Application of fungal communitiesfinger printings to determine the geographic origin of tropical timbers

Application of Fungal Communities Fingerprintings

to Determine the Geographic Origin of Tropical Timbers

Material and methods

Contacts: zaremski@cirad.fr, gaetan.lefloch@univ-brest.fr - IMC9 – The Bioloy of Fungi Edinburgh, UK – August 1-6, 2010

Table 1: Tropical species and origins of timber used in this study:

Limba (Terminalia superba), Limbali (Gilbertiodendron dewevrei) and Teak (Tectona grandis), from six different countries.

Introduction

from forest to its postmarketing use, tropical timber is subjected to colonization by many fungi (endophytes, wood rotting fungi, ...). Traceability for timber production through the world is a key resource for buyers looking to secure timber of verifiable origin from certified suppliers. The objective is to use the fungal community fingerprints as microbiological marker to

Wood Geographical origin Code

Limba

Fraké Central African Republic L RCA

Benin L Be

Côte d'Ivoire L Ci

Congo L Co

Cameroon L Ca

Limbali Central African Republic Li RCA (1 and 2)

Vaa _{Côte d'Ivoire Li Ci}

Cameroon Li Ca

Teck Polynesia T Pol

Côte d'Ivoire T Ci

Multivariate analysis seems to distinguish samples by wood type which are colonized by specific fungal communities. For origin determination using fungal communities, a focus on dominant taxa is needed. Further, it would be preferable to achieve cloning and sequencing of amplicons to determine the name of fungal taxa prior to use more accurate and informative fingerprint tool like D-HPLC (Maurice et al., 2010).

Results

Conclusions

Figure 2: CE-SSCP analysis of PCR fungal amplicons (ITS1 rDNA) of Limba from distincts origins.

A peak represents an Operational Taxonomic Unit (OTU). The y-axis value denotes the fluorescence intensity of each OTU which was used as quantitative data for the multivariate analysis (Principal Components Analysis).

References:

Maurice et al., 2010 Profiling fungal community in wood decay ecosystem by Denaturing

High-Performance Liquid Chromatography, 9–13 mai 2010, 41st International Research Group conference on Wood Protection. Biarritz, France.

White et al., 1990 Amplification and sequencing of fungal ribosomal RNA genes for

phylogenetics. pp.315–322.: PCR protocols: a guide to methods and applications, Academic Press, Inc., New York, N.Y.

Figure 3: PCA results extracted from CE-SSCP profiles of

fungal community from tropical timber. Only the first principal component was shown. The % variance explained by each component is given in parenthesis. Score plot showing the relations between samples. Communities seem to be structured according to the wood: Limba left of the F1 axis and Limbali to its right.

Samples (axis F1 et F2: 70 %) axis F2 (14 %) axis F1 (56 %) 15 10 5 0 -5 -10 -30 -25 -20 -15 -10 -5 0 5 10 15 20 Li RCA 2 _{Li Ci} Li RCA 1 L Ci L Be _{L Co} L RCA T Ci Li Ca L Ca T Pol

Figure 1: Technical procedure to profiling fungal biodiversity in tropical timber (adapted from

Maurice et al., 2010)

DNA Extraction (Fast DNA Spin Kit, MP biomedicals) DNA Quantitation (Nanodrop, Thermo)

Fungal ITS1 amplification (ITS1 and ITS2-6FAM, White et al., 1990)

Amplicons separation by Capillary Electrophoresis -Single Strand Conformation Polymorphism (CE-SSCP)

Electrophoregram alignment and comparison (Gene Mapper, Applied Biosystems) Multivariate Analysis (Principal Components Analysis)

Number of base pair

Flu or escencec in tensi ty L RCA L Ca L Ci L Be L Co

identify the geographical origin of wood imported from tropical countries. Many molecular tools have common approaches to understanding the genetic variation within microbial communities. Among these techniques, the CE-SSCP approach has been applied in order to profile fungal biodiversity colonizing tropical timber.

2

Université Européenne de Bretagne, France 3

Université de Brest, EA3882 Laboratoire Universitaire de Biodiversité et Ecologie Microbienne, IFR148 ScInBioS, ESMISAB, Technopôle de

Brest-Iroise, 29280, Plouzané, France 1_{Centre de Coopération Internationale en Recherche Agronomique}

pour le Développement, CIRAD, UPR 39 Génétique forestière, TA A-39/C, 34398 Montpellier Cedex 5, France

Alba Zaremski

, Sandy Maurice

, Renaud Henry

, Gaetan Le Floch

and Georges Barbier

AGRICULTURAL RESEARCH FOR DEVELOPMENT Conc ep tion et réali sation : Ci rad , Martin e Dupo rta l , juil le t 2 01 0