Nabil N. Youssef

The fine structure of the developmental stages of the microsporidian Nosema apis Zander

Schizonts, sporonts and sporoblasts of Nosema apis from honey bees collected in the summer and winter were studied with the electron microscope. The nuclei usually had a diplokaryon arrangement. Intranuclear spindles with polar vesicles were associated with division. Schizonts had a single limiting unit membrane, whereas sporonts had a two-layered wall. Sporonts from summer bees had only a thin single limiting membrane in some areas and evidence of endocytosis was sometimes seen in these. Sporonts from winter bees had branched tubular outpocketings from the wall. In sporoblasts, the development of the polar filament was closely associated with a network of dense structures interwoven with a system of tubules evidently of ER derivation; the Golgi complex was associated with this network.

Life Cycle of the Chalk Brood Fungus, Ascosphaera Aggregata, in the Alfalfa Leafcutting Bee, Megachile Rotundata, and Its Associated Symptomatology

ABSTRACTThe life cycle of Ascosphaera aggregata in Megachile rotundata and the symptomatology of the resultant disease are described. The formation of the ascogonium, and the process by which the a...

PRODUCTION OF MONOCLONAL ANTIBODIES SPECIFIC FOR EIMERIA TENELLA MICROGAMETOCYTES

A panel of 4 monoclonal antibodies specific for Eimeria tenella, the causative agent of cecal coccidiosis of birds in the genus Gallus, was produced by standard techniques. The indirect fluorescent antibody (IFA) test demonstrated specificity of these 4 antibodies for the microgametocytes. Hybridoma TIA3B9 secreted a monoclonal antibody of subisotype IgG2b that was used throughout the course of this study. Immunologic potency of this antibody was demonstrated by in vitro experiments that revealed a greater than 50% reduction in oocyst production, indicating an apparent inhibition of fertilization.

In vivo development and pathogenicity of Ascosphaera proliperda (Ascosphaeraceae) to the alfalfa leafcutting bee, Megachile rotundata

Abstract First instar larvae of the leafcutting bee, Megachile rotundata , were fed on either artificial or natural provisions containing spores of Ascosphaera proliperda . Two isolates were used as a source of inocula: one originated from in vitro isolates obtained while culturing what was thought to be pure spores of A. aggregata , the second originated from in vitro cultures from Denmark. Histological and scanning electron microscopy studies revealed that the spores germinated in the gut lumen and the developing hyphae invaded all tissues, after which they penetrated through larval integument and began the sexual phase of the life cycle aerially. Virtually all fungus-exposed larvae developed symptoms of disease regardless of source of inoculum, type of provision, and spore dose (1.5 × 10 3 to 3 × 10 6 ) per insect. It was concluded that the fungus was pathogenic to the alfalfa leafcutting bee under laboratory conditions and future studies should be conducted to determine its etiology, cross infectivity, and natural distribution in other bee taxa.

Fine structural aspects of the developing compound eye of the honey bee, apis mellifera L.

We studied the postembryonic development of the compound eye of the worker honey bee on the light and electron microscopy levels. During the late third and early fourth larval instars the cells of the optic primordium became organized into preommatidial cell clusters. Each cluster contained four prospective cone cells, nine prospective retinula cells (of which the ninth was centrally located and shorter than the other eight), and an undetermined number of prospective pigment cells. During ommatidial differentiation that was characterized by ommatidium and crystalline cone elongation, the ninth retinula cell moved laterally, and each retinula cell elaborated a rhabdomer. Possible roles of membrane junctional specializations and cytoplasmic microtubules in the processes of differentiation are discussed.

Biology and pathogenicity of the microsporidian Perezia hyperae sp. n., a pathogen of the alfalfa weevil, Hypera postica

Abstract Taxonomic identification, biology, and pathogenicity of a microsporidian recovered from a laboratory culture of the western variety of the alfalfa weevil, Hypera postica , were studied by means of light microscopy. The name Perezia hyperae sp. n. has been proposed for the organism. The developmental stages of the pathogen were characterized on the basis of cell division, nuclear arrangement, and measurements. The complete life cycle, spore-to-spore, including two stages of schizogomy, had a duration of about 100 hr at 22°C and 80% RH in experimentally inoculated fourth instar larvae. In some larvae, the infection induced the formation of irregularly shaped bulges and a reduction in silk production necessary for cocoon formation. However, adults developed from infected larvae did not show corresponding abnormalities. Malpighian tubules were found to be the primary site of infection in the larvae. In later phases of infection the microsporidian was found in the intersegmental muscles, abdominal nerve cords, epidermal cells of the integument, fat bodies, and tracheal systems of the larvae, but not in their alimentary canals.

Unique cell wall abnormalities in the putative phosphoinositide phosphatase mutant AtSAC9.

SAC9 is a putative phosphoinositide phosphatase in Arabidopsis thaliana involved in phosphoinositide signaling. sac9-1 plants have a constitutively stressed phenotype with shorter roots which notably accumulate phosphatidylinositol 4,5-bisphosphate and its hydrolysis product inositol trisphosphate. We investigated the primary roots of sac9-1 seedlings at the cytological and ultrastructural level to determine the structural basis for this altered growth. Despite the normal appearance of organelles and cytoplasmic elements, our studies reveal extreme abnormalities of cell wall and membrane structures in sac9-1 primary root cells, regardless of cell type, position within the meristematic area, and plane of section. Cell wall material was deposited locally and in a range of abnormal shapes, sometimes completely fragmenting the cell. Simple protuberances, broad flanges, diffuse patches, elaborate folds, irregular loops and other complex three-dimensional structures were found to extend randomly from the pre-existing cell wall. Abundant vesicles and excessive membrane material were associated with these irregular wall structures. We argue that a perturbed phosphoinositide metabolism most likely induces these observed abnormalities and hypothesize that a disorganized cytoskeleton and excessive membrane trafficking mediate the cell wall defects.

Development in cell cultures of Eimeria vermiformis Ernst, Chobotar and Hammond, 1971.

SummaryDevelopment of Eimeria vermiformis from sporozoite to mature first-generation schizonts in cultured bovine kidney cells, Madin-Darby bovine kidney cells, and primary cultures of whole mouse embryos is described. Intracellular sporozoites were seen at 5 min, and for as long as 120 h after inoculation. Sporozoites were observed penetrating cells, with uninucleate trophozoites and immature schizonts with 2–6 nuclei first appearing 24 h after inoculation. Schizonts with 6 or more nuclei, as well as mature schizonts containing first-generation merozoites, were first seen between 36 and 48 h after inoculation of all 3 cell types used. The first indication of merozoite formation was determined by the appearance of small protuberances of cytoplasm at the periphery of schizonts. Merozoites began development at the periphery of schizonts and were later observed radiating from a central body of cytoplasm, 14–20 merozoites being formed. Some mature schizonts retained a small spherical residual body after merozoite formation was completed. After the rupture of schizonts, intracellular merozoites, which contained anterior and posterior refractile granules, were seen at 48, 72 and 96 h postinoculation. Merozoites were not seen entering or leaving cells. No further development was observed.

Fine structure of the intersegmental membrane glands of the sixth abdominal sternum of female Nomia melanderi (Hymenoptera, Apoidea)

The fine structure of the intersegmental glands of the sixth abdominal sternum in 1‐week old females of Nomia melanderi is presented. The plasma membrane of the secretory cell is unfolded in many places and is covered by a basement membrane. The microvillous surface is invaginated to form a rather long sinuous cavity. The endoplasm is almost entirely filled by secretory granules. Many secretory granules are located close to the inner surface of the invaginated plasma membrane. The invagination contains a porous ductule, apparently of cuticulin origin, that is connected directly with the inner layer of the transport duct of the duct‐forming cell. This type of arrangement allows the direct flow of the secretory substance to the outside in a continuous way. The cylindrical duct‐forming cell, besides having typical cell organelles, contains a cuticular transport duct. This duct is composed of a thin cuticulin layer surrounded by a rather thick epicuticular one. The results suggest that the secretory cell has two secretory cycles. The first occurs while the gland is differentiating (at the pupal stage) and is involved in secretion of the cuticulin that forms the porous ductule. The second cycle, which starts by the beginning of nesting, is involved in the secretion of a substance that is carried to the outside via the transport duct of the duct‐forming cell.

A multi-gene phylogeny provides additional insight into the relationships between several Ascosphaera species

Ascosphaera fungi are highly associated with social and solitary bees, with some species being pathogenic to bees (causing chalkbrood) while others are not, and proper identification within this genus is important. Unfortunately, morphological characterizations can be difficult, and molecular characterizations have only used one genetic region. We evaluated multiple phylogenies of the Ascosphaera using up to six loci: the Internal Transcribed Spacer (ITS) region, 18S rRNA, 28S rRNA, Elongation Factor-1α (EF-1α) the RNA polymerase II largest subunit (RPB1), and the second largest subunit (RPB2). The ITS sequence alone produced an inadequate phylogeny, and the addition of both the 18S and 28S rRNA loci to the ITS sequence produced a phylogeny similar to that based on all six genetic regions. For all phylogenies, Ascosphaera torchioi was in a separate clade that was the most basal, with a strong genetic similarity to Eremascus albus, introducing the possibility of paraphyly within Ascosphaera. Also, based on this new phylogeny, we now suggest that the Apis mellifera (honey bee) pathogens arose within a group of saprophytes, and the Megachile (leafcutting bees) pathogens arose separately.