Horizontal chromosome transfer and independent evolution drive diversification in Fusarium oxysporum f. sp. fragariae.

The genes required for host-specific pathogenicity in Fusarium oxysporum can be acquired through horizontal chromosome transfer (HCT). However, it is unknown if HCT commonly contributes to the diversification of pathotypes. Using comparative genomics and pathogenicity phenotyping, we explored the role of HCT in the evolution of F. oxysporum f. sp. fragariae, the cause of Fusarium wilt of strawberry, with isolates from four continents. We observed two distinct syndromes: one included chlorosis ('yellows-fragariae') and the other did not ('wilt-fragariae'). All yellows-fragariae isolates carried a predicted pathogenicity chromosome, 'chrY-frag ', that was horizontally transferred at least four times. chrY-frag was associated with virulence on specific cultivars and encoded predicted effectors that were highly upregulated during infection. chrY-frag was not present in wilt-fragariae; isolates causing this syndrome evolved pathogenicity independently. All origins of F. oxysporum f. sp. fragariae occurred outside of the host's native range. Our data support the conclusion that HCT is widespread in F. oxysporum, but pathogenicity can also evolve independently. The absence of chrY-frag in wilt-fragariae suggests that multiple, distinct pathogenicity chromosomes can confer the same host specificity. The wild progenitors of cultivated strawberry (Fragaria × ananassa) did not co-evolve with this pathogen, yet we discovered several sources of genetic resistance.

Biological control of strawberry Fusarium wilt caused by Fusarium oxysporum f. sp. fragariae using Bacillus velezensis BS87 and RK1 formulation.

Two isolates, Bacillus sp. BS87 and RK1, selected from soil in strawberry fields in Korea, showed high levels of antagonism towards Fusarium oxysporum f. sp. fragariae in vitro. The isolates were identified as B. velezensis based on the homology of their gyrA sequences to reference strains. BS87 and RK1 were evaluated for control of Fusarium wilt in strawberries in pot trials and field trials conducted in Nonsan, Korea. In the pot trials, the optimum applied concentration of BS87 and RK1 for pre-plant root-dip application to control Fusarium wilt was 10(5) and 10(6) colony-forming units (CFU)/ml, respectively. Meanwhile, in the 2003 and 2005 field trials, the biological control efficacies of formulations of RK1 were similar to that of a conventional fungicide (copper hydroxide) when compared with a non-treated control. The RK1 formulation was also more effective than BS87 in suppressing Fusarium wilt under field conditions. Therefore, the results indicated that formulation of B.velezensis BS87 and RK1 may have potential to control Fusarium wilt in strawberries.

First Report of Dieback Caused by Lasiodiplodia theobromae in Strawberry Plants in Korea.

Dieback in strawberry (Seolhyang cultivar) was first observed during the nursery season (June to September) in the Nonsan area of Korea in the years 2012 and 2013. Initial disease symptoms included dieback on runners, as well as black rot on roots, followed by wilting and eventually blackened, necrotic discoloration in the crowns of daughter plants. A fungus isolated from the diseased roots, runners, and crowns is close to Lasiodiplodia theobromae based on morphological characteristics. Analysis of a combined dataset assembled from sequences of the internal transcribed spacer and translation elongation factor 1-alpha genes grouped nine fungal isolates with the type strain of L. theobromae. The isolates showed strong pathogenicity on strawberry cultivars Kumhyang, Seolhyang, and Akihimae, fulfilling Koch's postulates. Based on these results, the pathogen responsible for dieback on strawberry plants in Korea was identified as L. theobromae.

Cladosporium cladosporioides and C. tenuissimum Cause Blossom Blight in Strawberry in Korea.

Blossom blight in strawberry was first observed in a green house in Nonsan, Damyang, and Geochang areas of Korea, between early January to April of 2012. Disease symptoms started as a grey fungus formed on the stigma, which led to the blossom blight and eventually to black rot and necrosis of the entire flower. We isolated the fungi purely from the infected pistils and maintained them on potato dextrose agar (PDA) slants. To test Koch's postulates, we inoculated the fungi and found that all of the isolates caused disease symptoms in the flower of strawberry cultivars (Seolhyang, Maehyang, and Kumhyang). The isolates on PDA had a velvet-like appearance, and their color ranged between olivaceous-brown and smoky-grey to olive and almost black. The intercalary conidia of the isolates were elliptical to limoniform, with sizes ranging from 5.0~10.5 × 2.5~3.0 µm to 4.0~7.5 × 2.0~3.0 µm, respectively. The secondary ramoconidia of these isolates were 0- or 1-septate, with sizes ranging betweem 10.0~15.0 × 2.5~3.7 µm and 8.7~11.2 × 2.5~3.2 µm, respectively. A combined sequence analysis of the internal transcribed spacer regions, partial actin (ACT), and translation elongation factor 1-alpha (TEF) genes revealed that the strawberry isolates belonged to two groups of authentic strains, Cladosporium cladosporioides and C. tenuissimum. Based on these results, we identified the pathogens causing blossom blight in strawberries in Korea as being C. cladosporioides and C. tenuissimum.

Genetic Diversity and Pathogenicity of Cylindrocarpon destructans Isolates Obtained from Korean Panax ginseng.

We analyzed the genetic diversity of Cylindrocarpon destructans isolates obtained from Korean ginseng (i.e., Panax ginseng) roots by performing virulence tests and nuclear ribosomal gene internal transcribed spacer (ITS) and mitochondrial small subunit (mt SSU) rDNA sequence analysis. The phylogenetic relationship analysis performed using ITS DNA sequences and isolates from other hosts helped confirm that all the Korean C. destructans isolates belonged to Nectria/Neonectria radicicola complex. The results of in vivo and ex vivo virulence tests showed that the C. destructans isolates could be divided into two groups according to their distinctive difference in virulence and the genetic diversity. The highly virulent Korean isolates in pathogenicity group II (PG II), together with foreign isolates from P. ginseng and P. quinquefolius, formed a single group. The weakly virulent isolates in pathogenicity group I, together with the foreign isolates from other host plants, formed another group and exhibited a greater genetic diversity than the isolates of PG II, as confirmed by the mt SSU rDNA sequence analysis. In addition, as the weakly virulent Korean isolates were genetically very similar to the foreign isolates from other hosts, they were likely to originate from hosts other than the ginseng plants.

Taxonomic Re-evaluation of Colletotrichum gloeosporioides Isolated from Strawberry in Korea.

For the past two decades, the causal agent of anthracnose occurring on strawberry in Korea was considered Colletotrichum gloeosporioides. However, the recent molecular analysis has shown that the genus Colletotrichum has undergone many taxonomic changes with introduction of several new species. As a result, it revealed that C. gloeosporioides indeed consisted of more than 20 distinct species. Therefore, the Korean pathogen isolated from strawberry should be reclassified. The shape and size of the conidia of the pathogen were not distinctly different from those of C. gloeosporioides and C. fructicola, but it differed in shape of the appressoria. A combined sequence analysis of partial actin, glycer-aldehydes-3-phosphate dehydrogenase genes, and the internal transcribed spacer regions showed that the strawberry isolates formed a monophyletic group with authentic strains of C. fructicola. On the basis of these results, the anthracnose fungi of the domestic strawberry in Korea were identified as C. fructicola and distinguished from C. gloeosporioides.