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Microsporidia are obligate intracellular parasites, more closely related to fungi than protozoa on molecular phylogenetic analysis, and are known to be a rare cause of opportunistic infection in immune compromised patients including human immunodeficiency virus-positive patients and solid organ transplant recipients. We report the first case to our knowledge of microsporidial myositis in a lung transplant recipient. He was 49 years old and had received a lung transplant in 2000 for cystic fibrosis. He presented in 2009 with fevers, chronic diarrhea, myalgia, and pancytopenia, and developed progressive weakness and neurological symptoms before his death 35 days after hospital admission. Multiple investigations, including stool culture, rectal biopsy, colonoscopy, cerebrospinal fluid examination, bone marrow biopsy, lung biopsy, and bronchoalveolar lavage, failed to reveal a definite cause for the patient's deterioration. The diagnosis of microsporidial infection was made on post-mortem light microscopic examination of tissue sections of the tongue and deltoid muscle. Light microscopy diagnosed a microsporidial myositis, confirmed by transmission electron microscopy, which suggested that the organism was Brachiola species. The identity of the organism was confirmed by polymerase chain reaction as Brachiola algerae (recently renamed Anncaliia algerae). The case highlights the need to consider protozoal organisms in the differential diagnosis of myalgia and multisystemic infections in immune compromised patients., (© 2012 John Wiley & Sons A/S.)

The ultrastructure of a new microsporidian species Microgemma vivaresi n. sp. causing liver cell xenoma formation in sea scorpions, Taurulus bubalis, is described. Stages of merogony, sporogony, and sporogenesis are mixed in the central cytoplasm of developing xenomas. All stages have unpaired nuclei. Uninucleate and multinucleate meronts lie within vacuoles formed from host endoplasmic reticulum and divide by binary or multiple fission. Sporonts, no longer in vacuoles, deposit plaques of surface coat on the plasma membrane that cause the surface to pucker. Division occurs at the puckered stage into sporoblast mother cells, on which plaques join up to complete the surface coat. A final binary fission gives rise to sporoblasts. A dense globule, thought to be involved in polar tube synthesis, is gradually dispersed during spore maturation. Spores are broadly ovoid, have a large posterior vacuole, and measure 3.6 microm x 2.1 microm (fresh). The polar tube has a short wide anterior section that constricts abruptly, then runs posteriad to coil about eight times around the posterior vacuole with granular contents. The polaroplast has up to 40 membranes arranged in pairs mostly attached to the wide region of the polar tube and directed posteriorly around a cytoplasm of a coarsely granular appearance. The species is placed alongside the type species Microgemma hepaticusRalphs and Matthews 1986 within the family Tetramicridae, which is transferred from the class Dihaplophasea to the class Haplophasea, as there is no evidence for the occurrence of a diplokaryotic phase.

Reinvestigation of Cystosporogenes operophterae [J. Parasitol. 46 (1960) 755] by electron microscopy confirmed that development in host cells takes place in a vacuole with a single membrane at its boundary. Although ribosomes were not clustered on this membrane, it is hypothesised that it originates from host endoplasmic reticulum. The dome-shaped anchoring disc, the morphology of the polaroplast and the separation of the polar tube coils from the ribosome-packed cytoplasm are newly described details of spore structure. The polaroplast consists of an outer region of compact lamellae forming 'arms' surrounding an inner region of widely spaced lamellae. The 'arms' extends back into the region of an elongate nucleus. The genera Cystosporogenes and Endoreticulatus were differentiated by their positions in a previously obtained 16S rDNA phylogeny and on the new ultrastructural data.

Different courses of microsporidiosis, related to the route of infection, were observed in severe combined immunodeficient (SCID) mice inoculated with spores of the human microsporidian Trachipleistophora hominis (Phylum Microspora). After eye contamination by spores the mice became moribund within 7 to 8 weeks, showing severe infection in the conjunctiva and cornea, and lighter infections in the urinary bladder, liver and spleen. The mean survival time of intramuscularly inoculated SCID mice was 12 weeks, when heavy infection was found in muscles around the site of inoculation, and also in several viscera. Subcutaneously inoculated SCID mice developed skin lesions around the inoculation sites, and heavy urinary bladder infection, and died 6 or 7 weeks after inoculation. Intracerebrally inoculated SCID mice became moribund 5 or 6 weeks after inoculation with massive infection in the urinary bladder and liver, but none in the brain. Intraperitoneally inoculated SCID mice survived for 13 weeks and the urinary bladder and liver were the most heavily infected organs. The SCID mice, inoculated perorally and examined after 23 weeks, were uninfected. Infection was not detected in the brain of any of the inoculated SCID mice. Our results show that T. hominis has very little tissue specificity. Peroral infection seems to be ineffective in T. hominis, but eye conta mination or insect bite (as mimicked by injection) are suggested as possible routes of infection under natural conditions.

Glugoides intestinalis, Microsporidium sp., Ordospora colligata, Gurleya vavrai, Larssonia obtusa and Flabelliforma magnivora are microsporidian parasites of planctonic freshwater crustaceans Daphnia spp. We performed a phylogenetic analysis of the small subunit ribosomal DNA which revealed their positions as polyphyletic. G. intestinalis, Microsporidium sp. and O. colligata, which are horizontally transmitted gut parasites with small spores and low virulence, group with different lineages. G. intestinalis is related to 2 microsporidia infecting lepidopterans and to Vittaforma corneae, which has been described as a human pathogen. It is thought that V. corneae may have an invertebrate as its natural host. Microsporidium sp. is a relative of the genera Enterocytozoon and Nucleospora, pathogens of man and fish respectively. O. colligata is the first species found to be closely related to the genus Encephalitozoon, which is comprised of 3 species that are parasites of homeothermic vertebrates. G. vavrai and L. obtusa are sister taxa that branch close to the Amblyosporidae, the only microsporidia with known intermediate hosts. This finding supports the presumption of secondary hosts for G. vavrai and L. obtusa, as it has not been possible to maintain these species in Daphnia in the laboratory. F. magnivora roots deep at the base of the phylum microsporidia with no close relative found so far.

Myositis is a common clinical syndrome in advanced stages of AIDS. Trachipleistophora hominis (phylum Microspora) has been detected in several cases of painful, immobilising myositis in AIDS patients. Enzyme linked immunosorbent assays (ELISAs) and Western blotting of protein profiles separated by SDS PAGE were used to determine whether this species could be detected and differentiated by serology. Sixteen microsporidia, including several species known to infect man and species infecting fish, crustaceans and a mosquito, were used as antigen. Each species had a unique profile of SDS PAGE-separated proteins. In Western blots, mouse antiserum, raised to T. hominis and selected for its high ELISA specificity, bound to antigens ranging from less than 25 kDa to greater than 250 kDa with major bands at 39-44 kDa and 98-150 kDa on T. hominis protein profiles. The serum also recognised some high molecular weight antigens in the profiles of Vavraia culicis, Heterosporis anguillarum, and three species of Pleistophora but none in the remaining genera examined. It was concluded that ELISA and Western blotting could be used to detect and differentiate T. hominis in muscle biopsy tissue from patients with myositis. However, sera from T. hominis-infected patients in the terminal stages of AIDS would not be useful for detection of infections because of a sharp decline in antibody level.

Molecular data have proved useful as an alternative to morphological data in showing the relationships of genera within the phylum Microsporidia, but until now have been available only for ribosomal genes. In previous studies protein-coding genes of microsporidia have been used only to assess their position in the evolution of eukaryotes. For the first time we report on the use of a protein-coding gene, the A-G region of the largest subunit of RNA polymerase II (RPB1) from 14 mainly polysporous species, to generate an alternative phylogeny for microsporidia. Using the amino acid sequences, the genera and species fell into the same main groupings as had been obtained with 16S rDNA sequences, but the RPB1 data provided better resolution within these groups. The results supported the pairings of Trachipleistophora hominis with Vavraia culicis and Pleistophora hippoglossoideos with Pleistophora typicalis. They also confirmed that the genus Pleistophora is not monophyletic and that it will be necessary to transfer Pleistophora ovariae and Pleistophora mirandellae into one or more other genera, as has already been effected for Pleistophora anguillarum.

The microsporidium Trachipleistophora hominis Hollister, Canning, Weidner, Field, Kench et Marriott, 1996, originally isolated from human skeletal muscle cells, inhibited myotube formation from myoblasts when grown in a mouse myoblast cell line C2,C12. Uninfected cultures readily converted to myotubes. Albendazole, a drug with known antimicrosporidial activity, was tested against T. hominis in C2,C12 cells. The drug was added when infection had reached 75% of C2,C12 cells, a level comparable to that obtained in heavily infected muscle in vivo. Doses of 1 ng/ml and 10 ng/ml had no effect on merogony or sporogony. In cultures exposed to 100 ng/ml albendazole, the C2,C12 cells remained in good condition while infection levels dropped to 25% over 7 weeks. Drug doses of 500 ng/ml and 1,000 ng/ml were deleterious to the host cells but some spores retained viability and were able to establish new infections once albendazole pressure was removed. T. hominis meronts exposed to 100 ng/ml albendazole mostly lacked the normally thick surface coat and its reticulate extensions. Meronts were not seen in cultures exposed to higher drug doses. Albendazole at a concentration of 100 ng/ml and higher had a profound effect on spore morphogenesis. There was erratic coiling of the polar tube, often involving the formation of double tubes, and chaotic disposition of membranes which could have been those of polaroplast. The in vitro susceptibility of T. hominis to albendazole was low in comparison with in vitro susceptibility of other microsporidia of human origin.

Collections of Mansonia africana mosquito larvae were made at one site in N.E. Tanzania in 1985 and 1987 and from two additional sites, both within about 2 mi of the original one in 1987. An octosporous microsporidian, present at all three sites, was found in both years infecting from 7 to 22% of larvae. Spores (stained in Giemsa) measured 3.0 microns +/- 0.25 microns x 2.25 microns +/- 0.26 microns. Ultrastructurally, spores were seen to have an anterior rim surrounding a depressed area where the endospore was at its thinnest. In transmission electron microscopy section, the rim appeared as two processes into which all layers of the wall extended. At the posterior end all layers of the wall extended into a simple knob-like structure which could be interpreted as a section through a crest running longitudinally around the spore. The polar filament was anisofilar, with two anterior coils of greater diameter than the three posterior coils. Although most closely resembling the genera Amblyspora and Parathelohania in the family Thelohaniidae, the species in M. africana differs from the former, which has oval spores, broadly rounded at the ends, and from the latter, which has a prominent, ridged posterior extension to the spores. The new species has been placed in a new genus and the name Tricornia muhezae proposed.

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.