A New Species of Perezia (Microsporida: Pereziidae) from the Argentine Grasshopper Dichroplus elongatus (Orthoptera: Acrididae)1

The new microsporidium (Microsporida: Pereziidae), Perezia dichroplusae n. sp., infects the epithelial cells of the Malpighian tubules of the Argentine grasshopper Dichroplus elongatus. Characteristics of the pathogen include the following: development in direct contact with the host cell cytoplasm; bi-, tetra-, and sometimes multinucleate diplokaryotic meronts, rounded or elongate in shape; unikaryotic sporonts and sporogonial plasmodia, elongate in shape; sporoblasts and spores uninucleated; spores highly variable in size (1.6–6.7 by 1.0–2.7 μm, x?= 3.5 ± 0.09 by 1.5 ± 0.02, n = 100) with eight or fewer polar tube coils and showing a posterior electron-dense inclusion body.     

An Ultrastructural Study of Vairimorpha necatrix (Microspora, Microsporida) with Particular Reference to Episporontal Inclusions During Octosporogony

ABSTRACT. The life cycle of Vairimorpha necatrix was studied by electron microscopy. Disporous development has two distinct stages: 1) diplokaryotic meronts which are actively mitotic, and 2) diplokaryotic sporonts which are distinguished by reduced ribosome density and a thickened plasmalemma. After final division of the sporont, sporoblasts form spores which are ovocylindrical and measure 4.4 ± 0.08 × 2.3 ± 0.05 μ m (mean ± SE). Octosporous development results in eight haploid spores being formed in a sporophorous vesicle. The uninucleate octospores were smaller than the binucleate dispores and the exospore was thicker but less crenulate in outline. Early in octosporogony, tubules are produced from the sporont plasmalemma and electron-dense material accumulates in the episporontal space. The latter may be amorphous, vesiculated, or vacuolated in appearance and in later stages may take a stacked, lamellar form. At sporoblast formation, exospore material coats the plasmalemma and attached tubules; all inclusions in the episporontal space gradually disappear as spores are formed. These secretory products may have application to taxonomic distinction at the species level.     

Life Cycle of Culicosporella lunata (Hazard & Savage, 1970) Weiser, 1977 (Microspora) as Revealed in the Light Microscope with a Redescription of the Genus and Species1

Light microscopy studies of Culicosporella lunata (Hazard & Savage), a parasite of the mosquito Culex pilosus (Dyar & Knab), revealed two sporogonial sequences. One sequence begins with diplokaryotic meronts that undergo repeated nuclear divisions to produce sporogonial plasmodia with nuclei in diplokaryotic arrangement. These plasmodia form rosette-like clusters of sporoblasts during incomplete cytokinesis and, eventually, binucleate spores. These spores initiate infections in healthy larvae when they ingest spores. The second sequence begins with diplokaryotic meronts that undergo karyogamy and meiosis to form Thelohania-like sporonts and haploid spores. Anomalies are often observed in these sporonts which result in aberrant spores, usually fewer than eight, in an accessory (pansporoblastic) membrane. Normal haploid spores are morphologically similar to those of species of Amblyospora. The genus and the type species are redefined based on new information presented here and it and the type species are placed in the family Amblyosporidae.     

Ultrastructural description of the life cycle of Nosema monorchis n. sp. (Microspora, Nosematidae), hyperparasite of Monorchis parvus (Digenea, Monorchiidae), intestinal parasite of Diplodus annularis (Pisces, Teleostei)

Life cycle stages of a new species of the genus Nosema Naegeli, 1857 (Microspora, Nosematidae), were examined by light and electron microscopy. It parasitizes the gut and the uterus of the digenean Monorchis parvus (Monorchiidae), in Diplodus annularis (Pisces, Teleostei). All stages were in close contact with the cytoplasm of the host cell and were probably all diplokaryotic. The divisions of meronts and sporonts were recognizable by the formation of spindle plaques at the surface of the nucleus. Spores were oval, measured 3.2±0.3×2.5±0.2 μm on ultrathin sections, and had a polar filament with 16–17 coils. The polaroplast presented two parts: an anterior region with closely packed lamellae and a posterior part with wider lamellae. This Nosema species is compared with the other microsporidian parasites of digeneans. This new species is named Nosema monorchis n. sp., after the generic name of its host.     

Euplotespora binucleata n. gen., n. sp. (Protozoa: Microsporidia), a parasite infecting the hypotrichous ciliate Euplotes woodruffi, with observations on microsporidian infections in ciliophora

A new microsporidian species, Euplotespora binucleata n. gen., n. sp., from the brackish-water ciliate Euplotes woodruffi is described and defined on the basis of life history characteristics, light and electron microscopic features, and small subunit (SSU) ribosomal DNA (rDNA) sequencing. The life cycle of E. binucleata n. sp. probably has rather short merogonic and relatively long sporogonic phases. Some uninuclear meronts and sporonts, along with diplokaryotic sporoblasts and spores, were found in experimentally infected host cells. Such a peculiar life cycle has been induced experimentally in Euplotes eurystomus and constitutively microsporidian-free stocks of E. woodruffi. Spores of E. binucleata n. sp. are monomorphic, ovoid-cylindrical in shape, 3.44+/-0.17 x 1.65+/-0.22 microm in size, and characterized by a diplokaryotic condition and a large posterior vacuole. The polar tube is isofilar, 4.5-5.5 microm in length when ejected, and lacking a distinctive coiled region (half-coiled). The polaroplast is divided into two regions: the anterior part has a few lamellae close to the anchoring disc; and the posterior part is a rounded body (sack), about one-quarter of the spore length. Spores do not appear to cluster together as a group. Each spore is surrounded by a sporophorous membrane closely adjacent to the exospore layer. A phylogenetic analysis of SSU rDNA sequences by different methods placed E. binucleata n. sp. in a clade with representatives of the microsporidian genera Cystosporogenes and Vittaforma. Observations of microsporidia in several other ciliates are discussed in view of the microsporidian infection frequency in the phylum Ciliophora.     

Ultrastructural Study of Endoreticulatus durforti N. Sp., a New Microsporidian Parasite of the Intestinal Epithelium of Artemia (Crustacea, Anostraca)

ABSTRACT. A new microsporidian parasite of the Artemia intestinal epithelium has been studied. The microsporidium developed within a membranous parasitophorous vesicle from the host rough endoplasmic reticulum consisting of two membranes, with the proximal one usually lacking ribosomes.
All developmental stages had isolated nuclei. Unikaryotic meronts developed into merogonial plasmodia. Merogonial division occurred by binary fission and rosette-shaped fragmentation. In young sporonts, an electron-lucent space, corresponding to the developing endospore, was immediately observed between both the plasmalemma and the exospore primordium. Sporogonial division occurred also by rosette-shaped fragmentation, resulting in at least eight sporoblasts that developed directly into spores. Fresh spores were 1.7 × 0.9 μm in size and oval-shaped. The 8–11 coil isofilar polar filament was arranged in two rows. The polaroplast was bipartite. The nature of the parasitophorous envelope, host-parasite interaction, developmental cycle and taxonomy are discussed.     

Life cycle of a new species of Amblyospora (Microspora: Amblyosporidae) in the mosquito Aedes taeniorhynchus

A new species of Microspora, Amblyospora polykarya, is described from the mosquito Aedes taeniorhynchus. The parasite is transovarially transmitted for one generation only. Spores in adult females extrude binucleate sporoplasms which infect developing eggs. Merogony occurs in larval oenocytes with diplokaryotic stages in early instars giving rise to plasmodia with many diplokarya. Plasmodia undergo cytokinesis to form diplokaryotic sporonts. In fat body cells, these sporonts secrete pansporoblastic membranes and undergo two nuclear divisions to form octonucleate sporonts. Cytokinesis and differentiation result in uninucleate spores in packets of eight. These spores are not transmissible per os and are of different morphotype from those in adult females. Infected larvae die in the fourth stadium.     

Description of a New Species of Microsporidia from Muscidifurax raptor (Hymenoptera: Pteromalidae), a Pupal Parasitoid of Muscoid Flies

ABSTRACT. A microsporidian parasite, Nosema muscidifuracis n. sp., has been found in Muscidifurax raptor , a parasitoid of muscoid flies. Stages of the parasite developed in direct contact with the host cell cytoplasm and were detected in midgut epithelium, Malpighian tubules, ovaries (including oocytes) and fat body of larvae and adults. Spores were also detected within eggs deposited on the host. Light and electron microscopy revealed a developmental cycle with diplokaryotic stages dividing by binary fission and disporous sporulation sequences producing diplokaryotic spores of three morphological classes, differing significantly only in length of the polar filament. Two of the classes were found in larvae, pupae and adults. One of these, with about five turns in the coiled polar filament, is presumed to be responsible for transmission from cell to cell within the host (autoinfection) and the other, with about 10 turns, responsible for transmission from host to host. A third class, with about 15 turns in the polar filament, was found in eggs of M. raptor . It is, presumably, either involved in initiation and spread of the infection at eclosion or is responsible for horizontal transmission to a new host individual when eggs are cannibalized.     

Karyogamy and meiosis in an Amblyospora sp. (Microspora) in the mosquito Culex salinarius

Stages of merogony and sporogony are illustrated in photomicrographs of lacto-aecto-orcein stained smears of an undescribed species of Amblyospora in male Culex salinarius larvae. Karyogamy was observed to take place in diplokaryotic meronts followed by an unusual meiosis-like process in young sporonts. Karyogamy was confirmed by DNA measurements in the nuclei of meronts. DNA measurements at mingling indicate that tetrads may not be formed during “pachytene.” Therefore, structures previously reported to be synaptonemal complexes and meiotic configurations of chromosomes may not actually represent true pachytene chromosomes. It is further shown that karyogamy has been either misplaced or overlooked by numerous investigators and that the behavior of chromosomes during the meiosis-like process was misinterpreted by previous researchers studying chromosomes in microsporidian sporonts.     

Ultrastructural Study and Description of Ovavesicula popilliae N. G., N. Sp. (Microsporida: Pleistophoridae) from the Japanese Beetle,Popillia japonica (Coleoptera: Scarabaeidae)1

A new genus and species of microsporidia, Ovavesicula popilliae n. g., n. sp., is described from the Japanese beetle, Popillia japonica, on the basis of studies by light and electron microscopy. Parasite development primarily occurs within the Malpighian tubules of larvae, and spores are formed in a sporophorous vesicle. Meronts have diplokaryotic nuclei, develop in direct contact with the host cell cytoplasm, and divide by binary fission. Sporonts have unpaired nuclei, develop within a thick sporophorous vesicle, and undergo synchronous nuclear divisions producing plasmodia with 2, 4, 8, 16, and 32 nuclei. Cytokinesis of sporogonial plasmodia does not occur until karyokinesis is complete with 32 nuclei. Intact sporophorous vesicles are ovoid, containing numerous secretory products, and are surrounded by a persistent two-layered wall. The uninucleate spores are regularly formed in groups of 32, and the polar tube in each has six coils.     

The morphology and development of Nosema carpocapsae,a microsporidian pathogen of the codling moth,Cydia pomonella (Lepidoptera: Tortricidae) in New Zealand

The morphology of Nosema carpocapsae and its development in experimentally infected codling moth larvae are described. Spherical uninucleate meronts were the first stages. Nuclear division produced binucleate meronts which were the most abundant vegetative stage, although additional uninucleate and a few tetranucleate meronts were also observed at this time. All meronts were spherical and ranged from 2.8 to 5.8 μm in diameter. Uninucleate and binucleate fusiform sporonts then appeared followed by some tetranucleate and dividing forms. Oval sporoblasts developed after these and did not divide before maturing into spores. Sporonts were approximately 5.0 to 7.9 × 2.4 to 3.0 μm. Spores developed in all host tissues except the nervous tissue. The binucleate spores showed considerable variation in spore size, 2.4 to 3.9 × 1.3 to 3.1 μm (alcohol fixed, Giemsa stained). The polar filament was usually coiled 11 times (range 9 to 13) at an angle of 53° to the long axis of the spore. Its maximum observed length was 75 μm.     

Heterovesicula cowani N. G., N. Sp. (Heterovesiculidae N. Fam.), a Microsporidian Parasite of Mormon Crickets, Anabrus simplex Haldeman, 1852 (Orthoptera: Tettigoniidae)

ABSTRACT. Heterovesicula cowani , n. g., n. sp., is a dimorphic microsporidium described from the adipose tissue of the Mormon cricket, Anabrus simplex Haldeman. Proliferation of the microsporidium is by karyokinesis of uninucleate and binucleate cells to form binucleate and tetranucleate cells, respectively. These cells will undergo binary fission (merogony). Ultimately, the meronts undergo karyokinesis without subsequent cytokinesis producing spherical multinucleate plasmodia that are transitional to 2 types of sporogony. Transitional to disporoblastic sporogony, a fragile interfacial envelope delaminates from the plasmodium with morphogenesis to a monfiliform plasmodium consisting of fusiform binucleate diplokaryotic sporonts. These undergo karyokinesis to form tetranucleate diplokaryotic sporonts that undergo cytokinesis during disintegration of the plasmodium into isolated binucleate sporonts. Transitional to octosporoblastic sporogony, multinucleate plasmodia disintegrate into short monofiliform plasmodia of diplokaryotic sporonts which then segregate while undergoing gradual nuclear dissociation (haplosis by nuclear dissociation). These undergo two sequences of karyokinesis and subsequent multiple fission to form eight uninucleate (haploid) sporoblasts in a fusiform arrangement within a persistent envelope. Binucleate spores are ovocylindrical, about 5.4 × 1.7 μm (fresh), with an isofilar polar filament singly coiled about 11 turns. Uninucleate spores are ovoid to slightly pyriform, 4.0 × 1.7 μm (fresh), with an isofilar filament singly coiled about 9 turns. A new family, Heterovesculidae, is proposed for the new genus.     

Nosema tyriae n.sp. and Nosema sp., microsporidian parasites of Cinnabar moth Tyria jacobaeae.

Nosema tyriae n.sp. was found in 63% of a population of Cinnabar moth larvae (Tyria jacobaeae). The infection was found in the gut wall, silk glands, and fat body and was probably generalized but appeared to be of low pathogenicity. Merogony and sporogony were by binary fission of diplokaryotic stages. Fresh spores were elongate, slightly pointed at the anterior end, and measured 4.7 x 2.0 microm. Ultrastructural features of special interest were 20-nm tubules connecting the surface of sporonts with host cell cytoplasm and, in the spores, a deeply domed polar sac, polaroplast consisting of closely packed longitudinally arranged membranes and loosely packed horizontally arranged membranes, and 10.5-14 coils of the polar tube in a single rank. The 16S rRNA genes of N. tyriae and Nosema bombycis from silkworms, Bombyx mori, differed by only six nucleotides and N. tyriae spores gave a moderately positive reaction with a monoclonal antibody raised to N. bombycis. N. tyriae was infective to B. mori but was less virulent than N. bombycis. However, no amplification product was obtained by PCR using N. tyriae DNA and primers considered to be specific for N. bombycis. Also, the spores of the two species are of entirely different shapes. A second diplokaryotic microsporidium, Nosema sp., found as a light infection in only one of the larvae had much smaller developmental stages and spores measuring 3.8 x 2.0 microm (fixed). Ultrastructurally it was distinguished by an abundance of dense membranes in cytoplasmic vesicles in both meronts and sporonts. Spores with up to 15 coils of the polar tube in irregular clusters or with about 12 coils in a single rank were observed in the tissues fixed from the one larva infected with this parasite. As this larva had been kept with N. tyriae-infected larvae for a few days before examination, it is possible that the two types of spores resulted from a double infection.     

Redescription of Microsporidium takedai (Awakura, 1974) as Kabatana takedai (Awakura, 1974) comb. n

Ultrastructural study of the microsporidian Microsporidium takedai from the muscles of masu salmon Oncorhynchus masou proved that this species can be assigned to the genus Kabatana Lom, Dyková and Tonguthai, 2000. The parasites develop within disintegrated sarcoplasm without any delimiting boundary or cyst. Cylindrical multinucleate meronts proliferate by serial constrictions into uninucleate stages which repeat the process. Eventually, the uninucleate stages transform into uninucleate sporonts, which divide once to produce sporoblasts, thus functioning as sporoblast mother cells. Spores, with a subterminally located anchoring disc and 3 to 4 turns of the polar tube coil, average 3.3 by 1.9 microm in size. The exospore is divided into small fields; the endospore frequently makes small invaginations into the spore inside. Phylogenetic analysis using SSU rDNA sequence consistently placed Kabatana takedai in a group consisting of Microgemma sp., Spraguea lophii and Glugea americanus. The K. takedai could easily be separated from the other species in the same group by 2 inserts in the SSU rDNA sequence.     

Food utilization values of gypsy moth Lymantria dispar (Lepidoptera: Lymantriidae) larvae infected with the microsporidium Vairimorpha sp. (Microsporidia: Burenellidae)

Infection of the gypsy moth, Lymantria dispar, with the microsporidium Vairimorpha sp. strongly influences the development of the host in ways typical of many species of terrestrial entomopathogenic Microsporidia; growth is reduced while development time is extended in infected insects. The appearance of the different stages of the parasite in the host relative to the elapsed time after oral infection, as well as the influence of the parasite proliferation on food utilization of the host, were examined. At 3 days postinfection, midgut muscle cells were infected with primary spores, and the fat body tissues contained meronts, sporonts, and primary spores. Many more fat body cells contained vegetative stages and primary spores at 4 and 5 days postinfection, and diplokaryotic spores and immature octospores were also present. Approximate digestibility of infected larvae increased during this time period, whereas the conversion of ingested and digested food to body substance decreased. The relative growth rate of infected and uninfected groups did not differ significantly between 4 and 5 days postinfection, although the relative consumption rate in infected L. dispar larvae was higher. Between 8 and 10 days postinfection, the relative growth rate of uninfected larvae increased. The infected group did not demonstrate this increase at a time period characterized by maturation of diplokaryotic spores and octospores in larval fat body tissues. Total body weight of uninfected larvae remained higher than that of infected larvae after 8 days postinfection.     

The Light and Electron Microscopic Cytology of Janacekia adipophila N. Sp. (Microspora, Tuzetiidae), a Microsporidian Parasite of Ptychoptera paludosa (Diptera, Ptychopteridae)

ABSTRACT. The microsporidium Janacekia adipophila n. sp., a parasite of Ptychoptera paludosa larvae in Sweden, is described based on light microscopic and ultrastructural characteristics. Merogonial stages and sporonts are diplokaryotic. Merozoites are formed by rosette-like division. Sporonts develop into sporogonial plasmodia with isolated nuclei. These plasmodia give rise to 8–16 sporoblasts by rosette-like budding. A sporophorous vesicle is initiated by the sporogonial plasmodium. Sporoblasts and spores are enclosed in individual sporophorous vesicles. Granular inclusions of the vesicles, visible using light microscopy, discriminate sporogonial stages from stages of the merogony. The monokaryotic, fresh spores are oval with blunt ends, measuring 4.2-6.3 × 9.1-11.2 μm. Macrospores are formed in small numbers. The spore wall has three subdivisions and the exospore is electron-dense. The polaroplast has two parts: closely arranged lamellae anteriorly, wider sac-like compartments posteriorly. The isofilar polar filament, 191–264 nm wide, has 12-13 coils, which are arranged in one layer in the posterior half of the spore. The electron-dense inclusions of the sporophorous vesicle are modified during sporogony, and vesicles with mature spores are traversed by 21–27 nm wide tubules, which connect the exospore with the envelope of the vesicle. The walls of the tubules, the envelope of the vesicles, and the surface layer of the exospore are all identical double-layered structures. The microsporidium is compared to microsporidia of Ptychopteridae and Tipulidae and to related microsporidia of the family Tuzetiidae.     

A New Pathogen,Microsporidium itiiti n. sp. (Microsporida), from the Argentine Stem Weevil,Listronotus bonariensis (Coleoptera,Curculionidae)1

A new species of microsporidium (phylum Microspora) infecting the Argentine stem weevil, Listronotus bonariensis (Kuschel. 1955), is described on the basis of light and electron microscope observations. It has the following characteristics: nuclei always isolated; meronts spherical and sporonts ribbon-shaped, with variable numbers of nuclei; sporogony within a vacuole which is bounded by a thin membrane that usually breaks down before uninucleate spores mature; occasionally parts of the membrane remain so that clusters of variable numbers of spores may be seen in light microscopic preparations. Spores measure 2.5 × 1.4 μm (fresh) and development occurs mainly in the midgut, but also in the epidermis, fat body, muscle, and ovaries.     

Ultrastructural Characterisation and Molecular Taxonomic Identification of Nosema granulosis n. sp., a Transovarially Transmitted Feminising (TTF) Microsporidium

A novel microsporidian parasite is described, which infects the crustacean host Gammarus duebeni. The parasite was transovarially transmitted and feminised host offspring. The life cycle was monomorphic with three stages. Meronts were found in host embryos, juveniles, and in the gonadal tissue of adults. Sporoblasts and spores were restricted to the gonad. Sporogony was disporoblastic giving rise to paired sporoblasts, which then differentiated to form spores. Spores were not found in regular groupings and there was no interfacial envelope. Spores were approximately 3.78 x 1.22 microns and had a thin exospore wall, a short polar filament, and an unusual granular polaroplast. All life cycle stages were diplokaryotic. A region from the parasite small subunit ribosomal RNA gene was amplified and sequenced. Phylogenetic analysis based on these data places the parasite within the genus Nosema. We have named the species Nosema granulosis based on the structure of the polaroplast.