The Experts below are selected from a list of 276 Experts worldwide ranked by ideXlab platform
Jung-sheng Chen - One of the best experts on this subject based on the ideXlab platform.
-
Swimming Pool-Associated Vittaforma-Like Microsporidia Linked to Microsporidial Keratoconjunctivitis Outbreak, Taiwan.
Emerging infectious diseases, 2019Co-Authors: Jung-sheng Chen, Tung-yi Huang, Tsui-kang Hsu, Bing-mu Hsu, Shih-chun Chao, Pei-yu Yang, I-hsiu HuangAbstract:We analyzed 2 batches of environmental samples after a microsporidial keratoconjunctivitis outbreak in Taiwan. Results indicated a transmission route from a parking lot to a foot washing pool to a swimming pool and suggested that accumulation of mud in the foot washing pool during the rainy season might be a risk factor.
-
Molecular surveillance of Vittaforma-like microsporidia by a small-volume procedure in drinking water source in Taiwan: evidence for diverse and emergent pathogens
Environmental Science and Pollution Research, 2018Co-Authors: Jung-sheng Chen, Hsin-chi Tsai, Yu-pin Chen, Tung-yi Huang, Kuan-ying Li, Dar-der JiAbstract:Vittaforma corneae belongs to microsporidia, which include over 1500 species of opportunistic obligate intracellular fungi infecting almost all known animal taxa. Although outbreaks of ocular infections caused by waterborne V. corneae have been reported in recent years, little is known about the occurrence of this pathogen in aquatic environments. In this study, 50 water samples from rivers and reservoirs around Taiwan in two seasons were analyzed to explore the presence of this pathogen in natural aquatic environments. A high detection rate of Vittaforma -like amplicons (94%; 47/50) was observed in the water samples when examined by nested PCR with primer pairs specific to the small ribosomal subunit (SSU) rRNA gene. After electrophoresis, many lanes showed multiband patterns with expected molecular weights. After confirmation by DNA sequencing and by sequence alignment in the NCBI database, we identified a variety of Vittaforma -like microsporidia with weak sequence similarity, with approximately 85% identity to V. corneae , thus indicating high diversity of microsporidia in aquatic environments. Phylogenetic analysis showed clear-cut microsporidian clade classification and indicated that the most Vittaforma -like microsporidia in this study belong to clade IV and cluster into four major groups. The first group is similar to the microsporidia associated with ocular microsporidiosis. The second group is associated with the diarrheal pathogens, whereas the third and fourth groups are a novel group and a zoonotic group, respectively. This study provides abundant sequencing information, which will be useful for future molecular biological studies on microsporidia. Because microsporidia are important pathogens of animals and humans, it is urgently necessary to determine via a survey whether there are species with potential threats that have not yet been revealed.
-
Surveillance of Vittaforma corneae in hot springs by a small-volume procedure.
Water research, 2017Co-Authors: Jung-sheng Chen, Tung-yi Huang, Tsui-kang Hsu, Bing-mu Hsu, Yu-li Huang, Men-fang ShaioAbstract:Vittaforma corneae is an obligate intracellular fungus and can cause human ocular microsporidiosis. Although accumulating reports of V. corneae causing keratoconjunctivitis in both healthy and immunocompromised persons have been published, little is known about the organism's occurrence in aquatic environments. Limitations in detection sensitivity have meant a large sampling volume is required to detect the pathogen up to now, which is problematic. A recent study in Taiwan has shown that some individuals suffering from microsporidial keratitis (MK) were infected after exposure to the pathogen at a hot spring. As a consequence of this, a survey and analysis of environmental V. corneae present in hot springs became an urgent need. In this study, sixty water samples from six hot spring recreation areas around Taiwan were analyzed. One liter of water from each sample site was filtered to harvest the fungi. The positive samples were detected using a modified nested PCR approach followed by sequencing using specific SSU rRNA gene primer pairs for V. corneae. In total fifteen V. corneae-like isolates were identified (25.0% of sites). Among them, six isolates, which were collected from recreational areas B, C and D, were highly similar to known V. corneae keratitis strains from Taiwan and other countries. Furthermore, five isolates, which were collected from recreation areas A, C, E and F, were very similar to Vittaforma-like diarrhea strains isolated in Portugal. Cold spring water tubs and public foot bath pools had the highest detection rate (50%), suggesting that hot springs might be contaminated via untreated water sources. Comparing the detection rate across different regions of Taiwan, Taitung, which is in the east of the island, gave the highest positive rate (37.5%). Statistical analysis showed that outdoor/soil exposure and a high heterotrophic plate count (HPC) were risk factors for the occurrence of V. corneae. Our findings provide empirical evidence supporting the need for proper control and regulations at hot spring recreational waters in order to avoid health risks from this pathogen. Finally, we have developed a small volume procedure for detecting V. corneae in water samples and this has proved to be very useful.
Elizabeth S. Didier - One of the best experts on this subject based on the ideXlab platform.
-
Encephalitozoon cuniculi and Vittaforma corneae (Phylum Microsporidia) inhibit staurosporine-induced apoptosis in human THP-1 macrophages in vitro
Parasitology, 2018Co-Authors: Yuliya Y. Sokolova, Lisa C. Bowers, Xavier Alvarez, Elizabeth S. DidierAbstract:Microsporidia are ubiquitous fungi-related, obligate intracellular parasites that evolved with extremely reduced genomes and proteomes to become highly dependent upon the host cell biochemistry (Cuomo et al., 2012; Peyretaillade et al., 2011). This host cell dependency has been highlighted by the ability of several microsporidia species that secrete factors and appropriate host cell metabolism in favor of their infection and replication (Cuomo et al., 2012; Ferguson & Lucocq, 2018; He et al., 2015; Heinz et al., 2014; Luallen et al., 2015; Panek et al., 2014; Senderskiy et al., 2014; Watson et al., 2015; Wiredu Boakye et al., 2017). In addition, microsporidia can induce cell hypertrophy, giant cell formation, and increase lifespan of infected cells (Aoki Mdel et al., 2006; Desjardins et al., 2015; Leitch et al., 2005; Stentiford et al., 2014; Timofeev et al., 2016). Apoptosis of infected cells is a universal mechanism used by the host to eliminate pathogens and facilitate the release of pathogen antigens to then promote adaptive humoral and cell-mediated immune responses in mammals (Liu et al., 2009). Signaling pathways involved in regulating the host cell cycle and apoptosis however also are primary targets of intracellular prokaryotic and eukaryotic parasites for subverting innate immune responses (Faherty & Maurelli, 2008; Hay & Kannourakis, 2002; Huang et al., 2016; James & Green, 2004; Luder et al., 2001). A number of intracellular parasites, including several species of microsporidia, have evolved means to subvert apoptosis mechanisms to maintain or prolong infection (del Aguila et al., 2006; James & Green, 2004; Scanlon et al., 1999). For example, Anncaliia algerae inhibited induced apoptosis in human lung fibroblasts (Scanlon et al., 1999) and infection with Encephalitozoon spp. disrupted the host cell cycle of Vero cells (Scanlon et al., 2000). E. cuniculi also suppressed induced apoptosis in Vero cells and modulated the host cell cycle via inhibiting tumor suppressor factor p53 (del Aguila et al., 2006). V. corneae induced formation of multinucleated xenoma-like structures during infection of monkey kidney cells (Leitch et al., 2005) and Nosema bombycis suppressed apoptosis in a Bombix mori ovarian cell line (He et al., 2015). Furthermore, Nosema ceranae and Nosema apis can modulate the host cycle and suppress apoptosis of the ventricular epithelium cells of Apis melifera (Higes et al., 2013; Huang et al., 2016; Kurze et al., 2016; Kurze et al., 2015; Martin-Hernandez et al., 2017). Macrophages are key cells in the innate immune system of vertebrates and invertebrates and comprise the first line of defense against intracellular parasites for then transitioning to adaptive immune response in higher vertebrates. Macrophages can kill microsporidia via activation signaling processes yet also serve as host cells of infection for several species of microsporidia. Among these are the Encephalitozoon species, i.e. E. cuniculi, E. hellem, and E. intestinalis, as well as V. corneae, species that infect mammals, including humans, and that can be grown in tissue culture. Encephalitozoon spp. replicate within host cell-derived parasitophorous vacuoles while V. corneae replicates in direct contact with the host cell cytoplasm. Based on the earlier reports that microsporidia affect the host cell machinery and apoptosis pathways, the purpose of this study was to examine the effects of these two biologically dissimilar human-infecting microsporidia, E. cuniculi and V. corneae, on staurosporine-induced apoptosis in the human macrophage-differentiated cell line, THP-1. The results demonstrated that viable microsporidia, but not heat-killed organisms, inhibited apoptosis of the THP-1 macrophages.
-
Antimicrosporidial activity of (fluoro)quinolones in vitro and in vivo
2015Co-Authors: Elizabeth S. Didier, Derek Mittleider, Paul J. Brindley, Mary E. Stovall, Dorothy J. Kuebler, Lisa Bowers, Peter J. DidierAbstract:Abstract. Microsporidia are a cause of emerging and opportunistic infections in humans and animals. Although two drugs are currently being used to treat microsporidiosis, concerns exist that albendazole is only selective for inhibiting some species of microsporidia that infect mammals, and fumagillin appears to have been found to be toxic. During a limited sequence survey of the Vittaforma corneae (syn. Nosema corneum Shadduck, Meccoli, Davis et Font, 1990) genome, a partial gene encoding for the ParC topoisomerase IV subunit was identified. The purpose of this set of studies was to determine if fluoroquinolones, which target topoisomerase IV, exert activity against Encephalitozoon intestinalis (syn. Septata intestinalis Cali, Kotler et Orenstein, 1993) and V. corneae in vitro, and whether these compounds could prolong survival of V. corneae-infected athymic mice. Fif-teen fluoroquinolones were tested. Of these, norfloxacin and ofloxacin inhibited E. intestinalis replication by more than 70% compared with non-treated control cultures, while gatifloxacin, lomefloxacin, moxifloxacin, and nalidixic acid (sodium salt) inhibited both E. intestinalis and V. corneae by at least 60 % at concentrations not toxic to the host cells. These drugs were tested in vivo also, where gatifloxacin, lomefloxacin, norfloxacin, and ofloxacin prolonged survival of V. corneae-infected athymic mice (P < 0.05), whereas moxifloxacin and nalidixic acid failed to prolong survival. Therefore, these results support continued studies for evaluating the efficacy of the fluoroquinolones for treating microsporidiosis and for characterizing the target(s) of these fluoroquinolones in the microsporidia. Microsporidia are single-celled, obligately intracellu
-
Antimicrosporidial Activities of Fumagillin, TNP-470, Ovalicin, and Ovalicin Derivatives In Vitro and In Vivo
Antimicrobial agents and chemotherapy, 2006Co-Authors: Peter J. Didier, Paul J. Brindley, Lisa C. Bowers, Mary E. Stovall, Dorothy J. Kuebler, Jennifer N. Phillips, Mohamed Nasr, Elizabeth S. DidierAbstract:Therapies for microsporidiosis in humans are limited, and fumagillin, which appears to be the most broadly effective antimicrosporidial drug, is considered to be moderately toxic. The purpose of this study was to apply an in vitro drug screening assay for Encephalitozoon intestinalis and Vittaforma corneae and an in vivo athymic mouse model of V. corneae infection to assess the efficacy of TNP-470 (a semisynthetic analogue of fumagillin), ovalicin, and eight ovalicin derivatives. TNP-470, ovalicin, and three of the ovalicin derivatives inhibited both E. intestinalis and V. corneae replication by more than 70% in vitro. Another three of the ovalicin derivatives inhibited one of the two microsporidian species by more than 70%. None of the treated athymic mice survived the V. corneae infection, but they did survive statistically significantly longer than the untreated controls after daily treatment with fumagillin administered at 5, 10, and 20 mg/kg of body weight subcutaneously (s.c.), TNP-470 administered at 20 mg/kg intraperitoneally (i.p.), or ovalicin administered at 5 mg/kg s.c. Of two ovalicin derivatives that were assessed in vivo, NSC 9665 given at 10 mg/kg i.p. daily also statistically significantly prolonged survival of the mice. No lesions associated with drug toxicity were observed in the kidneys or livers of uninfected mice treated with these drugs at the highest dose of 20 mg/kg daily. These results thus support continued studies to identify more effective fumagillin-related drugs for treating microsporidiosis.
-
Antimicrosporidial activity of (fluoro)quinolones in vitro and in vivo.
Folia parasitologica, 2005Co-Authors: Elizabeth S. Didier, Derek Mittleider, Paul J. Brindley, Lisa C. Bowers, Mary E. Stovall, Dorothy J. Kuebler, Peter J. DidierAbstract:Microsporidia are a cause of emerging and opportunistic infections in humans and animals. Although two drugs are currently being used to treat microsporidiosis, concerns exist that albendazole is only selective for inhibiting some species of microsporidia that infect mammals, and fumagillin appears to have been found to be toxic. During a limited sequence survey of the Vittaforma corneae genome, a partial gene encoding for the ParC topoisomerase IV subunit was identified. The purpose of this set of studies was to determine if fluoroquinolones, which target topoisomerase IV, exert activity against Encephalitozoon intestinalis and V. corneae in vitro, and whether these compounds could prolong survival of V. corneae-infected athymic mice. Fifteen fluoroquinolones were tested. Of these, norfloxacin and ofloxacin inhibited E. intestinalis replication by more than 70% compared with non-treated control cultures, while gatifloxacin, lomefloxacin, moxifloxacin, and nalidixic acid (sodium salt) inhibited both E. intestinalis and V. corneae by at least 60% at concentrations not toxic to the host cells. These drugs were tested in vivo also, where gatifloxacin, lomefloxacin, norfloxacin, and ofloxacin prolonged survival of V. corneae-infected athymic mice (P < 0.05), whereas moxifloxacin and nalidixic acid failed to prolong survival. Therefore, these results support continued studies for evaluating the efficacy of the fluoroquinolones for treating microsporidiosis and for characterizing the target(s) of these fluoroquinolones in the microsporidia.
-
microsporidiosis an emerging and opportunistic infection in humans and animals
Acta Tropica, 2005Co-Authors: Elizabeth S. DidierAbstract:Microsporidia have emerged as causes of infectious diseases in AIDS patients, organ transplant recipients, children, travelers, contact lens wearers, and the elderly. These organisms are small single-celled, obligate intracellular parasites that were considered to be early eukaryotic protozoa but were recently reclassified with the fungi. Of the 14 species of microsporidia currently known to infect humans, Enterocytozoon bieneusi and Encephalitozoon intestinalis are the most common causes of human infections and are associated with diarrhea and systemic disease. Species of microsporidia infecting humans have been identified in water sources as well as in wild, domestic, and food-producing farm animals, raising concerns for waterborne, foodborne, and zoonotic transmission. Current therapies for microsporidiosis include albendazole which is a benzimidazole that inhibits microtubule assembly and is effective against several microsporidia, including the Encephalitozoon species, but is less effective against E. bieneusi. Fumagillin, an antibiotic and anti-angiogenic compound produced by Aspergillus fumigatus, is more broadly effective against Encephalitozoon spp. and Enterocytozoon bieneusi but is toxic when administered systemically to mammals. Gene target studies have focused on methionine aminopeptidase 2 (MetAP2) for characterizing the mechanism of action and for identifying more effective, less toxic fumagillin-related drugs. Polyamine analogues have shown promise in demonstrating anti-microsporidial activity in culture and in animal models, and a gene encoding topoisomerase IV was identified in Vittaforma corneae, raising prospects for studies on fluoroquinolone efficacy against microsporidia.
Tung-yi Huang - One of the best experts on this subject based on the ideXlab platform.
-
Swimming Pool-Associated Vittaforma-Like Microsporidia Linked to Microsporidial Keratoconjunctivitis Outbreak, Taiwan.
Emerging infectious diseases, 2019Co-Authors: Jung-sheng Chen, Tung-yi Huang, Tsui-kang Hsu, Bing-mu Hsu, Shih-chun Chao, Pei-yu Yang, I-hsiu HuangAbstract:We analyzed 2 batches of environmental samples after a microsporidial keratoconjunctivitis outbreak in Taiwan. Results indicated a transmission route from a parking lot to a foot washing pool to a swimming pool and suggested that accumulation of mud in the foot washing pool during the rainy season might be a risk factor.
-
Molecular surveillance of Vittaforma-like microsporidia by a small-volume procedure in drinking water source in Taiwan: evidence for diverse and emergent pathogens
Environmental Science and Pollution Research, 2018Co-Authors: Jung-sheng Chen, Hsin-chi Tsai, Yu-pin Chen, Tung-yi Huang, Kuan-ying Li, Dar-der JiAbstract:Vittaforma corneae belongs to microsporidia, which include over 1500 species of opportunistic obligate intracellular fungi infecting almost all known animal taxa. Although outbreaks of ocular infections caused by waterborne V. corneae have been reported in recent years, little is known about the occurrence of this pathogen in aquatic environments. In this study, 50 water samples from rivers and reservoirs around Taiwan in two seasons were analyzed to explore the presence of this pathogen in natural aquatic environments. A high detection rate of Vittaforma -like amplicons (94%; 47/50) was observed in the water samples when examined by nested PCR with primer pairs specific to the small ribosomal subunit (SSU) rRNA gene. After electrophoresis, many lanes showed multiband patterns with expected molecular weights. After confirmation by DNA sequencing and by sequence alignment in the NCBI database, we identified a variety of Vittaforma -like microsporidia with weak sequence similarity, with approximately 85% identity to V. corneae , thus indicating high diversity of microsporidia in aquatic environments. Phylogenetic analysis showed clear-cut microsporidian clade classification and indicated that the most Vittaforma -like microsporidia in this study belong to clade IV and cluster into four major groups. The first group is similar to the microsporidia associated with ocular microsporidiosis. The second group is associated with the diarrheal pathogens, whereas the third and fourth groups are a novel group and a zoonotic group, respectively. This study provides abundant sequencing information, which will be useful for future molecular biological studies on microsporidia. Because microsporidia are important pathogens of animals and humans, it is urgently necessary to determine via a survey whether there are species with potential threats that have not yet been revealed.
-
Surveillance of Vittaforma corneae in hot springs by a small-volume procedure.
Water research, 2017Co-Authors: Jung-sheng Chen, Tung-yi Huang, Tsui-kang Hsu, Bing-mu Hsu, Yu-li Huang, Men-fang ShaioAbstract:Vittaforma corneae is an obligate intracellular fungus and can cause human ocular microsporidiosis. Although accumulating reports of V. corneae causing keratoconjunctivitis in both healthy and immunocompromised persons have been published, little is known about the organism's occurrence in aquatic environments. Limitations in detection sensitivity have meant a large sampling volume is required to detect the pathogen up to now, which is problematic. A recent study in Taiwan has shown that some individuals suffering from microsporidial keratitis (MK) were infected after exposure to the pathogen at a hot spring. As a consequence of this, a survey and analysis of environmental V. corneae present in hot springs became an urgent need. In this study, sixty water samples from six hot spring recreation areas around Taiwan were analyzed. One liter of water from each sample site was filtered to harvest the fungi. The positive samples were detected using a modified nested PCR approach followed by sequencing using specific SSU rRNA gene primer pairs for V. corneae. In total fifteen V. corneae-like isolates were identified (25.0% of sites). Among them, six isolates, which were collected from recreational areas B, C and D, were highly similar to known V. corneae keratitis strains from Taiwan and other countries. Furthermore, five isolates, which were collected from recreation areas A, C, E and F, were very similar to Vittaforma-like diarrhea strains isolated in Portugal. Cold spring water tubs and public foot bath pools had the highest detection rate (50%), suggesting that hot springs might be contaminated via untreated water sources. Comparing the detection rate across different regions of Taiwan, Taitung, which is in the east of the island, gave the highest positive rate (37.5%). Statistical analysis showed that outdoor/soil exposure and a high heterotrophic plate count (HPC) were risk factors for the occurrence of V. corneae. Our findings provide empirical evidence supporting the need for proper control and regulations at hot spring recreational waters in order to avoid health risks from this pathogen. Finally, we have developed a small volume procedure for detecting V. corneae in water samples and this has proved to be very useful.
Men-fang Shaio - One of the best experts on this subject based on the ideXlab platform.
-
Surveillance of Vittaforma corneae in hot springs by a small-volume procedure.
Water research, 2017Co-Authors: Jung-sheng Chen, Tung-yi Huang, Tsui-kang Hsu, Bing-mu Hsu, Yu-li Huang, Men-fang ShaioAbstract:Vittaforma corneae is an obligate intracellular fungus and can cause human ocular microsporidiosis. Although accumulating reports of V. corneae causing keratoconjunctivitis in both healthy and immunocompromised persons have been published, little is known about the organism's occurrence in aquatic environments. Limitations in detection sensitivity have meant a large sampling volume is required to detect the pathogen up to now, which is problematic. A recent study in Taiwan has shown that some individuals suffering from microsporidial keratitis (MK) were infected after exposure to the pathogen at a hot spring. As a consequence of this, a survey and analysis of environmental V. corneae present in hot springs became an urgent need. In this study, sixty water samples from six hot spring recreation areas around Taiwan were analyzed. One liter of water from each sample site was filtered to harvest the fungi. The positive samples were detected using a modified nested PCR approach followed by sequencing using specific SSU rRNA gene primer pairs for V. corneae. In total fifteen V. corneae-like isolates were identified (25.0% of sites). Among them, six isolates, which were collected from recreational areas B, C and D, were highly similar to known V. corneae keratitis strains from Taiwan and other countries. Furthermore, five isolates, which were collected from recreation areas A, C, E and F, were very similar to Vittaforma-like diarrhea strains isolated in Portugal. Cold spring water tubs and public foot bath pools had the highest detection rate (50%), suggesting that hot springs might be contaminated via untreated water sources. Comparing the detection rate across different regions of Taiwan, Taitung, which is in the east of the island, gave the highest positive rate (37.5%). Statistical analysis showed that outdoor/soil exposure and a high heterotrophic plate count (HPC) were risk factors for the occurrence of V. corneae. Our findings provide empirical evidence supporting the need for proper control and regulations at hot spring recreational waters in order to avoid health risks from this pathogen. Finally, we have developed a small volume procedure for detecting V. corneae in water samples and this has proved to be very useful.
Govinda S. Visvesvara - One of the best experts on this subject based on the ideXlab platform.
-
Culture and Propagation of Microsporidia
The Microsporidia and Microsporidiosis, 2014Co-Authors: Govinda S. Visvesvara, Gordon J. Leitch, Hercules Moura, D A SchwartzAbstract:Isolation in culture should always be attempted even when a presumptive diagnosis has been made. This is particularly important for the purpose of establishing a bank of isolates to be used for antigenic, molecular, and biochemical analyses. The first attempt to culture microsporidia was made in 1937 by Trager, who was partially successful in establishing Nosema bombycis infection of a cell culture developed from the ovarian tube lining cells of the silkworm (Bombyx mori). Cultures of Encephalitozoon cuniculi were initiated in several different ways: by adding infected tissue explants to cultured cells, by allowing infected cells in explanted cells to grow, by allowing germination of spores in the presence of cells and thereafter infecting cells, and by scraping infected cells from infected cultures and adding them to fresh cell cultures. The first microsporidian isolated from a human, Vittaforma corneae (Nosema corneum) was cultured from a corneal biopsy that was shipped to the laboratory overnight in Hanks' balanced salt solution (HBSS). Only a few attempts have been made to isolate microsporidia into culture from feces because enteric bacteria and yeast usually overgrow the rich culture medium that is used, which impedes isolation of the fastidious microsporidia, especially Enterocytozoon bieneusi. Inoculated cell cultures should be examined frequently with an inverted microscope preferably equipped with phase-contrast or differential interference-contrast optics. One great advantage of obtaining spores from in vitro cultures is the absence of bacterial and fungal contaminants.
-
Multinucleate host cells induced by Vittaforma corneae (Microsporidia).
Folia parasitologica, 2005Co-Authors: Gordon J. Leitch, Andrew P. Shaw, Margaret Colden-stanfield, Mary Scanlon, Govinda S. VisvesvaraAbstract:The microsporidium Vittaforma corneae develops within the target cell cytoplasm. In the present study, green monkey kidney (E6) cells infected at 30 degrees C, 35 degrees C or 37 degrees C with V. corneae developed enlarged multinucleate structures of up to 200 microm in any horizontal dimension made up either of a single cell or of multiple fused cells. A number of epithelial cell types (SW-480, HT-29, Caco-2 and HCT-8) were infected with V. corneae but did not induce the same highly organized structures, suggesting that for the structure to develop, the host cell must be capable of continued mitosis, and not be differentiated or be detaching from the surface matrix. Live cell imaging of infected E6 cells revealed large, multinucleate infected cells characterized by a central focus from which radiated parasite stages and host cell mitochondria. Immunocytochemistry identifying gamma and alpha tubulin suggested that a single centrally-located microtubule organizing centre governed the distribution of parasite stages and host cell organelles, with mitochondria and parasites being eventually transported towards the periphery of the structure. Whole cell patch clamp analysis of infected cells indicated an average five-fold increase in total membrane capacitance, consistent with an enlarged single cell. Scanning electron microscopy revealed cell-like protrusions around the periphery of the structure with the intervening space being made up of parasites and cell debris. Clearly in the case of V. corneae-infected E6 cells the parasite-host cell relationship involves subverting the host cell cytoskeleton and cell volume control, providing the parasite with the same protected niche as does a xenoma.
-
Multinucleate host cells induced by Vittaforma corneae (Microsporidia). Folia Parasitol
2005Co-Authors: Gordon J. Leitch, Andrew P. Shaw, Margaret Colden-stanfield, Mary Scanlon, Govinda S. VisvesvaraAbstract:multinucleate giant cell Abstract. The microsporidium Vittaforma corneae (Shadduck, Meccoli, Davis et Font, 1990) develops within the target cell cytoplasm. In the present study, green monkey kidney (E6) cells infected at 30°C, 35°C or 37°C with V. corneae developed enlarged multinucleate structures of up to 200 µm in any horizontal dimension made up either of a single cell or of multiple fused cells. A number of epithelial cell types (SW-480, HT-29, Caco-2 and HCT-8) were infected with V. corneae but did not induce the same highly organized structures, suggesting that for the structure to develop, the host cell must be capable of continued mi-tosis, and not be differentiated or be detaching from the surface matrix. Live cell imaging of infected E6 cells revealed large, multinucleate infected cells characterized by a central focus from which radiated parasite stages and host cell mitochondria. Im-munocytochemistry identifying γ and α tubulin suggested that a single centrally-located microtubule organizing centre governed the distribution of parasite stages and host cell organelles, with mitochondria and parasites being eventually transported towards the periphery of the structure. Whole cell patch clamp analysis of infected cells indicated an average five-fold increase in total membrane capacitance, consistent with an enlarged single cell. Scanning electron microscopy revealed cell-like protrusions around the periphery of the structure with the intervening space being made up of parasites and cell debris. Clearly in the case of V. corneae-infected E6 cells the parasite-host cell relationship involves subverting the host cell cytoskeleton and cell volume control, providing the parasite with the same protected niche as does a xenoma
-
In Vitro Cultivation of Microsporidia of Clinical Importance
Clinical microbiology reviews, 2002Co-Authors: Govinda S. VisvesvaraAbstract:Although attempts to develop methods for the in vitro cultivation of microsporidia began as early as 1937, the interest in the culture of these organisms was confined mostly to microsporidia that infect insects. The successful cultivation in 1969 of Encephalitozoon cuniculi, a microsporidium of mammalian origin, and the subsequent identification of these organisms as agents of human disease heightened interest in the cultivation of microsporidia. I describe the methodology as well as the cell lines, the culture media, and culture conditions used in the in vitro culture of microsporidia such as Brachiola (Nosema) algerae, Encephalitozoon cuniculi, E. hellem, E. intestinalis, Enterocytozoon bieneusi, Trachipleistophora hominis, and Vittaforma corneae that cause human disease.
-
Detection by an immunofluorescence test of Encephalitozoon intestinalis spores in routinely formalin-fixed stool samples stored at room temperature.
Journal of clinical microbiology, 1999Co-Authors: Hercules Moura, Gordon J. Leitch, F. C. Sodre, Fernando J. Bornay-llinares, Thomas R. Navin, S P Wahlquist, Ralph T. Bryan, I. Meseguer, Govinda S. VisvesvaraAbstract:Of the several microsporidia that infect humans, Enterocytozoon bieneusi is known to cause a gastrointestinal disease whereas Encephalitozoon intestinalis causes both a disseminated and an intestinal disease. Although several different staining techniques, including the chromotrope technique and its modifications, Uvitex 2B, and the quick-hot Gram-chromotrope procedure, detect microsporidian spores in fecal smears and other clinical samples, they do not identify the species of microsporidia. A need for an easily performed test therefore exists. We reevaluated 120 stool samples that had been found positive for microsporidia previously, using the quick-hot Gram-chromotrope technique, and segregated them into two groups on the basis of spore size. We also screened the smears by immunofluorescence microscopy, using a polyclonal rabbit anti-E. intestinalis serum at a dilution of 1:400. Spores in 29 (24.1%) of the 120 samples fluoresced brightly, indicating that they were E. intestinalis spores. No intense background or cross-reactivity with bacteria, yeasts, or other structures in the stool samples was seen. Additionally, the numbers of spores that fluoresced in seven of these samples were substantially smaller than the numbers of spores that were present in the stained smears, indicating that these samples were probably derived from patients with mixed infections of Enterocytozoon bieneusi and E. intestinalis. Because a 1:400 dilution of this serum does not react with culture-grown Encephalitozoon hellem, Encephalitozoon cuniculi, or Vittaforma corneae or with Enterocytozoon bieneusi spores in feces, we concluded that an immunofluorescence test using this serum is a good alternative for the specific identification of E. intestinalis infections.
