Hellem Infection Research Articles

E. hellem Ser/Thr protein phosphatase PP1 targets the DC MAPK pathway and impairs immune functions.

Microsporidia are difficult to be completely eliminated once infected, and the persistence disrupts host cell functions. Here in this study, we aimed to elucidate the impairing effects and consequences of microsporidia on host DCs. Enterocytozoon hellem, one of the most commonly diagnosed zoonotic microsporidia species, was applied. In vivo models demonstrated that E. hellem-infected mice were more susceptible to further pathogenic challenges, and DCs were identified as the most affected groups of cells. In vitro assays revealed that E. hellem infection impaired DCs' immune functions, reflected by down-regulated cytokine expressions, lower extent of maturation, phagocytosis ability, and antigen presentations. E. hellem infection also detained DCs' potencies to prime and stimulate T cells; therefore, host immunities were disrupted. We found that E. hellem Ser/Thr protein phosphatase PP1 directly interacts with host p38α (MAPK14) to manipulate the p38α(MAPK14)/NFAT5 axis of the MAPK pathway. Our study is the first to elucidate the molecular mechanisms of the impairing effects of microsporidia on host DCs' immune functions. The emergence of microsporidiosis may be of great threat to public health.

Encephalitozoon hellem Infection Promotes Monocytes Extravasation.

Background: Microsporidia are a group of obligated intracellular fungus pathogens. Monocytes and the derivative macrophages are among the most important players in host immunity. The invasion of microsporidia may significantly affect the monocytes maturation and extravasation processes. Methods: We utilized a previously established microsporidia infection murine model to investigate the influences of microsporidia Encephalitozoon hellem (E. hellem) infection on monocyte maturation, releasing into the circulation and extravasation to the inflammation site. Flow cytometry and qPCR analysis were used to compare the monocytes and derivative macrophages isolated from bone marrow, peripheral blood and tissues of E. hellem-infected and control mice. Results: The results showed that the pro-inflammatory group of CD11b+Ly-6C+ monocytes are promoted in E. hellem-infected mice. Interestingly, the percentage of Ly-6C+ monocytes from E. hellem-infected mice are significantly lower in peripheral blood while significantly higher in the inflamed small intestine, together with up-regulated ratio of F4/80 macrophage in small intestine as well. Conclusions: Our findings demonstrated that E. hellem infection leads to promoted monocytes maturation in bone marrow, up-regulation of extravasation from peripheral blood to inflammation site and maturation into macrophages. Our study is the first systematic analysis of monocytes maturation and trafficking during microsporidia infection, and will provide better understanding of the pathogen–host interactions.

Molecular Identification of Encephalitozoon hellem from Companion Birds Kept in Pet Shops, Japan.

To evaluate the role of companion birds as a reservoir of Encephalitozoon hellem infection in humans, the present study determined the prevalence and genotypes of E. hellem from 269 birds in 4 pet shops using polymerase chain reaction (PCR) assay. E. hellem was identified in 4.8% (13/269) of the birds and was detected in all pet shops. Every positive sample corresponded to zoonotic genotype 1A. Considering the low prevalence of E. hellem infection, it is likely that the risk of zoonotic transmission from companion birds kept in pet shops to humans is low in Japan.

Characterization of a Murine Model for Encephalitozoon hellem Infection after Dexamethasone Immunosuppression.

Background: Encephalitozoon hellem (E. hellem) belongs to a group of opportunistic pathogens called microsporidia. Microsporidia infection symptoms vary and include diarrhea, ocular disorders and systemic inflammations. Traditionally, immunodeficient animals were used to study microsporidia infection. To overcome the difficulties in maintenance and operation using immunodeficient mice, and to better mimic natural occurring microsporidia infection, this study aims to develop a pharmacologically immunosuppressed murine model of E. hellem infection. Methods: Wild-type C57BL/6 mice were immunosuppressed with dexamethasone (Dex) and then E. hellem spores were inoculated into the mice intraperitoneally. Control groups were the Dex-immunosuppressed but noninoculated mice, and the Dex-immunosuppressed then lipopolysaccharide (LPS)-treated mice. Mice body weights were monitored and all animals were sacrificed at the 15th day after inoculation. Tissue fragments and immune cells were collected and processed. Results: Histopathological analysis demonstrated that E. hellem inoculation resulted in a disseminated nonlethal infection. Interestingly, E. hellem infection desensitized the innate immunity of the host, as shown by cytokine expressions and dendritic cell maturation. We also found that E. hellem infection greatly altered the composition of host gut microbiota. Conclusions: Dex-immunosuppressed mice provide a useful tool for study microsporidiosis and the interactions between microsporidia and host immunity.

Encephalitozoon hellem infection in aviary passerine and psittacine birds in Spain

A European goldfinch (Carduelis carduelis), a canary (Serinus canaria), and a lovebird (Agapornis roseicollis) captive-bred at three different private aviaries in Spain were submitted for necropsy with a history of weakness and ruffled feathers, weight loss associated with glossitis, and respiratory disease, respectively. Microscopically, enterocytes in the jejunum and ileum contained colonies of gram- and Stamp-positive, oval to elliptical microorganisms within parasitophorous vacuoles in the apical cytoplasm. Nested PCR using MSP primers that target microsporidian RNA genes produced amplicons of expected size for Encephalitozoon species, and analysis of forward and reverse DNA sequences confirmed the presence of Encephalitozoon hellem in all cases. The main cause of death of all three birds consisted of concurrent infections. However, intestinal encephalitozoonosis may have contributed to exacerbated catabolism. Encephalitozoonosis (or microsporidiosis) has been rarely described in passerine birds.

Encephalitozoon hellem Infection in a Captive Juvenile Freshwater Crocodile (Crocodylus johnstoni)

Microsporidiosis is reported rarely in reptiles and has never been reported in any species of crocodilian. Microsporidiosis was diagnosed histologically in a juvenile captive freshwater crocodile (Crocodylus johnstoni) that was found suddenly dead in its enclosure. Ultrastructural and molecular testing revealed infection to be due to Encephalitozoon hellem. This is the first report of E. hellem infection in any species of reptile.

Detection of Encephalitozoon spp. from human diarrheal stool and farm soil samples in Korea.

Microsporidia are eukaryotic organisms that cause zoonosis and are major opportunistic pathogens in HIV-positive patients. However, there is increasing evidence that these organisms can also cause gastrointestinal and ocular infections in immunocompetent individuals. In Korea, there have been no reports on human infections with microsporidia to date. In the present study, we used real-time PCR and nucleotide sequencing to detect Encephalitozoon intestinalis infection in seven of 139 human diarrheal stool specimens (5%) and Encephalitozoon hellem in three of 34 farm soil samples (8.8%). Genotype analysis of the E. hellem isolates based on the internal transcribed spacer 1 and polar tube protein genes showed that all isolates were genotype 1B. To our knowledge, this is the first report on human E. intestinalis infection in Korea and the first report revealing farm soil samples as a source of E. hellem infection. Because microsporidia are an important public health issue, further large-scale epidemiological studies are warranted.

Ceratoconjuntivite por Encephalitozoon hellem em periquitos agapornis (Agapornis spp.) no Brasil: relato de caso

Relata-se um caso de ceratoconjuntivite causada por Encephalitozoon hellem em agapornis (Agapornis spp.) adultos, provenientes de um criatório comercial. Cinco animais apresentaram sinais clínicos de ceratoconjuntivite, blefaroespasmo e blefaroedema bilateral, com presença de secreção seropurulenta. Amostras fecais foram colhidas e foi realizado exame coproparasitológico, com resultado negativo. Dois animais foram necropsiados, sendo detectados, em impressões de raspado de conjuntiva ocular, esporos e outros estádios evolutivos de Microsporidium. A confirmação do diagnóstico foi feita pela reação em cadeia de polimerase e sequenciamento de fragmentos amplificados, com utilização de primers específicos para o gene da subunidade 18S do rRNA de E. hellem. A análise dos fragmentos amplificados demonstrou 100% de similaridade com outras sequências de E. hellem publicadas no GenBank. Este é primeiro relato de infecção por E. hellem em aves no Brasil.

Humoral response of chicken infected with the microsporidium Encephalitozoon hellem

Chicken (Gallus gallus) were used as the experimental model for study of immune response against the microsporidium Encephalitozoon hellem (Didier et al., J Inf Dis 163:617-621, 1991) infection in birds. Two-day-old chicken were infected perorally or intraperitoneally with a dose of 10(7) spores of E. hellem. The anti-E. hellem immunoglobulin (Ig)A, IgY, and IgM antibody responses in sera and dropping sample extracts were determined by enzyme-linked immunosorbent assay. Results have shown specific antibody production in sera and intestinal secretions of infected birds. Chicken inoculated perorally developed the lowest antibody response. Microsporidian spores were not identified in the smears from cloacal swab samples of individual chicken. Intestinal segment cultures of perorally infected chicken cultivated in vitro showed the highest production of specific IgY and IgA antibodies in jejunum segments. In the further course of infection, the colon produced the highest amount of IgA, and the ileum and colon produced the highest amount of IgY.

Encephalitozoon helleminfection as the cause of a unilateral chronic keratoconjunctivitis in an umbrella cockatoo (Cacatua alba)

An umbrella cockatoo (Cacatua alba) was presented with a keratoconjunctivitis of 9 months' duration that was refractory to antibacterial and antifungal treatments. Microsporidian-type spores were observed in calcofluor M2R-stained cytological preparations of debrided corneal epithelium. Hematoxylin and eosin-stained sections of a conjunctival biopsy also showed a small number of intracellular parasites. Microsporidial DNA was detected by polymerase chain reaction in DNA extracted from the remaining biopsy material. Sequence analysis of the small subunit ribosomal RNA gene identified the microsporidium as Encephalitozoon hellem. The cockatoo was treated with oral albendazole for 3 months with resolution of the signs, and then again for 4 months when the signs returned. Since then, the bird has remained healthy for 2 years. This is the second report of a microsporidial keratoconjunctivitis in a bird, and the first report of an E. hellem infection in a cockatoo. Ophthalmologists should consider E. hellem as a possible cause of a keratoconjunctivitis in parrots that is refractory to treatment with traditional antimicrobials.

Glycosylation of the Major Polar Tube Protein ofEncephalitozoon hellem, a Microsporidian Parasite That Infects Humans

The microsporidia are ubiquitous, obligate intracellular eukaryotic spore-forming parasites infecting a wide range of invertebrates and vertebrates, including humans. The defining structure of microsporidia is the polar tube, which forms a hollow tube through which the sporoplasm is transferred to the host cell. Research on the molecular and cellular biology of the polar tube has resulted in the identification of three polar tube proteins: PTP1, PTP2, and PTP3. The major polar tube protein, PTP1, accounts for at least 70% of the mass of the polar tube. In the present study, PTP1 was found to be posttranslationally modified. Concanavalin A (ConA) bound to PTP1 and to the polar tube of several different microsporidia species. Analysis of the glycosylation of Encephalitozoon hellem PTP1 suggested that it is modified by O-linked mannosylation, and ConA binds to these O-linked mannose residues. Mannose pretreatment of RK13 host cells decreased their infection by E. hellem, consistent with an interaction between the mannosylation of PTP1 and some unknown host cell mannose-binding molecule. A CHO cell line (Lec1) that is unable to synthesize complex-type N-linked oligosaccharides had an increased susceptibility to E. hellem infection compared to wild-type CHO cells. These data suggest that the O-mannosylation of PTP1 may have functional significance for the ability of microsporidia to invade their host cells.

Pathologic Features and Immunofluorescent Antibody Demonstration of Ocular Microsporidiosis (Encephalitozoon hellem) in Seven Patients with Acquired Immunodeficiency Syndrome

We studied the clinicopathologic features of seven patients with acquired immunodeficiency syndrome (AIDS) and ocular microsporidiosis. All patients had decreased levels of CD4-positive cells (mean, 26/ml3) and ocular symptoms; five had bilateral punctate epithelial keratopathy, one had intermittent red eyes with conjunctivitis, and one had red eyes only. Light and electron microscopy of corneal and conjunctival biopsy and cytologic specimens and intact globes disclosed microsporidia belonging to the genus Encephalitozoon. Because E. cuniculi and E. hellem, the two species of the Encephalitozoon genus, are morphologically identical, an immunofluorescent antibody technique was used for species identification. In all seven patients, the agent was identified as E. hellem. Pathologic examination of globes obtained after autopsy disclosed E. hellem infection to be restricted to the corneal and conjunctival epithelium. We studied methods for the routine diagnosis of ocular microsporidiosis in patients with AIDS, including the role of immunofluorescent antibody staining.