¿ìè¶ÌÊÓƵ

SEEDS have the key to a strange world between life and death. Buried
in roadside verges, dormant poppy seeds lie in wait of some upheaval of
the soil that will expose them to light. Once at the surface, the seeds
need just a drizzle and in no time the roadsides are red with poppy flowers.
This is part of the poppy’s strategy for survival. The plants race to produce
seed while the soil is still relatively bare. By the time other plants have
established themselves and are forcing the poppies out, the poppy plants
have made, and probably shed, thousands of tiny seeds. Some germinate immediately
but others fall into deep cracks in the soil. Starved of light and oxygen,
the seeds remain dormant. They can stay in this state of suspended animation
for several years until they are exposed to the right conditions for germination.
The events that take place inside the waiting seeds keep them ticking over;
the seeds’ normal biochemistry continues, but imperceptibly slowly. Starved
of water as well, seeds enter an even deeper state of dormancy, governed
by the laws of physical chemistry. The conservation of most of the world’s
plant genetic resources relies on this metabolic sloth.

Seed physiologists found that most seeds remain viable for at least
15 years, and possibly for more than 200 years, if they are dried until
they contain only about 5 per cent water and if they are kept at -20 Degree
C. Botanists estimate that the seed of 80 per cent of the world’s flora
may, like the poppy seed, withstand such cooling and desiccation. Such seeds
are called orthodox.

The estimate of a 15-year shelf life comes from scientists at the Royal
Botanic Gardens’ gene bank at Wakehurst Place, in Sussex, which holds the
world’s largest collection of seeds of wild plants. The amount of moisture
in a seed is the single most important factor in maintaining the viability
of stored seed. ‘Orthodox seeds are hygroscopic. If you put them in an environment
of low humidity they will dry. It is only below a critical moisture content
that removing moisture improves longevity’, explained Roger Smith, head
of physiology at Kew’s Jodrell Laboratory. The second important factor is
temperature. A rule of thumb is that the lifespan of a seed doubles with
every 5 Degree C reduction in temperature or a fall in relative humidity
of 2 per cent.

The members of a species all appear to share the same response to moisture
content and temperature. There are differences between species, however.
Subjected to the same reduction of moisture content and temperature, cereal
grains live longest and the seeds of timber trees are the most short-lived.
‘But whatever you do to the environment only changes the rate at which seeds
will die,’ Smith said, ‘therefore the initial quality of your seed is very
important in terms of the length of its storage life.’

There is no way of telling if a seed is dead or dormant except by attempting
to make it germinate. You can never be sure whether you have killed the
seeds of, say, a new species of tropical legume in your attempts to break
dormancy or whether the seeds were dead to start with. Another indication
of the true state of the seed is to expose it to tetrazolium stain. This
colourless salt produces an insoluble red stain if it comes into contact
with active dehydrogenase enzymes, which are a sure sign of life. One visual
clue is that moulds tend to colonise dead seeds but seem to be repulsed
by seeds that are merely dormant. Another test rests on the tenuous principle
that if you don’t get an eyeful of rancid pulp from squeezing a seed that
has spent days on an agar plate then it is probably dormant. A nondestructive
test could take a lot of the anxiety out of trying to assess the viability
of small samples of rare seed.

A germination test estimates the initial quality of a seed lot. The
test simply counts the number of seeds that germinate from a sample of,
say, 100 seeds. If 97 seeds germinate, the sample is 97 per cent viable.
According to the standards set by the International Board for Plant Genetic
Resources (IBPGR), in Rome, keepers of gene banks should carry out germination
tests on each sample, or accession, every five years. Viability should not
drop below 85 per cent. Once the viability of the sample approaches 85 per
cent, it should be regenerated by growing the plants and collecting their
seed, in order to provide a new sample of seed for storing.

Seed banks hold the world’s future food supplies. They are more important
than money banks and more of a strategic liability than a harbour or airport.
Yet we ignore them most of the time. Curiously, they rise out of oblivion
every now and then as treasured national assets. All there is to see are
glasshouses and a glorified fridge whose temperature and relative humidity
can be preset and automatically controlled. The glasshouses are full of
plants going to seed, and the cold room is lined with drawers full of sealed
packets of seed and shelves of glass bottles and jars filled with still
more seeds. They are often understaffed and underfunded.

A good example is the Genetic Resources Unit at Wellesbourne, in Warwickshire,
which houses the Vegetable Gene Bank. This gene bank has world responsibility
for conserving certain vegetable crops, notably species of Brassica, the
cauliflowers, cabbages, sprouts and the like, and Allium, the onions, leeks,
chives and so on. Yet at Wellesbourne, the director and two assistant scientific
officers carry out the initial germination tests on samples sent in from
collecting missions, amateur gardeners, breeders and seed firms at home
and abroad. They carry out the mundane task of placing the seed in the drying
room at 15 per cent relative humidity and a temperature of 15 to 17 Degree
C. They clean the seed, separating it from the chaff and other paraphernalia
of the seed heads. Then they weigh them, and seal them into foil packets
lined with plastic. The packets are stored in drawers in a cold room at
a temperature of -20 Degree C. The scientists monitor the viability of existing
accessions by carrying out germination tests every five years as recommended.
One other assistant takes care of the 300 or so accessions identified each
year as in need of regeneration. Not surprisingly, there is always a backlog
of samples waiting to be regenerated.

So far, the monitoring at Wellesbourne has not identified any samples
that have deteriorated enough to seriously damage their chances of germinating.
Wellesbourne regenerates imported material mostly because the number of
seeds in the original sample is too low. Too small a sample carries the
risk of genetic drift. Genetic drift is a random change in the distribution
of genes in a population as the number of individuals in the population
decreases. It tends to whittle down the variability of a population and
should be avoided at all costs.

Regeneration is not straightforward. Plants have different strategies
for surviving in the wild and a seed physiologist may not always be aware
of that strategy or know how to mimic the particular environmental conditions
the plant has evolved to cope with. It can take years of frustrating experimentation
to find out how to crack a plant’s dormancy. Cereals, for example, ask only
a temperature of 10 Degree C to germinate. Less obligingly, shepherd’s purse
(Capsella bursa-pastoris) demands fluctuating temperatures, white or red
light and nitrate ions in its supply of water. The need for white or red
light, as it would receive from sunlight, suggests that shepherd’s purse
will not grow in the shade of other plants and will remain dormant until
exposed to direct sunlight. The seeds of the Rosaceae, apple for example,
have evolved to germinate only once winter has passed. They need a long
chill to coax them out of dormancy. Many species of wild plants have hard
seed coats. An incision can sometimes help them to take up moisture and
germinate.

Regenerating exotic species of plants is unsatisfactory but is sometimes
essential. It is unsatisfactory because it is better to grow the plants
in the same sort of climate and conditions as those in which they originated.
This would increase the likelihood that the genetic diversity in the regenerated
sample is the same as in the original. However, regeneration is most often
done in the country that has the gene bank, even though the seed may be
tropical and the gene bank may be in a temperate zone. ¿ìè¶ÌÊÓƵs can, however,
manipulate the temperature, humidity and lighting in glasshouses to mimic
the conditions in the country of origin.

In future, regenerating exotic material may be less of a problem. In
1985 the International Union for Conservation of Nature and Natural Resources
(IUCN) and the World Wide Fund for Nature (WWF) set up the Botanic Gardens
Conservation Secretariat, which aims to expand the role of botanic gardens
in conserving threatened species. Today the secretariat has more than 200
members, including gardens in 12 countries of Latin America, seven countries
in Africa and eight in Asia.

The secretariat holds meetings and workshops on conservation and maintains
a database on species conserved in botanic gardens around the world. According
to Vernon Heywood, the director of the secretariat, many of the species
conserved in botanic gardens are on the verge of extinction and botanic
gardens can play a crucial role in reintroducing them to the wild. ‘In 5
to 10 years we hope to have samples of half the world’s endangered species
in protective custody in botanic gardens,’ Heywood said. Perhaps by that
time botanic gardens will be able to take on more regeneration work.

The seeds that won’t conform

Gene banks can take care of most of what needs to be conserved. But
20 per cent or more of seeds are recalcitrant, that is, they cannot survive
desiccation. Many plants that have recalcitrant seeds are of economic importance;
they include mango, rubber, chestnut, oak, avocado, cacao, coconut and durian.
Recalcitrant seeds must be stored either in situ , in field gene banks such
as plantations, orchards and arboreta or in vitro, as tissue cultures. The
best solution would probably be to conserve in as many ways as possible.

Field gene banks are currently the safest solution to the problem of
conserving species with recalcitrant seeds. They are also the best method
of conserving the genetic material, or germ plasm, of species that produce
sterile seed or reproduce vegetatively. Examples are cassava, potato, yam,
sweet potato and sugar cane. Field gene banks usually hold cultivars or
breeding lines. Because the aim is to save as much diversity as possible,
ex situ gene banks will also hold collections of seed for these species.
Although potato seed, for example, is heterogeneous, and therefore of no
immediate use to breeders who need lines that will breed true, the seeds
of one plant will contain much more variability than a tuber and so are
a valuable addition to ex situ gene banks.

One problem with field gene banks is that they are labour-intensive
and demand large areas of land. The world potato collection at the International
Potato Centre (CIP) in Peru, for example, currently holds 4100 different
clones of potato all of which have to be planted annually. One researcher
estimates that conserving just 0.1 per cent of the estimated three million
almond trees in Turkey would mean planting 3000 almond trees on about 15
hectares of land. The development of techniques to grow tissue cultures
of plants may help to overcome some of these problems in the future.

Although it is possible to grow plants from protoplasts – plant cells
denuded of their cell walls – and from callus, or wound tissue, biotechnologists
prefer to use shoots and embryos for tissue culture. The reason is that
shoots and embryos are more organised structures and are more stable genetically.
There are two ways of conserving tissue cultures; the first is called slow
growth and the second is cryopreservation.

Slow growth relies on the fact that plants grow more slowly at abnormally
low temperatures and that certain compounds, such as the sugars mannitol
and sorbitol, inhibit the rate at which plantlets, growing on nutrient agar,
will absorb nutrients. The result is that the plantlets may need to be subcultured
only once a year. The main problem with this technique is that, at best,
it can offer only medium-term storage because the plantlets show varying
degrees of genetic instability. The technique is also labour-intensive and
expensive. Despite its drawbacks, the CIP has 3400 clones of its potato
collection in vitro.

Cryopreservation, which freezes plant tissue in liquid nitrogen, holds
more promise for conservationists. The technique, yet to be perfected, suspends
the metabolism of the plant tissue so that all the changes that happen with
time in a gene bank are suspended. ‘Time stands still for cryopreserved
tissues,’ said Lyndsey Withers of the IBPGR. ‘This means we can offer a
very high degree of stability.’ Cryopreservation is also a much cheaper
way to preserve material than by the slow growth method.

Withers and her colleagues at the IBPGR would like to be able to cryopreserve
crops such as coconut and banana. So far coconut embryos have proved resistant
to cryopreservation. Coconut embryos are large, about one centimetre long
and with the diameter of a pencil. They are composed of hundreds of millions
of cells all dividing according to an internal programme. Freezing disrupts
that programme and cells die. Callus, or wound tissue, forms when the embryos
thaw. Callus tissue is notoriously prone to genetic instability.

The IBPGR has at least managed to lighten the burden of the coconut
collector. It has devised a method of collecting coconut embryos and the
technique has been successfully tested in the Ivory Coast . ‘Ideally we’d
like to clone the material and cryopreserve it. At the moment, if we freeze
a coconut embryo and it dies, we’ve lost 100 per cent of the collected material,
100 per cent of the collector’s efforts have been wasted. But if we could
clone that embryo and, say, get 10 or 100 clonal replicates of it, we’d
have a lot more experimental material to store and we’d have a lot more
material to distribute to users,’ said Withers.

Recalcitrant seeds are the biggest challenge today. Conservationists
are confident that now they have successfully cloned oil palm embryos, it
is only a matter of time before those of recalcitrant seeds such as the
coconut succumb to their efforts.

* * *

Take it or leave it, gene banks versus conservation in the wild

ALTHOUGH everyone agrees that we have no choice but to conserve as much
plant genetic material as we can, as fast as we can, the use of gene banks
raises serious questions about our agricultural development. We have backed
ourselves into a corner as far as the genetic diversity of our crop plants
is concerned.

Our strategy of breeding improved varieties with narrow genetic backgrounds
and promoting their use globally, either directly by technology transfer,
or indirectly by economic pressure, forces us to rely more and more on gene
banks to safeguard genetic diversity. It is the same as saying ‘We no longer
need evolution. We have, inside our gene banks, enough genetic diversity
for now and forever.’ We have placed in the hands of scientists the responsibility
that nature once had for ensuring the evolutionary development of our food
crops.

Some people suggest that crops should be conserved in their natural
environment as well as in gene banks. The idea is to allow crops to evolve
alongside their wild relatives, in much the same way as they have since
Neolithic times. The suggestion is fundamentally flawed, however. The only
way that such a system could work would be to prevent communities from developing
their agricultural systems. They would have to remain stationary, using
only those varieties they have always used and capitalising on whatever
nature throws them. They would become anachronistic curiosities.

It would not be enough merely to pay farmers to grow traditional varieties
instead of improved modern ones. Evolutionary forces do not confine their
actions to fit neatly with the growing seasons of particular crops. Conservation
in situ makes sense only when the management of the whole ecosystem is left
to nature.

Conserving durians is a good example. The durian is an excellent candidate
for in situ conservation, not least because the fruit is such big business
in Southeast Asia, yet its continued survival is by no means secure. Durian
flowers are pollinated almost entirely by one species of fruit bat that
lives in the limestone caves of West Malaysia. As David Lee points out in
his book The Sinking Ark, if Malaysians wish to continue enjoying the fruits
of the durian tree they may well have to stop quarrying the caves where
the fruit bat roosts.

The Soviet Union has established reserves in the Caucasus mountains
to protect wild relatives of wheat and fruit trees and in the Kopet mountains,
north of the Iranian border, to protect wild pistachio, apricot and almond.
India’s first reserve is for wild relatives of citrus in the Garo Hills.
Influenced by the IUCN and WWF, many countries are now looking at existing
nature reserves as in situ gene banks for threatened and rare species. By
1988, there were more than 3500 protected areas in 125 countries and covering
an area of more than four million hectares.

Some information on what is held in those reserves is kept on a database
maintained by the World Conservation Monitoring Centre (WCMC), run by IUCN,
the United Nations Environment Programme and the WWF, based in Britain.
The WCMC’s Threatened Plants Unit has limited data on 51 000 species of
plants, of which about 19 000 are threatened. The information lists species,
which protected areas hold them, their world status and the sources that
provided the data. The database will become more useful as more studies
are carried out. Such studies would ideally be ‘ecogeographical’, looking
at the geographical distribution and ecological preferences of the plants.
Field surveys would add information on diseases, pests, resistances and
tolerances to environmental conditions. Laboratory studies of samples would
separate genetic variation within the species from environmental factors.

‘Biosphere reserves’ that conserve genetic resources and representative
examples of the world’s ecosystem types already exist. Since 1974, 269 protected
areas in 70 countries have been designated as biosphere reserves and are
part of UNESCO’s Man and Biosphere Programme. ¿ìè¶ÌÊÓƵs, the seeds industry
and conservationists reached an agreement last year at the Keystone International
Dialogue on Plant Genetic Resources, that a varied approach to conserving
diversity is the best one. Conservation in gene banks and conservation in
situ complement each other and should no longer compete.

* * *

Pick ‘n’ mix – the art of collection

THE AIM of any collector of germ plasm must be to gather as great a
range of genetic diversity as possible. The immediate problem is how to
judge genetic diversity merely by appearance, an impossible task except
in certain vegetatively propagated crops, such as the potato, where the
appearance of the tubers can give a good indication of range of genetic
variability that is available in a farmer’s store.

For crops propagated by seed, collectors aim to sample a field randomly,
collecting seed every few paces. Brian Ford-Lloyd and Michael Jackson, of
the University of Birmingham, recommend that selective samples are taken
as well. Collectors have to be alert, keeping their eyes open for interesting
characteristics, for example, an ear of corn that comes from an abnormally
tall or short plant or has a slightly different colour. If collectors are
after forage grasses they would do well to sample areas where animals water
because they might harbour varieties that stand up to trampling or tolerate
high concentrations of nitrates.

Other considerations must be climate, topography and aspect. Collectors
will obviously choose to visit areas in which the vegetation is threatened,
for example, because of changing land use, such as the conversion of agricultural
land to pasture. A recent IBPGR mission collected legumes and forages from
an area in Anatolia to be flooded by the reservoir of a proposed hydroelectric
dam.

Certain crops are easier to collect than others. The large fleshy tropical
fruits are particularly troublesome. Coconuts are clearly a collector’s
nightmare. They are bulky, fragile and have recalcitrant seeds. IBPGR, in
collaboration with a French agency called IRHO, has developed an easier
method of collecting coconuts. Collectors simply de-husk the coconut and
drill out a cylinder of endosperm containing the embryo. A sterilising agent
removes any bacteria and fungi that might have attached themselves to the
surface of the cylinder of flesh. The collector then places the embryo into
a glass container of sterile water or salts. Back at the laboratory they
can remove the embryos, culture them and plant them out into the field gene
bank.

Further reading Plant Genetic Resources: an introduction to their conservation
and use by Brian Ford-Lloyd and Michael Jackson, Edward Arnold 1986. See
also ‘The shrinking gene pool’, ¿ìè¶ÌÊÓƵ, 29 July.