Innovative timber tracking using genetic and isotopic fingerprints

Innovative timber tracking
using genetic and isotopic fingerprints
A demonstration project coordinated by Deutsche Gesellschaft für Internationale Zusammenarbeit (GIZ) GmbH
with its partners
U n i ve rs i tät H a m b u rg
kindly supported by the European Commission
European Commission
Fingerprinting - An innovative system
to verify the declared origin of timber
An increasing number of initiatives and legal acts aim to promote international trade in timber from
legally harvested forests and to exclude illegally harvested timber from the market:
Illegal logging takes place when timber is harvested in violation of national laws. So the knowledge about the origin of timber, i.e. its place of
harvest, is the first and indispensable step to identify the concrete circumstances of its harvest, such
as compliance with existing and locally relevant
laws and regulations and with voluntary certification standards (if applicable). To verify the origin of
timber a reliable tracking-system must be in place.
Existing timber tracking systems use
The EU recently has adopted the EU-illegal
timber regulation which comes into force in
March 2013. It prohibits the trade of illegally
sourced timber and obliges operators to seek
sufficient guarantees that the timber products they sell have been harvested according
to the relevant laws of the country of origin.
The EU is negotiating and concluding
EU-Voluntary Partnership Agreements
with individual timber-producing countries.
VPA-countries agree that only verified legal
timber with a FLEGT-license is exported to
the EU whereby the EU supports the
country inter alia in establishing Legality
Assurance Systems.
The US Lacey Act prohibits all trade in the
USA in plants and plant products including
timber and timber products that are illegally
sourced from any U.S. state or foreign country. It requires importers to declare the country of origin of harvest and species name of
all plants contained in their products.
The Convention on International Trade of
Endangered Species (CITES) regulates the
international trade in animal and plant species that are listed in one of three appendices.
Appendix II allows the trade in wild and “artificially propagated” species for commercial
and non-commercial purposes. In order to
bring these species under effective control,
look-alike species also have to be controlled.
Competent authorities need to identify species and sometimes need additional information on the origin to distinguish e.g. between
timber from a plantation and from natural
forests.
Some countries have established export bans
for raw or processed timber. Also the UN
sometimes imposes export bans for timber
which revenues are used to finance armed
conflicts. Exporting timber from such countries constitutes a legal offence which is illegal
trade of timber.
paper-based documentation of timber origin and
externally applied marks (e.g. ink, metal bands,
tags, barcodes) at the place of harvest. They bear
the risk of fraud as they can be applied to any piece
of timber, e.g. from illicit sources, and they are removed or changed in the course of processing timber. So they do not provide for a check of origin independent from the documentation system along
the full chain of custody. This is also true for certified timber.
To receive tamper-proof information about the
true origin of timber, systems are required that
use marks which are not externally applied but
are inherent in the wood itself and, thus, cannot
be manipulated externally. They should also be directly linked to the environment of the place of harvest to allow for an independent and reliable trackback of origin. Recent research proofed that such
inherent marks are the genetical and chemical characteristics of timber, the so called “genetic and
(stable) isotope fingerprints”. They can be used totally independent at any point of the chain of custody to verify if the declared origin or wood species
is right or not.
I Genetic fingerprinting
Trees differ genetically between species, populations
at different places and individuals. Some of the regions in the genome, especially at so called “microsatellites”, are highly variable and show a unique
pattern for each individual (genetic fingerprint).
The genetic composition of tree populations shows
a spatial pattern which is caused by extinction and
recolonisation in the past glacial times and by spatial limited pollen and seed dispersal.
fically planned sampling of plants. These samples
are screened for the genetic fingerprints and build
the reference data that are used for the timber
tracking. Here the timber origin can be controlled
by comparing the genotypes of wood samples with
the genotypes observed in sampled populations.
The comparison of a genotyped timber probe with
the reference data shows if the declared origin is
correct.
This spatial genetic pattern is the basis to control
the geographic origin of trees and the traded timber. The spatial genetic pattern is visible by a correlation of genetic differences and spatial distances
between tree populations. For a given landscape the
genetic pattern can be identified based on a scienti-
A small part of tree leaves or a small piece of cambium taken from the trunk serves as material for the
DNA extraction. This sampling does not cause any
severe damage to the trees. Together with the
sample the correct location of the sampled tree
(GPS-data) must be taken.
The statistical power of the timber tracking with
fingerprints depends on the number of sampled
populations and sampled trees per population as
well as on the genetic and isotopic variation. The
sampling needs to be adjusted to the requested
spatial resolution e.g. whether the control should
check the country of origin or concession of origin.
Both methods (genetic and isotopic fingerprinting)
can make a statement, if any piece of timber stems
from the declared origin which reference is known
or if it does not stem from there. This statement
can only be made if it is based on authentic refer-
Part of a punched-out bark: In the sample
a layer of cambium is visible. It will now
be dried in Silica-Gel for further analysis.
ence material from the place of interest. The reliability of the statement should be checked in advance through a so called “blind test”. In such a test
the scientists / laboratories that draw the reference
map get pieces of timber which were sampled by an
independent third party. The true origin is not
communicated to the scientists but is only known
to the third party. The scientists shall make a statement if or if not the piece of timber stems from the
place referenced or not. If the scientists have a success rate of close to 95% the blind test is said to be
successful and the reference map is assumed to be
reliable.
II Isotopic fingerprinting
Any organic material on earth is composed of chemical elements, especially Hydrogen, Oxygen, Nitrogen and Sulphur. Stable isotopes of atoms are natural variants which are not radioactive. They only
differ in their specific masses. Due to their different
weights they react a bit differently (are fractionated), illustrated by a different velocity of their molecules in the gaseous phase. Analytically they can be
measured in mass spectrometers, and natural variations can be observed.
Since the biomass is made up of the material in the
surrounding environment, the environment’s isotope composition is reflected by the biomass, i.e.
a product coming from a certain place may show
a significantly distinguishable isotope composition
when compared with the same product from another place. Hence, any biomass, and therefore e.g.
any timber item, exhibits a – non radioactive – natural tag, the so called “isotope fingerprint”. So,
plants take up different chemical elements through
water (Hydrogen, Oxygen), nutrition from soil
(Sulphur, Strontium, Nitrogen) and by photosynthesis (Carbon, Oxygen). The distribution of isotopes shows different patterns (following e.g. rain
fall patterns or geological formations). Different
elements are used to distinguish the origin on different regional scales:
The best known pattern is the fractionation of Hydrogen and Oxygen in the global water cycle. The
ratios of Hydrogen and Oxygen isotopes are used
for wider regions, for presently declarations consider the national level only.
The ratios of Carbon isotopes as a climate and
Strontium isotopes as a geological parameter differ
on regional level.
For the local level Sulphur and Nitrogen isotopes
and their ratios can be used because they reflect
geological / soil identities.
Through the combination of elements it is possible
to check the declaration of the origin of a product.
The method is commercially applied in the food
sector, it is used e.g. by big retailers to control their
food suppliers and is already accepted by court. The
European Community and the International Office
of Vine and Wine have officially accepted the stable
isotope method to control wines.
The costs of the timber tracking based on fingerprinting are estimated to be equal or below the
costs of other tracking systems (e.g. tag based
Helvetia timber tracking) and include fixed costs
for the elaboration of the reference data and variable costs for the genetic and isotopic analysis of
control probes along the chain of custody. The costs
for the creation of the reference data are quite different among species. This depends on the sample
area and the abundance of the tree species.
Case Study: GIZ-coordinated project in Cameroon –
application on concession level
An EU-co-funded project to demonstrate the applicability of these techniques was implemented by GIZ,
TFT – The Forest Trust TFT (facilitation of sampling),
the University of Hamburg / Johann Heinrich von
Thünen-Institute (vTI) – Institute of Forest Genetics
(genetic analyses) and the TÜV Rhineland / Agroisolab
(isotopic analyses).
It was conducted in South-East Cameroon for two timber species: Sapelli (Entandrophragma cylindricum) and
Iroko (Milicia excelsia). In the target concession (1,937
square kilometers = 193,700 hectares) a total of 5 populations of Sapelli and 1 Iroko population were sampled.
Additionally, samples of Sapelli and of Iroko were taken
outside the target concession from other concessions
within Cameroon to identify the spatial pattern.
In the concession there was a huge difference for the
abundance / density of Sapelli and Iroko: Sapelli is very
abundant whereas Iroko is very rare and occurred in
aggregates (clumps). Due to the height of the tress leaf
material was not available so all samples were taken
from the Cambium of the trunks and stored in SilicaGel. The scientists were able to identify microsatellites
and DNA sequences of the chloroplast that reveal a spatial pattern between the target concession and outside
that area. Also for the isotopic fingerprint the scientists
were able to identify spatial patterns using Hydrogen,
Oxygen, Carbon, Nitrogen, Sulphur and Strontium
isotopes.
A blind test was conducted for both species. Scientists
received samples without knowing the species and
origin of the timber. They were asked to identify if the
samples stem from the target concession or not. They
succeeded with 94% of the samples: 15 of 16 samples
were attached correctly by one or both techniques.
For Iroko only two different types of blind samples
were available: from very distant (Ivory Coast), this
sample was identified by the scientists as definitely not
deriving from the target concession. The other blind
samples stemmed from a concession area less than 100
km distant from the target concession. These were at
the few screened gene markers identical with results of
the target concession; the isotopic analyses excluded
them as originating from the concession.
Sapelli
(Entandrophragma
cylindricum)
Success rate of 94%
For Sapelli seven different blind test samples were available. For six of them the techniques truly identified if
the origin was the target concession (2 samples) or not
(4 samples). It is worth noting that one of these samples
was not a Sapelli sample but from Sipo timber which
is closely related with Sapelli and contained the same
microsatellite. Only one sample that stemmed from the
target concession was identified wrongly as having a
different origin.
The results for Iroko show that for the distinction on
regional level the existing markers can be used. For the
distinction on local level (less than 100 kilometers) additional markers are needed for a reliable distinction.
For Sapelli the results show that the genetic markers
identified can be applied for the distinction on all tested levels. The isotopic analyses showed good results.
Nevertheless, none of the techniques can be preferred
since both do not yet deliver results which are 100%
reliable. However, in combination they had a success
rate of 94%.
To present and discuss the results of this and other projects implemented by WWF Germany for Teak and
Mahogany an international conference “Genetic and
isotopic fingerprinting methods – practical tools to verify the declared origin of wood” was held in Eschborn/
Germany in November 2010 organised by GIZ – Sector Programme International Forest Policy (GIZ-IWP)
and WWF Germany. 69 participants from 19 countries
representing FLEGT- and CITES authorities, private
sector, science, certification organisations and consultancy gave a very positive feedback. The discussions
about the application spectrum, overlapping and extension to regulations like EU illegal timber regulation but
also on the development status of the methods have
been vivid and fruitful. Recommendations were formulated for practical application in producer and consumer countries and for future scientific research.
Related activities and next steps
A new activity is financed by ITTO and the German
government to further expand fingerprinting techniques on regional level. The ITTO-project “Development and implementation of a species identification and timber tracking system with DNA fingerprints and stable isotopes in Africa” will conduct further studies in the Congo Basin and Kenya as an important timber transit country. The expected outputs
of the final project are: (a) a timber tracking system
with DNA and stable isotopes working for 5 important timber species in Africa, (b) reference databases
on genetic and isotopic spatial patterns ready for
control uses, and (c) facilities for DNA-fingerprinting and stable isotopes with trained staff in timber
producer and timber consumer countries.
A new international facility “Identification of Timber Species and Origins” was recently established at
Bioversity International in Malaysia. Objectives are to
coordinate work on research, standard setting and to
establish an international open access database inter
alia through networking among research and implementing institutes. An international database is a
sensitive but important tool to further promote the
up-scaling of fingerprinting techniques. Lessons
learned from similar databases in the food sector
should be taken into account. Cooperation with
existing databases (GENEBANK, Barcode of Life)
could be sought. Private companies and competent
authorities from producer countries are invited to
support the work of the facility and the use of the
database by opening access to timber samples and
facilitating technical and scientific exchange. The
facility is supported by the German government
through the German Federal Ministry for Food,
Agriculture and Consumer Protection (BMELV).
Additional contact information
For FLEGT-activities at GIZ Sector Programme International Forest Policy (IWP):
Mr Herbert Christ ([email protected]), Dr Stefanie von Scheliha ([email protected])
For genetic analyses at vTI, the ITTO-project and the new international facility:
PD Dr Bernd Degen ([email protected])
For isotopic analyses at TÜV Rhineland / Agroisolab:
Dr Markus Boner ([email protected])
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