ALTERNATIVES IN FOREST INSECT CONTROL

Proceedings of a Symposium sponsoredby Algoma College, Sault Ste. Marie,

Ontario and published by:

GREAT LAKES FOREST RESEARCH CENTRE

AND

INSECT PATHOLOGY RESEARCH INSTITUTE

SAULT STE. MARIE, ONTARIO

SYMPOSIUM PROCEEDINGS O-P-2

CANADIAN FORESTRY SERVICE

DEPARTMENT OF THE ENVIRONMENT

JUNE 1973

Copies of this report may be obtainedfrom

Director,Great Lakes Forest Research Centre,or

Director,Insect Pathology Research Institute,Canadian Forestry Service,Department of the Environment,Box 490, Sault Ste. Marie, Ontario.

FOREWORD

Recognizing the widespread concern forpreservation of environmental quality, AlgomaCollege has instituted a series of symposia. Thepurpose of these is to bring to the attention ofthe public and of students informative and authoritative considerations of ecological matters.

Because Algoma College is situated inSault Ste. Marie, Ontario, which is also theheadquarters of the Great Lakes Forest ResearchCentre and the Insect Pathology Research Instituteof the Canadian Forestry Service, it was onlylogical that the expertise in these centres ofexcellence be called upon to contribute to thefirst symposium in this series -

"Alternatives in Forest Insect Control"

This Symposium was held in the Armoury Theatre inSault Ste. Marie, Ontario on November 18, 1972.

The Canadian Forestry Service is happy tomake available the papers presented at thisSymposium by publishing them as a joint reportof the Great Lakes Forest Research Centre and theInsect Pathology Research Institute, Sault Ste, Marie,Ontario.

TABLE OF CONTENTS

Page

INTRODUCTORY PAPER: AN ECOLOGICAL VIEW

Dr. Susan W. Sanders, Algonia College 1

THE USE OF INSECTICIDES IN THE CONTROL OF FOREST INSECT PESTS

Dr., G. M. Howse,Great Lakes Forest Research Centre . . . 5

DEVELOPING CONTROL OF INSECT VECTORS OF DUTCH ELM DISEASE

Dr, L. M»' Gardiner,Great Lakes Forest Research Centre . . . 10

INSECT SEX ATTRACTANTS: THEIR ROLE IN INSECT CONTROL

Dr. C. J. Sanders3Great Lakes Forest Research Centre

and

Dr» I. Weatherston5Insect Pathology Research Institute . . 19

CONTROL OF INSECT PESTS USING SYNTHETIC INSECT HORMONES

Dr. Ae Retnakaran,

Insect Pathology Research Institute 27

THE USE OF BACTERIAL INSECTICIDES IN FOREST INSECT PEST CONTROL

Dr. P. G. Fast,

Insect Pathology Research Institute ... 42

THE USE OF VIRUSES IN THE CONTROL OF FOREST INSECT PESTS

Dr. J. C. Cunningham,Insect Pathology Research Institute 46

ALTERNATIVES IN THE CONTROL OF FOREST INSECTS: FUNGI

Dr. D. Tyrrell and Dr. D. MacLeod,Insect Pathology Research Institute . . 53

INTRODUCTORY PAPER: AN ECOLOGICAL VIEW

Dr. Susan W. Sanders,

Algoma College,Sault Ste. Marie, Ontario

The word ecology has become a password of our times and to beconcerned about ecology is quite the "in" thing. When Algoma Collegefirst began considering material for this Symposium, we were concernedthat what we would offer you would be some light on what is actuallybeing done to contribute to the solution to pollution in the world andits wide-reaching effects on everyone.

Ecology, briefly, is the study of the relationships betweenliving organisms and their environment. Therefore, an ecologist is aperson who studies these relationships. In order to understand relationships between any two kinds of organisms, one must know a great dealabout each one — and if another organism is added to the group, thecomplexity increases enormously. Therefore, ecologists use in theirstudies the expertise of people who have studied the separate species.One of the things we hope to give you today is a bit of the knowledgeof the experts on the subject of controlling numbers of insects in theforest so that you may have an idea of what these people, ecologistsand foresters, are doing or proposing to do to your forests.

I have said that ecology is the study of relationships betweenlife and its surroundings. Ecosystem (short for "ecological system")is the term coined by ecologists for this inter-relationship of organisms and their nonliving, or abiotic, environment to produce anindependent, relatively stable system. An ecosystem, then, includesall the organisms (important or seemingly unimportant) which occur inan area. The area may be as large as the world, or as small as a woodplot or a farm pond.

All organisms, to be alive at all, depend on energy for theirlife processes, e.g., for breathing, eating, growing, moving, andreproducing. This energy comes ultimately from the sun and nowhereelse. Any synthesis of simple compounds (such as CO2, H20, or N2) intomore complex compounds (such as proteins or carbohydrates) takes energy.Animals cannot convert the sun's energy into needed chemical and mechanical energy and therefore are dependent on plants, which can do this.

In a forest, the trees are the primary convertors of energy;therefore, we tend to think of forests as being trees and nothing else.Even our idea of a forest, though, differs with the area in which welive. Someone from Geraldton would think of one or two species of tree

as constituting a forest (e.g., black spruce) while someone from southernOntario would think of a forest in terms of three or four species suchas beech, maple, and basswood. Someone from the tropics would have adifficult time naming the forest by any tree species, as there might beas many as 20-100 species in a square mile.

But the forest also includes all the other organisms, plant andanimal, in the region. And they are factors in the character of theforest.

The trees and the other plants in the forest convert sun energyto chemical energy and store it as sugars or starches. These plants arecalled producers. They use the nutrients in the area and the sun'senergy to build more complex compounds which have energy in their bonds.But an ecosystem has more than just producers; otherwise, the nutrientswould all be bound up in living material until they were used up andlife processes in that area would be no more.

In order to have a stable system, decomposers must break down theplant bodies, using the material for their own good, and in the processreleasing the nutrients back into the soil for use again by the producers.And the cycle goes on.

The previous example would be a very slow cycle, and probablyexists rarely in nature. Pioneer communities on bare rock might havethis sort of association.

In forest ecosystems, however, organisms called consumersmake use of the bound nutrients before the plants die. These consumers,usually animals, obtain their energy from plants or other consumers.The animals, such as deer, spruce budworm or woodpecker, all interrelatewith the plants in a food web.

Each producer and consumer eventually dies or is killed, itsbound nutrients are released by decomposers, and the cycle continues:producer to consumer to consumer, and each one to decomposer. Even thewaste products of living animals are decomposed and returned to thenutrient cycle.

If an ecosystem existed with only producers and decomposers, along time would elapse in an area before nutrients, once used, wouldagain be available. Add just one consumer and the recycling increasesa great deal. Add 100 consumers and nutrients are being exchanged at agreat rate! This makes a very stable ecosystem, because the loss of onemember is easily compensated for by others.

It is not even necessary to envision an entire forest to seethis mass of activity. Balch (1965) describes a tree as follows:

"...you find many hundreds of species, feeding, laying eggs, orsheltering on every part of a tree. On the leaves and shoots are allkinds of caterpillars, beetles> or sawflies chewing at their favouritetissues, some mining inside them. Aphids, leaf-hoppers, and scaleinsects have their beaks inserted wherever they can suck out the juicesthey need. Other beetles and caterpillars are boring in the bark, underthe bark, or in the wood. Dead trees are full of them. In the forestfloor insects are moulting, hibernating, or feeding on roots and organicmattero

"Among them move a still larger number of predaceous or parasitic species: flies, wasps, beetles, searching for their particularvictims, on which they must feed or lay their eggs. And, to completethe picturej there are the birds and small mammals that eat insects:chickadees and warblers searching the. branches; woodpeckers digginginto the stems; sparrows, squirrels, and skunks scratching in theduff. In the ground beneath the trees mice and shrews are foragingfor cocoons or pupae in a network of tunnels."

It is easy to see from the previous example that insects playa very important role as consumers in the forest (community ecosystem).They are small and have a short life span, a characteristic whichcontributes to rapid nutrient turnover and, therefore, stability. Anyone species is usually present in low numbers, its presence being regulated by weather, predators, parasites, disease and, in most forests,the wide spacing of food trees (to an insect's eye, anyway) whichwould cause a scarcity of food in an area.

In some areas, such as forests in which one or two species oftrees are frequent and food, therefore, is readily obtainable, extremelyfavorable weather or elimination of predators for several years in arow may cause an increase in numbers of a particular insect. An increase in numbers of adults means more eggs, more young, and even moreadults; the numbers increase exponentially and we have an outbreak.

After awhile these large numbers of insects eat all the food.Predators on the insects increase in numbers because their food sourceis readily available. These two factors then act on the insect population to reduce its numbers again to normal levels. In the case offorest trees, the numbers may remain low for a long while because treestake a long while to come back in normal succession.

This cycling goes on regularly and is a natural part of theecosystem. It is only as man has become a competitor with the insectfor the tree's energy and nutrients that we feel we cannot "allow"this "consumption of the forest by insects". We want to "consume it"in our own way, using its bound energy for firewood, houses, paper

products, or furniture. And we call this small competitor of ours apest!

Ecologists do not want to wipe out our insect competitor completely, as this would increase the instability of the forest ecosystem,and might have extremely far-reaching effects. They do want to keep thenumbers low, however, so that the usual controls on the insect population(e.g., predators, disease) can operate. In this way man obtains notperfect, but at least "normal", trees.

Ecologists can attempt to regulate numbers of insects in twobasic ways. First, they can use the insect's environment to keep itsnumbers low by managing its food supply (the forest) so as to avoid(a) too many food trees in one area, (b) too many trees of a similarage which would then be prey to the same insect, or (c) overmature and,therefore, less healthy trees. Control of physical surroundings suchas temperature, rainfall, wind, and other abiotic natural regulators isnot yet possible. Second, they can manage the insect populationdirectly or indirectly, using biological controls such as the insect'snatural regulators (predators or disease regularly occurring in itsenvironment) or introduced regulators (found by experiment). If theseare not feasible, chemical control of insect populations may be used.

And so the purpose of today's symposium is to expose you to someof the alternatives in the control of insect populations in the forestecosystem.

Reference

Balch, R.E. 1965. The ecological viewpoint. Can. Broadcast. Corp.,Toronto.

THE USE OF INSECTICIDES IN THE CONTROL OF FOREST INSECT PESTS

Dr. G. M. Howse,Great Lakes Forest Research Centre,Sault Ste. Marie, Ontario

A (general description is given of forest insectpests in Ontario and natural and applied insect control arediscussed. The use of insecticides is featured: the types,nature and action of chemical and biological insecticides,spraying by aircraft, problems and benefits of such spraying, types of aircraft and spraying equipment are discussed.Brief mention is made of spraying operations in Ontario inrecent years. Also outlined are decision-making processesand the roles of various federal and provincial agencies inthe above operations.

We are all aware these days that the subject of pesticides is acontroversial topic. The purpose of this paper is not to cause oravoid controversy, but to explain as clearly and simply as possible therationale for the use of insecticides in regulating the numbers offorest insect pests in Ontario. Entomology is the study of insects,and people who study insects are known as entomologists. Thus, forestentomology involves the study of insects that inhabit the forest.

One of the primary goals of entomologists, regardless of theirfield of specialization, be it forest insects, agricultural insects orinsects that affect man and animals, is to learn how to control insects,Insect control, defined in a broad sense, includes any natural orapplied process or method that regulates the numbers of insects inorder to prevent damage, to prevent an increase in numbers, to cause areduction in numbers or to prevent the spread of an insect species.Therefore, as a consequence, insect control is generally accomplishedby killing or destroying a sufficient number of insects, eitherdirectly or indirectly, to achieve desired goals.

As has been stated, control of insects may be broadly subdivided into (a) natural control and (b) applied control. Naturalcontrol is the sum of all the naturally occurring mortality factorsthat affect insect populations such as predators, parasites, disease,competition, starvation and climate (or weather). These natural mortality factors are operating continuously since they are part of theenvironment of an insect. However, there is very little that man can

do directly to manipulate these natural factors in his favour againstthe insect. Thus, some form of applied control becomes, in most cases,the only method available to man in his attempts to control insectswhen natural control factors fail to keep an insect population in check.

The term "forest insect pest" must first be defined. There aremany thousands of species of insects that live in our forests and byfar the majority of them are involved in the web of life in a veryunspectacular but vital fashion; unspectacular from the point of viewthat they never become numerous enough to cause outbreaks, or that theyare not obnoxious to man or animals; vital because they are an integralpart of the forest ecosystem. On the other hand, there are some insectspecies that are considered beneficial because they consume or killother insects. In contrast, however, there is a small number of forestinsect species that at some point in time we consider to be pests.

The effects that insects, termed pests, can have on the forestenvironment can be expressed in many ways. Several of the more obviousones are as follows: 1) there can be a direct loss owing to mortalityof harvestable timber or there may be mortality of replacement stockconsisting of seeds, seedlings, saplings or immature trees, 2) defoliating or leader-killing insects can cause increment reduction, or deformities, or retard tree growth, 3) bark-penetrating insects can damageor kill trees either directly or by introducing pathogens which mayeventually kill the tree, 4) if widespread tree mortality or defoliationof trees and stands occurs, an increased fire hazard may result (manyof the huge, destructive fires of past years in North America areconsidered to have been fuelled by insect-killed or damaged forests),5) tree mortality or damage can reduce or eliminate the aestheticvalues of the forest (This could affect a family out for a Sundaydrive, tourists, fishermen, hunters, wilderness enthusiasts, canoeists—in other words, virtually anyone who uses the forest for recreationalpurposes.).

An insect pest is generally defined within the context of economics; therefore, a forest insect becomes a pest when it causesdamage or is capable of causing damage to the forest or forest environment that is unacceptable to man. The key phrase is "unacceptable toman . Insect damage or potential for damage must be considered inrelation to forest management, forest protection or forest recreation.In other words, insect damage or potential for damage results in thatinsect becoming a pest in a situation in which man is competing withthe insect for the forest resource. Thus a definition of a forestinsect pest depends upon the degree of damage that is unacceptable incombination with plans for the present or future use of the forest.

A unit, known as the Forest Insect and Disease Survey, hasoperated as part of the federal Canadian Forestry Service (or its

various predecessors) across Canada for the past 35 or 40 years. Itspurpose has been to collect and identify forest insects and diseasesand to determine their relative abundance from year to year. Thus,specific forest insect pests are recognized primarily on the basis oftheir past history and performance. Outbreaks of insects, particularlythose that have been problems in the past and are capable of causingdamage to a forest resource required by man, are considered to be pestsand may need applied control. Any list of outstanding forest insectpests in Ontario must be headed by the spruce budworm. Other majorexamples are the white pine weevil, jack-pine budworm, larch sawfly,forest tent caterpillar, European pine sawfly and the native andEuropean elm bark beetles which are the vectors of Dutch elm disease.This is not a complete list but represents those forest insect peststhat are considered to be of greatest significance in Ontario.

As previously mentioned, when natural control factors fail tokeep an insect population in check, the only practical alternative issome form of applied control measure. These depend upon man for theirapplication and can be influenced by him to a great degree. The variousapproaches to applied control can be classified as follows:

1. Chemical control - insecticides, repellents, attractants,hormones

2. Biological control - involves the introduction and establishmentof biological enemies such as parasites, predators ororganisms that are pathogenic to insects (e.g., viruses,bacteria or fungi). These latter organisms can be usedas microbial insecticides; thus, an insecticide doesnot necessarily have to be a chemical poison.

3. Cultural control - There are various forestry practices suchas salvage, harvesting techniques or silviculturalmethods that can be useful.

40 Physical and mechanical control - low and high temperature,radiation, light traps, electric traps, etc. Overall,these approaches are not too practical for control offorest insects although radiation has proved successfulin some agricultural situations (e.g., screw worm in thesoutheastern United States).

5. Legal control - quarantines, embargoes, trade and commerceregulations designed to prevent the introduction orspread of native or foreign pests.

There have been control programs aimed at three forest insectpests in Ontario over the past 5 years. These insects are the spruce

budworm, jack-pine budworm and white pine weevil. The only practicalmethod of controlling or preventing damage by these insects has beenthrough the use of chemical insecticides applied from aircraft.Chemical insecticides can be grouped into classes depending upon theirmode of action: (a) stomach poisons (ingested by the insect),(b) contact poisons (absorbed through the integument or respiratorysystem) and (c) fumigants (respiratory system). Chemical insecticidescan also be classified by their chemical nature (e.g., inorganic ororganic compounds) or they can be classified by family groupings (e.g.,arsenicals, fluorines, pyrethrins, chlorinated hydrocarbons or organicphosphates).

Insecticides can be applied in a variety of ways. The propermethod of application depends on the properties of the insecticide,biology of the insect pest and the site to which the insecticide is tobe applied. Insecticides are generally applied as sprays but canalso be treated as dusts or fumigants. Equipment for application ofinsecticides includes hand dusters, squirt guns, aerosol bombs, mistblowers, hydraulic sprayers and aircraft. Aircraft are used for virtually all major forest spraying operations. A large variety ofaircraft, some of which are specially designed or converted, are usedfor forest spraying (Fig. 1). Two basic spraying systems are being

Figure 1. Stearman aircraft fitted with micronair units foragainst spruce budworm.

spraying

used at present on aircraft: the boom and nozzle, and micronairs forultra-low-volume applications (ULV). There are problems and benefitsarising from aerial spraying. Probably the major problem involved inusing chemical poisons is that of potentially harmful side effects. Achemical should be used in such a way that it will do the best possiblejob on the target insect and have the least harmful environmental sideeffects. The greatest benefit of the aerial spraying approach is thatit is the most efficient, practical and economical method of protectinglarge areas of forest.

Up to the present time, one of the major guidelines in decidingwhether or not to spray has been to compare the cost of spraying versusthe benefits to be derived from spraying. The point is that there islittle sense in protecting a resource if it costs more to protect itthan it is worth. This is a rather easy exercise if one measures thecost of the operation against the value of wood to be saved. However,at least in Ontario, other factors are being taken into considerationsuch as attempting to evaluate the preservation of the scenic conditionof a provincial park or the costs of preventing or fighting fires thatmay result from insect problems.

In Ontario, protection of the province's forest resource is theresponsibility of the Ontario Ministry of Natural Resources (formerlythe Ontario Department of Lands and Forests). The role of the CanadianForestry Service (federal) is to provide entomological information aboutpotential or actual forest insect problems in Ontario through its ForestInsect and Disease Survey Unit. Representatives of the respectiveprovincial and federal agencies then exchange their views of the situation and normally a consensus is reached. If the decision is to conducta control operation, then the operation itself (i.e., procurement ofinsecticide, aircraft, landing strips, etc.) is the responsibility ofthe province. However, the Forest Insect and Disease Survey Unitcooperates with the province to help conduct these operations. Morespecifically, the Forest Insect and Disease Survey provides the entomological input required to determine the necessity of carryingout an operation, to time the spray applications properly and todetermine the effectiveness of the operation. This type of approachand spirit of cooperation has, I believe, resulted in a wise and realistic approach to the immediate solution of forest insect problems inOntario„

In summation chemical insecticides will be used to prevent orremedy forest insect problems in Ontario until practical alternativesprovided by research and testing are available. We hope that this willoccur in the near future.

10

DEVELOPING CONTROL OF INSECT VECTORS OF DUTCH ELM DISEASE

Dr. L. M. Gardiner,Great Lakes Forest Research Centre,

Sault Ste. Marie, Ontario

The complicated nature of Dutch elm disease, whichinvolves the causal fungus, its insect carriers, and thetree 's reaction to both, suggests that "good" control willrequire several well-integrated methods. This paper dealswith the control of only one insect vector. The developmentof methods for reducing insecticide application to a minimum,using cacodylic acid to isolate community elm populations,and the introduction of more efficient parasites are discussed.The importance of seasonal history studies in developingeffective control programs is shown.

Introduction

The so-called Dutch elm disease was first reported in Romaniain 1910, then in France in 1918 and finally in Holland in 1920, where itis called simply the elm disease. It was recorded in Britain in 1927and first reports of North American incidences came from Ohio in 1930.In the last 40 years the disease has spread throughout most of the rangeof elm in the United States and has made serious inroads in Canada,stemming both from original importation from Europe and pincer-like,northward penetration through Quebec, eastern Ontario, Niagara Falls andWindsor. There is also a good possibility that it has crossed the borderat Sault Ste. Marie. It has now reached as far north as a line approximating the Trans-Canada Highway south and east of a point 40 miles northof Sault Ste. Marie.

The disease is caused by a fungus, Ceratocystis ulmi9 which inpart causes a reaction by the tree that results in a breakdown in transport of water and nutrients. The first symptom is wilting and this isfollowed by yellowing of the foliage on infected branches. Depending onthe initial severity of infection, and the treefs ability to counteractit, death usually follows in 1 or 2 years.

Fairly stringent quarantine regulations have failed to stop thespread of the disease. This is because the spores of the fungus areborne by insects from diseased to healthy trees. The two main culpritsare the smaller European elm bark beetle, Scolytus multistriatus, animported species, and the native elm bark beetle, Hylurgopinus rufipes.

11

Both species are known as secondary insects, i.e., they breedonly in dead or dying trees and are not known to kill trees by theirown activity. In starting a brood, the female excavates a gallery inthe bark and lays eggs along the sides. On hatching, the larvae fan outin the inner bark, making long feeding galleries, at the ends of trtiichthey pupate. When the resulting adults emerge, they feed on the innerbark of living elms before seeking out dead and dying trees for furtherbreeding. In the absence of disease, this adult feeding would constitute merely a nuisance to the tree; however, when the beetles emergefrom a tree killed by Dutch elm disease, they can carry the fungalspores to the living trees on which they feed. If, during feeding, theypenetrate the x*ood or xylem tissue, then the tree may be inoculated byspores from the insect. In this way, the beetles are transmitters orvectors of Dutch elm disease.

It is evident from the foregoing that Dutch elm disease is rather complicated, involving a fungus, at least two insect vectors, and thetree's reactions to both fungus and insects. The general health of thetree and its degree of resistance to the disease are also important. Itis also evident that satisfactory control of Dutch elm disease willnecessitate development of several well-incegrated methods of attackingthe various facets of the problem.

Two years ago, we began a research program at the Great LakesForest Research Centre to develop effective, economically and ecologically acceptable methods of controlling the disease. Work is in progresson methods of treating the disease directly (in other words, "curing" asick tree) by using fungicides, studying the physiology and chemistry ofinfection, and controlling the insect vectors. In regard to the latter,we have been concentrating on the native elm bark beetle which is theprincipal carrier of the disease in central Ontario.

This paper is concerned only with our attempts to develop control methods for this insect.

Seasonal History of the Native Elm Bark Beetle

One of the secrets of effective control of a pest is knowing thevulnerable periods in its life cycle and/or seasonal development. Atthe outset, we realized that we did not know enough about the seasonaldevelopment and movements of this species, particularly in centralOntario at the edge of disease distribution. The seasonal history hadbeen studied previously in the United States, Quebec, and southernOntario. These studies had shown that the beetle passes through onecomplete generation and part of a second generation per year. In thesouth most of the population apparently overwinters as adults m hibernation tunnels in the bark of living elms. During the tunnelling, thebeetles feed on the inner bark, both in the autumn and in early spring.

12

In May, the adults emerge and fly to suitable trees to start a new brood.Adults from the new generation begin emerging in late July and emergencecontinues until September. Some of the first beetles to emerge maybegin a second brood which overwinters as larvae, although most adultsproceed directly to excavate overwintering tunnels in living trees. Theoverwintered larvae, which can also include retarded larvae from thespring generation and some offspring from eggs laid in the summer, giverise to adults in June. These adults start a new brood in July whichproduces overwintering adults by early September, although late membersof the brood overwinter as larvae.

Most earlier workers have also reported that the adult beetlesfeed for a short period on the inner bark of smaller limbs and branchesof living trees before oviposition in dead trees. It is this period offeeding, particularly in the spring after growth commences, that issuspected as the time when inoculation of living trees by beetles occurs.

In the Sault Ste. Marie area, we have been following the seasonaldevelopment and movements of a beetle population for the past 2 years.The principal method used in this study is daily trapping of adultbeetles throughout the annual period of activity.

The trap used consists of a 6-inch-wide band of polyethylenesheeting wrapped tightly around the tree trunk or branch and coatedevenly with a thin layer of Tree-Stickem, a very sticky, stable substanceused by orchardists. This material appears to have no repellent effecton the beetles, which walk or fly into it quite readily. Traps wereinstalled at breast height on selected trees in a stand with manydiseased trees and a high population of beetles. Recording dailycatches afforded ameasure of seasonal beetle activity, as well as movements up or down the trunks and the relative attractiveness of differenttrees. Progress of seasonal development was determined by periodicremoval of. bark sections from selected trees.

In this area, and presumably throughout central and northernOntario populations of native elm bark beetle exhibit certain fairlyclear differences in behaviour from those in the south. It has beenreported from southern Ontario that the bulk of the population overwinters as adults. Our studies show that, locally, at least as manybeetles overwinter as larvae or pupae in the bark of brood trees as inthe adult stage. Also, local populations appear to be divided behaviourally into two groups fairly distinct from each other. Figure 1shows this dichotomy. One group overwinters as adults in shallow gal-inH16% V bark °f liVing °r dying elms- These adult* emerge in Mayand if the temperatures are seasonable, they crawl upwards on the treeand feed m the bark of the upper branches before flying to dying ordead trees to breed. If air temperatures are unseasonably warm, as in

13

May, 1972, the beetles fly immediately either to the upper branchesto feed or to breeding trees. The offspring of this group becomeadults in August and September. The earliest emergents may startanother brood, depending on weather conditions, which overwinters asyoung larvae. Most of this group, however, emerge too late for thisand overwinter as adults like their parents.

The other group overwinters as larvae in brood trees. In thespring, they complete development and emerge as adults from late Juneto late July (Fig. 1). Most of their offspring also pass the following

Aug. 28

1972. Daily Catches on 15 Trees

Dying or Dead i n 1971 .

Traps Installed 8 May

Figure 1. Daily catches on trees suitable for breeding. Solid areadenotes overwintering adult group; stippled area denotesoverwintering larval group.

14

winter as larvae or pupae, although a small number become adults andif the weather is warm may emerge as late as November and join the overwintering adult group. Probably all beetles caught in the traps in lateOctober and early November belong to this category; almost all werecaught at the top of the traps, on their way down the trees to hibernate.

No evidence has been found to suggest that adults from the overwintering larval group do any feeding in the upper branches as do theoverwintering adult group. Sticky traps set on limbs and higher branchesof elms at Ottawa in 1972 produced significant catches in late May butfew or none during the rest of the season.

Thus, their behaviour points to the importance of the overwintering adult group in regard to Dutch elm disease. Because of theirhabit of feeding on the inner bark of living trees, both in their overwintering niches and in the upper limbs and branches, they become theprime suspects in most of the transmission of Dutch elm disease, atleast in the northern part of its range, if not farther south as well.Also, their importance is amplified by the fact that the feeding andconsequent penetration of xylem tissue are done in the spring when newvessel growth renders the tree most susceptible to the fungus.

Implications for Control

(i) Chemical control. Thus far the principal control measure usedagainst both the European and the native elm bark beetle has been theapplication of high doses of insecticides to the entire tree ChemiLl.are applied by hydraulic sprayer, mist blower or helicopter a„?^ fare all of these methods wasteful and costly' but they £end'to conrl° *inate the surroundings. Originally, DDT was'used very liberally anf"certainly contributed to asignificant decrease in the incidence ofdisease in communities willing to pay the cost. Lately, DOThas beenreplaced by methoxychlor which, though reputedly safe/to the envJroLment, is much more expensive. environ-

*™ ^ disc?vei7 of the Prime importance of the overwintering adultgroup of the native beetle in disease transmission has interestingimplications m regard to chemical control from the standpoint of costand environmental pollution. It might boil down to asuccessful case ofdivide and conquer", because for along period each year the bulk ofthis whole group is concentrated in an accessible, relativelv vulnerL.position in the bark of the bottom 6-8 feet of livinftrels Sgests that minimal applications of insecticides tithe Wr trunksTelms in August and early May should be highly effective alainst thi*group as they are entering or leaving hibernation! *

A test of this method was tried in 1972. In 1971 adult*emerged in mid-May, climbed towards the upper part of the tree and were

15

caught readily along the bottom of sticky traps. We thought to usethis technique for evaluating chemical' treatment of the lower trunk.On May 10, 1972 several trees were sprayed with a 0.5% mixture ofmethoxychlor, and sticky traps were applied above the sprayed area.Other unsprayed trees were fitted with traps as controls. However,the test could not be evaluated because unseasonably hot weathercaused the emerging beetles to fly instead of crawl and none of thetraps caught any significant number along their bottom edges. Furthertests of the method are required as well as a search for effective butrelatively "safe" insecticides.

(ii) Sanitation. Sanitation cutting and removal of infected trees,though costly, has been a common method of controlling Dutch elmdisease, and there is no doubt that communities practising this methodthoroughly have cut their losses. The theory behind it is, of course,to remove and burn or bury the trees before they can give rise tospore-laden broods of beetles. Ordinarily, trees are marked in midsummer when symptoms of wilting and yellowing occur, and are removedduring the following winter or spring.

Our seasonal history studies have shown that it would be moreeffective to leave such trees to act as "trap trees" for beetles in thefollowing summer. Figure 1 shows that they may be left safely untilthe end of July at least before any emergence of fresh brood will occur.It also shows that, in order to take advantage of these trees as traptrees, one should not remove them before mid-June. Of course, thedeveloping brood of beetles must be destroyed by burning, burying orbarking the trunks.

(iii) Use of cacodylic acid - a tree killer. Another of our studieshas suggested a way in which infected trees might be enhanced as traptrees. This came about as an unexpected result of testing the commontree killer, cacodylic acid, as a method of controlling beetles.

Previous work by others has shown that the broods of some barkbeetle species fail to develop in trees killed with this chemical.Most of these species are classed as primary insects which attackliving trees. In trials, we have found that native elm bark beetlebrood development is not inhibited in our treated trees, except perhapsin the lower bole of some trees.

Two observations emerged from these trials, however, that maybe put to use. First, sticky traps on three killed trees yielded veryhigh catches indicating a high degree of attraction (Fig. 2) that waslater corroborated by the presence of high larval populations along

30-

25 —

^20-3

CO

a

a

10 —

cr

10

0)a

0)CO

16

1972. Daily Catches on 3 Trees

Treated with Cacodylic Acid.

Traps Installed 12 July

10 May 20

18 Aug. 28

30

1 " I r9 June 19

17 Sept. 27

29 9 July 19

I" ' I T |17 Oct. 27 6

Figure 2. Daily catches on trees treated with cacodylic acid on May 24.Comparison with Fig. 1 shows the greater attractiveness ofthe treated trees.

the trunk. Secondly, we found that the bark on treated trees could beremoved with extraordinary ease.

These observations suggest that it might be possible to create"super traps" out of infected trees by treating them in the spring withcacodylic acid. Destruction of the beetle brood would be simpler, as itwould be easy to remove bark and brood for quick disposal.

(iv) Biological control. The outlook for biological control of thenative elm bark beetle does not appear very promising. A small number ofnative parasite and predator species exists but these do not appear to

17

have a significant effect on beetle populations. Beetle populations inthis area, for example, appear to be almost unaffected by native parasites. They have appeared extremely infrequently in our rearings of manythousands of wild beetles.

A European parasite, Dendrosoter protuberans9 has been introducedin Ohio and Michigan against the European elm bark beetle, apparentlywith indifferent success except, possibly, in the Detroit area where itseems to have taken hold.

As a test in control of the native bark beetle, we have importedthis parasite from Austria and have been rearing it in the laboratorysince last spring using a new method. Previously, rearings have beencarried out by introducing parasites onto beetle-infested elm bolts incardboard boxes. We have had success using elm bark from infested treesin plastic boxes. Female parasites have been observed laying eggs inbeetle galleries through both the outer and inner surfaces of the bark.Also, many adult parasites emerge from the inner bark.

One parasite generation in the laboratory takes roughly a month.Four consecutive rearings have been completed in the laboratory, and agood supply of parasites has been developed in this way. These have beenused to infest material here and in southern Ontario for use in outdooroverwintering tests of parasite survival. Results of these tests willindicate the feasibility of proceeding with large-scale rearings forfree release in the future.

Two free releases of excess parasites have been made at Echo Bayand on St. Joseph Island. Attempts will be made to recover parasitesfrom these points next year.

Summary

The native elm bark beetle occurs in such high numbers, especiallyin areas of high Dutch elm disease infection, that effective controlmay seem impractical. However, there is good evidence that it takes alarge number of beetles to effect the inoculation of a single tree, andprevious success in reducing infection by use of insecticides and sanitation cutting suggests strongly that reduction of beetle populations isworthwhile, even though eradication is impossible.

Our studies of seasonal history and movements of beetle populations have suggested ways of controlling populations economically andwith minimal environmental pollution.

1. Treating the lower trunks of living trees with insecticide inAugust and early May should help to control overwintering adultbeetles, which appear to be the most important segment of thepopulation from the standpoint of spreading the disease organism.

18

2. In sanitation cuttings, trees may be left standing until atleast mid-July of the year in which they appear dead, to actas trap trees.

3. It may be possible to create "super trap trees" by killing thetree with the tree killer, cacodylic acid. Not only are suchtrees attractive to the bark beetle but they develop very loosebark which may be easily removed for destruction of brood.

In addition we are investigating the feasibility of introducingan exotic parasite which attacks larvae of the bark beetle. Results ofoverwintering survival tests will indicate the desirability of massrearing of parasites for free release.

19

LEPIDOPTEROUS SEX ATTRACTANTS: THEIR ROLE IN INSECT CONTROL

Dr. I. Weatherston,Insect Pathology Research Institute,and

Dr. C. J. Sanders,Great Lakes Forest Research Centre,Sault Ste. Marie, Ontario

Sex attractants, which bring the two sexes togetherfor mating, are widespread among insect species. InLepidoptera (including the spruce budworm), sex attractantsare produced by females to attract the males. As a controlmeasure, synthetic attractants can be used to lure males totraps or to disrupt normal mating behaviour by 1) confusingmales through dissemination of sufficient attractant toobscure location of individual females, or 2) jamming malereceptors so that the signal is not received. The mainadvantages over conventional insecticides are specificityto the target insect, the need for small quantities only, _nontoxicity, and application at the time when the insect isnumerically at its lowest (i.e., during the adult stage).

Insects are one of the most successful forms of animal life;they have penetrated every land and fresh-water habitat from thenorthernmost arctic to the hottest deserts. The incredible number andmass of insects give them a dominant role in the ecology of man senvironment - in North America there are about 90,000 species. Only byunderstanding this role can we cope with our deteriorating environment.

In 1963 two scientists of the United States Department ofAgriculture made the following statement: "Insects have managed topersist in hostile surroundings because they have developed extraordinary adaptations or abilities, one of which is ahighly specializedsense of smell. Some insects can follow unerringly an odour trail to asource of food, to host plants or animals, to the right place to laytheir eggs, and to the opposite sex." The chemicals utilized by theinsects in their communication systems are known as pheromones. Theword pheromone was coined in 1959 and used to designate "substances thatare secreted by an animal to the outside and cause a specific behavioural reaction in a receiving individual of the same species . To datethe various pheromones which have been differentiated are trail pheromones,alarm pheromones, aggregation pheromones, territorial-marking pheromonesand sex pheromones.

20

When an adult female moth emerges from her pupal case her onlyfunction is to entice a male to her, mate with him and lay viable eggsto ensure the perpetuation of the species. To communicate with a malethe female moth releases from an abdominal gland a minute amount of hersex pheromone. This volatile material is carried downwind and the molecules are received by the chemoreceptors of the male moth's antennae,from which a stimulus passes to the central nervous system, triggeringoff certain behavioural patterns and causing the male insect to flyupwind until it reaches the source of the odour. In this situation the

communication system of the insect is vulnerable and could be exploitedin an attempt to introduce some measure of control. Such control couldbe brought about if the material secreted by the female moth could becollected in sufficient quantities to allow isolation, purification andchemical elucidation of the biologically active compound. Once thechemical structure is known the compound would be synthesized and fieldtested to show attractancy and would then be available for utilizationin insect control.

The complete characterization of a sex pheromone involvesfive main steps:

1. Bioassay techniques2. Collection of the pheromone3o Purification and structural elucidation4, Synthesis5. Field trials

Bioassay techniques

Good, dependable bioassay techniques are of paramount importancesince it is imperative that at every stage of extraction and purification and after every chemical reaction, the materials involved arebioassayed, i.e., they are tested to see if they are still biologicallyactive. The types of bioassays used may be classified as (i) behavioural (ii) electrophysiological and (iii) field.

(i) Behavioural bioassay. This type of assay is dependent on asexual response from the male moths when they are exposed to femaleextracts, candidate pheromones, etc. The behavioural pattern in malemoths exposed to the specific pheromone is quite general: firstantennal movement is observed, then locomotor activity, intensive winevibration and finally attempted copulation. The intensive wing vibration is usually taken as the criterion of "activity" in behaviouralbioassays.

(ii) Electrophysiological bioassay (EAG). This assay involves themeasurement of the potential difference along a detached antenna while

21

a fast stream of air containing a small amount of test material ispassing over the antenna. The amplitude of a deflection, in millivolts,in relation to the concentration and structure of the test material isdiagnostic.

(iii) Field testing. This usually applies after the attractant hasbeen characterized and synthesized. It will be dealt with later.

Each technique has its advantages and disadvantages and theoptimum situation is a complementary use of all three.

Collection of the pheromone

Once the bioassay techniques have been set up and standardizedthe pheromone must be collected. Preliminary behavioural studies ofmating insects, both in the laboratory and in the field, will havedetermined, for the species being studied, the time of sexual activity,i.e., the hours between the "calling" (i.e., emitting of the pheromone)of the female insects and the corresponding flight and response periodof the males.

There are several methods of collecting the pheromone but thisdiscussion will be restricted to three.

(i) Excision of virgin female abdominal tips. This is the mostwidely used method and consists of extruding the abdominal tips andexcising the last two segments which contain the gland. The tips arestored in a solvent at freezer temperatures until they are required.

(ii) Air flow method. Adult female moths, during peak pheromoneemission period, are contained in a large vessel. Dry air is passedthrough the vessel and "volatiles" are trapped in suitable solvents atdry ice/methanol temperatures.

(iii) Washing. This method was necessary during spruce budworm studieswhen the two methods mentioned above failed to give physiologicallyactive materials. It has since been used successfully with three otherspecies. The method involves putting pupae or newly emerged femalemoths into large glass jars and leaving the moths there until theirpeak pheromone emission period is over. When the insects are removedthe jars are rinsed out with a suitable solvent.

Purification and structural elucidation

Once an active crude material has been obtained the first stagein the elucidation is determination of the class of compound to whichthe active constituents belong. Aliquots of the crude material aresubjected to hydrogenation, hydrolysis, reduction, esterification, etc.,

22

and the reaction mixtures are bioassayed. A second manipulation at thisstage makes use of a combined gas-chromatographic/EAG technique. Extractis injected into the gas chromatograph, the effluent split and the largerpart collected at 1-minute intervals in capillary tubes. The tubes arerinsed out and assayed by EAG, and the areas of activity are marked onthe chromatogram. This procedure is usually carried out on a polar andnonpolar chromatography column.

The data from these two experiments lead to deductions regardingthe class of compound and the carbon number of the active component orcomponents in the crude material.

Purification of a large amount of extract from several tens ofthousands of female tips is by conventional procedures and involves column, thin-layer and preparative gas chromatography and short-pathdistillation. The elucidation is based mainly on gas-chromatographyretention times and mass spectrometry followed, if necessary, by furtheranalytical techniques, e.g., ozonolysis, argentation or substractionthin-layer techniques.

Synthesis

To date approximately 30 sex attractants have been isolated fromfemale lepidopterous species and fully characterized. The variation inthe chemical structure of these pheromones places the consideration ofsynthetic methods outside the scope of a general discussion such as this.It is sufficient to say that the compound of the postulated structure issynthesized and compared chemically with the natural materials

Field trials

Before field testing, the synthetic pheromone will have beenassayed by EAG and behavioural techniques. Related compounds will alsohave been tested in this manner in order to ascertain if they are active.At this stage the sole reason for field testing is to obtain data onthe attractancy of the material under natural conditions. Strong evidence of attractant properties is the final criterion in the completecharacterization of a sex pheromone.

Summary

Bioassay techniques have been developed, the pheromone has beencollected, isolated and purified and the chemical structure elucidated.The synthetic pheromone has been prepared and shown to be chemically andbiologically identical to the natural material. At this juncture the

23

application of the pheromone to insect control may be considered.

Application of the pheromone to insect control

Insect control involves two steps: 1) the detection of thepest and 2) the regulation of its numbers; in both these areas there isa potential use for sex pheromones. In contrast to agricultural pestswhich are perennial problems for the farmer, most forest insects arecyclical, reaching epidemic proportions for several years, then virtually disappearing for a number of years before erupting again. Duringthe period of endemic populations the insects are still there, but insuch low numbers that the damage they do causes no concern. Ideally,outbreaks would be prevented by detecting ominous increases in theinsects1 numbers. This is where sex attractants can play an importantrole.

In early studies in Ontario on the monitoring of spruce budwormwith the sex attractant, square plywood boards coated with "tanglefoot",and having a central insert cut out to house a caged virgin female moth,were used to trap males. In an area of extremely low population (lessthan 1 budworm/100 branches by larval count) such traps regularlycaught two or three moths each. These are not very high catches butthey are sufficient to give a reliable population estimate from 10traps (see Table 1). It would take a three-man crew equipped withpole pruners several weeks to obtain the same number of larvae.

Table 1. Trapping data versus larval counts in spruce budworm populations at Black Sturgeon Lake, Ontario

Adults pervirgin-?-baited

Year Larvae per branch trap

1966 0.03 4.2

1967 0.01 31.6

1968 0.09 13.7

1969 0.03

1970 0.00 2.2

1971 0.00 1.6

1972 0.01 1.0

24

While this technique is very efficient it would be even moreso if live females could be replaced by "synthetic females" since livefemales have several disadvantages, viz., they must be reared, they varyin potency and they die after a very short period. With the identification and synthesis of the spruce budworm pheromone, a source of attractanthas been obtained and all that is required is a dispenser so that thepheromone is released at a slow, uniform rate similar to that of thefemale. Various workers have tried different dispensers such asimpregnated filter paper or dental floss wicks; however, in the sprucebudworm studies it has been found that small, hollow, polyethylene vialstoppers filled with small quantities of the attractant make excellentdispensers.

Like live female moths the sticky plywood boards have theirlimitations. They are large, bulky and, when coated with "tanglefoot",extremely messy and difficult to handle. They have now been supersededby commercially available, small, disposable cardboard traps.

A preliminary evaluation of these traps, baited with attractant-filled polyethylene stoppers, as monitors of spruce budworm populationswas made in 1972 across the whole of eastern Canada, and involved 272sampling locations. Owing to differences in the quality of the synthetic pheromone the catches were much lower than anticipated. Such teethingproblems are to be expected, however, and there is no doubt that in ayear or two the trapping technique will be very useful for monitoring thefluctuations in the normal low-level populations of the spruce budworm.

How can sex attractants be used in the control of pests? Thefirst obvious answer is by trapping male insects, reducing the probability of the females1 mating and hence the number of fertile eggs laid.This has been tried with some success in regulating the numbers oforchard pests, and the United States Department of Agriculture isattempting to stop the spread of gypsy moth by trapping the males alongthe advancing front of the infestation. (It should be noted that thefemale gypsy moth does not fly.) Unfortunately, the area susceptible toattack by the spruce budworm in Canada is not the 20-30 acres of a fruitgrower, or even the quarantine zone in front of an introduced pest suchas the gypsy moth. The spruce budworm is a native insect of the forestsof Canada endemic to millions of acres. The female insects are highlymobile so that even if enough males were trapped to prevent matings in asmall area, hundreds of females which mated elsewhere could fly intothe area or be carried there on wind currents. To regulate spruce budworm numbers, therefore, enormous areas would have to be treated withvery large numbers of traps, and as the data in Table 2 clearly show,this is impractical.

25

Table 2. Number of traps required per acre to effect control of sprucebudworm

% control

50

75

90

Trap efficiency (=female equivalent in trapping power)

1 5 10

*90,000

45

*180,000

90

*1,800,000

900

*30,000

L5

*36,000

JL8

*360,000

180

*9,000

5.

*18,000

9

*90,000

90

* These numbers represent traps required in "outbreak" densities.Underlined numbers represent traps required at endemic levels.

If trapping is impractical, what are the alternatives? One isto interfere with the insects1 communication system so that the malescannot find the females. If the forest is permeated with the attractantthe males may become confused and unable to find the females. If thechemical is present in sufficient quantities the males may lose theirability to perceive it and their nervous systems will become fatigued orhabituated. A second alternative is to use another chemical which masks

or inhibits the effect of the real attractant, thus blocking the message.In the spruce budworm studies two chemicals were discovered which inhibitperception of the pheromone by the males. Data are given in Table 3.

Preliminary trials this year, using insects in large cages inthe field, have shown that at least some matings can be prevented bypermeating the atmosphere with either the attractant or the inhibitors.Cages, however, create a very artificial situation; the insects areconfined in an unnaturally restricted area and many males and femaleswill meet accidentally. It is hoped that, at low population densitiesin the field where males have to find the female over distances of up

to 100 ft, the chemicals will be more effective in disrupting matingbehaviour.

Confusion, habituation and inhibition all involve the spreadingof chemicals uniformly through the forest in some kind of formulationwhich will ensure the stability of these rather unstable compounds,

26

Table 3. Percentage catch of male budworm in the field, using differentchemicals

Chemical Catch (%)

* Aldehyde 100

** Acetate 3

Acetate and aldehyde 8

*** Alcohol 0

Alcohol and aldehyde 4

* Aldehyde (E)-ll-tetradecenal (the sex pheromone of the spruce budworm)

** (E)-ll-tetradecen-l-ol acetate

*** (E)-ll-tetradecen-l-ol

while allowing them to diffuse gradually during the flight period of thespruce budworm, a period of 3-4 weeks. Further trials must await thedevelopment of a suitable formulation, but again it is hoped that this isa technological problem to which an answer will be found.

If these chemicals prove effective in disrupting mating and hencein regulating insect numbers, what will be their advantages over the moreconventional insecticides? First, they will be utilized during theadult stage when the insect is numerically at its lowest and where anyadditional mortality may be critical. Second, extremely small quantitiesof the chemical will be required - hundredths of a gram per acre. Third,they are essentially species specific and will affect only the targetinsect. Finally, since they are naturally occurring compounds whichbreak down quickly, they will be virtually nonpolluting.

References

Jacobson, M. 1972. Insect sex pheromones. Acad. Press, New York. 382 p.

Tahori, A.S. Ed. 1971. Chemical releasers in insects. Pesticide Chem.,Vol. 3. Gordon & Breach. 227 p.

Wood, D.L., R.M. Silverstein and M. Nakajima. 1970. Control of insectbehaviour by natural products. Acad. Press, New York. 345 p.

27

CONTROL OF INSECT PESTS USING SYNTHETIC INSECT HORMONES

Dr. A. Retnakaran,Insect Pathology Research Institute,Sault Ste. Marie, Ontario

A new avenue for insect control has opened withinduction of developmental abnormalities using syntheticinsect hormones that interfere with cellular differentiation.The most promising candidate is juvenile hormone, andalready many mimetic chemicals have been produced. Advantagesfor use as future control agents include biodegradabilityand insect specificity.

Introduction

Hormones are chemical messengers secreted predominantly byductless glands called endocrine glands. Most people have heard ofthe thyroid gland and the hormone it secretes. Excess or insufficientamounts of the hormone produce characteristic diseases. Synthetichormones are currently being used to cure such endocrine disorders.Early in the 1960fs it was found that estrogens (ovarian hormones)prevent the release of certain pituitary hormones (LS and FSH) that arenecessary for ovulation. On the basis of this finding, hormone preparations made of an estrogen and a gestagen were made available forcontraceptive use. This perhaps is the most significant contributionto human population control.

Many of the insect problems we have at present can be tracedback to the population explosion of a species. Control of such pestsdepended (and still does, to a great extent) on insecticides. Limitedbiodegradability and mammalian toxicity have made many insecticidesunsuitable for widespread use. A search for alternatives has, amongother things, stimulated intense research in insect physiology. Onepossible avenue of research that promises to be rewarding is the studyof insect endocrine glands.

Insect Hormones

The study of insect hormones is a relatively new field. Thedistribution of the various endocrine glands that have been described todate is shown in Figure 1. Below the brain there are two pairs of

28

glands called corpora cardiaca and corpora allata. Each corpus allatumsecretes a hormone called juvenile hormone. Above the esophagus is apair of glands called the prothoracic glands that secrete ecdysone. Thenervous system itself secretes several neurohormones. The brain, forexample, has been shown to secrete at least six hormones. The sub-esophageal ganglion of the silkworm secretes diapause hormone. Thebrain and the central chain of ganglia secrete diuretic and antidiuretichormones. The midgut of the European cornborer secretes a hormone calledproctodone. The apical cells of the testis in the glowworm secrete ahormone called the androgenic hormone.

Bursicon

Bursicon is the tanning hormone found in flies and cockroaches.The nature of its function is schematically shown in Figure 2. Theemerging blowfly is white and soft. If the neck is ligated the thoraxand the abdomen remain white and untanned. If the hemolymph from adarkened fly is injected into the abdomen of a ligated white fly theabdomen tans. The hormone is secreted by the brain and has been characterized as a protein.

Diapause Hormone

The silkworm has two generations per year. It overwinters as anegg. The eggs that go into diapause do so as a result of the influenceof photoperiod on the maternal moth. The effect of photoperiod in controlling the secretion of diapause hormone during the life of thisinsect is shown in Figure 3. The larvae emerge in spring and the adultmoths lay their eggs in summer. Influenced by the shortness of the daysin spring, the female moth lays nondiapause eggs. On the other hand themoths that lay eggs during fall are influenced by long day conditionsand therefore lay diapause eggs. The ecological adaptation of thisbivoltine (two generations per year) species becomes obvious.

Diuretic Hormone

The excess water in an insect is eliminated through the alimentary canal by the malpighian tubules. A schematic representation of theassay for diuretic hormone is shown in Figure 4. The malpighian tubulewith a part of the midgut is mounted in a drop of mineral oil. Twodrops of insect saline are placed on either end of the system. The volume of the drop in the midgut region is periodically measured with asyringe. It can be shown that the diuretic hormone induces excretion ofwater. Recently it was shown that some carbamate insecticides stimulatediuretic hormone production as a result of which the insect excretes

29

excessive amounts of water and dies of desiccation. Perhaps at a futuredate this will have practical implications.

Antidiuretic Hormone

The cockroach rectum resorbs water to prevent desiccation.Figure 5 shows how an isolated rectum resorbs water from the surroundingmedium.

Hyperglycemic Hormone

The function of the hyperglycemic hormone is shown in Figure 6.It is secreted by the corpus cardiacum and increases the blood sugarlevel of the insect. It is similar to the mammalian hormone glucagon.

Brain Hormone

The basic endocrine system in the head and thoracic regions of aninsect is shown in Figure 7. The two pairs of neurosecretory cells canbe seen in the brain. They secrete a hormone called ecdysiotropin whichstimulates the prothoracic gland to produce ecdysone.

Ecdysone

The hormone ecdysone has many functions as depicted in Figure 8.This is perhaps the most thoroughly studied of all hormones. It actsat the genetic level and induces gene activity. The giant salivarychromosomes (Polytene chromosomes) show characteristic puffs whentreated with this hormone. Wing buds require ecdysone in the medium inorder to differentiate. Dopa decarboxylase is an enzyme induced byecdysone and can be synthesized in an in vitro system consisting of ratliver microsomes and RNA extracted from ecdysone-treated insects.Ecdysone is necessary for the transport of the macromolecular factoracross the testis wall. This MFP stimulates spermatocyst differentiation. By far the most important function of ecdysone is the inductionof moulting in insects. It is necessary for larval-to-larval moulting,larval-pupal moulting, and pupal-adult moulting. The hormone is produced by the prothoracic gland only when the brain hormone is present.Recently it has been shown that ecdysone is produced even in the abdomen.The German physiologist Karlson isolated 15 mg of the hormone from halfa ton of silkworm pupae in 1965.

Many plants, especially the ferns, have phytoecdysones andpossess moulting hormone properties. Some people believe that these

30

plants developed, in the course of their evolution, the ability to produce the hormone-like substance as a means of protection againstinjurious insects.

Juvenile Hormone

The functions of juvenile hormone are represented diagrammaticallyin Figure 9. This hormone has four important functions: i) It inducesthe prothoracic gland to produce ecdysone. (This function is referred toas the prothoracotropic effect.) ii) It acts at the genetic level. Alarva remains as a larva under the influence of this hormone. Excessjuvenile hormone will produce supernumerary instars. (This function isreferred to as the morphogenetic effect.) iii) It is necessary for thefat body to synthesize vitellogenin (female protein) and transport thematerial into the ovary. In its absence the ovary fails to develop.(This function is referred to as the gonadotropic effect.) iv) Eggsnormally do not contain juvenile hormone. When juvenile hormone isapplied to eggs the embryo dies at a certain stage. (This function isreferred to as the ovicidal effect.)

Ecdysone and Juvenile Hormone During Metamorphosis

The titer of ecdysone and juvenile hormone during the life of theinsect is shown in Figure 10. These two hormones act together for theharmonious differentiation of the insect. Ecdysone is necessary formoulting. Juvenile hormone determines the outcome of the moult. Whena larva moults into another larva both hormones are present in substantial amounts. When the larva moults into a pupa the titer of juvenilehormone drops precipitously. The outcome is a pupa. Juvenile hormoneis absent for the most part during the pupal stage. In the adult it isnecessary for ovarian development.

Embryonic Blockage

The mechanism of the ovicidal action of juvenile hormone isrepresented in Figure 11. It is apparent that the time of applicationdetermines the stage at which embryonic blockage occurs. Many syntheticanalogs that have juvenile hormone properties have become available inthe last few years. Since this affords a method of control some of theanalogs were tested on forest insects in the laboratory. The eggs ofthe white pine weevil were treated with an analog of juvenile hormone,ihe development of the treated eggs was followed by dissection of thehead capsule. The results are summarized in Table 1. At the higherconcentrations only 50% showed head-capsule development. The ovicidaleffect of the analog is shown in Table 2. At higher concentrationsthere is a significant decrease in the number that hatched; none of the

31

treated embryos survived. The ovicidal effect of juvabione on thespruce budworm is summarized in Table 3. This compound is a naturalcomponent of balsam fir and has been extracted from pulp mill effluentsas well. The ovicidal activity of two other analogs along with juvenilehormone is summarized in Table 4. The dichloro analog was by far themost effective.

The United States Department of Agriculture came up with twocompounds that were considered 1000 times as active as the authentichormone. These aromatic terpenoid ethers were tested on the spruce budworm eggs. The results shown in Table 5 proved to be less dramaticthan expected.

Application of juvenile hormone to the last larval instarresults in the formation of a larval-pupal intermediate. Some larvaetreated with an analog of juvenile hormone (ZR 515) moulted into supernumerary instars. Normally the spruce budworm has six larval instars.The hormone induced two additional instars. The production of supernumerary instars, which was confirmed by measuring the head-capsulewidth, is summarized in Table 6.

The hemlock looper is extremely sensitive to juvenile hormoneanalogs. Application to the larvae at the level of 10 ppm inducesproduction of larval-pupal intermediates. In tests conducted in ourlaboratory, none of the larvae survived.

Some greenhouse experiments were conducted to determine thepersistence of the juvenile hormone analog (ZR 515). This analog iseasily broken down in water. When sprayed on the plants, however, itis absorbed by the waxes in the needles and survives UV exposure anddrenching with water. Spruce budworm larvae placed on these plants 21days after the treatment still showed the effects of the hormone.

The Colorado potato beetle overwinters as an adult. Duringthis period juvenile hormone is not produced (Fig. 12). During springwhen the days become longer the corpus allatum becomes active andsecretes juvenile hormone. As a result the ovary develops and theinsect, upon reaching sexual maturity, reproduces. Application of thehormone to the overwintering beetles breaks diapause. The insectreaches sexual maturity precociously.

The white pine weevil also has an adult diapause. The hormone,if applied to the insect in late fall when it goes into diapause, willinduce sexual maturity. Under the influence of the hormone the insectwill feed and reproduce and will not be aware of the onset of winter.As a result it will die. This hypothesis is being tested in our laboratory.

32

It is obvious from the foregoing presentation that insect hormoneswill play an important role in pest control. While it may not be apanacea for our insect problems it certainly is a harbinger of controlmeasures to come.

33

Table 1. Effect of a juvenile hormone analog onthe embryonic development of the headcapsule in the white pine weevil

Treatment

(yg in 1 yl)

No. of eggstested

Embryosshowing head

capsule developmentafter 6 days

(%)

Acetone control 50 50

10 50 40

50 50 32

100 50 24

200 50 24

34

Table 2. Ovicidal effect of a juvenile hormone on the white pineweevil

Treatment

Amount applied(yg in 1 yl/50

eggs)No. eggstreated

Hatched

after 8

days

(%)

Larvae

alive after

12 days(%)

Control (no treatment) 0 25 76 36

Control (1 yl acetone) 0 50 60 28

Juvenile hormone analog 10 50 60 32

ti ii n50 50 34 16

ii ii it100 50 26 4

" ii it200 50 16 0

35

Table 3. Effect of juvabione on the eggs of the sprucebudworm

No. eggs No. eggsTreatment Amount applied treated hatched Hatched

(yg/egg mass) (%)

Control 0 136 133 98

Juvabione 114 185 0 0

57 139 7 5

36

Table 4. Effect of juvenile hormone and two analogs on theeggs of the spruce budworm

No. eggs No.• eggsTreatment Amount applied treated hatched Hatched

(Mg/egg mass) (%)

Control 0 121 115 95

Juvenile hormone 100 197 0 050 153 0 025 137 18 1310 147 97 66

Dichloro analog 200 155 0 0100 131 0 050 141 0 025 147 8 5

Farnesyl methyl 200 1.61 0 0ether 100 139 0 0

50 157 17 1125 141 31 22

37

Table 5. Effect of two aromatic terpenoid ethers on the eggs ofthe spruce budworm

No. eggs No. eggsTreatment Amount applied treated hatched Hatched

(yg/egg mass)

Control

Methyl-aromaticterpenoid ether

Ethyl-aromaticterpenoid ether

0 216 211 98

100 199 12 6

50 210 27 13

25 213 59 28

100 186 0 0

50 191 17 9

25 207 32 15

38

Table 6. Increase of head-capsule width as a function ofinstar in Choristoneuva fumiferana larvae

Instar

Samplesize

Head-capsulewidth

(x ± S.D. mm) Range(mm)

Head-capsulewidth

(x 1.4)

1 18 0.2311 ± 0.0102 0.22 - 0.24 0.32

2 22 0.2918 ± 0.0133 0.26 - 0.32 0.40

3 12 0.4225 ± 0.0486 0.38 - 0.50 0.59

4 13 0.6257 ± 0.0562 0.56 - 0.76 0.88

5 20 0.9970 ± 0.0426 0.92 - 1.08 1.40

6 17 1.6752 ± 0.0705 1.52 - 1.76 2.34

7* 13 2.1084 + 0.0816 2.00 - 2.20 2.95

8* 2 2.7700 ± 0.1100 2.64 - 2.80 3.81

* Supernumerary instars induced with a juvenile hormoneanalog.

SITES OF HORMONE PRODUCTION IN INSECTS

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42

THE USE OF BACTERIAL INSECTICIDES IN FOREST INSECT PEST CONTROL

Dr. P. G. Fast,

Insect Pathology Research InstitutesSault Ste, Marie, Ontario

Preparations of Bacillus thuringiensis are at present being marketed commercially under various trade namessuch as Thuricide and Dipel. These preparations are specific stomach poisons for larvae, of Lepidoptera, many ofwhich are important agricultural and forest pests. Thecharacteristicsy benefits and disadvantages of the abovepreparations are described.

The possible relationships between insects and microorganismsrun the gamut from obligate symbiosis in which insects and microorganismsare dependent on each other, through the various facultative relationships to obligate parasitism. In the latter situation the hostfrequently suffers because of the association with the microorganism andin this situation the term "disease11 as we usually define it is entirelyapplicable.

The recorded history of insect diseases begins with Aristotlefsdescription of certain diseases of the honeybee in his Historia Animalium.In the 16th, 17th and 18th centuries numerous references were made todiseases of silkworm because of the great economic importance of thisinsect. The earliest association between microorganisms and diseasesof insects is attributed to Agostino Bassi who, in 1834, showed that afungus, now named Beauveria bassiana in his honor, was the cause of aninfectious disease in the silkworm. Louis Pasteur studied the diseases

of silkworms between 1865 and 1870 and devised methods to save the

French silk industry from almost certain ruin. But the recommendationthat insect diseases be studied to determine the most effective means ofusing them against noxious insects was not clearly stated until 1873.

No concerted effort was made, however, to follow up this suggestion until 1945 when a Department of Insect Pathology was established atthe University of California and, a year later, the Insect PathologyLaboratory was founded in Sault Ste. Marie. The development of resistance to chemical insecticides and the realization of their undesirableeffects on nontarget organisms have added impetus to the study of diseasesas control agents for insects. Over the years a considerable body ofinformation concerning organisms that cause disease in insects and theepidemiology and pathology of these diseases has accumulated.

43

As might be expected, bacteria, because of their associationwith disease in human beings and the availability of techniques fortheir identification and culture, have received considerable attention.Two techniques are used: the examination of insect cadavers, with cul-turing of any extraneous microorganisms and testing of them againsthealthy insects; and the study of bacterial fauna of supposedly healthyinsects to determine under what conditions these will cause disease.The first approach has more chance of yielding candidates for control.Of the many hundreds of such studies only two candidate species forcontrol of important insect pests have emerged.

The first of these is a bacterium that controls the larvae ofthe Japanese beetle, the white grub, notorious for causing damage inpastures and turf. The bacterium Bacillus popilliae causes disease onlyin one group of closely related beetles, the Scarabeidae. It cannotbe readily cultured artificially; hence, the disease organisms, likeviruses, are grown in mass rearings of the insect host. Diseased larvaecontaining millions of spores of the organism are ground up and mixedwith an inert filler. One such product, "Doom powder", applied as asurface dressing in areas where the insect occurs, is highly successful.

The second candidate for insect control is another bacillus,Bacillus thuringiensis. This bacterium is readily cultivated on artificial media and is presently being produced commercially in severalcountries. It is being used extensively on leaf vegetable crops and isbeing evaluated now, in Canada, for the control of insect pests offorests. The bacterium, Bacillus thuringiensis, is a long, rod-shaped,gram-positive, aerobic bacterium which grows very well in almost anyorganic medium and is very readily cultured in commercial fermentationprocesses. When the nutrients in the medium are exhausted the bacterium sporulates to form an environmentally resistant spore. At thesame time that the spore is formed a diamond-shaped body also forms inthe same cell. The cell wall then dissolves and the spore and thediamond-shaped body (which we call the crystal) are set free. It wasDr. T. A. Angus of the Sault Ste. Marie laboratory who showed back inthe 1950fs that it was this crystal, this diamond-shaped body, that wascrucial to the toxic effect of the bacteria.

These crystals are aggregates of protein molecules; there is noevidence of either carbohydrate or lipid being an essential part ofthem. The proteins comprising the crystals are of various sizesdepending on the strain of bacterium from which the crystals are derivedand the way in which they are dissolved. When the crystals are dissolvedin detergents, about 90% of their proteins are quite large and 10% aremuch smaller, about the size of an antibiotic molecule, but still toxic.The crystals are toxic at levels of 0.5 yg/insect, but only when theyare eaten by a susceptible insect. (Only certain species of insect, allof which are members of the order Lepidoptera, are susceptible.)

44

The sequence of events leading to the death of the larva havebeen studied in several laboratories. The details of this sequence varyfrom insect to insect but in the majority of the species both the sporearid crystal are required to kill the insect. The sequence can be visualized as beginning when the insect ingests amixture of spores andcrystals. The crystals dissolve in the gut and fragments of the crystals rapidly pass through the gut wall into the hemocoel or blood spaceof the insect. Appreciable quantities of the fragments are found inthe blood within 1minute after the spore crystal mixture is ingested.Within this same period of time uptake of glucose by the gut is increasedover the control condition but no other changes in permeability havebeen detected this soon. By 7minutes after ingestion, however, changesin permeability of the gut to potassium ions and to other compoundshave been observed. By 15 minutes gut metabolism has been badly distorted. Within 20 minutes after treatment the cells of the gut wall™?™ii SW^ ,f̂ ^ 3° mlnUteS many °f the Cells have burs* and thetllZl I ?? 7 dlsorSanized- At this stage the pH of the gut contents has fallen to a level at which the spore can germinate andmultiplication of the bacterium occurs. The insect appears to die as aresult of this bacterial infection.

i„co •/ con?omitant of the damage to the gut wall is that the insectloses its desire to feed. It is this characteristic which gives thedisease its commercial value. Insects are of economic importance primarily because they feed on plants or plant products important to thehuman economy either as food or as fiber, if feeding can be inhibitedxt matters little how long it takes the insect to die. Feedingstopswithin minutes after ingestion of crystals although the insect mav livefor days or even weeks thereafter. Protection of the crop or ProLc"thas thereby been achieved. P Product

ma,ro If t^ngiensis disease, milky disease organisms or, for thatmatter, any other disease organism is so effective why do insects stillremain aproblem? The basic reason is that individual insects rarelycome in contact with sufficient numbers of disease agents to induce thedisease state Most organisms have only arelatively^ short survivalattLkT ^^ h°St- SUnlight' ra±n Wash> -dement temperatures andraoidlv the rh°rga?iSmSKf *nvironmental chemicals all serve to reduceJ ^e nJ"nber <* viable disease organisms in the environment.Insects also have their own protective mechanisms.

The basic means of overcoming these drawbacks is the same forinLcCtLid°e T W\6thr ^ be.VirUS* b«*«*». ^ngus, or cWcalinsecticide In control operations sufficient disease organisms orchemical molecules are spread around that the target insect cannotavoid contracting the disease or contacting the chemical. Additivesto the spray formulation provide protection from sunlight, washine offand other hazards to the microorganism. wasnmg ott

45

The safety of commercial preparations has been demonstrated.Tests have been carried out on rats, mice, chickens and human beings.Feeding tests gave no evidence of untoward effects. Injection of commercial formulations into the animals, either into the abdomen or intothe blood stream, failed to kill the test animals or provide any indication of toxic effects. Nor did inhalation experiments in the testanimals or in human beings cause any difficulties. Acute and chronicfeeding tests on chicks, laying hens, fish, young swine and honeybeeadults and larvae were all negative. Serial passage of bacteria onblood-dextrose media failed to select strains of bacteria toxic to mice.

Among insects only the larvae of Lepidoptera are susceptible; no effectson other organisms have been detected. Thus the effects on the ecosystem in which Bacillus thuringiensis is used appear to be limited tothose affecting and affected by the pest species. In some strains ofB. thuringiensis a water-soluble exotoxin is produced which affectsother insect orders but this is removed from commercial preparations.

When conventional chemical insecticides are introduced into an

environment all or most insects are killed whether they are pests orbeneficial species. The predators and parasites that have been helpingto keep the pest population in check, as well as innocuous or beneficialspecies, are killed. The effect on the ecosystem is enormous, thoughnot necessarily immediately apparent. Massive rebound of the pestspecies to levels higher than those obtaining before the initial sprayor outbreaks of other pests as a consequence of spraying are commonplace.

Bacillus thuringiensis preparations attack only larvae ofLepidoptera, very few of which are beneficial. The populations ofpredators and parasites should therefore fluctuate under the influenceof natural controls and continue to hold residual pest populations incheck. No rebound should occur, and the ecosystem should be affectedonly by the reduction of the pest population.

The cost per acre for application of Bacillus thuringiensispreparations is still somewhat higher than for chemical pesticides.However, because of its safety, its specificity for pest insects andits lack of residual effect it may well prove to be more economicalthan chemical control when such factors are entered into the cost/benefit equation. Field tests of various formulations for ground andaircraft application are being evaluated at present.

Reference

Burges, H.D., and N.W. Hussey, Ed. 1971. Microbial control of insectsand mites. Chapters 10 and 11. Acad. Press, New York. 861 p.

46

THE USE OF VIRUSES IN THE CONTROL OF FOREST INSECT PESTS

Dr. J. C. Cunningham,Insect Pathology Research Institute,Sault Ste. Marie, Ontario

Many species of insects are susceptible to a variety of diseases caused by viruses> fungi, bacteria* protozoa,rickettsia or nematodes. Insect populations in the fieldoften collapse naturally from diseases. Epizootics can beinitiated in some species by the artificial disseminationof a virus. This provides an excellent means of biologicalcontrol as viruses are specific for the target insect pest(beneficial insects, other invertebrates, vertebrates andplants are not susceptible) and, once established in aforest insect population, persist from one year to the nextand spread from the original introduction. Most of thedifferent types of insect viruses contain virus particlesembedded in a protein inclusion body and are thus very stableand easy to handle. In advance of virus control projects,large numbers of insects are reared in the laboratory andvirus is propagated in them, stored, and sprayed in the fieldat the correct stage of insect development.

Introduction

Viruses are widespread in the ecosystem and cause diseases,sometimes mild and sometimes lethal, in almost every form of life.Even bacteria are susceptible to viruses called bacteriophage. Therelationship between insects and viruses 'is a very special one.Insect viruses can be divided into two categories: 1) viruses whichare exclusively infectious to Insects and 2) plant and mammalianviruses which are transmitted by insect vectors. It is only the firstgroup which will be discussed in this review.

Domestic insects as well as pest species are susceptible tovirus diseases and the first insect virus disease described in 1856 was

from the silkworm, although it was not recognized as a virus at thattime. Virus diseases of domestic insects, man, animals and plants arestudied with a view to developing therapeutic measures. Research onpest insects has a diametrically opposite goal: the initiation andspread of an epizootic as a means of biological control. Such anapproach has been used only once in mammalian virology when myxomatosisvirus was used to control rabbits in Australia and in Europe.

47

Viruses are prevalent in natural populations of insects andevery year new isolations are recorded. Viruses sometimes cause thenatural collapse of insect populations and scientists try to manipulateviruses to control pest species.

The very word "virus" alarms many people but viruses are highlyhost specific and insect viruses are safe to man, animals, plants andall other forms of life. When an insect population collapses naturallyfrom a virus disease the environment is very heavily contaminated withthe virus released from the dead insects and every animal and plant isexposed to it without adverse effect. There are published reports tha!tcabbage, as sold in supermarkets, is frequently contaminated with cabbagelooper virus an# those who eat coleslaw almost certainly at times ingestthis virus.

The Viruses Which Infect Insects

A virus is an obligate parasite which can reproduce only in aliving cell. In an infected cell the virus overrides the cell's geneticcode and instead of functioning normally its metabolism is directed toproducing more virus. A virus particle consists of nucleic acid withina protein coat. The nucleic acid, which contains the virus's geneticcode, may be ribonucleic acid (RNA) or deoxyribonucleic acid (DNA).Many insect viruses are morphologically similar to animal viruses buttheir genetic material only codes the information necessary to infectthe cells of the host insect.

There are different types of insect viruses which have a varietyof shapes and sizes. Most of the insect viruses recorded to date areinclusion body viruses; i.e., the virus particles are embedded withinrigid macromolecular paracrystalline protein bodies which are largeenough to be observed under a light microscope (Fig. 1). It is necessary to use an electron microscope to see the actual virus particles(Fig. 2). Such inclusion bodies are unique to insect viruses. Wheninclusion bodies are ingested by an insect larva they dissolve in thealkaline gut juices and the virus particles which are liberated penetratethe gut cells and cause infection.

Insect viruses are fairly host specific and one virus willprobably infect only the host from which it was isolated and perhapssome closely related species. For example, some viruses of the sprucebudworm will infect jack-pine budworm larvae and viruses of the westernhemlock looper and western oak looper will infect eastern hemlocklooper larvae. This specificity is both an advantage and a disadvantagealthough the former outweighs the latter. The advantage is that thetarget pest species is the only insect infected in the forest ecosystem.Predators, parasites, aquatic insects and other beneficial insects such

48

as pollinators are unaffected. The disadvantage is that a virus must bepropagated for each of our pest insects and, in most cases, can be usedonly against that one pest.

Some of the inclusion body viruses are referred to as poly-hedrosis viruses because the inclusion bodies appear to be many sidedAlthough incorrect in some cases the use of this word has remainedBrief descriptions of the different types of insect viruses follow.

1) Nuclear-polyhedrosis viruses have rod-shaped virus particleswith DNA as the nucleic acid. The nuclei of midgut, fat,and epidermal cells are the principal tissues infected.Exceptions are the nuclear polyhedrosis viruses of sawflieswhich infect the midgut only. This is the group of viruseswith the greatest potential for control.

2) Granulosis viruses are similar to nuclear-polyhedrosis virusesbut contain only one virus particle per inclusion body. Theinclusion body is capsule shaped. Some viruses in this grouphave been used to control important agricultural pests.

3) Cytoplasmic-polyhedrosis viruses have spherical virus particleswith RNA as the nucleic acid and they infect the cytoplasm ofgut cells only. *

4) Entomopoxviruses have large, oval virus particles embedded inoval inclusion bodies. Their nucleic acid is DNA and theyinfect the cytoplasm of gut, fat and epidermal cells.

5) "Nonoccluded viruses" is a convenient term to cover all insectviruses in which the virus particles are not embedded in inclusion bodies. Taxonomically they cannot be grouped together asthey differ in their size, shape and nucleic acid. One suchvirus has been used to control citrus red mites in Californiaand another to control the rhinoceros beetle in the SouthPacific but none has been used in Canada; hence these will notbe discussed further.

All the inclusion body viruses infect the larval stage of insectsand to date, there has been no record of the development of resistanceby the host to any of these viruses. Pesticide resistance is a seriousproblem in insect populations subjected to repeated applications ofcertain chemicals.

The Forest Insect Pests

Forest insect pests in Canada belong mainly to four orders ofinsects: Lepidoptera (butterflies and moths), Hymenoptera (sawflies)Coleoptera (beetles) and Homoptera (aphids). Some of the majorLepidopterous pests are the spruce budworm, eastern hemlock looper the

49

Douglas-fir tussock moth and the forest tent caterpillar. Viruseshave been found in all these species and are currently being investigated. Several species of sawflies are susceptible to virulentnuclear-polyhedrosis viruses; they include the European spruce sawfly,European pine sawfly and red-headed pine sawfly. No viruses have beenfound in economically important beetles in Canada or in the balsamwoolly aphid which is a very serious pest in some areas.

Mass Propagation of Viruses

Viruses, unlike Bacillus thuringiensis and entomophagous fungiwhich can be grown in broth cultures, must be grown in living cells.All mammalian viruses are now routinely grown in tissue cultures. Atpresent, insect tissue culture is still in its infancy and large-scalevirus production by this means cannot be considered practical at thistime.

Viruses must therefore be produced in insect larvae either inthe field or the laboratory. Field production is simple in species ofinsects which live in colonies, such as the European pine sawfly andthe red-headed pine sawfly. Colonies are sprayed with virus and whenmortality commences larvae are collected and brought to the laboratorywhere they are freeze-dried and stored. This method is very economical.

In the laboratory, insects are reared on their natural foodor on artificial diet, the latter being more convenient. Since sprucebudworm viruses cannot be produced in the field a large rearing programme was organized at the Insect Pathology Research Institute,Sault Ste. Marie, Ontario. In the winter of 1971-72 about 2,000,000budworm larvae were reared on artificial diet in plastic cream cups,infected with virus, harvested and processed.

Dissemination of Virus in the Field

There are two methods of using an insect virus for control.First, it can be used in the same manner as an insecticide or asBacillus thuringiensis to give immediate control. A sufficiently highdosage or repeated dosages must be applied so that insect mortality israpid and the crop protected. This method has been used on high-valueagricultural crops and two companies in the United States are marketingcorn earworm virus.

The situation in forestry is completely different. The crop isnot harvested at the end of the season and a certain amount of defolia

tion of the trees can be tolerated. Viruses can therefore be used as

true biological control agents in forests with long-term results as the

50

ultimate aim. Thus it is not important that viruses give a high level ofcontrol in the year of introduction provided that they persist in theenvironment from one year to the next and spread from their point ofintroduction.

Viruses are sprayed with the same equipment used for pesticidesand they may be applied from the ground or the air. Unlike modern contact insecticides, viruses must be ingested to cause mortality. Verygood coverage of the trees is therefore required and breakup of the sprayinto fine droplets is of paramount importance if low-volume applicationsare being made. Viruses have been applied effectively from the air at1 gallon per acre, a figure which is still considered high for chemicals.

Very little research has been done on the important topics ofpersistence, transmission and spread of viruses. Transmission from onegeneration of insects to the next can occur in three ways: 1) by trans-ovanal transmission (i.e., the virus is within eggs laid by chronicallyinfected adults), 2) by transovum transmission (i.e., the virus is onthe outside of eggs) and 3) by contamination of soil or foliage withvirus which escapes inactivation by solar radiation and persists fromone year to the next. Spread can be caused by chronically infectedor virus-contaminated adults which oviposit in unsprayed areas. Otheragents are parasites and predators which move from diseased to healthylarvae, and birds which ingest diseased larvae. Viable virus has beenrecovered from bird faeces.

Generally as insect larvae mature their susceptibility toviruses diminishes and it is usually desirable to spray the virus earlyin the season. Viruses are slow to kill when compared to chemicals orBacillus thunng%ensis. The most virulent ones kill larvae in about 5days but periods of 20 days or longer are frequently recorded betweenapplication and mortality. This period depends on the stage of development of the larvae, the dosage and the temperature in the field. Theuse of viruses is not recommended in situations where further insectdamage must be terminated immediately.

A virus is susceptible to inactivation by the ultraviolet radiation m sunlight and on exposed surfaces rapidly loses activity Viruson the undersides of leaves or in crevices shielded from sunlight isunaffected and the addition of sunlight protectants to virus formulations helps to reduce inactivation.

Research on insect viruses started in Canada in the 1940's whena nuclear-polyhedrosis virus accidentally introduced with parasites wasfound to control the European spruce sawfly. In the 1950*s nuclearpolyhedrosis viruses were found which could be used to control theEuropean pine sawfly and the red-headed pine sawfly. These viruses

51

persist in the field and spread rapidly when correctly applied. Theyshould be used as control agents whenever outbreaks of these sawfliesoccur.

Most of the research is currently centred on the spruce budworm. There are five viruses which will infect this species, anuclear-polyhedrosis virus, two granulosis viruses, a cytoplasmic-polyhedrosis virus and an entomopox virus. Short-term control of thispest can be obtained with nuclear-polyhedrosis virus but this approachis very expensive. Aerial spray trials are now in their second yearof study and, although results are encouraging, it is not yet possibleto evaluate the full impact of the long-term effects. Progress ininsect virus research is slow owing to the lack of research scientistsworking in this field. There are about 12 insect virologists in thewhole of Canada.

Summary

Insect viruses have great potential in the control of forestinsect pests. Their value lies in the fact that they are part of thenatural environment. They are highly specific and with correct manipulation they can reduce the abundance of a pest species with minimumdisturbance to the ecosystem. In contrast to insecticides, they aresafe to handle, and have no harmful side effects. To date there is norecord of any insect species becoming resistant to a virus.

When correctly applied some insect viruses can give biologicalcontrol in the true sense of the word. Once an epizootic is initiatedviruses may persist from one year to the next and spread from thepoint of application. A few viruses can be produced in insects in thefield at a cost which is competitive with that of chemical insecticides,Although laboratory-produced viruses used in a primary introductionmay be expensive, they are still economically attractive if long-termcontrol is obtained.

References

Burges, H.D. and N.W. Hussey, Ed. 1971. Microbial control of insectsand mites. Acad. Press, New York. 861 p.

Smith, K.M. 1967. Insect virology. Acad. Press, New York. 256 p.

Steinhaus, E.A., Ed. 1963. Insect pathology: an advanced treatise.Vol. 1. Acad. Press, New York. 661 p.

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Fig. 1 A squash preparation of gut tissue of an eastern hemlock lookerlarva infected with nuclear-polyhedrosis virus observed microscopically underphase contrast. The nuclei of the tracheal cells are packed with viralinclusion bodies. (900X.) p Wltn viralFig. 2. An electron micrograph of sectioned polyhedra in tissue of aWhii-Pvirus rods"? ^ ^ ^f^ W±th -clear-JolyhedrosS viru^f slnglTvirus rods (arrows) are embedded in the polyhedron protein matrix. (62,000X.)

53

ALTERNATIVES IN THE CONTROL OF FOREST INSECTS: FUNGI

Dr. D. Tyrrell and Dr. D. M. MacLeodInsect Pathology Research Institute,Sault Ste. Marie, Ontario

Fungi are probably the best-known organisms causingdisease in insects; dying insects tend to assume conspicuouspositions immediately before death, and subsequently arefrequently covered with fungal mycelium, presenting a somewhat unusual appearance. Species attacking insects belongpredominantly to two main groups of fungi, the Phycomycetesand the Fungi Imperfecti. Those in the former group tendto be more pathogenic and have narrow host specificity; thelatter are much easier to grow in artificial culture. Manyrecorded cases indicate collapse of insect outbreaks bynatural epizootics of entomogenous fungi; much more needsto be known of their physiology and factors governing growthand spread before optimum utility and effectiveness arepossible. Recent developments show promise for fungi- inintegrated insect control in forests and other habitats.

Fungi are a large, heterogenous group of microorganisms,none of which possesses the green colouring matter, chlorophyll, whichis found in most higher plants and in the algae. They range from verysimple, short-lived, single-celled structures whose single cell becomesthe organ of reproduction, to massive perennial mycelia giving rise togreat sporing structures, as in the puffballs, pore fungi, etc.

Because of their lack of chlorophyll, all fungi must obtain theirorganic food from sources external to themselves. Probably the majorityof fungi are saprophytic, i.e., they feed upon the organic products orremains of plants and animals, but not on the living organisms themselves. On the other hand, there are many fungi which secure theirfood ready made at the expense of some other living organism, and theseare termed parasites. Members of this group are among the chief causesof diseases in plants. Others are responsible for a great variety ofdiseases of animals, including insects. It is these latter fungi(which are parasitic on insects) that will be discussed in greaterdetail, since their importance lies in the fact that they help keep thepopulation of their insect hosts in check, and so are a factor in maintaining the biological balance.

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Some insect-infecting fungi, such as the Laboulbeniales, whichoften develop on adult beetles, flies and cockroaches, are aggressiveonly to the extent that they are able to colonize and assimilate nutrients from their hosts. They cannot be considered pathogenic since thisrelationship does not reslilt in death of the insect. Another remarkablegroup, the genus Septobasidium, lives in a mutualistic association withcolonies of scale insects, using some individuals for food and offeringshelter and protection for others. The pathogenic forms, on the otherhand, destroy the host internally, invading the tissues, blocking theblood circulation and finally breaking down and dissolving all the softparts of the insect.

There are 36 fungal genera containing entomogenous species, andof these there are eight in which the truly pathogenic species arefound. These include Beauveria^ Metarrhizium, Paecilomyces, HirsutellaCordycepsy Coelomomyces and Entomophthora and Massospora (Fig. 1-10).

Beauveria species (Fig. 1) appear to be a constant factor in thenatural control of many insect species, and one, Beauveria bassiana, isof interest historically in view of the fact that the Italian biologistBassi isolated and described the fungus from a diseased silkworm in1834. Strains of B. bassiana have been found on about 65 differentinsect species collected in various localities throughout Canada.Because of this wide geographical and host range, B. bassiana has beentested for pathogenicity against more pest insects than any other species. Another Beauveria species, B. tenella9 has also been the subjectof microbial control experiments in Europe against potato beetles.

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The commonest Metarrhizium species is M. anisopliae> the greenmuscardine fungus. It again has a very wide host range, having beenrecorded from over 200 insect species belonging to seven differentorders, of which the Coleoptera, or beetles, are the most common host.It is also second only to Beauveria in the frequency of its use formicrobial control trials.

Paecilomyces (Fig. 4)^ or Isaria as it is also known, andSpicaria are less commonly encountered than the previous two genera,but nevertheless are frequently encountered, as also are Beauveria andMetarrhizium species, as pathogens of soil-inhabiting insect larvae. Wehave recently isolated and cultured Paecilomyces farinosus from nymphsof the cicada Okanagana rimosa taken from an area of reforestation atSearchmont. In certain areas up to 16% of nymphs were killed by thisfungus.

The genus Cordyceps contains almost 140 different species, ofwhich no less than 125 have been recorded as parasitic on insects(Fig. 2). Insects from seven orders as well as spiders have beenreported as being parasitized by these fungi. It is perhaps interestingto note that in certain parts of the world lepidopterous larvae and

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pupae infected with Cordyceps species have long been used for medicinalpurposes and as food. Attempts to use fungi of this genus in controlexperiments date back as far as 1892, while more recently anotherCordyceps species has been isolated from the green-striped forest looperin British Columbia and is currently being field tested as a controlagent for that insect.

Members of the genus Coelomomyces are obligately parasiticaquatic fungi, the majority of which attack mosquitoes. There iscurrently a great amount of interest in members of this genus in view ofthe importance of their host as a disease vector throughout the worldand its increasing resistance to chemical insecticides, and severalcontrol experiments are underway to assess and increase their value asmosquito control agents.

Members of the genus Entomophthora are probably the most pathogenic of all entomogenous fungi. They are widely distributed in natureand approximately 140 different species have been described.

There are many cases on record where, under favourable conditions, epizootics caused by Entomophthora (Fig. 5-10) species havereduced large and destructive outbreaks of insect pests in variouscountries throughout the world. In Canada, Entomophthora species areindigenous and occur on a wide range of insect species. Perhaps one ofthe most dramatic examples was the collapse of the grasshopper invasionof western Canada in the early 1960's when sampling showed that up to95% of grasshoppers were infected with Entomophthora grylli (Fig. 8).Since then the grasshopper population in the western provinces hasremained relatively low until just recently.

In our own district of Algoma we have been able to study severaldifferent epizootics caused by Entomophthora species. Entomophthoraspecies played a large part in terminating the recent outbreak of theforest tent caterpillar in this area. During the last year of thiscaterpillar outbreak, at the height of the epizootic, over 150 deadlarvae were counted on 1 sq. ft of bark surface (Fig. 5) in one area.

Two Entomophthora species, Entomophthora aphidis andEntomophthora fresenii, together form a major controlling factor in theinfestation of red pine plantations by the woolly pine needle aphid. Anextended study of such an epizootic in a local red pine plantation hasyielded much valuable data on the interaction of the host, its parasitesand other environmental parameters such as weather.

In Newfoundland, where the eastern hemlock looper has caused somuch damage in recent years, the outbreak has similarly been brought downprincipally by two Entomophthora species, Entomophthora egressa andEntomophthora sphaerosperma.

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It is obvious, therefore, from the few examples I have justgiven, that we have an impressive list of fungi in our arsenal with somepretty impressive victories to their credit. Indeed, with that kind oftrack record, one might be tempted to assume that all the mycologist hadto do was to put his hand into the bag, select the most likely candidateand the problem would be solved. Unfortunately, this is not so, for anumber of very good reasons.

What, then, are the problems we face, and what are the prospectsfor fungi in pest control? Principally, the problems fall into twogeneral areas: 1) isolation of the fungus in pure culture and preparation of large quantities of the infective phase for dissemination inthe insect population, and 2) what has been called "the conceptualapproach to be used", i.e., whether we should aim for colonization ofthe fungus within the insect population or rely on a microbial insecticide operation.

Let us first consider the problem of isolation and growth ofthe fungus. Most of the imperfect fungi and the Cordyceps species arerelatively easily cultured on artificial medium with no special nutrientrequirements. However, the production of a form of the fungus whichwill withstand the rigours of spraying In the environment while stillremaining infective is more difficult. In this respect recent work bya group of French scientists in developing a large-scale method ofblastospore production of a Beauveria species should go a long waytowards overcoming these difficulties.

With the Entomophthoraceae and Coelomomycetaceae we have thereverse problem. They each form a thick-walled resting spore whichcarries the fungus through the winter and is an ideal candidate forfield dissemination (Fig. 9 and 10). Unfortunately, the Coelomomycesspecies are uniformly obligate parasites, i.e., they have never beengrown apart from their host insects, and a similar situation still holdstrue for many Entomophthoraceae. On the other hand, we have developedways by which considerable amounts of resting spores of the more easilycultured Entomophthora species can be produced, and indeed one worker isnow producing these spores in amounts of up to 10 grams a day. An alternative approach to the problem has also been developed in ourlaboratories. In our attempts to increase the growth rate ofEntomophthora egressa, the fungus which has been very successful in thecontrol of the eastern hemlock looper in Newfoundland as mentionedearlier, we discovered that the fungus will in certain media producefree-living, naked protoplasts. These wall-less protoplasts will growand multiply freely, and can readily be maintained by serial transfer;thus they are unique among fungi. They are also highly pathogenic to avariety of lepidopterous larvae when injected into the insect haemocoele,the insect dying about 5 days after injection. This allows us to do two*things. Either we can bypass the problem of dissemination of the fungus

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in the field by introducing laboratory-reared and infected Insects intothe environment and relying on these insects to introduce the fungusinto the natural insect population, or we can, by rearing the infectedinsects in the laboratory under controlled conditions, persuade thefungus to produce resting spores within the body cavity of the insectfollowing its death, thus building up a stock of resting spores forfuture dissemination from a fungus which will not produce these sporeson artificial media.

The second problem referred to a few moments ago is the typeof approach to be adopted. Basically, three different ways in whichfungi may be used in insect control are available to us. The first maybe referred to as the colonization approach. In this case the fungalagent is introduced into the pest population and natural disseminationand multiplication of the fungus are relied on to bring the insect downto endemic or economically acceptable levels. This is in essenceduplicating what happens in nature in the examples given earlier in thisdiscussion. In order that this approach may be successful, informationis badly needed in the following areas. Naturally occurring epizooticsmust be critically analyzed to define the essential factors that initiateand promote these disease outbreaks. In this respect our work at theInstitute on epizootics of Entomophthora on the woolly pine needle aphidand the forest tent caterpillar, and of Massospora species on the cicadaOkanagana rimosa has provided much definite information. The majorcomponents in determining the occurrence of an epizootic, despite theimportance of climate, are host density and fungal inoculum density.The number of colonization points needed in relation to host density andarea size, methods which enhance the rapid spread of fungi over a largearea and the development of fungal strains with high virulence, arealso areas in which more information is needed before we can hope forsuccess in this direction.

This method holds particular promise in the application offungi to the problem of pest control, especially in the forest, wherethe ability of a pathogen to multiply and spread from its point ofintroduction is of paramount importance in view of the sheer physicaland economic impossibility of treating vast areas of forest.

The other two approaches, i.e., repeated application of thefungus after the fashion of an insecticide, where initial high mortalityrather than gradually increasing mortality is the aim, and the combineduse of fungi and insecticides, are at the moment less promising, butfurther research may alter the situation. They are, however, more suitedto agricultural than to forest ecosystems.

The question of safety is one that deserves consideration in anydiscussion of microbial agents. While no biological control agent canbe considered safe in an absolute sense, experience to date has shownthat the probability of systemic infection of warm-blooded animals by

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insect pathogens is unlikely, because of the demonstrated inability ofthese fungi to grow at mammalian and avian body temperatures. Extensivetesting should be initiated, however, with candidate fungi wheneverlarge-scale dissemination of the fungus is contemplated.

In conclusion, therefore, we may predict that insect controlusing entomogenous fungi will become a practical possibility in thefuture. It is our task to gather the information to make it possibleto realize this potential.

References

Madelin, M.F. 1966. Fungal parasites of insects. Ann. Rev. Entomol._U: 423-448.

Miiller-Kogler, E. 1965. Pilzkrankheiten bei Insekten. Parey, Berlin.

Roberts, D.W. and W.G. Yendol. 1971. Use of fungi for microbial control of insects. In H.D. Burges and N.W. Hussey, Ed.Microbial control of insects and mites, p. 125-149. Acad.Press, London.

Steinhaus, E.A., Ed. 1963. Insect pathology: an advanced treatise.Vol. 2. Acad. Press, New York.

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Figure 1. A mummified larva of Malaoosoma amevicanum (F.) infectedwith a Beauveria strain. Note that the fungus is beginningto protrude from the intersegmental areas. (X 3)

Figure 2. An unidentified scale insect naturally infected withOphiocordyoeps olavulata (Schw.). (X 10.5)

Figure 3. A spruce budworm larva, naturally infected withEirsutella gigantea Petch, showing synnemata of varyinglengths. (X 3)

Figure 4. Paecilomyces farinosus (Dicks, ex Fr.) Brown & Smithemerging from the cocoon of a larch sawfly. (X 3)

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Figure 5. Naturally infected larvae of Malaoosoma disstria Hbn.destroyed by an Entomophthora species (X 0.7)

Figure 6. Actively discharged conidia forming an aureole around anadult Saroophaga aldriehi Park naturally infected withEntomophthora bullata Thaxter ex Povah. (X 2.6)

Figure 7. An adult of S. aldriohi Park naturally infected withE. bullata attached to a twig by rhizoids. (X 8.15)Specimen filled with resting spores.

Figure 8. Melanoplus bivitbatus Say destroyed by Entomophthora grylliFresenius. Several typical characteristics of the diseaseare shown, e.g., congregation in a vertical position nearthe top of the plant stem, clasping, and partial disintegration of some of the specimens. These insects containresting spores rather than conidia. (X 1.5) (Courtesyof CD.A. Research Station, Lethbridge, Alberta).

Figure 9. Cross section of abdomen of naturally infected adult inFig. 7 to show that body cavity is filled with restingspores of the pathogen E. bullata. (X 15.5)

Figure 10. Resting spores of E. bullata taken from the body cavitydepicted in Fig. 9 and shown at a higher magnification.(X 520)