The Structure and Development of Wax Glandsof Pseudococcus maritimus (Homoptera,

Coccidae).By

Priscilla Frew Pollister,

Department of Zoology, Columbia University.

With Plates 17-20, and 3 Text-figures.

INTRODUCTION.

ONE of the striking features of the family Coccidae is a uni-versal tendency to form extensive secretions of wax or wax-likesubstances on the surface of the body. This substance seems toserve a protective role, but in many instances the amount of itis so out of proportion to the necessity for this purpose thatmany entomologists have held that the exudation of this waxis more accurately to be regarded as a type of excretion of amaterial that is an inevitable by-product of the metabolism ofanimals feeding exclusively on plant juices.

The wax is secreted through pores in the cuticula, which arethe openings from underlying glands. The present study isconcerned with the distribution, minute structure, and develop-ment of these glands in one of the mealy-bugs, P s e u d o c o c c u sm a r i t i m u s Erhorn. There have been many descriptions ofthe structure and distribution of the external pores, and a fewstudies of the histology and development of the underlyingglands. Many of these observations have been incidental toa work primarily concerned with the taxonomy of the group.The present study attempts to deal with the problem primarilyfrom a histological and cytological point of view, since in thesestructures one finds some of the most elaborately differentiatedtypes of glands.

The problem was suggested by Professor Franz Schrader,and I am indebted to him for advice and encouragementthroughout the work.

128 PRISCILLA FREW POLLISTER

MATERIAL AND METHODS.

The original stock used in this investigation was obtainedfrom a commercial greenhouse in Falmouth, Massachusetts,where the mealy-bugs were growing on narcissus bulbs. A stockof these was kept on various bulbs during the winter monthsand on citrus fruits in the summer. The transfer to the fruiteach summer was indispensable to the continued reproductionof the stock. Specimens from the stock were identified asPseudococcus m a r i t i m u s Erh. through the kindness ofDoctor Harold Morrison of the Bureau of Entomology of theUnited States Department of Agriculture.

For the study of the external apertures of the glands pre-parations of the exoskeleton were made by the potash-magentamethod as described by Dietz and Morrison, 1916. The fixationof material for sectioning is difficult because of the covering ofwax, which must be softened and partially removed by touchingthe animals with a brush that has been dipped in absolutealcohol. Immediately after the application of the alcohol theanimals are dropped into the fixing fluid, which mus t behot. Good fixation apparently occurs when the alcohol hasaffected the wax just enough so that the fixing fluid will pene-trate readily, but has not acted so long that the alcohol hasreached the soft tissues of the animal. Many fixing fluids weretried and Allen's B 15 and Kahle's were found to give vastlysuperior results, so that the study was made entirely frommaterial fixed in one or the other of these mixtures. Thematerial was embedded in paraffin and sectioned at 6 or lOmicra,stained with iron haematoxylin, and in some cases counter-stained with either eosin or light green.

OBSERVATIONS.

Types and D i s t r i b u t i o n of Glands.In potash-magenta preparations of adult females of P s e u d o -

coccus m a r i t i m u s one can see hundreds of gland-pores.Each pore is at the top of a small papilla projecting above thelevel of the surrounding cuticula. As seen from above thesederm-pores are of three distinct types, to which, after Ferris

WAX-GLANDS OF PSEUDOCOCCUS 129

(1918), I shall refer as the triangular, cylindrical (or tubular),and the multilocular.

The triangular derm-pores show four openings, three elongatedones that form a triangle, and a fourth circular aperture withinthe triangular (Text-fig. 1 B). These pores are the smallest and

BECX CELL

B

TEXT-FIG. 1.

A, diagrammatic sagittal section of triangular gland. B, pore oftriangular gland as seen from above, c, sagittal section of neckregion of triangular gland drawn to the same scale as B.

most numerous. They are located on dorsal, ventral, and lateralsurfaces of all segments. They are especially concentratedaround the bases of the seventeen pairs of large lateral spines,with which they form the so-called cerarian complexes. Theyform small curled flakes of wax that are scattered over the sur-face of the body and are pushed outward along the large spinesto form the long wax filaments projecting from their tips—a feature very characteristic of the genus P s e u d o c o c e u s .

The tubular pores have a single large opening at the top ofthe papilla (Text-fig. 2). Like the first type, they are verynumerous and are found on all surfaces. They are especiallyabundant on the ventro-lateral parts of the body, and within

NO. 317 K

130 PRISCILLA FREW POLLISTBR

each segment are more numerous at the anterior and posteriorlimits. These glands form large solid cylinders of wax.

The multilocular type of pore is much larger than either ofthe others and consists of a shallow cup at the top of a lowpapilla. The margin of the cup overhangs the cavity, within

TEXT-PIG. 2.

A, diagrammatic sagittal section of tubular gland, B, pore of tubulargland as seen from above, c, sagittal section of neck region oftubular gland drawn to same scale as B.

which is a peripheral circle of ten triangular openings surround-ing a single central aperture. These pores are sometimes calledcircumgenital in allusion to the fact that they are especiallyconcentrated in a ring around the vulva—although they arealso found, in sparser number, on adjacent regions. The multi-locular pores are much less numerous than the other types andare restricted to the anterior and posterior regions of the ventralsurfaces of the last five segments. These glands form irregularmasses of wax. In Pseudococcus c i t r i Matheson (1928)seems to have made out a fibrillar texture to this wax, but sucha finer structure is not identifiable in Pseudococcus mar i -t imus.

WAX-GLANDS OF PSEUDOCOCCUS 131

H i s t o l o g i c a l S t r u c t u r e of t h e Glands .

T r i a n g u l a r Glands.

The triangular type of gland is composed of five cells. Asa whole the gland is roughly pear-shaped, with the narrowstem-like portion ending in a heavily chitinized triangularexternal pore at the top of a small papilla (fig. 13, PI. 17). Thestem or neck of the gland extends from the base of the epidermis(hypodermis) to the outer surface of the cuticula, while the bodyof the gland lies mainly below the epidermis (fig. 12, PL 17).The neck portion is formed from a single cell, the neck-ce l lor t e r m i n a l cell (fig. 17, PL 17). The main body of the glandis made up of four elements—a large pear-shaped c e n t r a lcell partially surrounded by three more slender p e r i p h e r a lcells. The latter are fitted into deep grooves in the former(figs. 1 and 12, PL 17). The central cell contains three nuclei,a large spherical one in the distal end of the cell and two smallones at the proximal end nearer the epidermis (figs. 2-4, 5, 6,and 18, PL 17). Above the distal nucleus one finds a large,globular, clear space, the rese rvo i r , and a system of tubeslined with chitin. This branched system of tubes leads into asingle efferent duct which emerges from the top of the reservoirand traverses the narrow upper part of the central cell, fromwhich it is continued through the centre of the neck-cell toterminate in the central aperture of the external pore (figs. 15,16, and 17, PL 17). The three peripheral cells are uninucleate.Directly above the nucleus are clear vacuoles (fig. 21, PL 17) inthe cytoplasm, and farther up these vacuoles run together toform a single space, the duct, which extends spirally throughthe upper end of each cell to the neck-cell (figs. 7-10,12,14,18,and 19, PL 17). From the base of the neck-cell the ducts ofthe peripheral cells are continued as three tubes, which pursuea slightly spiral course through the neck-cell to end in the threeperipheral openings that form the sides of the triangular ex-ternal derm-pore of the gland (figs. 10,11, and 13, PL 17). Thestructure of the triangular gland is indicated diagrammaticallyin Text-fig. 1 and PL 20.

132 PRISCILLA FREW POLLISTEE.

T u b u l a r Glands .The tubular type of gland is composed of twelve cells, and, as

a whole, is flask-shaped. The short neck portion is composedof a single cell that is located in the epidermis and containsa large central chitinous duct which opens to the exterior whereit is continuous with the exoskeleton. The bulk of the gland isbelow the level of the epidermis and consists of ten peripheralcells completely surrounding a much larger central cell. Thelatter has essentially the same shape and internal structure asthe central cell of the triangular gland. It contains three nuclei,one large and two small (figs. 55, 56, 57, and 64, PI. 18). In thelarger tubular glands the large distal nucleus of the central cellis shaped like a cup with one part of the rim higher than therest (fig. 65, PI. 18). The duct of the central cell emerges atits upper end and opens into the side of the large tube that isthe main duct of the gland as a whole (figs. 63 and 65, PI. 18).The internal structure of the peripheral cells is like that of thesame cells of the triangular glands. Above the single nucleusare vacuoles which become confluent to form a triangular ductthat fills the neck of the cell (figs. 51, 52, and 66, PL 18).The peripheral cells are fitted closely around the central celland they occupy grooves in the cytoplasm of the latter. Whenthere are spaces between the surrounding cells the cytoplasmof the central cell is pushed out to fill these spaces so that thegland as a whole has a smooth, globular outline (figs. 54-7, 61-2,PL 18).

As already stated, the neck of the tubular gland is composedof one cell, which is shaped like a truncated cone and whichcontains a chitinized tube which is of uniform diameter, exceptfor a slight flare at the basal end. Here it is joined by the ductof the central glandular cell (fig. 63, PL 18). The triangularducts of the peripheral cells do not open directly into thechitinized tube of the neck, but seem to end in a clear spacewithin the base of the neck-cell and just below the bottom ofthe chitinized tube (fig. 68, PL 18). Because of this relationshipto the peripheral cells, the base of the neck-cell, when seen inface view from above, appears to be a circular plate with tenperipheral triangular perforations (openings of the ducts of the

WAX-GLANDS OF PSEUDOCOCCUS 133

peripheral cells), and a single peripheral circular opening (ductof the central cell). The apices of the triangles face the slightly-raised centre of the base of the neck-cell, which is the pointwhere the central glandular cell abuts against this base. Thenucleus of the neck-cell is found outside the ring of ducts andclosely adjacent to the cell membrane (fig. 59, PL 18).

The complete structure of this complicated tubular type ofgland can now be clearly understood if one examines figs. 45-53,PI. 18, a series of slightly oblique sections from bottom to topof a single gland. These should at the same time be comparedwith the diagrammatic representations in Text-fig. 2 and PI. 20.The central glandular cell and its large nucleus project belowthe rest of the gland and thus are cut in the first section (fig. 45,PL 18). In the next sections appear the reservoir of the centralcell and, successively, the various peripheral cells and theirnuclei (figs. 46-8, PL 18). In fig. 49, PL 18, the section is abovethe large nucleus of the central cell and shows the two smallnuclei. The next section (fig. 50, PL 18) is through the chitinizedtubes in the upper part of the reservoir, and it likewise shows,for the first time, sections of all ten peripheral glandular cells.Next (fig. 51, PL 18) one sees the single efferent duct of thereservoir and the ducts of the peripheral cells, the latter on theright. Fig. 52, PL 18, is of a section which, on the upper right,passes through the base of the neck-cell (compare with figs. 58and 59, PI. 18). Just to the left of the neck-cell is the efferentduct of the central cell entering the tubular main duct of thegland. The lower half of this oblique section passes throughfive peripheral cells, as they curve up over the central cell andconverge to join the base of the neck-cell. Pig. 53, PL 18, showsthe tubular pore of the gland on the surface of the cuticula.

M u l t i l o c u l a r Glands .

The multilocular gland, like the tubular type, is formed fromtwelve cells, and is flask-shaped with a short, wide neck. Theneck is formed from a single cell, that is penetrated by the necksof the eleven basal cells, and it ends in a highly characteristic ex-ternal pore consisting of a single central aperture surrounded byten regularly arranged peripheral openings—the whole overhung

134 PRISCILLA FREW POLLISTER

by a scalloped border (figs. 40 and 42, PL 17). The generalplan of this gland is much like that of the tubular type, fromwhich it differs chiefly in the character of the external openingand in the relative sizes of the eleven glandular cells. For herethe ten peripheral cells are very large and the central cell, bycomparison, is dwarfed, is uninucleate, and is devoid of reservoirand complicated internal duct system. A cross-section throughthe lower part of the gland shows the peripheral cells as triangles,their rounded bases forming the outer circumference of thegland and their apices meeting in the centre (figs. 26, 27, and 33,PL 17). A higher cross-section, just below the neck region, dis-closes the small central cell wedged in between the apices of theperipheral triangles (figs. 29 and 34, PL 17). The ducts withinthe peripheral cells are like those in the other types of gland(figs. 28 and 29, PL 17). A section through the base of the neck-cell appears very similar to the base of the same cell of thetubular gland, with the circle of triangular peripheral ducts(figs. 24, 35, 39, and 44, PL 17). The nucleus of the neck-cellis seen on one side, fitted between the cell membrane and theperipheral ducts. The centre of the ring, however, in this glandis occupied by the upper part of the central cell, which projectsinto the neck-cell slightly. The nucleus of the neck-cell is seenin lateral view in figs. 40 and 41, PL 17. Unlike the situation inthe tubular gland where the ten peripheral cells terminate atthe base of the neck-cell—in the multilocular gland the basalglandular cells seem to continue up through the terminal celland to open to the exterior independently of one another, eachthrough a single member of the apertures in the pore (figs. 81,36, and 37, PL 17). See p. 138.

Figs. 25-31, PL 17, are from a series of sections, from bottomto top, of a multilocular gland, and should be compared withText-fig. 3 and PL 20. Figs. 25-7, PL 17, are through the basesand nuclei of the ten peripheral cells. Fig. 28, PL 17, shows,on the right, the supranuclear vacuolated zone of the peripheralcells. Fig. 29, PL 17, is from the next section, that passes throughthe nucleus of the central glandular cell and, on the right,through the base and nucleus of the neck-cell. Fig. 30, PL 17,is through the middle of the neck-cell, except on the left, where

WAX-GLANDS OF PSBUDOCOCCUS 135

several of the peripheral ducts are seen converging to the pointwhere they enter the neck-cell. Fig. 31, PI. 17, is from the upper-most section, an oblique view of the gland-pore and, on the left,the lower part of the neck-cell. A similar but less oblique seriesis illustrated in figs. 32-7, PL 17.

B

TEXT-FIG. 3.

A, diagrammatic sagittal section of multilocular gland. B, poreof multilocular gland as seen from above, c, sagittal section ofneck region of multilocular gland drawn to same scale as B.

Deve lopmen t of t he Glands.

It has been frequently pointed out that in the developmentof the female coccid there are three immature instars, whichare distinguishable from one another by such characteristics asthe number of antennal segments, of spines, and of gland-pores.The fourth stage, adult, of course, is very readily identified bythe presence of the vaginal opening. In potash-magenta pre-parations these exoskeletal characters are very easily studied.The data in the Table are from such material. It will be observedthat there are four types which are assumed to represent thefour stages of development. The first group, with six antennalsegments, a small number of spines, and only the triangular typeof derm-pore, is the first instar. The tubular and multilocular

136 PEISCILLA FREW POLLISTBR

glands are not present anywhere on these specimens. Asecond group, representing the second instar, likewise hassix antennal segments, but the number of spines and of tri-angular gland-pores is increased and the tubular type of poreappears. Other features found here, and not present in groupone, are a thickened chitinized region around the last cerariangroup of spines and a triangular area of thickened chitin on thedorsal surface of the last segment. These facts seem to justifyconsidering the specimens in group two to represent the secondinstar. In a third group there are seven antennal segments;the number of triangular glands is approximately tripled; andthe number of spines is typically doubled. A curiously incon-sistent feature is a considerable reduction in the number of tubularglands. This group of specimens represents the third instar.The adults, group four, are, of course, unique in the possession ofthe vulva and the multilocular glands. They also show a furtherincrease in the number of triangular and of tubular glands aswell as spines.

Although counts were made only on the penultimate segment,the data are roughly representative of the whole body. Casualexamination of any region shows that, with the exception ofthe tubular type between instars two and three, there is a pro-gressive increase of the glands and spines at each step in develop-ment. Accordingly the details of the formation of the glandscan be best studied in the post-embryonic period, especiallybetween instars three and four when hundreds of each type arebeing formed.

Studies of ecdysis in insects (see especially Wigglesworth,1933) have shown that the first stage in the process is theformation of a fluid between the cuticula and the epidermis,which separates these layers that were previously tightlyadherent. The epidermis is thus freed and contracts to forma thicker epithelium of higher cells, and its constituent cellscan then round up and divide independently of one another.Hence it is during this period that changes involving increasein the number of epidermal cells can occur. It seems that inP s e u d o c o c c u s this early stage of ecdysis must be relativelybrief, since in a great majority of the sectioned immature

WAX-GLANDS OF PSEUDOOOCCUS

TABLE.

137

Number of glands and spines of Pseudocoocus m a r i t i m u s foundon the penultimate segment in the various instars and the adult female.The number of segments to the antennae, which is constant for a giveninstar and the adult female, is also shown.

1st Instar

2nd Instar

3rd Instar

Adult

Segments ofAntennae.

6666

666666

7777

8888

TriangularGlands.

6666

273241303032

10810897

108

305253258303

TubularGlands.

0000

111114121212

3015

212195154186

MultilocvlarGlands.

0000

000000

0000

63585568

Spines.14141414

191919191919

38383838

89745577

specimens cuticula and epidermis seem continuous and there isno indication of cell division in the latter. In a few cases,however, a distinct space separates cuticula from epidermis,and in these specimens there are numerous mitotic divisions inthe epidermis, and one finds various stages in the developmentof glands and spines.

Most of the mitotic figures are oriented with the axis of thespindle parallel to the plane of the surface of the epidermis,and these are obviously going to give rise to two cells that willbe a part of the simple epithelial layer of the epidermis, increas-ing its area. Here and there, however, one finds mitotic figureswith the spindle axis perpendicular to the epidermal layer, andit is obvious that from such a layer should arise one epithelialcell and one that is sub-epithelial, i.e. sub-epidermal (fig. 67,

138 PBISCILLA FREW POLLISTER

P1.19). Such figures are possible first stages of development ofsub-epidermal structures, like the secretory cells of coecidglands, trichogen cells, and cenocytes.

Figs. 68 and 69, PI. 19, show a condition that seems to bea somewhat later stage in gland development, where there hasbeen an increase in number of sub-epidermal cells. Division ofthe cell continuous with the epidermis is contributing anothercell to those below the epithelial layer. Fig. 73, PI. 19, is evi-dently a stage slightly later than figs. 68 and 69, PL 19. Itappears that before cell-multiplication1 has been completed toattain the number characteristic of the mature functional gland,the uppermost cell, that is, within the epidermis, has differen-tiated the chitinized structures typical of the neck of the tubulargland. A similar condition for the multilocular gland is seen infigs. 72 and 81, PL 19, except that here it should be noted thatthe neck-cell has developed only the superficial opening of thegland.

It is well to digress somewhat at this point to emphasize thesignificance of the difference in development of these two typesto the interpretations of their structure when fully formed. Itwill be recalled that it was stated (p. 133) that the ducts of theperipheral cells of the tubular gland terminate below the baseof the neck-cell; while in the multilocular type (p. 134) thesecells penetrate the neck-cell to open independently on thesurface. This assumption concerning the latter type was pre-ferred to the alternative one that the continuation of the peri-pheral cell-ducts through the neck-cell were differentiationswithin the substance of that cell, largely because of the situationshown in figs. 71, 72, and 81, PL 19—since here there is no traceof differentiated structure in the neck-cell below the level of thesuperficial cup, into which the eleven ducts open in the fullyformed multilocular gland.

Appearances like figs. 70, 75, and 77, PL 19, are obviouslyslightly later than figs. 72 and 73, PL 19. Here the sub-epidermalglandular rudiment is represented by a mass of cytoplasmcontaining many nuclei, of uniform size and smaller than those

1 Cell-multiplication is shown only by the number of nuclei since it isnot possible to make out the cell boundaries in these early stages.

WAX-GLANDS OF PSEUDOCOCCUS 139

of earlier stages. Figs. 75, 77, and 78, PL 19, are all cross-sec-tions of the sub-epidermal part of the rudiments of multilocularglands, and are of special interest because of the fact that eachshows eleven nuclei. Since this is the number characteristicof the functional gland these figures must represent the end ofthe period of nuclear multiplication. (Note that in addition tothe eleven nuclei of the potential glandular cells in figs. 75 and77, PI. 19, the neck nucleus is also seen at a slightly higher focalplane.) There should be a similar time when the sub-epidermalpart of the rudiment of the developing tubular gland wouldshow thirteen nuclei, but it is very difficult to count them whenthey are closely crowded together and I have not been able todetermine one with more than the ten shown in fig. 70, PI. 19.

The stage just described requires little further change tobecome the mature multilocular gland, i.e. the appearance ofcell boundaries (fig. 78, PL 19), development of the secretorymaterial in the cytoplasm of the cells, and the growth of theglandular cells through the neck-cell to reach the apertures ofthe derm-pore. In the case of the tubular gland an extensivedifferentiation of the central cell must occur. Figs. 76, 79, and80, PL 19, show one later stage in the development of the tubulartype. The large nucleus must be that destined to be locatedat the base of the central cell. There is no reservoir in thecentral cell, nor are there vacuoles in those at the periphery—soapparently this gland is not yet functioning.

I have not definitely identified stages in the development ofthe triangular type of gland. These glands have but five cellsand it seems probable that development is much more rapidthan in the other two types. The ducts of all four gland-cellsseem to penetrate the neck-cell in the mature triangular gland,as do the cells of the multilocular gland, so that it seems likelythat here also the neck-cell gives rise to at most only the outer-most part of the gland-pore.

At an early stage of ecdysis, when the development of theglands is taking place, one finds numerous large irregularlyshaped cells lying in the body cavity, immediately under theepidermis (figs. 71 and 74, PL 19). At first they are so numerousas to form almost a continuous layer. In the late stage of

140 PRISCILLA FREW POLLISTER

ecdysis, after the glands and a new cuticula are formed, thenumber of these cells is much reduced. From their appearanceand behaviour these cells are undoubtedly the structures variouslyknown as oenocytes, coenocytes, ceridocytes, et als. Wiggles-worth, working on E h o d n i u s , has very carefully describedthe relationship of these cells to the course of ecdysis, showingthat in the later stages they become very much reduced in size,which leads him to believe them the source of the cuticularsubstance. Perhaps of more significance to the present studyis the observation of Eogojanu, 1935, that in the aphid E r i o -s o m a the oenocytes are, in some parts of the body, restrictedto the vicinity of the developing glands; where there is, in fact,a single oenocyte underlying each group of gland-cells that isa gland-field. Indeed, it is a remarkable fact that in E r i o s o m aEogojanu finds that he can, as it were, predict the site of deve-lopment of a gland by the presence of this cell below theundifferentiated epidermis. He suggests that one may regardthe oenocytes of E r i o s o m a as serving a nutrient functionfor the developing gland-cells. It was possible to establish theabove relationship in E r i o s o m a because the glands are fewin number and restricted to very definite areas. In P s e u d o -coccus the glands are so widely distributed that one cannotdetermine whether there is any definite correlation between thepresence of oenocytes and glandular differentiation.

DISCUSSION.

The only extensive comparative study of the glands inCoccidae has been made on the family Margarodidae, a differentgroup of coccids from that to which P s e u d o c o c c u s belongs.In this group Morrison (1928) describes and figures the appear-ance of the derm-pores as seen in potash-magenta preparationsof over a hundred species. It will be recalled that surface viewsof this sort of preparation of P s e u d o c o c c u s m a r i t i m u sshow two categories of pores: one which has a simple circularoutline (tubular, Text-fig. 1 B), and another with a marginalring of openings surrounding a single central aperture (includingthe triangular and multilocular types, Text-figs. 2 B and 8 B).

WAX-GLANDS OP PSEUDOCOCCUS 141

The former is what Morrison calls the simple type, and he findsit confined to four genera. He applies the term multilocular ina general sense to the latter type of pore, which is found in everyspecies of Margarodidae that he has examined. The derm-poresof the multilocular type show considerable variation within thisgeneral scheme of peripheral and central apertures. The numberof marginal openings surrounding a single central aperturevaries, in different species, and includes every number fromtwo to seventeen. Furthermore, the number of central openingsis variable, though within a narrower range. The most com-plicated derm-pore seems to be a type found in X y l o c o c c u s ,where there are two concentric circles, each of twelve pores,and two central apertures. Morrison's work on this family issubstantiated by numerous fragmentary observations of workerson other coccid groups. All the derm-pores of multicellularglands that have been described may be placed in one of Morri-son's two categories, i.e. either simple or multilocular. Thepresent study has demonstrated that as regards their sub-cuticular anatomy these two types are essentially identical,consisting of central and peripheral glandular cells. Hence onecan make a general statement that this plan of histologicalstructure is the rule for the wax-glands of Coccidae (see below,p. 142).

The histological structure of the wax-glands has hithertobeen most carefully studied by Matheson, 1923, working onP s e u d o c o c c u s c i t r i , a form also earlier examined byVisart, 1894. As far as Matheson's observations go they agreevery closely with the results of the present study on anothermember of the genus. In P s e u d o c o c c u s c i t r i are alsofound tubular, triangular, and multilocular glands that aredistributed as in the adult P s e u d o c o c e u s m a r i t i m u s ,and each of these types is made up of a single central cell sur-rounded by a ring of marginal cells. Matheson, however, believesthe number of the latter to be variable, but he agrees that asingle aperture in the derm-pore is always the opening of a singleperipheral glandular cell. In several other features Matheson'sresults differ from the present observations on P s e u d o c o c c u sm a r i t i m u s . He does not mention either the small nuclei or

142 PKISCILLA FREW POLLISTEB

the smaller ducts of the central cell, the spiral course of theperipheral ducts of the triangular gland, or the neck-cell.

In I ee ry a Murdock, 1928, has described the structure ofa multilocular gland, which has two central openings in thepore. There is but one central cell, which discharges throughthe two apertures. This cell is only a little larger than theperipheral cells, and it has neither reservoir nor duct system—features in which it resembles the small central cell in themultilocular gland of P s e u d o c o c c u s m a r i t i m u s .

The work of Teodoro, 1911, on P u l v i n a r i a is of interestto the present study chiefly because of the unique observationof the two small nuclei of the central cell. Otherwise his observa-tions are almost certainly incomplete, since he apparently notedno peripheral cells.

In S a i s s e t i a Marshall, 1929, has described glands whichevidently conform to the usual type, although cell boundariescould not be determined in his material. He mentions a largenucleus, above which is a 'vacuole' (reservoir?), 'accessorycells' (peripheral?), and other features that unmistakablysuggest the histological characteristics of wax-glands of otherCoccidae.

Very recently Eogojanu, 1935, has briefly described andfigured in O r t h e z i a a gland that is clearly very similar to thetubular type of P s e u d o c o c c u s m a r i t i m u s . Eogojanualso noted glandular areas containing but one type of cell, eachcell opening independently on the outer surface. It seems tothe author that these are more properly to be regarded as uni-cellular glands, and as such they constitute a unique exceptionto the usual method of wax production in Coccidae, resemblingmore closely the condition in Apis and some other insects.The case of O r t h e z i a would probably repay a re-examination,more thorough than the study of Eogojanu seems to be.

The literature contains numerous incomplete observationsshowing more or less of the histological structure of the multi-cellular wax-glands of Coccidae, which need not be discussedin detail (e.g. see Putnam, 1878; List, 1886; Berlese, 1898;Moulton, 1907; Fullaway, 1910; Johnston, 1912; Childs, 1914).With the possible exception of O r t h e z i a noted above, all of

WAX-GLANDS OF PSEUDOOOCCUS 143

the previous observations may be interpreted as consistent withthe view that wax-glands of Coceidae are all modifications ofone general scheme of histological structure in that they consistof two types of cells, distinguishable from one another by theirorientation within the glandular mass—either in a central ora peripheral position. These cell types frequently show markedcytological differences as well. The peripheral cells often containsmall scattered vacuoles which run together to form a large massin the end of the cell nearest the pore. These are not found inthe central cell. Instead, in those cases where its histologicaldifferentiation is marked it contains a single large vacuole orreservoir (presumably full of the special secretory product), andthis is drained by a complicated chitinized duct system.

In view of the wide occurrence of the above type of gland incoccids the question naturally arises of the significance of thesetwo types of cells in relation to the function of the wax-glands.In the present study the only possible method of approach tothis problem is a comparative one. When one considers wax-producing glands in insects other than coccids it is found that,in most cases, as in bees and aphids, the glands contain but onecellular type, a relatively unspecialized cell much like the peri-pheral glandular units of the Coceidae. In bees the glandsconsist of a layer of these cells in the form of an extensiveglandular field, on to the surface of which the wax exudes as anamorphous mass. This may be taken to indicate that the peri-pheral cells are the ones primarily concerned with the formationof wax in coccids; at least, it certainly shows that wax can beformed in the absence of the central cell type. Turning to otherinsect groups, one finds a wax-gland strikingly like those ofcoccids in an apparently distantly related group of Homoptera,the Fulgoridae. This was earlier described by Bugnion andPopoff, 1907, in F l a t a (Phromnia), and more recently Sulc,1928, has described these structures in the same form and inother Fulgoridae.1 Bugnion and Popoff considered the central

bulc's paper has not been consulted. It is written in Czech and is notreadily obtainable. The reference to it is given in Weber's 'Lehrbueh derEntomologie', where two of his figures are reproduced. The suggestionconcerning the function of the central cell is quoted in Rogojanu, 1935.

144 PEISCILLA FREW POLLISTBR

cell to be a nervous structure and Eogojanu (loc. cit.) hasadopted this suggestion as the most likely for the gland ofOrthezia. This view is manifestly absurd when applied to astructure like that of the central cell of the triangular andtubular glands of P s e u d o c o c e u s , and it seems to the writerthat this cell must be glandular in function. When comparisonbetween Fulgoridae and Coccidae is extended to the type ofwax produced it is found that these two groups are alike, anddiffer from other insects, in secreting the wax in the form ofaccurately moulded filaments instead of an amorphous mass.This correlation of histological structure with function has beenduly noted by Sulc, and he has suggested that the secretion ofthe central cell is not wax but a material that causes the waxelements produced by the peripheral cells to adhere to oneanother to form compound filaments having the dimensions ofthe pore. This hypothesis seems to the writer highly probable,and it is suggested that it be extended to apply to the centralcells of the coccid wax-glands. In the present study there isa slight further support for Sulc's theory in the correlation ofthe degree of development of the central cell with the type ofwax-form produced. It will be recalled that large wax filamentsare produced by the tubular and triangular glands, in both ofwhich there is a well-developed central cell. The multiloculartype, however, has a very small central cell that is only slightlydifferentiated. Indeed, when it is compared with the centralcell of the other two types it appears reasonable to consider itrudimentary and, in all probability, non-functional. In viewof this condition it seems highly significant that the wax ele-ments exuding from the apertures within the pore of this typeof gland remain distinct, that is, do not fuse to form a compoundproduct of specific shape, in marked contrast to the other twoglandular types.

If Sulc's suggestion be assumed to be correct, then one maysummarize the functioning of the typical coccid wax-gland asfollows: the peripheral cells produce wax, which is squeezedout of the peripheral apertures in the form of filamentousstructures. Simultaneously, from the central cell there comesa substance that will cause the filaments to adhere to one

WAX-GLANDS OF PSEUDOCOCCUS 145

another. In the tubular gland this mixture is squeezed withinthe tube and accurately moulded to form the uniform cylinderthat has been often figured. In the case of the triangular typethere is no mould, but the filaments must emerge in closecontact with one another and in parallel spiral courses, and itis not difficult to conceive that under such circumstances theywould adhere to form a definite compound structure, a largewax filament.

What is the nature of the material produced by the centralcell that has this property of causing the filaments to adhere ?The first thought, upon consideration of the properties of wax,is that this would be best accomplished by some volatile waxsolvent. This leads to a comparative consideration of cellshistologically similar to the central cell, a very specialized type.Cells with a similar chitinized internal duct system and reservoir(also sometimes chitinized) are also found in many Heteroptera,where they occur as unicellular glands that are generally be-lieved to produce the substance responsible for the scentcharacteristic of many of these insects (see Weber, 1933).Many scents are volatile oils, and the most common wax solventsbelong to this class of substances. To the extent that thiscomparison between cell types of rather remotely related insectsseems acceptable as valid one can perhaps tentatively concludethat the central glandular cells of coccids produce a volatileoil which, by virtue of a solvent action on the filamentous waxproducts of the individual peripheral glandular cells, causesthem to adhere to one another to form large compound filamentsof wax.

In none of the previous studies of the histology of coccidianwax-glands has there been a description of the neck-cell. Thisis probably because of the paucity of work on the developmentof glands, since, as will be recalled, it was this aspect of thepresent study that led to the concept of a special cell responsiblefor formation of more or less of the external cuticular orificeof the gland. The neck-cell seems closely analogous to the so-called tormogen cell that occurs in relation to developing spines.This cell, as has been most clearly shown by Wigglesworth,1933, is an individual of the epidermal layer, and in relation

NO. 317 L

146 PKISCILLA FREW POLLISTBE

to the outer layer of the tormogen cell there develops a cup-likedepression of the cuticula. A trichogen cell lies immediatelybeneath the tormogen cell, and the developing spine from theformer grows through the latter and emerges at the outersurface in the centre of the cuticular depression, so that the baseof the mature spine is encircled by a groove. The resemblanceof this course of development to the mode of origin of the rela-tionship between the pore of the multilocular gland and theducts of the glandular cells is obvious and striking.

The literature on the development of wax-glands of coccidsis very meagre, represented by but two papers. Eogojanu,1935, has studied the development of the glands of 0 r t h e z i a ,but since he was mainly concerned with cytological observationsindicating the onset of function his results need not be reviewedhere. His more significant observations on E r i o s o m a areconsidered on p. 140. The development of the glands of Sa i s -s e t i a was partially worked out by Marshall, 1929. Althoughhe does not recognize a neck-cell of the gland he apparentlyobserved the development of the tube before the differentiationof the glandular cells. His results are thus confirmatory of thisunique aspect of the present concept of the morphology of thegland.

SUMMARY.

The females of P s e u d o c o c c u s m a r i t i m u s have threetypes of multicellular wax-glands, one with a triangular externalpore, another opening through a long tube, and a third with amultilocular aperture. The first two are widely distributed onall surfaces of the adult. The third is restricted to the ventralsurfaces of the last five segments. This multilocular type isfound only in the adult. The triangular glands are found atall stages and these structures progressively increase in numberwith each successive instar. The tubular type appears first inthe second instar; the number is reduced in the third instar;and in the adult it is again increased to the largest numberfound at any stage.

The three glands are all modifications of one general plan ofhistological structure. The glandular elements are sub-epidermal

WAX-GLANDS OP PSBUDOCOCCUS 147

cells arranged in a ring of peripheral cells surrounding a singlecentral cell. There are three peripheral cells in the triangulargland and ten in each of the others. The peripheral cells areuninucleate and contain vacuoles of secretory material. Thecentral cell of the tubular and triangular glands has a large andtwo small nuclei and contains a large reservoir, from which achitinized duct system leads to the gland-pore. The centralcell of the multilocular gland is small and relatively undiffer-entiated. The author favours the view of Sulc that the wax isprobably secreted by the peripheral cells, while the central cellsecretes a substance that causes the wax filaments to adhere toform large cylinders.

The glands are developed by cell-multiplication from theepidermis at the time when it is freed from the cuticula at thebeginning of ecdysis. After the initial period of cell-multiplica-tion the first differentiation is the development of the externalpore within the neck-cell. Later in the development of multi-locular glands it is believed the glandular cells grow throughthe neck-cell to establish the functional relationship with thepore. It is suggested that this is analogous to the relationshipbetween tormogen cell and trichogen cell in the developmentof a spine.

REFERENCES.

Berlese, A., 1893.—"Le coociniglie Italiane viventi sugli agrumi", 'Riv.di Patologia Vegetale1, 2.

1894.—"Le coociniglie Italiane viventi sugli agrumi", 'Riv. di Pato-logia Vegetale', 3.

Bugnion, E., and Popoff, N., 1907.—"Glandes cirieres de Flata (Phroinnia)marginella", 'Bull. Soc. Vaudoise Sci. Nat.', 43.

Childs, L., 1914.—"Anatomy of the Diaspinine scale insect Epidiaspispiricola", 'Annals Ent. Soc. Amer.', 7.

Dietz, H. F., and Morrison, H., 1916.—"Coccidae or scale insects of In-diana", 'Publ. of State of Indiana'.

Ferris, 6. F., 1918.—'California species of mealy-bugs.' Leland StanfordJunior University Publications.

Fullaway, W. S., 1910.—"Description of a new coccid species, Ceroputoambigua", 'Proc. Davenport Aead. of Sciences', 12.

Johnston, C. E., 1912.—"Internal anatomy of Icerya purchasi", 'Ann.Entom. Soc. Amer.', 5.

148 PRISCILLA FREW POLLISTEK,

List, J., 1886.—" Orthezia cataphracta Shaw. Eine Monographie ", 'Zeitaeh.f. wissenschaftliche Zoologie', 45.

Marshall, W. S., 1929.—"Hypodermal glands of the black scale, Saissetiaoleae (Bernard)", 'Trans. Wise. Aead. Sci.', 24.

Matheson, R., 1923.—" Wax-secreting glands of Pseudococcus citri (Risso)",'Ann. Ent. Soc. Amer.', 16.

Morrison, H., 1928.—"Classification of the higher groups and general ofthe coccid family Margarodidae", 'U.S. Dept. Agr. Tech. Bull.', 52.

Moulton, Dudley, 1907.—"The Monterey pine scale, Physokermes insigni-cola (Craw)", 'Proc. Davenport Acad. Sci.', 12.

Murdock, G. E., 1928.—"Wax-secreting mechanism in the adult female ofIcerya purchasi", 'Pan-Pacific Entom.', 5.

Putnam, J. D., 1878.—"Biological and other notes on Coccidae. I. Pul-vinaria innumerabilis", 'Proc. Davenport Acad. Sci.', 2.

Rogojanu, P., 1935.—"Unters. u. d. Wachsdriisen u. d. Wachsabsonderungbei den Qattungen Sehizoneura und Orthezia", 'Zeit. f. mikr. anat.Porsch.', 37.

Teodoro, G., 1911a.—"Glandule ceripare della femmina della Pulvinariacamelicola", 'Redia', 7.

19116.—"Secrezione della cera nei maschi della Pulvinaria cameli-cola", ibid., 7.

1912.—"Glandulo laccipare e ceripare del Lecanium olea", ibid., 8.Visart, 0., 1894a.—"Contr. a. st. delle glandule ceripare delle coccinigli

(Dactylopius citri e Ceroplastes rusci)", 'Riv. di Patologia Vegetale', 3.18946.—"Contr. a. conosc. delle glandule ceripare negli afidi e nelle

cocciniglie", 'Bolletino Soe. Natur. Napoli', 8.Weber, 1933.—'Lehrbuch der Entomologie.' Jena.Wigglesworth, V. B., 1933.—"Physiology of the cuticle and of ecdysis

of Rhodnius prolixus (Triatoniidae, Hemiptera)", 'Quart. Journ. Micr.Sci.', 76.