A polyphasic characterization of small-spored Alternaria from Argentina

RODRIGUEIRO, MARCELA; PATRIARCA, A.; STENGLEIN, S.; DA CRUZ CABRAL, L.; NIELSEN, K.F.

Freising

Workshop; 2016 ICFM Workshop; 2016

Accurate identification of fungal contaminants in food is critical in the development of prevention strategies at pre- and post-harvest stages. Even though Alternaria is frequently isolated from Argentinean crops, little is known on the variability and differentiation of its populations in this country. The objective of this study was the characterization through a polyphasic approach of 45 small-spored Alternaria spp. obtained from edible parts from tomato fruits, pepper fruits, wheat and blueberries, involving morphological, metabolomic and molecular analyses. The isolates were grown on Potato Carrot Agar at 25ºC under an alternating light cycle (8 h light, 16 h darkness) for 7 days. Morphological classification was achieved according to Simmons (2007). Metabolite profiling was carried out in DRYES (14 days, 25ºC) by micro-extraction with ethyl acetate-1% formic acid. Detection was performed by HPLC-MS (ESI-TOF). Amplification and sequencing of a portion of the endopolygalacturonase gene (endoPG) and subsequent Bayesian analysis using a Kimura 80 model were carried out.Morphologically, isolates were classified as A. tenuissima sp.-grp. (29/45), A. arborescens sp.-grp. (11/45), and A. alternata sp.-grp. (2/45). The 3 remaining strains exhibited intermediate characteristics among the three mentioned groups and were referred to as Alternaria sp. By secondary metabolites analysis, no compound representative of morphological groups could be detected; all isolates showed overlapping profiles. Bayesian analysis from endoPG gene yielded two clades, both containing isolates belonging to the three species-groups and the four substrates with short branch length, indicating low number of substitutions per site. Even though morphological differences were observed, and most of the isolates could be assigned to a species-group based on them, the whole set of analysis suggested that they were strongly related. No correlation between morphospecies and phylogenetic or metabolomic studies could be established. These results are in agreement with recent Alternaria organization systems, which incorporate these 3 species-groups in a single section within the genus (Alternaria sect. Alternaria).