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Protistology 5 (4), 243-255 (2008) Pl'OtiStOlO&y
Overview of microsporidia and microsporidiosis 1
Elizabeth S. Didier 1 and Louis M. Weiss 2
1 Division of Microbiology, Tulane National Primate Research Center, Covington, LA, U.S.A.
2 Departments of Medicine and Pathology, Albert Einstein College of Medicine, Bronx, NY, U.S.A.
Summary
Microsporidia infections occur in virtually all invertebrate and vertebrate hosts, including mammals. In humans these small single-celled eukaryotic organisms have been recognized as emerging and opportunistic pathogens associated with a wide range of clinical syndromes in persons with HIV/AIDS, travelers, children, organ transplant recipients, and the elderly. The most common microsporidia infecting humans are Enterocytozoon bieneusi and members of the Encephalitozoonidae. These infections are often overlooked due to the small size of the infectious agents. Albendazole is effective for treating infections by Encephalitozoon spp., but no effective drug has been identified for treating E. bieneusi infections. Furthermore, it is difficult to study E. bieneusi because tissue culture and small animal models that simulate human infections are lacking. There is still debate about whether microsporidian infections remain persistent in asymptomatic immune-competent individuals, reactivate during conditions of immune-compromise, or are transmitted to other people under risk circumstances such as pregnancy or organ donation. Reliable serological diagnostic methods are needed to supplement PCR or histochemistry when spore shedding may be sporadic. The microsporidia have also generated much interest because of their reduced and compact genomes, and comparative molecular and phylogenetic studies continue to support a relationship between the microspo-ridia and fungi.
Key words: Microsporidia, Enterocytozoon, Encephalitozoon, opportunistic infection, emerging infection, therapeutics, diagnostic testing, genomics, proteomics
Introduction
The phylum Microsporidia comprises nearly 1200 species of single-celled, obligate intracellular, eukaryotic parasites that infect animals of virtually all animal phyla, particularly fish and insects (Canning et al., 1986; Larsson, 2005; Vossbrinck and Debrunner-Vossbrinck, 2005). Since they were discovered to be a cause of persistent diarrhea and systemic disease in persons with AIDS, the interest of the biomedical community in these organisms has grown tremendously (Desportes et al., 1985). Currently, 14
species have been identified as infectious agents of humans (Table 1). Enterocytozoon bieneusi and the Encephalitozoon spp. are the most prevalent microsporidia identified in humans (Weber et al., 2000; Didier, 2005; Didier and Weiss, 2006). Molecular epidemiology studies are generating a broader understanding about the wide demographic, geographic, zoonotic, and environmental range of the microsporidia that infect humans, and the detection of human-infecting microsporidia in water sources led to their inclusion into the NIH Category B list of biodefense pathogens and the EPA microbial con-
1 Materials presented on the V European Congress of Protistology (July 23-27, 2007, St. Petersburg, Russia).
© 2008 by Russia, Protistology
Table 1. Species of microsporidia infecting humans
Microsporidia species Anncaliia (syns. Nosema and Brachiola) algerae Anncaliia (syns. Nosema and Brachiola) connori Anncaliia (syns. Nosema-like and Brachiola) vesicularum Encephalitozoon (syn. Nosema) cuniculi
Encephalitozoon hellem a Encephalitozoon (syn. Septata) intestinalis
Enterocytozoon bieneusi Microsporidium africanum (syn. Nosema sp.) Microsporidium ceylonensis (syn. Nosema sp.)
Nosema ocularum
Pleistophora ronneafiei (syn. Pleistophora sp.) Trachipleistophora anthropopthera Trachipleistophora hominis Vittaforma corneae (syn. Nosema corneum)
Sites of infection
Eye, muscle
Systemic
Muscle
Systemic, eye, respiratory tract, urinary tract, liver, peritoneum, brain
Eye, respiratory tract, urinary tract, systemic Intestine, biliary tract, respiratory tract, bone, skin, systemic
Intestine, biliary tract, respiratory tract
Eye
Eye
Eye
Muscle Systemic, eye Muscle, eye Eye, urinary tract
a - Species that can be grown in long-term culture.
taminant candidates list of concern for waterborne transmission (Didier and Weiss, 2006). Completion of the Encephalitozoon cuniculi genome (Katinka et al., 2001) and the ongoing genome projects on Anncaliia (syn. Brachiola, Nosema) algerae, Spraguea lophii, Antonospora (syn. Nosema) locustae, and E. bieneusi are offering new insights into the genomics, proteomics, and basic biology of the microspo-ridia (Keeling et al., 2005; Texier et al., 2005; Tzipori, 2007). This review summarizes recent research on the microsporidia and microsporidiosis with an emphasis on infections of humans.
The microsporidian spore
General features. Unlike bacterial spores that are generated in response to environmentally stressful conditions, microsporidia spores develop as the mature and infectious stage of the life cycle. Similarly to bacterial spores, microsporidian spores are resistant and survive for long periods of time in the environment (Fayer, 2004). Spores of the microsporidian species that infect mammals are relatively small, measuring 1.0-3.0 ^M by 1.5-4.0 ^M. Spores are surrounded by a glycoprotein outer layer and a chi-tinous inner layer (Vavra et al., 1993; Metenier and Vivares, 2001; Southern et al., 2007). Several proteins have been identified in the spore wall and endospore including SWP1, SWP2, SWP3 (EnP2) and EnP1 (Peuvel-Fanget et al., 2006, Xu et al., 2006). Some of these proteins (i.e. EnP1) may be involved in spore wall adhesion to host cells or mucin, thereby playing a role in the process of invasion (Southern et al.,
2007). The cytoplasm of a microsporidian spore consists of a nucleus in a monokaryon or diplokaryon arrangement, an anterior anchoring disk, a membranous lamellar polaroplast that appears to include an atypical Golgi apparatus, polar vesicles that are likely to be reduced mitochondria called mitosomes, endoplasmic reticulum, ribosomes, and a poster vacuole (Vavra and Larsson, 1999; Vivares et al., 2002; Vavra, 2005; Burri et al., 2006; Beznoussenko et al., 2007).
Polar tube: Microsporidia possess a unique structure, the polar tube, which infects the host cell during germination (Keohane and Weiss, 1999). The coiled polar tube emanates from the anchoring disk and coils numerous times within the posterior region of the spore. A change in osmotic pressure results in swelling of the posterior vacuole and causes the polar tube to evert, followed by transfer of the cytoplasmic contents through the 50-500 ^M-long polar tube into the host cell (Weidner et al., 1994; Frixione et al., 1997) (Figure 1). The mechanism(s) of germination and polar tube formation remain to be determined. In addition, the mechanism by which the sporoplasm penetrates its host cell has not been resolved. At lease five polar tube proteins have been identified as components of the microsporidian polar tube using proteomic approaches (Polonais et al., 2007, Weiss et al., unpublished data). These proteins appear to be partially conserved among the microsporidia species studied to date but share little or no homology with other proteins researched through various databases, suggesting that the polar tube proteins define a novel family of proteins (Polonais et al., 2005). Ptp1, which is the most abundant component of the polar tube, is
Fig. 1. Life cycle of Encephalitozoon and Enterocytozoon species of microsporidia in humans. The majority of infections are believed to occur by ingestion or inhalation of infectious microsporidia spores which are the mature stages of these organisms. Infections are believed to occasionally occur by direct contact or trauma. Vertical transmission in humans has not been reported to occur. Spores are typically shed with the feces, urine, and possibly with respiratory secretions and mucus. This image is reprinted with permission from the DPDx website of the Centers for Disease Control and Prevention (http://www.dpd.cdc.gov/dpdx/).
modified by the addition of O-linked mannose residues, which probably play a role in the adhesion of the polar tube to host cells (Xu et al., 2003; Peek et al.,
2005). Ptpl and ptp2 both contain cysteine residues and appear to interact with each other. Ptp4 localizes to the end of the polar tube and may have a role in adherence to the host cell or in the final process of invasion (Polonais et al., 2005; Polonais et al., 2007). Serum from patients known to have been infected with microsporidia express antibodies which bind to the polar tube, suggesting that ptps may serve as serological diagnostic antigens (van Gool et al., 2004; Peek et al., 2005).
Phylogeny and taxonomy considerations
The taxonomic classification of Microsporidia and the species within this phylum were historically based primarily on morphology, ultrastructure, biology, and habitat features, but more recently molecular phylogenetics have been applied for this classification (Larsson, 2005; Vossbrinck and Debrunner-Vossbrinck, 2005). Early molecular phylogeny studies comparing rDNA sequences suggested that microsporidia were among the earliest or deep-branching eukaryotes because they lacked typical mitochondria, Golgi, and peroxisomes, and they possessed small ribosomes like those of prokaryotes (Vossbrinck et al., 1987). However, mi-crosporidia were found to exhibit many fast-evolving genes and a long-branch attraction artifact of such faster-evolving genes brought into question these early interpretations. The Microsporidia are now considered to be highly-diverged, well-adapted, and specialized parasites that are related or belong to the Fungi or perhaps represent a sister group to the Fungi (Fedorov and Hartman, 2004; Keeling and Slamovits, 2004; Thomarat et al., 2004; Gill and Fast,
2006). In a recent report, eight theorized placements within the Fungi have been rejected based on broad species sampling and comparisons with several genes, which has resulted in the hypothesis that the Microsporidia should be placed either at the base of the fungal tree within the Chytridiomycota or within the Entomophthorales (James et al., 2006).
Microsporidia are peculiar because they contain some of the smallest genomes of eukaryotes due to both gene reduction and compaction (Keeling and Slamovits, 2004; Keeling et al., 2005; Keeling and Slamovits, 2005; Texier et al., 2005). The E. cuniculi genome, for example, consists of 2.9 Mb on 11 chromosomes with approximately 2000 tightly packed genes, which have few introns, are shorter than the corresponding proteins in other eukaryotes and have
overlapping coding regions (Vivares et al., 2002; Keeling et al., 2005; Texier et al., 2005; Williams et al., 2005). The genome sizes ofthe Microsporidia are currently estimated to range from 2.3 Mb on 11 chromosomes for E. intestinalis to 19.5 Mb on 16 chromosomes for Glugea atherinae (Metenier and Vivares, 2001). Microsporidia lack many of the genes encoding proteins in metabolic and regulatory pathways, and retain those related to transport of energy sources and metabolites, presumably as a consequence of host cell dependence (Vivares et al., 2002; Keeling and Slamovits, 2004, 2005; Texier et al., 2005). In addition, sporoblast and spore stages were found to express proteins that are protective from environmental exposure (Brosson et al., 2006). More than a dozen genes encoding mitochondrion-derived proteins have been found and mitochondrial HSP70 has been localized to the mitosome, thus supporting the likelihood that microsporidia evolved from ancestors which contained mitochondria (Vivares et al., 2002; Williams and Keeling, 2003; Thomarat et al., 2004; Gill and Fast, 2006). Genome sequence projects are underway for several microsporidians, including E. bieneusi, Antonospora (syn. Nosema) locustae, and Anncaliia (syn. Brachiola) algerae; continued comparative genomic and proteomic analyses are expected to yield additional information about the phylog-eny and taxonomy of the microsporidia (Keeling et al., 2005; Texier et al., 2005; Tzipori, 2007). A web resource for microsporidia genomics has been developed (http://www.biohealthbase.org/) to assist the research community in the analysis of data generated on these pathogens.
Clinical characteristics
The relevance of microsporidiosis in humans was recognized in the mid 1980's in association with opportunistic enteric infections and persistent diarrhea in persons with HIV/AIDS (Orenstein et al., 1990). Debate followed about whether microsporidia were truly pathogenic, since they were also detected in persons without signs of diarrhea (Rabeneck et al., 1995; Didier, 2000). This was most likely a reflection that the immune status of the host plays a role in the expression of clinical signs during infection (Didier, 2000; Khan and Didier, 2004; Didier and Weiss, 2006). Immune-competent laboratory animals infected with E. cuniculi exhibited clinical signs during the early acute stage of infection that typically resolved even though the infection persisted, whereas immune-deficient athymic and SCID mice infected with E. cuniculi succumbed (Khan and Didier, 2004; Mathis et al., 2005). AIDS patients
with < 100 CD4+ T cells per mm blood, were most likely to experience persistent diarrhea, weight loss, and abdominal pain associated with E. bieneusi or E. intestinalis infections (Kotler and Orenstein, 1998). HIV-infected individuals receiving antiretroviral therapies, or non-HIV-infected individuals who were immunologically naive to microsporidia (i.e. children or travelers) initially developed diarrhea that subsequently resolved (Tumwine et al., 2005; Wichro et al., 2005). In humans, replication of mi-crosporidia (e.g. E. intestinalis, E. bieneusi) occurs in the villus epithelium of the small intestine, resulting in reduced villus height and surface area that appear to contribute to malabsorption and diarrhea (Kotler and Orenstein, 1998; Weber et al., 2000; Morpeth and Thielman, 2006; Wiwanitkit, 2006; Batman et al., 2007). E. bieneusi infections occasionally spread to the hepatobiliary system to cause cholangitis and a few pulmonary infections have been reported (Weber et al., 2000; Sodqi et al., 2004). Encephalitozoon species typically disseminate and infections have been identified in nearly every organ system including a recently described fatal pulmonary infection in a bone marrow transplant recipient (Orenstein, 2003; Orenstein et al., 2005). Interesting reports of human infection, although less common, include a case of Trachipleistophora anthropophthera cornea infection in an AIDS patient (Juarez et al., 2005) and a fatal case of myositis in a woman with rheumatoid arthritis, caused by Anncaliia (syns. Nosema, Brachiola) algerae, a microsporidian that typically infects mosquitoes (Coyle et al., 2004; Visvesvara et al., 2005; Franzen et al., 2006). This A. algerae case raises the possibility that some microsporidia infections in humans may occur through vector borne transmission in addition to the commonly accepted mechanism of water and food borne transmission of these pathogens. Emerging cases of microsporidia infections are being reported among contact lens wearers and cornea transplant recipients (Fogla et al., 2005; Joseph et al., 2005; Kodjikian et al., 2005; Vemuganti et al., 2005; Joseph et al., 2006; Kakrania et al., 2006).
Transplacental transmission of E. cuniculi has been reported in carnivores and laboratory rodents, and was recently considered to be responsible for the deaths of newborn emperor and cotton-top tamarins in Europe and the Americas (Guscetti et al., 2003; Reetz et al., 2004; Juan-Salles et al., 2006). Although transplacental transmission has not yet been reported in humans, it seems plausible that it may occur in them, since there are physiological similarities between humans and nonhuman primates, microsporidia species that infect nonhuman primates also infect humans, and microsporidia are ubiquitous in nature.
There still exist significant clinical questions about the persistence of microsporidian infections developing in humans. Microsporidia infections persist in immune competent mammals such as mice, rabbits, dogs, and rats, but no formal studies have been performed to document whether they also persist in immune competent humans or nonhuman primates. If microsporidia infections do persist in otherwise healthy individuals, it is reasonable to expect that the infections may reactivate and cause clinical signs of disease during conditions of im-mune-compromise (eg. aging, chemotherapy) and that persistently-infected individuals can transmit infections to others at risk. A case report of micro-sporidial keratoconjunctivitis being transmitted by the donor corneal graft supports the latter possibility (Kakrania et al., 2006). Microsporidiosis is also being reported more frequently in solid organ transplant recipients, but it is not clear if these infections are transferred by the donor organ, reactivate from latent infection in the recipient as a consequence of immunosuppressive therapy or are acquired from the environment by the immune compromised recipient (Barsoum, 2006).
Diagnosis
Prior to the widespread application of molecular diagnostics methods, transmission electron microscopy (TEM) was used to definitively confirm a diagnosis of microsporidiosis based on observing a polar filament within spores (Cali et al., 1991; Orenstein,
2003). TEM is still important for describing ultra-structural features that, along with newly-applied molecular biology approaches, contribute to the taxonomic organization of the microsporidia, as evidenced by the recent reclassification of Brachiola species to Anncaliia (Cali et al., 1991; Cali et al., 1993; Larsson, 2005; Franzen et al., 2006). Histochemical methods have also been applied for detecting micro-sporidia more efficiently in fluids (e.g. feces, urine, and mucus) and tissues. These methods included the application of fluorescent brighteners (e.g. Calcofluor White, Uvitex 2B, Fungifluor) that target the chitin-ous spore wall, modified (concentrated) trichrome staining used alone or in combination with Gram stain, and the Warthin-Starry silver stain (Weber et al., 2000; Garcia, 2002). Immunofluorescent antibody staining for species-specific identification has been applied using monoclonal and species-species absorbed polyclonal antibodies (Mo and Drancourt, 2004; Singh et al., 2005). Currently, PCR-based methods are commonly used in research laboratories for detecting microsporidia, but are less often used
in commercial diagnostics labs. PCR methods applied for diagnostics typically utilize primers that target microsporidian rDNA genes (Franzen and Muller, 1999; Weiss and Vossbrinck, 1999). Recently, an oligonucleotide microarrary system was reported for simultaneous detection of four species of human pathogenic microsporidia species in clinical specimens, which should increase diagnostic throughput (Wang et al., 2005).
Microsporidia infections are increasingly reported in relatively immune-competent individuals such as children, travelers, and the elderly, so serological tests are being developed to ascertain whether sub-clinical or asymptomatic infections can be detected. These approaches include using whole organisms or recombinant polar tube or spore wall proteins as antigens, especially in cases where the microsporidian species cannot be grown in culture (van Gool et al., 2004; Peek et al., 2005; Polonais et al., 2005; Xu and Weiss, 2005; Taupin et al., 2006). Interestingly, sera from humans infected with microsporidia bound to the polar tube and specifically to glycoepitopes found on this structure (Peek et al., 2005; Xu and Weiss,
2005).
Serology has not been used routinely for diagnosing microsporidiosis in humans because a correlation between antibody expression and concurrent infection has not yet been proven and variable antibody levels are observed in immune-deficient individuals. Many of the species of microsporidia that infect humans tend to disseminate and infect the kidneys, so examination of urine, in addition to feces, is likely to improve detection of these infections. Since the shedding of microsporidia spores in feces or urine may be intermittent or at levels below detection by histochemistry or PCR, serological approaches may become feasible for diagnosing infections in immune-competent individuals if it can be determined whether seropositivity is indicative of persistent infection.
Epidemiology and sources of infection
Infections due to microsporidia in humans have been reported world-wide and prevalence rates have ranged from 0 to 50% depending on the geographic region, method of diagnosis, and demographic characteristics of the population studied (Didier et al., 2004). Prevalence rates for microsporidiosis were highest among HIV-infected individuals with diarrhea and less than 100 CD4+ T cells per mm blood; the use of antiretroviral therapies has reduced the prevalence of microsporidiosis in persons with HIV/AIDS (Lewthwaite et al., 2005; Morpeth
and Thielman, 2006). In regions of South America, Africa, and Asia, where antiretroviral therapies are not readily accessible, microsporidiosis has been consistently identified in HIV-infected patients with AIDS and other risk factors including poor sanitary conditions and exposure to animals (Dascomb et al., 2000; Mak, 2004; Bern et al., 2005; Chacin-Bonilla et al., 2006; Morpeth and Thielman, 2006; Sarfati et al., 2006; Wiwanitkit, 2006). Since nowadays mi-crosporidia are specifically being looked for more often, they are increasingly recognized in travelers, children, the elderly, and organ transplant recipients (Abreu-Acosta et al., 2005; Leelayoova et al., 2005; Mungthin et al., 2005; Tumwine et al., 2005; Wichro et al., 2005; Barsoum, 2006; Nkinin et al., 2007). Microsporidiosis, however, is still probably overlooked since the causative organisms are quite small and their detection requires considerable expertise by the microscopist using histochemical methods. In the case of PCR, inhibitors commonly confound interpretation of results. In addition, microsporidia are often not included in the routine differential diagnoses for diarrhea, and urine specimens are typically not evaluated for microsporidia as a potential cause of systemic infections. With increasing awareness and sensitivity in the diagnostics methods, a rise in the reported prevalence rates of microsporidi-osis may be anticipated.
It is still unclear how most infections are transmitted to humans, but the genotypes of microsporidia that infect humans have been identified in domestic, farm, wild, and aquatic animals, supporting the likelihood that microsporidiosis is zoonotic (Deplazes et al., 2000; Graczyk et al., 2004; Mathis et al., 2005; Graczyk et al., 2007). Species of microsporidia that infect humans have also been identified in water sources, and risk associations for infection with mi-crosporidia included occupational and recreational contact with water (Didier et al., 2004; Graczyk and Lucy, 2007). These findings contributed to micro-sporidia being included in the NIH Category B biodefense list of pathogens (http://www3.niaid.nih.gov/ biodefense/bandcpriority.htm) and the list of EPA microbial contaminant candidates (http://www.epa. gov/safewater/ccl/ccl2_list.html) of concern for waterborne transmission. There also appears to be an association between microsporidia and food-borne transmission as a consequence of contaminated irrigation water, and organisms have been identified on lettuce, parsley, cilantro, and strawberries in Costa Rica (Calvo et al., 2004). These observations provided the rationale for studies of the transport of microsporidia through sandy porous media for developing mathematical models to assess the potential of mi-
crosporidia contamination of potable water supplies (Brusseau et al., 2005).
Immunology
Resistance to microsporidiosis depends upon functional T lymphocytes, a conclusion based on the greater severity of disease in AIDS patients with declining CD4+ T cell levels and the development of lethal microsporidia infections in experiments on mice depleted of CD4+ and CD8+ T cells (Hermanek et al., 1993; Didier, 2000; Khan and Didier, 2004; Moretto et al., 2004). Proinflammatory responses via Th1 cytokines, such as IFN-y, TNF-a, and IL-12, as well as reactive oxygen and nitrogen intermediates are important for early stages of resistance to Encephalitozoon infections, as shown in experiments using murine models and ex vivo human studies (Khan and Moretto, 1999; Khan and Didier, 2004; Franzen et al., 2005; Moretto et al., 2007). CD8aa+ intraepithelial lymphocytes were observed to increase rapidly after oral administration of E. cuniculi to mice and intestinal dendritic cells were observed to produce IFNy that subsequently led to cytotoxic lymphocyte activity and immune-regulation via IL-10 secretion (Moretto et al., 2004; Moretto et al.,
2007). Antibody responses seem to contribute to prolonging survival in SCID mice given E. cuniculi per os (Sak et al., 2006). Virtually nothing is known about protective immune responses to E. bieneusi infections, in part because a tissue culture system is lacking. SCID mice treated with anti-IFN-gamma that were inoculated with E. bieneusi became transiently infected but these severely immune-deficient animals eventually cleared the microsporidia infections and did not exhibit any signs of disease, unlike immune-deficient humans infected with the same microsporidian (Feng et al., 2006). This would suggest that mice are not natural hosts for E. bieneusi. Naturally occurring and experimental E. bieneusi infections that have been reported in SIV-infected and non-SIV-infected rhesus and pigtail macaques currently represent the only animal models that clinically simulate infections observed in immune-competent and immune-deficient humans (Tzipori et al., 1997; Mansfield et al., 1998; Sestak et al., 2003; Green et al., 2004; Drosten et al., 2005). Studies on immune responses to E. bieneusi, however, have not yet involved nonhuman primate models.
Therapy and disinfection
Antiretroviral therapies that inhibit HIV-infection and thereby partially reconstitute the immune
status have reduced the occurrence of opportunistic infections including microsporidiosis (Didier et al., 2005b; Morpeth and Thielman, 2006; Wiwanitkit,
2006). Furthermore, a recent study indicated that the aspartyl protease inhibitors used in the highly-active antiretroviral therapy cocktail also inhibited the growth of E. intestinalis in tissue culture (Menotti et al., 2005). Albendazole, a benzimidazole that inhibits microtubule assembly, is effective against Encephalitozoon, but not E. bieneusi infections (MacDonald et al., 2004; Tremoulet et al.,
2004). Fumagillin, an antibiotic and antiangiogenic compound produced by Aspergillus fumigatus, was more broadly effective against Encephalitozoon spp. and Enterocytozoon bieneusi, but was toxic when administered systemically (Molina et al., 2002). Current studies are focusing on compounds that target microsporidian polyamines (e.g. polyamine analogues), methionine aminopeptidase type 2 (e.g. fu-magillin-related compounds and analogues), chitin (e.g. nikkomycins), and topoisomerases (e.g. fluoroquinolones) (Bacchi et al., 2001; Didier et al., 2005a; Didier et al., 2005b; Zhang et al., 2005; Didier et al.,
2006). However, since E. bieneusi cannot be grown in long-term tissue culture, these studies have been based on cultivatable species of microsporidia (e.g. Encephalitozoon spp.).
Concerns exist about the potential of water-borne and food-borne transmission of microsporidia. Recent studies demonstrated successful disinfection of E. intestinalis in water by chlorine and ozone and successful disinfection of E. cuniculi in food by high pressure processing. It was also shown that the exposure of E. cuniculi to bleach, ethanol, HiTor, or Roccal was effective in reducing infectivity of these organisms in tissue culture model systems (Becnel et al., 1995; John et al., 2005; Jordan et al., 2005; Jordan et al., 2006).
Conclusion
The recognition ofmicrosporidia as causes ofop-portunistic infections in AIDS patients brought about a greater appreciation of these organisms and their ability to adapt and infect a wide range of animals, including humans. Improved diagnostic methods are likely to reveal a broader range of infections and to assist in establishing molecular epidemiological profiles for defining sources and modes of human infections' transmission. Current studies on comparative genomics and proteomics of the microsporidia are expected to provide new insights into microspo-ridia biology and to promote the development of effective preventive and therapeutic strategies.
Acknowledgements
The authors gratefully acknowledge research funding support from the National Institutes of Health, Bethesda, MD, USA (L.M.W. and E.S.D.) and the Tulane University Research Enhancement Fund, New Orleans, LA, USA (E.S.D.).
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Address for correspondence. L.M. Weiss. Departments of Medicine and Pathology, Albert Einstein College of Medicine, 1300 Morris Park Avenue Room 504 Forchheimer, Bronx, NY 10461, U.S.A. E-mail: lmweiss@aecom.yu.edu
