Life Cycles Of Microsporidia Research Articles

Immunolocalization of an alternative respiratory chain in Antonospora (Paranosema) locustae spores: mitosomes retain their role in microsporidial energy metabolism.

Microsporidia are a group of fungus-related intracellular parasites with severely reduced metabolic machinery. They lack canonical mitochondria, a Krebs cycle, and a respiratory chain but possess genes encoding glycolysis enzymes, a glycerol phosphate shuttle, and ATP/ADP carriers to import host ATP. The recent finding of alternative oxidase genes in two clades suggests that microsporidial mitosomes may retain an alternative respiratory pathway. We expressed the fragments of mitochondrial chaperone Hsp70 (mitHsp70), mitochondrial glycerol-3-phosphate dehydrogenase (mitG3PDH), and alternative oxidase (AOX) from the microsporidium Antonospora (Paranosema) locustae in Escherichia coli. Immunoblotting with antibodies against recombinant polypeptides demonstrated specific accumulation of both metabolic enzymes in A. locustae spores. At the same time comparable amounts of mitochondrial Hsp70 were found in spores and in stages of intracellular development as well. Immunoelectron microscopy of ultrathin cryosections of spores confirmed mitosomal localization of the studied proteins. Small amounts of enzymes of an alternative respiratory chain in merogonial and early sporogonial stages, alongside their accumulation in mature spores, suggest conspicuous changes in components and functions of mitosomes during the life cycle of microsporidia and the important role of these organelles in parasite energy metabolism, at least at the final stages of sporogenesis.

Laboratory identification of the microsporidia.

The microsporidia belong to the phylum Microspora within the taxonomic group Protozoa; there are more than 140 genera and 1,200 species that are parasitic in all major animal groups. Already, seven genera (Enterocytozoon, Encephalitozoon, Nosema, Pleistophora, Vittaforma, Trachipleistophora, and Brachiola) and some that are unclassified have been confirmed to cause human infection. Septata intestinalis, an organism that was identified in 1994 as belonging to a new genus, i.e., Septata (Encephalitozoon-like), has been reclassified as Encephalitozoon intestinalis, although there is not universal agreement on this generic change. The genus Microsporidium is essentially a catch-all genus for organisms that have not yet been classified (or may never be classified due to a lack of specimen). Depending on acceptance of these generic designations, there may be seven or nine microsporidial genera. The microsporidia are obligate intracellular, spore-forming protists with no active metabolic stages outside of the host cell. The life cycle involves a proliferative merogonic stage followed by sporogony, which results in spores containing a tubular extrusion apparatus (polar tubule) for injecting infective spore contents into the host cell (Fig. ​(Fig.11). FIG. 1. Life cycle of microsporidia. Reprinted from reference 9. Because microsporidia have a membrane-bound nucleus, an intracytoplasmic membrane system, and chromosome separation on mitotic spindles, they are considered to be true eukaryotes. However, they are somewhat unusual in that they have 70S ribosomes, have no mitrochondria or peroxisomes, and have simple Golgi membranes. Also, the microsporidial genome is smaller and less complex than those of other eukaryotes. Several characteristics, including the presence of chitin in the spore wall, suggest a potential link to the fungi. Microsporidiosis, a disease resulting from infection with these important emerging opportunistic organisms, has been noted in human immunodeficiency virus (HIV)-infected patients, as well as in other immunocompromised patients, including transplant recipients. The first human case was reported in 1959; since then, cases of microsporidiosis in both those who are immunocompromised and those who have no underlying immune problems and are considered to be immunocompetent have been reported (Table ​(Table1)1) (2, 7, 21). TABLE 1. Microsporidia causing human infection

Parasites of Larval Black Flies (Diptera: Simuliidae) and Environmental Factors Associated with Their Distributions

A total of 26,673 midto late-instar larval black flies were collected from 115 stream sites in South Carolina. Host larvae with patent, externally visible infections of nematodes, fungus, or microsporidia were identified to species either morphologically or cytologically. Six parasite taxa were identified: nematodes of the family Mermithidae; the fungus Coelomycidium simulii; and the microsporidia Janacekia debaisieuxi, Polydispyrenia simulii, Amblyospora bracteatalvarians, and Caudospora simulii. Of 43 species of black flies, 25 harbored at least one of these parasite taxa. Records from museum specimens increased the number of infected host species in South Carolina to 34 species. Prevalence of patent infections for each parasite taxon was low, ranging from 0.1 to 6.5% of all larvae. The distribution of nematodes among stream sites was nonrandom and correlated with season, host abundance, and stream-site conditions. The association of nematode distributions with stream parameters might reflect environmental influences on the nematodes during their free-living period and on host susceptibility to infection. Additional keywords: Mermithidae, Coelomycidium simulii, microsporidia Larval black flies (Diptera: Simuliidae) are a major component of the macroinvertebrate fauna in streams and rivers (Adler & McCreadie 1997). They anchor themselves to a silken pad spun on solid substrates (e.g., rocks, trailing vegetation) and filter food from the water column or graze it from the substrate. Larval and pupal development is temperature dependent and requires from one week to half a year (Crosskey 1990). After emergence from the pupa, females of most species mate, acquire sugar for energy and a vertebrate bloodmeal for egg maturation, and find a suitable stream for oviposition. Larvae are hosts of a variety of parasites, especially mermithid nematodes (Mermithidae), microsporidian protozoans (Microspora), and the chytrid fungus Coelomycidium simulii DEBAISIEUX. Preparasitic mermithid nematodes crawl about on the stream substrate and use a protrusible stylet to penetrate the host body (Molloy 1981). Infected host larvae are recognized by the presence of one or more coiled worms visible through the abdominal cuticle. Mermithids either exit and kill the host while it is still in the larval stage or pass into the adult stage, exiting shortly thereafter (Crosskey 1990). Postparasitic worms molt to adults, mate, and deposit eggs in the streambed (Poinar 1981). a Author for correspondence. E-mail: jmccrd@clemson.edu Microsporidian infections cause pathological changes, primarily to the host fat body (Weiser & Undeen 1981). Patent infections are recognized by the presence of large, irregular cysts that distend the host abdomen. Life cycles of microsporidia that attack black flies are not well understood, but larval hosts become infected both by transovarial transmission and by ingesting the free-living spores (Crosskey 1990). Spores might require some form of environmental conditioning before they become infective, and alternate hosts might be required to complete the life cycle (Jamnback 1973; Lacey & Undeen 1987). Larvae with patent infections of the fungus C. simulii are packed with minute, spherical thalli throughout the body cavity. Thalli give rise to spores that are released into the water column after the death of the host (Tarrant 1984). An intermediate host might be required to complete the life cycle (Lacey & Undeen 1987). In the present study, we survey selected parasites of larval black flies in South Carolina. Given that nematodes have free-living stages, we ask whether these parasites occupy a restricted range of stream conditions within the broader range of conditions under which their hosts are found. In other words, can occurrence (presence/absence) of nematodes in a particular host species, at a particular location, be predicted This content downloaded from 157.55.39.84 on Wed, 15 Jun 2016 06:30:20 UTC All use subject to http://about.jstor.org/terms Parasites of larval black flies by local stream conditions? Our objective is to examine factors that can explain parasite distributions, with the tenet that patterns of distribution can provide insight into the basic ecology of these organisms. Other parasites (microsporidia, C. simulii) were collected in numbers too low to address these questions.

Verification of intermediate hosts in the life cycles of microsporidia by small subunit rDNA sequencing.

Small subunit rDNA sequences were obtained from field-collected Amblyospora connecticus (Microsporida: Amblyosporidae) spores isolated from the mosquito, Aedes cantator (Diptera: Culicidae), and from field collected spores isolated from the putative intermediate host, Acanthocyclops vernalis (Copepoda: Cyclopidae). The ribosomal DNA sequences of the spores isolated from the two hosts were identical. These findings corroborate previous laboratory transmission studies and validate the intermediary role of A. vernalis in the life cycle of this microsporidium. These data represent the first comparative sequence analysis of a microsporidium isolated from its definitive and intermediate hosts. The results demonstrate the effectiveness of using rDNA sequence data for screening potential intermediate hosts. Unlike laboratory transmission tests, which can take months or years to complete, this technique can be completed in days and can be performed on a single infected organism.

Intermediate host for an Amblyospora sp. (Microspora) infecting the mosquito, Culex annulirostris

Experiments conducted in Australia are described which show that there is an intermediate host involved in the life cycle of an Amblyospora sp. infecting the mosquito, Culex annulirostris. Microsporidian spores produced in the cyclopoid copepod, Mesocyclops albicans, are infectious to laboratory colony C. annulirostris larvae and produce developmental stages identical to those of natural Amblyospora infections in this mosquito. These larval infections result in uninucleate spores in progeny mosquitoes which, in turn, are infectious to M. albicans. Subsequent studies conducted with Amblyospora, copepods, and mosquitoes in the United States to replicate and reconfirm results obtained in Australia were successful. This is the first substantiated evidence of the involvement of intermediate hosts in the life cycles of microsporidia, and the discovery now provides us with information needed to evaluate these organisms as biological control agents for mosquitoes and other disease vectors.

Meiosis in Microsporidia: effects on biological cycles

Synaptinemal complexes have been demonstrated in 7 microsporidian species belonging to 6 different genera (Gurleya, Thelohania, Pleistophora, Tuzetia, Baculea, Glugea). Thus, it can be presumed that a meiosis and consequently a karyogamy occur during their life cycle. Meisis occurs at the beginning of sporogony; therefore, karyogamy, must occur between spore and merogany, i.e. during the poorly known part of the life cycle. In the microsporidian species studied, with uninucleate spores and diplokaryotic merogony (Thelohania for instance), the 2 joined nuclei, each of them containing meiotic chromosomes, not only fail to fuse, but actually separate at the beginning of sporogony; afterwards, each of them undergoes meiosis. Their separation is accompanied by the appearance of an organelle whose structure and function are poorly understood. However, its structure resembles that of the kinetic center. The Nosema species studied do not have synaptinemal complexes; thus, their life cycle is difficult to understand: either karyogamy and meiosis occur during the unobserved part of the life-cycle, or sexual phanomena are absent altogether. In the latter case, the Nosema-type life cycle might be limited to vegetative multiplication which could be explained by the dimorphism theory of Microsporidia. It is shown also in the present study that the life cycle of Microsporidia does not involve haploid organisms which it might be thought to contain by comparing it with the cycles of sporozoa.