Spatial pattern of pine wilt disease caused byBursaphelenchus xylophilus (Nematoda: Aphelenchoididae) within aPinus thunbergii stand

To understand the mechanism of spread of pine wilt disease caused by the pinewood nematode, Bursaphelenchus xylophilus, which is vectored by a cerambycid, Monochamus alternatus, the spatial distribution of trees weakened by the nematode was examined within a Pinus thunbergii stand from June to October for 4 years. The weakened trees were distributed in a clumped pattern in 1980 and 1981, at an early stage of infestation. In many cases, they showed a double-clumped pattern. The degree of aggregation was higher in June or July than after August. They were uniformly distributed in June or July 1982 and in June 1983 whereas they showed a double-clumped pattern after August. The trees were frequently weakened in June or July when they were near the trees weakened during the previous year. At quadrat sizes of more than 25 m², spatial overlapping was pronounced between trees weakened during June–July of the current year and those weakened in the previous year. The seasonal changes in spatial distribution of weakened trees were explained by the interaction among M. alternatus, B. xylophilus and Pinus trees.     

Yearly changes in dispersal and life-history traits of Monochamus alternatus Hope with reference to its outbreak

The Japanese pine sawyer, Monochamus alternatus Hope, is the primary vector of the pinewood nematode, Bursaphelenchus xylophilus (Steiner et Buhrer) Nickle, the causative agent of pine wilt disease in East Asia. The range of B. xylophilus expands through the dispersal capability of its vectors and transport of host trees infested with the pathogenic nematode and its vector. Outbreaks of M. alternatus populations occur together with the epidemics of pine wilt disease, because the insect reproduces on host trees recently killed by the disease. We measured some dispersal and life-history traits of adults for four years to determine the change in flight capability and life history of a field population of beetles in relation to an outbreak. The population monitored exhibited an outbreak and subsequent collapse. The greatest mean body mass, largest area of hind wings, smallest wing load, and shortest preoviposition period were observed in the year of outbreak. By contrast, there was no difference in the ovariole number between pre-outbreak (latent) and outbreak years. The greatest mean hind wing area and smallest wing load suggest likely result in greater flight performance. As other studies showed, adult body mass is related positively to the flight performance and oviposition rate. Moreover, a shortened preoviposition period leads to a high reproduction rate. Thus, adults in outbreak populations are “superdispersers” because they are likely to have enhanced flight capability and reproduction power. This suggests that M. alternatus populations at the onset of a population outbreak enhance the expansion rate of B. xylophilus range more than those during the latent and pre-outbreak periods.     

金龟子绿僵菌在森林土壤中的分布及对松墨天牛致病性测定

松墨天牛是重大森林植物检疫性病害——松材线虫病的主要媒介昆虫。本研究于2005年8月～2006年8月从福建、江西两省共110个林分样区(其中松林88个样区)采集土壤样品330份,采用选择性培养基分离土壤中的金龟子绿僵菌。从21个样区的26份土样中分离出的金龟子绿僵菌占采集样区的19．1％和样品的7．9％,成菌落数(CFU)500～72500CFU/g,表明金龟子绿僵菌在森林土壤中有较为广泛的分布。对分离到的9个产孢量高的菌株,采用浸渍法(1×10~7孢子/mL)接种3～4龄健康松墨天牛幼虫,采用跗节接种法接种2～15日龄健康成虫,测定其致病力。结果表明,MaYTTR-03、MaYTTR-04菌株对松墨天牛幼虫和成虫均有较高致病力,表现出良好的生防潜力。     

金龟子绿僵菌在森林土壤中的分布及对松墨天牛致病性测定

松墨天牛是重大森林植物检疫性病害——松材线虫病的主要媒介昆虫。本研究于2005年8月～2006年8月从福建、江西两省共110个林分样区（其中松林88个样区）采集土壤样品330份,采用选择性培养基分离土壤中的金龟子绿僵菌。从21个样区的26份土样中分离出的金龟子绿僵菌占采集样区的19.1%和样品的7.9%,成菌落数（CFU）500～72500CFU/g,表明金龟子绿僵菌在森林土壤中有较为广泛的分布。对分离到的9个产孢量高的菌株,采用浸渍法（1×10⁷孢子/mL）接种3～4龄健康松墨天牛幼虫,采用跗节接种法接种2～15日龄健康成虫,测定其致病力。结果表明,MaYTTR-03、MaYTTR-04菌株对松墨天牛幼虫和成虫均有较高致病力,表现出良好的生防潜力。     

Temperature Effects on the Transmission of Bursaphelenchus xylophilus (Nemata: Aphelenchoididae) by Monochamus alternatus (Coleoptera: Cerambycidae)

The object of this research was to investigate the effects of ambient temperature on the transmission of the pinewood nematode, Bursaphelenchus xylophilus, by its vector, Monochamus alternatus. Ninety M. alternatus were reared individually at one of three constant temperatures (16, 20, and 25 °C). As the ambient temperature decreased from 25 to 16 °C, longevity of vectors decreased, nematode transmission efficiency decreased, and the peak period of nematode transmission was delayed and its peak height decreased. Low temperature may inhibit the transmission process, and this inhibition could be partially responsible for preventing pine wilt disease from devastating pine forests in cool regions.     

Optimization of a solid culture medium based on Monochamus alternatus for Cordyceps militaris fruiting body formation

Monochamus alternatus (Coleoptera: Cerambycidae; M. alternatus), popularly known as the Japanese pine sawyer, is a vector of pinewood nematode (Bursaphelenchus xylophilus) that causes pine wilt disease. A solid medium culture with M. alternatus produced Cordyceps militaris fruiting bodies with the longest strips and the highest biological efficiency. Supplementing the original form of M. alternatus with oats resulted in slightly enhanced fruiting body production. The original form of M. alternatus showed higher production than its powder form. The solid culture medium was optimized using a response surface methodology, and the optimal medium contained the following: 8·5 g per bottle of M. alternatus and 11·5 g per bottle of oats mixed with 22·4 ml of water in a 300-ml cylindrical plastic bottle. The optimal culturing period for the fruiting body formation was 37·1 days. Under these conditions, a fruiting body dry weight of 38·0 g per bottle (actual value) was attained. The fruiting body produced using a solid culture medium based on M. alternatus had a cordycepin content of about 25 µg g⁻¹. The solid culture medium containing M. alternatus is highly efficient and eco-friendly, and its effectiveness in large-scale fruiting body production from C. militaris has been demonstrated.     

Population dynamics of Monochamus carolinensis (Col., Cerambycidae) under laboratory conditions

The study of life history dynamics of the pine sawyer beetle Monochamus carolinensis (Olivier) is of interest because the beetles are vectors for the pinewood nematode, Bursaphelenchus xylophilus (Steiner and Buhrer) Nickle, the causal agent of pine wilt in susceptible pine trees. Previous observation of a laboratory colony of M. carolinensis suggested numbers of adult beetles reared from logs varied little in relation to oviposition densities among individual logs. Logs in laboratory colonies are routinely subjected to artificially high oviposition rates. The objectives of this study were to examine the relationship between initial oviposition rate, holding time (time between log cut and oviposition), log volume and area and adult survivorship in laboratory colony. This study examined the number of eggs, larvae, adults and generation survivorship in logs over a 2‐year period. The number of adults emerged per log was not related to the number of eggs laid. Late larval and adult numbers were significantly correlated with log size. Only 12% of the initial cohort completed development and emerged as adults. We attribute the high within‐log mortality of beetles to intraspecific competition and cannibalism.     

浙江省余姚地区松褐天牛的生活世代

松褐天牛Monochamus alternatus Hope是松材线虫(Bursaphelenchus xylophilus)病的主要传播媒介,明确松褐天牛的发生世代及成虫羽化规律对于松材线虫病的防控有着重要意义。本研究利用"塑料薄膜包裹松树树干、定期解除包裹供天牛产卵"的方法,系统研究了余姚地区松褐天牛的产卵规律及成虫的羽化规律。研究结果表明,成虫产卵期为5月上旬至8月下旬,高峰期出现在6月下旬至8月中旬。松褐天牛在浙江余姚多数为1年1代,部分为2年1代,95%以上的成虫集中在产卵后的第2年羽化,少数在第3年羽化。成虫自5月上旬开始羽化,高峰期出现在5月中旬到6月中旬,7月中旬羽化基本结束。     

Transmission of Bursaphelenchus xylophilus between Monochamus alternatus and Monochamus saltuarius through interspecific mating behaviour

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.     

Canopy-related adult density and sex-related flight activity of Monochamus alternatus (Coleoptera: Cerambycidae) in pine stands

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.     

Transmission of the Pinewood Nematode, Bursaphelenchus xylophilus,to Slash Pine Trees and Log Bolts by a Cerambycid Beetle,Monochamus titillator, in Florida

Field-collected adults of the southern pine sawyer, Monochamus titillator (F.) (Coleoptera: Cerambycidae), naturally infested with fourth-stage juveniles (dauerlarvae) of the pinewood nematode, Bursaphelenchus xylophilus (Steiner and Buhrer, 1934) Nickle, 1970, were maturation fed on excised shoots of typical slash pine, Pinus elliottii Engelm. var elliottii, for 21 days. During August 1981, a male and female adult beetle were held in a sleeve cage placed on the terminal of a side branch of each of seven replicate, healthy 10-year-old slash pine trees. All seven branch terminals showed evidence of beetle feeding on the bark after 1 week, and pinewood nematodes were present in wood samples taken near these feeding sites. Four of the seven trees showed wilt symptoms in 4-6 weeks and died about 9 weeks after beetle feeding. Pinewood nematodes were recovered from the roots and trunks of the dead trees. Each of seven replicate slash pine log bolts was enclosed in a jar with a pair of the same beetles used in the sleeve cages. After 1 week, wood underlying beetle oviposition sites in the bark of all replicate log bolts was infested with the pinewood nematode.     

Pine wilt disease in Yunnan,China: Evidence of non-local pine sawyer Monochamus alternatus (Coleoptera: Cerambycidae) populations revealed by mitochondrial DNA

Abstract Monochamus alternatus (Hope) specimens were collected from nine geographical populations in China, where the pinewood nematode Bursaphelenchus xylophilus (Steiner et Buhrer) was present. There were seven populations in southwestern China in Yunnan Province (Ruili, Wanding, Lianghe, Pu’er, Huaning, Stone Forest and Yongsheng), one in central China in Hubei Province (Wuhan), and one in eastern China in Zhejiang Province (Hangzhou). Twenty-two polymorphic sites were recognized and 18 haplotypes were established by analyzing a 565 bp gene fragment of mitochondrial cytochrome oxidase subunit II (CO II). Kimura two-parameter distances demonstrated that M. alternatus populations in Ruili, Wanding and Lianghe (in southwestern Yunnan) differed from the other four Yunnan populations but were similar to the Zhejiang population. No close relationship was found between the M. alternatus populations in Yunnan and Hubei. Phylogenetic reconstruction established a neighbor-joining (NJ) tree, which divided haplotypes of southwestern Yunnan and the rest of Yunnan into different clades with considerable bootstrapping values. Analysis of molecular variance and spatial analysis of molecular variance also suggested significant genetic differentiation between M. alternatus populations in southwestern Yunnan and the rest of Yunnan. Our research suggests that non-local populations of M. alternates, possibly from eastern China, have become established in southwestern Yunnan.     

三峡库区松材线虫病扩张速度对人为活动的响应

松材线虫病是我国森林重要的检疫性病害,除自然媒介天牛传播,人为因素介导的传播在松材线虫病扩散和蔓延中起着至关重要的作用。选择在我国松材线虫病的重点发生区域三峡库区,研究马尾松林景观格局和以人为活动强度为主要因子的松材线虫病扩散规律,尝试揭示松材线虫病扩散机制。结果显示三峡库区的疫点数在近年有上升趋势,各地区发病率得到一定控制;在疫区和非疫区,马尾松林斑块聚合度最大的是非疫区的大渡口,为98.6406;斑块分离度最大的是非疫区的秭归,为0.9318;方差分析结果显示马尾松林景观格局和松材线虫病之间没有相关性。进一步研究了松材线虫病和人为活动强度的联系,结果显示松材线虫病发病率和地区路网密度、人口总量具有较强的相关性,和地区GDP、货运周转量相关性小。宜昌的松材线虫病发病率和路网密度的相关系数为0.985,和人口总数的相关系数为0.866;重庆的松材线虫病发病率和路网密度相关系数为0.924,和人口总数相关系数为0.999;其次基于路网密度和人口总数,对三峡库区松材线虫病疫情在未来十年进行了预测,结果显示随着路网密度和人口总数的扩增,宜昌松材线虫病也将呈现逐步增大的趋势,R~2值为0.919和0.637;重庆发生趋势一致,R~2值为0.976和0.992。     

Individual-based modeling of the spread of pine wilt disease: vector beetle dispersal and the Allee effect

Pine wilt disease is caused by the pinewood nematode Bursaphelenchus xylophilus, which is vectored by the Japanese pine sawyer beetle Monochamus alternatus. Due to their mutualistic relationship, according to which the nematode weakens and makes trees available for beetle reproduction and the beetle in turn carries and transmits the nematode to healthy pine trees, this disease has resulted in severe damage to pine trees in Japan in recent decades. Previous studies have worked on modeling of population dynamics of the vector beetle and the pine tree to explore spatial expansion of the disease using an integro-difference equation with a dispersal kernel that describes beetle mobility over space. In this paper, I revisit these previous models but retaining individuality: by considering mechanistic interactions at the individual level it is shown that the Allee effect, an increasing per-capita growth rate as population abundance increases, can arise in the beetle dynamics because of the necessity for beetles to contact pine trees at least twice to reproduce successfully. The incubation period after which a tree contacted by a first beetle becomes ready for beetle oviposition by later beetles is crucial for the emergence of this Allee effect. It is also shown, however, that the strength of this Allee effect depends strongly on biological mechanistic properties, especially on beetle mobility. Realistic individual-based modeling highlights the importance of how spatial scales are dealt with in mathematical models. The link between mechanistic individual-based modeling and conventional analytical approaches is also discussed.     

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.     

Synergistic attraction of pine sawyer Monochamus saltuarius (Coleoptera: Cerambycidae) to monochamol and α‐pinene

Monochamus (Coleoptera: Cerambycidae) species are longhorn pine sawyers that serve as insect vectors of the pinewood nematode Bursaphelenchus xylophilus (Nematoda: Parasitaphelenchidae), which are responsible for debilitating pine wilt disease. An aggregation pheromone, 2‐(1‐undecyloxy)‐1‐ethanol (hereafter referred to as monochamol), was shown to be effective at attracting Monochamus species. However, attraction of the pine sawyers to aggregation pheromones varied depending on semiochemicals, including host plant volatiles and kairomones. In this study, we investigated the abilities of monochamol and the host‐plant volatiles α‐pinene and ethanol to attract M. saltuarius in a pine forest in Cheongsong, Gyeongsangbuk‐do, Korea. A total of 91 M. saltuarius (28 males and 63 females) were captured. The combination of monochamol (700 mg) with α‐pinene and ethanol exhibited a synergistic effect on attracting M. saltuarius (11.0 beetles per trap), whereas monochamol alone and a mixture of α‐pinene and ethanol resulted in the capture of 3.2 beetles and 3.6 beetles per trap, respectively. Our results suggest that multi‐funnel traps baited with a blend of monochamol, α‐pinene and ethanol are highly effective for monitoring M. saltuarius and M. alternatus in pine forests.     

Changes in spatial and temporal distributions of Monochamus beetles along the fire severity in burned Pinus densiflora forests

Wildfire is an important disturbance factor in forest ecosystem and could affect the distribution of insects. Because Monochamus beetles are known vector insects of pine wood nematodes in Korea and burnt pine trees can be used as oviposition habitats for Monochamus beetles, we investigated changes in the spatial (among fire severity classes) and temporal (between years) distributions of M. alternatus and M. saltuarius. From late May 2017 to early October 2018, four multi-funnel traps with pheromone lure were installed in each fire severity class (unburned, light, light-moderate edge, moderate-severe edge, and severe). All traps were replaced fortnightly from May to October for two years. The spatial distribution of the two Monochamus beetles was significantly affected by fire severity. In particular, M. alternatus showed higher catches in the severely burned sites and responded positively to the fire. A similar number of M. saltuarius individuals were observed between 2017 and 2018, while more M. alternatus were caught in 2018 compared to 2017. Consequently, it was confirmed that the dead wood of P. densiflora caused by the wildfire had a great influence on the density of insects, especially M. alternatus.     

松墨天牛对寄主树木的产卵选择

在室内采用选择行为方法,研究了松墨天牛Monochamus alternatus对几种松属植物的产卵选择行为。结果表明,松墨天牛对黑松Pinus thunbergii的选择性最强,对马尾松P.massoniana的选择性最弱,二者间差异显著,它们与火炬松P. taeda和湿地松P. elliottii间无明显差异。在黑松和马尾松的被松材线虫危害木与健康木之间,松墨天牛明显选择被害木产卵;具产卵痕枝段和虫粪处理过的枝段,对松墨天牛的产卵有显著抑制作用,松墨天牛明显地选择对照枝段产卵。     

Niche‐based relationship between sympatric bark living insect pests and tree vigor decline of Pinus yunnanensis

Forest stands of Pinus yunnanensis Franch in southwestern China are seriously damaged by several bark living insect pests. These pests commonly exist in the same host tree and exploit limited phloem resources. In this study, we hypothesized that sympatric pests would occupy different ecological niches to reduce interspecific competition, and their coordinated attack would aggravate the tree vigor decline of P. yunnanensis. To quantify the ecological niches, we used a sampling method involving three dimensional divisions of tree resource states: (a) sample plot dimension (to evaluate the extent of pest colonization at plot scale), (b) trunk height dimension, and (c) tree vigor dimension. Those attacked pine canopies and colonized trunks were analysed in the field study. The results showed that Tomicus minor and Tomicus yunnanensis both widely and uniformly distributed in lightly, moderately and heavily damaged canopies while they aggregated on particular trunks of dying and withered trees. Lower and middle trunk sections (1–4 m) were mainly occupied by Monochamus alternatus and T. minor, yet T. yunnanensis dominated the upper parts (5–6 m). There was an overlap of the pine shoot beetles in the middle sections (2–4 m), whereas all three species were ecologically segregated in other trunk sections. During the shoot feeding phase, tree vigor declined with pine shoot beetles' increasing shoot feeding density. They coexisted in the same host tree, while M. alternatus only attacked dying and withered tree trunks, causing a greater infestation. Colonization of pine shoot beetle then followed by M. alternatus could be the major causes of tree mortality. This study highlights the resource utilization pattern of sympatric bark living insect pests corresponding with tree vitality. Those findings would help to understand the mechanisms of pest outbreak in P. yunnanensis ecosystem and provide potential guidance for developing an early monitoring pest warning system.     

金龟子绿僵菌MaYTTR-04菌株对松墨天牛成虫的致病力

松墨天牛Monochamus alternatus Hope是重大森林植物检疫性病害——松材线虫病的主要媒介昆虫。采用跗节接种法测定金龟子绿僵菌Metarhizium anisopliae MaYTTR-04菌株对松墨天牛成虫的致病力。结果表明： 在恒温条件下,松墨天牛第6天开始死亡,第18天～21天为死亡高峰期,每头成虫接种孢子量为2.3×10⁶±0.2×10⁶时成虫的死亡率第18天达85%,第21天达95%; 时间效应指标值LT₅₀为14.7天。室内自然变温条件下,松墨天牛第3天出现死亡,第15天～21天为死亡高峰期,每头成虫接种孢子量为2.3×10.6±0.2×10⁶时的死亡率在第15天达85%,第21天达100%;时间效应指标值LT₅₀为12.9天。林间套笼21天后,菌液处理的成虫平均死亡率为60%,僵虫率为48.9%;而通过无纺布菌条的成虫死亡率为86.7%,僵虫率为75.6%。表明MaYTTR-04菌株对松墨天牛成虫有强的致病力,且在较高变温环境温度下致病力更强。结果说明该菌株可作为生产性菌株应用于林间防治松墨天牛。