This document characterizes Pseudomonadaceae populations in wild truffle fields over the course of one year. Soil samples were taken from two wild truffle fields every 40 days and cultured on a selective medium, revealing similar abundances of culturable Pseudomonadaceae strains over the year. Samples from non-producing trees showed similar morphologies but lower abundances. 16S rRNA gene sequencing of 30 samples showed low diversity, with most bands corresponding to Pseudomonas chlororaphis and Stenotrophomonas species.

1. Characterisation of Pseudomonadaceae populations in Tuber melanosporum
wild truffieres
Vilanova, X1*.; Sanchez, M1.; Viñas, M2. Morcillo, M. 1
1*
Micología Forestal & Aplicada S. L.
Address* Rambla Arnau de Vilanova , 6 local D 08800 Vilanova i la Geltrú (Barcelona-Spain).
E-mail* xaviervilanova@micofora.com
URL* www.micofora.com
2
Giro Joint Research Unit IRTA-UPC, IRTA, Torre Marimon, E-08140-Caldes de Montbui (Barcelona-Spain).
Abstract
In the last decade several surveys have revealed an important role of Pseudomonas genus as a Mycorrhizal
Helper Bacteria (MHB). Moreover, the bioaugmentation of environmental isolated Pseudomonas strains to the
substrate at inoculation time in different plant species throughout mycorrhization assays have been related
on the occurrence of a greater number of mycorrhiza of Tuber genus.
In this work, a selective culture medium was designed to isolate bacteria belonging to Pseudomonaceae
family from soil samples taken from two different wild truffières. The amount of culturable
Pseudomonadaceae populations was monitored using the selective medium throughout one year in different
wild truffières. A total of eight samples per monitored truffières were taken every 40 days for the two
monitored truffières. In the same area, non-truffle producing evergreen oaks were used as negative controls
being sampled simultaneously to the monitored truffières. The results revealed both a low diversity of
colonial morphology and a similar abundance of culturable Pseudomonadaceae strains on the selective-
designed agar medium throughout the one-year survey in both monitored truffières (6.44 x 104 ± 3.54 x 104
-1 -1
CFU · g and 5.71 x 104 ± 3.25 x 104 CFU · g respectively (± standard deviation of n=8). Non-producing trees
showed similar colonial morphologies to those found from the truffières, but less abundant (2.54 x 104 ±
2.54 x 104 CFU · g-1of soil (± standard deviation of n=8)).
Finally, thirty samples from both truffières and controls were characterized by means of 16SrRNA gene-based
PCR-DGGE (Polymerase Chain Reaction (PCR) with a subsequent Denaturing Gradient Gel Electrophoresis
(DGGE). We obtained a low diversity (1-4 DGGE bands) in all the studied samples (both truffières and
controls).
It is noteworthy that most of detected bands migrated mainly in two defined positions from the DGGE gel,
corresponding to a two different ribotypes belonging to Pseudomonas chlororaphis and Stenotrophomonas
spp. respectively.
KEYWORDS
Pseudomonas, MHB, Tuber melanosporum, wild truffieres.