Prevalence of Enterocytozoon bieneusi in Swine: an 18-Month Survey at a Slaughterhouse in Massachusetts

ABSTRACT Slaughterhouse pig samples were analyzed by PCR for Enterocytozoon bieneusi infection. Thirty-two percent were found to be positive, with rates being higher over the summer months. Three isolates from pigs were identical in their ribosomal internal transcribed spacer sequence to human E. bieneusi type D, two were identical to type F (from a pig), and nine were previously unreported. The viability of these spores was demonstrated by their ability to infect gnotobiotic piglets. The presence of the infection in liver was shown by in situ hybridization.

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Transmission and Serial Propagation ofEnterocytozoon bieneusi from Humans and Rhesus Macaques in Gnotobiotic Piglets

Infection and Immunity ◽

10.1128/iai.66.11.5515-5519.1998 ◽

1998 ◽

Vol 66 (11) ◽

pp. 5515-5519 ◽

Cited By ~ 34

Author(s):

Ivanela Kondova ◽

Keith Mansfield ◽

Michael A. Buckholt ◽

Barry Stein ◽

Giovanni Widmer ◽

...

Keyword(s):

Animal Model ◽

In Situ Hybridization ◽

Chronic Diarrhea ◽

Rhesus Macaques ◽

Scientific Progress ◽

Enterocytozoon Bieneusi ◽

Oral Challenge ◽

Infected Piglet ◽

Gnotobiotic Piglets

ABSTRACT For over a decade Enterocytozoon bieneusi infections in people with AIDS have been linked with chronic diarrhea and wasting. The slow scientific progress in treating these infections is attributed to the inability of investigators to cultivate the parasite, which has also precluded evaluation of effective therapies. We report here successful serial transmissions of E. bieneusi from patients with AIDS and from macaques with AIDS to immunosuppressed gnotobiotic piglets. One infected piglet was still excreting spores at necropsy 50 days after an oral challenge. Spores in feces were detected microscopically by trichrome stain and by PCR and within enterocytes by in situ hybridization and immunohistochemistry. E. bieneusiinfection induced no symptoms. The development of an animal model forE. bieneusi will open up new opportunities for investigating this parasite.

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In situ hybridization: A molecular approach for the diagnosis of the microsporidian parasite Enterocytozoon bieneusi

Human Pathology ◽

10.1016/s0046-8177(99)90300-3 ◽

1999 ◽

Vol 30 (1) ◽

pp. 54-58 ◽

Cited By ~ 15

Author(s):

Jorge N Velasquez ◽

Silvana Carnevale ◽

Jorge H Labbe ◽

Agustin Chertcoff ◽

Marta G Cabrera ◽

...

Keyword(s):

In Situ Hybridization ◽

Enterocytozoon Bieneusi ◽

Molecular Approach ◽

Microsporidian Parasite

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Molecular Cloning and Functional Analysis of Three Type D Endogenous Retroviruses of Sheep Reveal a Different Cell Tropism from That of the Highly Related Exogenous Jaagsiekte Sheep Retrovirus

Journal of Virology ◽

10.1128/jvi.74.17.8065-8076.2000 ◽

2000 ◽

Vol 74 (17) ◽

pp. 8065-8076 ◽

Cited By ~ 87

Author(s):

Massimo Palmarini ◽

Claus Hallwirth ◽

Denis York ◽

Claudio Murgia ◽

Tulio de Oliveira ◽

...

Keyword(s):

In Situ Hybridization ◽

Endogenous Retroviruses ◽

Open Reading Frames ◽

Lung Epithelial Cells ◽

Structural Genes ◽

Jaagsiekte Sheep Retrovirus ◽

Type D ◽

Viral Particles ◽

Reading Frames

ABSTRACT Integrated into the sheep genome are 15 to 20 copies of type D endogenous loci that are highly related to two exogenous oncogenic viruses, jaagsiekte sheep retrovirus (JSRV) and enzootic nasal tumor virus (ENTV). The exogenous viruses cause infectious neoplasms of the respiratory tract in small ruminants. In this study, we molecularly cloned three intact type D endogenous retroviruses of sheep (enJS56A1, enJS5F16, and enJS59A1; collectively called enJRSVs) and analyzed their genomic structures, their phylogenies with respect to their exogenous counterparts, their capacity to form viral particles, and the expression specificities of their long terminal repeats (LTRs). In addition, the pattern of expression of enJSRVs in vivo was studied by in situ hybridization. All of the threeenJSRV proviruses had open reading frames for at least one of the structural genes. In particular, enJS56A1 had open reading frames for all structural genes, but it could not assemble viral particles when highly expressed in human 293T cells. We localized the defect for viral assembly in the first two-thirds of thegag gene by making a series of chimeras between enJS56A1 and the exogenous infectious molecular clone JSRV21. Phylogenetic analysis distinguished five ovine type D retroviruses:enJSRV groups A and B, ENTV, and two exogenous JSRV groups (African versus United Kingdom/North America isolates). Transient transfection assays indicated that the LTRs of the threeenJSRVs were not preferentially active in differentiated lung epithelial cells. This suggests that the pulmonary tropic JSRV developed from a type D retrovirus that did not have lung specificity. Consistent with this, in situ hybridization of a panel of normal ovine tissues revealed high expression of enJSRV mRNA in the luminal epithelium and glandular epithelium of the uterus; lower expression was localized in the lamina propria of the gut and in the bronchiolar epithelium of the lungs.

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Localization of Persistent Enterocytozoon bieneusiInfection in Normal Rhesus Macaques (Macaca mulatta) to the Hepatobiliary Tree

Journal of Clinical Microbiology ◽

10.1128/jcm.36.8.2336-2338.1998 ◽

1998 ◽

Vol 36 (8) ◽

pp. 2336-2338 ◽

Cited By ~ 34

Author(s):

Keith G. Mansfield ◽

Angela Carville ◽

Daniel Hebert ◽

Laura Chalifoux ◽

Daniel Shvetz ◽

...

Keyword(s):

In Situ Hybridization ◽

Macaca Mulatta ◽

Rhesus Macaques ◽

Clinical Signs ◽

Enterocytozoon Bieneusi ◽

Proximal Jejunum ◽

Microsporidian Parasite ◽

Immunodeficiency Virus ◽

History Of

Enterocytozoon bieneusi is the most common microsporidian parasite recognized in human patients with AIDS. Recently, we identified a virtually identical organism causing a spontaneous infection associated with hepatobiliary and intestinal disease in simian immunodeficiency virus (SIV)-infected macaques. To examine the natural history of the infection, we examined captive rhesus macaques for E. bieneusi by PCR, in situ hybridization, and cytochemical techniques. PCR performed on fecal DNA detected enterocytozoon infection in 22 (16.7%) of 131 normal rhesus macaques (Macaca mulatta), compared to 18 (33.8%) of 53 rhesus macaques experimentally inoculated with SIV. In normal rhesus macaques, persistence of infection was demonstrated for up to 262 days and was usually not associated with clinical signs. In six of seven normal rhesus animals, E. bieneusi was detected by PCR in bile obtained through percutaneous cholecystocentesis but not by in situ hybridization performed on endoscopic biopsies of duodenum and proximal jejunum.

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DNA sequence mapping in interphase and metaphase chromosomes by fluorescence in situ hybridization

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100122885 ◽

1992 ◽

Vol 50 (1) ◽

pp. 496-497

Author(s):

Barbara Trask ◽

Susan Allen ◽

Anne Bergmann ◽

Mari Christensen ◽

Anne Fertitta ◽

...

Keyword(s):

In Situ Hybridization ◽

Dna Sequence ◽

Dna Sequences ◽

Dual Band ◽

Nick Translation ◽

Metaphase Chromosomes ◽

Band Pass ◽

Texas Red ◽

Fluorescent Spot

Using fluorescence in situ hybridization (FISH), the positions of DNA sequences can be discretely marked with a fluorescent spot. The efficiency of marking DNA sequences of the size cloned in cosmids is 90-95%, and the fluorescent spots produced after FISH are ≈0.3 μm in diameter. Sites of two sequences can be distinguished using two-color FISH. Different reporter molecules, such as biotin or digoxigenin, are incorporated into DNA sequence probes by nick translation. These reporter molecules are labeled after hybridization with different fluorochromes, e.g., FITC and Texas Red. The development of dual band pass filters (Chromatechnology) allows these fluorochromes to be photographed simultaneously without registration shift.

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Ultrastructural analysis of the spatial distribution of MRNA

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100123167 ◽

1992 ◽

Vol 50 (1) ◽

pp. 552-553

Author(s):

Gary Bassell ◽

Robert H. Singer

Keyword(s):

Electron Microscopy ◽

Spatial Distribution ◽

In Situ Hybridization ◽

High Resolution ◽

Nucleic Acids ◽

Colloidal Gold ◽

Dna Probe ◽

Nucleotide Analogs ◽

Gold Particles

We have been investigating the spatial distribution of nucleic acids intracellularly using in situ hybridization. The use of non-isotopic nucleotide analogs incorporated into the DNA probe allows the detection of the probe at its site of hybridization within the cell. This approach therefore is compatible with the high resolution available by electron microscopy. Biotinated or digoxigenated probe can be detected by antibodies conjugated to colloidal gold. Because mRNA serves as a template for the probe fragments, the colloidal gold particles are detected as arrays which allow it to be unequivocally distinguished from background.

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