A new species of Nosema from Hylobittacus apicalis (Insecta: Mecoptera: Bittacidae)

An undescribed Nosema was found infecting adults of the mecopteran Hylobittacus apicalis. This microsporidium is described herein as the first record of a microsporidium from the order Mecoptera. The slightly pyriform spores measured 4.5 × 2.4 μm. Mature spores had 9.5–10 polar filament coils irregularly grouped in the posterior end. The life cycle and ultrastructure of the developmental stages were described, and were typical of other Nosema spp. This microsporidium was regularly recorded from adult Hylobittacus apicalis populations over a 10-year period and tbe incidence of infection increased during the summer.     

Tuzetia boeckella sp. nov. (protozoa: Microsporida), a parasite of Boeckella triarticulata (Copepoda: Calanoidea) in Australia

A hitherto undescribed microsporidan has been found in the Australian freshwater copepod, Boeckella triarticulata, collected from Lake Burley Griffin, Canberra. We name this protozoan Tuzetia boeckella n. sp. and describe it in this paper. Large numbers of spores were found in the muscle of both sexes and all stages of the animals. The pyriform spores measured 5.1 × 2.7 μm with the extruded polar filament measuring 102 μm. Ultrastructural studies revealed the presence of a pansporoblastic membrane around each spore. The polar filament was arranged in a single row of 13–14 turns and decreased in diameter toward the posterior end. Few details of the life cycle were elucidated; however, evidence is presented for each sporont forming eight spores. Differentiating characters to distinguish this species from the six other known members of the genus are given.     

Transmission and infectivity of spores of Burenella dimorpha (Microsporida: Burenellidae)

Burenella dimorpha infects the tropical fire ant, Solenopsis geminata, producing two morphologically distinct types of spores. A binucleate, nonpansporoblast membrane-bounded (NPMB) spore develops in and destroys the hypodermis, rupturing the cuticle in the pupal stage. A uninucleate, pansporoblast membrane-bounded (PMB) spore develops in the fat body. Adult ants cannibalize ruptured pupae but do not ingest spores. Instead, the spores and particulate foods are diverted to the infrabuccal cavity, formed into an infrabuccal pellet, and fed to fourth-instar larvae only. This larval instar is the only stage in the life cycle of S. geminata that is vulnerable to infection. NPMB spores are infective, but PMB spores do not extrude their polar filaments in the larval gut and are expelled in the meconium upon pupation.     

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.     

A microsporidium,Nosema pilicornis sp. n., of the purslane sawfly,Schizocerella pilicornis

A new microsporidian species, Nosema pilicornis, which infects the purslane sawfly, Schizocerella pilicornis, is described. This microsporidium infects most body tissues of the host. N. pilicornis was compared to other microsporidian species infecting Hymenoptera and to a group of similar microsporidia infecting Lepidoptera. N. pilicornis could be distinguished from all other microsporidian species on the basis of host range and ultrastructural characteristics of the spore. Spores were oval, containing 11 to 12 polar filament coils, and the polar filament had an angle of tilt of about 80°. N. pilicornis infected lepidopteran larvae, but only when heavy spore dosages were fed to early larval instars. S. pilicornis is a good but sporadic biological control agent of common purslane, Portulaca oleracea, a pernicious weed of vegetable, ornamental, and orchard crops. N. pilicornis, which is transovarially transmitted and causes high mortality in infected larvae, affects the performance of S. pilicornis as a biological control agent.     

Novel Myxobolus and Ellipsomyxa species (Cnidaria: Myxozoa) parasiting Brachyplatystoma rousseauxii (Siluriformes: Pimelodidae) in the Amazon basin,Brazil

We describe two novel myxosporean parasites from Brachyplatystoma rousseauxii, an economically important freshwater catfish from the Amazon basin, Brazil. Myxobolus tapajosi n. sp., was found in the gill filaments of 23.5% of 17 fish, with myxospores round to oval in frontal view and biconvex in lateral view: length 15 (13.5–17) μm and width 10.7 (9.6–11.4) μm; polar capsules equal, length 5.8 (4.6–7.1) μm and width 3 (2.3–3.8) μm containing polar tubules with 6–7 turns. Ellipsomyxa amazonensis n. sp. myxospores were found floating freely or inside plasmodia in the gall bladder of 23.5% of fish. The myxospores were ellipsoidal with rounded extremities: length 12.8 (12.3–13.6) μm and width 7.6 (6.7–8.7) μm; with two equal, slightly pyriform polar capsules, length 3.8 (3.8–4.0) μm and width 3.1 (2.5–3.4) μm, containing polar tubules with 2–3 turns. We combined spore morphometry, small-subunit ribosomal DNA data, specific host, and phylogenetic analyses, to identify both of these parasites as new myxozoan species. Maximum likelihood and Bayesian analyses showed that Myxobolus tapajosi n. sp. clustered in a basal branch in a subclade of parasites from exclusively South American pimelodid fishes. Ellipsomyxa amazonensis n. sp. clustered within the marine Ellipsomyxa lineage, but we suspect that although the parasite was collected in freshwater, its hosts perform a large migration throughout the Amazon basin and may have become infected from a brackish/marine polychaete host during the estuary phase of its life.     

Description of Hyalinocysta expilatoria n. sp., a microsporidian parasite of the blackfly Odagmia ornata

Hyalinocysta expilatoria n. sp. is described from a larva of Odagmia ornata collected in Sweden. Infection was restricted to the adipose tissue which was transformed into a syncytium. The earliest stage observed was diplokaryotic merozoites, which mature directly into diplokaryotic sporonts. Each sporont produces a sporophorous vesicle (pansporoblast), which persists, also enclosing mature spores. Usually nuclear divisions result in a plasmodium with 8 nuclei, which fragments into 8 sporoblasts, each of which develops into a spore without further division. Occasionally an aberrant number of spores (2, 4, 6) is formed. The spores are pyriform with a flattened area at the posterior pole. Spores in sporophorous vesicles with 8 spores are 4.0–6.0 μm long, in vesicles with 4 spores 4.0–5.0 μm, and in vesicles with 2 spores 7.0–8.0 μm. In some vesicles the spores develop asynchronously, and 2, 4, or 6 mature spores are found together with 6, 4, or 2 immature. There was also a small number of vesicles with supernumerary spores, less than 8 normally developed. The 325–350 nm thick spore wall is composed of three layers. The polar filament is anisofilar with 7 coils in a single layer. The anterior 5–6 coils are wide, the posterior 2-1 thin. The angle of tilt of the anterior filament coil is approximately 50°. The spore has a single nucleus. The sporophorous vesicle is delimited by a thin membrane, also visible in haematoxylin stained preparations. Vesicles with mature spores are void of metabolic inclusions.     

Fine structure of Chloromyxum menticirrhi n. sp. (Myxozoa) infecting the urinary bladder of the marine teleost Menticirrhus americanus (Sciaenidae) in Southern Brazil

A myxosporidian was found in the urinary bladder of the teleost Menticirrhus americanus Linnaeus, 1758 (Sciaenidae) collected from the South Atlantic coast of Brazil. Polysporic amoeboid plasmodia containing sporoblasts, developing pansporoblasts and spores were free in the bladder lumen. The prevalence of infection was 17.64% (15/85). Unfixed spores were spherical to subspherical, on average 10.5 μm long, 9.8 μm wide and 10.1 μm thick (n=25), and fixed spores measured 10.1×9.5×9.7 μm. The two spore valves were of equal size and each possessed prominent sutural lines and about 41 (37–45) surface ridges aligned parallel with the suture line. These ridges gave transverse sections a cog-wheel-like outline. The spores contained four pyriform polar capsules of equal size (3.20×2.0 μm) (n=25) (fixed), each with a polar filament having 3–4 (rarely 5) coils. The binucleate sporoplasm was irregular in shape, with granular matrix and randomly distributed dense bodies. The shape and dimensions of the spore, as well as the number, position and arrangement of the surface ridges, polar capsules and polar filament indicate that this is a new species, herein designated Chloromyxum menticirrhi. The gill, liver, gall bladder and intestine of the host showed no abnormalities.     

More on the life cycle of Malamoeba scolyti (Amoebidae: Sarcomastigophora) parasitizing: the bark beetle Dryocoetes autographus (Scolytidae,Coleoptera)

Some new developmental features previously unreported in, the life- cycle of an amoeba Malamoeba scolyti, which cause cysts in Malpighian tubules of the bark beetle Dryocoetes autographus, are described by means of light, phase-contrast, Nomarski interference, and electron microscopy. Two developmental phases are presumed to take place in the life cycle of M. scolyti: a phase with large multinucleate amoebae (trophozoites) developing in the gut and caecum of the host, and a phase (parasitic phase) causing cysts in the Malpighian tubules. Some data on the host-parasite relationship and rate of infection are also discussed.     

Enterocytozoon hepatopenaei sp. nov. (Microsporida: Enterocytozoonidae), a parasite of the black tiger shrimp Penaeus monodon (Decapoda: Penaeidae): Fine structure and phylogenetic relationships

A new microsporidian species, Enterocytozoon hepatopenaei sp. nov., is described from the hepatopancreas of the black tiger shrimp Penaeus monodon (Crustacea: Decapoda). Different stages of the parasite are described, from early sporogonal plasmodia to mature spores in the cytoplasm of host-cells. The multinucleate sporogonal plasmodia existed in direct contact with the host-cell cytoplasm and contained numerous small blebs at the surface. Binary fission of the plasmodial nuclei occurred during early plasmodial development and numerous pre-sporoblasts were formed within the plasmodium. Electron-dense disks and precursors of the polar tubule developed in the cytoplasm of the plasmodium prior to budding of early sporoblasts from the plasmodial surface. Mature spores were oval, measuring 0.7 × 1.1 μm and contained a single nucleus, 5-6 coils of the polar filament, a posterior vacuole, an anchoring disk attached to the polar filament, and a thick electron-dense wall. The wall was composed of a plasmalemma, an electron-lucent endospore (10 nm) and an electron-dense exospore (2 nm). DNA primers designed from microsporidian SSU rRNA were used to amplify an 848 bp product from the parasite genome (GenBank FJ496356). The sequenced product had 84% identity to the matching region of SSU rRNA from Enterocytozoon bieneusi. Based upon ultrastructural features unique to the family Enterocytozoonidae, cytoplasmic location of the plasmodia and SSU rRNA sequence identity 16% different from E. bieneusi, the parasite was considered to be a new species, E. hepatopenaei, within the genus Enterocytozoon.     

Alternative development in Polystoma gallieni (Platyhelminthes,Monogenea) and life cycle evolution

Considering the addition of intermediate transmission steps during life cycle evolution, developmental plasticity, canalization forces and inherited parental effect must be invoked to explain new host colonization. Unfortunately, there is a lack of experimental procedures and relevant models to explore the adaptive value of alternative developmental phenotypes during life cycle evolution. However, within the monogeneans that are characterized by a direct life cycle, an extension of the transmission strategy of amphibian parasites has been reported within species of Polystoma and Metapolystoma (Polyopisthocotylea; Polystomatidae). In this study, we tested whether the infection success of Polystoma gallieni within tadpoles of its specific host, the Stripeless Tree Frog Hyla meridionalis, differs depending on the parental origin of the oncomiracidium. An increase in the infection success of the parasitic larvae when exposed to the same experimental conditions as their parents was expected as an adaptive pattern of non-genetic inherited information. Twice as many parasites were actually recorded from tadpoles infected with oncomiracidia hatching from eggs of the bladder parental phenotype (1.63 ± 0.82 parasites per host) than from tadpoles infected with oncomiracidia hatching from eggs of the branchial parental phenotype (0.83 ± 0.64 parasites per host). Because in natural environments the alternation of the two phenotypes is likely to occur due to the ecology of its host, the differential infection success within young tadpoles could have an adaptive value that favors the parasite transmission over time.     

A multiplex PCR assay to diagnose and quantify Nosema infections in honey bees (Apis mellifera)

Correct identification of the microsporidia, Nosema apis and Nosema ceranae, is key to the study and control of Nosema disease of honey bees (Apis mellifera). A rapid DNA extraction method combined with multiplex PCR to amplify the 16S rRNA gene with species-specific primers was compared with a previously published assay requiring spore-germination buffer and a DNA extraction kit. When the spore germination-extraction kit method was used, 10 or more bees were required to detect the pathogens, whereas the new extraction method made it possible to detect the pathogens in single bees. Approx. 4-8 times better detection of N. ceranae was found with the new method compared to the spore germination-extraction kit method. In addition, the time and cost required to process samples was lower with the proposed method compared to using a kit. Using the new DNA extraction method, a spore quantification procedure was developed using a triplex PCR involving co-amplifying the N. apis and N. ceranae 16S rRNA gene with the ribosomal protein gene, RpS5, from the honey bee. The accuracy of this semi-quantitative PCR was determined by comparing the relative band intensities to the number of spores per bee determined by microscopy for 23 samples, and a high correlation (R² = 0.95) was observed. This method of Nosema spore quantification revealed that spore numbers as low as 100 spores/bee could be detected by PCR. The new semi-quantitative triplex PCR assay is more sensitive, economical, rapid, simple, and reliable than previously published standard PCR-based methods for detection of Nosema and will be useful in laboratories where real-time PCR is not available.     

Nosema blissi sp. n. (Microsporida: Nosematidae), a pathogen of the chinch bug,Blissus leucopterus hirtus (Hemiptera: Lygaeidae)

Nosema blissi sp. n. is described from the Malpighian tubules of adults of Blissus leucopterus hirtus. Spores measured 6.5 ± 0.3 × 2.5 ± 0.1 μm in Giemsa-stained preparations. The polar filament lay in 37 to 40 coils, arranged in a single layer in the posterior portion of the spore, and in several layers in the anterior portion.     

Evaluation of some morphological characteristics of spores of two species of Microsporida by scanning electron microscope and freeze-etching techniques

Scanning electron microscopy revealed spores of Nosema apis and Thelohania fibrata to be egg-shaped, but only the mature spore of T. fibrata was shown to possess a horseshoe-like concavity at the posterior pole. Freezeetched preparations indicated that this concavity was due to a thin area of the spore coat. Freeze-etching studies also show spores of N. apis do possess an umbrella-shaped polaroplast, and a polar filament which is arranged in a double layer with over 30 coils. The spore of T. fibrata contains a pear-shaped arrangement of the polaroplast membrane, and a polar filament arranged in a single layer of 22 coils.     

Description of Chytridiopsis Trichopterae N. Sp. (Microspora, Chytridiopsidae), A Microsporidian Parasite of the Caddis Fly Polycentropus Flavomaculatus (Trichoptera, Polycentropodidae), With Comments On Relationships Between the Families Chytridiopsidae and Metchnikovellidae

ABSTRACT. The microsporidium Chytridiopsis trichopterae n. sp., a parasite of the midgut epithelium of larvae of the caddis fly Polycentropus flavomaculatus found in southern Sweden, is described based on light microscopic and ultrastructural characteristics. All life cycle stages have isolated nuclei. Merogonial reproduction was not observed. the sporogony comprises two sequences: one with free spores in parasitophorous vacuoles, the other in spherical, 5.6-6.8 μm wide, sporophorous vesicles which lie in the cytoplasm. the free sporogony yields more than 20 spores per sporont. the vesicle-bound sporogony produces 8, 12 or 16 spores. the envelope of the sporophorous vesicle is about 82 nm thick and layered. the internal layer is the plasma membrane of the sporont; the surface layer is electron dense with regularly arranged translucent components. Both spore types are spherical. They have an ～ 35-nm thick spore wall, with a plasma membrane, an electron-lucent endospore, and an ～ 14-nm thick electron-dense exospore. the polar sac is cup-like and lacks a layered anchoring disc. the polar filament is arranged in two to three isofilar coils in the half of the spore opposite the nucleus. the coupling between the polar sac and the polar filament is characteristic. the surface of the polar filament is covered with regularly arranged membraneous chambers resembling a honeycomb. There is no polaroplast of traditional type. the cytoplasm lacks polyribosomes. the nucleus has a prominent, wide nucleolus. the two spore types have identical construction, but differ in dimensions and electron density. Free living spores are about 3.2 μm wide, the diameter of the polar filament proper is 102-187 nm, the chambers of the honeycomb are 70-85 nm high, and the polar sac is up to 425 nm wide. Living spores in the vesicle-bound sporogony are about 2.1 μm wide, the polar filament measures 69-102 nm, the chambers of the honeycomb are about 45 nm high, and these spores are more electron dense. Comparisons of cytology (especially the construction of the spore wall and the polar filament and associated structures) and life cycles reveal prominent differences among the Chytridiopsis-like microsporidia, and close relationships between the families Chytridiopsidae and Metchnikovellidae.     

Light and Electron Microscope Observations on Nosema nelsoni Sprague, 1950 (Microsporida,Nosematidae) with Particular Reference to its Golgi Complex*

SYNOPSIS. A species of Nosema in the muscles of the North American white shrimp, generally known as Penaeus setiferus but also known as P. fluviatilis, appears identical with type specimens of N. nelsoni Sprague, 1950, in P. aztecus. Its Golgi apparatus, as seen in the sporoblast, is a complex system of cisternae, small vesicles and expanded sacs which plays a major role in spore morphogenesis. It transforms directly into the polaroplast complex, certain membranous investments of the polar filament, the polar sac and perhaps part of the posterior vacuolar system. Probably the polar sac contains the polar cap. The PAS-positive material in both the cap and the filament may be a component of the Golgi complex. This new concept of the Golgi complex supplements our earlier view of spore morphogenesis according to which the polar filament is of nuclear origin. It also reconciles the idea with Vávra's identification of Golgi vesicles associated with the developing polar filament.     

Wittmannia antarctica N. G., N. Sp. (Nosematidae), a New Hyperparasite in the Antarctic Dicyemid Mesozoan Kantharella antarctica

ABSTRACT. Collections of the dicyemid mesozoan Kantharella antarctica were made in the Weddell Sea during the Antarctic Expedition of the research vessel RV Polarstern in 1990 and 1991. A diplokaryotic microsporidian was found infecting all nematogens from all the samples taken in both years. The infected cells contained all developmental stages. Merogony initially was monokaryotic and spoorogony of diplokaryotic sporonts was by multiple fission. The stained ovoidal spores measured between 4.3-6 μm X 1.7-2.3 μm. The ultrastructural findings come from 11 specimens of Kantharella antarctica that were cut in serial sections. All developmental stages were noteworthy because of the myelinosomes situated adjacent to each diplokaryon. Similarly conspicuous were some organelles in the spore: a prominent, extraordinarily electron dense anterior portion of the polaroplast and the posterior vacuole. The isofilar polar filament with a diameter of about 115 nm showed 9-11 coils. The great number of empty spore cases together with an extruded polar filament are indicative of an autoinfection. Though these characteristics resemble in part those of the genus Nosema from the family Nosematidae, the species in Kantharella antarctica differs from the former by its unusual development, life cycle and unusual host. Thus, this new species has been placed in a new genus and the name Wittmannia antarctica proposed.     

Fine structure of the developing spore of Nosema apis zander

Summary The mature spore possesses a thick spore coat and a particle-bearing spore membrane. The highly laminated polaroplast membranes are located at the anterior pole of the spore. Close to its base, the polar filament is surrounded by the polaroplast membrane. The polar filament runs spirally towards the posterior pole of the spore. A large portion of the polar filament is arranged in two layers. A similar arrangement was also observed in immature spores and in the sporoblast stage, although it was not so orderly arranged in the latter. The developing polaroplast membrane was observed in the immature spore, but not in the sporoblast. The sporoblast wall is much thinner than the spore coat, but has the same texture. Endoplasmic reticulum is the most prominent cytoplasmic organelle in the developing stages of Nosema apis. Porous nuclear envelopes are also observed in developing stages. The role of the endoplasmic reticulum in the formation of the polar filament, polaroplast and spore coat, and the function of the spore membrane, are discussed.     

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.