Human microsporidial infections.

Microsporidia are obligate intracellular spore-forming protozoal parasites belonging to the phylum Microspora. Their host range is extensive, including most invertebrates and all classes of vertebrates. More than 100 microsporidial genera and almost 1,000 species have now been identified. Five genera (Enterocytozoon spp., Encephalitozoon spp., Septata spp., Pleistophora sp., and Nosema spp.) and unclassified microsporidia (referred to by the collective term Microsporidium) have been associated with human disease, which appears to manifest primarily in immunocompromised persons. The clinical manifestations of microsporidiosis are diverse and include intestinal, pulmonary, ocular, muscular, and renal disease. Among persons not infected with human immunodeficiency virus, ten cases of microsporidiosis have been documented. In human immunodeficiency virus-infected patients, on the other hand, over 400 cases of microsporidiosis have been identified, the majority attributed to Enterocytozoon bieneusi, an important cause of chronic diarrhea and wasting. Diagnosis of microsporidiosis currently depends on morphological demonstration of the organisms themselves. Initial detection of microsporidia by light microscopic examination of tissue sections and of more readily obtainable specimens such as stool, duodenal aspirates, urine, sputum, nasal discharge, bronchoalveolar lavage fluid, and conjunctival smears is now becoming routine practice. Definitive species identification is made by using the specific fluorescein-tagged antibody (immunofluorescence) technique or electron microscopy. Treatment options are limited, but symptomatic improvement of Enterocytozoon bieneusi infection may be achieved with the anthelmintic-antiprotozoal drug albendazole. Preliminary observations suggest that Septata intestinalis and Encephalitozoon infections may be cured with albendazole. Progress is being made with respect to in vitro propagation of microsporidia, which is crucial for developing antimicrosporidial drugs. Furthermore, molecular techniques are being developed for diagnostic purposes, taxonomic classification, and analysis of phylogenetic relationships of microsporidia.     

Morphological and ultrastructural description of Pleistophora dammami sp. n. infecting the intestinal wall of Saurida undosquamis from the Arabian Gulf, Saudi Arabia

Pleistophora dammami sp. n. is described from Saurida undosquamis from the Arabian Gulf in Saudi Arabia. Infection appeared as whitish cysts in the intestinal wall. Cysts ranged in size from 1 to 4 mm. The prevalence of the infection across both fish sexes was 17.5% (24/420). Two kinds of spores were recognized, microspores and macrospores, and each were ovoid in shape. The microspores measured ~2.5 × 2.0 μm in size, while the macrospores measured ~6.0 × 3.0 μm. Ultrastructurally, the parasite did not form xenoma but it formed cysts surrounded by thick cyst wall. All stages of development as meronts, sporonts, sporoblast and spores occurred in the cytoplasm of the host cells within sporophorous vesicles. The stages of development occurred asynchronously and thus all stages were randomly distributed within the cysts. Meronts were elliptical and multinucleated, with unpaired nuclei which constantly divided giving rise to new sporonts. During the transition to sporonts, the border of the meronts increased in thickness to form dense discontinuous cell coat. Later, the sporont divided into sporoblast cells which gradually differentiated the typical organelles of the spores. In mature spores, the polar filament was arranged in 20-24 coils in two rows either side of the posterior vacuole. All ultrastructural and morphological criteria indicate that the described species belongs to the genus Pleistophora.     

Chronic Shigella flexneri infection preceding development of acquired immunodeficiency syndrome

Shigella sp. is known to be an important cause of diarrhea in homosexual men, although chronic infection is infrequently recognized. We describe recurrent and relapsing symptomatic infection due to Shigella flexneri in a human immunodeficiency virus-infected homosexual man subsequently developed acquired immunodeficiency syndrome. Patients with acquired immunodeficiency syndrome may be prone to developing chronic shigellosis because of impaired intestinal cell-mediated immunity.     

Human microsporidiosis: Clinical, diagnostic and therapeutic aspects of an increasing infection

Human microsporidiosis is a parasitic infection due to species of four different genera: Encephalitozoon; Enterocytozoon; Nosema; and Pleistophora. Although well known as a cause of disease in animals, microsporidiosis was only occasionally reported in humans. Recently, in human immunodeficiency virus (HIV)-infected patients, microsporidia belonging to Encephalitozoon and Enterocytozoon species have proved to be important opportunistic pathogens. Enterocytozoon bieneusi is associated with chronic intermittent diarrhea, cholangiopathy and sinusitis whereas Encephalitozoon intestinalis, Encephalitozoon hellem and Encephalitozoon cuniculi , the three Encephalitozoon species found in humans, are associated with diarrhea, rhinosinusitis, keratoconjunctivitis, nephritis and hepatitis. Diagnosis of microsporidial infections in humans was until recently an invasive, laborious procedure including electron microscopy of small intestine biopsies. However, new simple staining methods using Uvitex 2B or modified trichrome stain for feces and other body fluids have facilitated clinical diagnosis as well as drug evaluation and epidemiological studies. The application of monoclonal antibodies and molecular techniques such as the polymerase chain reaction have further improved microsporidial diagnosis. Treatment of Entero. bieneusi has, until now, been unsuccessful whereas albendazole has proved to be an effective treatment for Encephalitozoon species infection. Identification of effective treatment for Entero. bieneusi infections and further study of the pathogenicity of these microsporidial infections in immunocompetent hosts are important future challenges.     

Intestinal microsporidiosis as a cause of diarrhea in human immunodeficiency virus-infected patients: a report of 20 cases.

Chronic diarrhea accompanied by weight loss is a common and often debilitating problem associated with human immunodeficiency virus (HIV) infection. Enterocytozoon bieneusi, a newly identified species of the phylum of protozoa, Microspora, has been reported associated with chronic diarrhea and wasting in 11 acquired immunodeficiency syndrome (AIDS) patients in the United States, Europe, and Africa. Diagnosis has been based solely on the ultrastructural identification of this small, intracellular parasite in bowel biopsies. Seventy-one small bowel biopsies from 67 homosexual AIDS and AIDS-related complex patients with chronic diarrhea and with no pathogens identified by light microscopy on paraffin sections, were embedded in plastic and studied by light and transmission electron microscopy. Enterocytozoon bieneusi microsporidiosis was diagnosed by electron microscopy in 20 (22 biopsies) of the patients. More jejunal biopsies (16 of 36) were positive than duodenal biopsies (six of 35). Parasites and spores were clearly visible at the light microscopic level in the semi-thin plastic sections from 17 and 21 of the biopsies, respectively. In retrospect, parasites could be identified by light microscopy in standard hematoxylin and eosin-stained paraffin sections. Infection was confined to enterocytes covering the villi, especially the tips, and was associated with villous atrophy and cell degeneration, necrosis, and sloughing. Release of spores into the bowel lumen was evident. Colorectal biopsies from two of the patients with small bowel microsporidiosis were negative for microsporidia. Enterocytozoon bieneusi infection of the small bowel may be an important cause of diarrhea in HIV-infected persons.     

A microsporidian previously undescribed in humans, infecting enterocytes and macrophages, and associated with diarrhea in an acquired immunodeficiency syndrome patient.

To date, the only microsporidian that has been associated with diarrhea and weight loss in acquired immunodeficiency syndrome patients is the newly identified Enterocytozoon bieneusi. A second species is now described that was associated with intestinal symptoms in a 32-year-old, human immunodeficiency virus- seropositive, Native American male homosexual. Stool studies and routine light microscopy of multiple small intestinal biopsies that showed atrophy with acute and chronic inflammation were without apparent pathogens. Light microscopy of semi-thin plastic sections, cytochemical stains of paraffin sections, and ultrastructural studies revealed extensive microsporidial infection of enterocytes and submucosal macrophages. No other pathogens were identified. Unlike E bieneusi, this microsporidian appeared to develop within septated parasitophorous vacuoles, and lacked polar disks and clear clefts. It most closely resembled, but was distinguishable from, members of the genus Encephalitozoon. Awareness of the microsporidia as potential opportunists in acquired immunodeficiency syndrome patients is increasing the incidence of identification of these organisms.     

Rapid identification of Mycobacterium genavense with a new commercially available molecular test, INNO-LiPA MYCOBACTERIA v2

We report a rare mesenteric localized Mycobacterium genavense infection in a severely immunocompromised human immunodeficiency virus-infected patient. An INNO-LiPA MYCOBACTERIA v2 test was performed directly on biopsy samples. This new molecular tool could be used for simultaneous identification of mycobacterium species from human specimens, but other studies are needed to validate our first results.     

Myositis due to the microsporidian Anncaliia (Brachiola) algerae in a lung transplant recipient

Microsporidia are obligate intracellular parasites, more closely related to fungi than protozoa on molecular phylogenetic analysis, and are known to be a rare cause of opportunistic infection in immune compromised patients including human immunodeficiency virus-positive patients and solid organ transplant recipients. We report the first case to our knowledge of microsporidial myositis in a lung transplant recipient. He was 49?years old and had received a lung transplant in 2000 for cystic fibrosis. He presented in 2009 with fevers, chronic diarrhea, myalgia, and pancytopenia, and developed progressive weakness and neurological symptoms before his death 35?days after hospital admission. Multiple investigations, including stool culture, rectal biopsy, colonoscopy, cerebrospinal fluid examination, bone marrow biopsy, lung biopsy, and bronchoalveolar lavage, failed to reveal a definite cause for the patient's deterioration. The diagnosis of microsporidial infection was made on post-mortem light microscopic examination of tissue sections of the tongue and deltoid muscle. Light microscopy diagnosed a microsporidial myositis, confirmed by transmission electron microscopy, which suggested that the organism was Brachiola species. The identity of the organism was confirmed by polymerase chain reaction as Brachiola algerae (recently renamed Anncaliia algerae). The case highlights the need to consider protozoal organisms in the differential diagnosis of myalgia and multisystemic infections in immune compromised patients.     

Ultrastructure of the development of a species ofEncephalitozoon cultured from the eye of an AIDS patient

Human fibroblast cell cultures inoculated with microsporidia-infected corneal scrapings from an AIDS patient were fixed in situ and examined by scanning and transmission electron microscopy. The parasite grew prolifically and all developmental stages were observed. Meronts underwent binary fission and the daughter cells transformed into clongate, chain-like sporonts that eventually separated into sporoblasts. The formation of components of the mature spores is described. The parasite, a species ofEncephalitozoon, underwent development both in the cytoplasm and within a parasitophorous vacuole, distinguishing it from the morphologically similar speciesE. cuniculi andE. hellem, both of which have been described from lesions in the human eye and have been reported to develop exclusively within a parasitophorous vacuole.This project was funded by Natural Sciences and Engineering Research Council of Canada Operating Grant 6965 (to S.S.D.)     

New mouse models for chronic Cryptosporidium infection in immunodeficient hosts.

Cryptosporidium sp. causes fulminant diarrhea and chronic infection in immunocompromised, particularly human immunodeficiency virus-infected, persons. The lack of in vitro cultivation and a suitable animal model has limited development of effective treatment. We describe two new mouse models of chronic symptomatic cryptosporidiosis in adult athymic mice and in T-cell subset-depleted mice. A progressive infection, fatal within 4 months, occurred in most adult athymic mice; a few developed stable infections. Symptoms included dehydration, weight loss, intermittent diarrhea, and jaundice. Pathologic abnormalities and organisms localized in the intestine in stable infections but involved the hepatobiliary tree and pancreas in others. Lymphoid cells from histocompatible, Cryptosporidium sp.-immune mice cured infected nude mice. Identical infections occurred in neonatally infected BALB/c mice treated with anti-CD4 monoclonal antibodies alone or also with anti-CD8 monoclonal antibodies; the mice were cured when the monoclonal antibody treatments were stopped. These models will be useful in definition of the immune defects that permit chronic cryptosporidiosis to develop and in assessment of treatment modalities.     

Scytalidium dimidiatum and Lecythophora hoffmannii: unusual causes of fungal infections in a patient with AIDS.

Immunocompromised patients are susceptible to infections by fungi that seldom cause disease in humans. We describe a human immunodeficiency virus-infected patient who had simultaneous infections with two fungi which are rare causes of serious infection: Lecythophora hoffmannii, causing chronic sinusitis, and Scytalidium dimidiatum, causing skin lesions, lymphangitis, and lymphadenitis. The clinical and pathologic findings are discussed.     

Ultrastructure and molecular phylogenetics of a new isolate of Pleistophora pagri sp. nov. (Microsporidia, Pleistophoridae) from Pagrus pagrus in Egypt

The spore morphology and molecular systematic of a new microsporidian which was isolated from the common sea bream Pagrus pagrus (F: Sparidae Linnaeus, 1758) from the Red Sea, Egypt have been studied. Fifty-six out of 300 (18.7?%) of this fish were infected with microsporidian parasites. The infection was appeared as whitish, ellipsoid, round, or elongated nodules embedded in the epithelial lining of the peritoneum and also in the intestinal epithelium. Light microscopic study revealed that nodules were encapsulated by a fibrous layer encircling numerous mature spores measuring 1.7?±?0.6 (1.5-2.7?μm)?×?1.5?±?0.3?μm (1.2-1.8?μm) in size. Ultrastructure of spores was characteristic for the genus Pleistophora: dimorphic, uninucleate spores (each spore possesses three to five polar filament coils) and a posterior vacuole. Also, the early recognizable stages of the parasite within nodules include uninucleated, binucleated, and multinucleated meronts followed by detachment of the plasmalemma of the sporont producing sporoblasts which mature to spores that consist of a spore coat and spore contents. Also, we analyzed the small subunit ribosomal gene (SSUrDNA) using PCR and sequencing specimens from the marine populations of P. pagrus fish from the Red Sea. From blast searches, sequence analysis, and phylogenetic analysis, we did not find corresponding GenBank entries to our species. Comparison of the nucleotide sequences showed that the sequence of our microsporidium was most similar to five Pleistophora species with degrees of identity (>91.5?%). It was most similar (97.8?% identity) to that of Pleistophora hyphessobryconis (account no. GU126672) differing in 19 nucleotide positions and with lower divergence value, Pleistophora ovariae (96.2?% identity, account no. AJ252955), Pleistophora hippoglossoideos (91.9?% identity, account no. AJ252953), Pleistophora mulleri (91.9?% identity, account no. EF119339), and Pleistophora typicalis (91.9?% identity, account no. AJ252956). So, they likely represent new species named Pleistophora pagri sp. n. with accession number JF797622 and a GC content of 53?%.     

Brain abscess caused by Nocardia cyriacigeorgica in a patient with human immunodeficiency virus infection

Nocardia cyriacigeorgica is a recently characterized species within the genus of Nocardia. We report a brain abscess, following a primary pulmonary colonization, due to this species in a human immunodeficiency virus-infected patient. This case confirms that isolation of Nocardia in sputum is associated with a high risk of disseminated infection in immunocompromised patients.     

Endocarditis caused by Rochalimaea quintana in a patient infected with human immunodeficiency virus.

Rochalimaea quintana and Rochalimaea henselae are closely related, fastidious, gram-negative rickettsiae. Thus far, the spectrum of human Rochalimaea sp. infections has not included endocarditis. We describe a 50-year-old human immunodeficiency virus-positive man who developed endocarditis caused by R. quintana. DNA relatedness studies, which compared our patient's blood culture isolate with known Rochalimaea species, identified the organism as R. quintana. Our report expands the spectrum of Rochalimaea sp. infections and identifies a new infectious cause of endocarditis.     

Sensitivity and specificity of a monoclonal antibody-based fluorescence assay for detecting Enterocytozoon bieneusi spores in feces of simian immunodeficiency virus-infected macaques

Enterocytozoon bieneusi is clinically the most significant among the microsporidia causing chronic diarrhea, wasting, and cholangitis in individuals with human immunodeficiency virus/AIDS. Microscopy with either calcofluor or modified trichrome stains is the standard diagnostic test for microsporidiosis and does not allow species identification. Detection of E. bieneusi infection based on PCR is limited to a few reference laboratories, and thus it is not the standard diagnostic assay. We have recently reported the development and characterization of a panel of monoclonal antibodies against E. bieneusi, and in this publication we evaluated the specificity and sensitivity of an immunofluorescence assay (IFA), compared with PCR, in simian immunodeficiency virus-infected macaques. The IFA, which correlated with the primary PCR method, with a detection limit of 1.5 x 10(5) spores per gram of feces, will simplify considerably the detection of E. bieneusi spores in clinical and environmental specimens and in laboratory and epidemiological investigations.     

Improved light-microscopical detection of microsporidia spores in stool and duodenal aspirates. The Enteric Opportunistic Infections Working Group.

BACKGROUND. The diagnosis of infection with Enterocytozoon bieneusi, a microsporidian organism that causes chronic diarrhea in patients infected with the human immunodeficiency virus (HIV), has depended on invasive procedures. We have developed a new method to detect microsporidia spores in feces and duodenal aspirates. METHODS. Stool was obtained from four HIV-infected patients with biopsy-confirmed intestinal microsporidiosis. Slides prepared from unconcentrated, formalin-fixed stool specimens were stained with a new chromotrope-based technique and examined by light microscopy. Methods of stool concentration were also compared. The technique was then evaluated by examining 215 specimens from 134 HIV-infected persons with or without diarrhea. In addition, duodenal aspirates from 10 patients with unexplained chronic diarrhea were examined by light microscopy after staining according to the new and the traditional techniques. RESULTS. E. bieneusi spores were found in all unconcentrated stool specimens from the four patients with microsporidiosis. The use of various methods of stool concentration did not improve the detection of microsporidia spores. In the prospective study, microsporidiosis was detected in samples from 6 of 27 patients with chronic diarrhea, but in none of those from 42 patients with acute diarrhea or 65 patients without diarrhea. The presence of microsporidia spores in stool specimens and duodenal aspirates allowed the successful prediction of the presence of microsporidia in small-bowel biopsy specimens from all four patients who subsequently underwent endoscopy. CONCLUSIONS. E. bieneusi is an important cause of chronic diarrhea in HIV-infected persons. This new diagnostic technique serves as a practical, noninvasive means to detect microsporidia spores in stool specimens and is also applicable to the examination of duodenal aspirates.     

Alga associated with diarrhea in patients with acquired immunodeficiency syndrome and in travelers.

Spherical bodies resembling coccidian oocysts and measuring 8.0 to 9.0 microns in diameter were seen in the stools of eight persons with explosive, watery diarrhea. Seven had recently traveled to tropical countries, mostly in the Caribbean, and four had acquired immunodeficiency syndrome. The structures were easily discernible in wet mounts by light microscopy and contained variable numbers of granular inclusions, but were refractory to, or stained partially with, 12 commonly used laboratory stains. Electron microscopy revealed an outer fibrillar coat, a thin cell wall, granules, and organelles which were not surrounded by membranes. One type of organelle was similar to the thylakoid photosynthesizing organelles of blue-green algae (cyanobacteria). These findings indicate that the bodies may be a species of blue-green algae.     

Methods for detection of an intestinal secretory immunoglobulin A response to Candida spp. and their preliminary application in human immunodeficiency virus-infected patients with chronic diarrhea.

Six of 11 human immunodeficiency virus (HIV)-infected patients with chronic diarrhea, shedding only Candida spp. in their stools, elicited a Candida-specific secretory immunoglobulin A response. Similar responses were identified in only 1 of 10 HIV-positive patients with chronic diarrhea but without Candida spp. and in none of 10 HIV-negative subjects without diarrhea. Candida spp. may play a role in the etiology of chronic diarrhea associated with HIV infection.     

Ultrastructural insights into morphology and reproductive mode of <Emphasis Type="Italic">Blastocystis hominis</Emphasis>

To understand well the morphology and reproductive mode of Blastocystis hominis, with the help of transmission electron microscopy and scanning electron microscopy the ultrastructural details of B. hominis from fresh diarrheal specimens and cultured strains were observed. In both fecal samples and culture conditions, there were vacuolar and granular forms. In diarrhea, it exists in multivacuolar, avacuolar, and amoeboid forms. In the in vitro culture, vacuolar form could transform to granular form. The most commonly noticed structure on the cell surface was surface coat with diversity in appearance (the funiform, lamellar, filiform, and floccose in different thickness) and distributions. Three modes of reproduction were confirmed, they were binary fission, plasmotomy, and budding. Under the impact of host’s response, the ultrastructures of surface coat, nucleus, and mitochondrion-like organelle sometimes changed.     

Abdominal visceral peliosis associated with bacillary angiomatosis. Ultrastructural evidence of endothelial destruction by bacilli.

Peliosis involving solid internal organs is a rare entity, and it has been reported in association with chronic debilitating diseases. Bacillary angiomatosis (BA), on the other hand, is a recently identified lesion found virtually only in individuals infected by the human immunodeficiency virus. We describe herein two cases of visceral BA and peliosis in human immunodeficiency virus-infected patients. Based on light and electron microscopic findings, we conclude that (1) BA bacilli present in the hepatic sinusoidal endothelial cells, in a suitable host milieu, may be the causative agents of peliosis hepatis; (2) BA bacilli can be found both intracellularly and extracellularly; and (3) peliosis is also identified in association with BA in abdominal lymph nodes.