Proceedings of a Workshop on Bark Beetle Genetics: Current Status of Research. May 17-18, 1992, Berkeley, California

Home , Bark (botany), Conophthorus ponderosae, Ips paraconfusus

United States Department of Agriculture Proceedings of a Workshop Forest Service Pacific Southwest on Bark Beetle Genetics: Research Station

General Technical Report PSW-GTR-138 Current Status of Research

May 17-18, 1992, Berkeley, California Hayes, Jane L.; Robertson, Jacqueline L, tech. coords. 1992. Proceedings of a workshop on bark beetle genetics: current status of research. May 17-18, 1992, Berkeley, California. Gen. Tech. Rep. PSW-GTR-138. Albany, CA: Pacific Southwest Research Station, Forest Service, U.S. Department of Agriculture; 31 p.

The Proceedings reports the results of a workshop focusing on the topic of bark beetle genetics. The workshop evolved because of the growing interest in this relatively unexplored area of bark beetle research. Workshop participants submitted brief descriptions of their views of the current status of bark beetle genetic research and needs for the future. Contributions vary from broad overviews to detailed descriptions of specific projects. A common theme is the potential for rapid advances in this area of research given recent technological advances in the field of genetics. Also included is an extensive reference list.

Retrieval Terms: Dendroctonus species, Ips species, scolytid genetics, taxonomy

Dedication

We dedicate this report to the memory of Gerald Lanier, a valued colleague to all who share an interest in bark beetle research, and a dear friend of many of the workshop participants. It is particularly fitting that we pay tribute to Gerry for his pioneering efforts in bark beetle genetics and in recognition of his lasting contributions in the area of scolytid systematics.

Technical Coordinators:

Jane L. Hayes is a research entomologist and Project Leader of the Ecology, Behavior, and Management of Southern Pine Beetle Unit of the Southern Forest Experiment Station, USDA Forest Service, 2500 Shreveport Highway, Pineville, LA 71360. Jacqueline L. Robertson is a research entomologist and Project Leader of the Insect Biochemistry, Genetics, and Biometry Unit of the Pacific Southwest Research Station, 800 Buchanan Street, Albany, CA 94710.

Acknowledgments:

We thank Sue Moore for handling the innumerable workshop arrangements and mailings, and for assembling the draft report. We are also grateful to David Wood and the School of Forestry, University of California, Berkeley, for accommodating the Workshop on Day 1 and to Garland Mason and the Pacific Southwest Research Station for accommodations on Day 2. We extend special thanks to Maureen Stanton for providing the "outsider's" point of view and invaluable insight during the course of the Workshop.

Publisher:

Pacific Southwest Research Station Albany, California (Mailing address: P.O. Box 245, Berkeley, California 94701-0245 Telephone: 510-559-6300)

November 1992 Proceedings of a Workshop on Bark Beetle Genetics: Current Status of Research May 17-18, 1992, Berkeley, California

Jane L. Hayes and Jacqueline L. Robertson, Technical Coordinators

CONTENTS

Preface...... iii

An (Ecologically Biased) View of the Current Status of Bark Beetle Genetics and Future Research Needs ...... 1 Jane L. Hayes and Jacqueline L. Robertson

Qualitative Genetics of Mountain Pine Beetle in Central Oregon ...... 3 Lula E. Greene

Biochemical Indicators of Population Status of Pine Bark Beetles ...... 4 M. Carol Alosi and Jacqueline L Robertson

Spatial Analysis in Bark Beetle Research ...... 5 Haiganoush K. Preisler

Bark Beetle Genetics ...... 6 Wyatt W. Anderson and C. Wayne Berisford

Isozyme Studies of Bark Beetle Population Genetics and Systematics ...... 7 Molly W. Stock, Gene D. Amman, and Barbara J. Bentz

Biosystematics of Ips mexicanus and Ips plastographus (Coleoptera: Scolytidae) and Their Fungal Symbionts ...... 10 David L Wood, Andrew J. Storer, Thomas R. Gordon, Steven J. Seybold, and Marion Page

The Status of Scolytid Genetics―A Reductionist's View ...... 12 Steven J. Seybold

Genetic Characters for Bark Beetle Phylogenies ...... 14 James H. Cane

Cuticular Hydrocarbon Research ...... 16 Marion Page CONTENTS

Bark Beetle Genetics―An Overview ...... 17 Stephen Teale and Barbara Hager

Current Status and Critical Research Needs in Scolytid Genetics ...... 18 Kenneth F. Raffa

Genetic Basis of Semiochemically Mediated Bark Beetle-Predator Coevolution: Implications for Developing Mass-Trapping Technologies ...... 20 Robert A. Haack, Daniel A. Herms, Bruce D. Ayres, Matthew P. Ayres, and M.L Doozie Snider

Population Genetics of Spruce Bark Beetle Ips typographus (Col., Scolytidae) and Related Ips Species...... 22 Christian Stauffer, Renate Leitinger, and Erwin Fuehrer

Novel Bark Beetle Research Possible with New Genetic Techniques ...... 23 Ken Hobson and Owain Edwards

Systematics Research ...... 25 Donald E. Bright

References ...... 26

Workshop Participants ...... 31

ii USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. Preface This report is the result of a workshop focusing on the To structure our general discussions, priority research needs topic of bark beetle genetics held May 17-18, 1992, in Berke- were identified by participants. The group recommended that ley, Calif. Bark beetles play a significant role in pine forest research be initiated or continued on these 15 topics: ecosystems throughout the world; this group of insects often • understanding ancestry of scolytid species causes severe economic damage and loss of wildlife habitat. In • biosystematics of important species, including symbionts general, bark beetles threaten forest health in the short term. In • monitoring endemic vs. epidemic conditions, and under- the long term, however, they have served as natural agents to standing processes that trigger changes thin overcrowded stands in pine forests. Although bark beetles • spatial statistical techniques for mapping geographic pat- have been the focus of intensive research efforts, relatively terns of gene frequency (and changes in relation to popu- little effort has been devoted to understanding bark beetle lation dynamics of bark beetles) genetics. Our Workshop evolved because of (and reflects) the • interaction between phylogeography and population growth growing interest in this relatively unexplored area of bark status beetle research. The objectives of this workshop were four- • genetics of variation in pheromone systems fold: • understanding individual variation in pheromone and other qualitative characteristics of individual beetles 1. To promote interaction among researchers and exchange • genetics of variation in interaction with natural enemies of ideas; genetics of cold-hardiness 2. To foster collaboration; • understanding selection pressure (i.e., stage and mecha- 3. To summarize the state of knowledge; and nism) 4. To identify research needs for the future. • artificial rearing techniques for major species (especially Dendroctonus spp.) To accomplish the last two goals and to set the stage for • molecular population analyses that focus on phylogeo- discussions, participants were asked before the workshop to graphy, gene flow, paternity submit brief descriptions of their views of the current status of • gene expression (description of genome, protein and bark beetle genetics research and the needs for the future. How enzyme level, transcription) for key traits this assignment was fulfilled was as diverse as the participants' • survey of polymorphisms identified from different ana- approaches to the topic. Contributions varied from broad over- lytical techniques views to detailed descriptions of specific projects. As a whole, • applying information from other well-researched sys- these statements provide an insightful description of the state of tems (e.g., Drosophila, Tribolium) to bark beetle knowledge and research in the area of bark beetle genetics. genetics Students of bark beetle biology will find this review and Because items on this list were intentionally expressed in the combined reference list a valuable source of current infor- relatively broad terms and covered multiple levels of analysis mation on this topic. Significant advances have been made in (i.e., molecular to population to phylogenic levels), we did not the available technology in the decade since the topic of bark attempt to refine or prioritize this general list any further. beetle genetics was first reviewed (Mitton and Sturgeon 1982). Instead, we sought to identify more specific questions and Our knowledge of bark beetle biology has advanced slowly; technological advances necessary to address these questions. however, as many contributors indicate, the potential for more This discussion is summarized in table 1. rapid advances is imminent. Through this report we seek to continue the productive During the two-day workshop, participants gave brief pre- efforts described at this workshop: to share information and sentations on the status of their ongoing research efforts. A ideas, and to encourage the pursuits of others with interests in general group discussion of research needs and goals followed. the challenges of research into genetics of bark beetles. All A topic of considerable concern to the group was the lack of participants welcome further inquiries―addresses and tele- recognition of the need and support for a continuous effort in phone numbers are provided. We hope that this publication will the area of systematics. Declining support for systematics work aid those seeking to learn more about the topic, and we encour- has resulted in lack of training and career opportunities for the age efforts in studying new topics that are not mentioned here. next generation of skilled systematicists. Without a commit- ment by academic and funding institutions to support this Jane L. Hayes fundamental research, we face a future without advances in this Jacqueline L. Robertson essential specialty area. Technical Coordinators

USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. iii Table 1―Specific questions and technological advances necessary to address priority research needs.

Levels of Analysis Specific Questions Techniques

1. Molecular genetics a. Study patterns of expression for genes involved in Determine more biochemical pathways. semiochemical pathways, e.g., detoxification, pheromones. b. Study genetics of development, e.g., stage-specific gene Develop DNA probes associated with key (adult vs. juvenile, overwintered vs. pre-winter, etc.) developmental events. c. Screen for specific patterns of expression associated with Develop systems to describe transformation of dispersal vs. feeding adults; cold hardiness; different seasonal trees or beetles. cohorts; host-tree species. d. Development of molecular markers for ecological genetics. RAPDS, VNTR loci, mtDNA e. Studies of sex ratio anomalies (e.g., in Ips spp. etc.) (see Level 2)

2. Quantitative genetics of Once appropriate rearing techniques are established for Need rearing techniques for studying similarity ecologically important controlling environmental and genetic background, focus on within "constant" environment and for traits. traits, establish role of genes and environment as influencing manipulation of environment. variations among individuals. a. Semiochemicals, correlations between them (production Artificial media vs. outdoor trees, logs. and response profiles) b. Cuticular hydrocarbons, e.g., changes during development Explore Kermit Ritland's technique for h2 and relationship with host compounds estimation based on electrophoretic data. c. Cold-hardiness: examine patterns of expression in larvae d. Microbial interactions: involvement with overcoming host defenses, increased attack success, pheromone synthesis e. Morphological/physiological traits (e.g., elytra loadings, lipid Fluctuating asymmetry analysis. contents) f. Genetics of host selection g. Trees: genetic and environmental determinants of susceptibility

3. Selection and population dynamics a. Stages and sources of selection in mortality during the life cycle due to host stress and predation and parasitism at various stages. b. Dynamics of beetle interactions (e.g., competition) as a function of population phase in the context of tree interactions. c. Do patterns of selection differ between population phase? (e.g., r- vs. k-type) Rearing techniques. d. Potential for selection experiments for key characterization (e.g., thick vs. thin phloem) (See level 4c and d). Techniques for following cohorts of beetles dispersal: density and "phase"-dependency below the bark (acoustics, radiography, infra- red detection)

population structure: “phase”-dependency

continued

iv USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. Table 1, continued

Levels of Analysis Specific Questions Techniques

4. Population structure and a. Calculate hierarchical F-statistics: between geographic regions, Also relevant to Biosystematics phylogeography between populations, within populations (between host plants, pheromone "groups", emergence groups, families) (See Level 5a) b. Exploit existing data sets: Dendroctonus spp., I. pini (requires systematic sampling and coordination between groups). Spatial statistics development. c. Spatial analysis, interaction with GIS data now available. d. Dependence of population structure on: 1. local density 2. stage of population growth (See Level 3d) Elemental labelling e. Direct analysis of dispersal, gene flow in relation to elemental (Mark-Release-Recapture). labelling rare or unique alleles, mutants (beetles or symbionts) (See Level 3d)

5. Phylogenetic reconstruction and a. Species taxonomy of bark beetles is relatively complete, but Develop discrete characters for: biosystematics. need phylogenetic reconstruction. 1. morphometrics 2. mtDNA, nuclear markers 3. sequence data on rib. b. Focus on Ips, Dendroctonus, especially 1) species relationships DNA (larger scale) within these groups, and 2) ancestral relationships between them. 1. develop discrete characters for cladistic analysis, 2. continue applying isozymes. Apply new methods of phylogeny c. Based on phylogeny, consider evolution of following traits: reconstruction and analysis. 1. pheromone production and composition (semiochemistry), 2. behavior, including isolation mechanisms, 3. aggregation response, and 4. cuticular hydrocarbons

USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. v An (Ecologically Biased) View of the Current Status of Bark Beetle Genetics and Future Research Needs1

Jane L. Hayes and Jacqueline L. Robertson2

Bark beetles are commonly considered the most destruc- designed experimentation. Ultimately, these results must be tive insect pests in pine forests in North America. Despite the tied to ecologically relevant characteristics (phenotypic expres- relatively intense interest in this group because of their pest sion). status and uniquely complex ecological and evolutionary rela- Emphasizing an ecological genetics approach, our (other tionships, many aspects of their biology, including genetics, are team members include Alosi, Greene, and Preisler, see contri- poorly understood. Most genetic research on bark beetles has butions this proceedings) investigations have involved esti- concerned inferences about the evolution of species, especially mates of both genotypic and phenotypic variation within and the phylogeny of important genera (e.g., Cane and others 1990b, among bark beetle populations and communities, as well as Lanier 1967, 1981, Lewis and Cane 1990b, Mitton and Stur- across ecologically similar species (mountain, western, and geon 1982). Relatively little emphasis has been placed on southern pine beetles). Our research includes aspects of the population genetics, particularly on genetic variation within technology such as starch gel electrophoresis that were used in and among populations over time and space (reviewed in Milton the earlier studies of differences between populations (e.g., and Sturgeon 1982, and ref. below). Roberds and others 1987, Stock and others 1979, 1984). Be- The natural history of this group is probably the single sides genetic inferences made by electrophoresis, we are exam- most significant deterrent to studies of population genetics, ining differences in the levels of the key enzymes that help to because sampling and laboratory rearing is either difficult or adapt beetles to the presence of toxic secondary chemicals in impossible. Improved sampling of adults has paralleled ad- their host trees (see Alosi this proceedings). In contrast with vances in semiochemical research, which in turn have prompted previous gross examinations of terpene levels in host trees and permitted recent investigations into variation in the produc- (Coyne and Lott 1976, Smith 1975)―their primary defense― tion and response to semiochemicals at the population level we are examining differences in terpene composition by means (e.g., Birgersson and others 1984, Herms and others 1991, of more sensitive analyses to detect and monitor changes con- Miller and others 1989, Teale and Lanier 1991). However, sistent with changes in population structure. most of the life cycle is spent under bark, limiting observations To date, information acquired from five years (1985-90, and necessitating destructive sampling of brood in order to see Greene this proceedings) of monitoring mountain pine obtain requisite estimates of fitness (e.g., fecundity, age to first beetle populations in Central Oregon suggests that patterns of reproduction). Most species are relatively intractable in the esterase enzymes detected by electrophoresis have shifted con- laboratory, thus precluding classical genetic studies, estima- comitantly with a shift in population status from endemic to tions of heritability, and development of molecular genetics epidemic and back to endemic. In addition, quantities of es- techniques based on phenotypic studies. It is not surprising that terases measured biochemically have changed, even for moun- recent advances in our understanding of both molecular and tain pine beetles in the same host species. Glutathion S-trans- phenotypic variation are concentrated in the more easily reared ferase levels also seem to be involved in the transition. We have species such as Ips spp. (e.g., Cane and others 1990b, Gast initiated (1991-present) comparable studies with western and 1987, Lanier and others 1972, 1980, Miller and others 1989, southern pine beetles and begun investigations of phenotypic Piston and Lanier 1974, Stauffer and others 1992a; but see variation between and within populations including analyses of Langor and Spence 1991, Teale 1990, Teale and Lanier 1991). behavioral (pheromone response), physiological (lipid content) The newest molecular techniques (e.g., Restriction Fragment and morphological (fluctuating asymmetry) variation. These Length Polymorphism and Randomly Amplified Polymorphic studies represent modifications to previous efforts to quantify DNA) may mitigate sampling deficits, are likely to lead to gene and relate phenotypic variations to genotypic variation among mapping, and vastly accelerate the molecular exploration of beetle populations (e.g., Langor and Spence 1991, Sturgeon genetic variation. Even with these advanced techniques, how- and Mitton 1986). For example, morphological variability ever, dramatic advances depend on novel application and well- within individuals may provide a valuable early guide to changes in beetle quality. Fluctuating asymmetry (nondirectional deviations from bilateral symmetry) has been shown to be an 1An abbreviated version of this paper was presented at the Bark Beetle effective and inexpensive tool for detecting stress in natural Genetics Workshop, May 17-18, 1992, Berkeley, California. populations (Leary and Allendorf 1989) and in cultures (Clarke 2Supervisory Research. Entomologist, Southern Forest Experiment Sta- tion, USDA Forest Service, 2500 Shreveport Highway, Pineville, LA 71360, and McKenzie 1992), because it provides a reasonably direct and Supervisory Research Entomologist, Pacific Southwest Research Station, measure of developmental stability (Palmer and Strobeck 1986). USDA Forest Service, Albany, Calif. Environmental and genetic perturbations could affect fluctuat-

USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. 1 ing asymmetry analogously, i.e., as the level of stress of any must be assessed to understand the sources of pest population kind increases, fluctuating asymmetry and its variability should variation. increase provided that the stress is sufficiently intense to pro- 3. Continuous rearing technology (e.g., artificial bark duce an effect (Parsons 1990a, b). beetle diet) must be developed for Dendroctonus spp. In addition, in most genetic studies done to date, standard statistical methods involving chi-square comparisons of ob- Needs in Quantitative Studies served gene frequencies versus those predicted by Hardy- 1. Estimate heritabilities of key traits over space and time Weinberg equilibrium have been used for data analyses. Our 2. Identify genetic markers and develop DNA/RNA probes, investigations emphasize the use of exploratory statistics to estimate frequencies, and study changes in population structure examine actual numbers rather than frequencies (see Preisler over space and time this proceedings). Uses of these new procedures will permit us 3. Having identified one or more markers that differ con- to model patterns in biotic communities of which bark beetles stantly with space, time (or both), quantitative investigations are a part, to estimate realistic models of species overlap, and to are needed to establish sources of variation (selection pres- formulate causal hypotheses about changes in bark beetle popu- sures), including lations in transition from endemic to epidemic states. a. Biochemical traits: levels of key enzymes such as Priority Research Needs esterases, mixed function oxidases, or any other enzyme that can be directly linked to survival 1. Long-term (multi-generation, multi-season) qualitative (fitness) and quantitative genetics studies are needed to detect genetic b. Phenotypic or quantitative traits: the expression of changes that result in changes on the population structure. life history traits (e.g., fecundity, longevity, age to 2. Likewise, changes in the component(s) that comprise first reproduction, phenology), or behavioral, physi- the gene-environment interaction (e.g., host-insect interactions) ological or morphological traits.

2 USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. Qualitative Genetics of Mountain Pine Beetle in Central Oregon1

Lula E. Greene2 The mountain pine beetle (MPB), Dendroctonus similarity coefficients suggested that they were closely related ponderosae, is a predator of all native and many introduced (Stock and Guenther 1979). Coastal and inland populations of pines in western North America (Furniss and Schenk 1969, MPB collected from varieties of lodgepole pine showed signifi- McCambridge 1975, Smith and others 1981, Wood 1982). The cant differentiation (Stock and others 1978). In Utah, geo- most important hosts of MPB are ponderosa, lodgepole, sugar, graphically separated populations of MPB from the same host and western white pine. Although these pine species occur were more similar than adjacent populations of MPB in diffe- sympatrically throughout portions of their ranges, MPB are ent hosts (Stock and Amman 1980). These studies showed that rarely found in more than one host species in any given locality. there is great genetic variation within and among the popula- Along the Front Range in the Colorado Rockies, this bark tions of MPB in the same host in addition to variation due to beetle is found primarily in ponderosa pine, but throughout the geography. Host tree species is implicated as a factor in northern Rockies, its preferred host is lodgepole pine. In maintaining differences among the MPB populations (Stock Oregon, the major hosts are both ponderosa and lodgepole and Amman 1985). However, the different host trees were not pines, but in California, the major host is sugar pine. Neverthe- sampled at the same site, so the significance of host variation as less, small areas can be found in which this beetle has colonized a factor cannot be separated from natural variation and geo- two or more hosts at the same time. graphic variation. No factor in the insect/host interaction influences the life Studies by Sturgeon and Mitton (1986) and Langor (1989) of an endoparasitic phytophagous insect more than its host indicate that significant genetic divergence does exist in co- (Diel and Bush 1984). Because plant species differ chemically, occurring populations of MPB in different hosts at the same physically, and biologically, they can present markedly differ- locality. These differences, they conclude, can be attributed to ent selective environments to insects feeding on them (Bush either selection, substructuring populations of MPB according 1969, Wood 1980). Differential selection pressures provided to host, or nonrandom mating in the hosts' population. Either by different host species can cause genetic differentiation of an of these events could lead to host-race formation, and ulti- insect species along host lines. Significant genotype variation mately, to speciation. in forest trees (the hosts) have been identified for bark beetles To gather more information to clarify insect/host interac- (Berryman 1972, Smith 1963, Smith 1972). Host species affect tions, we initiated a 10-year project on the Deschutes National emergence, egg gallery characteristics, fecundity, development, Forest in central Oregon in 1985. One of our objectives is to and mortality of MPB (Amman 1982, Amman and Cole 1983, determine the effects of secondary plant chemicals on MPB Amman and Pasek 1986, Schmid 1972). infesting five host trees over time and space. This should aid in Sturgeon and Robertson (1985) measured microsomal developing a basic understanding of host-induced genetic changes polysubstrate monooxygenase (PSMO) activity in western pine in bark beetles. Our long-term study will enable us to monitor beetle from ponderosa pine and in MPB from both lodgepole changes in allelic frequencies and genotype structuring of popu- and ponderosa pines. Although they detected no significant lations for trends that indicate changes in the population status difference in the PSMO activity between the two beetle spe- (endemic to epidemic). cies, PSMO activity in MPB from ponderosa pine was signifi- At present, we are analyzing data from the first five years cantly lower than in MPB from lodgepole pine. These results of our study. Esterase enzymes are involved in the insect's suggest that MPB in the two different host species might have ability to withstand toxins in its environment and its ability to qualities that vary with respect to their ecological genetics. survive. The esterase enzymes reveal four loci and show the Isozyme electrophoresis is another means that can be used most genetic variation. At the beta-esterase 1 locus, there is to track changes in beetle quality; in such studies, the genetic decided preference for heterozygotes, with one heterozygote structure of insect populations is used to measure genetic varia- genotype having a frequency that ranges 0.720 to 1.00 in 3 out tion. Data from electrophoretic surveys have provided direct of 39 samplings. This locus seems promising as a diagnostic measures of genetic variation within and genetic similarity indicator of host preference and perhaps, population status. among populations (Stock and others 1978). Stock and associ- Although our present focus is on MPB, information gained ates (1984) have surveyed much of the northern range of the can provide insights for research into other polyphagous insects MPB for electrophoretically detectable genetic variation. Beetles and other insects that persist at low levels for long periods of collected from populations in Oregon, Washington, Idaho, and time and suddenly explode to devastate entire forests. Montana could be differentiated at two enzyme loci, although No one biotechnology investigatory tool presently available can provide all of the information required to satisfy the 1An abbreviated version of this paper was presented at the Bark Beetle needs of entomologists, population geneticists, and systematicists, Genetics Workshop, May 17-18, 1992, Berkeley, California. 2Research Entomologist, Pacific Southwest Research Station, USDA but several used in concert can validate or bring a greater degree Forest Service, 800 Buchanan Street, Albany, CA 94710 of credibility to the information obtained through their use.

USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. 3 Biochemical Indicators of Population Status of Pine Bark Beetles1

M. Carol Alosi and Jacqueline L. Robertson2

Background

The processes by which insects cope with toxic chemicals In bark beetles, levels of GST may vary with host and with in their environment are commonly classified as class 1 and 2 population status (endemic versus epidemic) (Robertson and reactions (Dauterman and Hodgson 1978). Class 1 reactions others in progress). These observations have led us to propose include oxidations, reductions, and hydrolyses. By these pro- that a study of the structure and expression of GST genes in cesses, a hydrophilic group is added onto the foreign molecule beetles will provide us with molecular indicators of population and the molecule is then eliminated. Class 2 reactions consist of status. In addition, studies of GST genes in beetles will provide conjugations, i.e., glutathione conjugations, glucoside forma- insight into the mechanism of metabolic resistance in polypha- tion, and amino acid conjugation. gous insects. Sturgeon and Robertson (1985) examined the comparative activity of polysubstrate monooxygenases (PSMO = mixed function oxidases) of mountain pine beetle (MPB) in lodgepole Method of Study pine (the preferred host) and ponderosa pine (alternate host). The immediate goal of our investigation is to isolate cDNA PSMO activity of western pine beetle (WPB) in ponderosa pine clones of expressed GST genes from mountain, western, and was compared to that of MPB in lodgepole and ponderosa pine. southern bark beetles. In order to do this we will construct Results of this investigation, done with MPB in an endemic cDNA libraries of all three beetles and screen the libraries with population in the Deschutes National Forest in 1982, suggested heterologous probes derived from Drosophila and lepidopteran that host preference is reflected in levels of this detoxification cDNAs; alternatively, GST clones may be identified using enzyme. By 1985, the Deschutes population reached epidemic lepidopteran-derived GST antibodies. Positive GST isolates levels; MPB infested five pine species (lodgepole, ponderosa, from the libraries will provide us with homologous probes. western white, sugar, and whitebark). Homologous probes are necessary to attain the sensitivity nec- For the past seven years, we have collected emerging and essary to detect allelic variations of genes and to identify (for attacking MPB from a number of locations in the vicinity of La example) evidence of gene amplification in relationship to Pine, Oregon (Deschutes and Winema National Forests) for insect population dynamics. Presently, our work on this project biochemical and qualitative genetic studies. These studies have includes developing methodology for purification of RNA and been done to test the hypothesis that population status of this DNA from bark beetles. In addition, we are conducting hybrid- bark beetle species is associated with biochemical diversity ization studies between a Drosophila GST cDNA probe and (i.e., differences in class 1 and 2 reactions are associated with bark beetle RNA and DNA. These preliminary studies provide host tree species) that varies over time. We have quantified us with information useful for establishing experimental condi- levels of PSMO's and esterases in class 1, and glutathione S- tions for screening beetle cDNA libraries. transferase (GST) in class 2. We have detected changes in all three enzyme systems. During 1991, we expanded our attention to western and southern pine beetles, and to various Ips species. GST was chosen for study at the molecular level because methods have been developed for various Dipteran species and are being developed for a tortricid leafroller, Epiphyas postvittana, in New Zealand. GST are enzymes that enable cells to detoxify harmful compounds. Various forms of GSTs, representing allelic variation and sometimes complex gene families, are found in all eucaryotic organisms. There is some evidence that insects utilize the GST-enzyme systems to detoxify plant allelochemicals that they encounter during herbivory (Wadleigh and Yu 1987).

1An abbreviated version of this paper was presented at the Bark Beetle Genetics Workshop, May 11-18, 1992, Berkeley, California. 2Research Molecular Geneticist and Supervisory Research Entomologist, respectively, Pacific Southwest Research Station, USDA Forest Service, 800 Buchanan Street, Albany, CA 94710.

4 USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. Spatial Analysis in Bark Beetle Research1

Haiganoush K. Preisler2

Genetic and ecological information about bark beetle popu- models is quite flexible and very helpful as an aid for under- lations most often consists of data collected over space and standing spatial patterns of discrete data points and for relating time. However, when these data are analyzed, little attention is those patterns to specific environmental factors or tree charac- usually given to relationships operating in the spatial dimen- teristics. sion. One of the reasons for the small number of studies that The second technique that might be useful for the analysis have examined the spatial aspects of bark beetle populations is of spatial data is the methodology of Geographic Information the lack of statistical tools for the analysis of spatial-temporal Systems (GIS). GIS can be used to store spatial and temporal data. information on insect populations. Genetic information can One statistical technique that can be adapted for the analy- then be added to the system and used to analyze changes over sis of spatial data is the methodology of generalized linear space and time; interactions among and within various factors regression models. This method was used in a series of articles can be investigated. Although GIS allows entomologists to by Mitchell and Preisler (1991, Preisler [in press], Preisler and compile spatial data, deriving inferences from these data is still Mitchell [unpublished]). By fitting an auto-logistic model, we extremely difficult because appropriate statistical techniques were able to study the factors affecting the probabilities of trees are lacking. A few references that discuss statistical techniques being attacked by mountain pine beetle (MPB). Those factors for analyzing geographic information and gene-frequency maps included characteristics of individual trees (such as tree size are Brillinger (1990), Kemp and others (1989), Liebhold and and vigor), in addition to the locations of each tree within a Elkinton (1989), Piazza and others (1981). Further statistical stand. We have found that the framework of generalized linear research is needed in this area.

1An abbreviated version of this paper was presented at the Bark Beetle Genetics Workshop, May 12-18, 1992, Berkeley, California 2Research Statistician, Pacific Southwest Research Station, USDA Forest Service, 800 Buchanan Street, Albany, CA 94710.

USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. 5 Bark Beetle Genetics1

Wyatt W. Anderson and C. Wayne Berisford2

1. Previous work 3. Work to be done A. Protein electrophoresis the basic technique for most A. Long-term study to follow genetic variability and popu- work. lation structure in relation to population cycles of SPB. Electro- B. Genetic variability has been characterized. phoresis and perhaps restriction mapping techniques of choice. C. Population differences demonstrated and analyzed. B. Studies using mtDNA as molecular markers of popula- D. Genetic population structure assessed. tion history in relation to geographic location―phylogeography. E. Basic molecular systematics explored; phylogenetic Restriction mapping and sequencing of PCR-amplified sequences trees developed. techniques here. F. Attempts to link protein loci to phenotypic characters C. Molecular systematics of bark beetle taxa such as such as behavior. Dendroctonus species, using mtDNA and nuclear loci. Restric- 2. Our work at University of Georgia tion mapping and sequencing of PCR-amplified sequences A. Starch gel electrophoresis of Dendroctonus and Ips techniques here. populations, in terms of genetic variability, population structure, and population differences. B. mtDNA studies, using restriction site analysis and DNA sequencing. (1) Cloned part of mitochondrial DNA (2) Completed restriction site analysis of SPB populations (3) Begun sequencing of several mtDNA segments

1In lieu of a full narrative paper, the authors submitted this outline for publication. Any questions should be directed to the authors. 2Professor of Genetics, Dept. of Genetics, 101B Life Sciences Bldg., University of Georgia, Athens, GA 30602 and Professor of Forest Protection, University of Georgia, Athens, GA 30602

6 USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. Isozyme Studies of Bark Beetle Population Genetics and Systematics1

Molly W. Stock, Gene D. Amman, and Barbara J. Bentz2

Heterozygosity and Environmental Stress Electrophoretic analysis was used extensively in the 1970's to estimate genetic relationships among populations, species, Males were more differentiated than females among sites and closely related genera of many different plants and ani- and numbers of emerging males more variable in thin-phloem mals, including many insect groups. Some of the first electro- trees (Stock and Amman 1980). In a later study (Stock and phoretic work on bark beetles was done by Anderson and Amman 1985), males emerging from thin-phloem lodgepole others (1979), Anderson and others (1983), Florence and Kulhavy pine were found to be more heterozygous and more variable (1981), Florence and others (1982), and Namkoong and others from tree to tree, and fewer and more variable males (relative to (1979). These studies demonstrated that bark beetles are highly females) emerged from thin phloem. We suggested that genetic polymorphic and amenable to population studies using this diversity of MPB (as measured by average heterozygosity) technique. The first isozyme study of the mountain pine beetle might increase in response to increased severity of environ- (MPB) Dendroctonus ponderosae (Stock and Guenther 1979) mental conditions (i.e., stress), a response observed in numer- showed differentiation among geographically separated popu- ous other organisms by other workers (e.g., Samollow and lations. A subsequent study of MPB from four locations (Stock Soule 1983). Male MPB are generally thought to be more and Amman 1980) suggested that there might also be genetic sensitive to microenvironmental conditions than females, and differences at individual gene loci between MPB from lodge- therefore would be expected to have greater genetic diversity if pole and ponderosa pine, and greater overall heterozygosity in a generalized population-level response to increased selective beetles from ponderosa pine. Some preliminary data also showed pressures was an increased level of heterozygosity. Further differences between beetles emerging from trees with thin and evidence to support this hypothesis was provided by a study of thick phloem and between early- and late-emerging beetles isozyme variation observed over six consecutive generations of from the same tree. MPB laboratory-reared in thick and thin phloem; heterozygos- Inferences that could be made from these observations of ity increased in both groups over the generations, but more differences related to host species were, however, confounded rapidly in the beetles in thin phloem (Amman and Stock unpub- by geographic distance among sites. Sturgeon (1980) and Stur- lished). geon and Mitton (1986) studied sympatric groups of MPB In Ips pini (Gast and Stock unpublished), genetic diversity attacking different hosts in mixed pine stands and found signifi- was significantly higher in overwintered than in non-overwin- cant differences related to host tree species. Further evidence of tered beetles. However, Langor and Spence (1991) observed differentiation of MPB by host was found in a comparison of changes in gene frequencies but no consistent increase in het- isozyme frequencies of MPB collected from lodgepole and erozygosity in MPB with overwintering. These authors at- ponderosa pine at a single site in Utah (Stock and Amman tribute the genetic differentiation they observed to differential 1985). Deviations in genotype frequencies from Hardy-Weinberg survival in the hosts, rather than to differential host preference (random mating) expectations occurred when isozyme data of beetle genotypes. Thus, one might see, as did Gast and Stock from all beetles were pooled (Wahlund effect); frequencies (unpublished), greater differences between beetles that have were much closer to random mating expectations when sepa- lived for a longer time in different hosts or in the same host rate analyses were conducted for beetles from each of the host species under more severe environmental conditions. Other species. Once again, greater heterozygosity was observed in evidence supports this hypothesis. For example, Gast (1987) beetles from ponderosa pine. Overall, beetles from thin-phloem reported that overwintered I. pini responding to pheromone lodgepole were more heterozygous than those from thick ph- were significantly more heterozygous than overwintered beetles loem, though these differences were not detected in beetles responding to host volatiles. Heterozygosity levels in host- from ponderosa pine. responding, pheromone-responding, and nonresponding beetles that had not overwintered were very similar across response categories. Time of collection can therefore have an important influence on the type of genetic variation observed and interpretations that can be made from it. Langor and Spence (1991) and 1An abbreviated version of this paper was presented at the Bark Beetle the work of Gast (1987) suggest that laboratory rearing of Genetics Workshop, May 17-18, 1992, Berkeley, California. beetles collected from logs cut before winter reduces the differ- 2Professor of Forest Resources, Dept. of Forest Resources, College of Forestry, University of Idaho, Moscow, ID 83843; and Supervisory Research ential selective influence of host tree species. This possibility Entomologist and Research Entomologist, respectively, Intermountain Re- could help explain the fact that Sturgeon (1980) and Sturgeon search Station, USDA Forest Service, Ogden, Utah. and Mitton (1986) found genetic differences between MPB

USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. 7 from lodgepole and limber pine, whereas Langor and Spence unpublished). Thus, there may be a correlation between factors (1991) did not. Sturgeon's (1980) beetles were collected in of increased environmental stress that act to end an outbreak summer, just before emergence from their host trees; they and the higher levels of heterozygosity observed at this time. represented survivors of within-tree mortality estimated to av- One might infer a relationship between heterozygosity and erage 85 to 99 percent of the brood which had been established population size, but we need to keep in mind that the processes the previous fall. Langor and Spence (1991) used beetles reared underlying observed genetic variation are very complex and in the laboratory and collected in the fall and suggested that that factors that influence genetic variation under one set of data collected from laboratory-reared beetles better reflect pos- conditions (e.g., during outbreaks or epidemics) can be very sible effects of host selection behavior and the genetic makeup different under another set of conditions (e.g., during the low- of colonizing beetles. We compared heterozygosity of early- density or endemic population phase). For example, inbreeding and late-emerging MPB reared from fall-collected lodgepole in low-density populations might act to reduce heterozygosity logs from six sites (Stock and Amman unpublished). In beetles and counterbalance any tendencies toward increased heterozy- from four of the six sites, heterozygosity went up over the gosity related to environmental stressors that might be keeping emergence period. It is possible that such trends would be even the population at a low level. In the field, heterozygosity ap- more apparent in samples of beetles collected as they emerged pears to be lower in endemic than in epidemic populations in the field the following year. (Stock and others unpublished), and laboratory studies (Amman and Stock unpublished) showed that small numbers of parent Heterozygosity and Species Distribution beetles per log resulted in an F1 population with a much lower level of heterozygosity than the parental population. Langor and Spence (1991) attributed the low levels of On the basis of these ideas, Stock and others (unpublished) heterozygosity (10.0-11.2 percent) they observed in MPB from hypothesized that heterozygosity increasing over time might Alberta and British Columbia (compared to heterozygosity in indicate that a population was approaching the end of an epi- MPB from U.S. studies) to the location of their populations demic; conversely, a stable or decreasing heterozygosity in a near range margins. In contrast, our studies suggest that popula- high-density population could indicate that the population was tions near the margins of species distribution (presumably liv- unstressed and that population numbers would increase or stay ing under suboptimal conditions) could be expected to be more high. Predictions of population trend based on these hypotheses heterozygous. The highest level of polymorphism that we ob- for six MPB populations were good in four of six cases. served in a MPB population (more than 17 percent) occurred in a population from California, another area that might be con- Systematics of Dendroctonus Species sidered near the range margin for this species (Higby and Stock 1982). Heterozygosity levels of Langor and Spence's (1991) Using electrophoretic techniques, Higby and Stock (1982) Canadian populations were lower than in this California popu- identified diagnostic isozymes for sympatric populations of the lation, but approximately equal to heterozygosities in almost all sibling species D. ponderosae and D. jeffreyi in California. An other populations we studied―including a population from the earlier study (Stock and others 1979) showed very similar Black Hills (South Dakota), a third area that could be consid- levels of differentiation between D. pseudotsugae populations ered marginal for the species and which had an average het- from Idaho and Oregon, but the wide geographic separation erozygosity of 11.1 percent (Stock and others 1984). between these two populations precluded any definitive statements about species status. Genetic Indicator of Population Phase Wood (1963) used anatomical and biological characters to rank Dendroctonus species in order of increasing evolutionary Some very early electrophoretic work with the Douglas-fir specialization relative to other Scolytidae. Lanier (1981) also tussock moth (Stock and Robertson 1977) revealed an esterase suggested an order of specialization based on cytogenetic char- allozyme that appeared only in low-density populations. Popu- acteristics. Although Wood and Lanier placed the species into lations sampled two or more times during 1976 and 1977 nearly identical species groups, their postulated order of spe- showed changing frequencies of this allozyme (relative to the cialization of these groups were nearly opposite. Bentz and other two allozymes at that locus) that seemed to parallel Stock (1986) used electrophoretic data from 22 populations, changes in population density. At that time, we hypothesized representing 10 Dendroctonus species, in an attempt to unravel that relative frequencies of these three allozymes might be used the discrepancies in phylogenetic interpretations. Although the as a genetic indicator of future population trends in the Dou- species groups identified using electrophoretic data were simi- glas-fir tussock moth. Hayes and Robertson (these proceed- lar to those previously identified by both Wood (1963) and ings) describe allozyme variation at an esterase locus showing Lanier (1981), the implied phylogeny was independent of the similar correlation with population status in the MPB. species groupings. Some members of a species group were Average heterozygosity also appears to be related to popu- more specialized than other members in the same group. On the lation phase, reaching highest levels as epidemic populations basis of these data, it appears that no single species group can reach peak numbers and begin to decline (Stock and others be said to be more advanced or more primitive than another.

8 USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. The phylogenetic tree produced using electrophoretic data Research Needs suggests that D. rufipennis, D. adjunctus, and D. approximatus were the most primitive species, followed by D. ponderosae (1) Test the heterozygosity model of Stock, Schmitz, and D. brevicomis; D. valens, D. terebrans, D. simplex, D. and Amman for predicting phase (endemic, building, epidemic, pseudotsugae, and D. frontalis are among the more evolution- postepidemic) of MPB populations. arily advanced species in the genus. There were no discernible (2) Investigate further the change from endemic to trends in level of heterozygosity (by species) based on evolu- building phase: What are the influencing factors associated tionary position in the hypothesized phylogenetic tree. These with it (population size, mixing of demes, synchrony of emer- interpretations of the phylogenetic relationships among gence and attack, host characteristics, etc.)? Dendroctonus species could be useful in analysis of behavioral (3) Continue to investigate differences in MPB re- strategies used in scolytid genera, such as production and response to verbenone (and possibly other pheromones): Are sponse to semiochemicals, mating strategies, and host tree they genetically based and, if so, what factors are causing associations. selection? Stock and others (1987) compared the great European (4) Dispersal: Are there physiological and/or genetic spruce bark beetle, D. micans, to the 10 North American differences among individuals with different flight behavior Dendroctonus species studied by Bentz and Stock (1986). Av- (long vs. short fliers) and do these relate to population phase? erage heterozygosity was 5.3 percent in D. micans; North (5) The effect of temperature on MPB is very impor- American species ranged from 11.4 to 22.6 percent. Cluster tant in Rocky Mountain populations. Is cold tolerance geneti- analysis suggests that D. micans is most closely related to D. cally based, possibly showing up as geographic differences terebrans and D. valens, species with which it shares the char- associated with elevation and latitude? acteristic of gregarious larval feeding in the phloem of living (6) Continue study of bark beetle systematics using hosts. More recently, Gast and Furniss (unpublished) compared electrophoresis and other, newer, molecular genetic techniques. isozyme characteristics of D. micans with its morphologically similar, spruce-infesting, North American counterpart, D. punctatus.

USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. 9 Biosystematics of Ips mexicanus and Ips plastographus (Coleoptera: Scolytidae) and Their Fungal Symbionts1

David L. Wood, Andrew J. Storer, Thomas R. Gordon, Steven J. Seybold, and Marion Page2

Ips mexicanus (Hopk.) (concinnus group of S.L. Wood Very little is known about the fungal symbionts of these 1982) and I. plastographus (plastographus group) share com- species. Recently Parmeter and others (unpublished data) have mon coniferous hosts throughout western North America (S.L. isolated unidentified species in the genera Ophistoma, Wood 1982). In California their distribution in the coastal pines Leptographium, and Graphium from I.p.. maritimus and coastal (Pinus radiata, P. muricata, P, contorta contorta, and P. populations of I. mexicanus. These fungi were inoculated into contorta bolanderi) and in the high elevation P. contorta living trees. Through dye studies (Parmeter and others 1989) murrayana of the Sierra Nevada Mountains (and Cascade Moun- some isolates were shown to interrupt water transport. The role tains) is especially interesting because of their wide separation of symbiotic fungi in the biology of bark beetles has been by the Sacramento and San Joaquin Valleys. This separation is studied extensively, especially in the Ophistoma spp./ 2-400 km from Redding in the north to Bakersfield in the south, Dendroctonus spp. system (reviewed exhaustively in Schowalter a latitudinal distance of about 1200 km. Thus these populations and Filip 1992; see for example, Owen and others 1987, Parmeter have been noninterbreeding populations for thousands of years. and others 1989, 1992). In Ips spp./fungi associations, Furthermore, these coastal and montane populations probably Ophistoma spp. have been shown to inhibit oviposition but not come together where P. contorta and P. contorta murrayana or development (Yearian and others 1972) and to interrupt water P. c. contorta hybridize. Native Monterey pine (Pinus radiata) transport (Parmeter unpublished data, Homvelt and oth-ers is now represented by only three small separated coastal popu- 1983). Although no specialized morphological structures are lations. Other hosts of I. mexicanus and I. plastographus occur apparent in Ips spp. for carrying these fungi [such as the on small islands off the southern California coast and into the pronotal mycangium of Dendroctonus brevicomis and D. frontalis mountains of Mexico (Bright and Stark 1973, S.L. Wood 1982, and maxillary mycangia of D. jeffreyi and D. ponderosae Seybold and others 1992a). (Whitney 1982)], there is a consistent and ubiquitous associa- On the basis of morphological criteria and reduced hatch- tion between bluestain fungal invasion of the sapwood and ing in the F1, Lanier (1970b) defined subspecies of I. phloem and the gallery systems of these bark beetles. There- plastographus: I. plastographus maritimus Lanier for the fore, studies of relatedness and divergence in I. mexicanus and coastal form and I. plastographus plastographus (Leconte) for I. plastographus would be enhanced by similar analyses of the montane form. In limited cross-mating studies, he showed these phoretic symbionts. Evidence of divergence with either that both populations were interfertiles at the F1 and at the or both taxa would be of great value in understanding the parental backcrosses. Similar limited studies were conducted evolution of these species. with the cohabiting I. mexicanus (Lanier 1967). However, in Studies of relatedness in Ips spp. have focused on I. pini recent studies of the distribution of ipsenol (2-methyl-6- (eastern/western populations) and on the grandicollis group methylene-7-octen-4-ol) and ipsdienol (2-methyl-6-methylene- sibling species (paraconfusus, confusus, and hoppingi). In I. 2.7-octadien-4-ol), pheromone components in Ips spp., pini, no post-mating barrier has evolved between eastern popu- Seybold and others (1992b) found both components in lations (Ontario, Canada) crossed with western populations montane I. mexicanus but only ipsdienol in coastal I. (California and Arizona) (Lanier 1972). However, a premating mexicanus. The enantiomeric com-position of ipsdienol is ~90 barrier has evolved between these two populations. There phero- percent-(-) for both populations. Although the status of these mone component, ipsdienol, is produced by both populations. compounds as pheromone compo-nents for these species has However, in the California population (as well as in most not been established, they have been shown to be pheromone western North American populations―see Seybold and others components in all species studied to date (D.L. Wood 1982). 1992d), ipsdienol is produced as a mixture of 98 percent (-)- Nevertheless, the presence of ipsenol in only the montane and 2 percent (+-)-ipsdienol. The (+)- isomer is an interruptant, population offers considerable opportunity to investigate the whereas the (-)- isomer is a powerful aggregation attractant genetics of these two populations. (Birch and others 1980; Seybold and others 1992d). In the eastern population a mixture of about 40 percent (-) and 60 percent (+)-ipsdienol is produced. Here both isomers function 1An abbreviated version of this paper was presented at the Bark Beetle as an attractant. We have found only a few beetles responding Genetics Workshop, May 17-18, 1992, Berkeley, California. 2Professor of Entomology and Post-Doctoral Researcher, respectively, to the racemate in California (Seybold and others 1992d). Dept. of Entomological Sciences, University of California, 201 Wellman Hall, Eastern populations respond at low levels to 98 percent (-)- Berkeley, CA 94720; Assistant Professor, University of California, Berkeley, ipsdienol (Teale 1990). In addition to ipsdienol, lanierone (2- CA; Post Doctoral Scholar and Research Entomologist, Pacific Southwest hydroxy-4,4,6-trimethyl-2.5-cyclohexadien-l-ol) has recently Research Station, USDA Forest Service, 800 Buchanan Street, Albany, CA 94710.

10 USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. been isolated and shown to be an important component of the canker infections associated with attacking adults. This fungus, pheromone of the eastern population (Teale and others 1991). which is native to pines of the southern U.S. (Correll and others Although this compound caused an increased response to 1992), causes extensive tip dieback followed by numerous ipsdienol in California, it has not been found in California resinous bole-cankers. We believe that this fungus further weak- populations (Seybold and others 1992c). As with ipsenol in the ens trees so that they are more easily killed by these Ips species. montane population of I. mexicanus, the presence or absence of Our studies suggest that progress of this disease from lanierone and the differences in enantiomeric composition of urban plantings of Monterey pine in the Santa Cruz area into ipsdienol offer opportunities to investigate the genetics of these our native forests will likely be a consequence of propagule two populations of I. pini. transmission via I. plastographus maritimus and I. mexicanus. In the work with the grandicollis group, Lanier (1970a and We predict transmission from I. mexicanus to I. plastographus b) and Merrill (1991) established the presence of postmating in the northernmost native stands of Monterey pine (Ano Nuevo barriers among I. paraconfusus, I. hoppingi, and I. confusus. State Park) to urban plantings of Monterey and bishop pine (P. Premating barriers are weakly developed or not developed at all muricata), to native stands of bishop and shore pine (P. contorta (Cane and others 1990c, Fox and others 1991 a). Although there contorta) on the coast and ultimately inland to the Cascade and is pheromonal cross-attraction among these species (Lanier and Sierra Nevada Mountains, perhaps via P. ponderosa. Ips Wood 1975; Cane and others 1990a, 1990c), we have found mexicanus and I. plastographus cohabit these coastal pine nonreciprocal divergence in these pheromone-elicited behav- species as well as P. contorta murrayana (I.p. plastographus) iors, i.e., I. paraconfusus females do not discriminate between in the above mountain ranges. Thus our studies on the coastal conspecific and I. confusus males, whereas I. confusus females and montane populations of these Ips spp. will be important in prefer conspecific males in the host pine species (Cane and predicting the ultimate distribution of this newly introduced others 1990c). Furthermore, females join heterospecific males fungal pathogen. At the same time we are in a unique position in host and nonhost pines (Fox and others 1991 a). Morphomet- to investigate the coevolved system of these bark beetle species ric, electrophoretic (Cane and others 1990b), and cuticular with their symbiotic fungi. Such studies will provide baseline hydrocarbon (Page and others 1992) analyses support the spe- information which can be used to understand the new symbi- cies status of these sibling 5-spined Ips described by Lanier otic relationship that we expect will develop when F. subglutinans (1970b). becomes a part of this community. Thus we see the evolution of postmating and weak-to- nonexistent premating barriers in the sibling grandicollis group Research Objectives and the reciprocal scenario in I. pini. The studies we propose will add to the knowledge base developed over the past 20 1. Determine the extent of postmating barriers to gene flow years on the biosystematics of these two Ips spp. complexes. At between coastal and montane populations of I. mexicanus. the same time, expanding our investigation to include the sym- 2. Determine the extent of premating barriers to gene flow biotic fungi should give us greater insights into the evolution of between the above populations. this group of organisms. 3. Determine the presence or absence of ipsenol in the Also of importance to the proposed study is the recent above crosses of I. mexicanus and in the backcrosses. discovery of the pitch canker fungus, Fusarium subglutinans, 4. Compare the genetic constitution of the above popula- in Monterey pine (P. radiata) plantings in the vicinity of Santa tions using mitochondrial DNA and cuticularhydrocarbon analy- Cruz, Calif. Our studies have shown that I. paraconfusus and I. ses. mexicanus adults carry propagules of this fungus (Fox and 5. Compare the genetic constitution of the fungal sym- others 1990, 1991b). Furthermore we have demonstrated that bionts commonly associated with the above populations using attraction of I. mexicanus to Monterey pines results in pitch mitochondrial DNA analyses.

USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. 11 The Status of Scolytid Genetics―A Reductionist's View1

Steven J. Seybold2

As biologists considering studies of scolytid genetics we because of the stature of the insects in forest management, are fortunate that an insect model exists that has been the target minor scientific gains may have large economic impacts. of genetic analyses on both the individual and population levels However, rather than repeat the past 80 years of Droso- for most of the twentieth century. The knowledge base associ- phila genetics with the Scolytidae, we should apply the Droso- ated with Drosophila melanogaster3 and congenerics is stag- phila knowledge base to scolytids as often as possible. No gering, and it is a tribute to scientific inquiry that so much is scolytid is likely to become a model for eukaryotic biology, but known about organisms of such little economic importance. by applying the tools used with Drosophila, we may discover The goal of the classical genetic crosses of D. melanogaster some interesting and perhaps unique biological phenomena in begun in the lab of T.H. Morgan in 1909 was to infer the the Scolytidae. genetic basis of the phenotype (phenotype →a gene). The devel- For the purposes of organization, it is useful to divide opment of P-element-mediated transformation (Rubin and scolytid genetics into the reductional and holistic approaches. Spradling 1982) allows "reverse genetic" studies where the By reductional I mean the analysis of biochemical, morpho- gene of choice is placed into the organism and the resulting logical, or behavioral traits of individuals and the interpretation individual is assayed to infer the phenotypic product of the of the results from a molecular or mechanistic perspective. By gene (gene → phenotype). During the period between the holistic I mean the analysis of the same traits from pooled advent of each of these approaches with Drosophila, serendip- samples of individuals and the interpretation of the results from ity and intensive screening procedures revealed a wide array of an ecological or population perspective. The following are morphological, developmental, and behavioral mutants; bio- several reductional areas of study that seem relevant to scolytid chemical studies characterized nucleic acids and proteins; ge- genetics. netic studies led to the development of physical, cytological 1) Genetic analysis of morpho- and chemosystematic maps of polytene chromosomes; and, soon, the entire genome characters: A variety of useful morphological characters have will be sequenced (Merriam and others 1991). The phenom- been identified for distinguishing scolytid taxa (Bright 1976, enon of transposition ("jumping genes") has been actively Bright 1992, Bright and Stock 1982, S.L. Wood 1982). How- investigated in D. melanogaster (Green 1980, Engels 1983), ever, mutations in these characters have not been studied in a and the transposons themselves have been the targets of studies systematic fashion [e.g., red vs. white eye color, vestigial vs. of population genetics (Charlesworth and Langley 1989). In wild-type (normal) wing morphology in Drosophila]. Recently addition to the biochemical and molecular work, the drosophilids there has been considerable interest in establishing chemical have been the subject of studies of population biology, phylog- phenotypes for some species in the Scolytidae, and using the eny, and pheromone production (Bartlet and others 1986, newly discovered chemosystematic characters in constructing Kaneshiro 1988, Singh 1989). phylogenetic relationships (cuticular hydrocarbons―Page and In the Scolytidae, no mutants have been isolated, prospects others 1990a, b, 1992; Bright 1992; chirality of pheromone for long-term culture have not been developed, most crosses components―Seybold and others 1992b; and migration of have been performed to determine species status of wild popu- metabolic enzymes in starch gels―Cane and others 1990b, and lations rather than to assay for intraspecific phenotypic vari- other studies). ants, and few biochemical reactions have been characterized. However, while typical chemical phenotypes have been When evaluated against the background of Drosophila genet- characterized, nothing is known about the genetic regulation ics, the status of scolytid genetics can be described as primitive leading to these phenotypes and possible mutations that might at best. Thus, it is in an area of enormous opportunity, where, lead to altered chemical phenotypes. Furthermore, because of the insensitivity of the chemical assays, the chemical phenotypes have often been determined on pooled samples of individuals, whereas we ought to evaluate the phenotypes of indi- 1An abbreviated version of this paper was presented at the Bark Beetle viduals. It will be important to standardize laboratory and Genetics Workshop, May 17-18, 1992, Berkeley, California. 2Post Doctoral Scholar, Pacific Southwest Research Station, USDA statistical procedures used to evaluate a range of individual Forest Service, 800 Buchanan Street, Albany, CA 94710. normal chemotypes so that mutants can be recognized when 3An organization called the Drosophila Information Service catalogs and they are encountered. This is not an easy problem. For example, communicates biological (mostly, genetic) data about D. melanogaster and the cuticular hydrocarbon profile from a species typically con- related species. In addition, there is an encyclopedia of Drosophila mutations authored by Lindsley and Grell, and various stock cultures of wild type and tains 60 to 80 hydrocarbon components which vary in presence, mutant strains are maintained around the world.

12 USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. absence, and quantity. Here are two interesting problems that strain of I. mexicanus that is deficient in the ability to produce seem appropriate to this topic. ipsenol. (a) Chirality of ipsdienol as a heritable trait of indi- 2) Characterization of biochemical pathways: To ac- vidual male Ips pini. Ips pini occurs in two basic pheromone company classical genetic analyses, we need to expand our types (Birch and others 1980, Lanier and others 1972, Lanier knowledge of bark beetle biochemistry. Because of a series of and others 1980, Miller 1990, Teale 1990, Miller and others pheromone biosynthesis studies with I. pini and I. paraconfusus 1989). The eastern form produces and responds maximally to using labelled precursors (Fish and others 1979, 1984, Hendry ipsdienol with an enantiomeric composition of 30 percent to 60 and others 1980, Vanderwel and Oelschlager 1987, Vanderwel percent-(4R)-(-), whereas the western form produces and re- 1991), these two species make particularly good candidates for sponds maximally to ipsdienol with an enantiomeric composi- initial studies on the enzymes involved in pheromone biosyn- tion of 93 percent to 100 percent-(4R)-(-). The eastern form thesis. produces and responds strongly to lanierone (Teale and others 3) Sex ratio condition: One intriguing genetic problem 1991); the response of the western form is enhanced by lanierone, that bears further study is the sex ratio condition discovered in but males from this population do not appear to produce lanierone Ips latidens and other species (Lanier and Oliver 1966). In this (Seybold and others 1992c). Populations from North America phenomenon individuals from certain populations carried a have been surveyed (Miller 1990, Seybold and others 1992b), maternally transmitted factor that resulted in selective mortality and a zone of sympatry between the forms appears to occur in of male embryos. A similar sex ratio condition also occurs in British Columbia. Teale (1990) has crossed eastern (New York) Drosophila spp. (Sakaguchi and others 1965, Ikeda 1965, and western (Arizona) forms and demonstrated heritable pat- Novitski and others 1965), and in recent studies the causative terns to the chirality of male-produced ipsdienol. He has also microorganisms have been isolated and cultured (Hackett and found a correlation between the response to enantiomeric blends others 1986). Other sex ratio distortions in Drosophila spp. are by males and production in the eastern form of I. pini. We are caused by chromosomal and extrachromosomal factors. Mo- pursuing similar studies with the western population. lecular studies of a distortion in the latter group (hybrid (b) Ipsenol as chemotypic marker for two populations dysgenesis) led to the discovery of the P-element transposon of Ips mexicanus. Ips mexicanus occurs as three distinct popu- (Rubin and Spradling 1982). Application of modern biochemi- lations in western North America: coastal in Pinus radiata, P. cal and molecular biological techniques to the sex ratio condi- muricata, and P. contorta; montane in P. contorta murrayana tion in Ips may lead to similar discoveries. Lanier never pub- and P. contorta latifolia; and interior in other Pinus spp. The I. lished the fact that the condition also occurs in another species plastographus and I.integer complex can be broken down into of Ips [confusus (LeConte)] and in the scolytid genera three similar entities. In a survey of the enantiomeric composi- Pityophthorus and Dendroctonus. tion of ipsenol and ipsdienol produced by male Ips, we have 4) Construct gene libraries: With the cooperation of found that the coastal and montane populations of I. mexicanus Carol Alosi (Pacific Southwest Research Station, USDA Forest differ in their ipsenol-ipsdienol phenotype. Coastal I. mexicanus Service), we have begun to construct a c-DNA library of genes produce ~90 percent-(4R)-(-)-ipsdienol, whereas montane I. for I. pini. Large numbers of male and female western I. pini mexicanus produce the same ipsdienol and >99 percent-(4S)-(-)- have been frozen for this purpose. Genomic libraries for the ipsenol. We have not studied an interior population of I. major scolytid species should be constructed and made cen- mexicanus. Thus, interbreeding studies of coastal and montane trally available to all researchers (the beginnings of a Scolytid populations of I. mexicanus could be followed by traditional Information Service?). traits such as larval: egg niche ratios (Lanier 1967), as well as by ipsenol production. Coastal I. mexicanus could be treated as a

USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. 13 Genetic Characters for Bark Beetle Phylogenies1

James H. Cane2

The current taxonomic classification of scolytid bark beetles of evolutionary convergence, as in the stridulatory capabilities is based largely upon external morphological characters. This of taxonomically disparate bark beetle taxa. Even if characters will remain the status quo into the foreseeable future for rea shared by two taxa are indeed homologous, such as some of the sons of cost, time, convenience, and reproducibility. For genera saturated straight chain cuticular hydrocarbons of Dendroctonus of particular economic interest, notably Dendroctonus and Ips, spp. (Page and others 1990b), they are shared with many other additional taxonomic characters have been sought more or less insect genera and so are uninformative in describing phyloge successfully through karyological and electrophoretic studies netic relationships within any one lesser taxonomic unit. This and applied to teasing apart sibling species complexes and dilemma is best resolved using cladistics, an approach which circumscribing species groups (e.g., Lanier 1967, 1970a, 1970b, has emerged from the fires of heated debate over the past 20 1972, 1981; Cane and others 1990b). As noted by Bright (this years as the most appropriate method for reconstructing phylo workshop), this task of describing species of the North Ameri genetic relationships (Wiley 1981). Cladistic algorithms hierar can scolytid fauna is virtually complete, thanks to the works of chically arrange species into ancestor-descendant lineages (a S. L. Wood, D. E. Bright, the late G. N. Lanier and G. R. genealogical tree) supported by shared, derived characters, thus Hopping and many others (Bright 1992). Before us lies the task formalizing what good taxonomists have done all along. Re- of developing supportable phylogenetic hypotheses for key cent experimental work with a virus has shown that a known bark beetle taxa, essentially converting lists of species and ancestral genealogy can be fully recovered using a cladistic higher taxonomic groups into ancestor-descendant hierarchies analysis of nucleotide sequences of the terminal viral families or genealogical lineages. (Hillis and others 1992). How can we benefit from having rigorous phylogenetic Bark beetles had been too wee for molecular methods hypotheses for bark beetles? Certain bark beetle taxa offer us using DNA until the advent of the polymerase chain reaction, unusual opportunities for insights into the evolution of po or PCR. Now, isolated mitochondrial, ribosomal or nuclear lygyny, mate choice, pheromonal and stridulatory courtship DNA can, with skill, determination and some luck, be itera communication, host association, and predator-prey interac tively amplified to provide enough reliably replicated DNA for tions. For instance, does pheromonal specificity result from RFLP's (restriction fragment length polymorphisms) or actual ecological interaction of co-occurring species, or does it reflect nucleotide sequencing. These data can have advantages over the intervening time since common ancestry? For species of electrophoretic and morphometric data in that they provide the grandicollis group of Ips, at least, only the latter hypothesis discrete characters suitable for binary, presence-absence cod is supported (Cane and others 1990a, 1990c, Lewis and Cane ing. Because the cladistic approach works with patterns of 1990b). shared, derived characters and not clustering based upon over- Furthermore, it appears that divergence in specificity of all similarity, character states must be discrete rather than stridulatory communication parallels that of pheromone speci continuous so that they can be polarized as either ancestral or ficity (Lewis and Cane 1992) despite its role in defense (Lewis derived at every juncture or branch in the phylogenetic tree. It is and Cane 1990a). Evolutionary biologists and ecologists are comparatively easy to imagine an ancestral Ips either possess coming to appreciate the value of a proper phylogeny for the ing or lacking a third, hooked spine on its elytra. However, for testable evolutionary scenarios that it can offer to comparative a continuous measure such as size, is a length of 4.6 mm primi or evolutionary questions in chemical ecology, plant-herbivore tive or derived? The same problem can be encountered with interactions, parasitology, and studies of symbioses (Cane 1983, karyological, electrophoretic or biochemical data. Nucleotide Gittleman and Kot 1990, Mitter and others 1991, Wanntorp and sequences may provide us with the supplemental discrete, heri others 1990). table characters needed to establish trees of phylogenetic rela Rigorous, testable evolutionary hypotheses for bark beetle tionships for our evolutionary questions (Gittleman and Kot behavior and ecology require equally rigorous phylogenetic 1990, Cracraft and Helm-Bychowski 1991). statements that are defendable on their scientific merits. Unfor I have emphasized the application of genetic information tunately, mere character similarity is insufficient evidence for to the solution of phylogenetic and evolutionary questions for quantifying such phylogenetic relationships, as it confounds bark beetles because of the topic's familiarity to me and rel the possible sources of similarity. Similarity may be the result evance to this workshop. Obviously, there are a number of other prominent and perhaps more pressing genetic questions regarding bark beetles that demand our attention as well. 1An abbreviated version of this paper was presented at the Bark Beetle 1. What is the genetic structuring of select scolytid popula Genetics Workshop, May 17-18, 1992, Berkeley, California. 2Associate Professor Entomology, Dept. Entomology, 301 Funchess tions? In particular, what is the size of a deme for a given Hall, Auburn Univ., Auburn, AL 36849-5413. species during eruptive and quiescent periods of population

14 USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. cycles? What are the respective rates of gene flow during these 3. It is crucial that we gain an understanding of the relative periods? Do allele frequencies shift as a result of natural or contributions of environment and inheritance to the variance sexual selection during either part of these population cycles? we see in bark beetle phenotypes, particularly in their produc If there is an oscillation in selective regimes, is its effect tion, perception and response to semiochemicals. Teale and roughly reciprocal, such that selection in one phase cancels that Hager's work (this workshop) with heritability and covariance in the next? Or do populations gradually change genetically as in pheromone production and response, and its responsiveness a result of such cyclical selection? Or is it not selection at all to imposed selection, is some of the first work in this exciting but drift during the phase of population crash that fixes genetic and much-needed line of investigation. change in populations, the so-called "flush-crash" hypothesis 4. We must not allow ourselves to become complacent or of Hampton Carson? flip about the methodological hurdles that lie in wait of bark 2. Over what distances do some individuals of a species beetle biologists wishing to make use of the tools of molecular migrate? Are outbreaks intensified by an influx of beetles from biology and genetics. As noted by Seybold, by Alosi, and by neighboring populations, or do they result from expansion of Anderson (this workshop), we can benefit from the genetic and the local population? Do outbreaks spawn greater proportions molecular knowledge already accrued for other insect taxa in of dispersing individuals? our hunt for methods to answer our own genetic questions about scolytids.

USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. 15 Cuticular Hydrocarbon Research1

Marion Page2

We have been studying existing taxonomies of forest in- types. Two phenotypes within a species were so different that sects that are based on morphological, genetic and/or behav we used these data as a basis for proposing two subspecies, Z. ioral characteristics to evaluate the utility of cuticular (surface) nevadensis nevadensis and Z. nevadensis nuttingi. hydrocarbons as taxonomic characters (Haverty and others Confident that we were capable of proceeding with our 1988, 1989, Page and others 1990a, 1990b). Cuticular hydro- original purpose involving cone beetles, we embarked on stud carbons are relatively stable metabolic end products that appear ies of Conophthorus spp. and [testing hypothesis on] select to be genetically fixed. Because the insects studied so far species of forest insects. We conducted a study to determine the synthesize all or most of their hydrocarbon components, hydro- degree of similarity or diversity or both among eight of the 15 carbon composition and taxonomic grouping should be related. described species of Conophthorus (Page and others 1990a). Ideally, we would use these chemical characters much as clas From these species we identified 140 hydrocarbons occurring sical taxonomists use morphology, behavior or genetics, i.e., to as individual and isomeric mixtures. Many hydrocarbons were sort the groups of insects on the basis of surface chemical species specific. We discovered that the relatedness of hydro- characters first, rather than after groups have already been carbon profiles for each species parallels existing morphologi sorted on the basis of existing (nonchemical) character criteria. cal keys. These data support the synonymy of C. monticolae However, by comparing our taxonomic separations on the basis with C. ponderosae. Conophthorus from sugar pine could com of cuticular hydrocarbons with existing taxonomic divisions prise a sibling species. Hydrocarbon mixtures of two eastern based on other characteristics, we are broadening the data base species, C. resinosae and C. banksianae, are identical, support of cuticular hydrocarbons as potential taxonomic characters for ing the suspicion that C. banksianae may not be a valid species. forest insects. Closely related pinyon cone beetles, C. cembroides and C. We initially started our studies on the dampwood termites, edulis, have similar combinations of hydrocarbons except for a Zootermopsis, while trying to understand a synonymy of two unique and abundant alkene (C27:1) in C. edulis and two species of scolytid cone beetles, Conophthorus ponderosae and dimethylalkanes in C. cembroides. C. lambertianae. Since these beetles have few useful diagnostic To date we have published the only studies on the cuticular morphological characters, we decided to examine cuticular hydrocarbons of scolytid beetles (Page and others 1990a, 1990b). hydrocarbons as taxonomic characters. To test our understand We examined four species of Dendroctonus that comprise two ing of the current methodology, we repeated the results of sibling species and one pair of morphologically similar species Blomquist and others (1979) on Z. angusticollis. Our initial (Page and others 1990b). Each of these four species has an investigation produced hydrocarbon profiles that differed from abundance of information on behavioral classifications (i.e., those they had published. Had we made an error in methodol location and patterns of larval and adult galleries), host associa ogy? Was their hypothesis about species and caste-specific tions, and taxonomic classification based on host-finding and hydrocarbons correct? Had we observed population or colony mating behaviors, pheromone chemistry, and classical mor variation unreported by them? phological traits. The mountain pine beetle and Jeffrey pine On the basis of morphological characters, our termite speci beetle are sibling species and are difficult to separate on the mens were identified as Z. nevadensis, not Z. angusticollis. basis of morphological characters. The western pine beetle can Blomquist (personal communication) suggested that both our be distinguished from its close relative, the southern pine beetle, laboratories reevaluate our termite collections using identical by a few morphological characters, such as larger body size, methods and gas chromatography parameters. Surprisingly, and on the basis of geographical distribution. We were able to Blomquist's and our laboratory data were identical to those in ascertain that the cuticular hydrocarbon mixtures in these four our first trial but different than his published data. Had we species are species specific. The hydrocarbon patterns of the discovered a sibling species? Additional collections and analy sibling species corroborate their similarity, yet a few hydrocar ses led us to identify an "extra" hydrocarbon phenotype of bon components are unique enough to allow separation. West- Zootermopsis and to find a morphological character for un ern pine beetle and southern pine beetle have hydrocarbon equivocal identification of the three described species of mixtures that are not qualitatively identical. However, they are Zootermopsis (Thome and Haverty 1989). We have separated similar enough to each other to confirm that these species are all species of the dampwood termite by hydrocarbon pheno- closely related, as suggested by electrophoresis analyses.

1An abbreviated version of this japer was presented at the Bark Beetle Genetics Workshop, May 17-18, 1992, Berkeley, California. 2Research Entomologist, Pacific Southwest Research Station, USDA Forest Service, 800 Buchanan Street, Albany, CA 94710.

16 USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. Bark Beetle Genetics―an Overview1

Stephen Teale and Barbara Hager2

Our understanding of bark beetle genetics is fairly rudi ing that the pheromone systems of Ips pini is significantly mentary. A few studies have used enzyme variation to docu altered by predator eavesdropping. An understanding of the ment population structure of Ips and Dendroctonus species in genetic control of pheromone production and response is essen relation to geographic separation (Anderson and others 1979, tial to understanding these reciprocal predator-prey adapta 1983, Namkoong and others 1979), host trees (Langor and tions. Spence 1991, Sturgeon and Milton 1986), dispersal (Florence 2. Heritable variation in host selection traits of bark beetles and others 1982), and phenology (Gast 1987). Electrophoretic and how it can influence population dynamics through interac data have also been used to look at distance relationships of Ips tive selection between host trees and beetles under varying species (Cane and others 1990b). Preliminary studies have population densities. The groundwork in this area was laid by investigated genetics of morphological (Linton and others 1984) Raffa and Berryman (1987). The genetic component involved and pheromonal (Piston and Lanier 1974) traits. in interactions both among and within trophic levels has gener The three main priority areas that we see in need of par ally not been considered in bark beetle population biology. ticular attention (not necessarily in this order) are areas that, 3. The use of DNA techniques, particularly sequencing, in until recently, have not received attention: phylogenetic reconstruction. With an unparalleled array of mating 1. Heritable variation in traits affecting bark beetle-natural systems, the family Scolytidae begs to become a model for the enemy interactions, in particular, the use of bark beetle phero study of the evolution of mating systems as well as the evolu mones by predators and parasitoids in prey location and how it tion of host selection (i.e., which beetle taxa feed on which tree affects heritable qualities of bark beetle populations as well as taxa). This can be done only in a phylogenetic context, and population size. The basis for this research need is reports by DNA sequences yield the most robust phylogenies. Raffa and Klepzig (1989) and Herms and others (1991) show-

1An abbreviated version of this paper was presented at the Bark Beetle Genetics Workshop, May 17-18, 1992, Berkeley, California. 2Assistant Professor and Post Doctoral Associate, respectively, Dept. of Environmental and Forest Biology, Syracuse University of New York, College of Environmental Science and Forestry, Syracuse, NY 13210.

USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. 17 Current Status and Critical Research Needs in Scolytid Genetics1

Kenneth F. Raffa2

During the past two decades, our knowledge of bark beetle Host Selection and the Genetics behavior and ecology has greatly expanded. In particular, stud of Outbreak Behavior ies of pheromone communication, host plant relations, and population dynamics have increased our basic knowledge of Genetic traits associated with host selection may be useful bark beetle biology, improved our ability to manage pest spe in predicting the onset of bark beetle outbreaks. Low density cies, and provided strong models for research on other insect populations are typically concentrated within severely stressed groups. However, bark beetles remain our most damaging trees. Yet during outbreaks, almost all trees are attacked and forest pests nationwide. In addition, the various components of killed, regardless of their vigor. The mechanisms by which scolytid biology are usually studied in isolation, and some key high beetle numbers kill trees is fairly well understood: aggre gaps in our knowledge preclude effective synthesis. In my gation pheromones coordinate rapid mass attacks on entered opinion, a major reason for these difficulties is that insufficient trees, and the tree's resistance capabilities are exhausted. More attention has been given to bark beetle genetics. Increased beetles are required to kill vigorous trees than stressed trees. knowledge in this area could serve as a powerful unifying However, this tells us nothing about how beetles' decisions to theme among the various subdisciplines, and improve our man enter trees could be influenced by their population densities. agement capabilities. Each beetle undergoes a strict behavioral repertoire, which The lack of attention to scolytid genetics is well evidenced culminates in either host entry or resumed flight (Raffa 1988). by the major books and review articles on this family. Most This suggests that different selective pressures may operate on texts and chapters make no, or relatively brief, mention of individual beetles at different beetle densities. At low popula genetics or the role genetics could play in population behavior. tions, the likelihood of an entered beetle being joined by enough Some areas of scolytid genetics have received well-focused recruits to successfully colonize healthy trees is low, and so attention, in particular isozyme variation within and between relatively discriminating behavior may be favored. Conversely, species (Langor and Spence 1991, Stock and Guenther 1979, at high densities, beetles that enter healthy trees are more likely Stock and others 1984). These studies established polymor to be joined by sufficient recruits to kill the host, and so less phism, heterozygosity, and population structuring across geo discriminating behavior is adaptive (also, discriminating types graphic regions and sometimes host species. Some basic work may never find acceptable hosts, as the weakest trees are has been done on cytology (e.g., Lanier 1981), before the colonized by generations leading to the outbreak). Therefore, development of modern methods. However, a number of criti changes in gene frequencies associated with host acceptance cal areas, ranging from the molecular to population levels, could both result from, and partially cause, bark beetle popula remain unexplored. tion increases (Raffa and Berryman 1983, Raffa 1988). The A number of new tools are now available for studying relatively high levels of within-tree, compared to spatial and insect genetics. However, I feel the most critical need is to temporal, genetic (electrophoretic) variation in Dendroctonus generate testable hypotheses regarding how heritable aspects of frontalis (Florence and Kulhavy 1981), and the high levels of scolytid biology relate to their behavior and population dynam genetic diversity that can be maintained in dispersed popula ics. Milton and Schneider (1987) summarized two basic ap tions (Florence and others 1982) could lead to such directional proaches to the role of genetics in insect outbreaks: (1) start selections. Observed behavioral differences between outbreak with a heritable marker, and conduct a broad search for correla and nonoutbreak Ips typographus japonicus (Futura 1989) are tions between gene frequencies and population fluctuation, and consistent with such a pattern. (2) identify life history traits whose variation would greatly There are alternative explanations of how host selection influence population behavior, and determine heritable varia could change through time, such as individuals becoming less tion. Both approaches are valid, and both must be pursued to discriminating as their stores deplete, or pheromones eliciting make relevant gains. As a population ecologist, I will approach host entry in addition to attraction [although Hynum (1978) the issue from the latter perspective. Hopefully, this perspec observed the proportion of beetles resuming flight after landing tive will complement equivalent suggestions arising from the does not decrease during mass attack]. However, the genetics marker-up, and more molecular approaches. From an ecologi of host selection should be considered a high-priority area. This cal/population management view, I think that three critical can be approached by: (1) quantifying for heritable variation in genetic issues must be addressed. beetle selectivity, (2) quantifying genetic diversity throughout the endemic and epidemic phases within an area, and between endemic and epidemic populations at one time, using polymor phic DNA markers such as RFLP's or RAPD's, and (3) testing 1An abbreviated version of this paper was presented at the Bark Beetle Genetics Workshop, May 17-18, 1992, Berkeley, California. alternative hypotheses. 2Forest Entomologist, Dept. of Entomology, University of Wisconsin, Madison, WI 53706

18 USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. Role of Natural Enemies Pheromone Synthesis and Attraction Biological control of bark beetles has received only scant Pioneering work by Lanier (1970c) demonstrated Mende attention, largely because the major pests are native species. lian control over Ips pheromone production and response. Such However, geographic and local variation in predator behavior inheritance patterns can contribute to demic structuring (Cane may provide some avenues for better exploiting natural en and others 1990c). Recent studies of scolytid pheromone com emies. These include introducing predators from distant popu munication have shown strong inter- and intrapopulation dif lations, and biasing against natural enemy removal during ferences within species (Berisford and others 1990, Birgersson semiochemically based population control measures. and others 1988, Miller and others 1989, Seybold and others The major insect predators locate prey by responding to 1992c). These results could have strong implications for bark bark beetle aggregation pheromones. Prey mortality is often beetle biology and management, particularly in guiding extremely high. Thus, selective pressures exerted by predators semiochemically based management strategies and biological may favor alterations in prey semiochemistry that reduce their control (see above). attractiveness to predators, yet retain intraspecific functional Several factors have been proposed as possible contribu ity. Several lines of evidence support this view: (1) predacious tors to heritable genetic variation in scolytid semiochemical beetles, although attracted to scolytid pheromones, may show synthesis and response. For example, it has been proposed that greater affinity for stereoisomers that are relatively less attrac some beetles do not actively engage in host searching, but only tive to (and produced by) the prey (Herms and others 1991, react to pheromone plumes indicative of others' success. The Payne and others 1984, Raffa and Klepzig 1989); (2) minor frequency of such types may change with beetle density pheromone components can have markedly different effects on (Birgersson and others 1988), in a fashion similar to the host predator and prey species (Seybold and others 1992c, Miller selection model proposed in #1. If so, accompanying genetic or and others personal communication); (3) local predator guilds phenotypic markers may be useful in predicting outbreak be may be more attracted to distant than local populations of the havior. Genetically based variation in pheromone chemistry same prey species (Lanier and others 1972, Raffa and Dahlsten may also be important in maintaining the structure of scolytid personal communication). mating populations (Teale personal communication). Studies The underlying genetics of predator-prey interactions are should (1) conduct classical hybridization tests among beetles, completely unknown. No studies on the heritability of predator putatively divergent types (based on arrival time in aggregation responses to bark beetle pheromones have been conducted, sequence and relative responsiveness to host cues in laboratory and only recently has individual variation in bark beetle assays), (2) identify DNA markers associated with putatively semiochemistry been explored (see #3 below). Studies should different behavioral types, and (3) determine the level of gene be aimed at: (1) conducting classical hybridization studies of flow and demic integrity among putatively different types. divergent perception phenotypes within predator species, (2) identifying DNA markers associated with pheromone percep tion among both predators and prey, and (3) quantifying selec tive pressures imposed by predators on beetle reproductive success (at both the colonizing cohort and individual beetle levels) and on predators to locate a particular prey guild.

USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. 19 Genetic Basis of Semiochemically Mediated Bark Beetle-Predator Coevolution: Implications for Developing Mass-Trapping Technologies1

Robert A. Haack. Daniel A. Herms, Bruce D. Ayres, Matthew P. Ayres, and M.L. Doozie Snider2

1991, Raffa and Klepzig 1989). Such variation in the kairomone Background response of Thanasimus could lead to differential mortality of The pheromone chemistry of Ips pini (Coleoptera: Ips adults that respond/produce different enantiomeric blends Scolytidae) has been extensively studied in recent years. In of ipsdienol. Ips that respond/produce low-risk ipsdienol blends eastern North America, I. pini populations respond most strongly theoretically could escape intense predation and increase in to ipsdienol with an enantiomeric composition of about 40 numbers. This change in Ips pheromone production would, percent to 75 percent-(S)-(+) (Herms and others 1991, Lanier eventually exert a reciprocal selective force on its predators, and others 1980, Raffa and Klepzig 1989, Teale 1990). In resulting in a coevolutionary feedback system. western populations, I. pini typically responds maximally to Similar to the selective pressures described above for ipsdienol with an enantiomeric composition of 93 percent to Thanasimus, mass-trapping with a single enantiomeric blend of 100 percent-(R)-(-) (Lanier and others 1980, Miller and others ipsdienol may also cause differential mortality of Ips popula 1989, Seybold and others 1992d). tions and lead to an increase in Ips that respond/produce blends In eastern populations of I. pini, both isomers of ipsdienol of ipsdienol that differ from the pheromone bait being used. In function as attractants (Teale 1990), whereas in most western other words, there is potential for Ips populations to evolve populations the (-)-isomer serves as an attractant and the (+)- resistance to mass-trapping programs that utilize a single blend isomer functions as an interruptant (Birch and others 1980, of ipsdienol. Seybold and others 1992d). Recently, lanierone was demon strated to be an important component of the aggregation phero mone of eastern populations of I. pini (Teale and others 1991), Current Research Objectives but it was not found in western populations (Seybold and others 1992c). We are interested in developing mass-trapping technology Teale (1990) demonstrated in I. pini that the chirality of for managing I. pini populations in midwestern pine woodlots. male-produced ipsdienol was heritable. In addition, Teale and This will require an understanding of the relative genetic and others (personal communication) have shown a strong, positive environmental components to variation in the pheromone re correlation between male response to and production of enan sponses of I. pini and its specialist predators (Herms and others tiomeric blends of ipsdienol. If positive assortive mating were 1991). Heritable variation in beetle responses to different enan common among individuals with similar ipsdienol preference/ tiomeric blends of the aggregation pheromone ipsdienol im production, then genetic substructuring of I. pini populations plies the potential for Ips populations to evolve resistance to could easily occur. Moreover, if movement by Ips adults were mass-trapping programs that use individual blends of ipsdienol. limited between habitat patches (e.g., isolated pine woodlots), This risk is especially great if Ips populations are genetically then further genetic divergence could result on a spatial scale. substructured on the basis of pheromone preferences or if gene Thanasimus dubius (Coleoptera: Cleridae) is a specialist flow between pine woodlots is rare. The risks are complicated predator of Ips and other bark beetles, and can cause significant by secondary impacts on predators that exploit ipsdienol as a bark-beetle mortality (Riley and Goyer 1986, and references kairomone for prey location. Mass-trapping has potential as an therein). Thanasimus dubius uses the pheromone of I. pini as a economically viable, ecologically sound control measure, but kairomone in prey location, and like I. pini, demonstrates prefer its sustained effectiveness may be compromised by disregard ences among different blends of ipsdienol (Herms and others ing the above risks. We will develop models to predict evolu tionary responses of Ips and their predators to different mass- trapping scenarios. The expected product is an economical resistance management program, probably based on sequential or simultaneous use of different ipsdienol blends, and including considerations of predator-prey dynamics. 1An abbreviated version of this paper was presented at the Bark Beetle Genetics Workshop, May 17-18, 1992, Berkeley, California. 2Supervisory Research Entomologist, North Central Forest Experiment Station, USDA Forest Service, Stephen S. Nisbet Bldg., 1407 S. Harrison Road, Current Research Question East Lansing, MI 48823; Staff Entomologist, The Dow Gardens, Midland, Mich.; President, Woods Run Forest Products, Colfax, Wis.; Research Ento To what extent are populations of I. pini and T. dubius mologist, Southern Forest Experiment Station, USDA Forest Service, Pineville, genetically substructured within and between habitat patches La.; Research Technician, Michigan State University, East Lansing, Mich. (pine woodlots)?

20 USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. Experiments in Progress Future Research Plans 1. Electrophoretic studies to test for genetic differentiation We plan to initiate replicated mass-trapping studies in within and among populations of I. pini (and its clerid predator, several isolated red pine woodlots in Michigan in 1992. If there T. dubius) based on ipsdienol-blend preferences. is a genetic component to the observed variation in pheromone 2. Mark-recapture studies to test pheromone-response speci- response, we should be able to manipulate pheromone response ficity of I. pini. profiles by exerting selection through mass-trapping. Direc- 3. Comparison of pheromone-response profiles of I. pini tional selection (e.g., using one pheromone blend) should change populations from several isolated red-pine stands in Michigan. the response profile, whereas stabilizing selection (e.g., using Assuming a genetic component exists to the observed intrapo- several blends) should maintain it. If the genetic component to pulation variation, then genetic drift or differential selection pheromone response is weak relative to environmental influ- pressures could result in stand-to-stand differences on a local ences, or if there is substantial gene flow within and among scale. woodlots, then mass-trapping should have little effect on response profiles of local I. pini populations.

USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. 21 Population Genetics of Spruce Bark Beetle Ips typographus (Col., Scolytidae) and Related Ips Species1

Christian Stauffer, Renate Leitinger, Erwin Fuehrer2

Enzyme electrophoresis proved to be a useful method to them originating from different glacial refuge areas (Central study population genetic aspects in Ips typographus (L.) be- Russia, Carpathian Mountains, Dinaric Alps, respectively). Thus cause of three highly polymorphic loci. Aspartate aminotrans- it seems likely that I. typographus populations had been subject ferase locus-2 (Aat-2), physiologically involved in the transfer to genetic differentiation by long-term isolation during the of aminogroups, has six alleles per population. Amylase locus- glacial period and postglacial remigration to middle and north 1 (Amy-1), physiologically involved in the hydrolysis of starch, Europe, parallel to its host tree (Leitinger and others 1992). has 10 alleles per population, and esterase locus-2 (Est-2), Preliminary results indicate, moreover, that within physiologically involved in processes of detoxification and the I. typographus populations the genetic variance of samples metabolism of juvenile hormone, has 16 alleles per population. collected from pheromone traps differs more or less from those The allozymes, pattern, and the mode of inheritance of nearly which had been collected from infested logs. As to the genetic all alleles were genetically proved. The different amount of divergence between log-trapped and pheromone-trapped samples, alleles among allozymes could reflect the physiological role of a slight trend from middle European lowland populations to- the enzymes (Johnson 1974); therefore most studies were cal- ward upland and Scandinavian populations, respectively, was culated individually (Stauffer and others 1992b). ascertained (Leitinger 1991). Because subpopulations of A related but distinctly different species is the spruce bark I. typographus, which are colonizing different spruce logs, also beetle Ips amitinus. This relationship could be also demon- differ genetically, a genetically determined variance of orienta- strated by enzyme electrophoresis. The existence of I. amitinus tion patterns within this bark beetle species is assumed. This and I. amitinus var. montana (Fuchs 1913), which appeared to phenomenon may concern the pioneer problem too. Voltinism be doubtful, was investigated by behavioral, morphological is another property with epidemiological significance, which and biochemical (cuticular analysis and enzyme electrophore- seems to be genetically determined within populations of sis) techniques. Because no obvious differences between the I. typographus. Depending on different altitudes, the propor- two races could be found, the distinction of a var. montana tion of obligately and facultatively diapausing beetles is differ- becomes very unprobable (Zuber and others 1992). ent. First attempts to biochemically identify both diapausing Efficiency of a commercial pheromone product, which is types revealed promising results. commonly used in central Europe for trapping I. typographus, From the epidemiological point of view, work will be proved to be regionally different. A genetic basis of these focused on functional and ecological population genetics. Us- differences was assumed. Seventeen European populations of ing the allozyme method for identification of different "types," this species, collected from infested logs, were studied. The the functionality of enzymes as well as enzyme activity must be dendrograms and an Alscal analysis of Aat-1 and Est-2 clus- studied more in detail. We proposed to begin with the juvenile tered three groups―the northern (Scandinavian) populations, hormone esterase (Stauffer and others 1992a), the the southern (alpine) populations, and the middle European monooxygenase (Sturgeon and Robertson 1985), and the gen- (prealpine/Czechoslovakian) populations. Distribution of these eral esterases. three genetic groups corresponds to the three parts of natural distribution of the host tree, Picea abies, in Europe, each of

1An abbreviated version of this paper was presented at the Bark Beetle Genetic Workshop, May 17-18, 1992, Berkeley, CA. 2Institute of Forest Entomology, Forest Pathology and Forest Protection, Universität für Bodenkultur, A-1190 Vienna, Austria.

22 USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. Novel Bark Beetle Research Possible with New Genetic Techniques1

Ken Hobson and Owain Edwards2

Prospects ing enough polymorphism to conduct very fine-grained analyses of this sort. Now several of the new molecular techniques The new molecular techniques in genetics offer solutions permit rapid analysis of DNA with enough resulting informa- to questions about bark beetle biology in two principal areas. tion to carry out gene flow and dispersal analyses with dozens The first of these is dominated by questions of phylogenetic of markers resulting in more precise estimates. interest. For example, the unity of Dendroctonus valens as a Contrasts that exist between beetles in an endemic or early species (e.g., Pajares and Lanier 1990) could be explored with infestation population and an epidemic or late infestation popu- precision. Geographic substructuring of other scolytid taxa, lation may involve genetic drift or perhaps repeating cycles of including regional differences in pheromone production and shifting gene frequencies. response, partial mating barriers in sibling Ips and so on, share this phylogenetic approach. Much of this work is well under- way. Methods The second area, which is wide open for new ideas, uses Sequencing of both mitochondrial and genomic DNA has the ability of the new techniques to discriminate individuals provided information previously unattainable to systematists. and populations to address questions from population dynam- These data, in conjunction with other chemical phenotype ics and behavior (Slatkin 1987). It is possible now with mam- data, such as cuticular hydrocarbon analysis (e.g., Page and others malian and bird DNA to characterize individuals and verify 1990b), will provide answers to many bark beetle phylogenetic genetic contributions of parents to their offspring. Paternity questions. analyses of this sort have recently provided some surprising However, discrimination of bark beetle populations re- insights in ornithological literature where extra-pair copula- quires molecular techniques that detect higher levels of poly- tions have been found to be much more common than was morphism. DNA sequencing of highly polymorphic regions of previously recognized. Little is known about bark beetle be- DNA can be useful, but these regions must first be located. havior under the bark―e.g., whether supposedly monogamous Sequencing is costly and labor-intensive, so its application in Dendroctonus females sometimes mate with more than one population genetics can only be limited. Restriction Fragment male before ovipositing their first clutch of eggs, whether there Length Polymorphism (RFLP) has also been used for the detec- is selection by females for males with particular characteristics, tion of DNA polymorphism. However, RFLP analysis requires and so on. The ability to identify individuals and their descen- that knowledge of a polymorphic region to target exists, and its dants would allow paternity analyses of eggs from scolytid sequence must be known so that a DNA probe can be con- galleries. Male-male competition may be occurring in D. valens structed. RFLP analysis is much less labor-intensive than se- galleries where there are three adults present (Hobson 1992). quencing, but it is still quite costly. Radioactivity is also neces- Paternity analysis of egg clutches of even single pairs of beetles sary for both sequencing and RFLP. Because of the drawbacks could provide insights into the frequency of pre-emergence of these DNA techniques, protein allozyme analysis has been mating. the most popular method for performing genetic polymorphism The ability to identify beetles from subpopulations and studies. Allozyme analysis is much less expensive, but it is track their descendants may allow us to answer questions such often difficult to find polymorphic loci. as whether patch kills of trees by D. brevicomis are caused by The polymerase chain reaction (PCR) has made it much beetles which are the descendants of an earlier nearby infesta- simpler and more cost-effective to perform molecular analyses tion or instead are beetles which are drawn from the general at the level of the DNA (Arnheim and others 1990). Because population of all dispersing adults. PCR dramatically increases the copy number of specific DNA Similarly, typical bark beetle dispersal distances can be sequences, analyses using PCR can be performed with very assessed by looking at variability along transects through beetle little starting DNA. DNA sequencing of PCR-amplified DNA populations. The number of migrants per generation between eliminates the need for cloning, which dramatically reduces the populations can be estimated from FST statistics as estimated cost and effort. RFLP analysis using PCR-amplified DNA using the allele frequencies of several different genes. Studies eliminates the need for a probe, so less sequence information is employing electrophoresis have had difficulty in the past find- required, and the need for radioactivity is eliminated. However, the level of polymorphism detected using these techniques may still limit their usefulness in answering some population genet- 1An abbreviated version of this paper was presented at the Bark Beetle Genetics Workshop, May 17-18, 1992, Berkeley, California. ics questions. 2Graduate Students, Dept. of Entomology, Univ. of California, 201 Wellman Hall, Berkeley, CA 94720.

USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. 23 In the last two years, a PCR-based technique called Ran gions of DNA. In humans, this is accomplished by analyzing domly Amplified Polymorphic DNA (RAPD) has proven very Variable Number Tandem Repeats (VNTR's), which are highly successful for performing genetic analyses in plants (Williams mutable (10-3-10-4) regions of repetitive DNA (Jeffreys and and others 1990). Its advantages are that it detects higher levels others 1985). The major disadvantage of this technique is that of polymorphism than either isozyme or RFLP analyses, and few VNTR's have been found in insects (Bigot and others that no prior DNA sequence knowledge is necessary. A disad 1990, Blanchetot 1991, Moritz and others 1991), so it is likely vantage to this technique is that the electrophoretic bands it that the application of this analysis to the study of bark beetles produces are inherited as completely dominant traits as op would first require significant time and effort to locate the posed to semi-dominant traits as in RFLP or isozyme analysis, repetitive DNA. Once this was accomplished a very high de which can make genetic analyses somewhat more complicated. gree of polymorphism would be available for free grain dis It has also been reported that some RAPD bands are not inher persal studies and behavioral investigations requiring charac ited in a Mendelian fashion (Riedy and others 1992). Success terization of an individual and its progeny. with this technique has been reported with some insects (Edwards, Given the diversity of techniques for assessing polymor unpublished results), including haplodiploid species where the phism and discriminating groups under study, one of our first dominance problem can be avoided by performing the analysis goals might be to survey the level of polymorphism that is on males. found in the groups of interest. Then the appropriate technique The characterization of individuals, as needed for paternity for a particular problem can be chosen. analysis, requires an analysis that targets highly variable re

24 USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. Systematics Research1

Donald E. Bright2 that will improve understanding of phylogeny, evolution, ge neric and tribal relationships, and other such basic systematic The following comments about bark beetle genetics reflect questions. my current thoughts on the status of bark beetle systematics and Morphological studies have been the backbone of taxo present some of my ideas concerning future needs. nomic research for decades and will probably remain of pri The basic alpha taxonomy of North American bark beetles mary importance for decades to come. However, we now have is essentially finished. All of the species occurring in North an exciting array of other methods that may prove to be of America are, more or less, well characterized at the alpha level immense importance. Isozyme analysis, cuticular hydrocarbon and their taxonomy is well established. An authoritative name analysis and PCR techniques will be increasingly used in the can now be consistently given to almost any specimen or series future to help resolve some of the questions mentioned above. of specimens. Keys and descriptions to all species in all genera A molecular systematics laboratory is being established at of North American scolytids are now available. A catalog of the the Biological Resources Division, Canada Department of Ag world fauna will soon be available. It is extremely unlikely that riculture, in Ottawa, Ontario. I hope it will be able to address very many morphologically distinct, unnamed species of North some of the questions raised above. American Scolytidae remain to be discovered. Any newly dis One concern for the future of scolytid systematics is that in covered species will probably be found in either host plants of less than 10 years the currently working North American very restricted distributions (e.g., Santa Lucia fir), in the vari systemists will have all retired. I am not aware that any current ous desert shrubs or deciduous trees, or as currently unrecog graduate student is working on scolytid systematics, nor are nized sibling species. I would estimate that fewer than 10 any young, currently employed researchers working on this morphologically distinct, unnamed species remain undiscov group. I urge university faculty to seek out qualified individuals ered in North America. and guide them into the systematic study of the Scolytidae. This, however, does not mean that systematic investiga Forest entomology has been fortunate during the past three tions are no longer needed. Indeed, we are now able to embark decades in having several active individuals interested in sys on the most exciting aspect of systematic research. With all of tematic studies of bark beetles, and this interest needs to be this basic knowledge, we are now left with the task of resolving continued. a number of species groups, subspecies, host/geographic races, sibling species, and so on. Unresolved species groups are known to exist in Dendroctonus, Ips, Pityophthorus, probably Scolytus, and undoubtedly in a number of less studied genera. Also, on a more fundamental level, there is a need to develop information

1An abbreviated version of this paper was presented at the Bark Beetle Genetics Workshop, May 17-18, 1992, Berkeley, California. 2Research Scientist, Canada Dept. of Agriculture, Systematics Research, Centre for Land and Biological Resources Research, Ottawa, Ontario K1A 0C6

USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. 25 References

Alosi, M. Carol; Robertson, Jacqueline L. 1992. Biochemical indicators of Blanchetot, A. 1991. A Musca domestica satellite sequence detects indi population status of bark beetles [this proceedings]. vidual polymorphic regions in insect genome. Nucleic Acid Research 19: 929-932. Amman, G.D. 1982. Characteristics of mountain pine beetles reared in four pine hosts. Environmental Entomology 11: 590-593. Blomquist, G.J.; Howard, R.W.; McDaniel, C.A. 1979. Structure of the cuticular hydrocarbons of the termite Zootermopsis angusticollis (Hagen). Amman, G.D.; Cole, W.E. 1983. Mountain pine beetle dynamics in lodge- Insect Biochemistry 9:371-374. pole pine forest. Part II: population dynamics. Gen. Tech. Rep. WT-145. Ogden, UT: Intermountain Forest and Range Experiment Station, Forest Bright, D.E. 1976. The insects and arachnids of Canada, Part 2. The bark Service, U.S. Department of Agriculture; 59 p. beetles of Canada and Alaska. Publication 1576. Ottawa, Ontario, Canada: Canadian Department of Agriculture; 241 p. Amman, G.D.; Pasek, J.E. 1986. Mountain pine beetle in ponderosa: effects of phloem thickness and egg gallery density. Res. Paper INT-376. Ogden, Bright, D.E. Systematics of bark beetles. In: Schowalter, T.D.; Filip, G.M., UT: Intermountain Forest and Range Experiment Station, Forest Service, eds. Beetle-pathogen interactions in conifer forests. New York: Aca U.S. Department of Agriculture; 7 p. demic Press. [In press.]

Amman, G.D.; Stock, M.W. Effect of phloem thickness on heterozygosity of Bright, Donald E. 1992. Systematics research [this Proceedings] laboratory-reared mountain pine beetles (Dendroctonus ponderosae Bright, D.E., Jr.; Stark, R.W. 1973. The bark and ambrosia beetles of Califor Hopkins, Coleoptera: Scolytidae). Unpublished draft supplied by author. nia: Scolytidae and Platypodidae. Bulletin California Insect Survey 16. Anderson, W.W.; Berisford, C.W. 1992. Bark beetle genetics. [this Proceed Berkeley: University of California Press; 169 p. ings] Bright, D.E.; Stock, M.W. 1982. Taxonomy and geographic variation. In: Anderson, W.W.; Berisford, C.W.; Kimmich, R.H. 1979. Genetic differences Mitton, J.B., Sturgeon, K.B., eds. Bark beetles in North American coni among five populations of the southern pine beetle. Annals of Entomo fers. Austin: University of Texas Press; 46-73. logical Society of America 72: 323-327. Brillinger, D.R. 1990. Spatial-temporal modelling of spatially aggregate Anderson, W.W.; Berisford, C.W.; Tumbow, R.H.; Brown, C.J. 1983. Genetic birth data. Survey Methodology 16: 255-269. differences among populations of the black turpentine beetle, Dendroctonus Bush, G.L. 1969. Sympatric host race formation and speciation in frugivo terebrans, and an engraver beetle, Ips calligraphus (Coleoptera: Scolytidae). rous flies in genus Rhagoletis (Diptera, Tephritidae). Evolution 23: 237- Annals of Entomological Society of America 76: 896-902. 251. Arnheim, N.; White, T.; Rainey, W.E. 1990. Application of PCR: organismal Cane, J.H. 1983. Chemical evolution and chemosystematics of the Dufour's and population biology. BioScience 40: 174-182. gland secretions of the lactone-producing bees (Hymenoptera: Colletidae, Bartlet, R.J.; Schaner, A.M.; Jackson, L.L. 1986. Aggregation pheromones in Halictidae, Oxaeidae). Evolution 37: 657-674. five taxa of the Drosophila virilis species group. Physiological Entomol Cane, James H. 1992. Genetic characters for bark beetle phylogenies [this ogy 11: 367-376. Proceedings]. Bentz, B.J.; Stock, M.W. 1986. Phenetic and phylogenetic relationships Cane, J.H.; Merrill, L.D.; Wood, D.L. 1990a. Attraction of pinyon pine bark among ten species of Dendroctonus bark beetles (Coleoptera: Scolytidae). beetle, Ips hoppingi, to conspecific and I. confusus pheromones (Co Annals of Entomological Society of America 79: 527-534. leoptera: Scolytidae). Journal of Chemical Ecology 16(10): 2791-2798. Berisford, C.W.; Payne, T.L.; Berisford, Y.C. 1990. Geographical variation Cane, J.H.; Stock, M.W.; Wood, D.L.; Gast, S.J. 1990b. Phylogenetic rela in response of southern pine beetle (Coleoptera: Scolytidae) to aggregat tionships of Ips bark beetles (Coleoptera: Scolytidae): Electrophoretic and ing pheromones in laboratory assays. Environmental Entomology 19: morphometric analyses of the grandicollis group. Biochemical Sys 1671-1674. tematics and Ecology 18: 359-368. Berryman, A.A. 1972. Resistance of conifers to invasion by bark beetles- Cane, J.H.; Wood, D.L.; Fox, J.W. 1990c. Ancestral semiochemical attrac fungus associations. BioScience 22: 598-602. tion persists for adjoining populations of sibling Ips bark beetles (Co Bigot, Y.; Hamelin, M: H.; Periquet, G. 1990. Heterochromatin condensation leoptera: Scolytidae). Journal of Chemical Ecology 16: 993-1013. and evolution of unique satellite-DNA families in two parasitic wasp Charlesworth, B.; Langley, C.H. 1989. The population genetics of Droso- species: Diadromus pulchellus and Eupelmus vuilleti (Hymenoptera). phila transposable elements. Annual Review of Genetics 23: 251-287. Molecular Biology and Evolution 7: 351-364. Clarke, Geoffrey M.; McKenzie, Leslie J. 1992. Fluctuating asymmetry as a Birch, M.C.; Light, D.M.; Wood, D.L.; Browne, J.E.; Silverstein, R.M.; quality control indicator for insect mass rearing processes. Journal of Bergot, B.J.; Ohloff, G.; West, J.R.; Young, J.C. 1980. Pheromonal Economic Entomology 85: 2045-2050. attraction and allomonal interruption of Ips pini in California by the two enantiomers of ipsdienol. Journal of Chemical Ecology 6(3): 703-717. Correll, J.C.; Gordon, T.R.; McCain, A.H. 1992. Genetic diversity in Califor nia and Florida populations of the pitch canker fungus Fusarium Birgersson, G.; Schlyter, F.; Bergstrom, G.; Lofquist, J. 1988. Individual subglutinans f. sp. pini. Phytopathology 82: 415-420. variation in the aggregation pheromone of the spruce bark beetle, Ips typographus. Journal of Chemical Ecology 14: 1737-1761. Coyne, J.F.; Lott, L.H. 1976. Toxicity of substances in pine oleoresin to southern pine beetles. Journal of Georgia Entomological Society 11: 301- Birgersson, G.; Schlyter, F.; Lofqvist, J.; Bergstrom, G. 1984. Quantitative 305. variation of pheromone components in the spruce bark beetle Ips typographus from different attack phases. Journal of Chemical Ecology Cracraft, J.; Helm-Bychowski, K. 1991. Parsimony and phylogenetic infer 10: 1029-1055. ence using DNA sequences: some methodological strategies. In: Miyamoto, M.M.; Cracraft, J., eds. Phylogenetic analysis of DNA sequences. New York: Oxford University Press; 184-220.

26 USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. Dauterman, W.C.; Hodgson, E. 1978. Detoxification mechanisms in insects. Greene, Lula E. 1992. Qualitative genetics of mountain pine beetle in central In: Rockstein, M., ed. Biochemistry of insects. New York: Academic Oregon [this Proceedings]. Press: 541-577. Haack, Robert A.; Herms, Daniel A.; Ayres, Bruce D.; Ayres, Matthew P.; Diel, S.R.; Bush, G.L. 1984. An evolutionary and applied perspective of Snider, M.L. Doozie. 1992. Genetic basis of semiochemically mediated insect biotypes. Annual Review of Entomology 29: 471-504. bark beetle-predator coevolution: implications for developing mass-trapping technologies [this Proceedings]. Engels, W.R. 1983. The P family of transposable elements in Drosophila. Annual Review of Genetics 17: 315-344. Hackett, K.J.; Lynn, D.E.; Williamson, D.L.; Ginsberg, A.S.; Whitcomb, R.H. 1986. Cultivation of the Drosophila sex-ratio spiroplasma. Science Fish, R.H.; Browne, L.E.; Bergot, B.J. 1984. Pheromone biosynthetic path- 232: 1253-1255. ways: conversion of ipsdienone to (-)-ipsdienol, a mechanism for enantioselective reduction in the male bark beetle, Ips paraconfusus. Haverty, M.I.; Page, M.; Blomquist, G.J. 1989. Value of cuticular hydrocar- Journal of Chemical Ecology 10: 1057-1064. bons for identifying morphologically similar species of pine cone beetles. Proc. 3rd Cone and Seed Insect Working Party, IUFRO, Victoria, B.C., Fish, R.H.; Browne, L.E.; Wood, D.L.; Hendry, L.B. 1979. Pheromone Canada. 50-62. biosynthetic pathways: Conversions of deuterium labelled ipsdienol with sexual and enantioselectivity in Ips paraconfusus Lanier. Tetrahedron Haverty, M.I.; Page, M.; Nelson, L.J.; Blomquist, G.J. 1988. Cuticular hydro- Letters 17: 1465-1468. carbons of the dampwood termites, Zootermopsis: Intra- and intercolony variation and potential as taxonomic characters. Journal of Chemical Florence, L.Z.; Johnson, P.C.; Coster, J.E. 1982. Behavioral and genetic Ecology 14: 1035-1058. diversity during dispersal: analysis of a polymorphic esterase locus in southern pine beetle, Dendroctonus frontalis. Environmental Entomol- Hayes, J.L.; Robertson, J.L. 1992. An (ecologically biased) view of the ogy 11: 1014-1018. current status of bark beetle genetics and future research needs. [this Proceedings] Florence, L.Z.; Kulhavy, D.L. 1981. Genetic variation and population structure of the southern pine beetle, Dendroctonus frontalis. In: Stock, M.W., Hendry, L.B.; Piatek, B.; Browne, L.E.; Wood, D.L.; Byers, J.A.; Fish, R.H.; ed. Application of genetics and cytology in insect systematics and evolu- Hicks, R.A. 1980. In vivo conversion of a labelled host plant chemical to tion: proceedings of symposium, National ESA Meeting; December 1-2, pheromones of the bark beetle Ips paraconfusus. Nature 284: 485. 1980; Atlanta, GA. Moscow, ID: Forest, Wildlife, and Range Experi- ment Station, Univ. of Idaho; 141-152. Herms, D.A.; Haack, R.A.; Ayres, B.D. 1991. Variation in a semiochemical- mediated prey-predator interaction: Ips pini (Scolytidae) and Thanasimus Fox, J.W.; Wood, D.L.; Cane, J.H. 1991 a. Interspecific pairing between two dubius (Cleridae). Journal of Chemical Ecology 17: 1705-1714. sibling Ips species (Coleoptera: Scolytidae). Journal of Chemical Ecol- ogy 17: 1421-1435. Higby, P.K.; Stock, M.W. 1982. Genetic relationships between two sibling bark beetle species, the Jeffrey pine beetle and the mountain pine beetle, Fox, J.W.; Wood, D.L.; Koehler, C.S. 1990. Distribution and abundance of in northern California. Annals of Entomological Society of America 75: engraver beetles (Scolytidae: Ips Species) on Monterey pines infected 668-674. with pitch canker. Canadian Entomologist 122: 1157-1166. Hillis, D.M.; Bull, J.J.; White, M.E.; Badgett, M.R.; Molineux, I.J. 1992. Fox, J.W.; Wood, D.L.; Koehler, C.S.; O'Keefe, S.T. 1991b. Engraver beetles Experimental phylogenetics: generation of a known phylogeny. Science (Scolytidae: Ips species) as vectors of the pitch canker fungus, Fusarium 255: 589-592. subglutinans. Canadian Entomologist 123: 1355-1367. Hobson, K.R. 1992. Studies on host selection and biology of Dendroctonus Fuchs, A.G. 1913. Forstzoologiche Ergebnisse einer Sommerreise ins Engadin. valens and fungal host interactions of Dendroctonus brevicomis. Berke- III. Die Arven, Larchenund Fichtenborkenkafer des Engadin. ley: University of California. Dissertation. Naturwissenschaftliche Zeitschrift fur Forst-und Land-wirtschaft 11(2); Hobson, Ken; Edwards, Owain. 1992. Novel bark beetle research possible 65-86. with new genetic techniques. [this Proceedings] Furniss, M.M.; Schenk, J.A. 1969. Sustained natural infestations by the mountain pine beetle in seven new Pinus and Picea hosts. Journal of Homvelt, R.; Christiansen, E.; Solheim, H.; Wang, S. 1983. Artificial inoculation with Ips typographus-associated fungi can kill Norway spruce Economic Entomology 62: 518-519. trees. Meddelelser fra Norsk Instituut for Skogforskning 38(4): 1-20. Futura, K. 1989. A comparison of endemic and epidemic populations of the spruce beetle (Ips typographus japonicus Niijima) in Hokkaido. Journal Hynum, B.G. 1978. Migration phase interactions between the mountain pine beetle and lodgepole pine. Pullman: Washington State University. Dis- of Applied Entomology 107: 289-295. sertation. Gast, S.J. 1987. Factors affecting host and pheromone attraction and a comparison of genetic diversity in consecutive generations of Ips pini Ikeda, H. 1965. Interspecific transfer of the "sex-ratio" agent of Drosophila willistoni in Drosophila bifasciata and Drosophilia melanogaster. Sci- (Say)(Coleoptera: Scolytidae) in Idaho. Moscow: Univ. of Idaho. M.S. ence 147: 1147-1148. Thesis. Gast, S.J.; Furniss, M.M. Electrophoretic comparison of Dendroctonus Jeffreys, A.J.; Wilson, V.; Thein, S.L. 1985. Hypervariable "minisatellite" regions in human DNA. Nature 314: 67-73. punctatus LeConte and D. micans (Kugelann) (Coleoptera: Scolytidae). Unpublished draft supplied by author. Johnson G.B. 1974. Enzyme polymorphism and metabolism. Science 184: Gast, S.J.; Stock, M.W. Genetic diversity in consecutive generations of Ips 283-292. pini (Say) (Coleoptera: Scolytidae) in Idaho. Unpublished draft supplied Kaneshiro, K.Y. 1988. Speciation in the Hawaiian Drosophila. BioScience by author. 38: 258-263.

Gittleman, J.L.; Kot, M. 1$90. Adaptation: statistics and a null model for Kemp, W.P.; Kalaris, T.M.; Quimby, W.F. 1989. Rangeland grasshopper estimating phylogenetic effects. Systematic Zoology 39: 227-241. (Orthoptera: Acrididae) spatial variability: Macroscale population as- sessment. Journal of Economic Entomology 82: 1270-1276. Green, M.M. 1980. Transposable elements in Drosophila and other Diptera. Annual Review of Genetics 14: 109-120.

USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. 27 Langor, D.W. 1989. Host effects on the phenology, development, and mortal- Lewis, E.E.; Cane, J.H. 1992. Inefficacy of courtship stridulation as a premating ity of field populations of the mountain pine beetle, Dendroctonus ethological barrier for Ips bark beetles (Coleoptera: Scolytidae). Annals ponderosae Hopkins (Coleoptera: Scolytidae). Canadian Entomologist of Entomological Society of America 85(4): 517-524. 121: 149-157. Liebhold, A.M.; Elkinton, J.S. 1989. Characterizing spatial patterns of gypsy Langor, D.W.; Spence, J.R. 1991. Host effects on allozyme and morphologi- moth regional defoliation. Forest Science 35: 557-568. cal variation of the mountain pine beetle, Dendroctonus ponderosae Hopkins (Coleoptera: Scolytidae). Canadian Entomologist 123: 395-410. Linton, D.A.; Safranyik, L.; Whitney, H.S.; Spanier, O.J. 1984. Possible genetic control of color morphs of spruce beetles. Research Notes 4: 53. Langor, D.W.; Spence, J.R.; Pohl, G.R. 1990. Host effects on fertility and Victoria, B.C., Canadian Forest Service. reproductive success of Dendroctonus ponderosae Hopkins (Coleoptera: Scolytidae). Canadian Entomologist 121: 149-157. McCambridge, W.F. 1975. Scotch pine and mountain pine beetles. Green Thumb 32: 87. Lanier, G.N. 1967. Biosystematic, cytological, and sex ratio studies of closely Merriam, J.; Ashbumer, M.; Hard, D.L.; Kafatos, F.C. 1991. Toward cloning related bark beetles (Coleoptera: Scolytidae). Berkeley: University of California; 110 pp. Dissertation. and mapping the genome of Drosophila. Science 254: 221-225. Merrill, L.A. 1991. Biological barriers to hybridization in closely related Lanier, G.N. 1970a. Biosystematics of the genus Ips (Coleoptera: Scolytidae) species of Ips (Coleoptera: Scolytidae). Berkeley: University of Califor- in North America. Hopping's group III. Canadian Entomologist 102: 1404-1423. nia; 121 p. Dissertation. Miller, D.R. 1990. Reproductive and ecological isolation: community struc- Lanier, G.N. 1970b. Biosystematics of North American Ips (Coleoptera: ture in the use of semiochemicals by pine bark beetles (Coleoptera: Scolytidae): Hopping's group IX. Canadian Entomologist 102: 1139- 1163. Scolytidae). Burnaby, B.C., Canada: Simon Fraser University; 166 p. Dissertation. Lanier, G.N. 1970c. Sex pheromones: abolition of specificity in hybrid bark Miller, D.R.; Borden, J.H.; Slessor, K.N. 1989. Inter- and intrapopulation beetles. Science 169: 71-72. variation of the pheromone ipsdienol produced by male pine engravers, Lanier, G.N. 1972. Biosystematics of the genus Ips (Coleoptera: Scolytidae) Ips pini (Say) (Coleoptera: Scolytidae). Journal of Chemical Ecology in North America. Hopping's groups IV and X. Canadian Entomologist 15(1): 233-247. 104: 361-388. Mitchell, R.G.; Preisler, H.K. 1991. Analysis of spatial patterns of lodgepole Lanier, G.N. 1981. Cytotaxonomy of Dendroctonus. In: Stock, M.W., ed. pine attacked by outbreak populations of the mountain pine beetle. Forest Application of genetics and cytology in insect systematics and evolution. Science 37: 1390-1408. proceedings of symposium, National ESA Meeting; December 1-2, 1980; Atlanta, GA. Moscow, ID: Forest, Wildlife, and Range Experiment Sta- Mitter, C.; Farrell, B.; Futuyma, D.J. 1991. Phylogenetic studies of insect- plant interactions: insights into the genesis of diversity. Tree 6: 290-293. tion, University of Idaho; 33-66. Mitton, C.; Schneider, J.C. 1987. Genetic change and insect outbreaks. In: Lanier, G.N.; Birch, M.C.; Schmitz, R.F.; Furniss, M.M. 1972. Pheromones of Ips pini (Coleoptera: Scolytidae): variation in response among three Barbosa, P.; Schultz, J.C., eds. Insect outbreaks. New York: Academic Press; 505-532. populations. Canadian Entomologist 104: 1917-1923. Mitton, J.B.; Sturgeon, K.B. 1982. Bark beetles in North American conifers: Lanier, G.N.; Claesson, A.; Stewart, T.; Piston, J.; Silverstein, R.M. 1980. Ips pini: the basis for interpopulational differences in pheromone biology. a system for the study of evolutionary biology. Austin: University of Texas Press; 527 p. Journal of Chemical Ecology 6: 677-687. Moritz, R.F.A.; Meusel, M.S.; Haberl, M. 1991. Oligonucleotide DNA fin- Lanier, G.N.; Oliver, J.H. 1966. Sex-ratio condition: unusual mechanisms in bark beetles. Science 153: 208-209. gerprinting discriminates super- and half-sisters in honeybee colonies (Apis melifera L.). Naturwissenschaften 78: 422-424. Lanier, G.N.; Wood, D.L. 1975. Specificity of response to pheromones in the Namkoong, G.; Roberds, J.H.; Nunnally, L.B.; Thomas, H.A. 1979. Isozyme genus Ips (Coleoptera: Scolytidae). Journal of Chemical Ecology 1(1): 9- 13. variations in populations of southern pine beetles. Forest Science 25: 197-203. Leary, R.F.; Allendorf, F.W. 1989. Fluctuating asymmetry as an indicator of Novitski, E.; Peacock, W.J.; Engel, J. 1965. Cytological basis of "sex ratio" stress: implications for conservation biology. Trends in Ecology & Evo- lution 4: 214-217. in Drosophila pseudoobscura. Science 148: 516-517. Owen, D.R.; Lindahl, K.Q., Jr.; Wood, D.L.; Parmeter, J.R., Jr. 1987. Patho- Leitinger, R. 1991. Populationsgenetische Untersuchungen an Ips typographus genicity of fungi isolated from Dendroctonus valens, D. brevicomis, D. (Col., Scolytidae) outer geographischem and verhaltens-biologischem Aspekt. Dipl. Thesis BOKU Vienna. ponderosae to ponderosa pine seedlings. Phytopathology 77: 631-636. Page, Marion. 1992. Cuticular hydrocarbon research [this Proceedings]. Leitinger R.; Stauffer, Ch.; Fuehrer, E. 1992. Postglacial remigration of the European Ips typographus (Col.: Scolytidae) populations from three Page, M.; Nelson, L.J.; Blomquist; G.J.; Seybold, S.J. 1992. Cuticular hydro- refugial areas. Entomologia generalis. Unpublished draft supplied by carbons as chemosystematic characters of pine engraver beetles (Ips sp.) author. (Coleoptera: Scolytidae) in the Grandicollis subgeneric group. Unpub- lished draft supplied by author. Lewis, E.E.; Cane, J.H. 1990a. Stridulation as a primary antipredator defence of a beetle. Animal Behavior 40: 1003-1004. Page, M.; Nelson, L.J.; Haverty, M.I.; Blomquist, G.J. 1990a. Cuticular hydrocarbons of eight species of North American cone beetles, Lewis, E.E.; Cane, J.H. 1990b. Pheromonal specificity of Southeastern Ips Conophthorus Hopkins. Journal of Chemical Ecology 16: 1178-1193. pine bark beetles reflects phylogenetic divergence (Coleoptera: Scolytidae). Canadian Entomologist 122: 1235-1238. Page, M.; Nelson, L.J.; Haverty, M.I.; Blomquist, G.J. 1990b. Cuticular hydrocarbons as chemotaxonomic characters for bark beetles: Dendroctonus

28 USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. ponderosae Hopkins, D. jeffreyi Hopkins, D. brevicomis LeConte and D. Roberds, J.H.; Hain, F.P.; Nunnally, L.B. 1987. Genetic structure of southern Frontalis Zimmerman (Coleoptera: Scolytidae). Annals of Entomologi pine beetle populations. Forest Science 33: 52-69. cal Society of America 83: 892-901. Rubin, G.M.; Spradling, A.C. 1982. Genetic transformation of Drosophila Pajares, J.A.; Lanier, G.N. 1990. Biosystematics of the turpentine beetles D. with transposable element vectors. Science 218: 348-353. terebrans and D. valens (Coleoptera: Scolytidae). Annals of Entomologi Sakaguchi, B.; Oishi, K.; Kobayashi, S. 1965. Interference between "sex- cal Society of America 83: 171-188. ratio" agents of Drosophila willistoni and Drosophila nebulosa. Science Palmer, A.R.; Strobeck, C. 1986. Fluctuating asymmetry: measurement, 147: 160-162. analysis, patterns. Annual Review of Ecology and Systematics 17: 391- Samollow, P.B.; Soule, M.E. 1983. A case of stress-related heterozygote 421. superiority in nature. Evolution 37(3): 646-649. Parmeter, J.R., Jr.; Slaughter, G.W.; Chen, M.-M.; Wood, D.L. 1992. Rate and Schmid, J.M. 1972. Emergence, attack densities and seasonally trends of depth of sapwood occlusion following inoculation of pines with mountain pine beetle (Dendroctonus ponderosae) in the Black Hills. Res. bluestain fungi. Forestry Science 38: 34-44. Note RM-21. Fort Collins, CO: Rocky Mountain Forest and Range Parmeter, J.R., Jr.; Slaughter, G.W.; Chen, M.-M.; Wood, D.L.; Stubbs, H.A. Experiment Station, Forest Service, U.S. Department of Agriculture; 4 p. 1989. Single and mixed inoculations of ponderosa pine by fungal associ Schowalter, T.D.; Filip, G.M. 1992. Beetle-pathogen interactions in conifer ates of Dendroctonus sp. Phytopathology 79: 768-772. forests. New York: Academic Press. [In press]. Parsons, P.A. 1990a. Fluctuating asymmetry: an empigenetic measure of Seybold, Steven J. 1992. The status of scolytid genetics―a reductionist's stress. Biological Reviews 65: 131-145. view. [this Proceedings] Parsons, P.A. 1990b. Fluctuating asymmetry and stress intensity. Trends in Ecology & Evolution 5: 97-98. Seybold, S.J.; Milstead, J.E.; Wood, D.L. 1992a. Notes on the co-coloniza tion of Monterey pine, Pinus radiata by Ips sp. (Coleoptera: Scolytidae) Payne, T.L.; Dickens, J.C.; Richerson, J.V. 1984. Insect predator-prey coevo and new host associations for Ips mexicanus, I. paraconfusus, I. lution via enantiomeric specificity in a kairomone-pheromone system. plastographus maritimus and Dendroctonus valens. Unpublished draft Journal of Chemical Ecology 10: 487-491. supplied by author.

Piazza, A.; Menozzi, P.; Cavalli-Sforza, L. 1981. The making and testing of Seybold, S.J.; Ohtsuka, T.; Wood, D.L.; Kubo, J. 1992b. The enantiomeric geographic gene-frequency maps. Biometrics 37: 635-659. composition of ipsenol and ipsdienol from species in the subgeneric groups of Ips (Coleoptera: Scolytidae). Unpublished draft supplied by Piston, J.J.; Lanier, G.N. 1974. Pheromones of Ips pini (Coleoptera: Scolytidae). author. Response to interpopulational hybrids and relative attractiveness of males boring in two host species. Canadian Entomologist 106: 247-251. Seybold, S.J.; Teale, S.A.; Wood, D.L.; Zhang, A.; Webster, F.X.; Lindahl, K.Q., Jr.; Kubo, I. 1992c. The role of lanierone in the chemical ecology of Preisler, Haiganoush K. 1992. Spatial analysis in bark beetle research [this Ips pini (Say)(Coleoptera: Scolytidae) in California. Unpublished draft Proceedings]. supplied by author. Preisler, H.K. Modeling spatial patterns of trees killed in clusters by an Seybold, S.J.; Wood, D.L.; Ohtsuka, T.; Nomura, M.; Kubo, I.; Schultz, M.E. epidemic population of bark beetle. Applied Statistics [In Press]. 1992d. The response of the pine engraver, Ips pini (Say) (Coleoptera: Preisler, H.K.; Mitchell, R.G. 1992. Colonization patterns of the mountain Scolytidae) and its insect associates of the pure enantiomers of ipsdienol pine beetle in thinned and unthinned lodgepole pine stands. Unpublished in California. Unpublished draft supplied by author. draft supplied by author. Singh, R.S. 1989. Population genetics and evolution of species related to Raffa, K.F. 1988. Host orientation behavior of Dendroctonus ponderosae: Drosophila melanogaster. Annual Review of Genetics 23: 425-453. integration of token stimuli and host defenses. In: Mattson, W.J.; Levieux, Slatkin, M. 1987. Gene flow and the geographic structure of natural popula J.; Bernard-Dagan, C., eds. Mechanisms of woody plant resistance to tions. Science 236: 787-792. insects and pathogens. New York: Springer-Verlag; 369-390. Smith, R.H. 1963. Toxicity of pine resin vapors to three species of Dendroctonus Raffa, Kenneth F. 1992. Current status and critical research needs in scolytid bark beetles. Journal of Economic Entomology 56: 827-831. genetics [this Proceedings]. Smith, R.H. 1972. Xylem resin in the resistance of the Pinaceae to bark Raffa, K.F.; Berryman, A.A. 1983. The role of host plant resistance in the beetles. Gen. Tech. Rep. PSW-1. Berkeley, CA: Pacific Southwest Forest colonization behavior and ecology of bark beetles. Ecological Mono- and Range Experiment Station, Forest Service, U.S. Department of Agri graphs 53: 27-49. culture; 7 p. Raffa, K.F.; Berryman, A.A. 1987. Interacting selective pressures in conifer- Smith, R.H. 1975. Formula for describing effect of insect and host tree bark beetle systems: a basis for reciprocal adaptations. American Natural factors on resistance to western pine beetle attack. Journal of Economic ist 129: 234-262. Entomology 68: 841-844. Raffa, K.F.; Klepzig, K.D. 1989. Chiral escape of bark beetles from predators Smith, R.H.; Cramer, J.P.; Carpender, E.J. 1981. New record of introduced responding to a bark beetle pheromone. Oecologia 80: 556-569. hosts for the mountain pine beetle in California. Res. Note PSW-354. Riedy, M.F.; Hamilton, W.J.; Aquadro, C.F. 1992. Excess of non-parental Berkeley, CA: Pacific Southwest Forest and Range Experiment Station, bands in offspring from known primate pedigrees assayed using RAPD Forest Service, U.S. Department of Agriculture; 3 p. PCR. Nucleic Acid Research 20: 918. Stauffer Ch.; Huang, W.; Harshman, L.; Hammock, B.D. 1992b. Character Riley, M.A.; Goyer, R.A. 1986. Impact of beneficial insects on Ips sp. ization of the juvenile hormone metabolites of Ips typographus (Col., (Coleoptera: Scolytidae)bark beetles in felled loblolly and slash pines in Scolytidae) and Tenebrio molitor (Col., Tenebriodiae). Unpublished draft Louisiana. Environmental Entomology 15: 1220-1224. supplied by author.

USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. 29 Stauffer, Christian; Leitinger, Renate-, Fuehrer, Erwin. 1992a. Population Teale, Stephen A.; Hager, Barbara. 1992. Bark beetle genetics―an overview genetics of Ips typographus (Col., Scolytidae) and related Ips species [in Proceedings]. [this Proceedings]. Teale, S.A.; Lanier, G.N. 1991. Seasonal variability in response of Ips pini Stauffer, Ch.; Leitinger, R.; Simsek, Z.; Schreiber, J.; Fuehrer, E. 1992. (Coleoptera: Scolytidae) to ipsdienol in New York. Journal of Chemical Allozyme variation among nine Austrian Ips typographus populations. Ecology 17: 1145-1158. Journal of Applied Entomology 114: 17-25. Teale, S.A.; Webster, F.X.; Zhang, A.; Lanier, G.N. 1991. Lanierone: a new Stock, M.W.; Amman, G.D. 1980. Genetic differentiation among mountain pheromone component from Ips pini (Coleoptera: Scolytidae) in New pine beetle populations from lodgepole pine and ponderosa pine in north- York. Journal of Chemical Ecology 17: 1159-1176. east Utah. Annals of Entomological Society of America 73: 472-478. Thorne, B.L.; Haverty, M.I. 1989. Accurate identification of Zootermopsis Stock, M.W.; Amman, G.D. 1985. Host effects on the genetic structure of species (Isoptera: Termopsidae) based on a mandibular character of mountain pine beetle, Dendroctonus ponderosae, populations. In: Safranyik, nonsoldier castes. Annals of Entomological Society of America 82: 262- L., ed. The role of the host in the population dynamics of forest insects. 266. Victoria, B.C., Canada: Pacific Forest Research Centre; 83-95. Vanderwel, D. 1991. Pheromone biosynthesis by selected species of grain Stock, M.W.; Amman, G.D. Genetic diversity in early and late-emerging and bark beetles. Simon Fraser University; 230 p. Dissertation. mountain pine beetle. Unpublished draft supplied by author. Vanderwel, D.; Oehlschlager, A.C. 1987. Biosynthesis of pheromones and Stock, Molly W.; Amman, Gene D.; Bentz, Barbara J. 1992. Isozyme studies endocrine regulation of pheromone production in Coleoptera. In: Prestwich, of bark beetle population genetics and systematics [this Proceedings]. G.D.; Blomquist, G.J., eds. Pheromone biochemistry. Orlando, FL: Aca demic Press; 175-215. Stock, M.W.; Amman, G.D.; Higby, P.K. 1984. Genetic variation among mountain pine beetle (Dendroctonus ponderosae) (Coleoptera: Scolytidae) Wadleigh, R.W.; Yu, S.J. 1987. Glutathione transferase activity of fall army- populations from seven western states. Annals of Entomological Society worm larvae toward alpha, beta-unsaturated carbonyl allelochemicals of America 77: 760-764. and its induction by allelochemicals. Insect Biochemistry 17: 759-764. Stock, M.W.; Gregoire, J.C.; Furniss, M.M. 1987. Electrophoretic compari Wanntorp, H.-E.; Brooks, D.R.; Nilsson, T.; Nylin, S.; Ronquist, F.; Steams, son of European Dendroctonus micans and ten North American S.C.; Wedell, N. 1990. Phylogenetic approaches in ecology. Oikos 57: Dendroctonus species. Pan-Pacific Entomologist 63(4): 353-357. 119-132.

Stock, M.W.; Guenther, J.D. 1979. Isozyme variation among mountain pine Whitney, H.S. 1982. Relationships between bark beetles and symbiotic or beetle (Dendroctonus ponderosae) populations in the Pacific Northwest. ganisms. In: Mitton, J.B.; Sturgeon, K.B., eds. Bark beetles in North Environmental Entomology 8: 889-893. American conifers: a system for the study of evolutionary biology. Aus tin: University of Texas Press; 183-211. Stock, M.W.; Guenther, J.D.; Pitman, G.B. 1978. Implications of genetic differences between mountain pine beetle populations to integrated pest Wiley, E.O. 1981. Phylogenetics. New York: John Wiley and Sons; 439 p. management. In: Berryman, A.A.; Amman, G.D.; Stark, R.W.; Kibbee, D.L., eds. Theory and practice of mountain pine beetle management in Williams, J.K.G.; Kubelik, A.R.; Livak, K.J.; Rafalski, J.A.; Tingey, S.V. 1990. DNA polymorphisms amplified by arbitrary primers are useful as lodgepole pine forests―a symposium; April.25-27, 1978. Moscow, ID: genetic markers. Nucleic Acid Research 18: 6531-6535. University of Idaho, College of Forest Resources; 197-204. Stock, M.W.; Robertson, J.L. 1977. Genetic polymorphism in the Douglas-fir Wood, D.L. 1982. The role of pheromones, kairomones, and allomones in the host selection and colonications behavior of bark beetles. Annual Review tussock moth. USDA Forest Service Expanded Douglas-Fir Tussock of Entomology 27: 411-446. Moth Program, Final Report, December 31, 1977. Unpublished draft supplied by author. Wood, David L.; Storer, Andrew J.; Gordon, Thomas R.; Seybold, Steven J.; Page, Marion. 1992. Biosystematics of Ips mexicanus and Ips plastographus Stock, M.W.; Pitman, G.B.; Guenther, J.D. 1979. Genetic differences be- (Coleoptera: Scolytidae) and their fungal symbionts [this Proceedings]. tween Douglas-fir beetles (Dendroctonus pseudotsugae) from Idaho and coastal Oregon. Annals of Entomological Society of America 72: 394- Wood, S.L. 1963. A revision of the bark beetle genus Dendroctonus Erichson 397. (Coleoptera: Scolytidae). Great Basin Naturalist 23: 1-117.

Stock, M.W.; Schmitz; R.F.; Amman, G.D. Heterozygosity and density Wood, S.L. 1982. The bark and ambrosia beetles of North and Central change in mountain pine beetle (Dendroctonus ponderosae) (Coleoptera: America (Coleoptera: Scolytidae), a taxonomic monograph. Great Basin Scolytidae) populations. Unpublished draft supplied by author. Naturalist Memoirs No. 6. 1359 p.

Sturgeon, K.B. 1980. Evolutionary interactions between the mountain pine Wood, T.K. 1980. Divergence in the Enchenop binotata Say complex beetle, Dendroctonus ponderosae Hopkins, and its host trees in the (Homoptera: Membracidae) effected by host plant adaptation. Evolution Colorado Rocky Mountains. Boulder: University of Colorado, Disserta 34: 147-160. tion. Yearian, W.C.; Gouger, R.J.; Wilkinson, R.C. 1972. Effects of the bluestain Sturgeon, K.B.; Mitton, J.B. 1986. Allozyme and morphological differentia fungus, Ceratocystis ips, on development of Ips bark beetles in pine bolts. tion of mountain pine beetles Dendroctonus ponderosae Hopkins (Co Annals of Entomological Society of America 65: 481-487. leoptera: Scolytidae) associated with host tree. Evolution 40: 290-302. Zuber, M.; Stauffer, Ch.; Leitinger, R.; Nelson, L.; Page, M.; Bright, D. 1992. Sturgeon, K.B.; Robertson, J.L. 1985. Microsomal polysubstrate Biochemical, morphological and behavioural analysis of the relationship monooxygenase activity in western and mountain pine beetles. Annals of between Ips amitinus and Ips amitinus var. montana (Col., Scolytidae). Entomological Society of America 78: 1-4. Unpublished draft supplied by author. Teale, S.A. 1990. Ecology and evolution of pheromone communication in Ips pini (Coleoptera: Scolytidae). New York: State University of New York; 109 p. Dissertation.

30 USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. Workshop Participants Alosi, M. Carol, Pacific Southwest Research Station, USDA Forest Service, Mitchell, Russ, 20454 Steamboat, Bend, OR 97702, (503) 388-2869. (re- 1027 Trenton Ave., Bend, OR 97701; (503) 388-5422, FAX: (503) 383- tired) 5733. Page, Marion, Pacific Southwest Research Station, USDA Forest Service, Amman, Gene D., Intermountain Research Station, USDA Forest Service, 800 Buchanan Street, Albany, CA 94710; (510) 559-6300, FAX: (510) 507 25th Street, Ogden, UT 84401; (801) 625-5393, FAX: (801) 625- 559-6440. 5723. (retired) Preisler, Haiganoush K., Pacific Southwest Research Station, USDA Forest Anderson, Wyatt W., Department of Genetics, 101B Life Sciences Bldg., Service, 800 Buchanan Street, Albany, CA 94710; (510) 559-6300, FAX: University of Georgia, Athens, GA 30602; (706) 542-3326, FAX: (706) (510) 559-6440. 542-3910. Raffa, Kenneth F., Dept. of Entomology, University of Wisconsin, Madison, Bentz, Barbara J., Intermountain Research Station, USDA Forest Service, WI 53706; (608) 262-1125, FAX: (608) 262-3322. Forestry Sciences Lab, 860 No. 1200 East, Logan, UT 84321; (801) 752- 1311, FAX: (801) 753-8625. Robertson, Jacqueline L., Pacific Southwest Research Station, USDA Forest Service, 800 Buchanan Street, Albany, CA 94710; (510) 559-6300, FAX: Bright, Donald E., Canada Dept. of Agriculture, Systematics Research, Cen (510) 559-6440. tre for Land and Biological Resources Research, Ottawa, Ontario, Canada K1A OC6; (613) 996-1665, FAX: (613) 995-1823 or 7283. Seybold, Steven J., Pacific Southwest Research Station USDA Forest Ser vice, 800 Buchanan Street, Albany, CA 94710; (510) 559-6477, FAX: Cane, James H., Dept. of Entomology, 301 Funchess Hall, Auburn Univ., (510) 559-6440. Auburn, AL 36849-5413; (205) 844-5006, FAX: (205) 844-3005. Stanton, Maureen, Department of Botany, Univ. of California, Davis, CA Gordon, Thomas R., Dept. of Entomological Sciences, University of Califor 95616; (916) 752-2405, FAX: (916) 752-5410. nia, Berkeley, CA 94720; (510) 642-3874, FAX: (510) 642-3845. Stauffer, Christian, Institute of Forest Entomology, Forest Pathology and Greene, Lula E., Pacific Southwest Research Station, USDA Forest Service, Forest Protection, Universität für Bodenkultur, A-1190 Vienna, Austria. 800 Buchanan Street, Albany, CA 94710; (510) 559-6300, FAX: (510) Present address: Dept. of Entomology, Univ. of California, Davis, CA 559-6440. 95616; (916) 752-0492.

Haack, Robert, North Central Research Station, 1407 S. Harrison Road, East Stock, Molly W., Dept. of Forest Resources, College of Forestry, University Lansing, MI 48823; (517) 355-7740, FAX: (517) 335-5121. of Idaho, Moscow, ID 83843; (208) 885-7033, FAX: (208) 885-6226.

Hager, Barbara, Dept. of Environmental and Forest Biology, State Univ. of Storer, Andrew J., Dept. of Entomological Sciences, University of Califor NY, College of Environmental Science and Forestry, Syracuse, NY nia, 201 Wellman Hall, Berkeley, CA 94720; (510) 643-5806, FAX: 13210; (315) 470-6758, FAX: (315) 470-6934. (510) 642-7428.

Hayes, Jane L., Southern Forest Experiment Station, USDA Forest Service, Teale, Stephen, Dept. of Environmental and Forest Biology, State Univ. of 2500 Shreveport Highway, Pineville, LA 71360; (318) 473-7232, FAX: NY, College of Environmental Science and Forestry, Syracuse, NY (318) 473-7222. 13210; (315) 470-6758, FAX: (315) 470-6934.

Hobson, Ken, Intermountain Research Station, USDA Forest Service, For Wood, David L., Dept. of Entomological Sciences, University of California, estry Sciences Lab, 860 No. 1200 East, Logan, UT 84321; (801) 752- 201 Wellman Hall, Berkeley, CA 94720; (510) 642-5538, FAX: (510) 1311, FAX: (801) 753-8625. 642-7428.

USDA Forest Service Gen. Tech. Rep. PSW-138. 1992. 31 The Forest Service, U.S. Department of Agriculture, is responsible for Federal leadership in forestry. It carries out this role through four main activities: • Protection and management of resources on 191 million acres of National Forest System lands • Cooperation with State and local governments, forest industries, and private landowners to help protect and manage non-Federal forest and associated range and watershed lands • Participation with other agencies in human resource and community assistance programs to improve living conditions in rural areas • Research on all aspects of forestry, rangeland management, and forest resources utilization.

The Pacific Southwest Research Station • Represents the research branch of the Forest Service in California, Hawaii, American Samoa and the western Pacific.

Persons of any race, color, national origin, sex, age, religion, or with any handicapping conditions are welcome to use and enjoy all facilities, programs, and services of the U.S. Department of Agriculture. Discrimination in any form is strictly against agency policy, and should be reported to the Secretary of Agriculture, Washington, DC 20250. United States Department of Proceedings of a Workshop on Bark Beetle Genetics: Current Status of Research Agriculture

Forest Service May 17-18,1992, Berkeley, California

Pacific Southwest Research Station

General Technical Report PSW-GTR-138