[Ophthalmic findings caused by Encephalitozoon cuniculi (strain III) in a dog]. / Ophthalmologische Befunde durch Enzephalitozoon cuniculi (Stamm III) bei einem Hund.

A 2-year-old female mixed-breed canine patient from Namibia presented originally with chronic uveitis. A serum antibody titer and a PCR test performed on the aqueous humor were positive for encephalitozoon cuniculi. The left eye showed an immature anterior focal cortical cataract in the periphery with suspected lens capsule rupture and signs of chronic uveitis. An incipient anterior focal cortical cataract was also perceivable in the patient's right eye. Despite local treatment as well as systemic administration of carprofen, prednisolone, and fenbendazol recurrent uveitis occurred. The patient then underwent bilateral extracapsular lensextraction via phacoemulsification. A PCR test of the lens material was positive for encephalitozoon cuniculi strain III. Recurring uveitis and secondary glaucoma 10 months post-op resulted in permanent blindness of the left eye. The patient then continued to receive local anti-inflammatory treatment. The last recheck examination of both eyes, 31 month post-op, revealed no signs of uveitis. This is the first case reported of a cataract in a canine patient caused by encephalitozoon cuniculi strain III.

A massive systematic infection of Encephalitozoon cuniculi genotype III in mice does not cause clinical signs.

Encephalitozoon cuniculi genotype III disseminated intensively into most of the organs in all strains of mice, followed by a chronic infection with massive microsporidia persistence in immunodeficient mice and a partial decrease in C57Bl/6 mice. Treatment with 0.2 mg Albendazole/mouse/day temporarily reduces the number of affected organs in immunocompetent C57Bl/6 mice, but not in CD4-/- and CD8-/- mice. The application of medication temporarily decreased the spore burden at least by one order of magnitude in all groups. These results demonstrate that the E. cuniculi genotype III infection had a progressive course and surprisingly, Albendazole treatment had only a minimal effect. The E. cuniculi genotype III spore burden in individual organs reached up to 108 or 109 in immunocompetent or immunodeficient mice, respectively; however, these mice did not demonstrate any obvious clinical signs of microsporidiosis, and the immunodeficient mice survived longer. Our findings clearly show that the survival of mice does not correspond to spore burden, which provides new insight into latent microsporidiosis from an epidemiological point of view.

Encephalitozoon cuniculi Genotype III Evinces a Resistance to Albendazole Treatment in both Immunodeficient and Immunocompetent Mice.

Of four genotypes of Encephalitozoon cuniculi, E. cuniculi genotype II is considered to represent a parasite that occurs in many host species in a latent asymptomatic form, whereas E. cuniculi genotype III seems to be more aggressive, and infections caused by this strain can lead to the death of even immunocompetent hosts. Although albendazole has been considered suitable for treatment of Encephalitozoon species, its failure in control of E. cuniculi genotype III infection has been reported. This study determined the effect of a 100× recommended daily dose of albendazole on an Encephalitozoon cuniculi genotype III course of infection in immunocompetent and immunodeficient mice and compared the results with those from experiments performed with a lower dose of albendazole and E. cuniculi genotype II. The administration of the regular dose of abendazole during the acute phase of infection reduced the number of affected organs in all strains of mice and absolute counts of spores in screened organs. However, the effect on genotype III was minor. Surprisingly, no substantial effect was recorded after the use of a 100× dose of albendazole, with larger reductions seen only in the number of affected organs and absolute counts of spores in all strains of mice, implying variations in albendazole resistance between these Encephalitozoon cuniculi genotypes. These results imply that differences in the course of infection and the response to treatment depend not only on the immunological status of the host but also on the genotype causing the infection. Understanding how microsporidia survive in hosts despite targeted antimicrosporidial treatment could significantly contribute to research related to human health.

Encephalitozoon intestinalis: A new target for auranofin in a mice model.

Despite the fact that many approaches have been developed over years to find efficient and well-tolerated therapeutic regimens for microsporidiosis, the effectiveness of current drugs remains doubtful, and effective drugs against specific targets are still scarce. The present study is the first that was designed to evaluate the potency of auranofin, an anti-rheumatoid FDA approved drug, against intestinal Encephalitozoon intestinalis. Evaluation of the drug was achieved through counting of fecal and intestinal spores, studying the intestinal histopathological changes, measuring of intestinal hydrogen peroxide level, and post therapy follow-up of mice for 2 weeks for detection of relapse. Results showed that auranofin has promising anti-microsporidia potential. It showed a promising efficacy in mice experimentally infected with E. intestinalis. It has revealed an obvious reduction in fecal spore shedding and intestinal tissue spore load, amelioration of intestinal tissue pathological changes, and improvement of the local inflammatory infiltration without significant changes in hydrogen peroxide level. Interestingly, auranofin prevented the relapse of infection. Thus, considering the results of the present work, auranofin could be considered a therapeutic alternative for the gold standard drug 'albendazole' against the intestinal E. intestinalis infection especially in relapsing cases.

Effects of Homeopathic Phosphorus on Encephalitozoon cuniculi-Infected Macrophages In-Vitro.

INTRODUCTION: Encephalitozoon cuniculi (E. cuniculi), a fungus that acts as an intracellular pathogen, causes a marked neurological syndrome in many host species and is a zoonotic concern. Although no well-established treatment for this syndrome is known, previous successful clinical experience using homeopathic phosphorus has been described in which symptom remission with no mortality occurred in 40/42 animals by means of unknown immunological mechanisms. The latter observation was the main motivation for this study. OBJECTIVE: To verify, in an in-vitro model, if macrophages infected with E. cuniculi can change in function after treatment with different potencies of phosphorus. MATERIALS AND METHODS: RAW 264.7 macrophages were infected with E. cuniculi in-vitro and treated with various homeopathic potencies of phosphorus. The vehicle was used as a control solution (0.06% succussed ethanol). After 1 and 24 hours, the following parameters were analyzed: parasite internalization (by the Calcofluor staining method), lysosome activity (by the acridine orange method), cytokine/chemokine production (by the MAGPIX system), and cell ultrastructure. Automatic image analysis was used when applicable, and the experiments were performed in triplicate. RESULTS: Treatment with vehicle alone increased interleukin (IL)-6, tumor necrosis factor alpha and monocyte chemotactic protein -1 production (p ≤ 0.05) and reduced the number of internalized parasites (p ≤ 0.001). A progressive and time-dependent increase in RANTES (regulated on activation, normal T-cell expressed and secreted) and lysosome activity (p ≤ 0.002) was observed only after treatment with the highest potency of phosphorus (Phos 200cH), together with decreased apoptosis rate, intense parasite digestion, and the presence of non-internalized spores. CONCLUSIONS: Phos 200 cH has a modulatory action on the activity of infected macrophages, especially a specific increase in RANTES, a key element in the prognosis of E. cuniculi-infected and of immunosuppressed patients bearing infections.

The course of infection caused by Encephalitozoon cuniculi genotype III in immunocompetent and immunodeficient mice.

Encephalitozoon cuniculi is probably the most common microsporidia which infects a wide range of vertebrates, including human. So far, four genotypes of this parasite have been identified based on the rRNA internal transcribed spacer variations. The course of infection caused by E. cuniculi III had very massive onset in immunocompetent host characterized by the presence of this parasite in all organs and tissues within one week after peroral infection. Encephalitozoonosis caused by E. cuniculi III had very progressive spreading into all organs within first week post inoculation in immunocompromised SCID mice and led to the death of the host. The experimental treatment with albendazole of immunocompetent BALB/c mice infected with E. cuniculi III have shown very weak effect. Our findings clearly showed that the different course of infection and response to treatment depends not only on the immunological status of the host, but also on the genotype of microsporidia. It could be very important especially for individuals under chemotherapy and transplant recipients of organs originating from infected donors.

Effects of selected Indonesian plant extracts on E. cuniculi infection in vivo.

The present study was conducted to evaluate the methanolic extracts from several plant leaves widely used in traditional medicine to cure digestive tract disorders and in the self-medication of wild animals such as non-human primates, namely Archidendron fagifolium, Diospyros sumatrana, Shorea sumatrana, and Piper betle leaves, with regard to their antimicrosporidial activity against Encephalitozoon cuniculi in immunocompetent BALB/c mice determined using molecular detection of microsporidial DNA (qPCR) in various tissues and body fluids of infected, treated mice. Of the plant extracts tested, Diospyros sumatrana provided the most promising results, reducing spore shedding by 88% compared to untreated controls. Moreover, total burden per 1 g of tissue in the D. sumatrana extract-treated group reached 87% reduction compared to untreated controls, which was comparable to the effect of the standard drug, Albendazole. This data represents the baseline necessary for further research focused on determining the structure, activity and modes of action of the active compounds, mainly of D. sumatrana, enabling subsequent development of antimicrosporidial remedies.

Encephalitozoon cuniculi causes focal anterior cataract and uveitis in dogs.

Three mongrel dogs, aged 10 months (case 1), 14 months (case 2) and 7.5 years (case 3), were presented because of ophthalmologic disorders of 4 months, 6 months and 7 years duration, respectively. All three dogs were offspring of stray dogs from Hungary and Serbia and had positive serum antibody titres against Encephalitozoon (E.) cuniculi. The two young dogs showed unilateral, the older dog bilateral chronic anterior uveitis with posterior synechia and focal anterior cortical cataract. The fundi that could be evaluated developed focal tapetal hyporeflective lesions in the course of the disease. Dogs 1 and 2 underwent removal of the lens via phacoemulsification. PCR of the lens material was positive for E. cuniculi strains IV and II, respectively. In dog 2 findings suggestive of microsporidia were detected underneath the anterior lens capsule by immunohistochemical staining. In all cases medical treatment consisted of systemic fenbendazole, prednisolone, and topical anti-inflammatory drugs, and additional brinzolamid/timolol for dog 3. For the time being all cases (follow up 23 months, 6 months and 3 months, respectively) are still on topical anti-inflammatory therapy. It is concluded that E. cuniculi infections can cause cataract and chorioretinal lesions in dogs.

Microsporidiosis acquired through solid organ transplantation: a public health investigation.

BACKGROUND: Encephalitozoon cuniculi, a microsporidial species most commonly recognized as a cause of renal, respiratory, and central nervous system infections in immunosuppressed patients, was identified as the cause of a temporally associated cluster of febrile illness among 3 solid organ transplant recipients from a common donor. OBJECTIVE: To confirm the source of the illness, assess donor and recipient risk factors, and provide therapy recommendations for ill recipients. DESIGN: Public health investigation. SETTING: Two transplant hospitals and community interview with the deceased donor's family. PATIENTS: Three transplant recipients and the organ donor. MEASUREMENTS: Specimens were tested for microsporidia by using culture, immunofluorescent antibody, polymerase chain reaction,immunohistochemistry, and electron microscopy. Donor medical records were reviewed and a questionnaire was developed to assess for microsporidial infection. RESULTS: Kidneys and lungs were procured from the deceased donor and transplanted to 3 recipients who became ill with fever 7 to 10 weeks after the transplant. Results of urine culture, serologic,and polymerase chain reaction testing were positive for E. cuniculi of genotype III in each recipient; the organism was also identified in biopsy or autopsy specimens in all recipients. The donor had positive serologic test results for E. cuniculi. Surviving recipients received albendazole. Donor assessment did not identify factors for suspected E. cuniculi infection. LIMITATION: Inability to detect organism by culture or polymerase chain reaction in donor due to lack of autopsy specimens. CONCLUSION: Microsporidiosis is now recognized as an emerging transplant-associated disease and should be considered in febrile transplant recipients when tests for routinely encountered agents are unrevealing. Donor-derived disease is critical to assess when multiple recipients from a common donor are ill.

Effect of three drugs against Encephalitozoon cuniculi infection in immunosuppressed mice.

Microsporidia comprise a large group of obligate intracellular parasites. The microsporidian Encephalitozoon cuniculi causes disseminated infection in immunosuppressed patients with HIV, cancer, or transplants and in the elderly. In vivo and in vitro studies on the effectiveness of drugs are controversial. Currently, there is no effective treatment. We tested albendazole, albendazole sulfoxide, metronidazole, and cyclosporine in mice immunosuppressed with cyclophosphamide and inoculated by the intraperitoneal route with 10(7) E. cuniculi spores. One week after experimental inoculation, the mice were treated with albendazole, albendazole sulfoxide, metronidazole, and cyclosporine. Histological and morphometric analyses were performed to compare the treated groups. The state of immunosuppression was evaluated by phenotyping CD4(+) and CD8(+) T cells by flow cytometry. Nontreated mice showed acute disseminated and fatal encephalitozoonosis. The treatment with benzimidazoles significantly reduced infection until 30 days posttreatment (p.t.), but at 60 days p.t., the infection had recurred. Metronidazole decreased infection by a short time, and cyclosporine was not effective. All animals were immunosuppressed by all the experiments, as demonstrated by the low number of CD4(+) and CD8(+) T cells. We conclude that no drug was effective against E. cuniculi, but the benzimidazoles controlled the infection transiently.

Latent microsporidiosis caused by Encephalitozoon cuniculi in immunocompetent hosts: a murine model demonstrating the ineffectiveness of the immune system and treatment with albendazole.

BACKGROUND: Microsporidia are obligate intracellular parasites causing severe infections with lethal outcome in immunocompromised hosts. However, these pathogens are more frequently reported as latent infections in immunocompetent individuals and raises questions about the potential risk of reactivation following induced immunosuppression. AIMS: To evaluate the possibility latent microsporidiosis, efficacy or albendazole, and reactivation, the authors monitored the course of E. cuniculi infection in immunocompetent BALB/c mice and immunodeficient SCID mice using molecular methods. METHODS: Mice were per orally infected with 10(7) spores of E. cuniculi. Selected groups were treated with albendazole, re-infected or chemically immunosuppressed by dexamethasone. The presence of microsporidia in the host's organs and feces were determined using PCR methods. Changes in numbers of lymphocytes in blood and in spleen after induction of immunosuppression were confirmed using flow cytometry analysis. RESULTS: Whereas E. cuniculi caused lethal microsporidiosis in SCID mice, the infection in BABL/c mice remained asymptomatic despite parasite dissemination into many organs during the acute infection phase. Albendazole treatment led to microsporidia elimination from organs in BALB/c mice. In SCID mice, however, only a temporary reduction in number of affected organs was observed and infection re-established post-treatment. Dexamethasone treatment resulted in a chronic microsporidia infection disseminating into most organs in BALB/c mice. Although the presence of E. cuniculi in organs of albendazole- treated mice was undetectable by PCR, it was striking that infection was reactivated by immunosuppression treatment. CONCLUSION: Our results demonstrated that microsporidia can successfully survive in organs of immunocompetent hosts and are able to reactivate from undetectable levels and spread within these hosts after induction of immunosuppression. These findings stress the danger of latent microsporidiosis as a life-threatening risk factor especially for individuals undergoing chemotherapy and in transplant recipients of organs originating from infected donors.

Key diagnostic features of granulomatous interstitial nephritis due to Encephalitozoon cuniculi in a lung transplant recipient.

Microsporidia are increasingly recognized as opportunistic pathogens in immunocompromised organ transplant recipients (OTR). Disseminated infection due to Encephalitozoon sp. is reported mainly in human immunodeficiency virus (HIV)-positive patients and rarely in HIV-negative OTR. The clinical spectrum ranges from keratoconjunctivitis, to pneumonitis, to acute kidney injury. The kidney is a common site for disseminated infection; however, specialized techniques are required for definitive diagnosis. We report the first case of disseminated Encephalitozoon cuniculi infection in an HIV-negative lung transplant recipient diagnosed on renal biopsy. Five months after transplant, he presented with fever and a lung infiltrate and developed acute kidney injury. Renal biopsy showed granulomatous interstitial nephritis with gram-positive rod-shaped organisms with a "belt-like stripe" in tubular epithelial cells. Electron microscopy, polymerase chain reaction, and mammalian cell cultures of the urine sediment confirmed E. cuniculi infection. Retrospective review of a previous lung biopsy showed similar organisms. On the basis of electron microscopy findings, the patient was treated with albendazole, and immunosuppressive therapy was reduced. However, the patient expired due to Aspergillus pneumonia and disseminated E. cuniculi infection. Microsporidia should be considered in cases of fever of unknown origin and/or multiorgan infection in HIV-negative OTR when other causes have been excluded, as successful treatment requires early detection.

First case report of infection caused by Encephalitozoon intestinalis in a domestic cat and a patient with AIDS.

Microsporidia are eukaryotic, intracellular obligate parasites that infect invertebrate and vertebrate animals, and have emerged as important opportunistic parasites in AIDS patients. We used light microscopy to detect microsporidial spores in stool samples of a domestic cat confirmed as Encephalitozoon intestinalis by PCR, owned by an AIDS patient with chronic diarrhea and E. intestinalis infection. Cats can be considered hosts of E. intestinalis.

Disseminated microsporidiosis with Encephalitozoon species in a renal transplant recipient.

To our knowledge, 5 cases of disseminated microsporidiosis with Encephalitozoon species have been reported worldwide in transplant recipients. George et al. present the first such case in Australia, to be reported and treated with good clinical recovery.

Clinical evaluation of therapeutic success in rabbits with suspected encephalitozoonosis.

Between 2000 and 2008, 95 rabbits with suspected encephalitozoonosis and neurological symptoms were treated at the Clinic of Small Animal Medicine, Ludwig Maximilian University Munich. Standard treatment consisted of oxytetracycline (from 2000 to 2003; n=50) or fenbendazole and oxytetracycline (from 2004 to 2008; n=45), and the rabbits were randomly assigned to treatment groups with or without dexamethasone. Each therapeutic regime was given for 10 days, with fluids, B vitamins and nutritional support added as needed. Therapeutic success was evaluated by assessing the survival rate on day 10, the neurological score of the surviving rabbits and Kaplan-Meier curves for long-term survival past 10 days. Inclusion of fenbendazole in the treatment protocol was associated with increased survival rates on day 10 (p=0.043), better neurological scores (p=0.008), and improved long-term survival (p=0.025) based on the results of univariate analyses. Treatment with dexamethasone showed no effect on neurological score or on short- or long-term survival. The study did not provide any evidence that dexamethasone is an effective component of the treatment scheme.

Disseminated infection with a new genovar of Encephalitozoon cuniculi in a renal transplant recipient.

Disseminated microsporidiosis is a life-threatening opportunistic infection. Here, we report about a previously undescribed genovar of Encephalitozoon cuniculi causing disseminated infection in a non-HIV-infected renal transplant recipient. Disseminated microsporidiosis must be considered in the differential diagnosis of chronic fever in renal allograft recipients, even those without urinary symptoms.

Encephalitozoonosis in rabbits.

Encephalitozoon cuniculi is an obligatory intracellular microsporidian parasite that can infect a wide range of mammals, including rodents, rabbits, horses, carnivores and humans, in which the organism is known as an opportunistic pathogen of immunocompromised individuals. Nevertheless, the main host for E. cuniculi is the rabbit and infections usually have a sub-clinical course. However, severe disease is recognised in pet rabbits more frequently within the last years. As the central nervous system, the kidney and the eye are predilection organs for the organism, predominant histopathological alterations comprise granulomatous meningoencephalitis, chronic interstitial nephritis and phacoclastic uveitis. A definitive diagnosis of encephalitozoonosis in vivo is difficult, but it is important for specific treatment and the determination of possible zoonotic risks. This review article covers epidemiology, pathology, pathophysiology, immunology, clinical signs, differential diagnosis, diagnosis and treatment of encephalitozoonosis in rabbits.

[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.

Clinical symptoms and diagnosis of encephalitozoonosis in pet rabbits.

Infections with Encephalitozoon cuniculi in rabbits are observed at increasing frequency and are known as opportunistic infections in immunocompromised humans. 191 pet rabbits with suspected encephalitozoonosis, presented at the Animal Hospital of the Veterinary University of Vienna (Austria), were included in this study. Rabbits were serologically examined for antibodies against E. cuniculi (144 positive out of 184 rabbits with suspected encephalitozoonosis compared to 14 positive out of 40 clinically healthy rabbits tested as part of a standard health check) and Toxoplasma gondii (8 positive out of 157). Of the 144 seropositive rabbits with clinical signs, 75% showed neurological symptoms, 14.6% demonstrated phacoclastic uveitis and 3.5% suffered from renal failure. 6.9% of the animals had combined symptoms. Vestibular disease dominated within the rabbits that showed neurological symptoms. Polymerase chain reaction (PCR) could not detect parasite DNA in urine or cerebrospinal fluid (CSF), but did so in 4 out of 5 samples of liquefied lens material in cases with phacoclastic uveitis due to lens capsule rupture. Additionally further diagnostic procedures, such as inspection of the external ear canal (N=69), radiography of the tympanic bullae (N=65) were performed to rule out differential diagnosis. 54.2% of the patients exhibiting neurological symptoms recovered within a few days, while 87.5% of the rabbits suffering from renal failure died or had to be euthanized.