 26
3.4. Groundwater chemistry at trial sites.......................................................................................... 27
3.4.1. Girgarre groundwater ........................................................................................................ 28
3.4.2. Japarit groundwater ........................................................................................................... 30
3.4.3. Lake Charm groundwater .................................................................................................. 30
3.4.4. Donald groundwater .......................................................................................................... 31
3.4.5. Use of nutrients ................................................................................................................. 32
4. RESULTS AND DISCUSSION .......................................................................................33
4.1. Seaweed assessment trials ......................................................................................................... 33
4.2. Growth studies and nutrient use ................................................................................................ 34
4.2.1. Girgarre ............................................................................................................................. 34
4.2.2. Jeparit ................................................................................................................................ 39
4.2.3. Lake Charm ....................................................................................................................... 41
4.2.4. Donald ............................................................................................................................... 41
4.3. Agricultural technology............................................................................................................. 42
5. IMPLICATIONS AND RECOMMENDATIONS ...............................................................43
Implications for industry ................................................................................................................... 43
Implications for Communities........................................................................................................... 44
Implications for Policy makers.......................................................................................................... 44
Recommendations ............................................................................................................................. 44
6. GLOSSARY ....................................................................................................................46
7. REFERENCES................................................................................................................47

v
Executive Summary
What the report is about
This report summarises the results of studies leading to an agronomy of the seaweed Gracilaria
chilensis in saline groundwater evaporation basins. An understanding of seaweed farming, physiology
and nutrition is based on the existing literature, which is reviewed in this report. There is an enormous
difference between monitoring seaweed growth in a glass aquarium and in a basin excavated in the
soil of a salt affected paddock. Not only is there an increase in scale but there are chemical dynamics
between soil, saline groundwater and a range of environmental variables that cannot always be
predicted. This report is about the field trials for seaweed cultivation in saline groundwaters where the
goal has been to achieve a growth rate of Gracilaria greater than 3% per day.

Target audience
The research was done to assist those farmers disadvantaged by saline groundwater and to enable them
to consider these waters as a resource for a crop that can be cultivated in saline water. Although the
report could be used by academic agricultural scientists to gain some appreciation of the issues and
problems of seaweed agronomy, it is primarily written as a tool to interest and inform farmers who are
seeking some hopeful news about their salt affected paddocks. The results have not been sufficiently
conclusive to provide a manual of seaweed cultivation in inland saline waters but do show that the
agronomic principles being developed for seaweed crops that could make currently degraded lands
productive again using saline groundwater as a resource.

Background
Studies of the physiology and nutrition of seaweeds have shown that despite the specialised
environment in which they live, most of the macro-algae (as seaweeds are more precisely called)
require the same basic mix of nutrients and have the same metabolic processes as the terrestrial crops
upon which we depend for our traditional grains. Only some of the mineral concentrations vary.
Seawater is essentially the same all over the world and contains all of the elements necessary for
growth and maintenance of marine plants. The chemistry of saline groundwater, on the other hand,
varies in each aquifer depending on how long it has been underground, it’s history and the types of
rock and soil with which it has been in contact. In the Murray River basin where the ground waters
are essentially marine, recent geological history has modified these ground waters and many of the
changes in mineral concentration of the salts are beyond the physical or bio-chemical ability of the
plants to respond favourably with sustained growth. High concentrations of particular salts are toxic
to the macro-algae, even where the total salinity is within the plant’s range of viability.

Objectives
The specific aim of this research is to develop an agronomy and associated technologies for the
cultivation of of Gracilaria seaweed so that the cropping of this marketable species could be
introduced into regions where saline groundwaters have degraded farmland in Australia.
The major beneficiaries of the research are expected to be farmers who currently have salinity
problems and need a crop to substitute for the loss of traditional crops on their salt-degraded land. The
production of seaweed could enable farmers to take advantage of our saline groundwater resources to
produce export crops worth more per hectare than the current suite of grain crops. Gracilaria would
be dried and exported in bales at $500-1000 per tonne to processors for the production of agar gel,
which has a growing international market as an additive in the food and biotechnical industries and
takes more than 35,000 tonnes of dried Gracilaria per annum.

vi
Methods
The selection of Gracilaria chilensis as the species for cultivation was based on its large world market
for the extraction of agar. Several varieties and eco-types of the Gracilaria were used in this study
and one type was selected for its wide salinity tolerance, high gel strength and good growth compared
to other eco-types of this seaweed.

Due to wide variations in the chemical composition and salinity of the groundwaters between
catchments and between irrigated and dryland salinity regions, four sites in Victoria were selected for
field trials: two irrigated salinity sites at Girgarre and Lake Charm and two dryland salinity sites at
Jeparit and Donald. The waters in these trials were indeed dissimilar, the Jeparit being unable to
support Gracilaria because it was too rich in iron and others requiring supplements of potassium
necessary for plant survival.

Results/Key findings:

• The outcomes achieved have not yet established a new paradigm that saline ground water can be
used as a productive agricultural resource to provide a new income stream for farms that have lost
the ability to harvest traditional crops from salinsed land. Pilot trials in small on-farm evaporation
basins have begun to establish the base line requirements and preliminary growth rates for the
seaweeds in inland saline groundwaters. If the methods become commercially viable, seaweed
harvests could mitigate the costs of engineering investments and operating costs for salinity
rehabilitation programs as well as providing a profitable new export crop for Australia.
• Whilst commercial quantities have not yet been harvested, sufficient evidence has been collected
to support detailed seaweed physiological studies and encourage further field trials that can lead to
sustainable and profitable cropping of Gracilaria seaweed.
• There is a limited range of chemical composition of the saline groundwater medium in which
Gracilaria will flourish, with a baseline of “normal” seawater. Either critically limiting nutrients
or a super-abundance of certain minerals will result in the death of the plant in an active process of
chemical transfer.
• Growth rates in appropriate groundwaters have only reached an unsustained 2.5% per day, far
lower than the potential shown in the literature.
• Until now seaweed agronomy research has been structured around field trials that might have been
immediately transferable to farmers but the results have been disappointing and suggest that the
scientific literature consulted was not a sufficient basis for the proposed development.

Implications:

• The profitable and productive use of saline groundwater could provide a chance to recover lost
income where farmland and infrastructure are being degraded by salinisation of soils.
• The export market for seaweed has a potential value to farmers measured in the tens of millions of
dollars
• A factory to extract gel or add value to the product could possibly be established in small
communities to increase employment and support spin-off industries.
• Whilst commercial quantities have not yet been harvested, sufficient evidence has been collected
to support detailed seaweed physiological studies and encourage further field trials.

vii
Recommendations
Seaweed cultivation in inland saline groundwater still has agronomic potential; and future research
and development is required to understand the limitations of our current systems.

A systems research approach is recommended, including hydro-geology, agronomy, water chemistry

and soil-water interaction, extension, engineering and business skills to develop technologies,
agronomy and models for profitable cropping of seaweed in saline groundwaters.

Seaweed physiology, photosynthesis and diagnostic information on seaweed nutrition needs to be

studied in the laboratory. Results of seaweed growth trials under laboratory conditions would inform
field trials.

There is a need to maintain focus on closely managed field trials at salt-degraded farm sites at Donald
and Lake Charm (Victoria) and Morawa (WA) that involve farmers at the earliest stages of technology
development to establish reliable agronomic protocols for crop and water management that maximise
both growth and gel quality of the harvest.

Agronomic models should consider the whole of the value chain, starting with the market demand for
raw material and moving backward to plan harvests for seaweed gel quality and yield that best meet
market needs.

viii
1. Introduction: Background to the Project
1.1. Salinised farmland
Mobilisation of geologically stored salt and rising salt water tables in the Murray Darling Basin have
reduced the sustainability of farming where highly saline groundwaters are beyond the tolerance of
most non-marine plants and soil animals. The damages and costs of rising water tables and soil
salinisation were summarised in the 1999 Salinity Audit of the Murray Darling Basin
(Inter-governmental Basin Salinity Working Grou, 1999). Much of the mobilised salt remains in the
landscape or is diverted into irrigation areas where the total economic impact was estimated at
$46 million a year and is expected to rise over the next century (idem). Lands salinised and
waterlogged by both irrigation salinity and dry land salinity in the Murray Darling Basin and Western
Australian Wheatbelt comprise hundreds of thousands of hectares that no longer support traditional
crops. In some cases the salinity is too severe to support any terrestrial plants that can be used for
crops, wood or pasture and limits the potential for any return of the original ecological diversity.
Much of this land is considered to be irretrievable for farming.

Although somewhat slowed by the drought since 2000, the problem of groundwater salinity has been
getting worse over the last few decades and is revealed by rising saline ground water tables that come
close enough to the surface to destroy crops and pasture. Saline groundwaters can be divided into at
least two categories: irrigated salinity and dry-land salinity.

Irrigated salinity starts as surface drainage of non-salinised water and captures salts from the soil as it
passes the shallow root zone of crops. It may meet a shallow saline ground water layer adding further
to the salt load. Dryland salinity comes from ancient saline water aquifers that may be 10-40 metres
below the surface flowing through paleo-channels directly beneath or adjacent to current catchment
drains. Upstream hydrostatic pressure forces water to the surface downstream, where the catchment
land surface is lower than the upstream water surface. In many instances, removal of trees and/or
inappropriate farming methods have raised the water levels not only in discharge areas but also in
recharge areas, resulting in surface or near surface saline water discharges far downstream.

An engineering solution to reverse the rising salt water recommends the excavation of salt water
evaporation basins filled by pumping ground water to lower the water table below the root zone of
traditional crops. Disposal evaporation basins have been shown to be the most effective and least-cost
option (Evans 1989) for removing high salinity drainage water and have become necessary to protect
productive farmland, town infrastructure and downstream water supplies from rising saline water
tables. Both community scale and on-farm basins, together with deep drains and pumping from spear
points or bores are used to maintain water tables below the critical level for agriculture and to limit the
export of salt outside irrigation areas. Although adjacent land is recovered from salt danger, these
evaporation basins take other land out of production and incur operating costs for pumping and
drainage without providing any direct return to the farmers or irrigation companies from the land used
for basins.

There are currently more than 150 of these salt water interception and evaporation basins throughout
the Murray Basin from Griffith, NSW to Waikerie, SA (Hostetle and Radke 1994). Eleven large-scale
saline groundwater interception schemes and their associated disposal basins in the Murray River
Basin contain 6250 hectares, were built at a cost of more than $108 million and cost more than
$3 million a year to operate, disposing of nearly 50,000 Mega-litres of salt water each year. Another
eight large schemes for the disposal of an additional 13,000 ML have been planned (Allan et al 2000).
In addition, on-farm evaporation ponds have been proposed in the region to rehabilitate individual
properties.

1
1.2. Productive use of saline ground water
It is well known that seawater supports a very wide range of marine life. Even when enclosed in tidal
basins or other containers, seawater can still support marine organisms if other aspects of the
environment (temperature, pH, pollutants, etc.) are maintained within the range required by the
organisms being cultivated.

Nearly every element in the periodic table can be found in seawater. Less than twenty of these
elements are required by seaweeds (Lobban & Harrison, 1994) and most are found in sufficient
quantity in nearshore waters to support the plants found there. This does not mean that the elements
are always in a nutrient form immediately available to the plants. Seawater is a dynamic mixture of
dissolved and particulate organics, inorganic minerals and living organisms that may be limiting to
plant growth under a variety of environmental conditions. In some cases seaweeds must wait their
turn to grow after a succession of developments in surface waters between nutrients, micro-algae
photosynthesis, bacteria and organic compounds, such as vitamins. For background information on
the composition and fertility of seawater, see articles by Goldberg, by Redfield et al and by Provasoli
in (Hill 1963).

Where saline groundwater has the equivalent concentration of the minerals necessary for seaweed
growth, it could be useful for cropping of marine plants. In addition to the static concentration of
minerals, saline ground water could have certain dynamic properties that might make these waters not
immediately useful. One example is pH, which can be very acidic in ground waters that contain high
levels of carbon dioxide. This could result in the release of toxic metals from surrounding substrates
and prevent the healthy functioning of the plant.

Although saline groundwater in the Murray River basin is essentially marine water, in that it is
basically a mix of salts dominated by sodium and chloride like the sea, groundwater salinity varies
widely between catchments. Even within short distances in the same catchment, groundwater varies
and changes salinity and ionic composition in three dimensions, as aquifers pile one atop another,
separated by clay layers.

Nevertheless, the saline groundwater and the evaporation basins can be considered together as a
resource as well as an environmental expense. Use of these inland marine waters for aquaculture,
especially those waters with an ionic composition close to seawater, has been considered as a potential
use of saline evaporation basins for nearly a decade. RIRDC helped to sponsor a workshop in 1997
(Smith and Barlow 1999) at which representatives from four states (NSW, VIC, SA and WA)
discussed the aquaculture trials, plans and potential of more than three dozen species of fish, algae,
molluscs and crustaceans.

1.3. Comparison of groundwater with seawater

Currently saline groundwater is a major problem for traditional crops but were “normal” oceanic
seawater (see Glossary, page 45) available as ground water, it could be used to produce any of the
marine plants we currently harvest from the sea.

The first question that needs to be raised in considering the agricultural use of saline groundwater is,
how similar is it to seawater? Of the six elements that constitute >99% of oceanic salt (Na, Cl, SO4,
Mg, Ca, K) all are usually found in ionic (see Glossary, page 45) form and are necessary for plant
growth. Some groundwater, even saline groundwater, may have more bicarbonate (HCO3 -) than
sulfate (SO4++) and more silicon (Si) than potassium (K). All of the elements required for seaweed
growth are present in the oceans, in low concentrations that are not toxic, do not interfere with each
other and are present in forms that can be actively (and passively) concentrated to meet the needs of
the plant. This is not always the case with groundwaters, which may have toxic concentrations of
some elements and critically insufficient concentrations of others, pH not conducive to seaweed

2
 viability and may be lacking in organics that can provide chaelated (see Glossary, p 40) metals or
vitamins required by the plants.

The first limiting nutrient in seawater is usually nitrogen, then phosphorous and occasionally iron,
though iron is more likely to be limited in the open ocean rather than nearshore waters. The
combination of run-off from the land and a large component of organic material in nearshore waters
assists the bacterial breakdown of inorganic compounds and provides the organic matrices to make
some of the necessary minerals available to plants by chaelation. The enormous reservoir of water in
the ocean, its nutrient load and the movement of water past the seaweed, distributes sufficient nutrients
to them even where the concentration of the nutrient in seawater is only a few micrograms per litre
(parts per billion). Marine plants can actively concentrate some the micronutrients they need, like Zn,
Fe, Mn, and even N and P, more than 100,000 times their usual concentration in seawater (Lobban and
Harrison 1994).

Table 1 provides a comparison between seawater and some samples of saline groundwater in regions
being considered for seaweed cultivation. Where possible, comparisons were made of the minerals
necessary for seaweed growth. Most of the elements in seawater are found in their ionic form, though
when the water is evaporated, the elements come together to form salts such that we can speak about
NaCl (sodium chloride) when we discuss salt water.

Table 1. Comparison of the concentration of elements in seawater with that in saline

groundwater in regions where seaweed cultivation is under consideration (since the total
salinity varies among the waters tested, comparisons are made as percentages of total salinity)
Concentration Percent in Percent in Percent in
in seawater Percent in Morawa Peter's Paul's Ercent in
Element Ionic form mg/ l seawater WA Donald Donald Lake Charm
Cl Cl- 19000 55.1% nd nd nd nd
Na Na+ 10556 30.6% 36.1% 29.3% 29.9% 22.6%
S SO4= 2649 7.7% 9.0% 9.5% 11.5% 14.8%
Mg Mg++ 1311 3.8% 3.6% 3.7% 3.8% 3.0%
Ca Ca++ 400 1.2% 0.8% 1.3% 1.4% 1.6%
K K+ 380 1.1% 1.4% 0.45% 0.71% 0.14%
C HCO3 140 0.4% nd 11.10% 16.00% 8.10%
=
B as H3BO3 26 0.08% 0.2% 0.07% 0.12% 0.09%
Fe Fe++ 0.01 0.00003% 0.00004% 0.00008% 0.00010% 0.00016%
Mn Mn++ 0.002 0.00001% 0.00088% 0.00006% <5 ug/l <5 ug/l
Zn Zn++ 0.005-0.01 0.000022% 0.000035% 0.000010% 0.000003% 0.000010%
Cu Cu++ 0.003- 0.005 0.00001% 0.00002% <0.003% <0.003% <0.005%
N variable 0.35-0.5 0.00122% 0.00001%
P variable 0.05-0.07 0.00017% <0.00003% <0.00003% <0.00005%
0.0005 or
Mo MoO4= 0.012 0.00001% 0.00001% <0.00001% 0.00004% 0.00004%
salinity ppt 34.5 ppt 34.5 ppt 30 ppt 40 ppt 30 ppt 20 ppt
pH units 8.2 8.2 8.0 8.2 8.0
notes (1) (2) (3) (4) (5)
groundwater bores at sub-surface
Donald Victoria, separated irrigation
by 2000 m, measured after drainage,
residence in the basins. Lake Charm,
<1ug/l = <0.000003% Vic
1. see Goldberg in Hill 1963 or Sverdrup, et al 1947
2. unpublished 2005 data EPA (Western Australia) from a deep drain No. 29 at Morawa WA
3-5. unpublished 2005 data analysed by Analytical Services Tasmania, Hobart

3
Two of the most abundant elements Hydrogen (H) and Oxygen (O) are not listed in the table since
they are found as compounds. Much of the chemical dynamics of seawater is driven by bicarbonate
(HCO3-) through uptake and release of CO2 and O2 by plants, usually micro-algae and bacteria, which
changes the pH of the water and consequently the relationships among other elements and ions.
Although some of the oxygen is dissolved as a gas in the water (O2), much of it is tied up in ionic
compounds like carbonate, sulphate, borate, silicate, phosphate, molybdate and with hydrogen in
hydroxyl ions and, of course, water (H2O).

What we can see from Table 1 is that about 85% of seawater salt is composed of sodium (Na+) and
chloride (Cl-) and that the six most abundant ions constitute 99% of the elements. In the groundwater,
the data are not so clear. Na+ and Cl- are also the dominant ions but the major difference between
seawater and these inland waters is that seawater usually contains nearly twenty times more sulphate
than bicarbonate. Inland waters contain a much higher proportion of carbonate and Table 1 shows
that at the two Donald sites the water has more bicarbonate that sulphate. In every groundwater case,
the bicarbonate was from two to forty times greater than that in seawater.

When we compare major ions, which are also major plant nutrients, only potassium in the saline
groundwater is consistently lower than its concentration in seawater, sometimes less than one-tenth.
In this case the problem to plants may not be the direct availability of potassium for nutrition but the
toxic physical effects of sodium in the absence of sufficient potassium (Newman 2004). Comparing
the micro-nutrients in Table 1, copper and zinc are slightly depauperate in the groundwaters examined
with reference to seawater. Although seaweed actively concentrates these elements by a factor of
100,000 from the normal concentration in seawater, a reduced amount of copper and zinc may be
restricted in their activity by interference from competing minerals.

1.4. Why Seaweed

If the inland saline waters currently threatening traditional cropping in Australia can support marine
organisms, the next step to making these waters useful is to select a marine crop. As shown at the end
of section 1.2 above, some researchers have suggested the aquaculture of fish or shellfish or prawns.

Having discussed the potential for fish and prawn farming in inland saline water with scores of broad
acre farmers and pastoralists from the Murray Darling Basin and the West Australia wheat country, the
overwhelming majority considered fish and prawns to be too “high tech” and too financially risky for
them to trial. A major concern was the demanding timetable and labour requirements for feeding,
monitoring, harvesting, post-harvest handling and transport of fish or prawn harvests from farmers that
are fully employed with their primary and traditional activities and are uneconomically distant from
urban markets. Another concern was the cost of infrastructure for aeration, monitoring, feeding and
the post-harvest handling of seafood harvests. The establishment of a handling and marketing agency
for seafood and a supporting institutional infrastructure in inland Australia was considered to be well
beyond reason without existing production in the region.

It would therefore appear most sensible to find a species for cropping in inland saline waters that can
make use of the techniques and experience already developed for agriculture in these regions. The
crop should require the minimum amount of time and detailed attention for cultivation by farmers
already busy with other activities. It should require a minimum of investment in infrastructure and the
harvests should be easy to market without expensive processing and storage by the farmer.

With these criteria in mind, industrial seaweeds for hydrocolloids (see McHugh 2003, p.2) were
selected as a potentially suitable crop for saline basins with large and growing markets for specific
species, which were identified for consideration. The cultivation, harvest, drying and baling of
seaweed is parallel to the making of hay. The farming and harvesting timetables for seaweed would
be far more flexible than those required for the husbandry of fish or shellfish in salt water ponds and
the comparative infrastructure required is minimal. All farmers contacted with high water table

4
salinity problems were very interested in adding seaweed as a crop to maintain sustainability of
production on otherwise degraded lands.

1.5. Objectives of the research

The objective of the current research is to introduce and develop the agronomic tools and aquaculture
technologies for the cultivation of the marine gel- bearing seaweed Gracilaria chilensis. There is an
abundance of peer-reviewed scientific papers on Gracilaria physiology and nutrient use, its growth
under a range of environmental variables in seawater and descriptions of its response to farming in
seawater. It was thought that the literature contained sufficient background and supporting material to
enable field trials that would result in an agronomy of seaweed.

Our objectives have not been fully met, due in large measure to differences between seawater and
inland saline groundwater, which had never before been used to cultivate seaweeds.
Agronomic research was structured around field trials that might have been immediately transferable
to farmers but the results have been disappointing and suggest that the scientific literature consulted
was not a sufficient basis to scale up to the proposed development.

1.6. Markets for Gracilaria seaweed

Although most of the approximately 6.3 million wet tonnes of seaweed harvested around the world is
used for food, about a million wet tonnes (280,000 dry tonnes) is processed as a source of chemicals.
There are three major polysaccharides extracted from seaweeds: agar, alginate and carrageenan. These
compose a group of chemicals called hydrocolloids and their uses are based on their ability to thicken
aqueous solutions, to form gels and water-soluble films. They find applications in a wide variety of
industries such as food processing, pharmaceuticals, cosmetics and toiletries, tissue culture, paper,
welding and textile printing. Current production of agar-bearing seaweeds is about 180,000 tonnes
wet weight (McHugh 2003).

About 40% of the world’s supply of agar is extracted from Gracilaria chilensis. This species, native
to the temperate waters of the South Pacific Ocean is found both in the cooler waters of South
America and the southern coast of Australia. Chile has, over the past twenty years, built an industrial
capacity to produce and process seaweeds about half of which are cultivated at offshore production
sites and the rest harvested from natural production.

China, Indonesia, Viet Nam and Thailand together produce about 60,000 wet tonnes of Gracilaria, in
most cases a mixture of wild and cultivated harvests. About one-third of this production is used fresh
as a food vegetable or to locally process with sugar and colouring as sweets (idem.).

The value of dried and baled Gracilaria seaweed could range from US$250 to US$900 per tonne,
depending on the impurities, gel content and gel strength.
The following specifications were quoted (Neish 2003) as worth $700-900 per tonne:
Packing: Press Bale @80 - 100kg
Water: 30%
Impurities: 2-3%
Gel Content: 25%
Process: Sun Drying
Quantity: 500MT/Month Minimum.
Gel-Strength from 600-800-900-1000 gr/cm2

5
The seaweed industry in Australia is currently limited to the collection of wave torn Laminarai kelp
from the beaches of King Island for export to European alginate processors and Undaria, an accidental
introduction from Japan that is collected and processed into food supplements for export (Lee and
Momdjian 1997). There is also an industry that harvests single celled algae from large inland saline
lakes for processing as a food supplement (Browitzka in Smith and Barlow 1999).

1.6.1. Agar
Agar is a hydrophilic colloid extracted from certain red seaweeds (marine algae of the class
Rhodophyceae). It is insoluble in cold water but soluble in boiling water. A hot aqueous 1.5%
solution is clear, and when cooled below 40°C forms a firm, resilient gel which melts at temperatures
greater than 80°C. This difference between gelling and melting temperatures, known as hysteresis,
makes agar of great importance in the microbiological/biomedical, biopharmaceutical and food
industries (Marbio 2003).

One objective of the research is to develop protocols for the cultivation of seaweeds that will contain
the maximum value of gel (agar) for processing. The maximum economic return on the seaweed crop
is a function of the yield of seaweed per dollar of production cost, the yield of gel per kg of seaweed
and the quality of the gel.

The world market for agar in 2001 was about 7600 tonnes, with a value of $137 million. About 90%
of this was used by the food industry and most of the rest as bacteriological and tissue culture medium
(McHugh 2003).

The major commercial value of Gracilaria comes from the agar, which constitutes from 10-50% of the
dry weight of the seaweed. The agar content of seaweed changes with environmental and nutrient
conditions and varies from species to species. A collection of data from eight species of Gracilaria
grown in six countries clearly showed an inverse correlation between gel strength and agar yield from
Gracilaria (Freile-Pelegrin 2000). Producers will need to calculate the relative value of quantity and
quality of agar as part of their production strategy. Older Gracilaria usually has greater gel strength
than younger. Lower sulphur and higher nitrogen concentrations improve quality but stronger gel may
also result from nutrient starvation at the start of the rapid growth period. Some research has shown
that yields of agar are higher in the summer but of lower quality that that grown in colder water over
longer time (Lewis and Hanisak 1996). It has been suggested (Friedlander 2001) that high summer
growth builds up sulfated agar but the agar “matures” during cold weather, resulting in higher
strength.

Carbon dioxide and nutrient concentration, water temperature and intensity of solar radiation are all
factors in the growth of the seaweed which influence the agar content in a given plant. The presence of
epiphytes and undesirable associated algae can markedly affect the yield and even the toxicity of agar
from commercially collected seaweed. Life-cycle stage of the seaweed and the time of year when
harvested also have an impact on the yield and quality of the extracted agar. (Santilices and
Doty 1989). Seasonal and environmental variations, quality and yield of agar have been the subject of
numerous studies (see: Bird & Ryther 1990, Rebello et al 1997 and Villanueva et al. 1999). Other
studies suggest that agar yield and quality may be seasonal and that faster growth in high nutrient
conditions lowers the agar yield and quality.

The extraction of agar from Gracilaria is a traditional and well known process of boiling the seaweed
to separate the agar followed by a repeated series of freezing and thawing to remove the water.
Currently, to increase the gel strength, the seaweed is first cooked in an alkaline bath for an hour. The
gelling ability of agars from Gracilaria is considerably improved by pre-treatment with alkali before
extraction to convert alpha-L-galactose 6 sulphate into 3,6-anhydro-alpha-L-galactose (Murano 1995).
Full explanations of agar extraction are widely available from the literature (see: McHugh 2003, ch 3,
Singh 1992, and Coppen & Nambiar, 1991, ch 4). Gel quality is measured by gel strength, viscosity,
clarity, texture, melting and gelling temperatures and the proportion of particular polysaccharides in
the gel.

6
In two analyses of the Australian Gracilaria chilensis variety being cultivated in the current trials the
gel strength was 7% and 30% greater than an equivalent high gel strength commercial product.
(M Liao 2001 & 2003, pers. comm., 24 April & 20 Oct)

1.6.2. Commercialisation Strategy

Although the major focus of this research is on the agronomic development of Gracilaria seaweed, the
project is embedded in a market driven, whole-of-chain approach that includes interaction and
communication with the processing and marketing sectors. The species selected for production is the
one that provides over 40% of the raw material for the world agar market. The international market
for Gracilaria chilensis, is about 35,000 dry tonnes and growing at 1-2% per annum (Porse 1998).
Seaweed Trading has discussed the export of dried Gracilaria chilensis for agar production with
companies in Spain, India and China

In a RIRDC research paper on seaweed by Lee and Momdjian (1997) explained that there was a
fundamental lack of information about seaweed resources and market opportunities in Australia such
that the current industry is based primarily on kelp collected from the beaches of King Island. The
perception is that the collection and exploitation of Australian seaweeds would be incompatible with
fisheries, biodiversity and ecosystem management. No mention was made of seaweed cultivation but
recommendations were made to support the flow of information, research & development and
feasibility studies to identify market opportunities and baseline information on Australian seaweeds.

Linkages for domestic marketing are through Goodman Fielder (Germantown division), Scalzo Food
Industries and Uncle Ben’s of Australia, all of which have expressed interest in processing the raw
seaweed into industrial food thickeners (see section 1.7) that are currently all imported into Australia.
A consortium from Donald, Victoria is also prepared to establish an agar processing plant at Donald
when local farmers begin to produce seaweed. The Melbourne-based Cooperative Research Centre
(CRC) for Bioproducts has been working with industrial colloids, in partnership with industry since
1994. This CRC has agreed to work with Seaweed Trading to identify commercially useful seaweed
polymers and monitor the quality of seaweed produced. The CRC is also prepared to take the lead in
establishing an Australian based seaweed processing industry.

Discussions were held with abalone aquaculture farmers, who prefer raw Gracilaria chilensis as a feed
for their stock but are currently limited by state governments from harvesting sufficient quantities of
the seaweed. Abalone aquaculturists would welcome the availability of this seaweed for abalone feed
(Cooper 2003).

A chemical analysis of one of the weed species of seaweed (Enteromorpha intestinalis) found in the
basins, was shown to have a feed value, as measured by digestibility, crude protein and metabolisable
energy, similar to that of good quality pasture hays and some cereal grains (Downes 2004). This
suggests that that some seaweed species may be useful as a fodder for dairy cattle, especially during
drought when traditional pastures cannot be provided.

1.7. Potential benefits of seaweed cultivation

Seaweed sales could at the minimum, mitigate the costs of land rehabilitation and drainage basin
operations. It could help diversify production and increase profitability on farms with salinity
problems that have reduced potential to grow traditional crops. The initial focus is on the export of
raw seaweed to processors abroad and the annual international trade in the selected species is about
A$180 million (McHugh 2003). Seaweed production can make productive, profitable and sustainable
use of salt degraded land that is currently unproductive due to high saline water tables. The major
beneficiaries of seaweed cultivation will be farmers who are currently disadvantaged by the loss of
productivity due to salinisation.

7
Other beneficiaries of seaweed cultivation will include the irrigation companies that currently operate
saline water disposal basins. The annual cost of operation of the large basins is more than $3 million
and the groundwater interception schemes proposed, planned and under construction will incur annual
operating costs of perhaps an additional $2 million (see Table 8.3 in Allan 2000). Productive use of
these basins and their saline groundwater will help off-set their annual operating costs and they could
become profit centres while rehabilitating degraded land and maintaining their social responsibilities
to prevent the downstream exportation of salt.

The beneficiaries will not only be farmers and irrigation companies. Development of processing
plants near the source of raw materials can provide opportunities for value adding, for off-farm
employment and for the development of support industries. The CRC for Bioproducts estimated that
the international value of seaweed hydrocolloids is measured in hundreds of millions of dollars
(Thwaites 2001).

Australian industry has already recognized the benefit of domestic processing of seaweed into
colloids. Australia currently imports its entire requirement of agar and carrageenan for food
thickeners, textile and cosmetic manufacture. Several letters from industry attest to their:

“…significant interest in establishing processing plants designed to produce hydrocolloids of

value to local users and to service global markets (idem).”

Goodman Fielder (Ball 2000) would have a major interest in the establishment of a marine colloid
industry in Australia.

Uncle Ben’s of Australia wrote:

“Were the raw seaweed available in Australia, Uncle Ben’s would be interested in processing it
into the colloid. This would therefore act as an import replacement for use in several of our
products. This would result in cost savings to our production for both export and domestic
products and a consequent saving to Australia through import replacement of an expensive bio-
chemical, which could be manufactured here.” (Howard 2000)

The introduction of seaweed species could fill a currently empty agricultural niche on what is now
considered to be unrecoverable degraded land. More importantly, it could increase and diversify the
range of species cropped by farmers and increase the economic sustainability of farms through the use
of currently non-productive land. The net effect of seaweed cultivation can have positive
environmental effects since it will soak up excess nutrients from irrigation drainage water and help to
remove salt water from the environment.

Although there is some potential in seaweed production for biomass to produce green electricity from
methane gas, this would required a different branch of research. A more immediate benefit of
seaweed from saline drainage water evaporation basins is the removal of about 200 kg of salt with
each tonne of seaweed grown.

8
2. Literature Review
2.1. Current Gracilaria farming production methods
Gracilaria has many unique agronomic properties that make it a useful seaweed to farm. It can
tolerate a wide range of temperature, salinity and light intensity. Gracilaria can be propagated
vegetatively by sterile frond fragments. It responds quickly to pulses of nutrients and has been shown
to have the potential to increase in biomass by up to 24% per day (Israel et al. 1999). In nature it
grows in association with non-consolidated fine sediments in estuaries and slightly sub-tidal regions of
low water movement. This calm area of soft sediments where spores or tendrils cannot attach, limits
competition from other seaweeds. The occasional shell fragment or stone in the substrate can act as a
localised stable point for spore attachment and the development of spores on small substrates can be
used as a tool for stocking marine farms. Comprehensive reviews of Gracilaria farming, which
include diagrams and photos are found in Critchley (1993) and Oliveira et al. (2000).

Gracilaria cultivation in Asia

Commercial farming methods for Gracilaria in inter-tidal ponds were developed in Taiwan during the
mid 1960s to produce food grade agar. Fish and shellfish were cultivated in the same brackish water
ponds so that nutrients were abundant, seawater was regularly exchanged on the tides, fish grazed off
the epiphytes and the synergies of polyculture increased pond profitability. Ponds no larger than about
one hectare in area were easiest to manage. These ponds were stocked with about 5-6 tonnes of
Gracilaria frond fragments and were harvested of one-third to one-half of the standing stock every
4-6 weeks. Pond productivity ranged from 16-43 wet tonnes per hectare per year (Chiang 1981).

Brackish water pond cultivation later spread to Hainan Island and Guangxi Province, China
(Wu et al. 1993), where daily growth rates could be maintained at 2.7% per day at a culture density of
0.5-1kg per m2 (5-10,000 tonnes per ha). The Philippines also copied the Taiwanese methods
(Trono 1993), as did Viet Nam (Tran, 1992) where average annual production in 1988 was 18 tonnes
per ha, and Indonesia, where abandoned and less productive prawn farms were enhanced by seaweed
cultivation.

The Philippines was the first and still the largest producer of Eucheuma (now Kappaphycus) seaweeds
for Carrageenan marine colloid (gel) production. This seaweed is cultivated inter-tidally on hard
substrate reef tops and warmer, clearer water than would suit Gracilaria. Nevertheless, the long-line
method of seaweed cultivation, in which a rope is stretched between stakes in the substrate and
seaweed cuttings are inserted between the twists of the rope, is also used for Gracilaria. This method
and a variation, where the ropes are suspended from floating rafts in deeper water is used in several
countries (Trono, 1993) where it is felt that higher gel strength is realized from sea-grown Gracilaria
than from pond-grown seaweed.

The farming of Gracilaria has focused on providing habitat in near-shore waters in tropical and
temperate regions where this seaweed species is naturally found. There are many examples in the
developing world of village-scale coastal projects where plastic ropes are attached to posts driven into
the substrate of inter-tidal waters or suspended from floats and rafts in deeper near-shore waters to
cultivate seaweeds. Harvests are notoriously unreliable due to grazing by fish, destruction of
infrastructure by storms or sedimentation from nearby rivers or dynamic coastal processes (see
Kalkman et al. 1991). Nevertheless, suspended cultivation at sea in Chile and South Africa has been
reported to produce 34-45 tonnes (dry weight) of Gracilaria per ha per annum (Oliveira et al. 2000).

9
Gracilaria cultivation in Chile
Although Gracilaria cultivation and trials occur at some level in more than a dozen countries, the
major faming nation is Chile, where more than 100,000 tonnes were harvested annually from marine
sites since 1995, resulting in more than 2000 tonnes of agar (Table 4, idem). Chile is the world’s
largest producer of Gracilaria and it has since 1982 built an industrial capacity to produce and process
seaweeds. Farmed production of Gracilaria chilensis comprised about half the total harvest since the
Chilean Government limited the harvest from natural seaweed beds in 1986 (Buschmann et al. 2001).
Nearly all Chilean production is from the ocean, where the unpredictability of storms, competition
from other marine plants and herbivore pressure are major problems. Since farming of Gracilaria
chilensis occurs in the marine environment where environmental control is not possible, research
focuses on field cultivation, habitat manipulation and species interactions but not on agronomic factors
of seaweed production. During the decade of the 1990s most of the Gracilaria research came from
Chile, the major world producer of this seaweed and, together with Spain and Japan, produces about
60% of the world’s agar. (Porse, 1998).

One of the cultivation methods used in Chile is a long-line suspended from floating rafts (Santelices,
et al, 1993). Other farming methods have also been tried: (1) wire cages on the sea bed but competing
seaweeds grew better on the cages and storms washed the cages onto the shore (2) fragments of
Gracilaria pushed into the sandy substrate to “seed” new sub-tidal seaweed beds. Bundles of thalli of
about 400 g are set 20 cm apart in circles or rows spaced at 1-2 m (about 2 kg/m2). Seasonal
movement of sand and storms can quickly cover these beds, which then disappear. (3) Sand-filled
polyethylene tubes are used as anchors and seaweed fronds are attached with rubber bands. These are
scattered on the sea bed and periodically harvested. Despite annual production rates of more than 20
tonnes per ha (Oliveria et al, 2000), the sand filled tubes were discontinued, “as it facilitates the
development of pests and pollutes the farm and beaches” with plastic bags (Pizarro and Santelices,
1993).

The sea bed cultivation techniques and the problems of epiphytes, grazing organisms and
sedimentation were reviewed by Buschmann et al, (1995), including comparative benefits of inter-
tidal and sub-tidal stocking. He concluded that inter-tidal cultivation of Gracilaria would be a better
investment than sub-tidal despite the lower inter-tidal yield, because the initial investment is lower and
ultimately provides a better return.

Intensive Gracilaria cultivation

Growth of Gracilaria at high densities in tanks was attempted in the USA during the 1980s to develop
a renewable source of biomass for conversion to fuel and to “mop up” nutrient effluent from fish
farms (Hanisak, and Ryther, 1986). The economic and political realities of growing marine biomass at
high density for bio-conversion into methane and methanol to replace the consumption of fossil fuels
resulted in such farming being “an unrealised vision” (Hanisak, 1998). Nevertheless, high intensity
cultivation in a pulse fed nutrient regime was shown to be sustainable in a 15m3 raceway over four
years at an average of 175% increase in wet weight of Gracilaria per fortnight (Capo et al, 1999).
These experiments resulted in an improved understanding of the dynamics of seaweed growth under
prospective commercial conditions. At low water exchange and high density of seaweed, carbon
dioxide (CO2) and light became limiting as well as nutrients and subsequently all intensive cultivation
used aeration.

More recently the Israelis have begun to investigate intensive tank and pond cultivation (Friedlander
and Levy, 1995). They confirmed earlier studies that showed water motion to be the primary driver of
growth in intensive tank cultivation. Two processes seem to be involved, mass transfer of gasses and
nutrient minerals through the boundary layer and rate limiting enzymatic processes in the plant. The
main contributions of aeration to seaweed growth are the reduction of the boundary layer between the
water and the plant that increases the transfer of CO2 and nutrients, uniform mixing and access to

10
nutrients and the equal exposure of plants to light, especially at high densities (idem.). At densities of
4 kg wet weight m2, CO2 was limiting, especially at higher temperatures.

2.1.1. Constraints to Gracilaria Seaweed Farming

In Chile seasonal effects in the nearshore waters are the largest determinant of production, with growth
rates of 4.6 % per day in summer and only 1.8% per day in winter, suggesting a seasonal cycle
positively correlated with surface water temperature and light intensity. These seasonal effects include
blooms of epiphyte (see Glossary p.40) species on the seaweed and micro-algae, including red tides
and diatom blooms associated with Spring up-welling of nutrients and increased daylight. Chilean
seaweed researchers have modelled these environmental variations and planned their stocking density
and harvest frequency to maximise production (Pizarro and Santelices 1993).

The most common phenomena that constrain Gracilaria production can be characterised in terms of
light and temperature effects. Epiphytic weeds, which compete with Gracilaria for nutrients are more
abundant during the periods of high light intensity. Weeds like Ectocarpus fasciculatus and
Ceramium sp are the same as those associated with Gracilaria in Australia.

Epiphytes and competing algae are seen as a major obstacle to the economic production and
harvesting of seaweeds and many environmental or biological controls have been suggested, with
mixed effectiveness. In Taiwan seaweed is cultivated with herbivorous fish that graze on the
epiphytes. Others suggest snails, shading or avoiding the light-temperature optima to remove
epiphytes (idem. 1993), nutrient pulsing (Pickering et al. 1993 and Capo et al. 1999) and careful site
selection. The ability of Gracilaria to absorb nitrogen in the dark and concentrate large amounts for
rapid growth over one or two weeks is used as a mechanism to out-compete epiphytes and weed
species in cultivation (Oliveria et al. 2000). An overview of the epiphyte problem suggested that the
main adverse effects on the production of Gracilaria were the competitive removal of nutrients and
inorganic carbon from the water and the shading effect on the target plants (Fletcher 1995). His
review contains suggestions for control of epiphytes by biological, chemical and physical methods.

Epiphytic seaweeds and micro-algae are recognized as pest species under high nutrient conditions.
Fouling is considered the major problem for commercial Gracilaria culture due to increased
competition for nutrients, dissolved gasses and light. Harvests of Gracilaria with a high proportion of
attached weed species lowers the quality and value of the harvest. The ability of Gracilaria to store
nitrogen for at least a week of growth and to actively take up nitrogen in the dark can be used to
out-compete weed species that do not have this capability.

Control of green seaweeds and epiphytic diatoms on Gracilaria was attempted in Taiwan by reduction
of light, by short term drying of the seaweed, by increased water circulation, by low level aeration, by
pulse feeding of Gracilaria at night, by high ammonia (>0.5 m-mole/litre), by immersion in
hypochlorite solution and with grazing by small fish and prawns (Chen 1993). Some of these methods
were successful where Gracilaria was cultivated in closed tanks but where seaweed was cultivated in
the open sea, minimal control was possible and weeds continued to be a problem.

Gracilaria can survive buried in soft sediment for up to six months and produce seasonal crops when
they are uncovered. In polluted waters, high levels of one essential element can constrain the uptake
of others. Ca++, Na+, as well as Mg++ and K+ are mutually antagonistic (Lobband & Harrison 1994).
Copper, even at 0.2 mg/l (about 50 times its usual concentration) caused decrease in the growth of
Gracilaira tenuistipitata (Collen 2003). High calcium carbonate concentration from nearby reefs and
sediments can bind phosphate, making it unavailable to the seaweed (Hanisak 1987).

We have found competition by Enteromorpha intestinalis to be a problem that seriously constrained

seaweed cultivation in the long-established benthic ecosystem at Girgarre, Victoria. Control of
competing Enteromorpha by reduction of light, short term drying of the seaweed, increased water
circulation, low level aeration, pulse feeding of Gracilaria at night, especially with high ammonia
(>0.5 mmole/litre), immersion in 2-6% hypochlorite solution and grazing by small invertebrates was

11
not successful. We found that hypochlorite solution at these concentrations was toxic to the associated
Ceramium, Ectocarpus and Enteromorpha but also to the Gracilaria and pulse feeding of ammonium
assisted the Enteromorpha more than the Gracilaria.

Due to the dominance of Enteromorpha at Girgarre and the possibility of growing large volumes of
the seaweed in saline groundwater, tests were conducted to assess its potential as a stock feed.
Analysis of Enteromorpha showed that as dry matter it had an 88.2% digestibility, 13.1 MJ/kg dry
matter metabolisable energy and a crude protein level of 14.1% of dry matter (Downes 2004), almost
identical to wheat grain in these categories. This suggests its potential as a fodder, especially during
drought or to supplement poor pastures on saline soils.

There is no cultivation of seaweed in inland saline waters. Where it is farmed, seaweed is contained at
sea on lines or cages or in coastal waters or tidal ponds. Many of these inter-tidal ponds, especially in
Asia, were abandoned by prawn farmers after the prawn virus pandemic of the 1980s and 1990s forced
many investors to give up prawn farming or supplement it with fish and seaweed cultivation. Seaweed
cultivation in coastal waters is always subject to the vagaries of weather, marine pollution, fresh water
flooding and destruction by marine pests and weeds. The ability to cultivate seaweed in inland saline
waters would provide control over environment and nutrients now available to other farmed crops.

2.1.2. Spores
Most Gracilaria cultivation is propagated vegetatively from cuttings and some of the ‘varieties’ of
Gracilaria collected appear to be sterile populations that only produced vegetatively in nature.
Nevertheless, there are many advantages to seeding with spores, including selection of advantageous
agronomic traits, seeding new areas, seeding during seasons when natural production is low and
movement of new seed stock where transport costs are a factor. Chinese scientists have been
collecting and growing seaweed from spores since 1959. The collection and culture of Gracilaria
spores is described in an FAO training manual (Zhanjiang Fisheries College 1990) and it would appear
that spore collection is a regular part of the seaweed farming process there.

Gracilaria has a three phase life cycle that is typical of the red algae. It is comprised of an alternation
of generations as shown in Figure 1 on the next page. The diploid tetrasporophyte plant produces
male and female seaweed plants (haploid gametophytes), the spores of which develop as diploid
carposporophytes, living as parasites on the female gametophyte. This phase produces the non-sexual
diploid seaweed (tetrasporophytes), whose spores produce male and female plants. The
tetrasporophyte and gametophyte phases are morphologically (though not genetically) identical plants
and harvestable plants can be grown from either type of spore (Santelices and Doty 1989, and
Critchley 1993)

12
Figure 1. Life cycle of Gracilaria (Zjanjiang Fisheries College 1990)

Spores will develop and germinate in a relatively wide range of temperature and salinity,
corresponding to most of the range of the mature species during spring, summer and autumn (Yokoya
and Oliveira 1993) though very high temperatures and irradiance is as deadly to spores as very cold
temperatures (Orduña-Rojas and Robledo 1999). Despite the value of using mass seeding of
‘sporelings’, most seaweed farming, including the major producer, Chile, still cultivate seaweed from
vegetative cuttings (Santelices et al. 2004). A manual for spore culture is available (Glenn et
al. 1996).

Fertile Gracilaria plants of some species can be harvested to produce a source of spores, which are
settled onto gravel or shells before being broadcast onto the farm-site. Where the seaweed is grown on
lines or nets, lines placed intermittently enable spore settlement. The system is relatively simple.
Mature plants showing sporangia are collected, washed and allowed to dry on bamboo curtains for
2-4 hours until irregular corrugations appear on the plant surface. As soon as they are returned to
seawater they release spores which settle onto the substrate within a day and become dish-shaped
thalli (stems) in five days (Santilices 1996).

2.2. Seaweed Physiology and Nutrition

The marine habitat of seaweed is a complex and dynamic set of physical and chemical variables and
the response to changes in these variables is not always clear. Light, pH, temperature and salinity of
the water will affect nutrient availability and nutriton of the plants.

All plants need sixteen elements to grow and all of these elements are found in seawater (some plants
need more than 16 but those are in seawater too, though some in very low, even nano-molar
concentrations. Some of the “micro-nutrients” are concentrated by the plants to more than
10,000 times that of seawater and uptake of nutrients in seaweeds is primarily by active transport
(Lobban & Harrison 1994). So even a tiny trace of micro-nutrients in seawater may be sufficient to

13
support marine plant growth. On the other hand, concentrations of ions are almost never likely to be
toxic except in particularly polluted nearshore waters.

Of these 16 essential elements, Carbon and Oxygen enter as gasses (CO2 and O2) through the surface
of the water or from respiration and photosynthesis in the water or as (bi)carbonate ions dissolved in
the water. Carbon dioxide can occasionally be limiting to seaweed growth, not just as a nutrient but in
it’s role as a regulator of pH. Experiments with seaweed at high density in tank cultivation have
emphasized the need for supplemental CO2 and pH adjustment in the absence of water exchange. At
high experimental densities light becomes limiting as the seaweed becomes self-shading such that all
intensive tank cultivation of seaweed includes aeration and even CO2 injection. Note that Hydrogen is
also essential but is not found in an elemental form in seawater; it is always in compounds (H2O,
HCO3-, NH4+, etc).

Despite wide phylogenetic differences, seaweed has basically the same metabolic and nutritional
needs as terrestrial plants. While there have been some specialisations in seaweed physiology, due to
its evolution in a seawater medium, there is no reason that, apart from the method of seeding, that a
seaweed crop cannot be cultivated with methods equivalent or parallel to current methods for cropping
of grains and pasture.

The usual nutrient limitations to plant growth in the sea are nitrogen and phosphorous, both of which
occur at low concentration in normal oceanic water. Only occsionally or seasonally are they available
in sufficient quantities to produce a “bloom” of seaweed. Only in coastal waters does up-welling or,
more likely, run-off of nutrients from the land provide sufficient nitrogen and phosphorous to sustain
the growth of large standing crops of seaweed.

Nitrogen is most often the first driver of productivity since the other elements are more often in
surplus to plant needs in the absence of nitrogen (Friedlander 2001). The literature suggests that
ammonium (NH4+) is preferentially absorbed over nitrate (NO3=) as the most useful source of nitrogen
to seaweeds (Nelson et al. 1980) and the presence of ammonium may even supress nitrate up-take
(Smit, 2002). Pulse feeding of ammonium to Gracilaria on a seven day cycle has been shown in
several studies to provide a competitive advantage over epiphytes (Friedlander and Levy 1995),
especially if ammonium was pulsed at relatively high concentrations up to 150 μM (=2.7 mg/l)
(Pickering et al 1993, see also Buschmann et al. 1996).

After nitrogen, the second limiting nutrient is often phosphorous (P), which also available in various
forms but primarily as the phosphate ion HPO4= (97% at pH 8.2 and 20oC), though algae acquire
phosphorous principally as orthophosphate ions (PO43-), which account for less than 1% of free ions
(Lobban & Harison 1994). Phosphate dynamics were adequately reviewed about twenty years ago
(Cembella et al. 1982). At the pH of marine waters, about 8.2, PO4= is at its maximum availability to
seaweed. At high concentrations of either nitrate (>100 M) or phosphate (>1 M) these ions are
reportedly antagonistic and each may inhibit uptake of the other nutrient (Lobban & Harrison, 1994).
The median ratio of N : P in seaweeds is 30 : 1, about half the amount of P required by phytoplankton
algae (Atkinson & Smith 1983, in Lobban & Harrison 1994).

The other major ions required for plant growth are Calcium (Ca++), Magnesium (Mg++), Potassium
(K+), Sulphur (as the sulphate anion SO4=), Sodium (Na+) and Chlorine (Cl as the Chloride ion Cl-).
All these ions are abundant in seawater and the six ions together constitute more than 99% of the
composition of sea salt.

Amongst the remaining ions making up less than one-half of one percent of sea salts are other
essential plant nutrients: boron (B as borate H3BO3=), iron (Fe++), Manganese (Mn++), Copper (Cu++),
Zinc (Zn++) and Molybdenum (Mo as molybdate MoO4=). Borate and molybdate, like phosphorus, are
available at pH higher than 7.

14
Some species of seaweed also require Silicon, Cobalt, Nickel, Vanadium, Iodine or Selenium because
of special circumstances like enzyme co-factors or the silica shells of diatoms, but these elements too
are found in seawater and can be concentrated as necessary by active transport in the plants to
thousands of times their usual occurrence in seawater.

Iron, while one of the most abundant elements in the earth’s crust is one of the least soluble in marine
waters and at the pH of seawater (about 8.2) iron is virtually immobilised. Iron can be available to
seaweed when chaelated in an electron-donor organic molecule (ligand). Bacteria on sea grasses and
seaweeds secrete siderophores (another organic molecule) that chaelate and solublise iron. Although
not part of the chlorophyll molecule, iron is required as a co-factor in the synthesis of chlorophyll so
that nitrogen metabolism is seriously affected by the lack of iron (Jingwen et al, 2000). In an early
work, Palamas-Mordoyntseva (1968) found that 1 μg-Fe/g was the critical concentration of iron for
plants and showed a positive correlation between iron concentration and growth rate to 14 μg/g,
though 8-280 μg/g was toxic.

Some marine algae require organic nutrients, especially vitamins, like B12, thiamine and biotin, as co-
factors in enzyme reactions (Lobban & Harrison, 1994). Transfer through organic molecules may be
the major pathway for the inclusion of iron, copper, zinc and manganese into the plant cell; these ions
may otherwise be oxidised into unavailable minerals in the seawater environment. Many of these
transformations and organic nutrients and possibly growth regulators are produced by bacteria or
perhaps fungi and the full symbiotic relationships between flora and fauna are still at least partly a
mystery.

Seawater, especially close to shore, is not a stable solution; ions interfere with each other depending
on ion concentration, the physical environment or seasonal temperature, salinity and pH changes of the
water. Too much iron (Fe++) can prevent the plant’s use of manganese (Mn++). On the other hand,
sufficient manganese (and / or calcium) is necessary to overcome the toxic effect of high
concentrations of copper (ibid). Under marine conditions of high sulphate (SO4=), this ion can
interfere with the plant’s absorption of molybdate (MoO4=); required for nitrate reduction to
ammonium (Collier 1985). This may mean that under conditions of high ammonium concentrations,
molybdenum is not necessary for seaweed growth. At high concentrations of calcium, borate
(H3BO3=), necessary for plant growth, can be co-precipitated with CaCO3 and so become unavailable.
Calcium may also interfere with potassium absorption at high concentrations (Reuter et al. 1997).

The lack of water current can prevent the adequate exchange of gasses, minerals and waste products
past a thin boundary layer of water around the plant. The pH of seawater is usually about 8.2 and very
well buffered against rapid change. The major driver of pH change is carbon dioxide (CO2), which in
a biological system is a product of respiration and lowers the pH of the water. Alternatively, during
photosynthesis the CO2 is taken out of the water and the pH increases. This dynamic affects nutrient
availability, which decreases at high pH for zinc, copper, iron and manganese but increases at high pH
for calcium, molybdenum and phosphate.

2.2.1. Physiological limits in inland saline groundwaters

Inland saline waters in Australia are similar to seawater, in that they are dominated by the sodium and
chloride ions, but inland saline waters are not fully congruent with seawater in their ionic
concentrations. This is due in large part to the fossil nature of much groundwater, which is held
underground for millennia. Generally in underground fossil water the pH becomes more acidic due to
the influx of CO2 in rainwater, which helps to dissolve the rock formation in which the water lies,
adding those available surrounding minerals to the water and increasing its salt concentration.

When the water reaches the surface and the pH increases as it loses carbon dioxide to the air and
increases in dissolved oxygen, some of the ions become oxidised and precipitate out of the water as
iron, magnesium or calcium salts (eg, Fe(OH)3, MgSO4 or CaCO3) and other more complex minerals.
Some ions are adsorbed onto minerals in the soil, where they may be tied up and become unavailable
to plants- at least for a time.

15
The loss of dissolved potassium in saline ground waters in the Murray Darling basin results in a
serious deficiency of that element at concentrations often only 15% of that in seawater at the
equivalent salinity. Potassium is an important electrolyte used to replace sodium in cell metabolism
has been found to be a critical constraint to terrestrial plants in saline soils (Newman 2004). Very low
potassium concentrations in inland saline waters have been shown to contribute to poor seaweed
growth in the current study. Potassium has a more specific role in plant metabolism as an enzyme
activator (Lobban & Harrison 1994).

Although seaweeds can be sustainably grown in test tubes and petri dishes on a prepared medium,
there are problem in scaling up cultivation to commercial size in inland saline groundwater due to the
dynamic interaction or deficiency of minerals and organics that might not replicate conditions in the
sea.

Trace metals and organic compounds have been posited to be limiting to satisfactory marine plant
growth (Yoneshigue-brage and Baeta-neves 1981). Some of these trace elements in inland saline
groundwaters may be insufficient to sustain seaweed growth. Equally as important, fossil waters in
newly prepared or plastic-lined evaporation basins may not have the associated bacteria and mature
organic molecules necessasry for the production of critical vitamins or chaelators necessary for
seaweed growth.

The role of weed epiphytes and associated algal species have not been considered in the literature as
potentially symbiotic. The epiphyte seaweeds Ceramium, Ectocarpus Pachydictyon, and
Polysiphonia found in association with Australian Gracilaria may have nutrient exchange value, a role
in light attenuation or may simply be competitors. Ceramium has high vitamin B12 content
(Provasoli 1960) and perhaps makes this vitamin available as a growth factor to the Gracilaria. On
the other hand there may be some agronomic value to the commensal snails (prosobranch, gastropods)
and crustaceans (isopods and amphipods) that are associated with the Gracilaria in the Tasmanian and
Victorian estuaries where it is naturally found.

2.2.2. Growth rates of Gracilaria

Daily growth rates published in the literature are extremely variable, ranging from less than 1% per
day (Rebello, et al, 1996.) to more than 24% per day (Chirapart &. Ohno, 1993). A collection of
growth rate studies in the scientific literature (see Table 2 below) showed an average growth rate of
6.7% per day. A number of researchers have assessed Gracilaria growth rates against physical and
chemical variables and some of these are also shown in Table 2 below. These studies suggest a
number of variables controlling growth rates, including current flow and nutrient inhibition. Some of
the studies suggest that under the same conditions there are significant differences in growth between
Gracilaria species. Light, temperature, salinity and dissolved inorganic nitrogen species, phosphate,
potassium, iron and micronutrients all need to be considered for their contribution to seaweed growth
for any species in formulating a protocol for seaweed cultivation.

Growth rates can be correlated with environmental variables as well as nutrient concentration. Lee
(1998) demonstrated that maximum growth in summer (6%) was twice that in winter (3%) at the same
temperature range, suggesting day length as a factor. Gracilaria was unable to recover from
temperatures below 14oC or above 32oC. Xiu (2001) showed that nitrogen uptake was maximum at
the light range, 320-1600 μM photon/m2/sec at pH 8 and temperatures between 21-26oC. Data from
Friedlander et al (1990) suggested that water temperature affected the growth rate twice as much as
light irradiance, though it was not a simple linear relationship and mid-day irradiance and epiphyte
colonisation were major contributions to the model. The compensation pH (beyond which no net
photosynthesis occurs) has been reported to be 9.2 (Mercado, 2000), suggesting a CO2 limitation.

Lapointe & Duke, (1984) concluded that under light limitation, Gracilaria tikvahiae seaweed
accumulated nitrogen and was light limited in its growth rate at high nitrogen levels. At optimal light
intensity, growth was at its maximum until the carbon to nitrogen (C:N) ratio reached about 13:1,

16
when growth rate began to fall. That is, light (and temperature) limitations tend to uncouple the
growth rate from nitrogen uptake. Nitrogen accumulation in excess of immediate growth needs is
stored as phycobiliproteins (Ryther, et al, 1981). After ten days of nitrogen enrichment, the total
stored nitrogen became 5.78% of dry weight in G. tenuistipitata, which was then able to sustain rapid
growth for more than twenty days (Liu & Dong, 2001).

The wide variation in results between and among the published experiments has not been accounted
for and there has been no full and final analysis of the role of the multiple variables or the defining
combinations that enable sustainable high daily growth rates. For each species there is likely to be a
set of environmental and chemical variables that sustain maximum growth. The available studies
suggest that increasing light and temperature in the presence of high nitrogen loading may be the
starting point. Hence the rapid growth rate of Gracilaria in the Spring in southern Australia may be a
function of nitrogen (and other nutrients) accumulation during the low light and temperature period of
winter, with growth stimulated by increased light intensity and duration after about October to early
November.

17
 Table 2. Comparative growth rates of Gracilaria seaweed

Gracilaria Max. Location of Optimum Optimum Most Optimum reference

species growth experiment temp. light important Nitrogen
rate per growth
day variable
edulis 5.10% Indoor and 25-30oC 150-300uM temperature Gerung, et al
salicornia 4.90% outdoor tanks, photon m-2 1999,
Usa Japan, s-1
sp A 3.25% seaweed from
sp B 4.90% Indonesia
chilensis 3.17% 170 l plastic 20C 25uM photon nutrient 72- 143 Pickering et
bags indoors m-2 s-1 pulse uM NH4 al, 1993
New Zealand frequency
2x/ 14 days
edulis 7.55% Shallow reef 20C ammonium 1.02- Nelson SG et
flat Guam and light 3.95% dw al, nd
arcuata 3.50% 20C 2 species 2.2-4.3%
dw
foliifera 12% Raceway nd DeBoer, et al
Woods Hole 1978 in
Massachusetts Nelson
bursapastoris 12.24% Tanks Hawaii nd Hoyle, 1978
coronopifolia 6.41% nd in Nelson

bursapastoris 2.00% Field sites nd Hunt, et al,

coronopifolia 8.30% Hawaii 1979 in
Nelson
tenuistipitata 7% nd
chilensis 4.60% Suspended summer 1580 uE summer Santelices, et
ropes at sea 10-13C m-2 s-1 light and al,1993
Chile 12-14 hrs temperature

tenuistipitata 18.3% 40l outdoor 20oC >300 uM Salinity 0.5mM Israel et al,
var. liui (128% tanks Israel , photon m-2 39ppt & NH4+ 1999
per seaweed from s-1 Ammonium
week) Philippines
gracilis and 11% polypro. ropes open Wakiba, et al
(Gracilariopsis Saldanha Bay, water 2001
sp) South Africa

parvispora 4.60% net cages in Ammonium 2-5% N in Glenn, et al,

lagoon tissues 2002
Molokai,
Hawaii
tenuistipitata 3.30% outdoor ponds 20-30 C 250 uM salinity 21 4.36uM -N Wu, et al,
Guangxi , photon m-2 ppt, light 1993
China s-1
verrucosa 2.34% ~10-15 ha 20-25 C summer 10-22 ppt, manure Tran, 1993
(calc manured tidal 2-5 tn/
from ponds ha+ pulse
yields) @ 3-5 kg
(N:P, 2:1)
per ton
stock
G. species 10% Southern 30 C nutrient. 3.2% dry Chuan-
Taiwan varies wt Chuan Tsai,
winter/ 2005
summer
G. species 7% suspended ammonium Troell, et al ,
ropes at sea 1999
salmon farm
Sweden

18
Gracilaria Max. Location of Optimum Optimum Most Optimum reference
species growth experiment temp. light important Nitrogen
rate per growth
day variable
tikvahiae 5.76% outdoors 24-30 C 100 uE CO2 (pH) N >2% dry Hanisak,
(calc. Florida flow m-2sec-1 by current weight 1987
from through lined or aeration
yield) pond
balinae 1.12% Covered 200l 24-33 C 600-1600 uE light 0.5 mM Hurtado-
tanks outdoors m-2 sec-1 intensity urea Ponce et
Philippines al,1997
cornea 12.77% 1 litre flasks 27 C 100uM N:P ratio 824uM- N Navarro-
over 8 indoors photon m-2 10:1 any Angulo, et al,
weeks Yucatan, s-1 source, 1999
Mexico weekly

2.2.3. Nutrient media for algae

Most research on seaweed cultivation, even in small volumes of water have used natural seawater,
enriched with a few of the major nutrients, usually nitrogen and phosphate. Issues of varying water
quality, the need to control nutrient and trace element concentrations and the unknown seasonal
quality of many near shore waters have led researchers to use artificial seawater to explicitly define the
composition of their media for reproducibility of their experiments.

Artificial seawater has evolved over the last half century and improved its ability to support greater
numbers of previously hard-to-cultivate species. (Berges, et al 2001). Earlier attempts to improve
artificial seawater simply added an infusion of good quality soil to provide unknown organic factors
necessary for plant growth. As knowledge increased, this was replaced with vitamin stock containing
B12, Thiamine and Biotin, including the chaelator EDTA to assure the availability of iron and trace
metals. Since 1980, changed forms of nutrients have improved the ability of plants to use them from
the medium and new trace elements: nickel, selenium and molybdenum are now regularly added to
seawater media (idem). Due to interferences, anhydrous (without water in the molecule) and hydrated
(with water) salts are mixed as separate solutions, as is the chaelated iron, chaelated trace metals,
vitamins and major nutrients, before being prepared as an algal medium (idem).

These newest nutrient media can stand as a base line model for cultivation of seaweed as their
preparation and storage show the subtlety and delicate balance of mineral and organic species that are
contained in seawater. It is against this model that agronomists must try to modify and prepare the
solution of ions in sources of inland saline groundwater to support seaweed cultivation and to enrich
the supporting medium to enable commercial rates of growth and cultivation of the selected seaweed
species.

19
3. Methodology
The research objective of this project is to determine the set of agronomic and water quality criteria
that will provide the maximum economic yield of seaweed to meet the needs of export and domestic
market targets for raw and processed seaweed products. The strategy is part of a market-driven,
whole-of-chain approach that will include improvement of harvest quality for agar production, post-
harvest handling to market and domestic value addition using improved agar extraction technologies.

The target species for seaweed in cultivation is Gracilaria chilensis, a temperate water agarophyte
found in Australia and currently cultivated in the sea off Chile and having a world market value in
2001of US$137 million. (see section 3.1).

Pilot trials in small on-farm evaporation basins were used to develop agronomic methods for the
seaweeds and to establish protocols for crop and water management. These basins were used to
examine seaweed growth rates under ambient conditions equivalent to that expected under actual
farming circumstances. (see section 3.2)

Controlled experiments in smaller tanks and pools at the trial sites were used to examine more
precisely the growth factors necessary for seaweed cultivation that maximise plant growth. The
information collected in small pools was used to inform the field trials in evaporation basins. (see
section 3.3)

Wild stocks of Gracilaria were acclimated in Girgarre basin groundwater for two weeks before use in
nutrient or growth studies. After manipulation of water quality to meet base line criteria, seaweed
propagules were seeded to assess the success of the water treatments. Many of these tests were
non-quantitative bioassays to determine the ability of the saline ground waters to support live seaweed
(see section 3.4).

Since the natural medium of the seaweed is oceanic water, the ionic composition of the inland saline
waters was always compared to sea water for major ions and trace elements that might effect the
growth of seaweed and the production of colloids. Saline groundwater in the Murray Darling Basin
varies widely in ionic composition, even within a few hundred meters, due to multi-layered water
tables and paleo-channels that cross sedimentary layers; the minerals being dissolved into and
adsorbed or precipitated out of the entrained waters (see section 3.5).

A suite of chemical factors was regularly measured and correlated with growth of seaweed over a
series of fortnightly periods in 2.25 m3 pools. In evaporation basin field trials no controls were used
since only one of two basins at the field sites was ever ready at any one time. Growth was measured
fortnightly but calculted on a daily basis for comparison with growth rates in the literature (see
section 3.6).

During the course of the study several agricultural techniques were developed to support seaweed
cultivation in saline water evaporation basins. These methods focus on the needs of farmers to have
the appropriate tools to make seaweed cropping convenient. There is no doubt that farmers
themselves will develop further tools and technologies to meet particular needs of each farm.
(see section 3.7).

3.1. Seaweed species selection

A market-driven selection was made for Gracillaria chilensis from which the hydrocolloid agar is
commercially extracted. This seaweed is native to southern Australian coastal waters from at least
Geraldton, WA to Newcastle, NSW and including eastern Tasmania. Gracilaria chilensis is also
found in the Eastern Pacific. Chile harvests about 170,000 wet weight tonnes of this species for
processing into agar.

20
This species was discovered in several locations in southern Australia during the search for raw
materials for industrial gels by the CRC for Biopolymers (West and Kraft, 2002). Based on this
research (Byrne, et al, 2002) five geographical stocks (or eco-types) were investigated in this study for
cultivation in inland saline groundwater evaporation basins. Gracilaria shows morphological
variation with habitat, changing colour, length, thickness and form within the range of its taxonomic
status. Several varieties or eco-types of Gracilaria were collected from the waters of southern
Australia and were field tested at the Girgarre research facility before being used for agronomic
studies: type "F" collected as beach wrack from Flinders, Victoria, variety "H" from Hobson’s Bay,
Victoria, "J" from Westernport near Hastings Victoria, "M" from Marion Bay, Tasmania and “N”
from NorthWest Bay, Tasmania.

For on-farm field trials in this project type M was selected as a hardy and fast growing variety for
cultivation trials. This variety also exhibited the best growth performance in vitro (John West, 2003)
and in situ it has performed well in bioassays for viability in a wide range of conditions in northern
Victoria. Salinity limits of this variety in nature were very wide, from <20 to >80 parts per thousand
(ppt), an important tolerance to the wide range of salinity likely to be encountered in evaporation
basins.

Propagules of Gracilaria were selected for trials and seaweed was grown vegetatively from cuttings
rather than from spores to reduce the expense, time and technological input required to develop plants
from spores. Growth from cuttings also permits the greatest increase in biomass over the shortest time
and assures that the same genotypes are being tested in each trial.

Figure 2 shows the types J and M Gracilaria. Note that the thallus of type J is much longer that those
of type M, which are closely bunched and more turf-like. The bottom of type J is attached to a mollusc
shell where the spore will have landed and took hold. The darker parts at the bottom of type M have
been buried in soft mud and apparently grew vegetatively from a fragment of seaweed.

Figure 2. Gracilaria chilensis. Two varieties or eco-types from southern Australia, type “J” on
the left and type “M” on the right

3.1.1. Why Gracilaria?

This genus of seaweed grows in a wide range of latitudes, salinities and water conditions; hence it is a
good candidate for cultivation in inland saline ground-waters. It is of considerable economic
importance as a source of the hydrocolloid gel, agar, with a minor direct use as food; it has potential
use as in “mopping up” nutrients from other marine farming activities (Buschmann, et al, 1996) and as
a source of “biomass” for the production of methane gas (Hanisak, 1987). It is also a preferred feed
for abalone in aquaculture (Cooper, 2003). Understandably, there is a rich literature about Gracilaria
that serves as a base for agronomic studies.

21
Gracilaria chilensis is cultivated in the nearshore waters off Chile for industrial agar production.
Other species of Gracilaria are harvested or cultivated in coastal waters, primarily in Asia, the total
annual world harvest ranging around 240,000 tonnes wet weight, about 60,000 tonnes of which is
cultivated, either in the sea at intertidal and offshore sites in South America or intertidal ponds in Asia
(McHugh, 2003). Although other species of seaweed can be used for high quality agar extraction,
none of them grow as quickly or are as tolerant to as wide a range of environmental variables as is
Gracilaria (idem).

Several varieties and eco-types of the Gracilaria were used in this study and one type was selected for
its wide salinity tolerance, high gel strength and good growth compared to other available eco-types of
this species. Gracilaria is able to remain dormant and healthy in the dark and under the substrate for
months at a time and at Tasmanian winter temperatures. Due to the rate of growth shown in the
literature and the totipotent ability for growth from all parts of the plant, Gracilaria, in the appropriate
medium can be grown vegetatively from cuttings that are seeded into a basin, the increase regularly
harvested and cuttings from the harvest re-introduced as propagules.

G. chilensis did not have the highest agar yields among the few native Gracilaria species studied by
Byrne et al, (2002), but it was selected as the species of choice for agronomic research for cultivation
in inland saline ground-waters since it had a very high quality agar, had the best growth among the
species studied by West (2004) and it is such an important economic species with a large background
literature. G. cliftoni from the Byrne study had a very high agar yield but the gel strength was very
poor (Ming-Long Liao, 2003)

3.1.2. Taxonomy and local eco-types

Identification of taxa amongst gracilarioid algae has been notoriously difficult. These economically
important agarophytes of worldwide distribution have nearly 300 species assigned to the genus
Gracilaria of which about 110 species are currently recognized and new species are continuously
being identified (Rueness, 2005). The occurrence of similar morphologies in separate species and
genera, and of apparently sterile specimens collected in the field, makes assessment of systematic
classification extremely difficult or impossible. In many red algae, unattached forms are often found
in the vegetative state, and cultures may be necessary to induce formation of reproductive structures
and completion of the life history. Taxonomic problems are gradually being resolved by combining
studies of reproductive morphology with comparative analyses of selected DNA sequences (idem).
West (2003) was unable to find or induce spores on the type M Gracilaria and none have been
encountered during this research, either in culture or in nature.

The taxonomy of Gracilaria is poorly understood due in part to the difficulty in identifying
differences in members of the group, even at the microscopic level. The problem is sufficiently
serious that the agar processing industry prefers to label stocks according to their site of collection
rather than their scientific name. Interpretation of the genus has been described as chaotic. Adding to
the confusion, polymorphism and variation within a single species can be great enough to incorporate
descriptions of several species (Critchley, 1993).

About a dozen species have been described from Australian waters at different times, using both
morphology of the thallus (the stem of the seaweed) and the reproductive morphology (see Byrne et
al. 2002 for a review and references). During a taxonomic study of a new Australian Gracilaria
species using molecular tools (idem), G. chilensis from several locations and habitats was described
and compared for growth rates and polysaccharide (gel) content along with other species from
southern Australia. The morphology of Gracilaria is very variable. In an early study of Australian
species based on morphology, nine species, 24 forms, one variety and five uncertain taxa were listed
(idem). Within the chilensis species we have found that the morphology varies with habitat, shallower
waters producing a turf like form while deeper waters permit longer thallus growth (see figure 2). The
colour varies from golden yellow through dark green to black with nutrient concentration and
submergence in soft substrate.

22
3.2. Description of sites and basins
Disposal evaporation basins are used to collect saline groundwater and thereby maintain water tables
below the critical level for agriculture and to limit the export of salt outside irrigation areas. There are
currently more than 150 of these salt water interception and evaporation basins throughout the Murray
Basin from Griffith, NSW to Waikerie, SA, eleven of which are large-scale saline groundwater
interception schemes and their associated disposal basins (Hostetler & Radke, 1994).

Several field trials were established in evaporation basins in Victoria at Girgarre (2002), Jeparit (2003-
04, Donald and Lake Charm (2004-05). These sites exhibit a range of saline groundwater quality and
environmental characteristics that reflect both their catchment history and location in the landscape

3.2.1. Girgarre
Goulburn-Murray Water has operated a salt water interception and evaporation basin at Girgarre in
northern Victoria since 1987 to lower saline groundwater that threatens nearby farmland. A series of
pumps removes 375-400 megalitres of saline groundwater annually and delivers it to the 30 ha basin.
The basin is composed of three bays designed for a 0.5 m water level (see figure 3). Water from the
pumps is discharged into the 13 hectare Basin A at about 12,000 mg/l salinity (= 12 ppt). Water
evaporates as it flows for about a km into Basin B, where the salinity has increased to 30-50,000 mg/l.
Finally the saline water runs into the terminal Basin C where some salts precipitate and salinity ranges
about 110-140,000 mg/l. Water is pumped into the basin during summer since net rainfall in winter is
usually greater than evaporation and salinities decrease. The aquifer is in the top 12 m, where the
water layer is up to 10 m thick, with the salinity usually 4500 to 9000 mg/l (4.5-9 ppt).

Goulburn-Murray Water kindly made available eight trial basins with areas of 0.3 and 0.1 ha. These
basins were initially used by GM-Water to test pond linings for the facility. These trials ponds were
fortuitously situated to enable saline water for the seaweed experiments to be mixed from the bays to
provide a wide range of experimental salinities. Figure 3 is an aerial photograph of the Girgarre
Evaporation Basin facility showing the three bays and the trial ponds.

Bay B

Bay C
Trial
Bay A

Photo: Warwick

Figure 3. Aerial view of the Goulburn Murray Water Girgarre Evaporation Basin showing the
path of the evaporating water and the location of the trial ponds

The Girgarre Evaporation Basin is more than 15 years old and has developed a relatively stable but
seasonal saline ecosystem, characterised by various species of birds more often found around coastal
wetlands and a benthic flora dominated by the seagrass Ruppia sp. There are seasonal blooms of the
euryhaline (see Glossary page 40) green seaweed, Enteromorpha intestinalis. The hypersaline Bay C
has seasonal blooms of the red micro-algae, Dunalliela salina followed by a bloom of the zoo-
plankton Artemia salina.

23
The saline groundwater in the trial ponds comes from one or several pumps in the area that deliver
water to the shallow Bay A. During its passage toward the Trial Ponds the water is modified by the
wetland ecosystem consisting of sea grass and algae flora, small fish, insects, crustaceans and
especially bird fauna. The water leaves the bore at a pH of about 6.9 (State Water Lab, Rural Water
Corporation , Victoria , 2002) and by the time it arrives at the Trial Ponds the pH has increased to
more than 8 due to the diffusion of carbon dioxide into the air and its use by plants in Bay A

3.2.2. Jeparit
In 2003 the Victorian Department of Primary Industries, Horsham, initiated a seaweed trial alongside a
pre-existing groundwater pumping trial with the objective of making productive use of the waste
saline water (Hedt, 2005). The pumping trial was part of an attempt to save the adjacent cropping land
by lowering the water table. The project was established to remove groundwater using a windmill
driven pump, which delivered up to one litre per second, depending on wind strength, into a 150m
long drain that emptied into a series of two 33.75 m2 settling ponds. From there the water overflowed
into the 0.81 hectare shallow evaporation disposal basin.

Groundwaters were taken from the Parilla Sands aquifer, 6-18 m below the surface and this water was
used for seaweed trials before being emptied into the disposal basin. This is an entirely different
species of groundwater from the near surface drainage of Girgarre water with its irrigation salinity.
The two different water types made an interesting contrast for seaweed trials.

The Jeparit site is adjacent to a swampy Mellaleuca forest situated between ancient sand lunettes. The
forest had become salinised by a rising water table and all the trees are dead. The flora was dominated
by a variety of salt bushes and succulent salt-tolerant ground covers. A shallow water table was
perched above the Parilla Sands aquifer within about one meter of the surface. This water table had a
very different chemical signature to the Parilla sands and three small dams, each between 50 and
100 m2, that opened onto the perched near-surface aquifer were used for seaweed trials as well (idem).

This site was used for seaweed trials from May 2003 to December 2003 and then from July 2004 to
March 2005. Between December 2003 and July 2004, an additional pump was put into the system, the
drains were widened to permit better oxidation and precipitation of iron and a 1000m2 seaweed trial
pond was excavated between the settling ponds and the disposal basin.

3.2.3. Lake Charm

Lake Charm, Victoria is between Kerang and Swan Hill in the lower Lodden River (or North Central)
catchment which is plagued with soil mineralisation where saline water close to the surface is
degrading soils and limiting cropping and pasture capabilities. A farmer from the Lake Charm
Landcare group supported by the Victoria Department of Primary Industries constructed two basins
between his deep drainage system and the disposal of salt water in to the Lake Tutchewop saline water
disposal complex. The basins at Lake Charm were constructed to enable the adjustment of water
depth to maintain adequate light penetration, temperature stability and wind-driven movement of
water without disturbance of the seaweed. Dimensions were 20 m by 50 m by 0.5 m deep, set on a
North-South axis to make use of the wind for mixing but avoiding the long axis fetch that might push
seaweed to one end of the basin. One of the basins was lined with plastic to meet the requirements of
the Environmental Protection Agency to minimise saline water seepage.

The water source for the trial basins came from the sub-surface drainage of the farm, about one
kilometre upstream. This water, in which the saline groundwater was mixed with irrigation water, was
variable in its nutrient load dependent upon seasonal fertilisation of the farm. This provides another
important difference in water type for seaweed trials and a trial basin without the established
ecosystem that exists at Girgarre.

The site was used for seaweed trials from October to December 2004 and began again in November
2005 after re-installation of a plastic pond liner to limit seepage.

24
3.2.4. Donald
Two sets of seaweed trial basins were constructed under a National Landcare Program Innovation
Grant to the Donald and District Landcare Group for 2004-05. The area around Donald is subject to
“dryland salinity” which is explained by Dyson (2003-04) as paraphrased in the following two
paragraphs:

During a long geological history, oceanic incursions into Western Victoria have resulted in saline
water tables under the area now known as Buloke Shire. Discharges of saline groundwater from large
regional aquifers cause dryland salinity throughout much of the Shire. Over the course of a century,
un-informed farming methods and water management interventions have resulted in a rise of these
saline water tables sufficiently close to the surface to degrade soils and limit cropping. Changes to
landscape hydrology have also re-activated ancient saline groundwater discharge zones. In these cases
freshwater wetlands have reverted to somewhat barren saline swamps.

Salinity issues in the lower catchment are driven by the upward discharge of saline groundwater from
a regional alluvial aquifer comprising coarse-grained sands and gravels lying within the trench of an
ancient buried river system (deep lead). This aquifer forms a giant sub-surface conduit that drains
groundwater from most of the southern catchment into the lower floodplain. The regional nature of
this system mitigates opportunities for traditional farming system approaches to manage salinity that
try to reduce recharge to the groundwater system. Saline groundwater inflow is sustained irrespective
of immediate changes in the surface water balance.

Following the strategy that adaptation to saline conditions through the development and adoption of
innovative saline industries is more appropriate than trying to mitigate long term regional changes in
groundwater evolution, the Donald and District Landcare Group opted to run seaweed trials to learn
whether there was any commercial potential for a new industry.

There are two farms at Donald where a set of trial basins were constructed on paddocks where saline
water tables have degraded productive farm land. One set is adjacent to a saline discharge swamp that
had been a freshwater wetland little more than a generation ago. The second set is at the edge of a
flood plain that occasionally carries saline water from Lake Buloke.

Both sites lie atop an ancient deep lead of saline water 20-50 m below the surface and up to a
kilometre wide; which was part of a water course underlying the current Avon-Richardson system.
Reticulation is from two groundwater bores, which operate at 1 and at 3 liters per second, the higher
volume coming from the more southerly bore, which is closer to the centre of the deep lead. Both sets
of basins consist of two ponds of 1000 m2 each and an associated terminal disposal basin. They are
separated by about three km in a straight line along the axis of the ancient river bed (Dyson, 2004).

The sites were used for seaweed trials between 30 Dec 2004 - June 2005 and began again from
November 2005.

25
 photo: Trevor Campbell

Figure 4. Aerial view of one set of Donald seaweed trial basins and terminal disposal basin

3.3. Experimental methods and equipment

Gracilaria propagules collected from its coastal habitat were initially cleaned of epiphytes and
potential pests using 0.5-2% Cl-. This was not very successful since both the epiphyte species and the
target species were vulnerable at the same concentrations. It was later found that by holding 5-10 kg
of seaweed without water in 52 1 Styrofoam boxes (broccoli boxes from greengrocers) for 3-5 days
that the associated fine bodied epiphyte seaweeds: Polysiphonia, Ceramium and Ectocarpus would be
dehydrated while Gracilaria remained fresh. Under the same storage regime, the associated
zooplankton isopods and amphipods and most of the gastropod molluscs would not survive and could
be shaken out of the seaweed mass.

Collected seaweed material was held in 2.25 m2 plastic swimming pools (Clarke Rubber, Melbourne)
in Girgarre water at ambient salinity (usually 20-24 ppt) for inspection and removal of any foreign
matter before being used in trials. The pools were “aged” in water for two days before use in trials.

Small quantities of Gracilaria in closed containers permit precise control over variables supporting
viability and growth of the seaweed but do not provide information on those variables that would be
encountered in farm-scale evaporation basins where large quantities of seaweed are in contact with
soils and the open environments expected under agricultural conditions. Although some trials were
carried out in small containers with artificial seawater and controlled lighting, most trials were done in
situ, even where containers were placed into the larger trial basins.

Girgarre trial ponds of 1000m2 and 3000 m2 were used for bioassays and for comparative studies.
Polyethylene baskets 40 cm diameter by 35 cm deep (K-Mart, Shepparton) were distributed in an
orchard distribution across the 1,000 and 3,000 m2 trial basins. Treatments (eco-type variety of
Gracilaria and depth) were randomised.

For quantitative studies, twelve of the 2.25 m2 plastic pools (1.5 metre by 1.5 metre by 0.80 metre)
were used to compare nutrient concentrations in a series of trials at Girgarre. The pools were placed
into one of the 3000m2 trial basins and filled with 675 litres of water (30 cm depth) pumped from Bay
A, usually at 22 +/- 2 ppt salinity. Due to the lack of continuous water exchange between the plastic
pools and entire 3000m2 basin, steel mesh cages of the same dimensions as the pools but with 1 cm
mesh, were used to compare viability and growth rates of seaweed in the Girgarre basins.

Where studies in situ required quantification of growth rates in open water, a series of extruded
polyethylene tubular mesh bags were used to hold weighed amounts of seaweed. At first orange
coloured bags with 12mm x 12mm mesh were used but the mesh size permitted too much loss of
fragmenting Gracilaria material. Later more appropriate extruded bags with 7mm x 7mm mesh ,
450 mm wide (Type 44R Atlas Rope and Cordage, Wetherill Park, NSW) were prepared into 35 cm

26
long tubes and suspended from lines staked into the substrate of the trial basins. Using these bags,
trials were run to compare seaweed growth in mesh tubes suspended at the surface with those placed
on the substrate, 20-40 cm below the surface of the water.

Despite the excavation of trial ponds on salt degraded lands over saline water tables at Donald and
Lake Charm, the Victorian Government demanded that trial basins be lined with plastic to avoid
seepage of saline water back into the surrounding landscape. The liner reduced contact between water
and soil, though wind drifted soil and calcium precipitation quickly provided about one to two mm of
substrate. On one hand the reduced quantity of soil might limit the adsorption of dissolved minerals
from the water but on the other hand, it might not provide minerals to solution. The black
polyethylene liners (black for UV protection) did heat the water somewhat but this was not a problem
since waste water exchange was used during high evaporation periods to maintain stable salinity and
temperatures remained below 30oC.

3.4. Groundwater chemistry at trial sites

Saline groundwater in Australia can originate as fossil water from deep leads that may contain ancient
sea water and its chemically and biologically transformed descendants that have moved downstream in
great underground rivers 40-50 metres below the surface. Hydraulic pressure from higher elevations
upstream can push this saline water to the surface at lower elevations downstream. This is often the
case with dry land salinity where large volumes of saline groundwater may be available in regions
with little rainfall and only ephemeral surface water. Alternatively, there is irrigation salinity in which
non-saline surface water carries a load of agriculture nutrients past the root zone of pasture and annual
grain crops to add salts to the salinity of a rising water table. In some ways irrigation-enhanced saline
groundwater can be richer in nutrients and better able to support marine plants, though it is likely to be
more seasonally variable.

The basic ionic composition of the inland saline waters was compared to sea water for total dissolved
solids (salinity) and for trace elements that might effect the growth of seaweed and the production or
quality of colloids (see section 1.3 above). Some inland saline waters are overly rich in calcium or
iron and many of the groundwater samples in the Murray valley have been shown to be poor in
potassium compared to sea water.

Table 3. Comparative element concentration for waters in this study (in mg/ litre)

Lake
element ocean Donald 1 Donald 2 Charm Girgarre Girgarre Jeparit
salinity 32.6 ppt 40 ppt 30 ppt 20 ppt 16 ppt sediment 29.4 ppt
P 0.05 <0.01 <0.01 <0.01 0.013 170
S 884 1270 1150 987 512 75 1295
Cu 0.005 <0.001 <0.001 <0.001 <0.02 2 0.03
Fe 0.011 0.031 0.029 0.031 0.06 76 52
Mn 0.005 0.025 <0.005 <0.005 5 0.86
Mo x <0.005 0.011 0.008 5 <0.01
Ni 0.0001 <0.002 <0.002 <0.002 nd 0.06
Zn 0.005 0.004 0.001 0.002 0.09 11 0.09
Ca 400 502 423 326 76 46 700
K 380 179 213 28 17 19 170
Mg 1272 1460 1150 607 947 19 1600
Na 10561 11700 8960 4520 5210 58 10000
B 4.6 5.25 6.13 3.04 0.4 15 4.8

27
Table 3 shows the concentration of ions in mg per litre or parts per million (ppm) in the waters on trial
during this study. Seaweeds use active transport to concentrate minerals thousands of times their
concentration in the sea.

Comparing the above table to the list in Table 4, which compares the concentration of metal ions in
grain and seaweed, none of these ions dissolved in the saline waters being studied are higher than the
range of concentration in seaweeds and hence not expected to be toxic to the seaweeds. Conversely,
the sodium concentration in normal seawater is higher than the toxic concentration in barley and wheat
and hence seawater at 34 ppt is toxic to these crops (Reuter et al (1997) but not to seaweed.

The concentration of minerals in seaweed is within the range of those in terrestrial crops like barley or
wheat for N-P-K and the trace elements but for the other major nutrients: B, Ca, Mg., Na and S,
seaweed concentrations are 3-10 times higher than in crops.

Table 4. Concentration of mineral ions in seawater, crops and seaweed

concentration in mg/kg (ppm)
in seawater in grains (1) in seaweeds (2)
normal seawater critical mean in range in
element (3) deficiency toxic Seaweeds Seaweeds
B 4.6 3-5 >50 184 15-910
Ca 400 2000 >10000 14300 2-360000
Mg 1272 1000 3000 7300 1900-66000
Na 10556 5000 8000 nd nd
K 380 23000 60000 41100 30000-82000
Cu .005 1.2 15-75 15 0.6-80
Zn 0.005 15 300 90 2-600
Fe 0.011 35-100 ? 750 300 90-1500
Mn 0.005 10-15 150-400 50 4-240
N 0.35 1.5 >6.5 2.3 0.05-6.5
P 0.07 2000 7000 2800 2800-12000
S 884 1500 4000 19400 4500-82000
Mo 0.0005 .05-0.1 nd nd nd

Not shown in these tables are the “heavy metals, ”arsenic cadmium, lead, tin, strontium, et al, which
although not required by plants and usually occur in low concentrations in seawater, may occasionally
exist in ground waters at concentrations toxic to plants. Although tin and arsenic only occur at
0.003 mg/l, less than the micronutrients (recent studies show that Mo concentrations can be up to
0.05 mg/l), strontium is the fifth most abundant metal in the sea at 8 mg/l.

In field trials, salinity was occasionally adjusted higher by evaporation, where necessary to meet target
concentrations. No fresh water was used to lower salinity, though autumn and winter rains resulted in
lowered salinity in several experiments. The lowered salinity in at lest one case resulted in a bloom of
weed species that out-competed Gracilaria. Evidence from its natural estuarine habitat suggested that
Gracilaria type M can grow in a very wide range of salinities and may have a competitive advantage
in hyper-saline waters higher than 35 ppt.

The saline water ions in the trial ponds were regularly analysed and water chemistry dynamics
measured using a Lovibond PC 22 field photometer (Tintometer GmbH, Dortmund, Germany) for a
range of macro and micro-nutrients.

3.4.1. Girgarre groundwater

The Girgarre Evaporation Basin is more than 15 years old and has developed a relatively stable but
seasonal saline ecosystem, characterised by various species of birds more often found around coastal
wetlands and a benthic flora dominated by the seagrass Ruppia sp. There are seasonal blooms of the

28
euryhaline (see Glossary page 40) green seaweed, Enteromorpha intestinalis in Bays A and B and the
hypersaline Bay C has seasonal blooms of the red micro-algae, Dunalliela salina followed by a bloom
of the zoo-plankton Artemia salina.

Water at the Girgarre Evaporation Basin is pumped seasonally at 15-20 Ml per annum from several
bores in the area to maintain the groundwater level below about two metres so that crops and pasture
are not degraded by the salt.

Table 5 contains a comparison between “normal” (open ocean) seawater and that at the bore entering
the Girgarre basin. Note that the concentration of manganese, copper, iron and especially zinc are up
to 300 times greater than that of seawater but boron and potassium at Girgarre are one-third and one-
eighth the concentration of normal seawater. The high concentrations are not toxic to seaweeds; in
fact Gracilaria will actively concentrate zinc a further 150 times and some of the other metals more
than 1000 times their concentration in Girgarre water. The very low potassium concentration may be
limiting to plant growth, especially as the Na : K ratio may limit the active transport of potassium and
the passive release of sodium ions (Newman, 2004). The high concentrations of Ruppia seagrass is an
indication that these waters support plant growth.

From the table it can be seen that to make Girgarre water more ionically equivalent to seawater, it
would be supplemented with potassium and boron, in addition to the nitrogen and phosphate required
to support the targeted growth rates.

Table 5. Comparison of nutrient ions in seawater and saline groundwater at Girgarre,

presented as percentages of each element to allow for differences in salinity.
Natural "normal" seawater In Girgarre basin A (2)
total element %
dissolved as tds at % of element
solids (tds) 32.6ppt as tds at 11.9 mg/kg in
element in mg/ kg range salinity tds in mg/kg ppt salinity sediment (1)
Na 10556 32.40% 3200 26.94% 58
Mg 1272 3.90% 670 5.64% 19
S 884 2.71% 512 (1) 4.31% 75
by
SO4 2500 calculation 7.67% 1300 10.94%
Ca 400 1.23% 350 2.95% 46
K 380 1.17% 17 0.14% 19
C 28 0.09%
by
HCO3 142 calculation 0.44% 320 2.69%
Sr 13 0.04%
B 4.6 0.01% 0.4 (1) 0.003% 15
0.01 to 0.7 in
N 0.35 compounds 0.0011% 0.91 0.01%
Al 0.5 0.0015% <0.1 0.0006% 68
P 0.07 0.001 to 0.10 0.0002% 0.013 0.0001%
Fe 0.011 0.002 to 0.02 0.00003% 0.06 0.0005% 76
Cu 0.005 0.001 to 0.01 0.000015% <0.02 (1) 0.0001% 2
Zn 0.005 0.00002% 0.09 (1) 0.0006% 11
Mn 0.005 0.001 - 0.01 0.00002% <0.01 0.0001% 5
Cd 0.0004 0.000001% 0.02
0.0005-
Mo 0.005 0.000002% 5
(1) Cristen, et al,2000

29
3.4.2. Japarit groundwater
The Parilla Sands groundwater, pumped up from 6-18 metres below the surface, came up at 30 ppt
salinity and a very acidic pH of 5.2. This water carried a very high load of dissolved iron at 52 mg/l
nearly 5,000 times greater than is found in “normal” seawater at pH of 8.2. Iron is, after all, the fourth
most common element on the earth, but most of it is tied up as oxidized minerals. Upon reaching the
surface, the pH quickly increased to about 7, the iron became oxidized and precipitated out as Fe(OH)2
further oxidizing to Fe(OH)3 flocculent. By mid-August 2003 the bottom of the disposal basin was
covered by a thick slimy brown leathery skin of iron flocculent containing an iron-fixing bacterial
colony. Once this precipitated iron was fixed into the matrix, the dissolved iron remaining in the
water column was reduced to 0.02 mg/l.

Water came out of the bore at 19oC and in summer heated to 27oC by the time it reached the settling
ponds. Summer evaporation rates were greater than the net windmill pumping rate and until a second
pump was installed in 2004, water evaporated away in summer and the disposal basin became dry salt.

The pH remained at about 7 in the disposal basin, which did not support any plant communities apart
from the iron-fixing bacteria. The small dams, which did support plants, had a varying pH between
8.75 and 9.02 during the sunniest part of the day due to a reduction of CO2 by photosynthesis.

To improve the potential for seaweed cultivation at Jeparit the DPI installed a second groundwater
pump, widened the drain between the bore and the settling ponds and installed a seaweed trial basin
during 2004. Due to the lack of control over water flow, high concentrations of iron (4.3 to 20.5 mg/l)
and low pH (6.9) were still the norm entering the seaweed trial pond. This resulted in continual
precipitation of oxidized iron and a reduction of light intensity below 20 cm due to the flocculent.

3.4.3. Lake Charm groundwater

In November 2003 an analysis of the groundwater from a flow-though manhole in the drain between
the farm and Lake Kelly, showed that salinity was 20 ppt, alkalinity was high at 745 mg/l CaCO3 and
the pH was slightly alkaline at 7.42. Compared with seawater, potassium was very low at 27 mg/l-K
(seawater at that salinity would be about 230 mg/l-K), iron and copper were present in low quantities,
which is a good sign for plant cultivation. There was plenty of nitrate, which may have been run-off
from upstream paddocks and perhaps seasonal. PO4 was low as expected but not limiting at 0.3 mg/l.
The bore, which runs all summer, picks up saline groundwater seasonally, mixed with irrigation water
from 3 to 21 m depth.

During the next two years water quality was monitored to support seaweed trials. Salinity varied very
little from 20 ppt, increasing with evaporation and decreasing with rainfall when groundwater was not
exchanged, by only 4-5 ppt. pH remained stable at around 8+/- 0.2 throughout, which confirmed the
high alkalinity readings. Temperature also remained stable at about 20+/- 2oC when water was at least
occasionally exchanged. When water was not being exchanged, water temperature fell to 14oC in
winter and rose to 28oC in summer. The trace elements Fe, Cu, Zn and B were always in low but
sufficient quantity to support seaweed growth. Heavy metals and potential toxins were not measured.

The only unusual factors were potassium and calcium concentrations. Potassium ranged between one-
tenth and one-fifth that of seawater at equivalent salinities and is doubtless a constraint to seaweed
viability and growth. It was difficult to build potassium concentration in the water, suggesting that it
was being adsorbed or compounded into minerals unavailable to plant growth. Calcium concentration
was higher than expected in seawater. Measured as CaCO3 hardness and converted to calcium, its
concentration was variable but always 3-5 times greater than that expected in seawater at equivalent
salinities.

30
3.4.4. Donald groundwater
The groundwater at both the Donald seaweed trial sites was drawn from the same deep lead of ancient
saline water. Unlike the Parilla Sands groundwater at Jeparit these fossil waters were relatively low in
iron but high in calcium. Between the two sites is a near-surface deposit of Gypsum (CaSO4).

The following groundwater analyses were taken from saline water that had been standing in the trial
basins. Note that the calcium, magnesium and major nutrient concentrations are virtually the same as
in seawater. Note also how high the bi-carbonate concentration is; 30-40 times the concentration in
- =
seawater, which is typical of river waters, where HCO3- is the dominant anion, rather than Cl or SO4 .
At both sites in Donald thin flat crystals formed in sheets covering the bottom of the basins and
forming as thin crusts on plastic mesh bags, seaweeds and grasses that were blown into the basins.
Given the nature of the groundwater, the crystal is likely to have calcium and carbonate in its structure
(Chapman, 1992).

As at the other sites, nutrients in water were analysed regularly on-site with a Lovibond PC22 field
photometer. Most ionic species were about equivalent to seawater and gave no concern for plant
viability, including those trace elements like copper, zinc and boron. Only potassium concentrations
were cause for concern at less than 50 mg per litre, about one-tenth of the seawater concentration.
Potassium salts were added to the basin as nitrate or sulphate to bring concentrations back in line with
seawater. Table 6 provides a comparison of mineral ions between normal seawater and the two
Donald sites.

Table 6. Comparative ion ratios of elements in seawater and groundwater at Donald, Victoria
concentration in concentration in
element seawater Peter's Paul's seawater Peter's Paul's
comparison as % % % mg/l mg/l mg/l
Cl 55.1% nd nd 19000 nd nd
Na 30.6% 29.3% 29.9% 10556 11700 8960.00
S as SO4 7.7% 9.5% 11.5% 2649 3810 3450
Mg 3.8% 3.7% 3.8% 1310 1460 1150
Ca 1.2% 1.3% 1.4% 400 502 423
K 1.1% 0.45% 0.71% 380 179 213
HCO3 0.4% 11.10% 16.00%
B as H3BO3 0.08% 0.07% 0.12% 26 30 35
Fe 0.00003% 0.00008% 0.00010% 0.01 0.031 0.029
Mn 0.00001% 0.00006% <0.00002% 0.002 0.025 <0.005
Zn 0.00002% 0.000010% 0.000003% 0.007 0.004 0.001
Cu 0.00001% 0.000003% 0.000003% 0.004 <0.005 0.011
N (variable) 0.00122% 0.00001% ~ 0.4
P (variable) 0.00017% <0.00003% <0.0003% ~0.06 <0.01
Mo (query?) 0.00001% 0.00002% 0.00004% 0.0007

salinity 34.5 ppt 40 ppt 30 ppt 34.5 ppt 40 ppt 30 ppt

pH 8.2 8.0 8.2
percentage
major of each groundwater bores at
elements element Donald separated by about
required for from 2000 m. Sampled after
seaweed "normal residence in evaporation
growth seawater" basin.

31
3.4.5. Use of nutrients
Nutrients were supplemented in weekly or fortnightly pulses to the trials. Earlier experiments were
based on a supplement of 4% nitrogen content of the expected new seaweed biomass assuming a
growth of 3% to 5% per day. In some experiments this high nutrient load encouraged the growth of
competing algae. Since the literature suggests that epiphytes can be overcome by pulse feeding and
feeding at night, in some trials, the seaweed was soaked in a nutrient bath for up to two days before
setting it into the basins. Since nitrogen is posited as the limiting variable, where the water quality can
otherwise support seaweed viability, growth was correlated to nitrogen concentration. Precise
quantification was difficult in 1000 m2 trial basins due to the uncontrolled contribution of nitrogen by
birds, wind-blown soil particles and by run-off associated with rain.

The literature (Ryder & Nelson, 2004; Liu & Dong, 2001) shows that Gracilaria is saturated with
nitrogen at 4% to 5.8% of dry weight and can grow on that stored nitrogen for a fortnight. The
average concentration of nitrogen in seaweed by dry weight is 2.3% (Lobbana and Harrison, 1994).
Making an additional 4% of seaweed dry weight as nitrogen available to the plants should be sufficient
to support at least twice the standing crop, i.e. double the original mass, provided all other nutrients
are sufficient. At 5% growth per day, Gracilaria could achieve that doubling in 14 days. Trials for
the assessment of growth were based on this premise of adding 4% of the dry weight of the standing
crop as nitrogen, preferably as the NH4 ion, to double the standing crop in 14 days. Ammonium
nitrogen was used in the form of ammonium sulphate, containing 21% nitrogen.

Phosphate was supplemented to be available at one-tenth the molar concentration of Nitrogen since the
literature suggests that best growth is achieved at this ratio (Navarro-Angulo & Robledo, 1990),
though Tsai, et al (2005) achieved their best growth at a N:P ration of 20:1. Superphosphate was used
as the phosphate source with the understanding that only 7% is soluble and immediately available.
Potassium concentrations were supplemented to the equivalent of seawater at 1.16% of total salts. In
some inland saline waters large supplements are required to maintain the seawater Na : K ratio of
about 27 : 1 to replicate the natural environment of seaweed. Potassium sulphate and potassium
nitrate were used as the potassium source (and in the latter case as the source of nitrogen). Iron was
considered to be the fourth limiting variable but was in sufficient concentration in the waters at
Girgarre and was not supplemented. Hence the nutrients ratio used to stimulate the growth of
Gracilaria was N:P:K at 10:1:15.

Water depth was manipulated between 20-30 cm to maintain adequate light penetration, temperature
stability and to limit wind-driven movement of water so as not to destabilising the plants but avoid the
presence of a static boundary layer around each plant that might limit the passage of gasses, wastes
and nutrients across the cell walls.

32
4. Results and Discussion
4.1. Seaweed assessment trials
Much of the experimental seaweed work comprised bioassay trials to assess the ability of the species
(or eco-type) to live and grow in saline groundwaters and to evaluate the groundwater as a medium,
based on the literature reviewed in section 2 above.

Trial G1
In March 50 kg of type F Gracilaria collected by Kilkunda Abalone Farm (Phillip Island, Vic) at
Flinders Victoria was seeded into Girgarre trial Pond T-4A (1000m2). New water was mixed from
Basin A and Pond T3 to result in 26 ppt salinity and a depth of 50 cm. No additional nutrients were
used since NO3-N was about 0.1 mg/l and N-NH4 was about 1 mg/l in the ambient water. Phosphate
was 0.17 mg/l The day-night water temperature range was 14-26oC. Secchi disc (see Glosssary, p
45) reading was 15 cm due to brown colour of the incoming water. By 6 April, the water had cleared,
the min-max temperature was 13-23oC, seaweed was visible but the green seaweed Enteromorpha
intestinalis had become dominant. Temperature slowly fell during May to a min-max of 11-19oC

Results: During the week of 11 April, 75 mm of rain fell in the region, lowering the salinity to 18 ppt.

Table 7. Nutrient concentrations from 14th April to 3 June

Date nitrate Ammonium phosphate notes
14 April 0.10 0.89 0.17 Cladoceran zooplankton
24 April 0.09 0.75 0.00 Enteromorpha on the surface
2 May 0.08 0.62 0.15 Cladoceran population still high
18 May 0.08 0.32 0.09 Pond bottom becoming anoxic
3 June 0.08 0.21 0.13 Gracilaria wrapped in

Enteromorpha was dying. Analysis of the anoxic mud showed high iron (0.97 mg/l) and potassium
(1200 mg/l) that would have been available to plants at the low pH expected just below the surface of
this mud.

By 10 May there was a very high density of Cladoceran zoo-plankton and the green seaweed
Enteromorpha intestinalis had become dominant. Enteromorpha was observed growing around the
clumps of Gracilaria as if it were a nutrient source. Inside the mass of Enteromorpha, the Gracilaria,
like jam in a doughnut, was still alive but clearly cut off by the Enteromorpha from the light and any
transfer of nutrients, waste products or gasses to the free waters outside the Enteromorpha. In mid
May, the Gracilaria was black in colour and showed some growing tips but by early June it was clear
that Gracilaria was dying and disappearing. Salinity had increased with evaporation throughout May
and by June was 26 ppt again.

Trials G2
May – Sept: seventeen polyethylene baskets were set in an orchard distribution across the 1,000 and
3,000 m2 trial basins, which were covered with sea grass and benthic diatoms. Treatments (eco-type
F, J and M varieties of Gracilaria) were randomised. In each basket, 350 g of Gracilaria were stocked,
together with a mixture of N-P-K at 10:1:15 to stimulate a target growth of 5% per day for two weeks,
assuming no water exchange, to 660g wet weight. Seaweed was weighed each fortnight.

Growth never reached one percent per day in any of the baskets. Although there was some
Enteromorpha in the stock, this died over the winter as the minimum daily temperature fell from 11oC
to 6.5oC. There was no indication that any variety of Gracilaria used was superior to any other. Over
time most of the samples being tested fragmented into 0.5-1 cm pieces and floated off through the 2

33
cm by 7 cm openings in the baskets. Although some growth was noted and seaweed in the anoxic mud
did not die, much of the sample slowly changed colour from dark brown to light green, straw yellow
then white in the process of dying.

The dominant flora in the trial basin was the seagrass Ruppia sp which covered the bottom of the basin
at about 20 kg per m2 (wet weight). Beneath the Ruppia, the substrate was anoxic and occasionally
topped with benthic diatoms and bacteria. Movement of researchers in the basin always resulted in
suspended silt, some of which would invariably cover the seaweed in the basket with a thin layer of
fine silt. Analysis of silt-laden water in the basin did not show any significant difference in nutrient
concentration from clear water. Toxic metals were not examined.

A more oceanic species of Gracilaria called G. secundata was evaluated in the estuarine conditions of
Girgarre on two occasions but in both trials it died after one week. This would be typical of an
oceanic species that is adapted to a narrow range of temperature and salinity (Dawes, et al. 1999).

4.2. Growth studies and nutrient use

Growth rate was calculated as percent growth per day using the compound growth formula
Wf=Wo(1+r)n where Wf is final weight, Wo is original weight, r is the rate of growth per day and n is
the number of days of growth.

4.2.1. Girgarre
A range of nutrient concentrations taken from published studies, was used to assess the growth of
500g of two Gracilaria varieties of seaweed in 2.25 m2 plastic pools filled to 675 litres with 20ppt
saline water from Girgarre Bay A. Nitrogen was the focus nutrient and ranged from 2.1 to 43.6 mg/l
replicating concentrations shown in the research papers cited in Table 8. A control had only the
ambient nitrogen concentration which was 0.58 mg/l. Note that the difference between these
experiments and the previous sets is that nitrogen was supplemented in these to provide a measured
concentration per volume of water rather than to meet the projected nitrogen needs of new seaweed
growth. The first round of studies during September was contaminated by the green seaweed,
Enteromorpha intestinalis, indigenous to the Girgarre Basin at least since 1992 (Anon, 1992).
Lighting was ambient from 15 October to 02 December and temperatures ranged from 17o to 32oC
during this period. Phosphate was supplemented at 0.1M of the nitrogen molarity. Potassium was
supplemented to provide 100mg/l in each 675 litre pool. Table 8 shows the results of this experiment.

Table 8. Growth rate of two Gracilaria varieties in a range of nitrogen concentrations

Pickering, One Mole-
Media reference 1993 Provasoli, 1963 Nitrogen Glen, 2002 Control
Gracilaria variety M J M J M J M J M J
Mg/l- nitrogen 2.1 2.1 4.6 4.6 14 14 43.6 43.6 0.58 0.58
pool number 3 9 2, 5 10 4 11 6 12 1 8
days 15 15 15 15 15 15 15 15 15 15
1.4%, 2.11%,
growth /day I 0% 0.60% 2.04% 0.86% 0% 0.56% 0% 2.00% 0% 0%
days 42 31 42 31 42 31 42 31 nd nd
growth /day II 1.00% 1.48% 0.97% 0.85% 0% 1.41% 0% 0.60%
mean growth/day 0.50% 1.04% 1.47% 1.27% 0% 0.99% 0% 1.30% 0% 0%

In none of the trials did the Gracilaria growth rate reach the targeted 5% per day. This trial showed
that at very high concentrations of nutrients (14 and 43.6 mg/l), the Gracilaria did not begin to grow
for at least 15 days. The best growth was not at the highest concentration of nitrogen but at 4.6 mg/l,
used in the Provasoli medium, which was designed to culture algae in vitro. It also suggests that the
J type Gracilaria grows better than the M type in field conditions, a contradiction of the careful in

34
vitro studies shown in Byrne, et al. (2003). Additionally the growth rates are consistently very low
despite the conducive temperature and light conditions of Spring.

There are other contradictions between these results and the literature. Hypothetically, If 4% N was
sufficient to permit the doubling of biomass, then 4.4 mg/l- N would provide that quantity for 500 g of
seaweed. Hanisak (1987) and Chuan-Chuan Tsai (2005) reported that the critical amount of nitrogen
in Gracilaria was 2% of dry weight. That means that at 2% new nitrogen, over a 31 day period, the
maximum new growth of Gracilaria possible would be less than 57g and a growth rate over the
31 days from the 500 g introduction could be no more than 0.32%. The Pickering data in Table 8
shows a growth rate double that in the first 15 days and nearly five times the potential after 31 days.
Why such disparity?

As early as the first week of the trial, feathers were noted in the pools and by the measurements on day
15, the water in the pools were dark brown with phytoplankton. After a month the green weed algae
Enteromorpha was plentiful in most pools and dominant in some. Where phytoplankton appeared to
be the dominant user of nutrients, the Gracilaria was healthy but where Enteromorpha was dominant,
the Gracilaria had started to turn white and die. In addition to the feathers, bird droppings collected in
the corners of the pools, where it was obvious that birds had been perching.

The Girgarre evaporation basin is a wetland environment that draws hundreds of birds of at least six
species, some as large as pelicans and black swans. Bird droppings were an uncontrollable variable in
the pools that made quantitative nutrient studies impossible at Girgarre.

Growth experiments in pools

Throughout 2004 a long series of growth trials were continued using 1.5m by 1.5m plastic pools and
1 cm mesh cages of the same dimension set into a 3000m2 Girgarre trial basin. Water was taken from
the adjacent Bay A and pools were fertilised with NPK to sustain growth of 2.5% per day for 14 days.
The cages were set into pools of fertilised water fortnightly to “pulse fertilise” the Gracilaria. Due to
the lack of control over the addition of bird manure, nutrients (N, P, K and Fe) were monitored only to
assess whether they were between the probable limits of deficiency and toxicity. The pH and
alkalinity were noted as an indication of photosynthesis and the availability of carbon dioxide. Two of
the containers (one cage and one pool) were covered with shade cloth in an attempt to limit the bird
droppings.

Table 9 shows the results of growth experiments at Girgarre from March through October 2004. As
the data shows, daily growth rates never approached the targeted rates from the scientific literature
(see Table 2, page 16), which range from 2% to 24% per day and averaged 6.7% per day for the cited
studies.

Girgarre growth rates averaged less than 1% per day and T-tests showed that there were no growth
differences (at >95%) between fertilised and unfertilised sets, higher or lower concentrations of
ammonium, between closed waters of the 2.25m2 pools and the open waters of cages in the 3000 m2
trial basin or with the seasonal change in temperature and salinity. Although the Gracilaria in the
open cages looked better (qualitatively by smell, growth points, crispness, colour and lustre) than
many of the trial samples in the plastic pools, there was still fragmentation of the thallus and loss of
material through the mesh of the cages. Qualitatively, the Gracilaria in the shade-cloth-covered pool
continued to look healthy even though there was no growth under the 70% shade cloth.

35
Figure 6. Filling experimental pools at Girgarre.

36
 Table 9. (part a) Results of growth trials at Girgarre March to July 2004.

kg
growth rate

growth rate

growth rate

growth rate

growth rate
days from
cage/pool

Growth 7
treatment

18 days

18 days

18 days

27 May

15 days

16 days
12 May
19 Mar

24 Apr

17 Jun

10 Jun
24 Jun

10-Jul
6 Apr
kg

kg

kg

kg

kg

kg
3

4 mud on bottom 5.2 start 4.75 0% 4.92 0.17% Enteromorpha 12.8 new
5
6 8.75 new
8 shade cloth 2.0 2.00 0.00% 1.50 0.00% 1.70 0.70% 1.50 0.00% new 9.15 new 9.15 9.65 0.33%
9 2.0 2.35 0.90% 2.85 1.08% 2.39 0% Enteromorpha
10 0.5 0.66 1.55% 0.80 1.07% 0.62 0.18% 0.65 0.32% Enteromorpha
37

11 0.5 0.60 1.02% 0.62 0.18% Enteromorpha

12 5.0 5.80 0.83% 6.00 0.19% 6.45 0.40% Enteromorpha
control no
13 fertiliser 2.00 start 2.00 0.00% 2.63 1.53% 2.10 0.00% Enteromorpha new 10 11.25 0.74%

A mesh cage 5.0 4.25 0.00% 3.40 0.00%

mesh cage
B 2.0 1.54 0.00% 1.30 0.00%
mesh cage
C shade cloth 2.0 1.73 0.00% 1.35 0.00%
orange
mesh
bag open water 0.20 new
temp 14 to 30 15 to 21 7 to 18 8 to 17 6 to 11-13
17
o
salinity pools /oo 21ppt 21 ppt 21 ppt 20-21 ppt
cages 30-31 ppt 25

(a) the yellow pool was 7.1 m2 and contained 2800 litres
healthy Gracialria Enteromorpha contamination Enteromorpha dominant
 Table 9. (part b) Results of growth trials at Girgarre July-October 2004

kg
cage/pool

treatment

17 Sept
rate 16

rate 20

rate 18

rate 15

rate 21
15 Aug
growth

growth

growth

growth

growth

growth
rate 16
02 Sep

24 Oct
26 Jul

3 Oct
days

days

days

days

days

days
kg

kg

kg

kg

kg
5.0 mostly
3 3.83 new Enteromorpa 1.28%
mud on 4.8 mostly
4 bottom 12.90 0.00% 3.75 new Enteromorpa 1.18%
5 9.50 new
6 11.05 0.65% 13.25 0.55%
8 shade cloth 8.40 0.00% 8.3 0.00%
9 5.40 new 4.20 0.00%
10
38

11 2.85 5.5
12 11.20 new 13.2 0.53% 8.0 0.00%
4 bags
floating 1.00 1.25 1.12% 1.49 0.98%
yellow pool (a)
8 bags on
bottom 2.00 2.30 0.70% 2.53 0.53% 2.65 0.31%
orange mesh
bag open water 0.26 0.73% 0.20 0.00%
20 orange mesh open water
bags horizontal 7.0 new 8.51 1.09% 8.83 0.25% 8.71 0%
12 orange mesh open water
bags vertical 4.7 new 4.67 0.00% 4.77 0.14% 4.5 0%
17 orange mesh
bags 6.21 new 5.39 0.00% 4.82
temp 5 to 12 6 to 13 7 to 20
salinity pools 25 ppt 25 ppt 35 ppt
cages
a) the yellow pool was 7.1 m2 and contained 2800litres
Some of the pools that initially seemed healthy for a few weeks and showed some growth, later
became weaker (sensu: loss of colour and turgor, slower growth rate) and were fully overtaken by
Enteromorpha after about six weeks. An example of this is shown in Table 9 (part a) by the coloured
boxes. The blue boxes show healthy pure Gracilaria culture and the pink boxes indicated where
Enteromorpha was present as a contaminant. In every case, where the Gracilaria became
contaminated with Enteromorpha, the latter species quickly became dominant.

Since the comparison between the closed pools and open mesh cages seemed to favour the cages, new
trials were used to examine this further. Mesh bags containing Gracilaria were laid out in the open
water, attached to stakes and lines, either horizontally (ie, maximising the area at the surface of the
water) or vertically (less open to the sun). Again there was no difference in growth rates (>95%
confidence) from the mean daily growth rate between the closed pools and the open cages or mesh
sacks.

The major constraint to Gracilaria growth appeared to be competition by the green seaweed
Enteromorpha. From the end of May Enteromorpha began to contaminate Gracilaria cultures and
soon became dominant. It was not gone by mid October when the trials at Girgarre were abandoned.
There was no correlation between Enteromorpha abundance and either temperature or salinity. It
seems to require a nutrient rich substrate and settled on and out-grew Gracilaria, at times totally
covering the Gracilaria like the dough around a sausage roll. Under some conditions Enteromorpha
seemed more like a predator than a successful competitor, taking nutrients out of the Gracilaria.

The 2003 studies had suggested that there was some competitive advantage over Enteromorpha to
having greater concentrations of Gracilaria stocked per unit of area. During the height of the
Enteromorpha invasion in June 2004, three small clumps of Enteromorpha (210g, 290g and 290g)
were placed in containers with holes to permit water exchange and set into a plastic pool containing 18
kg of Gracilaria to assess whether there was any competitive advantage to a large mass of Gracilaria
that would enable it to resist or out-compete Enteromorpha. After 23 days the Enteromorpha had not
spread to the Gracilaria, and had neither died away or grown further.

4.2.2. Jeparit
Seaweed bioassays in the saline water disposal basin at Jeparit during 2003 were constrained by the
very high iron concentration in the groundwater and insufficient water flow from the windmill to
cover evaporation during the late Spring and Summer. Sufficient oxygen to precipitate all the iron
was not available until after the water reached the evaporation basin. In precipitating, the iron bound
into a flocculent that covered any seaweed placed in the water and formed large leathery sheets on the
floor of the evaporation basin. The iron hydroxide/ iron oxide coating did not permit the seaweed to
exchange gases, waste products and nutrients and resulted in the death of the seaweed.

From October 2003 to January 2004, as water dried up in the evaporation basin, seaweed trials were
carried out in a small dam that accessed a shallow aquifer with salinity that rose from 11 to 30 ppt and
had more oceanic levels of iron (0.01 mg/l). Growth rates were between 1-2% per day, in baskets
fertilised with NPK at the same levels as used at Girgarre.

During the period between July 2004 and March 2005 seaweed trials were run in the new seaweed
trial basin using 12 mm open mesh bags as at Girgarre but these were placed just under the surface of
the water on the mesh floor of a floating raft and at 40 cm depth at the bottom of the basin. Table 10
shows the results of these trials.

39
Figure 7. Seaweed with too much iron from Jeparit

Table 10. Daily growth rates for Gracilaria at Jeparit 2004

% growth
% growth

% growth

% growth
20 days

19 days

35 days

18 days
13-Aug

per day

per day

per day

per day
01-Sep

bottom
05-Oct

23-Oct
24-Jul

mean

mean
top
Weight
Position (g) (g) (g) (g) (g)

sta 3 float
(n=3) intake 750 1090 1.89% 1090 0 1100 0.03% 1100 0 0.4218
sta 3 bottom
(n=1) 250 330 1.40% 400 1.00% 400 0.00% 400 0 0.5178
sta 4 float
(n=3)
outflow 750 1090 1.89% 1010 0 1250 0.61% 1150 0 0.4708

sta 4 bottom
(n=1) 250 330 1.40% 350 0.31% 375 0.20% 350 0 0.3704
disposal
basin float
(n=3) 750 790 0.26% 890 0.63% 950 0.19% 1120 0.92% 0.4416
disposal
basin
bottom
(n=1) 250 380 2.12% 430 0.65% 450 0.13% 530 0.91% 0.8292
unweighted
mean
growth per * *
day 1.49% 0.43% 0.19% 0.30% 0.5725 0.4447

* means are different

with 98% confidence

The data in Table 10 was taken from both the seaweed trial pond and the disposal basin, both holding
Gracilaria in floats at the surface and anchored to the bottom. The results show relatively good
growth during the first twenty days of the trial and significantly decreasing afterward. This may have
been due to the loss of fragmented seaweed though the 12 mm mesh or more likely an insufficiency of
nutrients in the low nutrient water which was continually passing through the seaweed pond on its way
to the evaporation basin. The Jeparit seaweed pond had no gates or valves that would permit it to hold
water and there was no control of the volume of water flow, which was brought to the surface by an
air pump and a windmill. Any periodic introduction of nutrients would be quickly washed out of the
trial pond and high volumes of water would carry dissolved iron into the seaweed pond.

40
Although precipitation of oxidized iron did reduce light penetration to the bottom, there was a
significantly higher mean growth by seaweed held under 40 cm of water than those plants floating
higher in the water column (t test: 98% confidence level).

4.2.3. Lake Charm

A one month bioassay trial using 4.1 kg of Gracilaria at the Lake Charm basin in Sept-Oct 2004
resulted in the total loss of the seaweed. An attempt was made to mitigate the low potassium
concentration in the water when the seaweed was put into the basin, by adding four perforated plastic
containers, each with two kgs of potassium nitrate. Unfortunately the perforations were insufficient to
permit the potassium nitrate to enter into solution and supplement the potassium concentration in the
water. We assumed (without further knowledge) that the problem was critical deficiency of
potassium.

In a later trial in May - June 2004, with sufficient potassium added as K2NO3, an eight kg sample of
the N variety of Gracilaria together with 5 kg of J type didn’t grow but remained viable over 12 days.
Some of the material was taken from wild stock that that appeared starved of nutrients when collected.
The seaweed was distributed in the open basin and permitted to roll along the bottom with the wind.

A repeat of this trial was made in Nov-Dec 2005 with M type Gracilaria but the results were not
favourable, the Gracilaria dying by the 17th day. An analysis of the dry material before and after the
trial showed that both potassium and copper had decreased in the seaweed to less than 8% of it’s initial
dry weight concentrations to levels that were lower than critical deficiency levels in terrestrial crops
(Reuter et al, 1997). Copper is an important co-enzyme in seaweed and acts in photosynthesis.
Phosphate also decreased to 857 ppm, close to the critical level of 700 ppm found by Lewis and
Hanisak (1996). Potassium had been added to the basin as K2SO4 to increase its concentration in the
water. During the trial calcium increased by more than ten times in the Gracilaria dry matter, and
aluminium increased four times. Some of the heavy metals increased even more: arsenic 5x, nickel
6x, lead 20x, tin 47x and strontium 62x.

These results suggest that there may be two mechanisms acting on the seaweed. On the one hand, the
deficiency of potassium and copper may have limited Gracilaria growth and on the other hand such
high levels of strontium and perhaps tin may have been toxic to the seaweed. The very high levels of
calcium may have interfered with the uptake of potassium and copper and could have carried in the
strontium, which has the same valence (+2) as calcium. Minerals are often concentrated in Gracilaria
by active transport, often to 10,000 times their ambient concentration in the sea to meet the needs of
the plants. Minerals do interfere with each other but most often when the species are similar (like
PO4= and SO4= or Fe++ and Mn++). Potassium may act to protect the plant from sodium toxicity (both
are monovalent, K+ and Na+) but it is unclear why calcium (Ca++) should concentrate so strongly.
There are calcareous red seaweeds but Gracilaria is not one of them. It is not clear whether toxicity of
a heavy metal was the cause of death of the Gracilaria or the deficiency of potassium or copper.
Malea and Haritonides (1999) showed that live Gracilaria concentrated heavy metals in a polluted
Gulf. Seasonal concentrations of Pb, Zn, Cu and Cd were higher at Lake Charm than reported by
Malea and Haritonides, suggesting that these elements were not at toxic levels in Lake Charm
seaweed. It should be noted that copper toxicity decreases by orders of magnitude when calcium
concentrations are very high, as they are here and at Donald.

It is also not clear whether the very high concentration of heavy metals (and/or calcium) occurred by
active transport and storage or were passively accumulated during and after death. Gracilaria may be
able to bind heavy metals even after death, similar to the brown seaweeds as has been claimed for
Laminara japonica as a health product (NaturoDoc, 2005) and shown in the biosorption of zinc by non
living Sargassum biomass (da Costa, et al, 2001).

4.2.4. Donald
Parallel growth experiments were run at the two sites in Donald from April to May when temperatures
were falling to 12 oC. Both sites were low in potassium, as appears to be typical in the saline

41
groundwaters of the lower Murray River plain in Victoria. Phosphates were adequate at 2 mg/l so
potassium nitrate (K2NO3) was added to the basins sufficient to raise the potassium to seawater
equivalent and provide an over-abundance of nitrogen. The growth rate of one kg of Gracilaria in one
basin was 2.5% per day over 14 days in April-May at 12oC while the other basin showed no growth at
all. None of the chemical species analysed showed any growth-limiting difference between basins that
would account for the growth rate differences, though both iron (0.10 vs. 0.03 g/l) and phosphate
(2.25 vs. 0.7 mg/l) were higher in the pond that had growth. Further trials are planned.

4.3. Agricultural technology

Warwick Noonan developed a mechanical harvester under this RIRDC grant for use in shallow saline
water basins by making innovative modifications to a high pressure fire pump for use as a low-labour-
intensive harvesting tool. High pressure water is injected into a venture system mounted beneath the
water surface. Water and seaweed are “vacuumed” into a 100 mm flexible PVC pipe and discharged
with the water to the edge of the basin and into drying racks. Water flowing through the discharge
pipe cleans the seaweed of mud and deposits a stream of clean seaweed onto the bank while water
drains back into the basin. The entire unit is mounted on floatation tanks. The harvest takes two
workers, one to vacuum the seaweed from the bottom and the second to move the discharge hose
along the bank to evenly deposit the harvest into drying racks.

Gracilaria is sold in bales dried to 15-20% moisture. Two days of sun drying with one turning of the
harvest mass is sufficient to remove the moisture required before baling like hay or wool for shipment.

Figure 5. Noonan’s venturi pump seaweed vacuum system on left and the discharge end on right

CRC for Biopolymers technicians tested a naturally grown Gracilaria chilensis from the Melbourne
area for this project and showed that the gel strength was very high (1291 g) compared to
commercially available “high gel strength agar,” which was only 1207 g. Gel setting temperature was
40oC and the melting temperature was 85-90oC. Although the yield was only 10.3%, rather low for
commercial production, “on the whole, the results are quite encouraging Ming (2001).”

42
5. Implications and Recommendations
The objective of the study was to establish an agronomy for Gracilaria seaweed, which would enable
farmers to use land and saline water currently considered degraded and non-productive. Although it
has been shown that Gracilaria can at least remain viable in some of the inland saline waters studied,
and at best can grow at 2.5% per day, the information collected by the field methods used were not
sufficient to establish commercial growth rates or to support the preparation of a full scale economic
and financial feasibility study.

These studies have demonstrated that saline groundwaters in the Murray Basin are not homogeneous
but have a wide range of chemical compositions and chemical signatures, only some of which may be
useful for saline water agriculture. This conclusion applies to waters both in salinised irrigation
regions and for fossil saline groundwaters in dryland salinity areas.

The outcomes achieved have not yet established a new paradigm that saline ground water can be used as a
productive agricultural resource to provide a new income stream for farms that have lost the ability to harvest
traditional crops from salinsed land. Pilot trials in small on-farm evaporation basins have begun to establish the
base line requirements and preliminary growth rates for the seaweeds in inland saline groundwaters. If the
methods become commercially viable, seaweed harvests could mitigate the costs of engineering investments and
operating costs for salinity rehabilitation programs as well as providing a profitable new export crop for
Australia.

Whilst commercial quantities have not yet been harvested, sufficient evidence has been collected to
support detailed seaweed physiological studies and encourage further field trials that can lead to
sustainable and profitable cropping of Gracilaria seaweed.

Preliminary field trials in three rural communities have confirmed a keen interest in seaweed farming
that would ensure wide uptake of the technologies being developed. Further interest has been
expressed by wheat farmers at Morawa, WA and preliminary assessments have shown that the saline
groundwater there can support Gracilaria cultivation. Farmers have reported that they are
comfortable with seaweed as an agricultural crop since seaweed farming methods are expected to
parallel current cropping methods and there is a large degree of flexibility in production timetabling
and post-harvest handling. The social benefits of community involvement in their seaweed projects
and the hope it engenders to keep these salt and drought-stricken communities from dying are already
evident at Donald, Lake Charm and Morawa. Once the technology and economic viability have been
proven, more than twenty farmers are ready to leap into production.

Experimental agronomic work is continuing at Lake Charm and Donald with private financing from
Seaweed Trading and small grants from Victoria DPI (Kerang) and the North Central (Victoria)
Catchment Management Authority. The Morawa Farm Improvement Group has applied for a NLP
Natural Resources Innovation Grant for seaweed cultivation trials. The outcome of the seaweed
agronomy research will be a field-tested farm model for the profitable and productive use of saline
groundwater to harvest a market-driven agar gel-bearing seaweed.

Implications for industry

As yet there is no certainty that a commercial crop of Gracilaria can be harvested from basins of
inland saline groundwater. Successful yields of this seaweed would have an international market
value dependent on the agar gel quality but should be greater than $500 per dry tonne (about 6 wet
tonnes). Market demand is currently greater than 35,000 dry tonnes and inland cultivation in Australia
will have competitive advantages over the current South American supply in both quality of product
and the cost of transport of raw material to the major Asian markets.

The process of extracting agar gel from the raw material seaweed is in the public domain and could
easily be improved upon using existing Australian technologies. Small processing plants have been

43
shown to be profitable in Thailand and Indonesia and capital investment needs not be huge to produce
a series of value added products, some of which can be import replacement for marine gels imported
into Australia and valued in 1998 at $8 million.

Implications for Communities

The profitable and productive use of saline groundwater provides a chance to recover income lost to
communities where farmland is being degraded by salinisation of soils and infrastructure. A factory to
extract gel or add value to the product can be established in small communities to increase
employment and support spin-off industries.

Implications for Policy makers

Both state and national governments have supported ideas to mitigate the damage done by inland
salinisation and saline groundwater. Were seaweed cultivation proven to have commercial potential, it
would need to be supported with:

Export marketing assistance to access European and Asian markets for agar-bearing seaweed
Assistance and a positive attitude from AQIS to import cultivars of commercially useful seaweeds that can be
shown to not be invasive pest plants.
Support from the states for industry to collect some Gracilaria material from coastal waters consistent with
biodiversity and ecosystem protection.
Protection for seaweed cultivation against revenue raising by State agencies through permits, licences and fees,
which erode investor confidence and constrain development.
A clear policy for Environmental Protection Agencies that will enable seaweed producers to use and deal with
saline groundwater disposal in a realistic and cost effective way.
Insistence that seaweed cultivation remain “agriculture” to protect against state aquaculture programs that might
restrict, limit or restrain commercial development.

Recommendations
Seaweed cultivation in inland saline groundwater still has agronomic potential despite the slow
development of its agronomy over the past four years. More controlled research in Gracilaria
physiology and nutrition would be a good investment and a synergistic approach to seaweed agronomy
is recommended, including hydro-geology, agronomy, water chemistry and soil-water interactions,
plant physiology, extension, engineering and business skills to develop technologies, agronomy and
models for profitable cropping of seaweed in saline groundwaters.

Saline groundwater resources should be mapped and quantified for each region to assess the regional
potential for seaweed cropping. The extraction and environmental control of the saline groundwater
must be carefully planned and monitored to minimise any environmental damage.

The linkage between the water resources and primary production, including analysis of water quality
and nutrient chemistry should be continued for each site, since water characteristics differ in different
aquifers and over various soil types. The concentration of dissolved minerals and their dynamics
needs to be monitored both in situ and under laboratory conditions so that sustainable seaweed growth
can be optimised in the field.

Seaweed physiology, photosynthesis and nutrition needs to be studied in the laboratory. Results of
seaweed growth trials under laboratory conditions would inform field trials.
Continued focus on field trials at salt-degraded farm sites at Donald and Lake Charm (Victoria) and
Morawa (WA) to involve farmers at the earliest stages of development.

More controlled and closely managed field experiments to establish reliable growth rates and
agronomic protocols for crop and water management that maximise both growth and gel quality of the
harvest.

44
Agricultural extension, best management practices and the translation of university research into
practical application manuals can be used to transfer technology to interested stakeholders to adopt the
farming methods.

Once Gracilaria seaweed agronomy is sufficiently advanced, a full financial and economic feasibility
study will be required to inform investors and producers about realistic infrastructure and operating
costs and the triple bottom line benefits of seaweed cultivation.

Agronomic models should consider the whole of the value chain, starting with the market demand for
raw material and calculating back to plan harvests for seaweed gel quality and yield that best meet
market needs.

45
6. Glossary
• CHAELATE- The process by which a large organic molecule entraps and holds a metal ion (c.f.),
which can become available to plants. Several metals necessary for plant life become oxidised at
the high pH (c.f.) of seawater and therefore unavailable to plants. Chaelation enables those metals
to remain available to plants in seawater.
• EPIPHYTE- a plant that lives atop another plant. An orchid is epiphytic on trees.
• EURYHALINE- able to live in a wide range of salinities.
• IN VITRO- Latin for “in glass” i.e. in a test tube or closed and controlled conditions.
• ION– Atoms of elements and compounds in an electrically charged state, either positive like K+
and Ca++ or negative like Cl- or SO4=. The charge enables them to bond with other ions, opposites
attracting, to result in an electrically neutral compound, for example K2SO4. The electrical
properties of water favours the formation of ions so that seawater is filled with ions rather than
salts.
• “NORMAL” SEAWATER - The concentration of minerals and the composition of oceanic
seawater is relatively constant in the well-mixed oceans around the globe. A Standard for salinity
was established in 1902 by an international commission so that all ocean scientists could calibrate
their instruments and work from the same baseline (Sverdrup, et al 1947). The figures are
periodically updated and now include a total analysis of all salts in a defined “normal” seawater.
Only close to land masses where there is a contribution of fresh water from rivers or drainage of
nutrients, organics, and other farm effluent, industry or domestic pollutants, is there a change in
concentration and composition from the “normal” mix of oceanic salts.
• SECCHI DISK- A black and white chequered disk about 20 cm diameter that is lowered into the
water until it disappears from sight to find the depth at which the light no longer penetrates. This
is considered to be the compensation depth at which photosynthesis no longer provides a net gain
to the plant.
• THALLUS- the “stem” of the seaweed. In Gracilaria there are no leaf blades, holdfasts or other
differentiated parts so that any part of the plants is it’s thallus. Plural is thalli

46
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