The Chilopoda possess a bewildering array of muscles and no attempt will be made here to catalogue them. Meinert (1883) described the head muscles of Scolopendra and Bekker (1926, 1949) the trunk muscles of a geophilomorph and a scolopendromorph and compared the dorsal musculature of Cryptops and Lithobius. Bücherl (1940) described the musculature of Scolopendra viridicornis Newport, and Jangi (1966) that of Scolopendra morsitans though the trunk musculature was dealt with only briefly. The extrinsic and intrinsic musculature of the antennae of S. morsitans and the musculature of its terminal legs were described by Jangi (1960, 1961). Applegarth (1952) described the musculature of the head of Pseudolithobius megaloporus (Stuxberg), Rilling (1960, 1968) the head and trunk musculature of Lithobius forficatus in great detail and Füller (1963a) the musculature of Scutigera, Necrophloesphagus and Cryptops. Manton (1965) investigated the musculature of members of all five chilopod orders: she found Bekker's descriptions of the musculature of geophilomorphs and scolopendromorphs accurate but she could not entirely confirm his findings on Cryptops. She confirmed Rilling's (1960) observations on Lithobius.

Trunk muscles

The arrangement of the trunk musculature in myriapods has been summarised by Manton (1973). She distinguished six groups of trunk muscles: superficial, dorsal, lateral and sternal longitudinals, deep oblique and deep dorso-ventral muscles (Fig. 46).

European centipedes are usually brown or yellowish in colour but tropical scolopendromorphs show a remarkable range of bright colours, Ethmostigmus trigonopodus (Leach) being almost black with orange legs, Rhysida nuda togoenis deep violet and Asanda sokotrana Pocock red.

The purple pigment of Lithobius

Plateau (1878) noted, that the mandibular glands of Lithobius forficatus were often violet and Duboscq (1898) noted that the connective tissue contained a violet pigment and the blood of this species had a violet tint. The pigment is water soluble and present in granular form in asteroid cells underlying the epidermis and following the tracheal system (Needham, 1960). When Lithobius forficatus is dissected the intensity of the violet colour increases implying that in vivo the pigment is in a colourless, reduced form (Needham, 1945). The reduced pigment is pale yellow. It is rapidly and reversibly reduced by chemical reducing agents, including ascorbic acid, but remains oxidised down to very low oxygen tensions (Needham, 1958).

The purified pigment has a molecular weight of about 4000 and contains no detectable copper so that the chromophore is not a copper-protein or biuret type compound as Needham (1960) had suggested. The absorption spectrum, the nature of the redox and pH colour changes and some other factors suggest that the pigment may be a hydroxyquinone (Bannister & Needham, 1971). For a quinone to be so soluble in water and so insoluble in all lipid solvents tested, as is this pigment, it would have to be very firmly bound to some very hydrophilic conjugant.

Opinions differ as to the correct classification of the Chilopoda. A simple classification is given here to show the family and order to which the genera mentioned in the text belong. All families are listed but not all subfamilies.

Subclass Epimorpha

Order Geophilomorpha (about 1000 species)

The classification of the Geophilomorpha given here is based on that of Attems (1929) with modifications based on Crabill (1960c, 1970). Attems distinguished ten families, namely the Himantariidae, Schendylidae, Oryidae, Mecistocephalidae, Geophilidae, Soniphilidae, Gonibregmatide, Sogonidae, Neogeophilidae and Azgethidae. Within the Geophilidae he distinguished the subfamilies Geophilinae, Dignathodontinae, Pachymerinae, Chilenophilinae and Aphilodontinae. Crabill (1960c) showed that Azygethus atopus Chamberlin for which the family Azygethidae was created is Orphnaeus brevilabiatus and thus the Azygethidae is a junior synonym of Oryidae. Crabill (1970) added a new family, the Eriphantidae and proposed several changes in the classification which are adopted here. The data on distribution are largely from Attems (1929).

Family Himantariidae

Family Schendylidae

Family Oryidae

Family Mecistocephalidae

The internal organs of all centipedes except the Scutigermorpha are supplied with oxygen by tracheae, spirally thickened chitinous tubules of ectodermal origin, which originate from laterally placed openings, the spiracles. In the Scutigeromorpha the spiracles are situated dorsally on the tergites and open into ‘tracheal lungs’. Manton (1965) suggested that primitively each lateral spiracle may have had branching tracheae supplying its own segment, the head deriving its tracheal supply from the anterior pair of trunk spiracles. In addition a third, essentially pericardial respiratory system may also have been present, possessing a mid-dorsal spiracle from which tracheae extended into the pericardium dorsal and lateral to the heart and from which the scutigeromorph system evolved.

The terminology relating to the structure of the lateral spiracles has become very confused. The word spiracle (Stigma of German authors) will here be used for external openings of the tracheal system. The spiracle is often surrounded by a sclerotised rim or peritrema (Stigmaring) and leads into the spiracle cup or atrium whose wall is usually sculptured into trichomes, otherwise termed tubercles, pillars or cuticular lappets (Fig. 128). The tracheae, characterised by spiral thickenings (taenidia) may open directly into the atrium or, as in many geophilomorphs, into an inner atrial or substigmatic pocket by a slit (Stigmamund). In Scolopendridae the atrium is subdivided horizontally by flaps (valves) or a diaphragm.

Geophilomorpha

Spiracles

The spiracles of the Geophilomorpha are borne laterally on the stigmatopleurites of all leg-bearing segments except the first and last.

This book is an attempt to bring together information on centipedes hitherto to be found only in old texts or scattered through the literature and in unpublished theses.

My interest in the group is due to Sir Eric Smith, FRS who, as Professor J. E. Smith, suggested that I carry out postgraduate research on littoral centipedes. Since that time I have had a great deal of advice and encouragement from Mr. J. Gordon Blower, Dr E. H. Eason, Dr R. E. Crabill, Jr and Professor J. L. Cloudsley-Thompson. Over the years, I have received much help from members of the Arachnid Section of the British Museum (Natural History) most notably, Mr K. H. Hyatt, Mr D. Macfarlane, Mr F. R. Wanless and the late Mr D. J. Clark and also from the library staff of the Museum.

The Headmaster and Governors of Dover College generously allowed me a term's leave of absence to take up a School Teacher Fellow-Commonership at Sidney Sussex College, Cambridge, in order to complete the book. I am greatly indebted to the Master and Fellows of Sidney Sussex for their help and hospitality and to the librarians of the Cambridge Zoology Department Library for their assistance. I am grateful to Dr D. L. Gunn, CBE and the Royal Society Research in Schools Committee for their support during the preparation of this book.

Geophilomorpha

Secondary sexual characters

In most geophilomorphs the number of pairs of legs, which is always odd, varies and where this is so, the females tend to have a greater number of pairs of legs than the males, for example, in British Schendyla nemorensis there are 37–41 pairs of legs in males and 39–43 pairs in females. In the larger species Haplophilus subterraneus, there are 77–81 pairs of legs in males and 79–83 in females (Eason, 1964). In many species of the family Mecistocephalidae, however, the number of legs does not vary and is the same in both sexes, for example, 49 pairs in Mecistocephalus insularis (H. Lucas) from Africa and India and 51 pairs in M. evansi Brölemann from Iran. Analysis of broods of Henia illyrica (Meinert), Pachymerium fernigineum, Strigamia acuminata and S. crassipes has shown that female larvae have the same number of legs as the mother, male larvae two pairs less (Prunescu & Capuşe, 1972).

In many geophilids, for example species of Chaetechelyne, Strigamia and Geophilus the last pair of walking legs in males (Fig. 14) is tumescent and much more setose than that of the female which more closely resembles the normal walking legs. In some schendylids, for example Hydroschendyla submarina the last pair of legs are swollen and densely setose in both sexes. The setae are presumably sensory and the enlarged legs presumably contain glands but these have yet to be described.

introduction

Many technical advances have been made and new techniques developed in recent years in the field of microscopy, especially electron microscopy (EM), resulting in a tremendous expansion of our knowledge of cell substructure and function. Examples are freeze-cleave technique and improved methods for observing cells with scanning electron microscopy (SEM). Unfortunately, these new techniques have often been underutilized or sometimes ignored in the study of hemocyte structure and function. In this chapter I call attention to some of these new techniques and their essential features that should be applicable to future research on hemocytes, especially membrane studies. A comprehensive review of theory and practice of these new methods is beyond the scope of this book, and the reader is referred to the text and list of references for some of the many excellent works that cover these topics in depth.

Techniques for observing live hemocyte activities

Insect hemocytes are notorious for their ability to lyse and coagulate soon after withdrawal from the host insect. However, it is still possible to view live hemocyte activities in fresh slide preparations (Baerwald and Boush, 1971) and in tissue culture (see Chapter 9). Under ideal optical conditions, when a thin polished glass slide, ultrathin coverslip, and thin (25 μm) tissue spacers are used, a surprising amount of cell detail can be observed and photographed (Baerwald and Boush, 1971) in those hemocytes that attach to the glass and spread out completely (Fig. 21.1). Motile hemocytes do not reveal nearly as much detail as sessile hemocytes that spread completely.

Introduction

Histochemical techniques are designed to give information about the chemical composition of tissues. There is a vast range of techniques that can be used for the location and identification of nucleic acids, carbohydrates, lipids, proteins, enzymes, and some inorganic ions. The subject is complex, and there are many pitfalls for the inexpert. For many biologists, to do some histochemistry is merely to follow the procedures, as outlined, at the end of a histochemistry textbook–it all seems so simple! But many procedures are inadequately described; often, for example, no source is given for the vital reagent that it is essential to purchase from a particular supplier, and the interpretation of a positive result may be described so categorically as to be misleading or only half true.

The techniques that have been most frequently used on smears of insect hemocytes are primarily those for localizing carbohydrates and lipids, as the main purpose has been the identification of the chemical nature of the many granules and droplets that can be seen in certain types of hemocytes in histological preparations or by phase-contrast microscopy. For this reason, I shall concentrate on these methods in this chapter.

Fixation

An important prerequisite for the majority of histochemical tests is the fixation of the tissue. In general, only proteins are completely fixed, i.e., rendered insoluble and resistant to subsequent procedures, by the commonly used fixatives. Carbohydrates are not fixed, but are retained in the tissues by virtue of the enmeshing proteins. Lipids, in general, are not fixed, and most are still readily soluble in alcohols and other organic solvents.

Introduction

It seems strange that after almost a century of research in insect hemocytology, there is still no consensus on one of its most basic questions: Do the different forms of cells we find in the blood represent transient morphological variants of one kind of cell or constitute a number of distinctive and immutable types? If our indecision on this point seems strange, perhaps it can be forgiven if we consider that the answer requires at least some knowledge of the origins and biology of the hemocytes in a wide range of insect species. Such knowledge cannot yet be claimed, for in only a few species among about 100 genera have even superficial studies been made, and in only about 20 others has the research continued in any depth. Surprisingly, almost the entire body of research on insect hemocytes is concentrated among only 6 orders (Coleoptera, Dictyoptera, Diptera, Hemiptera, Lepidoptera, and Orthoptera) of the 25 or more in the class, and there are widely varied opinions on the status of hemocyte types in any one of them. Little wonder, then, that the subject is a contentious one.

Although this question cannot be answered to everyone's satisfaction even now, it can be examined fairly in the light of recent observations and with an attempt to eliminate ambiguities and misinterpretations that are perpetuated in the literature. This will be the approach taken here.

The status of the hemocyte types is inseparable from hemocyte classification and is bound to studies of the nature of the cells. Unfortunately, hemocyte classification is constantly in disrepute.

Introduction

The literature on the coagulocyte (CO) has been reviewed in several papers (Grégoire, 1951, 1964, 1970, 1971, 1974; Hinton, 1954; Wigglesworth, 1959; Jones, 1962, 1964; Arnold, 1974; Ratcliffe and Price, 1974). The most recent review is by Crossley (1975).

The CO is a newcomer in the considerable and highly controversial literature on insect hemocytes. Mentioned without details by Yeager and Knight (1933) in the hemolymph of Belostoma fluminea, this labile hemocyte has been missed in works in which old, classic methods of fixation and staining were used. The phase-contrast microscope (PCM) made it possible to observe in thin films of hemolymph the specificity of certain hemocytes in initiating plasma coagulation in the form of circular islets around themselves (Grégoire and Florkin, 1950). We named these cells coagulocytes; they were called cystocytes by Jones (1954, 1962). Further PCM studies in vitro on the hemolymph of about 1,600 species of Palearctic, African, and neotropical arthropods (Grégoire, 1951, 1955a,b, 1957, 1959a,b, and unpub. observ.) led to the recognition of four patterns of CO reactions in different taxonomic groups. Hemocyte studies by transmission electron microscopy (TEM) (especially those of Marschall, 1966; Hoffmann and Stoeckel, 1968; Stang-Voss, 1970; Moran, 1971; Scharrer, 1972; Ratcliffe and Price, 1974; Goffinet and Gregoire, 1975) furnished information on the ultrastructure of the cell components, but also opened new controversies. As pointed out by Ratcliffe and Price (1974), without integrated light and electron microscopic studies, correlation of results obtained by the two methods (PCM and TEM) is difficult or impossible.

Introduction

Hemocytes of arthropods and several other invertebrate groups have been studied (see Gupta, 1979). Among arthropods, they have been most extensively studied in insects, followed by crustaceans, arachnids, and myriapods. Hemocytes of a few onychophorans also have been described. It is not surprising, therefore, that the need for a reliable, uniform classification of various hemocyte types has been felt more keenly by insect hematologists than by those of other arthropod groups. Fortunately, a generally acceptable hemocyte classification in insects, based largely on morphological characteristics, now exists.

Hemocyte classifications both in insects and other arthropods have been variously based on morphology, functions, and staining or histochemical reactions of hemocytes. Thus, it is not unusual to find the same hemocyte type or its various forms being referred to by different names in various arthropods, by different authors – a situation that has inevitably resulted in a confusing mass of terminology. Consequently, it becomes very difficult to compare hemocytes of one species with those of others. This has particularly hindered any phylogenetic consideration of the evolution of hemocyte types in various arthropod groups and the Onychophora. Clearly, there is a need for a uniform hemocyte classification for insects as well as other arthropod groups. The insect hemocyte classification that is generally used has evolved over more than half a century. According to Millara (1947), Cuenot (1896) was the first to classify insect hemocytes into four categories and was later followed in this attempt by Hollande (1909, 1911) and others. Wigglesworth (1939) summarized most of the earlier classifications and presented a classification that was widely accepted. He modified it later (Wigglesworth, 1959).

Introduction

Although embryonic hemocytes, or blood cells as previously termed, have been described in papers dealing with organogenesis, the available information is fragmentary. The origin of hemocytes has been more or less closely observed, but the cytological features of hemocytes have not been adequately described. Furthermore, the development of hemocytes has hardly been noted. Functions and activities of embryonic hemocytes have been neglected almost completely.

Perhaps the reason why insect embryologists have not given much attention to embryonic hemocytes is because the role of these free cells in embryogenesis is quite obscure. No direct participation of hemocytes in the formation of any of the embryonic organs seems to have been observed. Embryologists have been devoting much attention to the details of midgut epithelium formation, because this elucidates the mode of entoderm formation. The metameric segmentation of the embryos also has been closely followed to discover the number of segments in the insect head. However, the unique nature of the embryonic hemocytes seems to have completely escaped the attention of embryologists.

The embryonic hemocytes are unique among embryonic cells because these cells are single and may circulate in the embryo even during the final phase of embryogenesis; they may move by themselves or be moved by the hemolymph. As embryogenesis proceeds, morphogenetic movement of each cell becomes slow, and the topographical positions of various cells in the embryo become gradually fixed. For example, the drastic shift in the positions of cells seen at the stage of germ band formation seems to be lost when the embryo is very close to hatching.

Introduction

Over three-quarters of animal species known to man are insects, and it has been estimated that there may be as many as 3 million different species (Williams, 1960) living in almost every habitat on earth. During their evolution and colonization of the terrestrial niches, the insects became open to attack from a formidable array of predators, parasites, parasitoids, and microorganisms, and recent research indicates that they have effective defense mechanisms capable of resisting, containing, and even eliminating many would-be invaders (Whitcomb et al., 1974). These defenses include the physicochemical barriers formed by the tough outer cuticle and gut and the internal humoral and cellular defense mechanisms. The cellular defenses are mainly mediated by the hemocytes and include the processes of phagocytosis, nodule formation, encapsulation, and hemolymph coagulation (Salt, 1970), which together form the main subject of this review. However, because most of our knowledge of insect defense mechanisms has been derived from laboratory investigations and little is known about the effectiveness of these defenses in the field, brief consideration must also be given to the natural biological control agents themselves, their mode of entry into the host, and the type of host response most likely to be elicited. This approach, it is hoped, will give the results of laboratory experiments a new dimension and prompt future investigators to accord more consideration to the defense mechanisms of natural insect populations.

Diseases of insects

These are produced by the pathogenic viruses, bacteria, fungi, protozoa, rickettsiae, and nematodes. In addition, predatory and parasitic insects are of enormous significance in controlling vast numbers of pest and potentially injurious insect species (van den Bosch and Messenger, 1973) (see also Chapter 16).

Introduction

The blood cells of many higher animals are formed in specialized tissues called hemocytopoietic tissues (blood-cell-making tissues; poiesis = “making”) or hemopoietic tissues. Although less accurate, the latter term is generally used to mean the same thing as hemocytopoietic tissues and will be preferred in this chapter.

Hemopoietic tissues are concerned with producing different lines of blood cells and delivering them into the blood; in some animals they may also function to remove dying or dead cells and histolytic debris from circulation. This function is assumed by phagocytic cells that originate from a particular cell lineage, generally called reticular or reticuloendothelial cells; it is this cell type that gives to hemopoietic tissues a characteristic reticular arrangement. It must be stressed that a third major function of hemopoietic tissues in higher animals may be immunologic, involving specialized cells such as lymphocytes and plasmocytes.

That in at least some insects definite hemopoietic tissues are present was reported near the end of the last century (Kowalevsky, 1889). However, discrepancies obviously exist between the organizational level of hemopoietic tissues (or their equivalents) in different insect orders; in addition, this organization may change during the development in the same species, as will be explained for Calliphora.

These differences are the major source of the controversies about the postembryonic origin of hemocytes that existed between Kowalevsky (1889) and Cuénot (1896). Another source is the lack of quantitative data.

Jones (1970) published an exhaustive report on these controversies (250 references), and it is unnecessary to duplicate such a review here (see also Chapter 3).

Introduction

The purpose of this chapter is to review the contribution of the hemocytes to the processes, and particularly to the control, of insect growth and metamorphosis. But there are certain ancillary matters that must be briefly considered.

Hemocytes as a source of “embryonic cells”

Hemocytes arise as residual mesodermal cells at the conclusion of embryonic growth. Berlese (1900, 1901) regarded their differentiation to form muscles and other mesodermal tissues as being their main function during postembryonic growth in higher Diptera. Likewise, in Galerucella (Col.: Chrysomelidae), Poyarkoff (1910) included the imaginal myoblasts among the hemocytes. Nowadays, although embryonic cells (notably myoblasts, derived from imaginal histoblasts) may be moving freely in the hemocoel at certain stages in development, it is usual not to include them among the hemocytes (Crossley, 1964, 1975). However, Penzlin (1963), studying regeneration of limbs in Periplaneta, concluded that the new muscles are formed by blastema cells derived from the hemocytes, not from the epidermis. And there are many claims for fat body formation during postembryonic development from “mesodermal leucocytes” (see Wigglesworth, 1959, and Chapter 1).

Hemocyte types

There is a vast descriptive literature on the varied types of hemocytes in different groups of insects (Wigglesworth, 1959; Jones, 1962; Gupta, 1969; Crossley, 1975). In recent years the tendency has been to simplify these descriptions and to put the emphasis on a limited number of more or less well-defined types. It is hoped that this procedure will lead to the recognition of true homologies between blood cells of different insects, with the allocation of definite functions to common types of cells.

Introduction

The postembryonic origin and multiplication of insect hemocytes continue to provide food for thought and material for research projects for many entomologists, comparative physiologists, and comparative pathologists. Review articles discussing insect hemocytes, total and differential cell counts, and hemopoiesis include Jones (1962, 1970), Wigglesworth (1959), Arnold (1974), and Crossley (1975). One should consult these articles for the historical development of the topic as well as for an extensive review of the literature.

Because the terms “hemopoiesis,” “hematopoiesis,” and “hemocytopoiesis” seemed to be used interchangeably in the literature, I decided to do a little checking before using one term or the other. According to Webster's Third New International Dictionary (1965) “hemopoiesis” and “hematopoiesis” are equivalent in meaning and in derivation and therefore can be used interchangeably. Both mean formation of blood or blood cells within the body. Webster does not list “hemocytopoiesis,” but does list “hemocytogenesis” as being the part of hematopoiesis concerned with the formation of blood cells. I will use “hemopoiesis” because it is the shortest of the terms and includes the formation of blood cells in its definition (see also Chapter 10).

The process of hemopoiesis includes both cell proliferation and cell differentiation. As cell differentiation proceeds, the capacity for cell proliferation decreases. Mature cells are incapable of proliferation (Best and Taylor, 1966). Even though I will be principally concerned with cell proliferation and multiplication in this chapter, I will also cover some aspects of cell differentiation because it bears an important relationship to the amount of cell proliferation in the normal animal.