scholarly journals Diagnosis of Enterocytozoon bieneusi by PCR in Stool Samples Eluted from Filter Paper Disks

2000 ◽  
Vol 7 (3) ◽  
pp. 504-506 ◽  
Author(s):  
Silvana Carnevale ◽  
Jorge N. Velásquez ◽  
Jorge H. Labbé ◽  
Agustín Chertcoff ◽  
Marta G. Cabrera ◽  
...  
Keyword(s):  
Filter Paper ◽  
Enterocytozoon Bieneusi ◽  
Sample Collection ◽  
Fresh Feces ◽  
Stool Samples ◽  
Formalin Fixed

ABSTRACT We report a PCR-based assay for the detection ofEnterocytozoon bieneusi. We extracted DNA from feces which had been applied to filter paper disks and evaluated four preserving solutions. Infected specimens were identified by electrophoresis of amplicons from concentrated formalin-fixed samples and unconcentrated fresh feces. Our findings demonstrate that this methodology is effective for sample collection, mailing, and diagnosis of this pathogen.

scholarly journals Detection by an Immunofluorescence Test of Encephalitozoon intestinalis Spores in Routinely Formalin-Fixed Stool Samples Stored at Room Temperature

1999 ◽  
Vol 37 (7) ◽  
pp. 2317-2322 ◽  
Author(s):  
H. Moura ◽  
F. C. Sodre ◽  
F. J. Bornay-Llinares ◽  
G. J. Leitch ◽  
T. Navin ◽  
...  
Keyword(s):  
Gastrointestinal Disease ◽  
Good Alternative ◽  
Cross Reactivity ◽  
Enterocytozoon Bieneusi ◽  
Clinical Samples ◽  
Immunofluorescence Test ◽  
Encephalitozoon Intestinalis ◽  
Staining Techniques ◽  
Stool Samples ◽  
Formalin Fixed

Of the several microsporidia that infect humans,Enterocytozoon bieneusi is known to cause a gastrointestinal disease whereas Encephalitozoon intestinalis causes both a disseminated and an intestinal disease. Although several different staining techniques, including the chromotrope technique and its modifications, Uvitex 2B, and the quick-hot Gram-chromotrope procedure, detect microsporidian spores in fecal smears and other clinical samples, they do not identify the species of microsporidia. A need for an easily performed test therefore exists. We reevaluated 120 stool samples that had been found positive for microsporidia previously, using the quick-hot Gram-chromotrope technique, and segregated them into two groups on the basis of spore size. We also screened the smears by immunofluorescence microscopy, using a polyclonal rabbit anti-E. intestinalis serum at a dilution of 1:400. Spores in 29 (24.1%) of the 120 samples fluoresced brightly, indicating that they were E. intestinalis spores. No intense background or cross-reactivity with bacteria, yeasts, or other structures in the stool samples was seen. Additionally, the numbers of spores that fluoresced in seven of these samples were substantially smaller than the numbers of spores that were present in the stained smears, indicating that these samples were probably derived from patients with mixed infections of Enterocytozoon bieneusi and E. intestinalis. Because a 1:400 dilution of this serum does not react with culture-grown Encephalitozoon hellem,Encephalitozoon cuniculi, or Vittaforma corneaeor with Enterocytozoon bieneusi spores in feces, we concluded that an immunofluorescence test using this serum is a good alternative for the specific identification of E. intestinalis infections.

scholarly journals Comparative Assessment of In-House Real-Time PCRs Targeting Enteric Disease-Associated Microsporidia in Human Stool Samples

Pathogens ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 656
Author(s):  
Konstantin Tanida ◽  
Andreas Hahn ◽  
Kirsten Alexandra Eberhardt ◽  
Egbert Tannich ◽  
Olfert Landt ◽  
...  
Keyword(s):  
Real Time ◽  
Indigenous People ◽  
Real Time Pcr ◽  
Latent Class ◽  
Enterocytozoon Bieneusi ◽  
Enteric Disease ◽  
Immunosuppressed Patients ◽  
Study Population ◽  
Stool Samples ◽  
Pcr Assays

Microsporidiosis is an infection predominantly occurring in immunosuppressed patients and infrequently also in travelers. This study was performed to comparatively evaluate the diagnostic accuracy of real-time PCR assays targeting microsporidia with etiological relevance in the stool of human patients in a latent class analysis-based test comparison without a reference standard with perfect accuracy. Thereby, two one-tube real-time PCR assays and two two-tube real-time PCR assays targeting Enterocytozoon bieneusi and Encephalocytozoon spp. were included in the assessment with reference stool material (20), stool samples from Ghanaian HIV-positive patients (903), and from travelers, migrants and Colombian indigenous people (416). Sensitivity of the assays ranged from 60.4% to 97.4% and specificity from 99.1% to 100% with substantial agreement according to Cohen’s kappa of 79.6%. Microsporidia DNA was detected in the reference material and the stool of the HIV patients but not in the stool of the travelers, migrants, and the Colombian indigenous people. Accuracy-adjusted prevalence was 5.8% (n = 78) for the study population as a whole. In conclusion, reliable detection of enteric disease-associated microsporidia in stool samples by real-time PCR could be demonstrated, but sensitivity between the compared microsporidia-specific real-time PCR assays varied.

Techniques' Comparison and Report of the North Carolina Experience

PEDIATRICS ◽  
1989 ◽  
Vol 83 (5) ◽  
pp. 843-848
Author(s):  
Thomas R. Kinney ◽  
Martha Sawtschenko ◽  
Mary Whorton ◽  
Jean Shearin ◽  
Christy Stine ◽  
...  
Keyword(s):  
North Carolina ◽  
Filter Paper ◽  
Screening Program ◽  
Medical Center ◽  
Sample Collection ◽  
Capillary Tubes ◽  
Blood Samples ◽  
Screening Programs ◽  
The North ◽  
Duke University

Controversy still exists as to the best laboratory method to use to screen newborns for sickle cell disease and other hemoglobinopathies. The proposed methods include hemoglobin electrophoresis, column chromatography, isoelectric focusing, and high performance liquid chromatography. There is also debate concerning the preferred method of sample collection. The proposed methods of sample collection include cord blood or blood obtained from the infant collected in a tube with anticoagulant or on filter paper. We compared hemoglobin electrophoresis patterns from infant blood samples collected in heparinized capillary tubes and on filter paper. This comparison was performed because hemoglobin electrophoresis of dried blood samples collected on filter paper has been advocated as a practical, reliable, and inexpensive method for mass screening programs, although the limitations of this technique have not been explored fully. We also summarize data from the North Carolina Newborn Hemoglobinopathy Screening Program, which relates to the advantages and limitations of hemoglobin electrophoresis from filter paper blood specimens. MATERIALS AND METHODS Specimens Four sets of specimens were used for this study: (1) specimens collected at Duke University Medical Center to compare hemoglobin electrophoresis patterns of hemolysates from filter paper and heparinized capillary tubes, (2) specimens collected by the North Carolina program for hemoglobinopathy screening, (3) specimens routinely collected at Duke University in heparinized capillary tubes for newborn hemoglobinopathy screening, and (4) samples for retesting to examine the error rate of the state program and to confirm screening results compatible with a hemoglobinopathy. Samples for Direct Comparison Between Filter Paper and Heparinized Specimens

Urine and fecal sample collection on filter paper for ovarian hormone evaluations

1995 ◽  
Vol 37 (4) ◽  
pp. 305-315 ◽  
Author(s):  
S. E. Shideler ◽  
C. J. Munro ◽  
H. K. Johl ◽  
H. W. Taylor ◽  
B. L. Lasley
Keyword(s):  
Filter Paper ◽  
Fecal Sample ◽  
Ovarian Hormone ◽  
Sample Collection

Use of blood specimens collected on filter paper in screening for abnormal hemoglobins.

1976 ◽  
Vol 22 (5) ◽  
pp. 685-687 ◽  
Author(s):  
R M Schmidt ◽  
E M Brosious ◽  
S Holland ◽  
J M Wright ◽  
G R Serjeant
Keyword(s):  
Disease Control ◽  
Cellulose Acetate ◽  
Filter Paper ◽  
Whole Blood ◽  
Sample Collection ◽  
Capillary Tubes ◽  
Cellulose Acetate Electrophoresis ◽  
Blood Samples ◽  
Blood Specimens ◽  
The U.S

Abstract Both cellulose acetate electrophoresis and citrate agar electrophoresis were performed on 834 blood samples collected on filter paper in Jamaica and shipped for testing to the National Hemoglobinopathy Standardization Laboratory at the U.S. National Center for Disease Control. Additionally, 30 blood samples collected locally were stored on filter paper, in microhematocrit capillary tubes, and as whole blood specimens; at selected times the samples were tested for stability to determine the best sample-collection technique for hemoglobin electrophoresis. Results were most nearly accurate when both cellulose acetate electrophoresis and citrate agar testing were used. The methods are easy to perform, but results are unreliable if the blood samples on filter paper are stored at 4 degrees C for longer than two weeks before they are tested.

scholarly journals Detection and species identification of microsporidial infections using SYBR Green real-time PCR

2011 ◽  
Vol 60 (4) ◽  
pp. 459-466 ◽  
Author(s):  
Spencer D. Polley ◽  
Samuel Boadi ◽  
Julie Watson ◽  
Alan Curry ◽  
Peter L. Chiodini
Keyword(s):  
Real Time ◽  
Species Identification ◽  
Real Time Pcr ◽  
Simultaneous Detection ◽  
Enterocytozoon Bieneusi ◽  
Sybr Green ◽  
Pcr Assay ◽  
Real Time Analysis ◽  
Highly Sensitive ◽  
Stool Samples

Diagnosis of microsporidial infections is routinely performed by light microscopy, with unequivocal non-molecular species identification achievable only through electron microscopy. This study describes a single SYBR Green real-time PCR assay for the simultaneous detection and species identification of such infections. This assay was highly sensitive, routinely detecting infections containing 400 parasites (g stool sample)−1, whilst species identification was achieved by differential melt curves on a Corbett Life Science Rotor-Gene 3000. A modification of the QIAamp DNA tissue extraction protocol allowed the semi-automated extraction of DNA from stools for the routine diagnosis of microsporidial infection by real-time PCR. Of 168 stool samples routinely analysed for microsporidian spores, only five were positive by microscopy. By comparison, 17 were positive for microsporidial DNA by real-time analysis, comprising 14 Enterocytozoon bieneusi, one Encephalitozoon cuniculi and two separate Pleistophora species infections.

Spore shedding pattern of Enterocytozoon bieneusi in asymptomatic children

2005 ◽  
Vol 54 (5) ◽  
pp. 473-476 ◽  
Author(s):  
Mathirut Mungthin ◽  
Ittisak Subrungruang ◽  
Tawee Naaglor ◽  
Pote Aimpun ◽  
Wirote Areekul ◽  
...  
Keyword(s):  
Light Microscopy ◽  
Enterocytozoon Bieneusi ◽  
Detection Methods ◽  
Asymptomatic Group ◽  
Source Of Infection ◽  
Asymptomatic Children ◽  
Immunodeficiency Virus ◽  
Spore Shedding ◽  
Stool Samples ◽  
Stool Specimens

Stool samples from seven human immunodeficiency virus (HIV)-negative and two HIV-positive children with asymptomatic Enterocytozoon bieneusi infections were daily examined to quantify spore shedding using Gram-chromotrope staining under light microscopy. The spore shedding pattern and intensity in these children was variable. Mean spore concentrations in the stool samples from these children ranged from 2.4 × 102 to 1.2 × 105 spores per gram. Light microscopy could detect spores in stool specimens for 9–33 days, while PCR was able to detect E. bieneusi in stool specimens for 3–40 days longer. This suggests that light microscopy may not detect low levels of spore shedding. Considering that the asymptomatic group are a potential source of infection, detection methods with a higher sensitivity should be used.

scholarly journals Plasmodium falciparum Allelic Diversity: A Comparison of DNA Extraction from Isolates Collected on Rapid Diagnostic Tests (Rdts) and Filter Paper

2021 ◽  
Author(s):  
Denise Patricia Mawili-Mboumba ◽  
Marie Louise Tshibola Mbuyi ◽  
Noe Patrick M’bondoukwe ◽  
Marielle Karine Bouyou-Akotet
Keyword(s):  
Plasmodium Falciparum ◽  
Nucleic Acids ◽  
Filter Paper ◽  
Diagnostic Tests ◽  
Allelic Diversity ◽  
Storage Conditions ◽  
Rapid Diagnostic Tests ◽  
Sample Collection ◽  
Filter Papers

Background: To perform molecular epidemiologic studies based on large cohorts, material such as RDTs or filter papers are useful for biological sample collection and extraction of RNA or DNA of good quality. Thus, we aimed to assess the quality of DNA extracted from malaria rapid diagnostic tests (RDTs) stored at various temperatures for the analysis of Plasmodium falciparum genetic diversity. Methods: Febrile patients benefitted from free malaria diagnosis using microscopy in a malaria sentinel site, at the Regional Hospital Estuaire-Melen, in Gabon, in 2015. P. falciparum isolates were collected onto one filter paper and 2 similar RDTs devices (Acon®) per patient. Nucleic acids were extracted with QiAmp Qiagen kit from paper and RDTs and the quality of the DNA was analyzed by msp1 gene amplification. Results: Msp1gene amplification was achieved in nucleic acids extracted from all filter papers and RDTs devices (n = 45, 100%). K1 alleles were detected in 93.3% (n = 42/45) of the samples and Mad20 alleles in 73.3% (n = 33/45). The number and the intensity of K1 and/or Mad20 fragments were comparable according to the sample collection material and the storage conditions (room temperature vs -20°C) of the samples. The size of the fragments indicating allelic diversity was comparable in 80% (n=36) of the samples. Conclusion: These data show that RDTs are a valuable source of DNA for malaria parasite genetic polymorphism analysis. Storage conditions of the devices did not influence the quality of DNA extracted from RDTs device, although some alleles may be missed.

scholarly journals Prevalence of microsporidiosis in human and cattle

2017 ◽  
Vol 40 (2) ◽  
pp. 147-154
Author(s):  
Kadum N. E.
Keyword(s):  
Rural Areas ◽  
Common Species ◽  
Enterocytozoon Bieneusi ◽  
Human Infection ◽  
Urine Samples ◽  
Milk Samples ◽  
Encephalitozoon Intestinalis ◽  
Blue Stain ◽  
Stool Samples ◽  
Apparently Healthy

     In order to identify microsporidia and other fungi  in  stool and  urine samples of human, and in fecal and milk samples of cattle, 100 stool samples with or without diarrhea and 50 urine samples, human fecal and urine samples were obtained from certain Baghdad hospitals and certain rural areas surroundings Baghdad city, in addition to 50 fecal and 56  milk samples  of cattle apparently healthy were collected  from Alshula Slaughter House and  directly from  anal of the animal field  of College of Veterinary Medicine/ Baghdad University. All samples were collected during six months from 1/10/2014 to 1/4/2015. Thin films were formed and stained by Webers Modified Trichrom stain and Modified Trichrom-Ryan Blue stain. The results showed that (23%) 23 out of 100 stool samples of human were positive for Microsporidia spp. and (16%) 8 out of 50 urine samples of human were positive for this fungus. While the result revealed (18%) 9 out of 50 fecal samples and (7.14%) 4 out of 56 milk samples of cattle were positive for Microsporidia spp. The result also explained that (25.3%) 19 cases of patients suffering from diarrhea expressed   Microsporidia spp. after the examination of 75 stool samples, while (16%) 4 persons without diarrhea showed positive Microspordia, through the examination of 25 stool samples. The study explains that the  Enterocytozoon bieneusi is a common species associated with human infection and  Encephalitozoon intestinalis is  a common Microsporidia associated with  cattle infection  whereas Encephalitozoon cuniculi is rarely identified in human  but recorded in cattle.

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