Extent of colonization by Raffaelea quercivora of artificially inoculated living and gamma‐ray‐sterilized seedlings of two Japanese and three American oak species

Abstract

Mass mortality of Fagacean tree species caused by Raffaelea quercivora has occurred widely in Japan. Because conidia or other propagules of the pathogen have not been found in infected trees, pathogen spread is assumed to occur primarily by hyphae. To clarify the relationship between hyphal growth of the pathogen within trees and their vessel arrangements, we examined two native Japanese oaks, Quercus crispula and Quercus glauca, and three exotic American oaks, Quercus coccinea, Quercus palustris and Quercus rubra. Quercus glauca is a radial-porous species, whereas the other four species have a ring-porous wood structure. Hyphal growth within inoculated potted living seedlings and in cut, sterilized stem segments of these species was examined microscopically after fungal inoculation. Water conductance in the seedlings was examined using transverse stem sections. The proportion of non-conductive sapwood in Q. crispula, Q. coccinea and Q. palustris differed between inoculation and control treatment, being much higher in inoculated seedlings. The proportions were positively correlated with the extent of the hyphal growth. In sterilized stem segments, the extent of fungal colonization varied among the foreign ring-porous species Q. coccinea, Q. palustris and Q. rubra. It is hypothesized that the extent of colonization by R. quercivora reflects the extent of non-conductive sapwood irrespective of tree species, but is little affected by vessel arrangements.

Variations in virulence and hyphal growth of four Raffaelea quercivora isolates within Quercus crispula

“…In addition, the fungus appears to spread primarily via hyphal growth within the host trees Takahashi et al 2010) rather than via conidial spread, which is a more common phenomenon in vascular wilt diseases such as Dutch elm disease, caused by Ophiostoma ulmi (Buisman) Nannf. Moreover, a previous study using two Japanese and three non-native oak species showed that the extent of the horizontal hyphal growth within trees was positively correlated with the area of non-conductive sapwood in transverse sections (Torii et al 2014a). Hunt (Jacobi and MacDonald 1980); and laurel wilt, caused by Raffaelea lauricola T.C.…”

“…To visualize the distribution of the fungal hyphae, three of the four transverse sections were stained with fluorescein-conjugated wheat germ agglutinin (Sigma Aldrich, Tokyo, Japan) (Takahashi et al 2010;Torii et al 2014a). In each treatment, we created four transverse sections (20 lm thick) from the distal segments at the inoculation point using a sliding microtome (NS-31, Yamato Kohki Industrial, Asaka, Japan) equipped with a refrigeration unit (Electro Freeze MCR802A; Kelk, Hiratsuka, Japan).…”

“…To test the relationship between the proportion of non-conductive sapwood and hyphal growth for each isolate, we used Pearson's product-moment correlation coefficient using the data from each isolate and from the control (Torii et al 2014a). We assumed a binominal distribution for mortality rates because the variable is calculated as a percentage.…”

“…In JOW, microscopic observations of transverse and longitudinal sections suggested that most hyphae of the fungus under both artificial and natural conditions were detected in non-conductive sapwood, irrespective of the time elapsed since the inoculation or infection (Kuroda 2001;Takahashi et al 2010;Murata et al 2011;Torii et al 2011). Moreover, a previous study using two Japanese and three non-native oak species showed that the extent of the horizontal hyphal growth within trees was positively correlated with the area of non-conductive sapwood in transverse sections (Torii et al 2014a). These results suggested that the extent of the mycelial colonization would be proportional to the extent of non-conductive sapwood, irrespective of the tree species, and may determine the probability of mortality.…”

“…The mycelial distribution of R. quercivora in artificially inoculated seedlings and naturally infected trees was limited to tissues around the inoculation points and tunnels bored by P. quercivorus (Takahashi et al 2010;Torii et al 2011Torii et al , 2014a. Thus, multiple-inoculation tests of R. quercivora or mass attacks by P. quercivorus appear to be necessary to induce tree mortality Takahashi et al 2010).…”