Genotype distribution of Raffaelea quercivora in the oak galleries and its composition in the mycangia of Platypus quercivorus

Raffaelea quercivora is the pathogenic fungus that causes Japanese oak wilt. The female monogynous ambrosia beetle, Platypus quercivorus, carries this fungus in mycangia on the pronotum. These beetles bore galleries in oak trees with their partners to produce offspring, and they deposit fungus on the gallery walls from their mycangia. The offspring mature in the gallery, before loading the fungal pathogen and flying from the gallery to other healthy trees. To investigate the unloading and loading modes of the fungus within the gallery, we developed four polymorphic microsatellite markers for R. quercivora and identified the fungal genotypes in the galleries and mycangia of the beetles. Small wood chips were sampled at 5–10‐mm intervals from the walls of five galleries in a dead Quercus serrata tree. The pronota were also sampled from five female adult beetles. The genotypes of the R. quercivora isolates from the wood chips and pronota were identified using the microsatellite makers. The genotypic analysis showed that each gallery was inhabited patchily by 5–10 genotypes of R. quercivora, and the mycangia of each beetle contained 3–6 genotypes. These results indicate that diverse R. quercivora genotypes are unloaded repeatedly from the mycangia of female beetles onto the gallery wall, which results in their patchy distribution on the walls. When the offspring leave the host tree, the fungal clones that proliferate in the walls are also loaded repeatedly into the mycangia of the mature beetles.     

Aggregation ofPlatypus quercivorus (Murayama) (Coleoptera: Platypodidae) on oak logs bored by males of the species

To provide evidence for the aggregation ofPlatypus quercivorus (Murayama) (Coleoptera: Platypodidae) in association with the mass mortality of oak trees in Japan, we compared the number of beetles attracted to oak logs bored by males with the number of beetles attracted to logs that had not been bored. Large numbers of males and females were attracted to the bored logs, and the total numbers increased as the number of entry holes in the logs increased. This result shows that both male and female beetles aggregate on logs bored by the males. We collected a large number of beetles attracted to living trees bored by the beetles, and the total number of beetles collected increased as the number of entry holes/m² on the tree trunk increased. Beetles also aggregated on living trees bored by the males.     

The survival of Ceratocystis fagacearum in South Carolina

This research, which involved observing and sampling artificially inoculated oak-wilted turkey oak trees, substantiates previous field observations that C. fagacearum does not quickly kill oak trees in South Carolina. C. fagacearum seems to quickly lose virulence during the first growing season following inoculation. Rather than being quickly killed, as occurs with infected red oaks in more temperate climates, oak trees in South Carolina either survive infection or die much more slowly. Moisture, total carbohydrate, and reducing sugar contents of infected trees indicate that these trees suffer an initial physiologic trauma. Branches die first, followed by upper stems, and in few cases lower stems. Some trees, however, have survived inoculation, and initial partial wilting, but have since recovered.     

Discrimination among host tree species by the ambrosia beetle Platypus quercivorus

The ambrosia beetle Platypus quercivorus is a vector for the fungus that causes Japanese oak wilt, and susceptibility to infestation by P. quercivorus varies by tree species. We postulated that P. quercivorus discriminates among host tree species differing in susceptibility to attack. To test this postulate, we counted the number of flying male beetles (NFM), the number of holes bored by male beetles (NH), and the number of flying female beetles (NFF) per unit area of bark surface in three fagaceous tree species: Quercus crispula (with high susceptibility to infestation) and Q. serrata and Castanea crenata (both with low susceptibility). NFM and NH were used to calculate the proportion of male beetles that bored holes out of those that flew to the tree (PBM). We used generalized additive models to predict NFM, NFF, and PBM. The locations of trees, expressed as x and y coordinates, numbers of weeks after the first male beetle’s flying (WEEK), diameters of trees 130 cm above ground (DBH), and tree species (SP) were incorporated into the models as candidate explanatory variables. The best-fit models for NFM and NFF included WEEK and DBH and the effect of location; SP was not included in the models. For PBM, the best-fit model included WEEK, DBH, and SP. The results indicate that male P. quercivorus prefer Q. crispula to Q. serrata and C. crenata and that selection is made before boring holes on trees, but that P. quercivorus do not discriminate among host species when they fly to trees.     

Oak mortality associated with crown dieback and oak borer attack in the Ozark Highlands

Oak decline and related mortality have periodically plagued upland oak–hickory forests, particularly oak species in the red oak group, across the Ozark Highlands of Missouri, Arkansas and Oklahoma since the late 1970s. Advanced tree age and periodic drought, as well as Armillaria root fungi and oak borer attack are believed to contribute to oak decline and mortality. Declining trees first show foliage wilt and browning, followed by progressive branch dieback in the middle and/or upper crown. Many trees eventually die if severe crown dieback continues. In 2002, more than 4000 living oak trees ≥11 cm dbh in the relatively undisturbed mature oak forests of the Missouri Ozark Forest Ecosystem Project (MOFEP) were randomly selected and inventoried for tree species, dbh, crown class, crown width, crown dieback condition (healthy: <5% crown dieback, slight: >5–33%, moderate: 33–66%, and severe: >66%) and number of emergence holes created by oak borers on the lower 2.4 m of the tree bole. The same trees were remeasured in 2006 to determine their status (live or dead). In 2002, about 10% of the red oak trees showed moderate or severe crown dieback; this was twice the percentage observed for white oak species. Over 70% of trees in the red oak group had evidence of oak borer damage compared to 35% of trees in the white oak group. There was significant positive correlation between crown dieback and the number of borer emergence holes (p < 0.01). Logistic regression showed oak mortality was mainly related to crown width and dieback, and failed to detect any significant link with the number of oak borer emergence holes. Declining red oak group trees had higher mortality (3 or 4 times) than white oaks. The odds ratios of mortality of slightly, moderately, and severely declining trees versus healthy trees were, respectively, 2.0, 6.5, and 29.7 for black oak; 1.8, 3.8, and 8.3 for scarlet oak; and 2.6, 6.5 and 7.1 for white oaks.     

The role of isolated old oak trees in maintaining beetle diversity within larch plantations in the central mountainous region of Japan

Old oak trees (Quercus crispula Blume) that are remnants of former old-growth forests have been isolated singly or as small patches within a matrix of conifer plantations in the central mountainous region of Japan. Fifty-six aerial Malaise traps were deployed around seven isolated oak trees within larch [Larix kaempferi (Lamb.) Carrière] plantations and at seven larch plots within larch plantations. The species richness and composition of beetles (Cerambycidae, Curculionidae, Elateridae, Chrysomelidae, and Lycidae) around the oak trees were compared to those in the larch plantations. Species richness was higher around the oak trees than in the larch plantations, and the species composition differed. A number of saproxylic beetles were characteristic of isolated old oak trees. These results emphasize the importance of isolated old oak trees for maintaining beetle diversity in larch plantations and raise the possibility that further losses of these isolated oak trees could eliminate many individuals and reduce beetle diversity in larch plantations.     

Reduction of isoprene emissions from live oak (Quercus fusiformis) with oak wilt

Many plants emit isoprene, a hydrocarbon that has important influences on atmospheric chemistry. Pathogens may affect isoprene fluxes, both through damage to plant tissue and by changing the abundance of isoprene-emitting species. Live oaks (Quercus fusiformis (Small) Sarg. and Q. virginiana Mill) are major emitters of isoprene in the southern United States, and oak populations in Texas are being dramatically reduced by oak wilt, a widespread fungal vascular disease. We investigated the effects of oak wilt on isoprene emissions from live oak leaves (Q. fusiformis) in the field, as a first step in exploring the physiological effects of oak wilt on isoprene production and the implications of these effects for larger-scale isoprene fluxes. Isoprene emission rates per unit dry leaf mass were 44% lower for actively symptomatic leaves than for leaves on healthy trees (P = 0.033). Isoprene fluxes were significantly negatively correlated with rankings of disease activity in the host tree (fluxes in leaves on healthy trees > healthy leaves on survivor trees > healthy leaves on the same branch as symptomatic leaves > symptomatic leaves; isoprene per unit dry mass: Spearman's rho = -0.781, P = 0.001; isoprene per unit leaf area: Spearman's rho = -0.652, P = 0.008). Photosynthesis and stomatal conductance were reduced by 57 and 63%, respectively, in symptomatic relative to healthy leaves (P < 0.05); these reductions were proportionally greater than the reductions in isoprene emissions. Low isoprene emission rates in symptomatic leaves are most simply explained by physiological constraints on isoprene production, such as water stress as a result of xylem blockage, rather than direct effects of the oak wilt fungus on isoprene synthesis. The effects of oak wilt on leaf-level isoprene emission rates are probably less important for regional isoprene fluxes than the reduction in oak leaf area across landscapes.     

The oak buprestid beetle,Agrilus biguttatus (F.) (Col., buprestidae), a recent factor in oak decline in Europe

During last years, the populations of the oak buprestid beetleAgrilus biguttatus (Fab.) in Europe have increased. In several countries, the beetles’ attacks have become a new component in oak dieback. The larvae excavate galleries under the bark of weakened trees, which are killed through the girdling. Because of the difficult diagnosis it is necessary to draw more attention to the infestations. Therefore, this paper presents a review on the literature concerning the recognition of the infestations by the insect and its bionomics.     

Factors affecting post-fire crown regeneration in cork oak (Quercus suber L.) trees

Cork oak (Quercus suber) forests are acknowledged for their biodiversity and economic (mainly cork production) values. Wildfires are one of the main threats contributing to cork oak decline in the Mediterranean Basin, and one major question that managers face after fire in cork oak stands is whether the burned trees should be coppiced or not. This decision can be based on the degree of expected crown regeneration assessed immediately after fire. In this study we carried out a post-fire assessment of the degree of crown recovery in 858 trees being exploited for cork production in southern Portugal, 1.5 years after a wildfire. Using logistic regression, we modelled good or poor crown recovery probability as a function of tree and stand variables. The main variables influencing the likelihood of good or poor crown regeneration were bark thickness, charring height, aspect and tree diameter. We also developed management models, including simpler but easier to measure variables, which had a lower predictive power but can be used to help managers to identify, immediately after fire, trees that will likely show good crown regeneration, and trees that will likely die or show poor regeneration (and thus, potential candidates for trunk coppicing).     

Occurrence of Phytophthora cinnamomi in cork oak forests in Italy

An increasing decline and mortality of cork oak trees have been recently observed in central Italy and Sardinia Island. Following surveys conducted in three declining cork oak forests, a Phytophthora species was consistently isolated from soil samples collected from trees displaying different level of decline. Based on morphological features, growth rates at different temperatures and analysis of DNA sequences of the ITS region, all isolates were identified as Phytophthora cinnamomi Rands. This pathogen caused large brownish lesions on inoculated freshly cut branches of cork oak. It was re‐isolated from all infected tissues. These findings represent the first report of P. cinnamomi on cork oak trees in Italy.     

Field trap test for bioassay of synthetic (1<Emphasis Type="Italic">S</Emphasis>,4<Emphasis Type="Italic">R</Emphasis>)-4-isopropyl-1-methyl-2-cyclohexen-1-ol as an aggregation pheromone of <Emphasis Type="Italic">Platypus quercivorus</Emphasis> (Coleoptera: Platipodidae)

An ambrosia beetle, Platypus quercivorus, is a vector of Japanese oak wilt, which causes massive mortality of oak trees in Japan. Previous studies have identified (1S,4R)-4-isopropyl-1-methyl-2-cyclohexen-1-ol ((−)-IMCH) as the major component of the aggregation pheromone of P. quercivorus. We tested the ability of synthetic (−)-IMCH to attract P. quercivorus adults. The synthetic compound attracted 14.4 times as many P. quercivorus, of both sexes, as control traps. Control and (−)-IMCH-baited traps both captured 3.32 times as many males as females, probably because of the nature of Lindgren funnel traps and of male-biased emergence sex ratio early in the season of adult flights. (−)-IMCH was proved to be a major component of the aggregation pheromone of P. quercivorus.     

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

Mass mortality of fagaceous trees caused by Japanese oak wilt has occurred widely in Japan. Although virulence of the causal fungus, Raffaelea quercivora, appeared to differ among isolates, its relation to the fungal growth within trees was unknown. To clarify the differences in fungal virulence against susceptible Quercus crispula, we examined fungal growth of four R. quercivora isolates within trees and the resulting virulence. In our study, the isolates were multiple‐inoculated in seedlings and single‐inoculated in twigs of mature trees. In the multiple‐inoculation test, mortality rates were examined by the observation of external symptoms. In the single‐inoculation test, water conductance and hyphal growth within the trees were examined by applying aqueous dyes and fluorescence microscopy, respectively. Mortality rates, the proportion of the cross‐sectional area comprising non‐conductive sapwood and horizontal hyphal growth differed significantly among the isolates. Univariate logistic regression analyses showed that both the proportion of non‐conductive sapwood and hyphal growth were significantly positively related to mortality rates. For three isolates, hyphal growth was significantly positively correlated with the proportion of non‐conductive sapwood. These results suggested that the virulence against Q. crispula varies among R. quercivora isolates and that the extent of fungal colonization of the tree determines fungal virulence.     

Platypus cylindrus,a vector of Ceratocystis platani in Platanus orientalis stands in Greece

The ambrosia beetle Platypus cylindrus was very common in stands of Platanus orientalis trees in Greece, infesting trees already infected by Ceratocystis platani. The fungus C. platani was isolated from 59% of adult beetles of P. cylindrus collected on the wing in a heavily infected stand of P. orientalis. Transmission of C. platani was demonstrated in 80% of P. orientalis seedlings challenged with P. cylindrus beetles, which were previously fed in cultures of the fungus. In most of the cases, the beetles had bored tunnels in stems, where abundant perithecia of C. platani were observed. Platypus cylindrus adults regularly visited artificially wounded P. orientalis trees in a natural stand. Although this ambrosia beetle normally infests stressed or dead trees, it appears to play a role as a vector of C. platani, especially by visiting wounded trees.     

Integrated management of the charcoal disease by silviculture,chemical and biological methods in forest parks

Biscogniauxia mediterranea (De Not.) Kuntze, is a widespread fungus, causes outbreaks of Charcoal disease on cork oak and dieback thousands oak trees in recent years in Iran. Yet no efficient and effective management methods have been found to control the disease. The present study aimed to provide a suitable method for charcoal disease management through silviculture operations, systemic pesticides and biological compounds. The results showed that, simultaneous implementation of silvicultural practices (scarification+sanitation), use of systemic fungicides and Trichoderma compounds, reduce the number and depth of cankers, increase callus thickness, and ultimately improve health of the diseased trees. But disease severity and fungi activity were increased when scarification operation implements alone. The greatest effectiveness of the treatments and health condition improvement of the diseased trees was observed 12 months after applying the treatments. Then, preventive and therapeutic effectiveness of the treatments reduced over time. Sanitation and scarification practices can increase soil permeability and also destroy the larvae, pupa or adult insects and oak borer beetles that play an important role in disease outbreaks. Therefore, simultaneous implementation of sanitation and scarification practices with application of biological and systemic compounds can reduce primary source of pathogen inoculum and help trees to recover their health.     

Effects of root damage associated with Phytophthora cinnamomi on water relations,biomass accumulation,mineral nutrition and vulnerability to water deficit of five oak and chestnut species

The effects of root damage associated with Phytophthora cinnamomi on water relations, biomass accumulation, mineral nutrition and vulnerability to water deficit were investigated in pedunculate oak (Quercus robur), red oak (Quercus rubra) and holm oak (Quercus ilex) saplings over two years. Comparison was made with sweet chestnut (Castanea sativa), a susceptible species to infection by P. cinnamomi, and with a resistant hybrid chestnut (Castanea crenata × C. sativa). Trees were inoculated in 1998 and were subjected to water shortage in 1999. All inoculated sweet chestnuts died before the application of water shortage. Hybrid chestnut, pedunculate oak and red oak displayed low root susceptibility to P. cinnamomi. In these species, water relations, aerial growth and mineral nutrition were slightly affected by inoculation. By contrast, holm oak was the most susceptible oak species to P. cinnamomi as inoculated well‐watered trees displayed the highest root loss (67%) and a 10% mortality. Root loss was associated with a decrease in predawn leaf water potential, a 61% reduction in stomatal conductance, a 55% reduction in aerial biomass, a decrease in leaf carbon isotope discrimination and reduced leaf N and P contents in comparison with controls. In hybrid chestnut and pedunculate oak, water shortage resulted in a similar decrease of predawn leaf water potential, stomatal conductance and aerial biomass in inoculated and non‐inoculated trees. In red and holm oaks, soil volumetric water content of inoculated trees subjected to water shortage remained high. The effects observed in those trees were similar to those of inoculated well‐watered trees and were probably the result of root infection only.     

Survival of the oak wilt fungus in air-dried lumber

The survival of the oak wilt fungus, Ceratocystis fagacearum (Bretz) Hunt, in air-dried red, white, and scarlet oak lumber was studied in South Carolina and West Virginia. The position of boards in the piles did not have a significant effect on culture recovery of C. fagacearum. The fungus was not recovered after the initial sample at set-up time in the summer harvested oaks in South Carolina nor beyond 28 days in West Virginia. In autumn harvested material recoverability was slightly longer, 28 days in South Carolina with one positive recovery at 70 days, and much longer in West Virginia with some recovery of C. fagacearum occurring at 140 days. Recovery of C. fagacearum from white oak at both locations was generally much lower than from red or scarlet oak during any given sample period.     

The larger,the better? Effects of delayed diameter-limit cutting on old-growth attributes and saproxylic beetle diversity in temperate oak forests

Intensive management implies harvesting large, old trees, which reduces the old-growth attributes. This negatively affects biodiversity, especially saproxylic beetles. In managed temperate oak forests, rotation extension induced by increasing the diameter threshold of final harvest by about 10 cm compared to conventional practices (i.e. DBH around 70 cm) might mitigate this negative effect. Here, we used a gradient of the proportion of overmature trees (DBH?≥?80 cm) among mature trees (DBH?≥?70 cm) across plots of high oak French forests to test the potential effect of increasing diameter threshold on (1) structural features and (2) species composition and diversity of saproxylic beetles communities. We assessed deadwood and microhabitats availability (i.e. volume/density and diversity) and canopy openness in 81 1-ha plots across eleven French forests. Results highlighted that a larger proportion of overmature trees, for a given density of mature trees, had limited effects on structural features: only cavities density showed a significant positive response, with no cascading effect on cavicolous beetles. Moreover, the proportion of overmature trees had no significant effect on the composition of saproxylic beetles communities (and ecological variables altogether explained only 17% of the composition inertia). By contrast, mature tree density enhanced microhabitat density and indirectly increased the abundance of rare species. Thus, shifting DBH from 70 to 80 cm in high oak forest could have no or limited effect on saproxylic beetles conservation. Improved strategies might rather stem from combining (1) longer rotation extension and (2) less intensive management practices in extended rotation stands.     

Isolation and characterization of 10 new microsatellite loci in the ambrosia beetle <Emphasis Type="Italic">Platypus quercivorus</Emphasis>

The ambrosia beetle, Platypus quercivorus, is an insect vector for the causal agent of Japanese oak wilt disease, Raffaelea quercivora. We have developed 10 new polymorphic microsatellite markers from Type 1, one of the genetic types of the beetle, using a biotin-enrichment procedure. Fifty-three adult beetles were used for characterization of the markers. The number of alleles per locus ranged from two to nine (average 4.9). The observed and expected heterozygosity ranged from 0.49 to 0.77 and from 0.44 to 0.80, respectively. None of the loci deviated significantly from Hardy–Weinberg equilibrium. No linkage disequilibrium was detected in any pairwise comparisons of loci. Most of the markers successfully amplified in other genetic types of the beetle. These markers could become useful tools for population genetic studies and for parentage and sibship analysis for this beetle.     

Comparisons of hyphal growth of Raffaelea quercivora among four Japanese Fagaceae species

Japanese oak wilt causes widespread oak mortality in Japan. Possible differences in susceptibility to the causal fungus, Raffaelea quercivora, may be due to vessel arrangements in host trees. To clarify whether constitutive defence mechanisms including vessel arrangements or induced defence mechanisms are the main determinants of host susceptibility, we inoculated the fungus into living seedlings or sterilized stem segments of four Japanese fagaceous species. In seedlings, water conductance was assessed with dye. In both seedlings and stem segments, hyphal growth was examined by fluorescence microscopy. In seedlings, the area of non‐conductive sapwood in stem cross sections and hyphal growth differed significantly among species. In the susceptible species Quercus crispula and Quercus serrata, hyphal growth was significantly and positively correlated with the proportion of non‐conductive sapwood. In stem segments, hyphal growth was not significantly different among species or between vessel arrangements and was similar to or greater than that in seedlings. These results suggest that the extent of sapwood colonization by R. quercivora could be used as a marker for susceptibility and that susceptibility is determined mainly by induced defence responses.     

Identification and distribution of Armillaria species associated with an oak decline event in the Arkansas Ozarks

Forests in the Ozark Mountains of northern Arkansas recently experienced a widespread oak decline event. Armillaria, a root rot fungus, has been associated with other oak decline events and may have been an important contributing factor to tree mortality in this event. Although Armillaria has been identified from the Ozark Mountains in Missouri, it has never been investigated in the Arkansas Ozarks. Molecular diagnostic techniques were used in this study to identify species of Armillaria present on roots removed from dead trees of two common oak species, northern red oak, Quercus rubra L., and white oak, Q. alba L., from three geographic areas and on three topographic positions – ridges, south‐ and west‐facing benches. Armillaria(A. mellea, A. gallica or A. tabescens) was identified from 31% of root samples taken from 102 trees in seven of nine sample plots. Armillaria mellea, occurred most often (20 samples, both oak species on seven plots) followed by A. gallica (10 samples, northern red oak only on four plots), and A. tabescens occurred twice (on northern red oak in a single plot). Thus, all three Armillaria species occurred on northern red oaks while A. mellea was the only species recovered from white oaks. Results varied by topographic position with samples from tree roots on ridges having the fewest positive identifications, one of 29. West‐facing benches had the highest positive samples with 20 of 41 testing positive and trees on south‐facing benches were intermediate with 11 of 32 samples from infected trees. This study documents the occurrence of three species of Armillaria in the Arkansas Ozarks and their association with oak mortality resulting from an oak decline event coupled with a red oak borer, Enaphalodes rufulus, outbreak. Further, it documents some potential variation in host/pathogen combinations and forest site conditions.