Development of New Applications for 'Rhizoid Regeneration Technique'
for use in Monitoring Pesticides and Herbicides in Vineyards
Jane Shields developed a 'Rhizoid Regeneration Technique' during post-
graduate research at Heriot-Watt University, Edinburgh. The aim
of the original research was to develop an effective treatment system for
the marine environment using algae. However, there was no effective
method for assessment of the contaminants in the water without using marine
animals in a LD50.test, (the concentration of a
substance required to kill 50% of the organisms). This is
a crude measurement and Jane wanted to find an alternative to animal experimentation.
Methods with multicellular algae used dead material or staining, both ineffective
and crude. All the available methods focused on the concentrations required
to kill a certain type of organism. Her thesis became the
development of this technique.
Algae form the first link in many food webs, oxygenate the
water and are important in the cycling of dissolved organic and inorganic
substances. Toxicants accumulated by algae may be passed up the whole
food web, and if stored rather than metabolised may be concentrated
and have effects at higher trophic levels.
For this reason the focus was on the development of a sensitive
technique that could detect the first signs of stress in algal populations
which required the algae to remain alive. Another difficulty to overcome
was the variability and steepness of physio-chemical gradients in estuarine
water, the continuous and frequently unpredictable oscillations of salinity and dissolved
and particulate matter in the water. It was also necessary for
the technique to not be affected by reproduction.
The genus, Enteromorpha (Chlorophyta, Ulvales), a green tubular
parenchymatous alga was chosen as it has a wide geographical distribution (
both northern and southern hemispheres, well know nutrient requirements, small
genetic and phenotypic variation, good taxonomic characterisation of the strains,
a high growth rate and ease of handling. The habitat ranges
from fresh water to seawater but it is more common in estuarine and coastal habitats.
One of the most precarious stages in the life history of a
marine benthic organism is the colonisation of new substrata, for which
highly specialised processes of settlement and early growth have developed. In
Enteromorpha, a primary rhizoid develops which penetrates a substrate on structures
being colonised and other rhizoids soon follow the point of entry.
Secondary rhizoid regeneration occurs in nature when the alga is removed from
the substrate by either predation or wave action and must reattach.
The technique is based on the secondary rhizoid regeneration process that can
be simulated by cutting 5 mm off the basal end of the
algal filament which removes the specialised cells that grow rhizoids. This
technique has proved to be extremely sensitive as the algal filiments must
be in good health in order to regenerate the specialised cells and
produce rhizoids. Assessment is based on the number of algal filaments
that have regenerated in a given substance or natural waters and the
density and number of the rhizoids which is a finer indicator of
the degree of stress on the organism.
This technique has been used to assess water quality along an estuary
by studying different algal populations located in different parts of an estuary
in combination with testing samples along an estuary tested against a base
population used as a control, using standard statistical analysis. It
has also been shown to be effective in testing particular contaminants at
known concentrations, e.g. copper, biocide and salinity.
ITV in France has asked Jane Shields to see if the rhizoid
regeneration technique can be developed to monitor pesticides and herbicides. There
are hundreds of pesticides and herbicides in use but only a few
of these have a test that can analyse it so it is
virtually impossible to understand the full impact of these contaminants in the
soil or water. Also most are used in mixtures making analysis even more difficult.
We are starting to work on the development of the rhizoid regeneration
test for this purpose. Initial funding is expected from ITV.