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Ultrastructural morphology of the male and female

genital tracts of Psoroptes spp. (Acari: Astigmata:

Psoroptidae)

MIREILLE LEKIMME1,*, MICHELE LECLERCQ-SMEKENS2,CHANTAL DEVIGNON3, THIERRY LECLIPTEUX4, YVESPOUMAY2,3 and BERTRAND LOSSON1Service of Parasitology and Pathology of Parasitic Diseases, Department of Infectious Diseases,

Faculty of Veterinary Medicine, University of Liege, Boulevard de Colonster 20, B43, 4000 Liege,

Belgium; 2Service of Histology-Embryology, Faculty of Medicine, Facultes Universitaires Notre-

Dame de la Paix of Namur, Namur, Belgium; 3Unity of Electronic Microscopy, Facultes Universi-

taires Notre-Dame de la Paix of Namur, Namur, Belgium; 4Coris Bioconcept, Parc Crealys, rue Phocas

Lejeune 30, 5032 Gembloux, Belgium; *Author for correspondence (e-mail: mireille.lekimme@ulg.ac.be;

phone: +32-4-366-40-93; fax: +32-4-366-40-97)

Received 1 November 2004; accepted in revised form 19 April 2005

Key words: Acari, Functional morphology, Genital system, Psoroptes, Ultrastructure

Abstract. The structure of the male and female genital systems of the astigmatid mite Psoroptes ovis

(Hering) is described. The male genital system is composed of a paired testis, fused at its proximal

part, two vasa deferentia, an ejaculatory duct, into which a single accessory gland opens, and a

copulatory organ. The testis is characterized by a peripheric syncytial cell surrounding spermato-

gonia, spermatocytes, spermatids and spermatozoa which are distributed regularly in the gonad

according to the sequence of spermatogenesis. The female genital system consists of a copulatory

pore (the bursa copulatrix), a seminal receptacle, paired ovaries and oviducts, a glandular uterus

and an ovipositor which leads to the oviporus. Ovaries are composed of somatic cells, germ cells

and a central cell, with a multilobular nucleus, connected to oocytes by a stalk. Similarities with

other astigmatic mites belonging to Psoroptidia and Acaridia are also discussed.

Introduction

Species of the genus Psoroptes are astigmatid, obligatory non-burrowing ec-toparasites that cause various forms of psoroptic mange in mammalian hostsincluding sheep, cattle, goats, horses and rabbits. As many as five species ofPsoroptes mites have been described (P. cuniculi, P. cervinus, P. equi, P. ovisand P. natalensis), primarily on the basis of their localization and their hosts(Sweatman 1958; Strong and Halliday 1992). However, because cross-mating(Wright et al. 1983), antigenic (Boyce and Brown 1991) and genetic (Zahleret al. 1998; Ramey et al. 2000) studies indicate that Psoroptes from varioushosts and geographic areas are identical, some authors suggest that all host-related species described by Sweatman (1958) should be described as Psoroptesovis (Hering) (Bates 1999; Sanders et al. 2000). In this context, the following

Experimental and Applied Acarology (2005) 36:305–324

DOI 10.1007/s10493-005-6076-8 � Springer 2005

observations performed on Psoroptes originated from the ears of rabbits couldbe extended to Psoroptes from other hosts.

Psoroptic mange is present on mainland Europe and has considerable eco-nomic importance in animal husbandry (Meleney and Roberts 1979; Wrightand Guillot 1984; Wright 1986). Bovine psoroptic mange is particularlyimportant in Belgium (Pouplard et al. 1990; Lonneux and Losson 1996).This mite has therefore been the subject of major interest over the past decades(e.g. Downing 1936; Guillot and Meleney 1982; Kirkwood 1986; Boyceet al. 1990; Perrucci and Macchioni 1995; Coles and Stafford 1999; Rameyet al. 2000; Smith et al. 2000; Brooks and Wall 2001).

In contrast, few papers have been dedicated to the anatomy and finestructures of the mite. Until now, anatomy and general morphology of thedigestive tract have been investigated in P. ovis with light microscopy byBeetham (1997) and by Mathieson and Lehane (2002) by electron microscopy.The structure and function of the gnathosoma have been studied electronmicroscopally by Mapstone et al. (2002). The nervous system and the femalereproductive tract have briefly been described by Beetham (1997) while thebursa copulatrix, genitalia and associated suckers were investigated by Guillotand Wright (1983) and Krishna Rao et al. (1974); the spermatozoa weresuccinctly depicted by Alberti (1984).

The present study on P. ovis from rabbits aimed to consider the entire maleand female reproductive systems using the analysis of serial semi-thin sectionsfollowed by ultrastructural observations.

Materials and methods

Psoroptes mites were obtained from the ears of experimentally infected rabbitsheld at the Faculty of Veterinary Medicine in Liege, Belgium.

Transmission electron microscopy

The legs of adult mites were removed to facilitate penetration of the fixative forTEM investigation. This was performed at 4 �C to suppress mite motility.Adult males and females were then fixed for 48 h at 4 �C in either 2% glu-taraldehyde or a solution of 2.5% glutaraldehyde and 2% formaldehyde in0.1 M cacodylate buffer, following the technique of Karnovsky (Hayat 1986).Specimens were then washed in 0.2 M cacodylate buffer and postfixed for 1 hat 4 �C in 2% OsO4 solution in cacodylate buffer. After a final wash, the miteswere slowly dehydrated at room temperature in increasing concentrations ofethanol and propylene oxide and then embedded in Epon 812.

Serial semi-thin (2 lm) sections (eight females and six males), stained withtoluidine blue, were examined to reconstruct the reproductive systems. Ultra-thin sections (12 females and six males) were contrasted with uranyl acetate

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and lead citrate (Reynolds 1963) and examined in a Philips Tecnai 10 electronmicroscope at 60 kV.

Scanning electron microscopy

The external genital organs of Psoroptes males and females were examined bySEM. In this case, unfixed males and females were dehydrated in graded seriesof acetone and critical-point-dried. The dry samples were stuck on double-sided adhesive tape, coated with gold and examined with a Philips XL20scanning electron microscope.

Results

The male reproductive system

The male reproductive system of Psoroptes consists of a unique testis, two vasadeferentia, an accessory gland, a median ejaculatory duct and the copulatoryorgan (aedeagus or penis) (according to Alberti and Coons 1999) (Figure 1).The latter is situated ventro-medially between the third and the fourth pairs oflegs (Figure 2b). Functionally related structures include one pair of adanalsuckers (Figure 2d) and two pairs of small suckers on the tarsi IV (Figure 2c).

TestisThe unpaired testis is situated in the hemolymphatic cavity of the opisthosomaand does not show any suspension. In fact, it can be assumed that primarilypaired testes have fused because the testis comprises two units interconnectedby a narrow bridge (not illustrated), situated symmetrically in the posteriorpart of the body, on both sides of the rectum. The testis is bordered by anepithelium that seems to consist of a syncytium. Cell borders within the syn-cytium have not been identified and nuclei are encountered in clusters atirregular intervals (Figure 3g). Cytoplasm extensions of this syncytium (spurs)are visible between the germ cells (Figure 3a). The nuclei of the syncytiumcontain little condensed chromatin, measure approximately 3 lm and arecharacterized by one or more distinct nucleoli. Different stages of the sper-matogenesis are evenly distributed between the spurs of the syncytium: thespermatogonia are clustered in the dorsal region and the central part is occu-pied by the spermatocytes and spermatids. The ventral area is filled with spermcells (Figure 3a).

Spermatogonia are polygonal cells, approximately 4 – 5 lm in diameter,which adhere tightly to each other. The nucleus is of 2.5 – 3.5 lm in diameter,contains few patches of electron-dense chromatin and is limited by a nuclearenvelope that possesses some scattered nuclear pore complexes where extra-nuclear electron-dense material accumulates (Figure 3b). The nucleolus, mea-

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suring approximately 1.5 lm, is often located centrally and its three majorstructural components (granular and fibrillar regions and fibrillar centers) areclearly visible that could show a great activity in protein synthesis of the cell.Mitochondria are 220 – 260 nm wide, possess an electron-dense matrix withlongitudinally orientated cristae and are located close to the nuclear envelope.It can be assumed that they represent rather long mitochondria as observed insome sections (more than 2 lm in length). The cytoplasm further containsmany free ribosomes responsible for the dark aspect at low magnification andrare dispersed dictyosomes.

Early spermatocytes (young spermatocytes) are very similar to spermato-gonia. They differ by their size (8 – 10, 3.5 – 4 and 1.3 – 2.1 lm in diameter forthe cell, the nucleus and the nucleolus, respectively), their paler cytoplasm andthe increasing number of Golgi complexes situated at the periphery (notillustrated) (Figure 3c). Mitochondria are evenly distributed in the cytoplasmcomparing to what has been observed in spermatogonia while the rough

Figure 1. Schematic drawing of the ventral (a) and sagittal (b) aspects of P. ovis male genital

system. AG = accessory gland; AS = adanal sucker; ED = ejaculatory duct; Gn = gnatho-

soma; P = penis; T = testis; VD = vasa deferentia; I, II, III, IV = legs. (scale = 100lm).

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endoplasmic reticulum cisternae are often located beneath the plasmalemma.In more advanced (mature) spermatocytes (Figure 3d) which measure 8 –12 lm in diameter, the electron-dense granular material adhering to the nuclear

Figure 2. (a–d) Various SEM aspects of P. ovis male; (a) ventral view; (b) copulatory organ

(expanded); (c) adanal suckers; (d) small suckers present on the tarsi IV; (e) SEM of the docking

papillae (copulatory lobes) of the female tritonymph of P. ovis. A = anal slit; AS = adanal

sucker; CS = copulatory sac (genital papilla); DP = docking papilla; Gn = gnathosoma;

P = penis; PL = posterior lobes; S = sucker ; I, II, III, IV = legs.

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pore complexes on their cytoplasmic side becomes pyramidal (basal diameter isabout 1 nm and height ranges from 0.5 to 0.8 nm). The nucleolus becomeseccentric and seems to be smaller than observed in the early spermatocytes.Mitochondria with transverse cristae are either dispersed, either clustered in thecytoplasm (Figure 3e). Accumulation of large amounts of cisternae of theendoplasmic reticulum can be observed beneath the cell membrane and formspongy layers (Figure 3d).

Spermatids show circular or oval sections and have a diameter of 4 lm(Figure 3f). They occur in groups composed of several closely apposed cellsresulting from the spermatocyte division. The early spermatid is characterizedby the absence of a nuclear envelope. The nuclear material is organized in aform of fine threads spreading throughout the cytoplasm. Mitochondriameasure 150 – 210 nm in width. Spongy layers get thicker and mitochondriatend to assemble.

Spermatozoa are located in the distal part of the testis and near the accessorygland. In cross-sections, they are polygonal (2 – 3 lm · 8 – 10 lm) (Figure 3g).The cytoplasm is electron-transparent and contains slightly eccentrically lo-cated 40 nm chromatin threads and peripherally located aggregations ofmitochondria (110 – 140 nm in thickness) – often near the chromatin threads.The R.E. cisternae-derived membranes (spongy layers) detach themselves fromthe cell surface and differentiate into lamellae in the cytoplasm.

Vasa deferentiaAt its anterior end, the testis gives rise to the paired vasa deferentia (Figure 1).No clear border can be drawn between testis and sperm ducts. The vasa de-ferentia are packed with spermatozoa and also with late stages of spermato-genesis (Figure 4b). They fill completely the ventral part of the body betweenthe gut, the accessory gland and the testis and serve as seminal vesicles. Tinycircular muscle fibers surround the wall of the vasa deferentia. Their con-traction makes possible the sperm transportation. The vasa deferentia mergeinto the ejaculatory duct at the level of the fourth pair of legs, ventrally to theaccessory gland.

Accessory glandA large, single accessory gland is located in a median position, above the vasadeferentia (Figure 1). This gland consists of a sac, surrounded by muscles,

Figure 3. (a) TEM view of the organization of the testis of P. ovis male; (b–g) TEM aspects of the

male germ cells; (b) spermatogonium; (c) early spermatocyte; (d) advanced spermatocyte; (e) higher

magnification of the nuclear envelope of the advanced spermatocyte; (f) spermatid; (g) spermatozoa

close to the syncytium. BL = basal lamina; Ch = chromatin threads; D = dictyosomes;

EM = accumulation of extranuclear material; FC = fibrillar center; FR = fibrillar region;

GR = granular region; L = lamellae; Mi = mitochondria; Mu = muscle; N = nucleus; Np =

nuclear pore; Nu = nucleolus; R = ribosomes; S = spur of the syncytium; Sc = spermatocyte;

Sd = spermatid; Sg = spermatogonium; SL = spongy layers; Sz = spermatozoon.

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Figure 4. (a) Sagittal semi-thin section of pairing between a female tritonymph and a male of

P. ovis. (b–c) TEM aspects of the accessory gland of P. ovis male; (b) general aspect; (c) magni-

fication of the epithelium and lumen of the gland. (d–e) TEM aspects of the uterus of P. ovis female;

(d) general aspect; (e) magnification of the apical region of the epithelium. AG = accessory gland;

AS = adanal sucker; BL = basal lamina; C = cuticule; Co = colon; DP = docking papilla;

FT = female tritonymph; Lu = lumen; M = male; Mu = muscle; Mv = microvilli;

N = nucleus; Nu = nucleolus; Sc = spermatocytes; SG = secretory granule; Sz = spermato-

zoa; V = ventriculus; VD = vas deferens; ZO = zonula occludens.

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more or less distended depending on its degree of activity and repletion. Its wallis made of an epithelium with very large cells showing large nuclei (5 – 6 lm indiameter) and electron-light cytoplasm (not illustrated) that become flattenedwhen secretion occurs in the lumen (Figure 4b). The cytoplasm of the epithelialcells becomes electron-dense with granular vesicles up to 1 lm in diameter(Figure 4c). The lumen of the gland is filled with small electron-dense granulesof approximately 64 nm in diameter.

Ejaculatory duct and copulatory organDirectly after the junction of the vasa deferentia with the accessory gland, thecuticule-lined ejaculatory duct (not illustrated) gives rise to the copulatory organ(aedeagus or penis). The latter, consisting of a telescopic tube coated by a rel-atively thick cuticule, is very delicate and tears easily in ultra-thin sections.Whennot in use, it is invaginated, hidden in the body and points anteriorly. Duringpairing, the penis is evaginated and points posteriorly (as illustrated in(Figure 2b) to make the retroconjugate mating possible. The penis is protectedand maintained in this position owing to the genital papillae (copulatory sacs)that may be expanded considerably when hydrostatic pressure rises (Figure 2b).

Accessory copulatory and pairing organsThe male possesses a pair of ventral copulatory suckers situated posteriorly(Figure 2d). They are directed ventrally and are cup-shaped with a flat surface.During the process of pairing, they protrude caudally with their openings di-rected backward (as illustrated in Figure 2d). Attachment takes place betweenadult males and female tritonymphs. The latter possess docking papillae(copulatory tubercules) (Figure 2e) on which the male’s copulatory suckers getfixed (Figure 4a). In this position, the opisthosomal lobes of the male overlapthe dorsal surface of the posterior part of the female. Although the adult maleand female tritonymph are found in such a pairing condition, the actualinsemination takes place only when the female tritonymph moults and reachesthe adult stage, as the insemination pore is formed only then. Adult males alsoappear to use their short fourth pair of legs, which possesses a pair of smallsuckers to grasp and hold the females (Figure 2c).

The female reproductive system

The female reproductive system of Psoroptes possesses two openings, theinsemination pore and the oviporus (Figure 6a). Starting from the insemina-tion pore (bursa copulatrix, copulatory pore, sperm induction pore), it consistsof a single inseminatory canal (sperm access tube, canalis copulator), a singleseminal receptacle (receptaculum seminis, sacculus), paired efferent spermducts (ducti conjunctivi), paired ovaries, paired oviducts, a single glandularuterus (chorion gland) and a single ovipositor joining the oviporus (accordingto Alberti and Coons 1999) (Figure 5).

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Insemination poreThe insemination pore is situated at the end of the anal plate near the anal slitand has a raised, cone-shaped structure (Figure 6a) of approximately 10 lm inlength.

Seminal receptacleThe inseminatory pore joins the seminal receptacle via a slightly chitinizedinseminatory canal (Figure 5). The seminal receptacle is situated dorsallyabove the rectal cavity. It is an oval sac, up to 50 lm in length and 40 lm inwidth when filled with sperm and lying laterally at the left side of the body(Figure 6b). In gravid females, as observed in this study, the receptacle is filledwith spermatozoa (Figure 6c). For this reason, epithelial cells appear either

Figure 5. Schematic dorsal (a) and sagittal (b) drawings of the female genital system of P. ovis.

A = anal slit; ED = efferent sperm duct; Gn = gnathosoma; IC = inseminatory canal; IP

= insemination pore; Od = oviduct; Op = ovipositor; Ov = ovary; SR = seminal receptacle;

U = uterus; I, II, III, IV = legs. (scale = 100lm).

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flattened, with electron-dense cytoplasm extensions inserted between sperma-tozoa. They seem organized in a monolayer. Spermatozoa do not seem to differfrom those observed in the male genital tract. The chromatin threads measure30 – 50 nm in width and the mitochondria are 90 – 130 nm wide.

OvariesThe ovaries are paired and located on both sides of the digestive tract, beneaththe rectum (Figure 6d). Each ovary is surrounded by a flattened epitheliumwhose cells are rich in mitochondria (not illustrated). It is bordered by muscles.Anterodorsally, they contain a large 50 lm central cell surrounded by the germcells (Figure 6d). The central cell contains a large multilobular nucleus withnumerous clumps of condensed chromatin and nucleoli. On histological serialsections, up to 15 lobuli, of 8 – 11 lm in diameter, are visible (Figure 7a). Thecenter of the nuclei always harbors a large area of electron-dense chromatinmass surrounded by an electron-lucent area. The nuclear membrane presentsnumerous folds where moderate electron-dense material accumulates(Figure 7a). The cytoplasm is finely granular and rich in free ribosomes. Feworganelles such as Golgi apparatus, polysomes and mitochondria are visible inthe periphery. Germ cells of different sizes are connected with the central cellvia nutritive cords (stalks) (Figures 6d and 7b). These stalks are filled withgranular electron-dense rods (Figure 7c). Three developmental stages of germcells can be distinguished and each germ cell is surrounded by somatic cells.The first ones are small ovoidal cells, 4 – 6 lm in diameter, containing a nucleusof approximately 3 lm in diameter, numerous mitochondria (0.5 – 0.7 lm indiameter), free ribosomes and Golgi complexes (Figure 7d). They are to beregarded as oogonia; sometimes, in their cytoplasm, filaments of unknowncomposition and function can be observed. The cells were closely linked andsome junctions were also observed between them. The second type is repre-sented by cells of 8 – 10 lm in diameter, containing a nucleus of approximately5 lm in diameter, dispersed mitochondria (0.7 – 1.2 lm in diameter), numerousfree ribosomes and dispersed Golgi elements (Figure 7b). They are consideredoocytes. The nucleus of these two types of cells shows a dispersed chromatinwith electron-dense aggregates at the periphery. The third type is representedby the early vitellogenic oocytes (Figure 6d). They are characterized by anundulating outline and a swelling of their cytoplasm that becomes granular andelectron-dense due to the presence of great amounts of free ribosomes. Theirregular nucleus is electron-lucent with electron-dense chromatin patches inthe periphery. At this stage, the cells detach from the central cell and move tothe oviducts where the vitellogenesis takes place.

OviductsThe oviducts arise from the ventral part of the ovaries, at the level where theefferent sperm ducts terminate (Figure 5). The oviduct’s wall consists of amonolayer epithelium, with little electron-dense Golgi elements and sur-rounded by muscles, that becomes flattened when an oocyte passes through

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(not illustrated). The oocyte in vitellogenesis increases in size (up to 90 – 150 lmin diameter) and contains a nucleus similar to the one of previtellogenic oocyte(Figure 7e). It contains also two types of granules, electron-dense ones of 1 –8 lm in diameter and electron-lucent ones of 0.3 – 0.9 lm in diameter formingthe yolk. Cellular organelles are observed in the peripheral cytoplasm. Asobserved in proximal and distal parts of the oviduct, the vitelline envelopeappears homogenous and lucent, and measures 0.2 – 0.3 lm in thickness.

UterusThe uterus or chorion gland is situated at the junction of the two oviducts andis a sac-like organ which lies dorsally to the terminating oviducts (Figure 5).The epithelial cells of this organ are prismatic with apical microvilli and aretightly connected together at the apical area with gap junctions and zonulaoccludens (Figure 4e). At their basal part, these cells contain a large sphericalnucleus of approximately 3 lm in diameter, with patches of condensed chro-matin, numerous dispersed mitochondria, rough endoplasmic reticulum andGolgi complexes (Figure 4d). The apical two-third of the cells is filled withnumerous secretion granules up to 1 lm in diameter. The granules are electron-lucent and merge by groups of two or three before entering an exocytosis event.After secretion, they give rise to the egg chorion. The uterus lumen is narrowwhen no egg is present. Circular muscles are attached to the outside part of thegland. They are responsible for propelling the eggs toward the ovipositor.

Ovipositor and oviporusThe ovipositor consists of a broad, thinly sclerotized tube underlaid by a thinepithelium (not illustrated) and terminates at the oviporus. The lumen is broadin the females which have already laid eggs and the terminal section of the tubecan be evaginated to form a short tube (Figure 6a).

Discussion

The male and female genital tracts of Psoroptes spp. are very similar to thosedescribed in other Astigmata. Nevertheless, differences exist in their organi-zation or structure.

Figure 6. (a) SEM of P. ovis female, ventral view; insert = insemination pore and anus at higher

magnification; (b–c) semi-thin section and TEM of the seminal receptacle of P. ovis female; (d)

semi-thin section (transversal view) of the organization of the ovaries and oviducts of P. ovis

female. A = anal slit; C = cuticule; CC = central cell; Ep = epithelium; EVOc = early

vitellogenic oocyte; Gn = gnathosoma; IP = insemination pore; Oc = oocyte; OcV = oocyte

in vitellogenesis; Od = oviduct; Op = ovipositor; Ov = ovary; R = rectum; SR = seminal

receptacle; Sz = spermatozoa; I, II, III; IV = legs; * = stalk.

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In the majority of astigmatic mites, testes are paired. In Psoroptes, like inDermatophagoides (Walzl 1992) and Notoedres (Witalinski 1988), the testes arelinked by a narrow bridge filled with spermatogonia. In these species, and alsoin Sarcoptes (Witalinski and Afzelius 1987), the testes are surrounded by aperipheric syncytium in opposition with the central cell observed in someAcaridae (Prasse 1968; Heinemann and Hughes 1970, Vijayambika and John1975; Alberti 1980; Witalinski et al. 1990). Spermatogenesis occurs from theposterior end to the anterior part of the testis and mature spermatozoa arestored in the vasa deferentia. One of the steps occurring during spermato-genesis includes the formation of spongy layers. Various functions have beensuggested for this layer (Witalinski et al. 1986). In Psoroptoidea, spongy layersseem to be involved in the extrusion of excessive cytoplasm (Witalinki andAfzelius 1987; Witalinski 1988).

Spermatozoa, as described by Alberti (1980, 1984), and generally recognizedin the Astigmata, are characterized by a polygonal form, absence of flagellumand lack of an acrosome. This latter characteristic can be explained by the earlypenetration of the spermatozoa in the previtellogenic oocytes, before they ac-quire an egg envelope (Witalinski et al. 1986). The nuclear material is orga-nized in a form of chromatin threads not limited by a nuclear membrane. Thepartial or total disappearance of the nuclear envelope during spermatogenesisis a characteristic of the Actinotrichida (Alberti 1980). The thickness of thechromatin threads of Psoroptes is similar to the values measured in Acarus siro(30 – 35 nm) and Tyrophagus putrescentiae (42 – 45 nm) (Witalinski et al. 1986).But they are much thinner than those recorded in Sarcoptes (110 – 120 nm)(Witalinski and Afzelius 1987), although the two species are phylogeneticallycloser. The cytoplasm contains lamellae the role of which is unknown. Theywere observed in the spermatozoa of all acarid mites investigated so far andprobably originate from ER cisternae (Witalinski et al. 1986). As recorded inSarcoptes (Witalinski and Afzelius 1987), no morphological modifications havebeen observed in the mitochondria during the spermatogenesis as described inNotoedres cati (Witalinski 1988), A. siro and T. putrescentiae (Witalinski et al.1986).

Only a few accessory glands are present in Astigmata. They may be paired(one or two pairs) or single organs (Evans 1992; Witalinski and Walzl 1995) asobserved in Psoroptes. This mite does not produce a spermatophore like some

Figure 7. (a–e) Various TEM aspects of the ovary and germ cells of P. ovis female; (a) central cell

in the ovary surrounded by oogonia and oocytes; (b–c) oocyte and stalk; (d) oogonium; (e) oocyte

in vitellogenesis in the oviduct. CC = central cell; Cy = peripheral cytoplasm; EM = accumu-

lation of extranuclear material; F = fibrillae; Mi = mitochondria; N = nucleus; NLo = nuclear

lobulus; Oc = oocyte; OdE = oviduct epithelium; Og = oogonium; OvE = ovary epithelium;

SC = somatic cell; St = stalk; VE = vitelline envelope; Y = yolk; * = patches of electron-dense

material.

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Acaridae or Glycyphagidae and the secretions of the accessory gland flow up tothe ejaculatory duct probably to form the sperm fluid.

The ejaculatory duct is lined with a thin cuticule and the end of the ductis protrusible. Like in other Astigmata, the terminal section of this duct isenclosed in a strongly sclerotized intromittent organ, the aedeagus or peniswhich is situated between the bases of legs III and IV. It is directed posteriorlywhen extruded. Like some Acaridia (Rhizoglyphus, Acarus) and Psoroptidia(Dermatophagoides, Listrophorus), Psoroptes has a pair of adanal copulatorysuckers and two pairs of small suckers on the tarsi IV which increase theattachment to the female during pairing.

In the female genital tract of the Astigmata, a sperm access system uncon-nected with the genital orifice is present and acts for the reception of sperm(and in some cases also the maturation and storage) (Evans 1992). In fact, thesperm is not introduced through the primary genital orifice but through aninsemination pore (bursa copulatrix) reaching the seminal receptacle by asimple cuticule-lined inseminatory canal. P. ovis does not depart from the rule.Prasse (1970) described the wall of the seminal receptacle of another astigmaticmite, Caloglyphus, as an epithelial layer with cells having large nuclei and longmicrovilli. This description is similar to the structure observed in Psoroptes,A. siro (described with more details in Witalinski et al. 1990) and Dermato-phagoides (Walzl 1992). Spermatozoa contained in the receptaculum seminis ofPsoroptes female do not seem to differ from those found in the male genitaltract. This suggests that a capacitation phase is lacking in Psoroptes species,although it is mentioned in A. siro (Alberti 1980) and N. cati (Witalinski 1988)and usually observed in ticks and other mites (Evans 1992; Alberti and Coons1999). Another characteristic of the Astigmata is the presence of paired ovaries,that are found in both Acaridia and Psoroptidia, in contrast to the Prostigmataand Oribatida (Evans 1992). Each ovary is connected by a narrow duct to themedian sac-like seminal receptacle of the sperm access system. These ductshave not been observed in Psoroptes despite the number of mites consideredbut it can be supposed that they would be similar to those described in otherastigmatic mites. The ovaries are composed of a central cell as observed in A.siro (Witalinski et al. 1990), Dermatophagoides (Walzl 1992) and, maybe,Caloglyphus mycophagus where Heinemann and Hughes described a centralstructure (1970). This central cell contains a multilobular nucleus like in A. siroand is not a syncytial cell like in Dermatophagoides since connections have beenobserved between lobuli. The central cell is connected with the oogonia andoocytes by nutritive cords. This feature has also been observed in Varroajacobsoni by Alberti and Hanel (1986) and Alberti and Zeck-Kapp (1986). Inthis species, the nutritive cords originate in a syncytial nutritive tissue and areresponsible for the transport of mainly ribosomes and mitochondria tooocytes. The role of this central cell is not fully known but it could play somepart in the organization of the ovary or in the nutrition and induction ofmaturation of the oogonia/oocytes. It would be interesting to perform bio-chemical studies and to examine nymphs to elucidate the development of this

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structure and understand better its relationship with the germ cells. This cell, asessential for mite reproduction, could become a potential target of new selec-tive acaricides, as happens with molting hormones in other arthropods(Halliwell and Carlotti 1998). Germ cells attached to the central cell wereconsidered to be oogonia and oocytes, according to their size, the volume oftheir cytoplasm and the number of obvious organelles. In fact, Walzl (1992)described oocytes of different sizes as being germ cells in Dermatophagoides buthe did not describe oogonia in the ovaries. Witalinski et al. (1990), for theirpart, indicated that oogonia and oocytes of Acarus are connected to the centralcell, but they did not supply any detailed description of these two cell types.The germ cells are surrounded by somatic cells that escort them until theprevitellogenic stage as described in Dermatophagoides (Walzl 1992). Theyprobably play a role in the maturation and in the induction of vitellogenesisbut detailed biochemical studies should be carried out to confirm thishypothesis.

Fertilization takes place before the passage of the oocyte through the oviductand before the vitellogenesis (Prasse 1968; Witalinski et al. 1986). Unfortu-nately, no accurate data are available on the mode of vitellogenesis in the genusPsoroptes. By analogy with observations realized in other Psoroptidia (Wita-linski 1993), it can be supposed that the vitelline envelope is produced by theoocyte itself and that exochorion, proposed to be an adhesive layer which fixedthe eggs to the substratum, is of uterus (chorion gland) secretion. No secretoryaspect was observed in the oviduct of Psoroptes. Since all mites examined weregravid females whose oviducts were filled with an oocyte in vitellogenesis, theepithelium was flattened. Oviducts may have, in fact, a secretory role andvitellogenesis may take place or be completed there as described in otherAcaridida (Witalinski et al. 1990; Walzl 1992; Desch 1998). In contrast, theuterus is well developed and contains numerous secretory granules. It joins thegenital orifice through the ovipositor of ectodermal origin. Its epitheliumcomprises flattened cells which secrete the thin, pliable cuticule lining the lu-men. Like in Acaroidea (Acarus, Lardoglyphus, Tyrophagus), this duct iscapable of considerable expansion to accommodate the large egg (approxi-mately 115 · 260 lm in Psoroptes) during its passage to the exterior. Theanterior region of the ovipositor in many Acaroidea forms an egg-laying organ.In its simplest form, as seen here, the anterior region of the genital tract iseverted by the egg and forms a short tube.

In conclusion, the present study provides additional information for theclassification of Psoroptes spp., assuming that the observations made on P. oviscan be transposed to Psoroptes from other hosts. In fact, this study demon-strates that the male of Psoroptes is more similar to Dermatophagoides than toSarcoptes or Notoedres, even through it is placed in the same superfamily. Thisis also true as far as the female is concerned; morphologically it appears to becloser to Dermatophagoides than to Knemidocoptes. Consequently, one couldargue that the genus Psoroptes is more closely related to the Pyroglyphidaethan is the family Sarcoptidae. Witalinski and Afzelius (1987) even consider

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that the Psoroptidae are more closely related to the Acaridia than are theSarcoptidae, and this view is strengthened by the presence of adanal suckers inthe male. However additional information on the female genital tract of Not-oedres and on the male genital tract of Knemidocoptes, are needed to validatethis assertion. This could also be confirmed by performing studies on theoogenesis and spermatogenesis in Psoroptes.

Acknowledgements

We want to thank Dr. D. Gridelet-de Saint Georges (University of Louvain-la-Neuve, Belgium) for her help in mites phylogeny and Mr. Y. Houbion(Facultes Universitaires Notre-Dame-de-la-Paix of Namur, Belgium) for histechnical support in SEM. This work was financially supported by the Min-istere de la Sante Publique, Brussels, Belgium (Convention S-6085).

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