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Timing of an early sporulation sequence of microsporidia in the genus vairimorpha (Microsporidia: burenellidae)

An early sporulation event in the host midgut tissues has been reported for several species of microsporidia infecting Diptera, Hymenoptera, and Lepidoptera. The role of these primary spores, formed between 35 and 96 h postinfection per os, has been suggested to be the cell to cell spread of infection within the host, but the sequence of events during the early sporulation stages has been reported for only a few species of microsporidia. We investigated these early life cycle events for two species of microsporidia, Vairimorpha necatrix and Vairimorpha sp. from Lymantria dispar, tested in the laboratory hosts Spodoptera exigua and L. dispar, respectively. We injected hemolymph drawn from orally infected host larvae into uninfected larvae at time periods from 2 to 96 h postinfection to determine the timing of infection of the hemolymph and target tissues. Our studies demonstrated that, for both Vairimorpha species, the early sporulation in the midgut tissues is a discrete first stage of infection. Invasion of the hemolymph and infection of the target tissues follows maturation of "primary spores" in the midgut and coincides with the germination of these spores beginning approximately 30 h postinfection for hosts held at a constant 24 degreesC. The developmental stages of the microsporidia observed in the target tissues at specific time periods postinfection corresponded to the stages observed in the hemocytes, suggesting that sporoplasms from the germinating primary spores are directly injected into the target tissues and the hemolymph. Copyright 1998 Academic Press.

Journal Article↗

Evolutionary strategies and adaptations for survival between mosquito-parasitic microsporidia and their intermediate copepod hosts: a comparative examination of Amblyospora connecticus and Hyalinocysta chapmani (Microsporidia: Amblyosporidae).

The epizootiology, transmission dynamics, and survival strategies employed by two mosquito-parasitic microsporidia that utilize copepods as intermediate hosts are examined in relation to the biological attributes of their hosts and the environments in which they inhabit. Amblyospora connecticus Andreadis, 1988, a parasite of Ochlerotatus cantator (Coquillett) and Acanthocyclops vernalis (Fischer) is found in an unstable salt marsh environment that is subject to periodic flooding and drying. Both hosts have distinct non-overlapping generations. A. connecticus exhibits a well-defined seasonal transmission cycle that relies heavily on maternal-mediated transovarial transmission by female O. cantator during the summer, and horizontal transmission via the copepod host during the spring (copepod to mosquito) and fall (mosquito to copepod). Its survival strategies include: delayed virulence, low pathogenicity and high tissue specificity that allow for transstadial transmission of horizontally acquired infections and maximum spore production, reliance on living hosts throughout most of its life cycle with overwintering in the copepod, polymorphic development that is well synchronized with host physiology, and production and dissemination of infectious spores that are coincident with the seasonal occurrence of susceptible stages in each host. Hyalinocysta chapmani Hazard et Oldacre, 1975, a parasite of Culiseta melanura (Coquillett) and Orthocyclops modestus (Herrick) is found in a comparatively stable, subterranean habitat that is inundated with water throughout the year. Copepods are omnipresent and C. melanura has overlapping broods. H. chapmani is maintained in a continuous cycle of horizontal transmission between each host throughout the summer and fall but lacks a developmental sequence leading to transovarial transmission in the mosquito host. It relies on living hosts for most of its life cycle and overwinters in diapausing mosquito larvae. Transstadial transmission does not occur and there is no dimorphic development in the mosquito host. The spatial and temporal overlap of both mosquito and copepod hosts during the summer and fall affords abundant opportunity for continuous horizontal transmission and increases the likelihood that H. chapmani will find a target host, thus negating the need for a transovarial route. It is hypothesized that natural selection has favoured the production of meiospores in larval female mosquitoes rather than congenital transfer of infection to progeny via ovarian infection as a strategy for achieving greater transmission success. Analysis of the molecular phylogeny data suggest that (1) transovarial transmission and the developmental sequence leading to ovarian infection have been secondarily lost in H. chapmani, as they occur in all other closely related genera, (2) the ancestral state included complex life cycles involving transovarial transmission and an intermediate host, and (3) mosquito-parasitic microsporidia are adjusting their life cycles to accommodate host ecological conditions.

Adaptation, Biological↗

Molecular phylogeny and evolution of mosquito parasitic Microsporidia (Microsporidia: Amblyosporidae).

Amblyospora species and other aquatic Microsporidia were isolated from mosquitoes, black flies, and copepods and the small subunit ribosomal RNA gene was sequenced. Comparative phylogenetic analysis showed a correspondence between the mosquito host genera and their Amblyspora parasite species. There is a clade of Amblyospora species that infect the Culex host group and a clade of Amblyospora that infect the Aedes/Ochlerotatus group of mosquitoes. Parathelohania species, which infect Anopheles mosquitoes, may be the sister group to the Amblyospora in the same way that the Anopheles mosquitoes are thought to be the sister group to the Culex and Aedes mosquitoes. In addition, by sequence analysis of small subunit rDNA from spores, we identified the alternate copepod host for four species of Amblyospora. Amblyospora species are specific for their primary (mosquito) host and each of these mosquito species serves as host for only one Amblyospora species. On the other hand, a single species of copepod can serve as an intermediate host to several Amblyospora species and some Amblyospora species may be found in more than one copepod host. Intrapredatorus barri, a species within a monotypic genus with Amblyospora-like characteristics, falls well within the Amblyospora clade. The genera Edhazardia and Culicospora, which do not have functional meiospores and do not require an intermediate host, but which do have a lanceolate spore type which is ultrastructurally very similar to the Amblyospora spore type found in the copepod, cluster among the Amblyospora species. In the future, the genus Amblyospora may be redefined to include species without obligate intermediate hosts. Hazardia, Berwaldia, Larssonia, Trichotuzetia, and Gurleya are members of a sister group to the Amblyospora clades infecting mosquitoes, and may be representatives of a large group of aquatic parasites.

Animals↗

[The artificial infection of the cladoceran Moina macrocopa (Crustacea: Phyllopoda) with the microsporidia Gurleya sp. (Microsporidia: Gurleyidae)].

Microcrustaceans Moina macrocopa (Straus, 1820) have been collected in a small vernal pond and cultured under laboratory conditions. The infection of this culture with the microsporidia Gurleya sp. was detected and high virulence of this microsporidian species allowed us to carry out the transmission experiments. Horizontal transmission, the high host specificity and maximal spore production in the hypodermal and fat body cells through 5-6 days after per oral experimental infection have been revealed. Histological investigations demonstrated that Gurleya sp. reduces drastically host reproductive success without a considerable influence on the microcrustaceans' mortality.

Animals↗

Microsporidia of the genus Trachipleistophora--causative agents of human microsporidiosis: description of Trachipleistophora anthropophthera n. sp. (Protozoa: Microsporidia).

Trachipleistophora anthropophthera n. sp., was found at autopsy in the brain of one and in the brain, kidneys, pancreas, thyroid, parathyroid, heart, liver, spleen, lymph nodes, and bone marrow of a second patient with AIDS. The parasite is similar to the recently described T. hominis Hollister, Canning, Weidner, Field, Kench and Marriott, 1996, in having isolated nuclei, meronts with a thick layer of electron dense material on the outer face of their plasmalemma and sporogony during which spores are formed inside a thick-walled sporophorous vesicle. In contrast to T. hominis, this species is dimorphic as it forms two kinds of sporophorous vesicles and spores: Type I--round to oval polysporous sporophorous vesicle, 7-10 microns in size, usually with eight spores (3.7 x 2.0 microns), thick endospores, subterminal anchoring disc and anisofilar polar filaments forming seven thicker and two thinner terminal coils. This type of sporophorous vesicle is associated with 25-30 nm filaments extending into the host cell cytoplasm. Type II--smaller, bisporous sporophorous vesicle (4-5 x 2.2-2.5 microns) with two, nearly round, thin-walled spores, 2.2-2.5 x 1.8-2.0 microns in size, having 4-5 isofilar coils. No outside filamentous elements are associated with the bisporous sporophorous vesicle. Both types of sporophorous vesicles were common in the infected brain tissue and could be found within the same cell. The newly described species, together with T. hominis and previously reported Pleistophora-like parasites from human muscle, likely represent a group of closely related human microsporidia.

AIDS-Related Opportunistic Infections↗

Blinded, externally controlled multicenter evaluation of light microscopy and PCR for detection of microsporidia in stool specimens. The Diagnostic Multicenter Study Group on Microsporidia.

The quality parameters for the detection of microsporidia in identical sets of 50 stool samples were determined for six laboratories where technicians used light microscopy and for six laboratories where technicians used PCR. The average overall sensitivities were 67% (89% for patient samples only) for the PCR laboratories and 54% (80% for patient samples only) for the light microscopy laboratories. Specificities were 98 and 95%, respectively. Differences in results were most apparent between the individual laboratories rather than between the two major methods used.

Animals↗

[New species of Microsporidia--Thelohania jungarica SP. N. (Microsporidia, Nosematidae) from larvae of black flies].

A new species of microsporidians, Thelohania jungarica sp. n., was described from larval populations of Odagmia sp. inhabiting the mountain rivers of the northern spurs of Tien-Shan, south-eastern Kazakhstan. The larval infection rate is 3 to 16%. The pupal infection was reported from single individuals. The species in question differs from other members of the genus in the morphology of spores, ecology and host. Sizes of measured 50 spores varied in the following ranges: live ones--5.1-7.6 x 2.5-3.8, fixed and stained--3.8-6.4 x 1.8-3.8 mkm. The parasite causes pathological and morphological changes in injured tissues.

Animals↗

[New species of Microsporidia - Thelohania assovi Sp. N. (Microsporidia, Nosematidae) from the larvae of black flies].

A new species of microsporidians, Thelohania assovi sp. n., was found in population of larvae of Odagmia sp. inhabiting mountain rivers of Tien-Shan and south-eastern Kazakhstan. The infection of larvae with this species varies from single individuals to 18-20% while pupae are infected only with single speciments. The species differes from all known members of this genus in a number of characters. Size of 50 measured spores varied from 4.4-7.0 X 2.5-3.8 in live specimens to 3.8-6.4 X 2.3-3.8 mem in fixed ones. The parasite causes patho--morphological changes in injured tissues.

Animals↗

Evidence from beta-tubulin phylogeny that microsporidia evolved from within the fungi.

Microsporidia are obligate intracellular parasites that were thought to be an ancient eukaryotic lineage based on molecular phylogenies using ribosomal RNA and translation elongation factors. However, this ancient origin of microsporidia has been contested recently, as several other molecular phylogenies suggest that microsporidia are closely related to fungi. Most of the protein trees that place microsporidia with fungi are not well sampled, however, and it is impossible to resolve whether microsporidia evolved from a fungus or from a protistan relative of fungi. We have sequenced beta-tubulins from 3 microsporidia, 4 chytrid fungi, and 12 zygomycete fungi, expanding the representation of beta-tubulin to include all four fungal divisions and a wide diversity of microsporidia. In phylogenetic trees including these new sequences, the overall topology of the fungal beta-tubulins generally matched the expected relationships among the four fungal divisions, although the zygomycetes were polyphyletic in some analyses. The microsporidia consistently fell within this fungal diversification, and not as a sister group to fungi. Overall, beta-tubulin phylogeny suggests that microsporidia evolved from a fungus sometime after the divergence of chytrids. We also found that chytrid alpha- and beta-tubulins are much less divergent than are tubulins from other fungi or microsporidia. In trees in which the only fungal representatives were the chytrids, microsporidia still branched with fungi (i.e., with chytrids), suggesting that the affiliation between microsporidian and fungal tubulins is not an artifact of long-branch attraction.

Animals↗

Evaluation of methodologies including immunofluorescent assay (IFA) and the polymerase chain reaction (PCR) for detection of human pathogenic microsporidia in water.

Microsporidia is a term used to describe a group of emerging protozoan pathogens whose environmental occurrence has only recently been documented due to lack of detection methodologies. This study evaluates and describes current methods for detection of microsporidia in water. Standard methods, for the collection and processing of large volumes of water to detect protozoa, showed only a 4.8% recovery, of microsporidia spores, from 100 l volumes of tap. Immunofluorescent assay (IFA) analysis was assessed using two different antibodies specific for human pathogenic microsporidia. Results indicated that the use of IFA for routine screening of water for microsporidia was not an acceptable approach. The antibodies tested for the IFA resulted in false positives and false negatives and did not react with Enterocytozoon bieneusi, which is an important human pathogenic microsporidia. Finally, the small sizes of the human pathogenic microsporidia prevent confirmation and species determination by light microscopic methods. Two methods for isolating microsporidia DNA from water for use in polymerase chain reaction (PCR) amplification of microsporidia target sequences were assessed. Both of these DNA isolation methods when combined with the PCR showed the ability to detect less than ten spores in purified water concentrates. Thus, this study represents the first documentation and evaluation of current methods for the detection of human pathogenic microsporidia in water.

Animals↗

Fish microsporidia: fine structural diversity and phylogeny.

Structural diversity of fish microsporidian life cycle stages and of the host-parasite interface is reviewed. In the infected cell of the fish host, microsporidia may either cause serious degradation of the cytoplasm and demise of the cell, or they may elicit host cell hypertrophy, producing a parasite-hypertrophic host cell complex, the xenoma. The structure of the xenoma and of its cell wall may differ according to the genus of the parasite, and seems to express properties of the parasite rather than those of the host. In merogony, the parasite cell surface interacts with the host cell in diverse ways, the most conspicuous being the production of thick envelopes of different types. Sporogony stages reveal different types of walls or membranes encasing the sporoblasts and later the spores and these envelopes may be of host or parasite origin. Nucleospora differs from all other fish microsporidia by its unique process of sporogony. Except for the formation of conspicuous xenomas, there are no essentially different structures in fish-infecting microsporidia compared with microsporidia from other hosts. Although the structures associated with the development of fish microsporidia cannot be attributed importance in tracing the phylogeny, they are relevant for practical determination and assessing the relation to the host. The possibility of the existence of an intermediate host is discussed. Higher-level classification of Microsporidia is briefly discussed and structure and evolutionary rates in microsporidian rDNA are reviewed. Discussion of rDNA molecular phylogeny of fish-infecting microsporidia is followed by classification of these parasites. Most form a rather cohesive clade. Outside this clade is the genus Nucleospora, separated at least at the level of Order. Within the main clade, however, there are six species infecting hosts other than fish. Based on data available for analysis, a tentative classification of fish-infecting microsporidia into five groups is proposed. Morphologically defined groups represent families, others are referred to as clades. Group 1, represented by family Pleistophoridae, includes Pleistophora, Ovipleistophora and Heterosporis; Vavraia and Trachipleistophora infect non-fish hosts. Group 2, represented by family Glugeidae, is restricted to genus Glugea and Tuzetia weidneri from crustaceans. Group 3 comprises three clades: Loma and a hyperparasitic microsporidian from a myxosporean; Ichthyosporidium and Pseudoloma clade and the Loma acerinae clade. For the latter species a new genus has to be established. Group 4 contains two families, Spragueidae with the genus Spraguea and Tetramicridae with genera Microgemma and Tetramicra, and the Kabatana and Microsporidium seriolae clade. Group 5 is represented by the family Enterocytozoonidae with the genus Nucleospora and mammal-infecting genus Enterocytozoon.

Animals↗

The First United Workshop on Microsporidia from Invertebrate and Vertebrate Hosts.

The phylum Microsporidia is a large group of parasitic unicellular eukaryotes that infect a wide range of invertebrate and vertebrate taxa. These organisms are significant human and veterinary pathogens with impacts on medicine, agriculture and aquaculture. Scientists working on these pathogens represent diverse disciplines that have had limited opportunities for detailed interactions. A NATO Advanced Research Workshop 'Emergent Pathogens in the 21st Century: First United Workshop on Microsporidia from Invertebrate and Vertebrate Hosts' was held July 12-15, 2004 at the Institute of Parasitology of the Academy of Sciences of the Czech Republic to bring together experts in insect, fish, veterinary and human microsporidiosis for the exchange of information on these pathogens. At this meeting, discussions were held on issues related to taxonomy and phylogeny. It was recognized that microsporidia are related to fungi, but the strong opinion of the participants was that the International Code of Zoological Nomenclature should continue to be applied for taxonomic descriptions of the Microsporidia and that they be treated as an independent group emerging from a paraphyletic fungi. There continues to be exponential growth in the pace and volume of research on these ubiquitous intracellular protists. The small genomes of these organisms and the reduction in the size of many of their genes are of interest to many disciplines. Many microsporidia are dimorphic and the mechanisms underlying these morphologic changes remain to be elucidated. Epidemiologic studies to clarify the source of human microsporidiosis and ecologic studies to understand the multifaceted relationship of the Microsporidia and their hosts are important avenues of investigation. Studies on the Microsporidia should prove useful to many fields of biologic investigation.

Animals↗

How do microsporidia invade cells?

Microsporidia are obligate intracellular eukaryotic parasites that utilize a unique mechanism to infect host cells. One of the main characteristics of all microsporidia is that they produce spores containing an extrusion apparatus that consists of a coiled polar tube ending in an anchoring disc at the apical part of the spore. With appropriate conditions inside a suitable host, the polar tube is discharged through the thin anterior end of the spore, thereby penetrating a new host cell for inoculating the infective sporoplasm into the new host cell. This method of invading new host cells is one of the most sophisticated infection mechanisms in biology and ensures that the microsporidia enter the host cell unrecognized and protected from the host defence reactions. Recent studies have shown that microsporidia gain access to host cells by phagocytosis as well. However, after phagocytosis, the special infection mechanism of the microsporidia is used to escape from the maturing phagosomes and to infect the cytoplasm of the cells. Gaining access to cells by endocytosis, and escaping destruction in the phago-/endo-/lysosome by egressing quickly from the phagocytic vacuole to multiply outside the lysosome, is a common phenomenon in biology that has been evolved several times during evolution. How this is put into execution by the microsporidia is an inimitable principle by which an obligate intracellular organism has managed this problem. The extrusion apparatus of the microsporidia has obviously ensured the success of this phylum during evolution, resulting in a group of obligate intracellular organisms, capable of infecting almost any type of host and cell.

Animals↗

Review of microsporidia-mosquito relationships: from the simple to the complex.

Microsporidia in mosquitoes can be divided into two categories based on their life cycles and host-parasite relationships. Some species of microsporidia exhibit simple life cycles with one spore type responsible for oral (horizontal) transmission. They affect only one generation of the mosquito and are not usually host or tissue specific. Brachiola algerae and Vavraia culicis are examples of species isolated from mosquitoes with relatively straightforward life cycles (one spore type) and simple host-parasite relationships. B. algerae and a close relative of V. culicis have also been isolated from a vertebrate (human) host but sources for these infections are unknown. In contrast to B. algerae and V. culicis, polymorphic (heterosporous) microsporidia in mosquitoes are characterized by complex life cycles involving multiple spore types responsible for horizontal and vertical transmission. They affect two generations of the mosquito and some involve an obligate intermediate host. These microsporidia are generally very host and tissue specific with complex developmental sequences comprised of unique stages and events. The microsporidium Edhazardia aedis is a pathogen of Aedes aegypti and does not require an intermediate host. The developmental cycle of E. aedis is characterized by four sporulation sequences, two in the parental host and two in the filial generation. Recent speculation relative to the source of B. algerae human infection have implicated infected mosquitoes and raised concerns about the safety of mosquito microsporidia in general. The subject of this review is to compare and contrast three species of microsporidia from mosquitoes, two with broad host ranges (B. algerae and V. culicis) and one specific to mosquitoes (E. aedis). This review describes features that distinguish mosquito-parasitic microsporidia with simple life cycles and broad host ranges from truly mosquito-specific microsporidian parasites with complex life cycles.

Animals↗

Physiological host specificity of microsporidia as an indicator of ecological host specificity.

For most groups of biological control agents the relationship between laboratory (physiological) host range and the host range in the field (ecological host range) has not been explored empirically. The objective of our study was to investigate this relationship using the North America gypsy moth, Lymantria dispar, as a model nontarget host for microsporidia from native North American Lepidoptera. The gypsy moth, L. dispar, a native of Europe, has been established in North America for nearly 130 years and presumably exposed to many species of microsporidia from sympatric native Lepidoptera. Nevertheless, microsporidia have never been observed in North American populations of L. dispar. We conducted traditional laboratory feeding experiments using microsporidia from 20 lepidopteran host species and 1 coleopteran host species against L. dispar. Microsporidia from 18 native hosts infected L. dispar larvae. Although some of the infections were not typical of infections in the indigenous natural hosts, mature spores were produced in most of these infections. Horizontal transmission experiments, based on exposure of uninfected L. dispar larvae to infected L. dispar larvae, demonstrated that the microsporidia were far more host specific than the direct feeding experiments suggested. Of the three microsporidian biotypes that were horizontally transmitted between the nontarget L. dispar larvae, all were transmitted at very low levels. The results of our experiments provide additional evidence that the ecological host specificity of terrestrial microsporidia is much narrower than the physiological host specificity. Our studies establish the validity of using nonindigenous insect species with long-term data sets on natural enemies associated with them as a tool for testing hypotheses about host specificity.

Animals↗

Evidence from small-subunit ribosomal RNA sequences for a fungal origin of Microsporidia.

The phylum Microsporidia comprises a species-rich group of minute, single-celled, and intra-cellular parasites. Lacking normal mitochondria and with unique cytology, microsporidians have sometimes been thought to be a lineage of ancient eukaryotes. Although phylogenetic analyses using small-subunit ribosomal RNA (SSU-rRNA) genes almost invariably place the Microsporidia among the earliest branches on the eukaryotic tree, many other molecules suggest instead a relationship with fungi. Using maximum likelihood methods and a diverse SSU-rRNA data set, we have re-evaluated the phylogenetic affiliations of Microsporidia. We demonstrate that tree topologies used to estimate likelihood model parameters can materially affect phylogenetic searches. We present a procedure for reducing this bias: "tree-based site partitioning," in which a comprehensive set of alternative topologies is used to estimate sequence data partitions based on inferred evolutionary rates. This hypothesis-driven approach appears to be capable of utilizing phylogenetic information that is not available to standard likelihood implementations (e.g., approximation to a gamma distribution); we have employed it in maximum likelihood and Bayesian analysis. Applying our method to a phylogenetically diverse SSU-rRNA data set revealed that the early diverging ("deep") placement of Microsporidia typically found in SSU-rRNA trees is no better than a fungal placement, and that the likeliest placement of Microsporidia among non-long-branch eukaryotic taxa is actually within fungi. These results illustrate the importance of hypothesis testing in parameter estimation, provide a way to address certain problems in difficult data sets, and support a fungal origin for the Microsporidia.

Animals↗

Detection of Microsporidia, cryptosporidia and Giardia in swimming pools: a one-year prospective study.

In order to estimate the rate of microsporidia, cryptosporidia and giardia contamination of swimming pools, sequential samples of water were collected during a one-year period in six different swimming pools in Paris, France. Fourty-eight samples were submitted to filtrations. Eluates were examined for microsporidia using polymerase chain reaction (PCR) and for cryptosporidia and giardia using immunofluorescence staining. One of 48 specimens was positive for microsporidia. Using DNA sequence analysis, unknown microsporidia species were identified, which were close to an insect microsporidia Endoreticulatus schubergi. One sample was positive for cryptosporidia and none were positive for giardia. This study shows a low level of swimming pool water contamination by microsporidia, cryptosporidia or giardia, demonstrating the efficacy of cleaning filtration and disinfection procedures used in French swimming pools.

Animals↗