Caste Differentiation in Social Insects

Science

Section A

In Memoriam

CHAPTER 1

In Memoriam Martin Lüscher (1917–1979): His Contributions to Insect Physiology and Sociobiology

J. DE WILDE*,     Department of Entomology, Agricultural University, Wageningen, The Netherlands

Publisher Summary

This chapter presents the scientific work of Martin Luscher and his contribution to insect physiology and sociobiology. It discusses his contributions to the knowledge of polymorphism and polyethism in termites and the honeybee. It also discusses his work on communication within the termite colony and its importance to the regulation of caste development. The chapter additionally focuses on the significance of the grassland termite programme of the International Centre of Insect Physiology and Ecology and its impact on the knowledge of foraging, nutrition, building activity and the regulation of caste development in the higher termites. It also reviews the research of Luscher and his co-workers on the endocrine regulation of growth and reproduction in the Blattidae and its importance for the understanding of similar processes in termites is stressed.

CONTENTS

1.1 Introduction

1.2 Earlier Works

1.3 Termite Biology

1.3.1 Communication—the pheromone concept

1.3.2 Caste determination

1.3.3 Functions of termite colonies

1.3.4 The ICIPE programme on grassland termites

1.4 Endocrine Regulation in Blattidae

1.5 Honey Bee Worker Functions—Their Endocrine Control

1.6 An Evaluation of Lüscher’s Contributions to Insect Physiology, Especially Regarding Social Insects

1.7 Summary

1.8 References

1.1 INTRODUCTION

It is certainly fitting that, on an evening during our symposium, we sit together and commemorate Professor Martin Lüscher, whose untimely death on 9 August 1979, at the age of 62, has been such a shock to his relatives and friends, but no less to the International Community of Insect Physiologists.

He was at the height of his performance, sometimes attending two, three or more International Meetings, Symposia or Congresses, during one summer, releasing a constant flow of publications, and at the same time preparing for a more quiet phase of his life, when more and more of his work would be shifted

from the laboratory to his home. He had gradually surrounded himself with a number of collaborators who had grown to the level of independent and internationally renowned scientists.

Had his institute been built out to a size to accommodate several chairs, several professors of animal physiology and behaviour would now have continued the work he had initiated; but as it was, until the last day, he had to be satisfied with a minimum of staff and administrative help.

Nevertheless, it would seem to the outer world that the flow of scientific output by this group could only be performed by a larger personnel and sophisticated housing provisions, so varied was his approach in concept as well as in technology. But Lüscher was far from being an empire builder. His belief was in research rather than administration, and when technology was not available at his institute, he preferred to seek cooperation with specialized laboratories elsewhere. And in his thematical approach he found constant inspiration, as Leuthold (1980) has stated, in the awe and respect at the incomprehensible and beautiful features of the organisms of his study. Those were mainly termites, and later on he extended his interests to honey bees.

1.2 EARLIER WORKS

It would certainly not be due to dwell in my survey on the details of Martin Lüscher’s personal life and his development as a biologist, as so much more qualified accounts have been given by Mrs Lüscher (1980) and by Leuthold (1980). I may only state that his Ph.D. in Zoology was guided by Rudolf Geigy in Basel. In his thesis he discussed his experiments on the determination of larval and adult features in the cloth moth, Tineola biselliella. This study of developmental physiology was certainly an excellent way to be introduced into the problems of insect differentiation and development, a field in which Lüscher remained interested for the rest of his life.

It may be remembered that Rudolf Geigy (1931), who had an unconventional approach to the study of insect metamorphosis, had been able to demonstrate that in Drosophila, localized irradiation of the cleaving egg, results in either larval or adult defects, depending on the time of treatment. Lüscher (1944) confirmed these data for Tineola, and thereby showed that the early programming of larval and adult features is of wider application within the domain of the Holometabola.

As an assistant of Lehmann in Berne, he successively worked on topics of amphibian developmental physiology, and especially regeneration, a subject he continued to study during his stay with Wiggleworth in Cambridge, England, in the years following the end of the Second World War. But this time the experimental animal was Rhodnius prolixus (Lüscher, 1948) and its study marked the beginning of a long-lasting scientific relation and friendship with the nestor of Insect Physiology, whose standard treatise, The Principles of Insect Physiology, he later translated into German.

1.3 TERMITE BIOLOGY

1.3.1 Communication—the pheromone concept

Following the stay in Cambridge, Lüscher went to Paris where a sojourn at the Laboratoire d’Evolution des Etres Organisés with P. P. Grassé introduced him into the domain of the termites. Grassé and Noirot (1946a, b) had started work on social polymorphism in the colonies of the European termite Kalotermes flavicollis, and had developed some interesting hypotheses on the way castes were determined. Lüscher immediately recognized that the numerical regulation of caste individuals required an extensive system of communication and feedback mechanisms and, as optical stimuli were excluded, the probability of chemosensory communication was evident.

This was in contrast with the prevailing conceptions centring around the state of nutrition, or around the so called group effects of a hypothetical nature (Grassé, 1949). The most likely mechanism, best supported by experimental evidence, is the effect of token substances released by the differentiated castes, which have been referred to in literature as ectohor–mones, sociohormones, and, most recently by the more generally-accepted term of pheromones, proposed by Karlson & Lüscher (1959).

Extending the experiment of Light (1944) with Zootermopsis, Lüscher could show that in Kalotermes, the inhibitory pheromone of the primary sexuals, preventing the formation of replacement reproductives, is released by the anus. He established by further experiments that the larvae pass on the pheromone by oral intake and anal release, in the frame of proctodaeal trophallaxis common in termites (Lüscher, 1955a, b).

The inhibitory effect of reproductives on the development of their own castes finds a parallel in the development of soldiers.

Stimulating tokens are also present. In Kalotermes, male sexuals stimulate the transformation of female larvae into replacement reproductives, and this effect can even be obtained by extracts of the heads of functioning males (Lüscher, 1964). Together with Springhetti (1970), Lüscher could demonstrate that in the same species, reproductives activate the formation of soldiers.

The isolation and identification of the various pheromones instrumental in caste regulation of termites is certainly a challenging task for future workers. The regulative effects I discussed were established in the lower termites, but recent observations on the higher termites point to similar mechanisms. With Macrotermes michaelseni, in the framework of the termite programme of ICIPE, Lüscher tried to test the inhibitory effects of the royal pair on nymph formation. But these effects may be somewhat obscured by the huge size of the colonies (Lüscher, 1976).

1.3.2 Caste determination

Caste differentiation depends on specific trigger stimuli acting during sensitive periods. Their relation to the determination of patterns is not a direct one. Reprogramming is part of ontogenetic determination, leading to the development of caste features. According to Hadorn (1967) determination is the programming of the developmental potential by activation of specific groups of genes. Juvenile hormone (JH) is at the base of this control in caste polymorphism. Extrinsic control of caste differentiation is mediated therefore, through the environmental impact of JH levels. Much of the above statement is based on work done by Lüscher and his co-workers, and although recent work on social Hymenoptera was necessary to allow for generalization of the mechanism, Lüscher’s work on Kalotermes has provided the primary impetus (Lüscher, 1974a).

It also gave a physiological explanation for the phenomenon of competence. It had struck the investigators that the larvae or nymphs of Kalotermes are not always able to develop, under the proper stimulations, into replacement reproductives, but can only do so during early spring and summer. The volume of their corpora allata (CA) is much reduced during these periods, and Lüscher (1974b) could show that a low CA activity is a requirement for competence. After competence has been obtained and the adult moult is induced, the CA increase in size, which is related to their activity in reproduction.

With soldier formation it is different. Larvae and nymphs can be induced to change into presoldiers by increasing their JH levels. Competence is generally prevailing, and is only suppressed when soldiers are already abundant. After presoldiers are formed, they automatically moult into soldiers, a caste with reduced CA activity.

I only briefly relate Lüscher’s experiments with prothoracic glands and their hormone, ecdysone; they did not reveal any specific effect on caste induction, but merely interfered with the time of moulting. They thereby sometimes interfered with the above-mentioned effects of JH, in an indirect manner.

As regards the higher termites, the ICIPE Termite Programme has marked a period rich in new and interesting findings, though sometimes baffling and at the same time puzzling.

I omitted in my former discussion the ingenious techniques developed by Lüscher for rearing termite colonies in captivity in such a way that their functions were open to biological observation and experiment. This was already a feature of his earliest work with Kalotermes, but became even more essential in his studies of Macrotermes, where huge mounds and stone-hard building elements are prohibitive of any detailed work on caste determination and behaviour.

Bühlman (1977a, b) has been instrumental in solving the essential problems and devising the in vitro culture of microcolonies showing all the essential features of the natural colony.

A first and all-important question was, whether caste determination in the higher termites is blastogenic, i.e. already prevailing in the egg stage.

Termite eggs are rich in JH, but their content is varying throughout the seasons. These fluctuations are paralleled by the haemolymph JH titre of the physogastric queen varying more than tenfold in the course of the year. Such queens have multilobed CA, divaricating in the cavities of neck and thorax.

Caste formation in the higher termites is much more rigid than in the lower groups, and regressive moults are absent. In some cases it has been established that the decision to develop into one of the neuter forms (soldier, worker) occurs in the first larval instar.

Lüscher (1976) initiated research to test several hypotheses with respect to the relation between the JH content of the egg and the formation of nymphs in Macrotermes. His aim was to decide whether or not, as suggested by some authors, the induction of nymphs would already occur during embryonic development, a process known as blastogenic caste formation. With a great deal of care he started collecting data on JH content of the egg following a seasonal pattern; he passed away from life before this work could be completed.

It has to be understood that in this line of thought, endocrine induction of blastogenic caste formation would be similar to caste determining endocrine induction in a growing larva. This comparison is purely speculative and a complete fixation of a developmental program as is present in holometabolous insects has never been demonstrated in termites. In fact, even in the social Hymenoptera, caste induction takes place in the course of larval life (de Wilde & Beetsma, 1982). In higher termites, deviation from the developmental programme is observed under strenuous conditions such as removal of reproductives (Bordereau, 1975). Also, there are several ways in which a first instar larva can still increase JH content, and already by this fact we may conclude that blastogenic caste determination in termites has not been proven.

Beautiful additions to our knowledge of endocrine caste determination have been given by some of Lüscher’s African students. Wanyonyi (1974), whose untimely death has been such a loss, was able to demonstrate that external application of increasing doses of JH and a JH analogue to larvae of Zootermopsis subsequently leads to the inhibition of reproductive development, to regressive development, and to presoldiers and worker-like forms. We are now waiting for the determination of JH titres in normal development of these castes. During this Symposium, another African student, Dr Okot-Kotber has presented more evidence on the higher termite, Macrotermes michaelseni.

1.3.3 Functions of termite colonies

It has been said that a colony of social insects is in a sense a superorganism. In these creatures, the physiological functions of the individual are paralleled by social functions. During his stay in Ivory Coast, on one of the foreign visits he made before settling in Berne, Lüscher became acquainted with the elaborate nest structures built by Macrotermes natalensis. In the chimneys and inner channels of the termite mound he recognized a system of ventilation with regulation of oxygen tension, temperature and humidity, functioning through the temperature gradient at the outline of the nest (Lüscher, 1955a, b). But the Termite Programme he developed with Sands at ICIPE provided further opportunities, and foraging and nest building were two of the fields in which outstanding progress was made.

Grassland termites were found to compete with cattle or wild herbivores, and food intake by a full-sized colony of Macrotermes michaelseni was found to be equivalent to that of a large antelope. Leuthold and his students found that harvester termites orientate their way home by means of their optical sense (Leuthold et al., 1976).

Building behaviour was studied under Lüscher’s guidance by Bruinsma, and very remarkable facts were found, especially in the construction of pillars and queen cells. This work suggests that the fatbody of a termite queen is in fact functioning as a pheromone gland, the building pheromone being released through the abdominal stigmata. A gradient is thereby created at which the building termites measure the distance at which the cell wall has to be constructed, with respect to the body of the queen (Bruinsma & Leuthold, 1977).

I could go on entertaining you with the remarkable outcome of their observations, made possible by Bühlmann’s microcolony rearing method mentioned before.

1.3.4 The ICIPE programme on grassland termites

It was in the beginning of 1972 that some members of the ICIPE Board were meeting in Oxford with John Pringle to evaluate research proposals, and among them a rather elaborate one by Martin Lüscher, to be supported by the Swiss National Science Foundation Fund to study communication and caste development in grassland termites. It was clear right from the first moment that the new Director of Research had rather strict financial and other requirements, and we were worried whether he was planning to go his own way or was prepared to integrate his work in the kind of scientific developmental aid which we had in mind when founding ICIPE.

It has been our lucky fate that we decided to go along with Lüscher. Though strictly adhering to his principles of responsibility in guiding the research, he has been collaborating with the Board in an excellent way, and has managed to extend the termite work both in the physiological and the ecological sphere. Sharing the directorship of research with a termite ecologist such as Sands and assisted by a behaviour specialist such as Leuthold, he was able to attract very competent people and at the same time to do a considerable job in training young African biologists in the painstaking research he had developed. This group has been one of the best organized at ICIPE, and his critical attitude has resulted in a very high quality of work. For example, Darlington’s and Lapage’s contributions towards the understanding of termite populations and their foraging activity in a semi-arid ecosystem (Kajiado, Kenya) have been enormous and unparalleled in this region. Oloo, another young African scientist contributed much towards the understanding of mechanisms of foraging behaviour of grass-feeders, Trinervitermes etc.

Lüscher annually spent several months at ICIPE, and among his efforts is a film on colony life in Macrotermes which has become famous. Due acknowledgement should be made to Mrs Lüscher who has assisted her husband during these stages and shared his work in every respect. When in 1970 Lüscher ended his role as a Director of Research, his task was taken over by Noirot.

During the 7 years, Lüscher has given very dedicated guidance and has obtained very considerable support from outside, to the benefit of ICIPE. It is therefore due that a statue has been erected here to commemorate his great merits to this Institute.

1.4 ENDOCRINE REGULATION IN BLATTIDAE

Termites are uniquely unsuitable for the study of the endocrine regulation of body functions. In isolation they survive for only a short time, and when released in a colony after having been operated on, they are invariably eaten by the other members. Even to fix a termite in a colony for the purpose of individual observation, leads to a considerable mortality. Therefore, Lüscher chose to study endocrine processes in the related group, the Blattidae. For several reasons, this group can be considered to be taxonomically very close to termites. Their individual development, their symbiont-dependent digestive functions, their nocturnal habits with corresponding tegumental features are all in line with this concept. Nauphoeta cinerea and Leucophaea maderae became the species of his study. First in association with Lüscher and Engelmann (1955), later on with Wyss-Huber and Lüscher (1972) and finally with Lüscher and Lanzrein (1976), he embarked on a series of studies which mainly concerned the regulation of CA function and the endocrine control of metabolism, vitellogenesis, and the differential effect of JH in ontogenesis and reproduction.

The fact that three and later even more JH were found to exist, and their presence at various rates in different developmental stages, led Lüscher to investigate whether these hormones had a differential effect on larval development and adult reproduction with some initially positive results, which are now questionable in the light of more recent experience. But very interesting was Lüscher’s finding (Lüscher et al., 1971) that the fatbody of the cockroach after allatectomy starts to synthesize a protein not found under normal conditions. In my laboratory, similar results had been obtained by De Loof in the Colorado beetle, and the term short-day proteins had been applied to denote this category.

In this research, Lüscher profoundly proved to be an insect physiologist. Here he could bring his endocrine work in level with the international progress, and participate in the study of problems belonging to the frontiers of our science. It was especially Lanzrein among his students who gradually carried this line on her own and is now setting forth the rich tradition. Inevitably, the factors regulating the JH titre came under study, and an overall picture was drawn of morphometrical and physiological parameters of the endocrine regulation of oocyte maturation (Lanzrein et al., 1978; Lanzrein et al., 1981). Also, the ecdysteroids and JH present in the egg, and their role in embryogenesis were given due attention (Imboden et al., 1978).

1.5 HONEY BEE WORKER FUNCTIONS—THEIR ENDOCRINE CONTROL

Through his relations with the Swiss Bee Research Station at Liebefeld-Berne, Lüscher became acquainted with social functions in honey bees analogous to those he had studied in termites. Gradually, the termite work more and more enriched the bee research, and this has led to very interesting discoveries. As worker bees are morphologically identical, but show a successive shift in behavioural functions, the term polyethism was introduced, and the physiological states concerned were subjected to endocrine studies. Publication of this work started in 1974.

It so happened that in my department in Wageningen, Wirtz (1973) had shown in his thesis that caste differentiation in the honey bee was based on the haemolymph titre of JH, the sensitive period being at the end of the third day of larval life. Low JH titres result in the development of worker bees, high titres in queen development.

After Lüscher’s findings in termites, this was the second case in which JH was shown to be involved in caste differentiation. But this time the situation was more complicated. After the sensitive period has elapsed, several larval moults take place, and only at the subsequent pupal and adult moult, there are time lapses and caste features showing the fulfilment of the programme. Normally, for the pupal and adult moult to take place, a low JH titre is required. It thus appears that in the honey bees, JH induces programmes comprising various activities of the CA.

During adult life, in the honey bee, as said above, a series of functions is performed, from cell cleaning to foraging. With some variability, this program is carried out in every worker bee and it is this form of age-polyethism that Lüscher with research workers from Liebefeld-Bern such as Rutz, Gerig and Wille, started out to investigate endocrinologically. They thereby found that a high JH titre is responsible for the transition from hive bee to field bee and suppresses the activity of glands characteristic for hive bees, such as the hypopharyngeal glands and the wax glands (Rutz et al., 1977).

Interesting enough, in the Wageningen laboratory the matter was approached from a different direction, but with the same outcome. After discovering the JH-induced queen differentiation, we realized that here a real danger was presented by the juvenoid insecticides we helped to develop for many years. We therefore fed whole colonies with diets containing JH analogues and found that some analogues would not do any harm, but others disorganized the colony, the worker bees ceasing to feed the larvae. This brought us to making independently the same discovery as reported by the Swiss group, providing a beautiful confirmation of this very important effect (Beetsma & ten Houten, 1975).

In subsequent work, the Swiss group found that winter bees, which are in a diapause-like condition, are characterized by a very low JH titre in the hemolymph (Fluri et al., 1977). Further work was on queen pheromones (Lüscher & Walker, 1963), vitellogenic protein synthesis and its dependence on JH, and some nutritive aspects.

1.6 AN EVALUATION OF LÜSCHER’S CONTRIBUTIONS TO INSECT PHYSIOLOGY, ESPECIALLY REGARDING SOCIAL INSECTS

When I try to survey the panorama of Lüscher’s work and Lüscher’s impact on the work of others, my feelings are of respect, understanding and sympathy. I may freely say so, since my own approach to insect physiology has grown from the same sources and in many cases had led to results similar to his own. In the foregoing account the audience has found some examples.

I may therefore state that Lüscher was a typical organismal physiologist, who drew his inspiration from the insect as a whole, and, in his case, from the insect as a member of a society. He had the strength of mind to fix the goals of his research, and to continue his approach for a lifetime.

He thereby has enlightened our picture of communication and caste formation in the considerable complexity of colonies even of the lower termites, and when he passed away, he had already a strong foothold in the higher termites.

If he had lived, promises were that he would have succeeded in unravelling the considerably complicated relations in this group a good deal further.

Lüscher and his group deepened their endocrinological knowledge by the study of Blattidae and subsequently broadened our knowledge on endocrine regulations in termites and their dependence on stimuli inside and outside the colony. In doing so, their contribution to environmental endocrinology has been considerable.

Despite of a great deal of continuity, Lüscher’s approach had much flexibility, and this was especially shown in the way he built out and helped to conduct the termite programme of ICIPE. His attitude was not at all alien to the ecologist and the taxonomist, with whom he shared on the one hand the great inspiration provided by working in an ecosystem and on the other a considerable precision in his performance.

Lüscher has pursued the role of hormones in termite polymorphism and the role of pheromones, of which he was a nominator, in termite communication. He subsequently closed the bridge by showing that pheromones have their impact on termite differentiation via their effect on the endocrine system. In doing so, Lüscher stretched the impact of hormones to the utmost. He showed the vapour tension of JH to have important effects and he tried to study the hypothesis that JH, next to being a hormone, is also a pheromone. It is characteristic of his approach that he also studied unlikely possibilities and unattractive hypothesis, which gave his work a great deal of objectivity and candour. And when the outcome was negative, he drew his conclusions.

In my account, I have purposely concentrated on Lüscher’s share in the modern development of termite biology, and have not named the many important contributions of workers from different nations, without whose impact his work would have been less fruitful or perhaps non-existing. But today I want to put Martin Lüscher in the limelight.

If our handbooks nowadays contain less vague conceptions, less unproven theoretical mechanism, and less incomprehensible relations in the field of termite biology and physiology, this is for a considerable part due to the virtue of Martin Lüscher.

1.7 SUMMARY

An account is given of the scientific work of Martin Lüscher and its importance to insect physiology and sociobiology. Mention is made of his contributions to our knowledge of polymorphism and polyethism in termites and in the honey bee and his work on communication within the termite colony and its importance to the regulation of caste