The structure of the egg-shell of Porrocaecum ensicaudatum (Nematoda: Ascaridida)

Wharton D. A. 1979. The structure of the egg-shell of Porrocaecum enslcaudatum (Nematoda: Ascaridida). International Journal for Parasltology9: 127–131. The egg-shell of Porrocaecum ensicaudatum is oval with an opercular plug at either end. The shell consists of three layers: an inner lipid layer, a middle chitinous layer and an outer vitelline layer. The vitelline layer has strands of particulate material attached to its outer surface. The chitinous layer consists of 8.5 nrn fibrils which are made up of a chitin microfibril core surrounded by a protein coat. The fibrils are oriented randomly or in parallel, there being no indication of helicoidal architecture.The chitinous layer varies in thickness to form a pattern of interconnecting ridges on the surface of the egg. This pattern presumably increases the shell's structural strength.     

The egg-shell of Labiostrongylus eugenii (nematoda,strongyloidea): Structure and function

The egg-shell of Labio-strongylus eugenii consists of three layers; an outer vitelline layer, a middle chitinous layer and an inner lipid layer. The presence of chitin and protein in the middle layer was demonstrated by the use of chitinase and differential staining. The lipid layer was found to be made up of two layers, the innermost having large globules on its outer face. The shell was found to be permeable to liquid water, as demonstrated by the penetration of vital dyes. Eggs were able to develop normally with little or no loss of volume during periods of moisture stress when either osmotic or suction pressures were applied. The survival of eggs, as measured by hatching success, over periods of time at a range of saturation deficits and osmotic pressures was measured, and compared with known survival rates for other nematode species. The problems of working with nematode species whose life cycles have not been established are discussed. Possible survival strategies for Australian strongylids during periods of moisture stress are outlined.     

Cold tolerance of hatched and unhatched second stage juveniles of the potato cyst-nematode Globodera rostochiensis

Perry R. N. and Wharton D. A. 1985. Cold tolerance of hatched and unhatched second stage juveniles of the potato cyst-nematode, Globodera rostochiensis. International Journal for Parasitology15: 441–445. Hatched second-stage juveniles of Globodera rostochiensis can survive sub-zero temperatures by supercooling when not in contact with water. When frozen in water free juveniles cannot survive ice seeding across the cuticle and concomitant freezing of their body contents. Unhatched juveniles can survive in water, probably because the egg-shell protects the juvenile from ice seeding across from the medium; in this state juveniles survive by supercooling.     

Oogenesis and egg-shell formation in Aspiculuris tetraptera Schulz (Nematoda: Oxyuroidea).

The ovary of Aspiculuris tetraptera has a prominent terminal cap cell. This is considered to be part of the ovarian epithelium. Oogonia detach from the short rachis and increase in size from 6 to 60 microns; accumulating hyaline granules, shell granules and glycogen. The hyaline granules persist in the eff cytoplasm after shell formation has been completed and are considered to be lipoprotein yolk. The shell granules contribute to the non-chitin fraction of the chitinous layer. A classification of the cytoplasmic inclusions of the nematode oocyte is proposed. Upon fertilization a vitelline membrane is formed which constitutes the vitelline layer of the egg-shell. The chitinous layer is secreted in the perivitelline space, between the vitelline layer and the egg oolemma. Upon completion of chitinous layer synthesis, the egg cytoplasm contracts away from its inner surface. The material of the lipid layer is secreted at the surface of the egg cytoplasm and adheres to the inner surface of the chitinous layer. During secretion of the chitinous and lipid layers by the egg cytoplasm, the uterine cells secrete the unit membrane-like external uterine layer and the crystalline internal uterine layer. A complex system of interconnecting spaces develops in the internal uterine layer. This system is open to the exterior via breaks in the external uterine layer. There is no direct involvement of the uterine cells in the formation of this structure.     

Heterodera glycines: eggshell ultrastructure and histochemical localization of chitinous components

The eggshell in most nematodes consists of an outer vitelline layer, a middle chitinous and an inner lipid layer. Earlier work with eggs of Heterodera glycines suggests the presence of two chitinous layers but the vitelline layer was not observed. From our observation the outer chitin layer described in past literature is actually a vitelline layer. Histochemical analysis has demonstrated that chitin is absent from the outer envelope. Electron microscope observations of the eggshell show a waxy appearance and osmium staining consistent with that of the proteinaceous vitelline layer found in other nematodes. Lectin localization also shows that the eggshell continues to develop past fertilization with the delivery and integration of eggshell precursors. Contrary to previous reports, we propose that the ultrastructure of the eggshell H. glycines follows the common three-layer structure observed in other nematodes.     

The structure and formation of the egg-shell of Syphacia obvelata Rudolphi (Nematoda: Oxyurida)

The egg of Syphacia obvelata is a flattened elipsoid. The egg-shell consists of 5 layers: external uterine layer, internal uterine layer, vitelline layer, chitinous layer and lipid layer. An operculum is present at one pole of the egg. The opercular groove consists of a break in the uterine layers and the modification of the chitinous layer by the deposition of lipoprotein material. On the curved side of the egg the uterine layers are modified to form alternate ridges and depressions. Discrete spaces are present in the internal uterine layer between the ridges. These are open to the exterior via pores in the external uterine layer. The structure of the uterine layers is quite different on the flattened side of the egg. The morphology of the reproductive system and the formation of the egg-shell is described. It is suggested that the complex structure of the uterine layers of oxyurids forms by a self-assembly process.     

Ultrastructural studies on the nematode Xiphinema diversicaudatum: Egg shell formation

The ultrastructure of the formation of the egg shell in the longidorid nematode Xiphinema diversicaudatum is described. Upon fertilization a vitelline membrane, which constitutes the vitelline layer of the egg shell, is formed. The chitinous layer is secreted in the perivitelline space, between the vitelline layer and the egg cell membrane. On completion of the chitinous layer, the material of the lipid layer is extruded from the egg cytoplasm to the outer surface, through finger-like projections. Both chitinous and lipid layers are secreted by granules in the egg cytoplasm that disappear as the layers are completed. Chitinous and lipid layers are formed during the passage of the egg through the oviduct. The vitelline layer is enriched with secretions produced by the oviduct cells and then by phospholipids secreted by the cells of the pars dilatata oviductus. The inner uterine layer is also formed by deposition of secretory products apposed on the egg shell in the distal uterine region and Z-differentiation. In the proximal part of the uterus, the egg has a discontinuous electron-dense layer, the external uterine layer. Tangential sections between chitinous and uterine layers revealed the presence of holes, possibly egg pores, delimited by the two uterine layers.     

Egg-shell calcium and the hatching of Globodera rostochiensis

The Ca²⁺ content of Globodera rostochiensis egg-shells was investigated by X-ray microanalysis. When intact eggs were treated with 5 mM sodium 1, 2-di (2-aminoethoxy) ethane-N,N,N',N',-tetra-acetate (EGTA) only part of the egg-shell Ca²⁺ was removed. Similarly treated opened egg-shells lost almost all their Ca²⁺. We think that Ca²⁺ of intact egg-shells which is accessible to EGTA is in the outer part of the shell and that the inaccessible Ca²⁺ is in the inner lipoprotein layer. Much Ca²⁺ was removed from opened egg-shells by the hatching agents Zn²⁺, La³⁺ and decationised potato-root exudate, but none by dilute acetic acid or Mg²⁺. Hatching agents, by binding to or replacing Ca²⁺, may change lipoprotein membrane structure. Eggshells treated with potato-root exudate contained about 3 times as much Ca²⁺ as untreated shells, because the treatment makes additional binding sites available. Our results suggest that three types of Ca²⁺ -binding sites occur in the egg-shell.     

Observations on the morphology of Toxocara pteropodis eggs

Fertile eggs of Toxocara pteropodis, passed in the faeces of juvenile flying-foxes, were ovoid to spheroid in shape with a diameter range of 80-110 microns. The shell was often seen to comprise 4 layers: a fine inner lipid layer, a thicker clear chitinous layer, an equally thick outer vitelline layer and a pitted outermost, proteinaceous uterine layer of variable thickness. Infertile eggs were less uniform in shape and generally did not have well-defined shell layers, the formation of which is triggered by sperm penetration of the oocyte. The eggs of this species are bulkier than those of related ascaridoids, apparently because of a thicker external coat which, while not providing mechanical strength, is thought to protect against desiccation. Scanning electron microscopical findings suggest that the outer layer is not applied directly by uterine cells, but forms by the gradual deposition of secretions in the uterine lumen, regardless of whether the oocyte has been fertilized.     

The Influence of Potato Cultivar on Lipid Content and Fecundity of Bolivian and British Populations of Globodera rostochiensis

The influence of host cultivar on the lipid levels provided by a female to her progeny was investigated with Oil Red O stain and a quantitative image analyzer. A population of Globodera rostochiensis was multiplied at Toralapa Field Station in Bolivia on 25 different potato cultivars grown in that country. The mean neutral lipid content of newly formed second-stage juveniles varied significantly with cultivar over a 200% range. The corresponding range was only 18% and 28% for the same Bolivian and a UK population of G. rostochiensis, respectively, when both completed reproduction concurrently on 10 pot-grown European cultivars in the United Kingdom. Egg numbers per female varied with host for Bolivian cultivars that lack known partial resistance to Globodera spp. There was a 15-fold range between the most and least fecund nematode-host combinations (Kosi and Gendarme). The Bolivian G. rostochiensis population showed only a 2-fold range in mean eggs per cyst when grown on European cultivars in the UK. The fatty acid profiles of lipids from Bolivian G. rostochiensis cysts reared on Bolivian potato cultivars were dominated by C₂₀ (37-64%) and C₁₈ (28-46%) fatty acids and ranged from C₁₄ to C₂₂. The three major fatty acids detected were C_20:4:, C_20:1, and C_18:1. Few differences between cultivars were observed. For a UK population of G. rostochiensis reared on ssp. tuberosum, higher relative percentages of C₁₈ and monounsaturated fatty acids and lower relative percentages of C₂₀ and polyunsaturated fatty acids were found.     

The role of eggshell and underlying vitelline membrane for normal pattern formation in the early C. elegans embryo

Summary The embryo of the nematode Caenorhabditis elegans is surrounded by an inconspicuous inner vitelline membrane and a prominent outer chitinous eggshell proper. We demonstrate that the complete removal of the chitinous eggshell does not interfere with successful development to yield a normal worm. The same result can be obtained when the vitelline membrane is penetrated with laser microbeam irradiation of only the eggshell proper, gently enough to permit its resealing after a while. However, when large holes are made into the eggshell the concomitantly penetrated vitelline membrane does not reseal. While early development is quite normal under these conditions, gastrulation is defective in that gut precursor cells do not migrate in properly, eventually leading to embryonic arrest. This suggests a crucial role for pattern formation of the micro-environment around the embryo preserved by the intact vitelline membrane. Removing both eggshell and vitelline membrane results in a string-like arrangement of founder cells and subsequent grossly abnormal cell patterns. Our experiments support the idea that the prominent eggshell proper just functions as a mechanical protection while the thin vitelline membrane directly or indirectly serves as a necessary control element affecting the positions of cells which to begin with are determined by the orientation of the cleavage spindle. Correspondence to: E. Schierenberg     

The chemical nature of the egg-shell of helminths. I. Absence of quinone tanning in the egg-shell of the liver fluke, Fasciola hepatica

Ramalingam K. 1973. The chemical nature of the egg-shell of helminths—I. Absence of quinone tanning in the egg-shell of the liver fluke, Fasciola hepatica. International Journal for Parasitology, 3: 67–75. The mode of stabilization of the egg-shell protein of Fasciola hepatica has been studied. It was observed that quinone tanning is absent in the egg-shell. Unlike quinone-tanned proteins, egg-shell proteins show auto-fluorescence. Investigation on the fluorescent compounds present in the egg-shell protein shows the presence of di-tyrosine. In addition to the presence of di-tyrosine, the egg-shell protein is stabilized by —S—S—bonding. The resistant properties of the egg-shell of F. hepatica are attributed to the presence of the above types of cross-links.     

Major Sperm Protein Genes from Globodera rostochiensis

Three genes in the major sperm protein (MSP) gene family from the potato cyst nematode Globodera rostochiensis were cloned and sequenced. In contrast to the absence of introns in Caenorhabditis elegans MSP genes, these genes in G. rostochiensis contained a 57 nucleotide intron, with normal exon-intron boundaries, in the same relative location as the intron in Onchocerca volvulus. The MSP genes of G. rostochiensis had putative CAAT, TATA, and polyadenylation signals. The predicted G. rostochiensis MSP gene product is 126 amino acids long, one residue shorter than the products in the other species. The comparison of MSP amino acid sequences from four diverse nematode species suggests that O. volvulus, Ascaris suum, and C. elegans may be more closely related to each other than they are to G. rostochiensis.     

Ultrastruktur-Untersuchungen zur Eischalenbildung bei Microdalyellia fairchildi (Turbellaria)

Zusammenfassung Die Zellen des Uterus von Microdalyellia fairchildi, die durch ein reichlich entwickeltes ER ausgezeichnet sind, produzieren ein bläschenartiges Sekret, das am Beginn der Eibildung ausgeschieden wird. Die innere Oberfläche der Zellen bildet zahlreiche, teils verzweigte lamellenartige Auffaltungen, die wechselseitig ineinandergreifen. Nach Einfließen von Eizelle und Dotterzellen in den Uterus verstreichen die Auffaltungen zu einer glatten inneren Begrenzung des Lumens. Anschließend werden zahlreiche Schalentropfen aus den Vesikeln der Dotterzellen freigesetzt. Sie fließen an der dünnen Sekretschicht der Uteruszellen, die die Innenseite des Uterus bedeckt, zu einer Eihülle ineinander. Bei geringen Mengen Uterussekret entsteht keine geschlossen einheitliche Schicht. Aus der anfänglich dünnen Eihülle geht durch Aufnahme weiterer Schalentropfen und anschließende Sklerotisierung die braune Eikapsel hervor.

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Ultrastructural studies of egg-shell formation in Microdalyellia fairchildi (Turbellaria)

Summary The uterus cells of Microdalyellia fairchildi which are rich in agranular ER produce a vesicular substance released from the cells at the beginning of egg-shell formation. The lumenal border of the cells is thrown into lamellae like and partly branched outfoldings which inter-digitate. After filling of the uterus with the oocyte and several vitelline cells the outfoldings are stretched to form a smooth inner wall. A plenty of shell droplets are set free by the vitelline cells. The droplets accumulate at the inner surface of the uterus and coalesce along a thin layer of uterus secretion. Lower amounts of uterus secretion do not achieve a uniform inner lining of the uterus wall. The initially thin capsule wall is thickened by fusing with further shell droplets and subsequently becomes resistent by sclerotization.

     

The structure of the egg-shell of Aspiculuris tetraptera Schulz (Nematoda: Oxyuroidea).

The egg of Aspiculuris tetraptera is an ellipsoid measuring 93 x 40 microns. The shell consists of 5 layers: the external uterine layer, internal uterine layer, vitelline layer, chitinous layer and the lipid layer. This nomenclature is based upon the formation and histochemistry of the shell layers. The internal uterine layer contains a system of interconnecting spaces, partly filled by uterine secretion, which open to the exterior of the egg via breaks in the external uterine layer. The surface of the egg is covered by a system of interconnecting grooves. Freeze-etching reveals that the internal uterine layer is open to the exterior via pores, which open into the grooves. Rod-shaped particles are also revealed in the external uterine layer. The operculum of the egg consists of a modification of the uterine and chitinous layers of the shell.     

Fine structure of the eggshell of Ommatissus binotatus Fieber (Homoptera,Auchenorrhyncha, Tropiduchidae)

The external morphology and fine structure of the eggshell of Ommatissus binotatus Fieber (Homoptera : Tropiduchidae) was investigated by light, scanning and transmission electron microscopy. The egg surface has 2 main regions: a specialized area and an unspecialized egg capsule. The specialized area is characterized by a large respiratory plate containing the operculum and a short respiratory horn. The latter consists of an external hollow tube and an internal coneshaped projection hosting a micropylar canal. The eggshell has 4 layers: the vitelline envelope, a wax layer, the chorion and an outer mucous layer. The chorion has inner, intermediate and outer parts. The functions of the different parts of the eggshell are discussed. Characters useful to define the eggs and the oviposition habit in the family Tropiduchidae were provided. The size and morphology of the egg, plate, respiratory horn and operculum are suggested as useful characters for ootaxonomic analysis.     

In situ incorporation of Fatty acids into lipids of the outer and inner envelope membranes of pea chloroplasts

When incubated with [1-¹⁴C]acetate and cofactors (ATP, Coenzyme A, sn-glycerol-3-phosphate, UDPgalactose, and NADH), intact chloroplasts synthesized fatty acids that were subsequently incorporated into most of the lipid classes. To study lipid synthesis at the chloroplast envelope membrane level, ¹⁴C-labeled pea (Pisum sativum) chloroplasts were subfractionated using a single flotation gradient. The different envelope membrane fractions were characterized by their density, lipid and polypeptide composition, and the localization of enzymic activities (UDPgalactose-1,2 diacylglycerol galactosyltransferase, Mg²⁺-dependent ATPase). They were identified as very pure outer membranes (light fraction) and strongly enriched inner membranes (heavy fraction). A fraction of intermediate density, which probably contained double membranes, was also isolated. Labeled glycerolipids recovered in the inner envelope membrane were phosphatidic acid, phosphatidyl-glycerol, 1,2 diacylglycerol, and monogalactosyldiacylglycerol. Their ¹⁴C-fatty acid composition indicated that a biosynthetic pathway similar to the prokaryotic pathway present in cyanobacteria occurred in the inner membrane. In the outer membrane, phosphatidylcholine was the most labeled glycerolipid. Phosphatidic acid, phosphatidylglycerol, 1,2 diacylglycerol, and monogalactosyldiacylglycerol were also labeled. The ¹⁴C-fatty acid composition of these lipids showed a higher proportion of oleate than palmitate. This labeling, different from that of the inner membrane, could result either from transacylation activities or from a biosynthetic pathway not yet described in pea and occurring partly in the outer chloroplast envelope membrane. This metabolism would work on an oleate-rich pool of fatty acids, possibly due to the export of oleate from chloroplast toward the extrachloroplastic medium. The respective roles of each membrane for chloroplast lipid synthesis are emphasized.     

Possible involvement of a sperm-associated body in the process of fertilization in quail

The present paper describes a novel structure, termed the sperm-associated body, which is found both in the lumen at the oviductal infundibulum and in the vitelline membrane of the ovum in the quail Coturnix japonica. The fully developed sperm-associated body, which is about 100 microm long, consisted of two parts; a core of concentric-circular appearance and a cortex of needle-like projections. The outer surface of the body was coated with CaCO3. The body was always accompanied by spermatozoa. About 70 sperm-associated bodies were observed in a single ovum. Electron-microscopically, small numbers of holes were detected in the vitelline membranes of a fertile ovum, and the sperm-associated bodies were always present in these holes. Frequently observed in the vitelline membranes was a disk speculated to be a portion of the inner layer of the membrane partially affected by spermatozoa. However, neither sperm-associated bodies nor spermatozoa were observed there. It was suggested that the sperm-associated bodies assist fertile spermatozoa in binding the inner layer of the vitelline membrane and penetrating it.     

Oogenesis of Cardiochiles nigriceps viereck (Hymenoptera : Braconidae): Histochemistry and development of the chorion with special reference to the fibrous layer

A fibrous layer on the surface of eggs of the parasitoid, Cardiochiles nigriceps (Hymenoptera : Braconidae), has been implicated by earlier studies in the evasion from encapsulation by host hemocytes. The present histochemical and ultrastructural study was undertaken to characterize fibrous layer material and to determine the source of fibrous layer and other components of the eggshell. The fibrous layer contains neutral glyco- or mucoprotein; acidic mucoproteins or glycosaminoglycans are absent. The mature eggshell is resolved into 5 morphologically distinct layers by electron microscopy: (from inner to outer) vitelline envelope, endochorion, an electron-dense “irregular layer”, papilliary layer and fibrous layer. During oogenesis each eggshell layer is laid down sequentially in the order mentioned above. Eggshell material appears to be produced by the follicle cells because these develop extensive rough endoplasmic reticulum and golgi apparatus and exhibit apparent exocytotic activity at the plasma membrane adjacent to the egg.     

Effects of Initial Nematode Density on Population Dynamics of Globodera rostochiensis on Resistant and Susceptible Potatoes

The influence of resistant and susceptible potato cultivars on Globodera rostochiensis population density changes was studied at different nematode inoculum levels (Pi) in the greenhouse and field. Soil in which one susceptible and two resistant cultivars were grown and fallow soil in pots was infested with cysts to result in densities of 0.04-75 eggs/cm³ soil. A resistant cultivar was grown in an infested field with Pi of 0.7-16.7 eggs/cm³ soil. Pi was positively correlated with decline of soil population densities due to hatch where resistant potatoes were grown in the greenhouse and in the field but not in fallow soil. However, Pi was not correlated with in vitro hatch of G. rostochiensis cysts in water or potato root diffusate. Under continuous culture o f a resistant cultivar, viable eggs per cyst declined 60-90% per plant growth cycle (4 weeks) and the number of cysts containing viable eggs had decreased by 77% after five cycles. The rate of G. rostochiensis reproduction on both resistant and susceptible cultivars was negatively correlated with Pi. These data were used to predict the effect of resistant and susceptible potato cultivars on G. rostochiensis soil population dynamics.