Katsunori Nakamura

Susceptibility of adult trees of the endangered species Pinus armandii var. amamiana to pine wilt disease in the field

Adult trees of Pinus armandii var. amamiana (PAAm) and P. thunbergii grown in the field were inoculated with 100 000 or 1000 of the nematode Bursaphelenchus xylophilus to evaluate their susceptibility to pine wilt disease. PAAm trees inoculated with 100 000 nematodes started to show disease symptoms 2 weeks after inoculation, and all died within 29 weeks. Although the PAAm trees inoculated with 1000 nematodes tended to show delayed disease symptoms compared with those inoculated with 100 000 nematodes, all of them died within 33 weeks after inoculation. All P. thunbergii trees inoculated with 1000 nematodes had died 6 weeks after inoculation. In the nematode-inoculated PAAm trees, death of branches distal to the nematode inoculation site was the first visible symptom, followed by the systemic discoloration of needles, whereas the whole tree wilted simultaneously in P. thunbergii trees. In nematode-inoculated PAAm trees, the period from inoculation to death was longer than that in P. thunbergii. These results suggest that adult PAAm trees are susceptible to pine wilt disease, but are less vulnerable than P. thunbergii.

AN ENTOMOPARASITIC ADULT FORM IN BURSAPHELENCHUS DOUI (NEMATODA: TYLENCHOMORPHA) ASSOCIATED WITH ACALOLEPTA FRAUDATRIX

Abstract:  The nematode family Aphelenchoididae (Rhabditida: Tylenchomorpha) includes species with various feeding habitats. Bursaphelenchus, a member of the family, has for a long time been considered as a home for plant parasitic or mycophagous species (or both). However, recent intensive biological studies on the family revealed that the genus contains several insect parasitic species. Dauer juveniles of Bursaphelenchus doui were isolated from Acalolepta fraudatrix during a field study of longhorn beetle–Bursaphelenchus nematode associations. Two different insect-associated forms, an “entomoparasitic adult form” and a regular dauer juvenile, were isolated from a single individual beetle in a subsequent laboratory investigation of the B. doui–A. fraudatrix relationship. Thus these 2 distinct, insect-associated forms were confirmed to occur simultaneously. The entomoparasitic form is morphologically similar to that of Bursaphelenchus luxuriosae, with a dome-shaped head and vacuole-like spots assumed to be an internal structure of sensory organ, a stylet, a metacorpus (median bulb), and a moderately-developed and seemingly fully functional reproductive system. It is distinguishable from B. luxuriosae based on male spicule morphology and female tail morphology. A degenerate ingestive–digestive system distinguishes the entomoparasitic form from the propagative form and, unlike dauer juveniles, it has a moderately-developed reproductive system. The presence of this characteristic parasitic adult form is known only in these 2 Bursaphelenchus species. However, these 2 species did not form a clear monophyletic clade within the Bursaphelenchus xylophilus group and, thus, this characteristic parasitic form may occur independently in each species.

Pine wilt disease as promising causal agent of the mass mortality of Pinus armandii Franch. var. amamiana (Koidz.) Hatusima in the field

The causal agent of the mass mortality of field populations of Pinus armandii Franch. var. amamiana (Koidz.) Hatusima (PAA) was investigated with special respect to the involvement of pine wilt disease. Wood chips, branches and/or increment cores for detecting the pinewood nematode, feeding marks of the vector insect and environmental stress in the past, respectively, were taken from live and dead PAA trees grown in three locations, Yaku-shima and Tanega-shima Islands and a plantation in Kagoshima City, from 1997 to 1998. Five trees died after the spring of 1996 and, of these, four were inhabited by the pinewood nematode. Feeding marks of the vector insects were found on the branches of all dead trees and most of the live trees investigated. These results suggest that the infection of pine wilt disease in PAA trees occurs in the field. Annual ring growth of the sample trees showed neither intervention nor growth reduction, which implies strong environmental stress that may cause mortality in PAA trees.

Virulence of Bursaphelenchus xylophilus Isolated from Naturally Infested Pine Forests to Five Resistant Families of Pinus thunbergii

Pine wilt disease is one of the most serious epidemic tree diseases in Japan, and resistant pine trees have been developed through a breeding program. To evaluate resistance of resistant families of Japanese black pine, Pinus thunbergii, to the pinewood nematode, Bursaphelenchus xylophilus, isolated from the field, and to determine whether differentiation of pathogenicity to resistant pine families appears in the nematode isolates, seedlings of five resistant pine families were inoculated with 25 nematode isolates. Disease incidence 18 weeks after inoculation was significantly different among nematode isolates and among pine families but there was no interaction effect between nematode isolate and pine family. This indicates that nematode isolates did not have differential host specificity to resistant families of P. thunbergii. Isolate Shimabara, a test isolate of the breeding program, showed the same degree of virulence as the highly virulent isolates frequently used in experiments. However, more virulent isolates than Shimabara were found among the isolates collected from natural pine forest. This indicated that B. xylophilus populations with higher virulence than Shimabara exist in the natural population. These findings are important in development of more efficient breeding procedures for resistant pine trees.

The process of decline of an endangered tree species,Pinus armandii Franch. var.amamiana (Koidz.) Hatusima, on the southern slope of Mt. Hasa-dake in Yaku-shima Island

The process of decline of an endangered tree species,Pinus armandii var.amamiana, was monitored on the southern slope of Mt. Hasa-dake in Yaku-shima Island from 1994 to 1998. There are 163 trees ofP. armandii var.amamiana over 1.3 m in height. They are distributed on steep ridges and rocks with a thin soil layer mostly consisting of friable granite. During the monitoring period, 21 trees ofP. armandii var.amamiana died and the mortality rate was 12.9%. Dead trees were categorized into three types: standing, uprooted and landslide. The uprooted- and landslide-typed dead trees were found only after a severe typhoon struck Yaku-shima Island. This suggests that the combination of fragile site conditions and severe typhoons play an important role in the process of decline ofP. armandii var.amamiana. The standing-typed dead trees were presumed to have been killed by pine wilt disease, accounting for 71.4% of the dead trees. However,Bursaphelenchus xylophilus, the cause of pine wilt disease, was not detected from any of the wood chips or branch samples from the standing-typed dead trees ofP. armandii var.amamiana. This might indicate that some factor(s) other than pine wilt disease could be responsible for the standing-typed death ofP. armandii var.amamiana in natural habitats.

Different Development of Pine Wilt Disease in Artificially Infected Pinus thunbergii Seedlings Potted together with Different Tree Species

To evaluate the effect of adjacent tree species on the susceptibility of Japanese black pine (Pinus thunbergii) to pine wilt disease, an inoculation experiment was conducted usingP. thunbergii seedlings potted with seedlings of six tree species,i. e. Alnus sieboldiana, Eurya japonica, Lespedeza bicolor formacutifolia, Pinus thumbergii, Robinia pseudo-acacia andSarothamus scoparius. About ten months after planting, they were inoculated with the pinewood nematode (Bursaphelenchus xylophilus) in early July 1992. After that, the proportion of pine seedlings with completely discolored foliage increased more quickly when the seedlings were potted withR. pseudo-acacia, S. scoparius orA. sieboldiana than when potted withP. thunbergii, L. bicolor orE. japonica. At the end of the study period, 17 weeks after inoculation, it reached 90.6%, 90.0%, 87.5%, 72.7%, 63.3%, and 59.4% when the pine seedlings were potted withR. pseudo-acacia, S. scoparius, A. sieboldiana, P. thunbergii, L. bicolor andE. japonica, respectively. This indicated that the susceptibility ofP. thunbergii seedlings to pine wilt disease was influence by the species of adjacent trees.

Distance effect of co-occurring tree species on pine wilt disease incidence in Pinus densiflora seedlings inoculated with Bursaphelenchus xylophilus

Field-grownPinus densiflora seedlings were inoculated withBursaphelenchus xylophilus and the incidence of pine wilt disease was analyzed with respect to the spatial relationships between pine seedlings and adjacent seedlings of other tree species in a pure pine stand and three stands mixed withAlnus sieboldiana, Sarothamus scoparius or naturally associated species. The disease incidence was 60.9% in a 0–40 cm distance class from the nearestA. sieboldiana seedlings and then decreased with increasing minimum distance. The mean minimum distance between pine seedlings andA. sieboldiana was significantly shorter in diseased seedlings than in healthy ones. The highest disease incidence (53.9%) was found in a 0–40 cm distance class from the nearestS. scoparius, although the relationship with minimum distance was blurred by a high incidence in the 80-cm distance class. The mean minimum distance between pine seedlings was significantly short in diseased seedlings in a pure stand, whereas it was not so short between pine seedlings and other tree species in the two stands mixed withS. scoparius and the naturally associated species. The distance effect on disease incidence was noticeable inA. sieboldiana andP. densiflora at high density. This was not so clear inS. scoparius and was not found in the naturally associated species.

Effects of low-temperature summer nights on adults of Monochamus alternatus (Coleoptera: Cerambycidae)

ABSTRACT To clarify the factors determining the northern distribution limit of Monochamus alternatus Hope, which vectors the pinewood nematode, we examined our hypothesis that “low-temperature summer nights in cool regions affect the survival, maturation feeding, and ovary development of M. alternatus, and determine its distribution limit.” We measured summer air temperatures in three pine forest stands in northern Japan. Based on the results, we examined the effects of low nighttime temperatures on survival, maturation feeding, and ovary development in the laboratory. Low nighttime temperatures had little effect on the survival and the total amount of maturation feeding of M. alternatus adults. A daytime temperature of 25°C and nighttime temperature of 15°C did not affect ovary development. In contrast, 25°C in the daytime and 10°C in the nighttime delayed ovary development and reduced the number of mature eggs, but some ovary eggs were able to mature under this condition. We concluded that the effect of low-temperature summer nights on ovary development alone could not explain the northern distribution limit of M. alternatus.