title: Highly specific detection of Cryptosporidium spp. oocysts in human stool samples by undemanding and inexpensive phase contrast microscopy.
abstract: To compare phase contrast microscopy (PCM) of unstained slides for the detection of Cryptosporidium spp. oocysts with a commercially available enzyme immunoassay (EIA) for the detection of cryptosporidial antigen in human stool samples, we prospectively analysed by both methods 463 fresh human stool samples obtained from diarrhoeic patients between July and October 2014. Compared with the EIA, the sensitivity, specificity, positive and negative predictive value of PCM were 88.9 % (95 % confidence interval (CI), 66.0-98.1 %), 100 % (95 % CI, 99.0-100 %), 100 % (95 % CI, 77.3-100 %) and 99.6 % (95 % CI, 98.3-100 %), respectively. Additionally, we retrospectively examined with PCM 65 fixed stool samples that had been collected in 2010 from mostly asymptomatic Rwandan children <5 years of age; 14 of these samples had previously yielded positive results with a highly sensitive real-time (RT)-PCR. PCM detected cryptosporidia in 5/14 RT-PCR-positive samples, and notably, also in one of 51 RT-PCR-negative samples, which was subsequently confirmed by acid-fast staining. Positive and negative percent agreement of PCM with RT-PCR were 35.7 % (95 % CI, 16.2-61.4 %) and 98.0 % (95 % CI, 88.7-100 %), respectively. Positive PCM results were associated with higher RT-PCR cycle threshold values (p = 0.044). In conclusion, PCM offers a highly specific, undemanding and inexpensive method for the laboratory diagnosis of acute human cryptosporidiosis independent of the causative Cryptosporidium species.
authors.affiliation: Laboratory Enders and Partners, Rosenbergstr. 85, 70193, Stuttgart, Germany. ralf.ignatius@charite.de.|Department of Microbiology and Hygiene, Charité-Universitätsmedizin Berlin, Berlin, Germany. ralf.ignatius@charite.de.
authors.forename: Ralf
authors.initials: R
authors.lastname: Ignatius
authors.affiliation: Laboratory Enders and Partners, Rosenbergstr. 85, 70193, Stuttgart, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Klemm
authors.affiliation: Institute of Tropical Medicine and International Health, Charité-Universitätsmedizin Berlin, Berlin, Germany.|Agents of Mycoses, Parasitoses and Mycobacterioses, Robert Koch-Institute, Berlin, Germany.
authors.forename: Steffen
authors.initials: S
authors.lastname: Zander
authors.affiliation: University Teaching Hospital of Butare and School of Medicine and Pharmacy, University of Rwanda, Butare, Rwanda.
authors.forename: Jean Bosco
authors.initials: JB
authors.lastname: Gahutu
authors.affiliation: Laboratory Enders and Partners, Rosenbergstr. 85, 70193, Stuttgart, Germany.
authors.forename: Peter
authors.initials: P
authors.lastname: Kimmig
authors.affiliation: Institute of Tropical Medicine and International Health, Charité-Universitätsmedizin Berlin, Berlin, Germany.
authors.forename: Frank P
authors.initials: FP
authors.lastname: Mockenhaupt
authors.affiliation: Laboratory Enders and Partners, Rosenbergstr. 85, 70193, Stuttgart, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Regnath
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000818:Animals; D000953:Antigens, Protozoan / Q000302:isolation & purification*; D002648:Child; D002675:Child, Preschool; D003457:Cryptosporidiosis / Q000175:diagnosis*; D003458:Cryptosporidium / Q000276:immunology / Q000302:isolation & purification*; D003967:Diarrhea / Q000469:parasitology; D005243:Feces / Q000469:parasitology*; D005260:Female; D006801:Humans; D007124:Immunoenzyme Techniques; D007223:Infant; D008297:Male; D008858:Microscopy, Phase-Contrast; D008875:Middle Aged; D033661:Oocysts; D011446:Prospective Studies; D060888:Real-Time Polymerase Chain Reaction; D012189:Retrospective Studies; D012432:Rwanda; D012680:Sensitivity and Specificity; D055815:Young Adult
chemical_list: D000953:Antigens, Protozoan
keywords: Cryptosporidium; ELISA; Phase contrast microscopy; RT-PCR; Rwandan children