Disseminated Infection of Encephalitozoon cuniculi Associated With Osteolysis of Hip Periprosthetic Tissue.

Background: Among patients with hip joint endoprosthesis, periprosthetic osteolysis is the most common complication following primary arthroplasty, and subsequent implant loosening is the leading cause of arthroplasty revision. Causes of stability loss, though not always evident, can be mechanical, allergic, or infectious (bacterial and fungal agents) in nature. Microsporidia, widespread opportunistic fungal pathogens that infect most human tissues, are a potential infectious cause of stability loss. Infections caused by Encephalitozoon species-one of the most common microsporidial pathogens in humans-primarily localize to intestinal and respiratory tracts, but can disseminate to tissues throughout the body. Methods: We examined 53 immunocompetent patients, 23 after revision and 30 after primary hip arthroplasty, for infection by Encephalitozoon species. Periprosthetic tissue, urine sediments, and stool samples were tested by microscopic examination and genus-specific nested polymerase chain reaction followed by genotyping. Results: Ten patients had Encephalitozoon-positive periprosthetic tissues, 9 (39%) after revision and 1 (3.3%) after primary hip arthroplasty. Among the tissue-positive postrevision patients, 7 had a positive urine sample and 1 had a positive stool sample. Encephalitozoon cuniculi genotype II was identified in 88.8% (16/18) of samples. Two urine samples were positive for a novel Encephalitozoon species. Conclusions: Encephalitozoon cuniculi should be considered as a cause of osteolysis in hip periprosthetic tissue, leading to a loss of implant stability.

[Prevalence of Encephalitozoon intestinalis and Enterocytozoon bieneusi in cancer patients under chemotherapy]. / Kemoterapi alan kanserli hastalarda Encephalitozoon intestinalis ve Enterocytozoon bieneusi prevalansi

Microsporidia species are obligate intracellular parasites and constitute one of the most important opportunistic pathogens that can cause severe infections especially in immunocompromised patients. Enterocytozoon bieneusi and Encephalitozoon intestinalis are the most common species among 14 microsporidia species identified as human pathogens. The aim of this study was to investigate the prevalence of E.intestinalis and E.bieneusi in cancer patients under chemotherapy by immunofluorescent antibody and conventional staining methods. A total of 123 stool samples obtained from 93 patients (58 male, 35 female) with cancer who were followed in oncology and hematology clinics of our hospital and 30 healthy volunteers (13 male, 17 female) were included in the study. Fifty-one (55%) of the patients had complain of diarrhea. The presence of E.intestinalis and E.bieneusi were investigated by a commercial immunofluorescence antibody test using monoclonal antibodies (IFA-MAbs; Bordier Affinity Products, Switzerland) in all of the samples, and 50 of the samples were also investigated by modified trichrome, acid-fast trichrome and calcofluor staining methods. A total of 65 (69.9%) patients were found positive with IFA-MAbs method, including 43 (46.2%) E.intestinalis, 9 (9.7%) E.bieneusi and 13 (14%) mixed infections. In the control group, 5 (16.7%) subjects were positive with IFA-MAbs method, including 2 (6.7%) E.intestinalis, 1 (3.3%) E.bieneusi and 2 (6.7%) mixed infections. The difference between the positivity rate of the patient and control groups was statistically significant (p< 0.05). Of the patients with diarrhea, 68.6% (35/51) were infected with microsporidia, and the difference between cases with and without (48.6%) diarrhea was statistically significant (p< 0.05). When 50 samples in which all of the methods could be performed were evaluated, the frequency of microsporidia were detected as follows; 66% (n= 33) with IFA-MAbs, 34% (n= 17) with modified trichrome staining, 24% (n= 12) with acid-fast trichrome staining and 42% (n= 21) with calcofluor staining methods. Our data indicated that the use of IFA-MAbs method along with the conventional staining methods in diagnosis of microsporidia will increase the sensitivity. As a conclusion, the prevalence of E.intestinalis and E.bieneusi in cancer patients under chemotherapy was detected quite high (69.9%) in our study, it would be appropriate to screen these patients regularly in terms of microsporidian pathogens.

Disseminated microsporidiosis in a renal transplant recipient: case report and review of the literature.

Microsporidia are opportunistic pathogens that usually cause a limited disease in the gastrointestinal tract. Occasionally, they can cause disseminated disease. In solid organ transplant recipients, disseminated disease has been reported only rarely. We describe a 68-year-old woman who presented with fever, cough, and acute kidney injury 6 months after kidney transplantation. Dissemination was confirmed by identification of microsporidial spores in urine and bronchoalveolar lavage fluid. Polymerase chain reaction analysis identified the species as Encephalitozoon cuniculi.

Detection of Encephalitozoon cuniculi in the feline cataractous lens.

PURPOSE: Identification of Encephalitozoon cuniculi (E. cuniculi) as a possible causative agent for cataracts and uveitis in cats. METHODS: Within a 12-month study period, cats that were presented with focal anterior cortical or mature cataract and secondary uveitis underwent a complete ophthalmic examination, complete blood count, serum biochemistry, serologic tests for E. cuniculi and tests for feline immunodeficiency virus (FIV), feline infectious peritonitis (FIP), feline leukemia virus (FeLV) and Toxoplasma gondii (T. gondii). PCR for DNA detection of E. cuniculi and T. gondii as well as cytologic examination of aqueous humor after paracentesis and phacoemulsified lens material were also performed. In addition histopathologic examination of the resected anterior lens capsule and attached lens epithelial cells was performed. Serologic testing for antibodies against E. cuniculi was also performed in 100 ophthalmologically healthy cats. RESULTS: Eleven (19 eyes) European shorthair cats with a median age of 3.5 years were included. Nine of 11 cats had bilateral cataracts, with 12/19 eyes having focal anterior cortical cataracts and 7/19 eyes having mature cataracts. In 14/19 eyes anterior uveitis was present. All cats had a positive antibody titer (1:80-1:10,000) for E. cuniculi. Encephalitozoon cuniculi DNA was detected by PCR and sequencing in 18/19 lenses and in 10/19 aqueous samples. Five tentative positive results were detected by cytologic examination. Spores were detected in 15/19 samples of lens material with histopathologic staining. Only 2/100 ophthalmologically healthy cats showed a positive antibody titer for E. cuniculi. CONCLUSION: Encephalitozoon cuniculi is a cause of focal anterior cortical cataract and anterior uveitis in cats.

[Systemic microsporidiosis and toxoplasmosis in a patient with T prolymphocytic leukemia]. / Microsporidiose et toxoplasmose disséminées chez une patiente présentant une leucémie prolymphocytaire T.

We report a case of microsporidiosis in a 72-year-old woman presenting with prolymphocytic leukemia. The underlying conditions 7 months after leukemia was diagnosed were pancytopenia and immunosuppression due to alemtuzumab and pentostatin. The patient's status had worsened and she presented with dysuria. Urine cultures for bacteria were repeatedly negative. She was first empirically treated with broad-spectrum antibiotics. Three months later, urinary symptoms were persisting. Her blood lymphocyte count was 90/microl. Urine examination was positive for microsporidia using modified trichrome staining and Uvitex 2B fluorescence. Microsporidia were also detected in stools. The patient was cured by albendazole. This was consistent with an infection due to Encephalitozoon sp. Concurrently, disseminated toxoplasmosis was diagnosed. Toxoplasma gondii was detected in bone marrow, broncho-alveolar lavage and cerebrospinal fluid. She was successfully treated with sulfadiazine-pyrimethamine. Four cases of microsporidiosis in myeloid leukemic patients have been already described. The present case in a patient with lymphoid leukemia is the first to be reported.

Dual infection of urinary tract with Enterocytozoon bieneusi and Encephalitozoon cuniculi in HIV/AIDS patients

Microsporidia are emerging pathogens which cause an opportunistic infections in immunocompromised patients, especially those with AIDS. Intestinal microsporidiosis is the most recognized infection, whereas urinary tract infections caused by microsporidia are rarely paid attention to either due to their subclinical course or diagnostic difficulties. In this report dual microsporidial infection of urinary tract, caused by Enterocytozoon bieneusi and Encephalitozoon cuniculi was described in HIV/AIDS patients under cART therapy. Since microsporidiosis can cause severe complications or even death in immunosuppressed patients, our results suggest that microsporidial infection should be included in routine investigation of HIV-positive patients, even asymptomatic.

Radiographic signs of renal disease in rabbits.

The radiological features of 65 rabbits with suspected renal disease are reviewed. The radiological features included a generalised increase in bone opacity (osteosclerosis), renomegaly, nephroliths, ureteroliths and soft tissue mineralisation. One or more of these changes were present on radiographs of 57 of the 65 rabbits. Renal disease was suspected because of the clinical signs and the presence of kidney stones and/or high blood concentrations of urea and creatinine. Significant renal disease was confirmed in 14 cases that were examined postmortem. Blood urea and creatinine concentrations were measured in 47 cases but not all the rabbits had high levels of both. Blood calcium concentration was high in 33 of the 38 rabbits in which it was measured. Serum phosphate was high in 17 and low in five of 34 rabbits in which it was measured. Hyperphosphataemia was associated with generalised osteosclerosis and aortic calcification. Rabbits with osteosclerosis were thin, depressed and unwilling to move. Thirty-eight of 41 rabbits that were tested were seropositive for antibodies to Encephalitozoon cuniculi. Histological lesions suggestive of E cuniculi infection were found in all 13 cases that were examined postmortem, although the organisms were visible in only one case.

A case of equine abortion caused by Encephalitozoon sp.

A Lippizan mare aborted a male fetus a few days before the expected foaling date without showing any clinical sings. Focal lympho-histiocytic hepatitis in the foal and multiplex focal lympho-histiocytic villitis accompanied by villus necroses and marked hypertrophy of chorionic epithelial cells in the arcades were observed. Elongated nucleated organisms were seen in groups in vacuoles or solitarily located in the cytoplasm of the chorionic epithelial cells. The organisms were in large numbers and often extracellularly in areas of villitis and villus necroses. They were Gram-positive, stained with haematoxylin and eosin (HE), periodic acid-Schiff (PAS) and Giemsa, weakly with Warthin-Starry silver stain but not with Gömöri's methenamine-silver stain. By ultrastructural and immunohistochemical examinations, the organisms were identified as microsporidia belonging to the genus Encephalitozoon. No Encephalitozoon organisms were detected in the fetal organs. This is the first reported case of equine abortion induced by Encephalitozoon sp. in Europe. Although abortion induced by Encephalitozoon is rare, microsporidia should be considered a differential diagnosis for intracellular organisms observed in the chorionic epithelial cells of horses.

Diabetes mellitus increases the susceptibility to encephalitozoonosis in mice.

Microsporidiosis are diseases caused by opportunistic intracellular fungi in immunosuppressed individuals, as well as in transplanted patients, the elderly and children, among others. Diabetes mellitus (DM) is a metabolic disease characterized by hyperglycemia and decreased T cell response, neutrophil function, humoral immunity failure, increasing the susceptibility to infections. Here, we investigated the susceptibility of streptozotocin (STZ)-induced type I diabetic and/or immunosuppressed mice to encephalitozoonosis by Encephalitozoon cuniculi. Microscopically, granulomatous hepatitis, interstitial pneumonia and pielonephritis were observed in all infected groups. STZ treatment induced an immunossupressor effect in the populations of B (B-1 and B2) and CD4+ T lymphocytes. Moreover, infection decreased CD4+ and CD8+ T lymphocytes and macrophages of DM mice. Furthermore, infection induced a significant increase of IL-6 and TNF-α cytokine serum levels in DM mice. IFN-γ, the most important cytokine for the resolution of encephalitozoonosis, increased only in infected mice. In addition to the decreased immune response, DM mice were more susceptible to encephalitozoonosis, associated with increased fungal burden, and symptoms. Additionally, cyclophosphamide immunosuppression in DM mice further increased the susceptibility to encephalitozoonosis. Thus, microsporidiosis should be considered in the differential diagnosis of comorbidities in diabetics.

Reversible renal failure caused by a microsporidian infection.

OBJECTIVE: To report a case of renal failure associated with microsporidian infection in an HIV-seropositive patient. DESIGN: Case report. SETTING: Chelsea and Westminster Hospital, London, England, UK. PATIENT: An HIV-seropositive patient presented febrile with abdominal pain who developed renal failure. Renal biopsy and urinalysis showed infection with a microsporidian of the genus Encephalitozoon. INTERVENTION: Treatment with albendazole (400 mg) twice daily was associated with disappearance of infection from the urine, clinical improvement and return of renal function virtually to normal. CONCLUSION: HIV-seropositive individuals with renal failure should have urine screened for microsporidia. The administration of albendazole in such cases may reverse renal failure.

Identification of a human isolate of Encephalitozoon cuniculi type I from Italy.

A microsporidial strain, obtained from a person with AIDS living in Italy was isolated and cultivated on RK13 (rabbit kidney) cell monolayers. Identification at the species level was performed by immunological and molecular methods. Western blot analysis showed that the human isolate and the Encephalitozoon cuniculi reference strain had similar banding patterns. The small subunit rRNA sequence analysis confirmed the identification of the isolate as E. cuniculi, which is a widespread microsporidian species infecting a wide range of natural hosts, including humans. Moreover, based on the sequence of the rDNA internal transcribed spacer region, this isolate was classified as E. cuniculi type I (rabbit strain), previously reported in six persons with AIDS living in Switzerland. These results provide further information on the geographical distribution of E. cuniculi types.

Microsporidia: emerging advances in understanding the basic biology of these unique organisms.

Microsporidia are long-known parasites of a wide variety of invertebrate and vertebrate hosts. The emergence of these obligate intracellular organisms as important opportunistic pathogens during the AIDS pandemic and the discovery of new species in humans renewed interest in this unique group of organisms. This review summarises recent advances in the field of molecular biology of microsporidia which (i) contributed to the understanding of the natural origin of human-infecting microsporidia, (ii) revealed unique genetic features of their dramatically reduced genome and (iii) resulted in the correction of their phylogenetic placement among eukaryotes from primitive protozoans to highly evolved organisms related to fungi. Microsporidia might serve as new intracellular model organisms in the future given that gene transfer systems will be developed.

Isolation and identification of Encephalitozoon hellem from an Italian AIDS patient with disseminated microsporidiosis.

Microsporidia are primitive mitochondria-lacking spore-forming eukaryotic protozoa that infect a wide variety of animals and also humans. Of the five genera (Encephalitozoon, Enterocytozoon, Septata, Nosema and Pleistophora) that cause infections in humans, Enterocytozoon bieneusi, Septata intestinalis, and Encephalitozoon hellem are being increasingly identified in patients with acquired immunodeficiency syndrome (AIDS). E. bieneusi causes gastrointestinal disease, S. intestinalis causes gastrointestinal and disseminated disease, and E. hellem causes ocular as well as disseminated disease. We have established in continuous culture a strain of microsporidia isolated from the urine and throat washings of an Italian AIDS patient and identified it as Encephalitozoon hellem, based on its ultrastructural morphology, antigenic pattern, and polymerase chain reaction-amplified small subunit ribosomal RNA. We believe that this is the first time that a strain of microsporidia has been isolated from the throat washings of a patient with microsporidiosis.

Renal Encephalitozoon (Septata) intestinalis infection in a patient with AIDS. Post-mortem identification by means of transmission electron microscopy and PCR.

We describe the occurrence of renal Encephalitozoon (Septata) intestinalis infection in a 35-year-old AIDS patient who died with disseminated tuberculosis. The patient did not complain of specific symptoms involving the kidney or lower urinary tract during life, but at autopsy, light microscopic examination of the kidney revealed numerous small round or oval bodies in the tubules and tubular cell cytoplasm that were interpreted as intracellular protozoa. Transmission electron microscopy of tissue retrieved from paraffin-embedded samples identified these organisms as microsporidia belonging to the Encephalitozoonidae family, but did not allow definitive identification of the species of infecting parasite. This was made possible only by means of Southern blot hybridization after the polymerase chain reaction, which recognized the micro-organism as E. intestinalis.

Microsporidial keratoconjunctivitis in a patient with AIDS.

A male patient is described with acquired immune deficiency syndrome (AIDS) who developed chronic keratoconjunctivitis and chronic sinusitis due to infection with the microsporidian Encephalitozoon cuniculi. Diagnosis was confirmed by electron microscopic examination of conjunctival epithelial cells and nasal polypectomy specimens. Treatment with propamidine isethionate 0.1% (Brolene) eye drops six times daily led to a prompt resolution of the keratoconjunctivitis.

Autopsy verification of Encephalitozoon intestinalis (microsporidiosis) eradication following albendazole therapy.

Microsporidian infections are increasingly recognized as an important cause of morbidity for persons infected with the human immunodeficiency virus. Encephalitozoon (formerly Septata) intestinalis is a recently described microsporidian that causes intestinal and disseminated infections in severely immunocompromised patients with acquired immunodeficiency syndrome. Several studies suggest that albendazole is an effective therapy for E intestinalis infection. However, relapses of symptoms and reappearance of microsporidian spores in diagnostic specimens have been reported following treatment in some cases. Because these results are based on examination of feces or cytologic specimens with an inherent sampling bias, it would be ideal to have autopsy data on the complete tissue evaluation of major organ systems of patients who had antemortem E intestinalis infection treated with albendazole. This report describes an acquired immunodeficiency syndrome patient with diarrhea and wasting syndrome associated with E intestinalis infection. Treatment with albendazole produced relief of his clinical symptoms and eliminated microsporidian spores in his feces. Following his death from other causes, an autopsy was performed. Comprehensive microscopic examination of all major organs revealed no evidence of residual microsporidian infection, suggesting parasitologic cure of E intestinalis with albendazole. The postmortem finding of complete clearance of microsporidia from body tissues is significant for future albendazole treatment of patients infected with E intestinalis and provides strong support for the value of the autopsy in evaluating the therapeutic efficacy of antimicrobials in emerging infections.

Encephalitozoon cuniculi placentitis and abortion in a quarterhorse mare.

Encephalitozoon cuniculi is a microsporidial parasite, which has rarely been reported to cause placentitis in animals. A late-term aborted fetus and placenta from a Quarterhorse were presented to the Livestock Disease Diagnostic Center, University of Kentucky, for diagnostic examination. There was a necrotizing placentitis, with distension of many chorionic epithelial cells by intracytoplasmic vacuoles containing 1-2-microm-diameter, elongated, gram-positive organisms. The organisms were identified as E. cuniculi by electron microscopy and by polymerase chain reaction using primers to microsporidial ribosomal DNA. Joints of the fetus were swollen, with gross and microscopic lesions of synovitis; however, E. cuniculi DNA was not detected.

Western blot and immunofluorescence analysis of a human isolate of Encephalitozoon cuniculi established in culture from the urine of a patient with AIDS.

Microsporidia spores, identified as Encephalitozoon cuniculi (CDC: V282), were isolated from the urine of a patient with acquired immunodeficiency syndrome and disseminated microsporidiosis, established in continuous culture on monkey kidney cells (E6), and antiserum was produced in rabbits. Immunoblot studies that used the patient serum and the rabbit sera against CDC:V282, Encephalitozoon hellem (CDC:0291:V213), and Encephalitozoon intestinalis (CDC:V297) revealed that CDC:V282 and the rabbit isolate of E. cuniculi (ECLD) reacted intensely with the patient's serum and the rabbit anti-CDC:V282, producing a number of bands ranging from 200 to 15 kDa. By contrast, the heterologous antigens (CDC:0291:V213 and CDCV297) reacted minimally. Both CDC:V282 and ECLD isolates of E. cuniculi reacted minimally with the rabbit anti-E. hellem and the rabbit anti-E. intestinalis sera. In the immunofluorescence test, performed on the lung biopsy section of the patient, the rabbit anti-CDC:V282 serum reacted extensively with the spores in the tissue section and produced bright apple green fluorescence. These studies demonstrated that the human (CDC:282) and the rabbit (ECLD) isolates of E. cuniculi were similar in their antigenic profiles but differed considerably from E. hellem and E. intestinalis, and that the patient's serum reacted specifically, strongly, and with equal intensity, with the 2 isolates of E. cuniculi.

Bovine leukemia virus-induced clinical signs and morphological changes of encephalitozoonosis in rabbits.

Fourteen three-month-old rabbits spontaneously-infected with the microsporidium Encephalitozoon cuniculi Levaditi, Nicolau et Schoen, 1923 were inoculated intravenously with lymphocytes (Ly) from seropositive bovine leukemia virus infected cattle (Ly/BLV) or with fetal lamb kidney cells infected with bovine fetal leukemia (FLK/BLV). Thirteen rabbits were seropositive to BLV at least for a period of three months. Six rabbits died of pulmonary lesions. Chronic inflammatory lesions of encephalitozoonosis were found in six rabbits killed between 454 and 548 days of the observation period. Five animals bore subcutaneous granulomas. Immunohistochemically, E. cuniculi was demonstrated in the inflammatory lesions of rabbits studied. Control animals also spontaneously infected with E. cuniculi did not show clinical signs of encephalitozoonosis. Morphological changes were found incidentally in the form of small glial foci and focal interstitial nephritis in these animals. The combined action of BLV-E. cuniculi on the bodies of rabbits is proposed as a suitable model for the study of encephalitozoonosis in man with human immunodeficiency virus (HIV) infection.