The ultrastructure of spores (Protozoa: Microsporida) from Lophius americanus, the angler fish

Spinal and cranial ganglia of American angler fish, Lophius americanus, are often infected with microsporidia. This protozoon elicits the formation of large, spore-filled, hypertrophied host cells, cysts. Previous reports of microsporidia in European lophiids identify the parasite as Spraguea lophii, a genus which has recently been shown to be dimorphic. The spores from L. americanus are monomorphic (2.8 X 1.5 micron) and uninucleate. Each spore contains a polar tube that forms six to nine coils. Spraguea lophii differs from the microsporidium described in L. americanus in several ways. Spraguea lophii has two spore types: a large spore (4.0 X 1.25 micron) containing a diplokaryon and three to four polar tube coils and a smaller uninucleate spore (3.5 X 1.5 micron) with five to six polar tube coils. Because of these major differences, the microsporidium from L. americanus is removed from the genus Spraguea and returned to its original genus, Glugea, as a new species, G. americanus n. sp. Other ultrastructural characteristics of G. americanus are included: the posterior vacuole encloses two distinct membranous structures; one is tubular and resembles a "glomerular tuft" and the second is lamellar and composed of concentric membrane whorls, additionally, the straight or manubroid portion of the polar tube proceeds beyond the posterior vacuole before it turns anteriorly and begins to coil.     

New Family,Genus, and Species of Microsporida (Protozoa: Microsporida) from the Tropical Fire Ant,Solenopsis geminata (Fabricius) (Insecta: Formicidae)*

SYNOPSIS. A new species of Microsporida, Burenella dimorpha sp. n., representing a new family, Burenellidae fam. n. and genus, is described on the basis of light- and electron-microscope observations. The family is characterized by 2 sequences of sporogony, each sequence having morphologically different sporonts and spores. The parasite infects the tropical fire ant, Solenopsis geminata (Fabricius), producing distinct pathologic manifestations (clearing of the cuticle and eye malformation) and death in the pupal stage of development. Transmission of the infection per os to healthy S. geminata, to the Southern fire ant, Solenopsis xyloni McCook, and to the red and black imported fire ants, Solenopsis invicta Buren and Solenopsis richteri Forel, is reported.     

The Ultrastructure of Encephalitozoon cuniculi (Microsporida,Nosematidae) and its Taxonomic Significance*

SYNOPSIS. This study augments our knowledge of several ultrastructural features of Encephalitozoon cuniculi and provides evidence that this species is disporous. We support Cali's view that Encephalitozoon is distinct from Nosema and should be treated as a valid genus. We compare these with 2 other disporous genera, Glugea and Perezia, and conclude that Glugea is also distinct but Perezia is a junior synonym of Nosema.     

The Effect of Ultraviolet Radiation on the Germination of Nosema algerae Vávra and Undeen (Microsporida: Nosematidae) Spores1

Spores of Nosema algerae Vávra and Undeen were subjected to various dosages of 254 nm ultraviolet radiation (UV). Very high dosages of UV were required to block germination. Germination was normal immediately after UV dosages of 0.2 to 1.0 J/cm², followed by a delayed effect in which both percentage germination and the intrasporal concentration of trehalose decreased with time after UV exposure. Although a few spores were germinated, most of them were inactivated (rendered temporarily unable to germinate) by exposure to UV of 1.1 J/cm². Ultraviolet radiation between 1.1 and 3.4 J/cm² stimulated spores to germinate. However, spores were completely unable to germinate immediately after exposure to dosages above 3.8 J/cm². Ammonia had little effect on stimulation by UV but was inhibitory to germination after stimulation had occurred. These results demonstrate that UV behaves like a germination stimulus and are discussed in terms of the hypothesis that germination is initiated by the breakdown of barriers between trehalose and trehalase.     

Pleistophora oncoperae sp.n. (Protozoa: Microsporida) from Oncopera alboguttata (Lepidoptera: Hepialidae) in Australia

Pleistophora oncoperae sp.n. is described from adults and larvae of Oncopera alboguttata and O. rufobrunnea. The main site of infection was muscle, though fat body and connective tissue were also infected. Fresh pansporoblasts measured about 25 μm in diameter and contained 16 to 32 or more spores with a mean size of 5.9 × 3.1 μm. Macrospores measuring 7.7 × 4.4 μm were also seen. The mean polar filament length was 158 μm; ultrastructural studies showed that the filament is normally arranged in 14 coils (range, 13 to 20) at an angle of 53.5° to the axis of the spore. The species was found to be distinct from all previously described Pleistophora reported from Lepidoptera.     

Experimental infection of American winter flounder (Pseudopleuronectes americanus) with Glugea stephani (Microsporida)

Young-of-the-year Pseudopleuronectes americanus were captured in Sandy Hook Bay, New Jersey and maintained in aquaria at 16–19° C for 115 days in 1982 and 127 days in 1983. Half of them were experimentally exposed to Glugea stephani and the others were sham exposed. During the course of these experiments fish from both groups died as a result of G. stephani infection, demonstrating Glwgea -induced mortality in this species. Infections were found in both control and experimentally exposed fish. Mortality of Glwgea -infected fish in the exposed group was 63% while that in the controls was 28.5%.     

Ultrastructure and Cortical Morphogenesis in the Euplotine Hypotrich Certesia quadrinucleata Fabre-Domergue, 1885 (Ciliophora,Protozoa)1

The cortical development during cell division and the interphase ultrastructure of the marine interstitial hypotrich Certesia quadrinucleata is described using light microscopy and both scanning and transmission electron microscopy. Membranelles are paramembranelles; postciliary microtubules from rightmost membranellar kinetosomes line the buccal cavity and separate parallel arrays of pharyngeal discs that border the cytopharynx. A large paroral membrane is present; an endoral membrane is absent. Alveolar plates lie within alveolar membranes except in regions where organelles and organellar complexes (cirri, the condylopallium, dorsal bristles, membranelles, and the paroral membrane) emerge from the cortex. Muciferous-like bodies attach to the plasma membrane in these regions. Dorsal bristles possess transverse and postciliary microtubules as well as kinetodesmal fiber like those of other hypotrichs. Lasiosomes are present. A unique bulbous structure—the condylopallium—protrudes from the anterior right of the cell. The morphogenetic pattern is euplotine in that cortical development begins in one latitudinal zone, and the oral primordium of the opisthe develops within a subsurface pouch apart from the frontal primordia. Microtubular bundles appear beside (later attached to) developing frontal anlagen; they disappear after cirri are in final interphase locations. Although possessing unique characters, Certesia shares a close phylogenetic relationship with Euplotes.     

Fine Structure of Developing Spores of Thelohania bracteata (Strickland, 1913) (Microsporida,Nosematidae) Emphasizing Polar-Filament Formation*

SYNOPSIS. In the microsporidian, Thelohania bracteata, the polar filament, as it starts to develop in the sporoblast, apparently receives material synthesized by the granular endoplasmic reticulum and Golgi vesicles. In immature spores many dilated sacs are observed in areas where there is less endoplasmic reticulum. These sacs, that persist into the almost mature spore, are probably Golgi-type vesicles and may be related to the formation of the spore coat. The polar filament of the mature spore possesses 8 coils and in cross section or cross-fractured face the electron-dense central portion of the polar filament contains a tubular structure, ringed by 12–14 cylindrical structures. In thin sections, an electron-lucid zone is observed between the core and membrane of the polar filament. The polar filament runs through the highly laminated polaroplast which occupies the anterior portion of the spore. In cross-fractured face the lamellae of the polaroplast are arranged like the petals of a flower. The basal portion of the polar filament is enlarged, appearing arrow-shaped in thin sections and pear-shaped in frozen-etched preparations. Frozen-etched membranes differ in the size and distribution of the surface particles.     

Distribution of peptidyl-glycine alpha-amidating monooxygenase immunoreactivity in the brain,pituitary and islet organ of the anglerfish (Lophius americanus)

Peptidyl-glycine alpha-amidating monooxygenase (PAM; EC 1.14.17.3) is an enzyme that catalyzes conversion of glycine-extended peptides to alpha-amidated bioactive peptides. Two peptides that are processed at their carboxyl-termini by this enzyme are neuropeptide Y and anglerfish peptide Y, both of which possess a C-terminal glycine that is used as a substrate for amidation. Results from previous reports have demonstrated that neuropeptide Y-like and anglerfish peptide Y-like immunoreactivities are present in the brain of anglerfish (Lophius americanus). Furthermore, neuropeptide Y-like peptides, namely anglerfish peptide Y and anglerfish peptide YG (the homologues of pancreatic polypeptide) are present in the islet organ of this species. Neuropeptide Y has also been localized in the anterior, intermediated and posterior lobes of the pituitary gland in a variety of species. In order to learn more about the distribution of the enzyme responsible for alpha amidation of these peptides in the brain and pituitary and to specifically investigate the relationship of this enzyme to peptide synthesizing endocrine cells of the anglerfish islet, we performed an immunohistochemical study using several antisera generated against different peptide sequences of the enzyme. PAM antisera labeled cells in the islet organ, pituitary and brain, and fibers in the brain and pituitary gland. The PAM staining pattern in the brain was remarkably similar to the distribution of neuropeptide Y immunoreactivity reported previously. Clusters of cells adjacent to vessels in the anterior pituitary displayed punctate PAM immunoreactivity while varicose fibers were observed in the pituitary stalk and neurohypophysis. Endocrine cells of the islet organ were differentially labeled with different PAM antisera. Comparison of the staining patterns of insulin, glucagon, and anglerfish peptide Y in the islet organ to PAM immunoreactivity suggests a distribution of forms of PAM enzyme in insulin and anglerfish peptide Y-containing cells, but no overlap with glucagon-producing cells. The results also indicate that PAM immunoreactivity is widely distributed in the brain, pituitary and islet organ of anglerfish in cells that contain peptides that require presence of a C-terminal glycine for amidation.     

Further Observations on the Fine Structure of the Spores of Glugea weissenbergi (Microsporida,Nosematidae)*

SYNOPSIS. A Glugea xenoma sectioned and viewed with the electron microscope contained many spores with everting polar filaments. Several details not seen in previous studies of this species were observed. A specialized area with the appareance of a lattice was commonly present near the anterior end of the polaroplast. The external portion of a partially everted polar filament appeared to have about twice the diameter of the part remaining within the spore. No membrane was seen limiting the external surface of the everted portion. The everting filament had pushed thru the polar cap and the adjacent thin area of the spore wall, making the polar cap into a ring. The ring connected the proximal end of the everting filament to the inner spore membrane, thereby anchoring the filament to the spore. The electron density of some of the membranous organelles of the spore was enhanced by the use of ruthenium red.     

Interactions between Oncopera alboguttata (Lepidoptera: Hepialidae) and its microsporidan pathogen, Pleistophora oncoperae (Protozoa: Microsporida)

Infection by Pleistophora oncoperae is widespread in populations of Oncopera alboguttata. Detailed study of insects from one site indicated that 85% of the 131 pupae and 65% of the 403 adults were infected. The infection did not adversely affect duration of larval and pupal development, adult life span, the number of eggs laid, or fecundity. Infected females were able to transmit the disease to their progeny, though the degree of transmission ranged from 0 to 92%, depending on the severity of the parental infection. Transmission of the disease by infected males could not be detected. It was estimated that 49% of the progeny from this site would have been infected by transovum transmission. Details of the incidence of P. oncoperae infection from this area during the period 1972 to 1979 are given to emphasise the possible role of this disease in O. alboguttata population dynamics.     

Infection and development of Nosema bombycis (Microsporida: Protozoa) in a cell line of Antheraea eucalypti

Spores of Nosema bombycis derived from diseased insects were highly purified by Urografin density gradient centrifugation. Antheraea eucalypti cells were inoculated with the purified spores primed with 0.1 n KOH solution to start a continuous propagation of N. bombycis in cell culture. The first increase in the number of infected A. eucalypti cells was observed at 48 hr postinoculation, and it was caused by the secondary infective forms of N. bombycis. The secondary infective forms were produced during the course of sporoblast differentiation. The parasites in cell cultures divided synchronously until 36 hr postinoculation. Mature spores were observed initially 6 days postinoculation at 27°C. The infected cultures were subcultured extensively for more than 1 year with the addition of healthy A. eucalypti cells.     

The storage of infective spores of Vairimorpha necatrix (Protozoa: Microsporida) in antibiotic solution at 4 degrees C.

The interactions between the Diacrisia virginica granulosis virus (DGV) and the Hyphantria cunea baculovirus isolates were determined, utilizing defined differences between the time-mortality responses of these viruses fed to H. cunea larvae. The DGV, having a prolonged incubation period, when given an advantage in time or in the number of capsules, was able to prevent the expression of the more lethal H. cunea granulosis virus (HcGV) isolate. The time-mortality response of test larvae simultaneously fed equivalent dosages of HcGV and DGV was intermediate to that achieved with HcGV alone or DGV alone. Larvae infected with the DGV isolate were still susceptible to double infection by the nucleopolyhedrosis virus. The time-mortality response demonstrated that the development of nucleopolyhedrosis was only partly inhibited by preinfecting the test larvae with the DGV isolate.     

Ultrastructure of Sarcocystis spp. (Protozoa: Apicomplexa) in rodents from North Sulawesi and West Java, Indonesia

Tissue cysts of the protozoan genus Sarcocystis were detected in the skeletal muscles of 16 (40%) of 40 wild rodents captured in North Sulawesi and West Java, Indonesia. Two types of cysts were found to differ in their morphological characteristics. Macroscopic and microscopic cysts bounded by thick radially-striated cyst walls were detected at both locations in a total of 13 rodents belonging to seven different species (Bunomys chrysocomus, B. fratrorum, Maxomys bartelsii, M. musschenbroekii, Paruromys dominator, Rattus xanthurus and R. exulans). The primary cyst walls contained numerous broad spatula-like protrusions and the cysts were identified as S. singaporensis Zaman and Colley, 1976. In contrast, microscopic cysts bounded by thin smooth cyst walls were detected in seven rodents belonging to three different species captured at Toraut in North Sulawesi (B. chrysocomus, B. fratrorum and P. dominator). Ultrastructural examination revealed numerous slender hair-like protrusions of their primary cyst walls. It is proposed that these cysts be named S. sulawesiensis sp. n. on the basis of their unique morphological characteristics, their intermediate host range and their limited geographic distribution. Mixed infections by both species were found in three rodent species (B. chrysocomus, B. fratrorum and P. dominator).     

The preservation of infective spores of Nosema necatrix (Protozoa: Microsporida) in Spodoptera exempta (Lepidoptera: Noctuidae) by lyophilization

Cadavers of Spodoptera exempta infected with Nosema necatrix were lyophilized. The lyophilized material was tested against a standard spore preparation 1 day after lyophilization and retested 2 yr later. There was some loss of spore viability. The feasibility of using dose-mortality experiments to estimate the loss in viability is discussed.     

Localization and characterization of neuropeptide Y-like peptides in the brain and islet organ of the anglerfish (Lophius americanus)

Summary Results from a previous report demonstrate that more than one molecular form of neuropeptide Y-like peptide may be present in the islet organ of the anglerfish (Lophius americanus). Most of the neuropeptide Y-like immunoreactive material was anglerfish peptide YG, which is expressed in a subset of islet cells, whereas an additional neuropeptide Y-like peptide(s) was localized in islet nerves. To learn more about the neuropeptide Y-like peptides in islet nerves, we have employed immunohistochemical and biochemical methods to compare peptides found in anglerfish islets and brain. Using antisera that selectively react with either mammalian forms of neuropeptide Y or with anglerfish peptide YG, subsets of neurons were found in the brain that labelled with only one or the other of the antisera. In separate sections, other neurons that were labelled with either antiserum exhibited similar morphologies. Peptides from brains and islets were subjected to gel filtration and reverse-phase high performance liquid chromatography. Radioimmunoassays employing either the neuropeptide Y or peptide YG antisera were used to examine chromatographic eluates. Immunoreactive peptides having retention times of human neuropeptide Y and porcine neuropeptide Y were identified in extracts of both brain and islets. This indicates that peptides structurally similar to both of these peptides from the neuropeptide Y-pancreatic polypeptide family are expressed in neurons of anglerfish brain and nerve fibers of anglerfish islets. The predominant form of neuropeptide Y-like peptide in islets was anglerfish peptide YG. Neuropeptide Y-immunoreactive peptides from islet extracts that had chromatographic retention times identical to human neuropeptide Y and porcine neuropeptide Y were present in much smaller quantities. These results are consistent with the hypothesis that peptides having significant sequence homology with human neuropeptide Y and porcine neuropeptide Y are present in the nerve fibers that permeate the islet.     

Renal tubular transport of inorganic divalent ions by the aglomerular marine teleost,Lophius americanus

A characterization was attempted of the mechanisms involved in the tubular transport of inorganic divalent ions by the aglomerular kidney of Lophius, attention being paid particularly to the possible existence of transport maxima (Tm) and to competition for transport among related substances undergoing tubular excretion. Excretory rates of divalent ions in non-treated fish during standard laboratory conditions paralleled spontaneous changes in urine flow. Tm rates of excretion were reached for magnesium, sulfate, and thiosulfate with corresponding plasma levels of 2 to 5, 5 to 17, and 4 to 12 microM/ml. respectively. Elevation of magnesium chloride levels in plasma markedly depressed calcium excretion; sodium thiosulfate similarly depressed sulfate excretion. Experimental observations suggest the existence of a transport system for divalent cations separate from another for divalent anions. Within each transport system the ion with the higher excretion rate depressed competitively transfer of the other ion. Neither system was influenced by probenecid (benemid) in doses which markedly depressed the simultaneous excretion rate of p-aminohippuric acid.