Intraspecific diversity of Erwinia amylovora strains from northern Algeria

Abstract

Background: Fire blight, caused by Erwinia amylovora, is the most destructive bacterial disease affecting plants in the Rosaceae family, leading to significant economic losses. In Algeria, this disease has been reported since 2010. This study aimed to investigate the origin of fire blight in Algeria, in order to increase knowledge of the epidemiology of this serious disease and contribute to its management. A comprehensive characterization of 18 E. amylovora isolates recovered from northern Algeria between 2016 and 2021 to evaluate their phenotypical and genotypical diversity was conducted.

Results: Phenotypic differences, particularly in growth kinetics, virulence, and fatty acid profiles, allowed differentiation of strains into five groups, possibly indicating distinct introduction events. Genetic characterization revealed that only one strain lacked the ubiquitous plasmid pEA29, which is correlated with reduced virulence, while none harbored the pEI70 plasmid. Phylogenetic analysis using concatenated sequences of the recA, groEL, rpoS, ams, and hrpN genes grouped Algerian strains with those from a broadly prevalent clade. CRISPR genotyping identified a novel CR1 pattern and three genotypes, two of them previously unreported.

Conclusions: This study represents the first phenotypic, genetic, and phylogenetic investigation of E. amylovora strains in the region, and provides valuable information on the possible pathways of the introduction of this fire blight pathogen in northern Africa. The findings suggest one or more introduction events from a common ancestor, likely originating in northern Italy, followed by dispersal in various regions of Algeria.

Keywords: Erwinia amylovora; CRISPR; Diversity; Fatty acid profile; Fire blight; Growth kinetics; MLST; Plasmids; Virulence.

Fig. 1

Locations of the orchards of pear and apple trees in different provinces of Algeria from which the 18 Erwinia amylovora strains used in this study were isolated. Dashed areas denote locations where fire blight was previously (prior to this study) reported. The map was edited using QGIS

Fig. 2

Dendrogram of cluster analysis of fatty acid compositions of the Algerian Erwinia amylovora strains assayed in this study using Ward’s method

Fig. 3

Kinetic parameters obtained from the growth curves of the Erwinia amylovora strains tested in this study: (a) latency phase (λ); (b) maximum growth rate (μ) estimated for the exponential period; and (c) growth potential obtained by calculating the area under each curve (AUC)

Fig. 4

Symptoms induced by the Algerian tested Erwinia amylovora strains on immature pear fruits of cv. Blanquilla 7 days after inoculation. The severity index (SI) scale is shown in brackets [78]

Fig. 5

MFA analysis based on fatty acid content, virulence, growth kinetics, and biochemical characteristics of the studied Algerian strains of Erwinia amylovora. (a) Factor map illustrating the distribution of strains on the 1st and 2nd dimensions, with distinct colors representing different clusters. (b) Correlation circle plot showing the associations between the groups and the MFA dimensions. (c) Projection of the studied groups of parameters onto the first two dimensions of the MFA

Fig. 6

Phylogenetic tree of Algerian tested Erwinia amylovora strains (in bold) according to the concatenated sequences of the recA, ams, groEL, hrpN, and rpoS genes by using neighbor-joining reconstruction based on a Tamura 3-parameter model and bootstrap analysis (1000 replicates)

Fig. 7

Patterns/genotypes from CRISPR arrays CRISPR 1, CRISPR 2, and CRISPR 3 found in the Algerian Erwinia amylovora strains tested in this study (a–c), compared to the reference strain ATCC49946, Ea1/79Sm, and TS3128 (d–f) (a) The most common genotype (4, 24, 38/A a α) found in this study, also present in strain CFBP1430; (b) The new genotype (NP, 24, 38/naα) identified in strain EA54; (c) The new genotype (4, 29, 38/Aeα) identified in strain EA55; (d) genotype (1, 21, 38/Bbα) found in strain ATCC49946; (e) genotype (5, 24, 38/Daα) previously found in strain Ea1/79Sm; (f) genotype (2, 22, 38) found in the Korean strain TS3128 CRISPR spacers are represented by boxes, with the spacer positions numbered at the top of the columns. Spacers that differed from others by more than 5 nucleotides were given a specific color, while white boxes indicate the absence of the spacer within the pattern/genotype. The spacers were numbered, and the patterns or genotypes were named according to the nomenclature presented by [17, 18]