Interactions between twenty-two Arabidopsis thaliana accessions and

Xanthomonas campestris strains

Phenotypic variation was observed in interactions between 22 A. thaliana accessions (core collection, Table 4) and each type strain of Xcr races 1, 2 and 3 (HRI6490, 8305, 6519), Xcc races 1, 4 and 6 (HRI3811, 1279A, and 6181) and the X. campestris strain HRI8506. The maximum and the minimum (or unique) phenotype scores observed among four plant replicates within each combination of A. thaliana accession, bacterial strain and inoculation method used, are presented in Table 6.

The interaction phenotypes (IPs) were characterized according to the symptoms observed using different inoculation methods for different pathogens: spraying for interactions with Xcr strains and wounding for interactions with Xcc strains and the X. campestris strain HRI8506 (which produced symptoms identical to those caused by the Xcc strains). Spray inoculations with Xcr strains allowed reproducing typical leaf spot symptoms. Moreover, the spray method allowed to obtain more reproducible results than the wound method as some accessions exhibited leaf spot symptoms in all plant replicates tested by spray inoculation, but did not show symptoms in all wound-inoculated plants. Wound-inoculations with Xcc strains and strain HRI8506, allowed to easily identify each inoculated leaf and detect the vascular symptoms starting from the wounding sites. In sprayed plants with the Xcc race 6 strain and strain HRI8506, it was difficult to distinguish between disease symptoms and natural senescence of older leaves, particularly in the accessions Bur-0, Can-0 and Ct-1.

Table 6. Interaction phenotypes of twenty-two Arabidopsis thaliana accessions and seven Xanthomonas campestris strains a

Abbreviations: HRI, formerly Horticulture Research International, now University of Warwick Crop Centre, Wellesbourne, UK; na, not available; r, race; s, spray inoculation; w, wound inoculation; Xc, Xanthomonascampestris; Xcc, X.campestris pv. campestris; Xcr, X. campestris pv. raphani.

a _{The interaction phenotypes (IPs) are presented as follows: maximum and minimum scores of four} plants examined in two experiment repeats, with each of the two inoculation methods, spray (s) and wound (w), 10 or 14 days after inoculation with Xcr or Xcc strains and Xc strain HRI8506, respectively; IPs were scored using a six-point scale (see Figures 4 to 7). IPs were characterized according to the results obtained by spray-inoculation for Xcr strains and wound-inoculation for Xcc strains and Xc strain HRI8506, as follows: incompatible (resistance; highlighted in green) when no symptoms were observed in any plant tested (IP 0 to 1); compatible (susceptibility; highlighted in yellow) when all plants tested showed typical disease symptoms (IP 3 to 5); inconclusive when, either at least one plant showed lesions restricted to a small area of the leaf (IP 2), or susceptible and resistant phenotypes were observed among individual plants (not highlighted).

b _{X. campestris strain from Mauritius reported to cause leaf spot and blight symptoms in cabbage (R.} Lobin, pers. comm.).

3811 (r1) 1279A (r4)

s w s w s w w w s w s w

Incompatible interaction phenotypes with all Xcr strains (resistance)

Col-0 0 0-1 0-1 0-1 0-1 0-1 0 0-1 1-5 3-4 0-1 0-1

Col-5 0 0-1 0-1 0-1 0-1 0-1 0 0-1 0-3 3-4 0 0

Kn-0 0 0-1 1 1 0-1 1 0 0-3 0-2 0-3 1-3 0-5

Wil-2 0-1 0-1 0-1 0-1 0-1 1 0 0-1 0-1 0-1 0 0-1

Compatible interaction phenotypes with all Xcr srains (susceptibility)

Nd-1 3-4 3 3-5 3-4 3-4 3-4 0-1 3-5 0-1 0-2 1-3 3-4

Zu-0 3 1-3 3-4 4 3-4 3-4 0 1-4 0-2 2-5 2-4 0-2

Differential interaction phenotypes with Xcr strains

Oy-0 0-1 0-1 0 0-1 3 3 3 0 0 0-1 0-1 0-1

Ws-0 0-1 0-1 0-1 1 3 1-3 0-3 0-1 0-3 0-1 0-1 0-1

Ct-1 0-1 0-1 3 1-3 3 1-3 0-2 0-2 2-3 0-2 2-4 2-3

Po-0 0-1 0-1 3 1-4 3 3 0-3 3 3-4 3-4 3-4 3-4

Rsch-4 0-1 0-1 3 1-4 3 2-3 0-2 0-2 0-2 0-3 0-2 0-2

Inconclusive interaction phenotypes with at least one Xcr strain

Edi-0 0-1 0-1 0 0-1 2-4 1-3 0-1 3-5 0-2 0-5 0-2 0-1 No-0 0-1 0-1 1 0-1 2 1-2 0 0-1 0 1-3 0-2 0-2 Tsu-0 0-1 0-1 1 0-1 1-2 1 2 0-1 0-1 0-5 0 0-1 Ler-0 0-1 0-1 1-5 1-5 0-1 1 3-5 3-5 2-5 5 5 4-5 Bur-0 0-1 0-1 2 2-3 1-2 1-3 0-2 0-3 1-3 0-3 0-2 0-3 Can-0 0 1 2 2-4 1-2 2-3 3-5 3-5 3-4 3-4 3-4 4-5 Mt-0 0 0-1 1-2 0-2 1-2 0-1 0 0 0-5 3-5 0-1 0-1 Ws-3 1 0-1 2 1-2 2 1-3 0 0-2 0-3 0-3 0-4 0-3 Sf-2 1-2 0-1 3-5 3-4 3-4 3 0-5 0-4 0-2 0-2 2-5 0-2 Hi-0 2-3 0-2 2-3 2-4 2 1-3 1-4 3-4 0-2 1-3 0-4 3 Wu-0 1-2 0-1 2 1-2 2 1-2 0-1 1-3 0-2 0-3 0-3 0-2 6181 (r6) 8506 Inoculation method A. thaliana accession

HRI strain accession

Xcr Xcc Xc

The IPs were considered: incompatible (plant resistance) when no typical symptoms were observed among all plants tested (IP 0-1) and compatible (plant susceptibility) when typical disease symptoms affecting a substantial area of the leaf were observed in all plants tested (IP 3-5). The IPs were considered inconclusive when at least one plant showed lesions affecting a small area of the leaf (IP 2) or when compatible and incompatible phenotypes were observed among plant replicates. These restrictive criteria aimed to highlight the most clear compatible and incompatible interaction phenotypes that were reproducible in all plants tested.

In interactions with Xcr strains, a few accessions showed clearly distinct phenotypes as illustrated in Figure 8. Four accessions (Col-0, Col-5, Kn-0 and Wil-2) were resistant to all strains tested (incompatible interaction); two accessions (Nd-1 and Zu-0) were susceptible to all strains (compatible interaction); and five accessions (Oy-0, Ws-0, Ct-1, Po-0 and Rsch-4) showed differential responses to the three strains tested representative of each known Xcr race. The results suggested that the latter accessions might distinguish the three known races of Xcr. This hypothesis was investigated further in this Chapter by testing more strains of each race and the results are presented in Section 2.3.4. The remaining 11 accessions tested showed inconclusive results in interactions between at least one plant and at least one strain (Table 6).

In interactions with Xcc strains, Wil-2 was resistant to all strains tested and Can-0 and Ler-0 were susceptible to all strains tested (Table 6). Differential responses of individual accessions to different Xcc strains were also identified. Col-0, Col-5 and Mt-0 were resistant to all strains except to Xcc race 6 (strain HRI6181) that produced disease symptoms in these accessions. Nd-1, Oy-0 and Edi-0 showed differential responses to Xcc race 1 (strain HRI3811) and Xcc race 4 (strain HRI1279A). Nd-1 and Edi-0 were susceptible to HRI1279A, but resistant to HRI3811 while Oy-0 was resistant to all strains except to HRI3811. The X. campestris strain HRI8506 was similar to Xcc race 4 in the interactions with brassica differentials of Xcc races (according to the results presented in Chapter 4), but it differed from the type strain of Xcc race 4 (HRI1279A) in the interaction phenotype with the A. thaliana accession Edi-0. This accession was resistant to the X. campestris strain HRI8506 and susceptible to the type-strain of Xcc race 4 (Table 6).

Figure 8. Differential interaction phenotypes of Arabidopsis thaliana accessions spray- inoculated with Xanthomonascampestris pv. raphani (strain HRI8305). Photographs were taken 10 days after inoculation: a, Col-0 incompatible phenotype (only minor lesions);

b, Nd-1 compatible phenotype (typical leaf spot symptoms).