Screening of antagonist soil bacteria to control Phytophthora cinnamomi in Montado

MARCIA DA CASTRO, S.; COLAVOLPE B.; MACHADO, H.; COSTA, A.; RUIZ, OSCAR A.; VIDEIRA E CASTRO, I.

SANTAREM

Congreso; 11º Encontro Nacional de Proteção Integrada; 2017

Escola Superior Agrária de SANTARÉM, Portugal

Mediterranean holmoak (Quercus rotundifolia Lam.) woodlands are important forest ecosystems in Iberia. In Portugal, specifically at Alentejo, these woodlands have been threatened by the environmental constraints aggravated under global climate change and under anthropogenic pressure, mainly related to inadequate practices. Under these global changes, the susceptibility to pathogens is increasing. One of the most destructive of all tree root pathogens is Phytophthora cinnamomi, a soil-borne pathogen that causes root rot and is involved in decline of Quercus suber and Q. rotundifolia in the Mediterranean region (OEPP/EPPO, 2004). Taxonomically is an Oomycetes, a cellulosic fungus-like, with glucose as the main monosaccharide source and about 25% of cellulose in the cell walls (Bartnicki-García, 1966). Natural pastures, under trees canopy layer, are typically unproductive and generally are endangered by shrub encroachment and the success of the sown pastures is frequently reduced. Moreover, often the persistence of the sown species does not exceed two years. In this context, the improvement of the pastures it is urgently needed within the management practices. A strategy for improving pastures in the Montado has been developed and is based on the establishment of biodiverse permanent pastures including selected and improved plant species in which inoculated legumes with high efficiency rhizobia have an important role (Ferreira & Castro, 2011).The commoncagriculture practice of Rhizobium inoculation is an environmental safer sourcecof nitrogen for the improvement of pastures, especially in poor soils ascMontado, increasing soil fertility and reducing soil erosion. This practice cancbe useful in conjugation of a large diversity of Rhizobium strains and ancabundance of native clovers in Portugal (Ferreira & Marques, 1992) in a search of Rhizobium antagonism to pathogenic microorganisms.Thus the main objective of this work is to find out antagonism activity in Rhizobium strains involved in the control of P. cinnamomi. Soil samples werecollected in a holm oak woodland in southeastern Portugal (Barrancos). Samples were put in sterile bags and then transported to the laboratory. For the isolation of plant-associated microorganisms 10 g each of soil were transferred separately to sterile Erlenmeyer flasks containing 90 ml of sterilized H2Od.The samples were shaken for 10 min and serial dilution were done. Trifolium subterraneum seeds were added to flasks with medium Jensen (1941) agar and 1 mL of soil solutions were incorporated to flasks. Control treatments without soil solution and with KPNO3 solution were also done. Nodules from T. subterraneum plants were then removed for the isolation of bacteria following the methodology described by Vincent (1970) and using Yeast Mannitol Agar (YMA) medium. Plates were incubated for 5 days at 27°C and after that colony forming units were visually observed and purified taking into account colony morphology. All bacterial isolates were screened for their activity against  P. cinnamomi isolated from soil samples of Montado and routinely grown on Potato Dextrose Agar (PDA) medium. Isolates were assessed in a dual culture in vitro: on PDA and YMA (Vincent, 1970), after 2 days of a previous inoculation with P. cinnamomi in plates. After bacteria incubation at 27°C during 3 days, zones of inhibition of P. cinnamomi were measured according to Berg et al. (2000). A total number of 121 autochthonous bacteria from Montado soil were isolated from nodules of T. subterraneus and we found four rhizobia bacteria with antagonist activity against P. cinnamomi. Many Rhizobium bacteria shown celulase activity what represents a protection behaviour. Apart from that many bacteria have capacity of phosphate solubilization which increases soil fertility and reduces soil erosion. This exploratory soil bacteria screening work may generate new bioinoculants with antagonistic and high efficiency nitrogen fixing activity by rhizobia as new promising solutions to improve the decline of Mediterranean holm oak.