Search

Summary Bursaphelenchus xylophilus, the causative agent of pine wilt disease, and B. mucronatus are transmitted by Monochamus adults to host trees. Feeding and oviposition wounds made by vectors are the primary transmission pathways to trees. Monochamus saltuarius female adults carrying B. mucronatus were reared singly and allowed to mate with nematode-free males at 5-day intervals, to determine the ratio of nematodes transmitted via the two different pathways. The survival time, lifetime fecundity and other reproductive traits decreased with increasing initial nematode load (number of nematodes carried by a newly-emerged adult). Model selection indicated that numbers of B. mucronatus departing from M. saltuarius and of those transmitted to pine via oviposition and feeding wounds were closely related to the initial nematode load, which was affected by survival time and number of oviposition wounds constructed. Most temporal patterns of nematode departure and transmission via oviposition or feeding wounds from individual vectors had a peak in the medium and heavy initial nematode loads. The nematode departure curve was significantly similar in shape to each of the nematode transmission curves via oviposition and feeding wounds, which were not significantly similar to each other, for individual vectors. After M. saltuarius females began to construct the oviposition wounds, it was estimated that B. mucronatus was transmitted via the oviposition wounds at a probability of 0.767. If B. xylophilus is transmitted in the same way as B. mucronatus, the incidence of pine wilt disease by mature vectors would be much lower than in the case of no oviposition-related transmission.

Summary The pine wood nematode (PWN), Bursaphelenchus xylophilus, is the causative agent of pine wilt disease and is transmitted primarily by cerambycid adults of the genus Monochamus. It is believed to have been introduced into Japan from North America in the 1900s and confirmed to have invaded western Europe in 1999. The nematode has been devastating pine forests in invaded areas because the native pine species are susceptible. This study examined the number of PWN carried by individual M. carolinensis adults (initial PWN load) immediately after emerging from dead Pinus sylvestris and P. strobus trees in Illinois, USA. The data were compared with the initial PWN load on M. alternatus adults emerging from dead P. thunbergii trees in Japan ca 80 years after the PWN invasion. When the zero-inflated negative binomial model was fitted to the initial PWN loads it provided three results. First, the PWN exhibited clumped distributions among vectors in the PWN-M. carolinensis-P. sylvestris or P. strobus system in Illinois and the PWN-M. alternatus-P. thunbergii system in Japan. Second, there was no difference in the frequency distribution of the initial PWN load between the three disease systems. Third, the initial PWN load increased as the overwintering PWN density in wood or the adult body mass increased, whereas it decreased as the emergence date of vectors increased. Consequently, the frequency distribution of initial PWN load exhibited no evolutionary change in Japan and the initial PWN load was considered to be determined by environmental factors and conditions.

Summary The pine wood nematode (PWN), Bursaphelenchus xylophilus, is the causative agent of pine wilt disease and is transmitted by cerambycid adults of the genus Monochamus. It is believed to have been introduced into Japan from North America in the 1900s and since then it has been devastating the native pine forests. By contrast, the epidemics of the disease have never occurred in pine forests native to North America in the original habitats, indicating no expression of virulence. To determine the evolutionary change in the transmission traits of PWN, the temporal pattern of PWN transmission to fresh branch sections of Pinus sylvestris was studied using 38 Monochamus carolinensis adults in Illinois, USA, and compared with previous studies on the PWN-M. alternatus and B. mucronatus-M. saltuarius systems in Japan. A great difference was observed in the initial nematode load among vectors irrespective of the three systems. The maximal numbers of PWN transmitted (5 days)−1 by individual vectors were greater and the efficiencies of nematode transmission and invasion were higher for the Japanese PWN-M. alternatus system than for the North American PWN-M. carolinensis system in two heavy classes of initial load of over 1000 nematodes. The proportion of PWN transmitted to healthy pine trees by reproductively immature female vectors was small (2.5-11.5%) in the three nematode-vector systems in most cases. The evolutionary changes in the transmission traits and the control of the range expansion of the PWN were discussed in relation to the expression of virulence.

Summary An Oryza sativa cultivar inoculated with the white tip nematode, Aphelenchoides besseyi, was cultivated over 18 years to investigate the population dynamics of the nematode and the change in the virulence of nematodes and tolerance/resistance of plants. The mean number of living nematodes per seed (S) showed two peaks, each of which was followed by a 3-year decrease in the S value, during the initial 10 years. The annual reproduction curve between in year and in year t had a peak in a range of of −1 in the previous year, whereas the proportion of seeds with living nematodes increased with increasing mean number of living nematodes seed−1 in the current year. Inoculation tests of the nematode cohorts on the seedling cohorts derived from seeds harvested in the same or different years indicated that the initial 6-year interaction between the rice and nematode populations caused the nematodes to increase the incidence of white tip disease and to decrease the degree of seed swelling and the proportion of heavy seeds. On the other hand, the interaction caused the rice plants to prevent the incidence of disease from increasing and the degree of seed swelling and the proportion of heavy seeds from decreasing.

Monochamus alternatus alternatus Hope is distributed in China and Taiwan and transmits the pinewood nematode, the causative agent of pine wilt disease. Unlike univoltine and semivoltine Monochamus ...

Monochamus alternatus alternatus Hope (Coleoptera: Cerambycidae) is the primary vector of pine wilt disease in China and Taiwan. It has a partially bivoltine life cycle with facultative diapause at the final larval instar that is mediated by the initial larval density. To evaluate the effect of density-related induction of diapause on population fluctuation, one, two or four newly hatched larvae were inoculated on individual Pinus thunbergii bolts and the life-history traits including diapause were investigated. This experiment indicated that as initial population size/bolt (crowding) increased, the diapause incidence, larval stage survival, adult body size, development time of nondiapause individuals and pupal period of diapause individuals decreased. The analysis indicated that larval interference as well as food shortage suppressed diapause induction. Increasing larval density decreased the probability of diapausing, survival, adult body mass and other traits. Using the data obtained from this study, a simple mathematical model indicated that inverse density-dependent induction of diapause in the first generation had a destabilizing effect on the population size of overwintering generation, which was composed of diapause larvae of the first and second generations; whereas, direct density-dependent induction of diapause exerted a stabilizing effect on it.

Summary The white-tip nematode, Aphelenchoides besseyi, is a seed-borne ectoparasite of the rice plant Oryza sativa. Nematodes in dried rice seeds are in dehydrated quiescence or anhydrobiosis and are reactivated by rehydration. To understand the persistent parasitism better, the effects of temperature on nematode survival were examined during prolonged periods of seed storage. There was a marked difference in the survival rate among different temperatures; survival was lower at 20 and 25°C than at −5 to 10°C after storage for 190 days. No nematodes survived a storage period of 1313 days at 20 and 25°C. When rice seeds were held at 5°C, the survival rate of nematodes decreased during a period of 7315 days. The estimated instantaneous rate of mortality per month was higher in the early and late parts of the period than in the middle part. Another nematode population also showed a reduction in the survival rate during 6553 days storage at 5°C. In storage at −30°C, survival was not affected during a period of 6485 days. After storage at −30°C for 6546 days, 96% of rice seeds germinated, whereas 60-96% of seeds of two groups germinated after storage at 5°C for 6610-7373 days. Therefore, prolonged storage of rice seeds at 5 or −30°C was unlikely to disrupt the connection between rice plants and A. besseyi.

Macrodasyceras hirsutum Kamijo (Hymenoptera: Torymidae) has been considered to be a specialized seed parasitoid wasp of Ilex integra Thunb. since the description as a new species. A previous study showed that they deposited eggs in seeds of the closely related tree I. latifolia when I. integra was experimentally absent. However, natural occurrence of oviposition into I. latifolia or successful emergence from I. latifolia as adults had not been investigated. Thus, to determine whether M. hirsutum utilizes I. latifolia seeds in the fields, we dissected 536 I. integra seeds and 501 I. latifolia seeds. Macrodasyceras hirsutum larvae were found in 152/307 fertilized seeds of I. integra, while found in 16/285 fertilized seeds of I. latifolia. Additionally, 12 male and 13 female M. hirsutum adults emerged from I. latifolia berries collected from field. The present study confirmed natural occurrence of oviposition by M. hirsutum females in I. latifolia seeds and the emergence of adults from berries.

Natural selection operates on a suite of life-history traits. Those traits are under constraints and trade-offs which often differ among species. Thus, there are an array of suites found in insects. Studies conducted to date predict two reproductive strategies in the family Cerambycidae: body-size dependent ovariole number and pro-ovigeny versus body-size independent ovariole number and synovigeny. To investigate these relationships, we examined the elytral length, the body mass, the number and development of ovarioles, and the content of alimentary canals for adult females of three cerambycid species captured in mulberry orchards. The results confirmed synovigeny in two lamiines Apriona japonica Thomson (Coleoptera: Cerambycidae) and Psacothea hilaris (Pascoe) (Coleoptera: Cerambycidae) and no adult feeding in the cerambycine Xylotrechus chinensis (Chevrolat) (Coleoptera: Cerambycidae). We also found the dependence of ovariole number on body size and pro-ovigeny in X. chinensis, whereas invariable numbers of ovarioles irrespective of body size and synovigeny were found in A. japonica and Ps. hilaris. Two likely different suites of reproductive traits including size dependence of ovariole number and ovigeny are discussed by comparing life-history traits of nine cerambycids reported by the present and previous studies.

Monochamus alternatus endai Makihara is the primary vector of pine wilt disease in Japan and South Korea. Larvae of this cerambycid beetle exhibit induced diapause in the final instar. To d...

Monochamus alternatus Hope adults vector the pinewood nematode, which is the causal agent of pine wilt disease. Larvae of this cerambycid beetle enter diapause in the final instar. To deter...

Based on DNA sequences of mitochondrial cytochrome oxidase subunit I (mtCOI) genes, three PCR forward primers specific for Bursaphelenchus mucronatus mucronatus, B. m. kolymensis, and B. xylophilus (Nematoda: Aphelenchoididae), and one common PCR reverse primer were developed to determine the interspecific hybrids in a population. PCR amplification with the new primer pairs was expected to produce DNA fragments of 286 bp, 117 bp and 479 bp for B. m. mucronatus, B. m. kolymensis, and B. xylophilus, respectively. Electrophoresis of PCR products with the new primer pairs distinguished the two B. mucronatus subspecies and B. xylophilus collected in Japan. It also exhibited a band-free image for Bursaphelenchus doui, B. niphades, B. sinensis, B. firmae, Bursaphelenchus sp. (NK230), and Aphelenchoides stammeri, whereas it exhibited one (ca. 100-bp DNA fragment) and two (ca. 100 and 500-bp DNA fragments) bands for B. poligraphi and B. yongensis, respectively. As PCR products with other primer pairs for nuclear ribosomal RNA (rRNA) gene distinguished B. mucronatus (210 bp DNA fragment), B. xylophilus (557 bp fragment), and the eight other nematode species (no amplification), primer pairs for mtCOI and rRNA genes can determine whether B. poligraphi resides in specified trees or in study pine stands. Thus, those primer pairs would be helpful to find out hybrids between B. mucronatus and B. xylophilus and to analyze the dynamics of hybrids and introgressants in hybrid-derived populations in field and laboratory.

Aphelenchoides besseyi Christie (Aphelenchida: Aphelenchoididae) is a seed-borne ectoparasite of rice Oryza sativa Linnaeus and the causative agent of white tip disease. To evaluate effects that the amount of substances translocated to seeds and the consumption by nematodes exert on seed size, 20 μL water containing 100 living or dead nematodes or no nematodes was inoculated individually into florets in bloom that appeared on culms with the amount of foliage modified: culms with no blades, half-cut blades, or intact blades of the three uppermost leaves. Inoculation of living A. besseyi nematodes into rice florets resulted in the highest proportion of light seeds (specific gravity

To evaluate the effects of Taiwanese Monochamus alternatus alternatus (Coleoptera: Cerambycidae) (T) on native M. a. endai populations (J) and pine wilt disease epidemics in Japan, the biological traits of reciprocally crossed lines between two subspecies were compared with parental lines. No significant differences were observed in egg viability among four crosses in the F1 generation. Hybridized lines had greater survival rates than inbred lines of the J × J cross. The incidence of larval diapause was lower in the inbred lines of M. a. alternatus (0.176 and 0.361) than in the inbred lines of M. a. endai (1.000). Hybridized lines showed a high diapause incidence of 0.896–1.000 in the F1 and F2 generations, indicating that diapause induction was dominant over nondiapause induction. Large numbers of ovarioles dominated over small numbers of ovarioles. Hybridized lines revealed greater adult body sizes than the inbred lines of parental subspecies in the F1 generation (heterosis), but not in the F2 generation. Heterosis in adult body size may contribute to pine wilt disease epidemics because of the greater oviposition rate and initial load of pathogenic nematodes. Two unlinked loci are estimated to control diapause induction. Gene introgression and the dynamics of nondiapause-inducing alleles are discussed.

To establish a simple, time-saving method for selecting Pinus tree candidates tolerant to the infection of Bursaphelenchus xylophilus, the causative agent of pine wilt disease, three experiments were conducted. When two virulent isolates Sc-9 and T-4 were inoculated on cut ends of branch sections, more nematodes were inhibited from passing through P. taeda branch sections than P. thunbergii sections. Sc-9 tended to pass through P. thunbergii sections more easily than T-4. The probability of nematodes passing through 5-cm-long P. thunbergii branch sections was greater when 200 nematodes were inoculated. When 200 Sc-9 nematodes were inoculated on 5-, 10- or 15-cm-long branch sections of P. taeda trees and P. thunbergii clones of different tolerance classes in late August, an insignificant correlation was found between the tolerance class of P. thunbergii clones and the number of nematodes passing through 5-cm-long branch sections, although nematode dispersal ability could distinguish between tolerant and susceptible pine clones in mid-August, suggesting it was a helpful method for selecting tolerant tree candidates. Abrupt seasonal increases in tolerance level suggested that weather conditions, such as hot temperatures and drought, may have disrupted the tolerance mechanism of resistant species of P. taeda.

Akbulut, Suleyman/0000-0002-1497-3562 WOS: 000465174800019 Cerambycid adults of the genus Monochamus transmit the pine wood nematode (Bursaphelenchus xylophilus [Steiner et Buhrer] Nickle), the causative agent of pine wilt disease. To confirm the transmission of B. xylophilus between vector species, nematode-infected Monochamus alternatus Hope and nematode-free Monochamus saltuarius (Gebler) adults were paired and their behaviour was observed for 107-220 min. In three pairs that exhibited mounting without copulation, nematode transmission occurred, whereas it was not found in another pair without mountings. The effect of nematode transmission between different vector species on the invasion biology of B. xylophilus is discussed. Japan Society for the Promotion of ScienceMinistry of Education, Culture, Sports, Science and Technology, Japan (MEXT)Japan Society for the Promotion of Science [26292080] Japan Society for the Promotion of Science, Grant/Award Number: JSPS KAKENHI grant 26292080

Semanotus japonicus larvae feed on the phloem of living trees of Cryptomeria japonica and Chamaecyparis obtusa, and newly emerged adults stay in xylem from August through the following April when they emerge from the trees. A positive correlation is known between the female body mass and lifetime fecundity in this insect. To determine the timing of maturation in reproductive organs and the relationships of female body size to the total number of ovarioles of a female (ovariole number) and egg production per ovariole, we dissected adults taken from dead trees from October through the following April. Dissecting females showed that well-developed eggs began to appear in the calyx in April, that the ovariole number ranged from 34 to 79, and that the ovariole number per female and egg production per ovariole increased as the female body mass increased, indicating that the body size-dependent lifetime fecundity was attributed to greater ovariole number and greater egg production per ovariole for large ...

To determine the effect of larval crowding on induction of diapause in Monochamus alternatus alternatus Hope (Coleoptera: Cerambycidae), newly hatched larvae were inoculated singly (density 1) or in pairs (density 2) on Pinus thunbergii Parlatore (Pinaceae) bolts and reared at 25 °C and 16 light:8 dark hour photoperiod. Adults emerged at density 1 and 2 between 75 and 139 days after the inoculation. About 150 days after larval inoculation, dissection of pine bolts indicated the incidence of larval diapause was greater at density 1 (0.536) than at density 2 (0.222), indicating inversely density-dependent induction of diapause. For both larval densities, the probability of entering diapause increased as bark surface area per larva (a proxy for per capita amount of food available) increased, showing that diapause induction can be explained by the amount of food available per larva. Adults that emerged at density 1 were heavier than those at density 2. When diapause and nondiapause insects occurred in the same pine bolts, six heavy adults had experienced diapause whereas three heavy adults had not, indicating that bark surface area affected the induction of diapause indirectly through larval interaction. The relationship between two different manners of density-dependent diapause induction and food supply is discussed.

Mandibles and genae of adult females of most members in the genus Doubledaya (Coleoptera: Erotylidae: Languriinae) exhibit distinct left–right asymmetry, i.e., the left side of the traits is larger than the right, which may facilitate boring into hard materials for oviposition; females of D. bucculenta and D. sinuata excavate holes or relatively deep depressions with their mandibles on internodes of dead bamboos to deposit the eggs on the inner surface of internode cavities. To determine the relationship between the degrees of mandibular and genal asymmetry and hardness of oviposition substrates, we measured the mandibular length and genal width of D. tonkinensis and D. sinuata , both of which oviposit on Sinobambusa sat bamboos, and examined the structure of their oviposition holes. For D. tonkinensis , we additionally examined the features of host culms. D. tonkinensis females showed a slight degree of left-directional asymmetry in the mandibles compared to males. By contrast, D. sinuata females showed distinctly marked left-directional asymmetry in the mandibles and genae. Females of D. tonkinensis excavated soft walls of internodes of weakened, young bamboo culms, whereas D. sinuata excavated hard internode walls. Cross sections revealed that the holes of D. tonkinensis and D. sinuata were funnel-shaped, indicating that the females first excavate a cone-shaped depression in the internode using mandibles followed by ovipositor’s penetrating wall from the bottom of the depression to the inner surface of the internode cavity. The results strongly suggest that the markedly asymmetric mandibles of Doubledaya females facilitate oviposition into hard substrates.

1. Farming by non-human organisms has arisen independently in several animal lineages, allowing them to survive on food sources that are otherwise difficult to access. However, agricultural gardens are prone to invasion by parasites that overgrow cultivars in the absence of host animals. The presence of garden parasites and associated host adaptations are well studied in advanced fungal agriculture practised by social insects (ants, termites, and ambrosia beetles), but the impact of garden parasites in more primitive forms of agriculture is poorly known. The aim of the present study was to report the presence of weed fungi in fungal gardens of the non-social lizard beetle Doubledaya bucculenta Lewis (Coleoptera: Erotylidae: Languriinae), which cultivates the yeast Wickerhamomyces anomalus (E.C. Hansen) Kurtzman, Robnett & Bas.-Powers in the internodes of recently dead bamboo culms. 2. The filamentous fungi Arthrinium spp., Fusarium spp., and an unidentified species were isolated from the inner wall and insect-made wall debris of bamboo internodes inhabited by the beetle. 3. When cultured together with the yeast in the absence of host larvae, Arthrinium sp.1 outcompeted the yeast, suggesting that this filamentous fungus can invade yeast gardens but is normally suppressed under natural conditions. 4. Rearing experiments showed that beetle larvae grew faster when grown on yeast cultures than when fed only Arthrinium sp.1. 5. These results suggest that Arthrinium sp.1 acts as a weed in the fungal gardens of D. bucculenta, inhibiting both growth of the beetle larvae and proliferation of the beneficial yeast.

Bursaphelenchus xylophilus (Steiner et Buhrer) Nickle causes pine wilt disease and is transmitted by Monochamus alternatus Hope adults in Japan. To determine the effects of the beetle larvae on the nematode population and the effects of the nematode population on beetle larval growth, we inoculated fresh pine bolts with various combinations of B. xylophilus, M. alternatus, and the blue-stain fungus Ophiostoma minus (Hedgc.) Syd. & P. Syd., the food of the nematodes. We also compared the content of ergosterol among the phloem, xylem, and larval frass. The nematode density was 22 or 25 times higher in frass than in phloem and the lowest in xylem, indicating positive effects of beetle larvae on the nematode population growth. Frass had twice higher ergosterol content than the phloem, indicating the fungal mycelial content was greatest in the frass. The difference in the content of fungal mycelia could explain in part the difference in B. xylophilus density between the frass and phloem. By contrast, B. xylophilus did not affect the growth and developmental rate of M. alternatus. This study showed that the relation between the beetle and nematode populations changes from mutualism to commensalism corresponding to the dispersal to reproduction stages of the nematode population.

Monochamus alternatusadults carry the pine wood nematodeBursaphelenchus xylophilus. The nematodes are present in the tracheal system of vector beetles. It is known that carrying more than 10 000 nematodes causes drastic reduction in the lifespan and flight activity of beetles. To understand the effects of heavy nematode loads, 41 newly emergedM. alternatusadults were examined for lateral distribution of nematodes within the insect body. The nematodes were distributed unevenly between right and left sides of the beetle body at small nematode loads, whereas they were distributed evenly between them at heavy nematode loads. An analysis using Hagen-Poiseuille flow in a thin tube suggested that the rate of air flow decreased exponentially as the number of nematodes increased in a trachea. Thus, even distribution of nematodes between right and left sides of the vector body was considered to be responsible in part for the effects of heavy nematode loads on beetle activity.

To determine the difference in egg-period length and its effects on postembryonic development in Psacothea hilaris (Pascoe), 14 adult females, which were captured in Yayoi, Tokyo, and their offspring were reared at 25 °C under a photoperiodic regime of 16-h photophase and 8-h scotophase. When 1,486 eggs were placed on wet tissue paper within a day of oviposition, they took from 7 to >40 days to hatch, although 85.9 % of larvae hatched between 7 and 13 days following oviposition; 71 % of females produced larvae with long egg periods of 21 or more days. When 78 newly hatched larvae with different egg-period lengths were reared singly in sections of mulberry shoots, 39 emerged as adults between 57 and 117 days posthatching. Dissection showed two feeding larvae and a diapause pupa in shoot sections 141–168 days after larval hatching. A tradeoff was observed between the egg-period length and the time taken for postembryonic development in females, but not in males. Egg-period length did not affect adult body size. An increased amount of available food (volume of shoot sections) increased adult body size but did not affect the time for postembryonic development.

Lateral gene transfer is any process in which an organism incorporates genetic material from a phylogenetically distant and reproductively isolated organism. A previous survey demonstrated that two Japanese populations of Monochamus alternatus Hope carried many Wolbachia genes that had been acquired by lateral gene transfer in their own genomes, but were not infected with Wolbachia. To understand the prevalence of Wolbachia infection and laterally transferred Wolbachia genes in this beetle, we performed a broader survey of natural populations covering two subspecies of M. alternatus from Japan (M. alternatus endai) and Taiwan (M. alternatus alternatus). We detected laterally transferred Wolbachia genes in all Japanese and Taiwanese populations of M. alternatus, but no Wolbachia infection in any population. In addition, we confirmed the absence of Wolbachia infection and of transferred Wolbachia genes in two Japanese populations of Monochamus saltuarius Gebler, which is a congeneric relative of M. alternatus. Our findings suggest that the Wolbachia endosymbiont as gene donor has disappeared from M. alternatus, and that the transfer of part of its genome to M. alternatus occurred in the ancestor of M. alternatus before the subspeciation event.

To confirm the facultative diapause of Monochamus alternatus alternatus Hope (Coleoptera: Cerambycidae) and to determine the relationships between available larval food resources, diapause, and adult traits, newly hatched larvae were inoculated singly on 98 Pinus densiflora Siebold & Zuccarini (Pinaceae) bolts and reared at 25 °C, 100% r.h., and L16:D8. Fifty adults emerged from them, between 70 and 126 days after larval inoculation. The remaining 48 bolts that did not produce adults were divided into two groups. One group was transferred to 10 °C, 100% r.h., and L8:D16, and returned 140–154 days later to the original conditions, resulting in adult emergence. The other group was maintained under the original conditions for a mean of 358 days. These bolts did not produce adults. Dissection revealed that development was arrested at final instar in pine bolts. The larvae developed into adults after being exposed to 10 °C, 100% r.h., and L8:D16 for 146 days. Consequently, this species has facultative diapause. Diapause incidence was estimated to be 0.42. Non-linear model and one-way ANOVA showed a positive correlation between adult body size and available food resources under conditions of food shortage, and no effects of diapause or available food resources on the ovariole number, respectively. When larvae were inoculated on 28 pine branch sections, the results were similar to those obtained from pine bolts and led to estimation of a low diapause incidence of 0.045. The combined data showed the inhibitory effect of food shortage on diapause induction. Diapause of M. a. alternatus, especially reduced diapause induction in response to environmental deterioration (food shortage), is discussed in relation to risk-spreading.

Bursaphelenchus xylophilus causes pine wilt disease in Pinus trees whereas B. mucronatus has no or little virulence to the trees. Interspecific crossing experiments conducted so far suggest reproductive interference between the two nematode species. Theory predicts that one of the two competing species populations quickly displaces the other through reproductive interference in a frequency-dependent manner. Thus, it is anticipated that B. mucronatus suppresses the virulence of B. xylophilus against pine trees when B. mucronatus heavily outnumber B. xylophilus. To determine the suppressive effects of B. mucronatus, the two nematode species were inoculated simultaneously on 30 3-year-old Pinus thunbergii seedlings at three combinations of different numbers, and B. xylophilus alone was inoculated on 30 other seedlings at the corresponding numbers in early August. Seedlings were observed at intervals of 4 or 6 weeks and two stem sections were sampled from each seedling to determine the density and species composition of nematode populations after death or in December. Inoculation of B. mucronatus significantly retarded the speed of foliage discolouration from 0.170 ± 0.024 week−1 to 0.061 ± 0.017 week−1 and significantly prolonged the survival time of seedlings. Bursaphelenchus mucronatus significantly reduced the nematode density from 1799.7 ± 305.0 to 521.0 ± 148.4 (g dried seedling stem)−1. Analysis of rDNA genotype showed 1846 B xylophilus, no B. mucronatus and one hybrid.

Macrodasycerashirsutum Kamijo is the seed parasitoid wasp of the bird-dispersed, dioecious tree, Ilexintegra Thunb. The wasp reduces the level of dispersal mutualism between the Ilex tree and its frugivorous birds by manipulating the color of mature berries. The female trees do not blossom every year and sometimes change sex. Thus, the reproduction biology of I. integra affects the population size and structure of M. hirsutum in a forest and consequently influences the seed dispersal mutualism between the tree and birds, because of limited ability of adult locomotion. To investigate the wasp population structure with reference to the dispersal mutualism between trees and birds, we isolated 14 microsatellite loci of M. hirsutum wasps. Every locus was polymorphic among 20 females, with 3–13 alleles per locus, without linkage disequilibrium. The observed and expected heterozygosities ranged from 0.100 to 0.900 and 0.099 to 0.818, respectively, indicating their utility in molecular analyses of the wasp population.

Preference-performance hypothesis in host selection by herbivorous insects predicts selective oviposition on plant on which the offspring maximise fitness. The seed parasitoid wasp, Macrodasyceras hirsutum Kamijo (Hymenoptera: Torymidae) selectively lays the eggs into the fertilised seeds of Ilex integra Thunberg (Aquifoliaceae). Only one larva develops in a seed. Therefore, the hypothesis predicts a uniform distribution pattern of wasp eggs among fertilised seeds. Dissection of 531 berries showed that M. hirsutum deposited one to five eggs into a fertilised seed. Iwao's patchiness regression suggested a uniform distribution pattern of M. hirsutum eggs among fertilised seeds at the scales of tree and berry and their random distribution pattern among berries at the tree scale. Destroying the connection of seeds within berries revealed that female wasps randomly selected berries for oviposition in most trees. Generalised linear mixed models showed that the number of fertilised seeds in a berry could not explain the number of eggs in a seed but could explain the number of eggs in a berry. Therefore, this study shows that the wasp females do not distinguish between berries with different numbers of fertilised seeds but distinguish between seeds harbouring different numbers of eggs, which supports the hypothesis.

Mouthparts and genae of the adult females of Doubledaya bucculenta Lewis (Coleoptera: Erotylidae: Languriinae) exhibit distinct directional asymmetry. Females excavate holes with their mandibles on hard internodes of bamboos to deposit their eggs on the inner surface of internodes. To determine the relationship between the spatial distribution pattern of completed oviposition marks (COMs) and the degree of mandibular asymmetry, COMs were counted on the culms and internodes of the bamboo Pleioblastus simonii (Carriere) Nakai (Poaceae), and the survival rate of immature stages was investigated. Another languriine beetle, Anadastus pulchelloides Nakane, exhibits a smaller degree of mandibular asymmetry. For this species, COMs were also investigated on culms and internodes but of the perennial grass Arundinella hirta (Thunb.) Tanaka. Females of the two beetle species selected large basilar internodes of host plant culms for oviposition in the field. The mean number ±SD of COMs per COM-containing inte...

Monochamus alternatus Hope is an important vector of the pinewood nematode Bursaphelenchus xylophilus (Steiner et Buhrer) Nickle, which causes pine wilt disease. To determine canopy-related differences in the activity density of M. alternatus adults in pine stands, sticky screen traps with no lure were suspended under and in the canopy at 3.8 and 10 m above the ground of a Pinus densiflora Siebold et Zuccarini stand during a flight season. The five upper and five lower traps caught 338 and 54 adults, respectively, during a flight season from 9 June to 12 October 2011, indicating that activity density was six times as high in the canopy as under it. Male-biased sex ratio was observed for trap catches during a season: The sex ratio was male-biased in the early half of the flight season and 1:1 in the late half. There was no difference in sex ratio between the inside and outside of the canopy. To determine the sex-related difference in flight activity, the adults were captured by two different measures in two other mixed stands of P. densiflora and P. thunbergii Parlatore. Kicking tree trunks to cause beetles to drop revealed a 1:1 sex ratio of catches, whereas sticky screen traps tended to show a male-biased sex ratio in a 4-year study, suggesting that males were more active fliers than females.

Pine wilt disease (PWD) is caused by the pinewood nematode (PWN), Bursaphelenchus xylophilus (Steiner et Buhrer) Nickle, and is transmitted by cerambycid beetles. In some pine trees infected with the PWNs in Japan, foliage changes from green to brown in summer to autumn of a nematode infection year (summer- autumn-killed trees) and the others in the following spring of a nematode infection year (spring-killed trees). The vector beetles require 1 or 2 years for development in cool summer areas and 1 year in warm summer areas. To evaluate the effects of the spring-killed trees and vectors with a long developmental time on the PWD epidemics, we presented simple mechanistic mathematical models. The models showed that it was possible for spring-killed trees to cause PWD epidemics when the transmission rate was high, and the efficacy of spring-killed trees as infection source was similar to that of summer– autumn-killed trees. Spring-killed trees and vector beetles with a developmental time of 2 years harbored in summer- autumn-killed trees delayed epidemic timing by 3–10 years or actually suppressed epidemics.

1. Frugivorous and seed-feeding insects may alter the traits of fruits, such as shape and size, which may influence fruit attractiveness to frugivorous birds. Consequently, trait-mediated interactions may occur in systems where plants, seed-dispersing frugivorous vertebrates, and frugivorous or seed-feeding insects interact. We investigated colour manipulation in Ilex integra Thunb. berries caused by the seed parasitoid wasp Macrodasyceras hirsutum Kamijo and how that manipulation relates to fruit attractiveness for frugivorous birds. 2. In winter, the colour of I. integra berries varied from green to red, but most berries were greenish, indicating that the berries were immature. Berry dissection indicated that the number of live parasitoid larvae present within each berry was closely related to berry colour – the greater the number of live larvae, more intense is the green colour of the berry. However, the wasp larvae did not modify the shape or size of the berries. More than 98% of berries that were protected from the insects by gauze bags ripened and turned red. In the present study, berries with unfertilised seeds alone turned red. Field-feeding preference tests showed that the brown-eared bulbul Hypsipetes amaurotis Temminck preferred red berries to green berries. 3. We demonstrated that the seed parasitoid wasp manipulates the berry colour, but not its shape or size, in a density-dependent manner. Because green berries suffered less from bird foraging, we believe that this colour manipulation helps the wasps to avoid being killed by the birds. The present study indicates that manipulation by wasps may reduce the level of mutualism between the tree and seed-dispersing birds.

Sex change has been recognised in some plants. The sex change in dioecious woody plants includes a monoecious state, which means that the timing of the change is synchronous in branches or shoots but not synchronous among plants, such as in partial dioecious woody plants. To determine whether such a phenomenon is common in woody plants, we investigated the sex expression in two populations of the bird-dispersed, dioecious woody plant Ilex integra Thunb. (Aquifoliaceae), one of the climax forest species in East Asia, over 3 years. No monoecious trees or hermaphrodite flowers were observed. Sex changes were observed at low frequencies, i.e., both female to male and male to female. This study suggests that I. integra may be a complete dioecious woody plant species and that the sex change occurs at the individual level, indicating that the timing of sex change is synchronous in an individual tree.

Bursaphelenchus xylophilus causes pine wilt disease in susceptible pine species. To determine whether B. xylophilus persists in forests containing Abies sachalinensis and Picea jezoensis, we performed inoculation experiments using ten or 15 seedlings of A. sachalinensis, P. jezoensis, Pinus densiflora and P. thunbergii. Inoculation of 15 000 B. xylophilus caused 20 and 7% mortality in A. sachalinensis and P. jezoensis seedlings, respectively, in the nursery, indicating low susceptibility of A. sachalinensis and a lack of susceptibility of P. jezoensis. By contrast, B. xylophilus caused 40 and 70% mortality in P. densiflora and P. thunbergii, respectively. Bursaphelenchus xylophilus was recovered at extremely low densities from stems of externally asymptomatic seedlings of the first two tree species 9.5 months after inoculation. Inoculation of B. xylophilus on excised stem sections of A. sachalinensis and P. jezoensis seedlings showed a greater increase in population density in the bark than xylem after 3 weeks, whereas that on P. densiflora stem sections showed no difference in nematode density between the two tissues. The results suggest that B. xylophilus may persist in forests containing A. sachalinensis and P. jezoensis, when there are insect vectors with affinity for the nematode.

To determine the relationship between resistance to pine wilt disease and the inhibition of nematode systemic dispersal in Pinus densiflora, a suspension of 200 Bursaphelenchus xylophilus was placed on the upper cut end of 5-cm-long, living or boiled branch sections of 17 clones of pine that had different resistance levels. Significantly more nematodes passed through boiled sections than living sections during 24 h. Living branches of the resistant P. densiflora clone group significantly suppressed the dispersal of B. xylophilus compared with those of the susceptible group, suggesting that the inhibition of nematode systemic dispersal was involved in the resistance mechanism of selected disease-resistant pine clones. However, there was no significant correlation between the resistance class and the mean number of nematodes passing through live branch sections within the resistant clone group. The reason for the lack of correlation is discussed in relation with the resistance mechanism.

To determine the likelihood of introgressive hybridisation between Bursaphelenchus xylophilus and B. mucronatus, interbreeding experiments were performed. Of 240 trials involving 24 reciprocal crosses (ten trials per cross, ten virgin females and ten virgin males per trial) between three B. xylophilus isolates and four B. mucronatus isolates under conditions that allowed F1 hybrids to backcross with parents, 174 produced hybrid juveniles. The others failed to generate juveniles. Twenty trials in four crosses successfully established populations which persisted for 65.8-179.5 days, corresponding to 10-40 generations. Analyses of ribosomal DNA (rDNA) fragments for 20 hybrid-derived populations and analysis of inheritance mode of mitochondrial DNA indicated that three crosses produced populations whose members had the B. mucronatus nuclear genome and B. mucronatus or B. xylophilus cytoplasm, whereas the remaining interbreeding produced populations in which 79% of members examined had the B. xylophilus rDNA and cytoplasm and 17% had rDNA fragments of the two species. Populations composed of heterospecific cytonuclear individuals took longer to eat up Botrytis cinerea mycelial mat in a rearing flask than others. Formation of a tail tip mucro in adult females, a characteristic of B. mucronatus, was suggested to be controlled by nuclear and cytoplasmic genes. The results suggested that the introgression of B. xylophilus genes into the B. mucronatus genome was easier than introgression in the opposite direction.

Carabid beetles can escape from pitfall traps. To determine whether or not a female-biased sex ratio for catches of Carabus lewisianus Breuning from pitfall traps could be explained by sexual differences in the rates of capture by and escape from traps, we performed two laboratory experiments to estimate these rates using mathematical models. The results indicated that the two sexes dropped into the pitfall traps at almost equal rates (0.0456/min for females and 0.0366/min for males) and that males escaped from the traps at rates of 0.0591 and 0.0889/min, but no females managed to escape. The bias of escape rates in the field is shown by the higher captures of female C. lewisianus by pitfall traps. Calibration of capture and escape rates suggested similar activity densities of the two sexes in the field. Development of more accurate measures of the number of the relevant sex on the soil surface, and of the methodology for estimating the values of parameters in the field is discussed.

It is speculated that the wasps that attack the seeds of gymnosperm trees (conifers) before they are fertilized can induce unfertilized seeds to accumulate storage material whereas those that attack after the seeds are fertilized selectively oviposit in fertilized seeds. Moreover, in the case of the wasps that oviposit after fertilization of seed, the presence of unfertilized seeds and seedless fruit may increase plant fitness via reduced parasitism of the viable seed. To determine the relationship between the two strategies, host manipulation or selective oviposition, and the time of fertilization of the seeds of angiosperm host plants, fertilized seed of Ilex integra Thunb. was dissected out of berries either immediately after the flight of the seed wasp Macrodasyceras hirsutum Kamijo in the field or the death of adults in the laboratory. The wasps oviposited mostly in fertilized seeds and rarely in unfertilized seeds. Unfertilized seeds, produced by flowers enclosed in pollen exclusion bags, and then exposed to wasps did not contain immature wasps or storage material, which indicates that the wasp did not oviposit in unfertilized seeds. These results support the above mentioned hypothesis and indicate that the substantial proportion of seedless berries do not function as an egg sink.

Most wood-feeding insects need an association with microbes to utilize wood as food, and some have special organs to store and convey the microbes. We report here the discovery of the microbe-storage organ (mycangium) in stag beetles (Coleoptera: Lucanidae), which develop in decayed wood. The mycangium, which was discovered in the abdomen, is present in all adult females of 22 lucanid species examined in this study, but absent in adult males. By contrast, adult insects of both sexes of selected Passalidae, Geotrupidae, and Scarabaeidae, which are related to Lucanidae, lacked mycangia similar to those of the lucanid species. Yeast-like microbes were isolated from the mycangium of five lucanid species. DNA sequence analyses indicate that the microbes are closely related to the xylose-fermenting yeasts Pichia stipitis, Pichia segobiensis, or Pichia sp. known from the gut of a passalid species.