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.     

Growth of Nosema algerae in Pig Kidney Cell Cultures*

SYNOPSIS. Nosema algerae, a microsporidan parasite of mosquitoes, can infect pig kidney cell cultures. Spores germinated in the culture medium, infected the cells within 30 min of germination, multiplied, and produced spores. The early developmental stages in the N. algerae life cycle are described.     

Establishment of the new genus Paranosema based on the ultrastructure and molecular phylogeny of the type species Paranosema grylli Gen. Nov., Comb. Nov. (Sokolova, Selezniov, Dolgikh, Issi 1994), from the cricket Gryllus bimaculatus Deg

The ultrastructure of the microsporidian parasite Nosema grylli, which parasitizes primarily fat body cells and haemocytes of the cricket Gryllus bimaculatus (Orthoptera, Gryllidae) is described. All observed stages (meront, meront/sporont transitional stage ("second meront"), sporont, sporoblast, and spore) are found in direct contact with the host cell cytoplasm. Nuclei are diplokaryotic during almost all stages of the life cycle, but a brief stage with one nucleus containing an abundance of electron-dense material is observed during a "second merogony." Sporogony is disporous. Mature spores are ovocylindrical in shape and measure 4.5+/-0.16micromx2.2+/-0.07 microm (n=10) on fresh smears and 3.3+/-0.06 micromx1.4+/-0.07 microm (n=10) on ultrathin sections. Spores contain 15-18 coils of an isofilar polar filament arranged in one or two layers. Comparative phylogenetic analysis using rDNA shows N. grylli to be closely related to another orthopteran microsporidian, Nosema locustae, and to Nosema whitei from the confused flour beetle, Tribolium confusum. Antonospora scoticae, a parasite of the communal bee Andrena scotica, is a sister taxon to these three Nosema species. The sequence divergence and morphological traits clearly separate this group of "Nosema" parasites from the "true" Nosema clade containing Nosema bombycis. We therefore propose to change the generic name of N. grylli and its close relative N. locustae to Paranosema n. comb. We leave N. whitei in former status until more data on fine morphology of the species are obtained.     

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.     

Ultrastructural characteristics and small subunit ribosomal DNA sequence of Vairimorpha cheracis sp. nov., (Microspora: Burenellidae), a parasite of the Australian yabby, Cherax destructor (Decapoda: Parastacidae)

This is the first record of a species of Vairimorpha infecting a crustacean host. Vairimorpha cheracis sp. nov. was found in a highland population of the Australian freshwater crayfish, Cherax destructor. The majority of spores and earlier developmental stages of V. cheracis sp. nov. were found within striated muscle cells of the thorax, abdomen, and appendages of the crayfish. Only octosporoblastic sporogony within sporophorous vesicles (SPVs) was observed. Diplokaryotic sporonts separated into two uninucleate daughter cells, each of which gave rise to four sporoblasts in a rosette-shaped plasmodium, so that eight uninucleate spores were produced within the persistent ovoid SPV. Ultrastructural features of stages in the octosporoblastic sequence were similar to those described for Vairimorpha necatrix, the type species. Mature spores were pyriform in shape and averaged 3.4x1.9 microm in dimensions. The anterior polaroplast was lamellar in structure, and the posterior polaroplast vesicular. The polar filament was coiled 10-12 times, lateral to the posterior vacuole. The small subunit ribosomal DNA (SSU rDNA) of V. cheracis sp. nov. was sequenced and compared with other microsporidia. V. cheracis sp. nov. showed over 97% sequence identity with Vairimorpha imperfecta and five species of Nosema, and only 86% sequence identity with V. necatrix. The need for a taxonomic revision of the Nosema/Vairimorpha group of species is discussed.     

萨梅诺娃新佩雷斯虫中国新记录

研究报道了中国首例摇蚊微孢子虫, 结合各发育阶段形态特征、生态学特征及分子特征, 鉴定其为萨梅诺娃新佩雷斯虫Neoperezia semenovaiae Issi, et al. 2012, 系我国新记录。萨梅诺娃新佩雷斯虫寄生于羽摇蚊幼虫脂肪体组织, 导致其体表呈白浊状。成熟孢子呈卵圆形, 孢子长(5.7±0.2) μm (5.3—6.3 μm), 宽(3.7±0.1) μm (3.4—4.0 μm)。透射电镜观察显示各发育阶段均为离核, 发育不同步, 与宿主细胞质直接接触。早期发育阶段为高电子密度的多核裂殖体阶段, 经原生质团分裂形成单核或多核产孢体, 进一步发育为单核孢子母细胞。孢子母细胞形状不规则, 周围被内质网环绕, 并逐渐形成微孢子虫的典型结构如极丝、极质体和三层孢壁等。成熟孢子卵圆形, 离核, 细胞核较大, 位于孢子正中央, 被大量核糖体包围。极质体分为两部分, 前半部分为海绵状, 后半部分薄膜状。锚状盘位于孢子前端, 呈蘑菇状。孢壁三层, 外层为高电子密度层, 厚26.5—62.7 nm, 中间层为电子透明层, 厚151.8—236.1 nm, 里层为质膜层。同型极丝, 30—31圈, 分2—3列排列。扩增获得其小核糖体序列为1356 bp, 序列比较发现其与俄罗斯列宁格勒区羽摇蚊的N. semenovaiae相似性为99.1%。系统发育关系分析表明N. semonovaiae与Neoperezia、Bryonosema、Schroedera属种类聚为一独立进化枝, N. semonovaiae种群出现明显的地理分化。     

Fine structure of the microsporidan Abelspora portucalensis gen.n., sp.n. (Microsporida) parasite of the hepatopancreas of Carcinus maenas (Crustacea,Decapoda)

A new species of a microsporidan, Abelspora portucalensis, was found in the hepatopancreas of Carcinus maenas, forming white xenomas. Each xenoma seems to consist of an aggregate of hypertrophic host cells in which the parasite develops and proliferates. This cytozoic microsporidan being characterized by one uninucleate schizont giving rise to two sporonts, each originating two sporoblasts, resulting in two spores within a persistent sporophorous vacuole (pansporoblast) should be included in a new family Abelsporidae. In fresh smears most spores were 3.1–3.2 μm long and 1.2–1.4 μm wide. Fixed, stained, and observed in SUS mature spores measured 3.1 ± 0.08 × 1.3 ± 0.06 μm (n = 25 measurements). Spore cytoplasm was dense and granular, polyribosomes were arranged in helicoidal tape form. The polar filament was anisofilar and consisted of a single coil with 5–6 turns. The anchoring disc and and the anterior zone of the filament are surrounded by the polaroplast composed of two usual zones. In the anterior zone, the membrane of the polar filament is in continuity with the membranes of the polaroplast. The appearance of a microsporidan with described nuclear divisions in life cycle, spores shape and size, polaroplast and polar filament morphology and identity of the host suggests that we may erect a new genus Abelspora and a new species A. portucalensis (Portugal = Portucalem).     

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.     

Growth of Nosema algerae in pig kidney cell cultures.

Nosema algerae, a microsporidan parasite of mosquitoes, can infect pig kidney cell cultures. Sores germinated in the culture medium, infected the cells within 30 min of germination, multiplied, and produced spores. The early developmental stages in the N. algerae life cycle are discribed.     

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.     

Nosema chrysorrhoeae n. sp. (Microsporidia), isolated from browntail moth (Euproctis chrysorrhoea L.) (Lepidoptera, Lymantriidae) in Bulgaria: characterization and phylogenetic relationships

A new microsporidian parasite Nosema chrysorrhoeae n. sp., isolated in Bulgaria from the browntail moth (Euproctis chrysorrhoea L.), is described. Its life cycle includes two sequential developmental cycles that are similar to the general developmental cycles of the Nosema-like microsporidia and are indistinguishable from those of two Nosema spp. from Lymantria dispar. The primary cycle takes place in the midgut tissues and produces binucleate primary spores. The secondary developmental cycle takes place exclusively in the silk glands and produces binucleate environmental spores. N. chrysorrhoeae is specific to the browntail moth. Phylogenetic analysis based on the ssu rRNA gene sequence places N. chrysorrhoeae in the Nosema/Vairimorpha clade, with the microsporidia from lymantriid and hymenopteran hosts. Partial sequences of the lsu rRNA gene and ITS of related species Nosema kovacevici (Purrini K., Weiser J., 1975. Natürliche Feinde des Goldafters, Euproctis chrysorrhoea L., im Gebiet von Kosovo, FSR Jugoslawien. Anzeiger fuer Sch?dlingskunde, Pflanzen-Umweltschutz, 48, 11-12), Nosema serbica Weiser, 1963 and Nosema sp. from Lymantria monacha was obtained and compared with N. chrysorrhoeae. The molecular data indicate the necessity of future taxonomic reevaluation of the genera Nosema and Vairimorpha.     

A Microsporidan Infection of Anopheles gambiae Giles, from Tanzania, Interpretation of Its Mode of Transmission and Notes on Nosema Infections in Mosquitoes

SYNOPSIS A microsporidan found parasitic in Anopheles gambiae in Tanzania was identified as Nosema algerae Vavra & Undeen, 1970. Its development was traced in laboratory-bred Anopheles stephensi. Spores were oval, 3.0-4.3 μ× 1.8-2.5 μ and had filaments 40-60 μ long; schizonts were uni-, bi- or tetranucleate; elongate sporonts gave rise to 2 sporoblasts. The parasite attacked almost every tissue of the body of the larvae, pupae and adults including the ova, but transovarial transmission was thought to be unimportant as infected ova did not develop into viable larvae. Hosts reacted to the presence of the parasite by encapsulating some spores in a coat of melanin.
The parasite was lightly infective to several species of culicine mosquitoes, but the infection was limited mainly to the nervous system, mid-gut epithelium and Malpighian tubules.
The species of Nosema parasitizing mosquitoes are reviewed.     

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.     

Morphological and molecular investigations of Tubulinosema ratisbonensis gen. nov., sp. nov. (Microsporidia: Tubulinosematidae fam. nov.), a parasite infecting a laboratory colony of Drosophila melanogaster (Diptera: Drosophilidae)

A new species of microsporidia from Drosophila melanogaster was investigated by light and electron microscopy and by ribosomal RNA (rRNA) sequencing. This microsporidium and the previously described Nosema kingi and Nosema acridophagus have been transferred to the new genus Tubulinosema gen. nov. with the following characters: nuclei are in diplokaryotic arrangement during the life cycle. All stages are in direct contact with the host cell cytoplasm, slightly anisofilar polar tube with the last coils being smaller in diameter arranged in one or two rows on both sides of the diplokaryon and small tubuli on the surface of late meronts. Spores are oval or slightly pyriform. Thick endospore wall, thinner over anchoring disc. This new genus and the genus Brachiola have been placed in a new family Tubulinosematidae fam. nov. Phylogenetic analysis of small subunit rRNA sequences by different methods placed Tubulinosema spp. in one clade with the genus Brachiola forming its sister clade, which is distant from the clade containing the true Nosema spp. including Nosema bombycis.     

Amblyospora sp. (Microspora, Amblyosporidae) infecting nerve ganglia of Culex pipiens (Diptera, Culicidae) from Egypt.

A species of Amblyospora-infecting neurones of Culex pipiens is described. Diplokaryotic meronts, which divided by binary fission, were distinguished at the electron microscope level by their unthickened plasma membranes. Sporonts with an electron-dense surface coat gave rise to eight uninucleate sporoblasts within a sporophorous vesicle, cytoplasmic division occurring at the quadrinucleate or octonucleate stages. Indications that nuclear fusion and chromosome reorganization occurred in merogony and sporogony were obtained by light microscopy but meiosis was not detected at the ultrastructural level. Spores were typical of Amblyospora, being ovoid when fresh, truncate when stained, and having an exospore of two membranous layers subtended by a thick amorphous layer, an electron-lucent endospore, an anisofilar polar filament, and a polaroplast comprised of an anterior region of close-packed lamellae and a posterior region of expanded sacs. The metabolic products in the sporophorous vesicle took the form of large globules, small globules with electron-dense borders, and fine granules. These were depleted in mature sporophorous vesicles, though a surface layer of fine granules on the spores may have been derived from them. Many stages were degenerate and it is suggested that C. pipiens may be an accidental host in which the parasite could develop suboptimally in nervous tissue only. Infections in larvae hatched from eggs in the laboratory indicate that vertical transmission occurs.     

Amazonspora hassar n. gen. and n. sp. (phylum Microsporidia,fam. Glugeidae), a parasite of the Amazonian teleost Hassar orestis (fam. Doradidae)

We describe the microsporidian Amazonspora hassar n. gen., n. sp. from the gill xenomas of the teleost Hassar orestis (Doradidae) collected in the estuarine region of the Amazon River. The parasite appeared as a small whitish xenoma located in the gill filaments near the blood vessels. Each xenoma consisted of a single hypertrophic host cell (HHC) in the cytoplasm of which the microsporidian developed and proliferated. The xenoma wall was composed of up to approximately 22 juxtaposed crossed layers of collagen fibers. The plasmalemma of the HHC presented numerous anastomosed, microvilli-like structures projecting outward through the 1-3 first internal layers of the collagen fibrils. The parasite was in direct contact with host cell cytoplasm in all stages of the cycle (merogony and sporogony). Sporogony appears to divide by plasmotomy, giving rise to 4 uninucleate sporoblasts, which develop into uninucleate spores. The ellipsoidal spores measured 2.69 +/- 0.45 x 1.78 +/- 0.18 microm, and the wall measured approximately 75 nm. The anchoring disk of the polar filament was subterminal, being shifted laterally from the anterior pole. The polar filament was arranged into 7-8 coils in a single layer in the posterior half of the spore, surrounding the posterior vacuole. The polaroplast surrounded the uncoiled portion of the polar filament, and it was exclusively lamellar. The spores and different life-cycle stages were intermingled within the cytoplasm of the HHC, surrounding the central hypertrophic deeply branched nucleus. The ultrastructural morphology of this microsporidian parasite suggests the erection of a new genus and species.     

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.