Assessment of the first commercial multiplex PCR kit (ParaGENIE Crypto-Micro Real-Time PCR) for the detection of Cryptosporidium spp., Enterocytozoon bieneusi, and Encephalitozoon intestinalis from fecal samples

2019 ◽  
Vol 95 (1) ◽  
pp. 34-37 ◽  
Author(s):  
Florent Morio ◽  
Philippe Poirier ◽  
Yohann Le Govic ◽  
Adrien Laude ◽  
Stéphane Valot ◽  
...  
Keyword(s):  
Multiplex Pcr ◽  
Real Time ◽  
Real Time Pcr ◽  
Enterocytozoon Bieneusi ◽  
Fecal Samples ◽  
Cryptosporidium Spp ◽  
Encephalitozoon Intestinalis

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.

scholarly journals A survey on the occurrence of Brachyspira pilosicoli and Brachyspira hyodysenteriae in growing-finishing pigs

F1000Research ◽  
2019 ◽  
Vol 8 ◽  
pp. 1702
Author(s):  
Arkadiusz Dors ◽  
Ewelina Czyżewska-Dors ◽  
Grzegorz Woźniakowski
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Finishing Pigs ◽  
Lawsonia Intracellularis ◽  
Fecal Samples ◽  
Brachyspira Pilosicoli ◽  
Brachyspira Hyodysenteriae ◽  
Veterinary Research ◽  
Two Samples ◽  
Intestinal Pathogens

Background: The major pathogenic intestinal spirochetes affecting pigs during the growing- finishing stage of production include Brachyspira hyodysenteriae and Brachyspira pilosicoli. Infections by these pathogens, which affect the economics of pig production, can result in mortality, growth rate losses and substantial antibiotic costs. The aim of this study was to assess the current occurrence of B. hyodysenteriae and B. pilosicoli in Polish pig herds. Moreover, associations between the presence of diarrhea or other intestinal pathogens and occurrence of B. hyodysenteriae and B. pilosicoli in pigs were investigated. Methods: Between January 2017 and August 2019, a total of 401 samples of pig feces from 95 different herds were submitted to the National Veterinary Research Institute of Poland. These samples were obtained from pigs older than 7 weeks. All the received fecal samples were examined for the presence of B. hyodysenteriae, B. pilosicoli and Lawsonia intracellularis by real-time PCR. Results: For B. pilosicoli, 4.5% (95% CI, 2.5–7.0%) of samples and 13.7% (95% CI, 7.5–22.3%) of herds were positive. Out of 12 samples, B. pilosicoli was detected simultaneously with L. intracellularis, B. hyodysenteriae and B. pilosicoli were detected alone in two samples each. In terms of B. hyodysenteriae, 7.0% of samples (95% CI, 4.7–9.9%) from 18.9% of herds (95% CI, 11.6–28.3%) were positive in real time PCR. The presence of B. hyodysenteriae in fecal samples was associated with the presence of diarrhea in pigs. Conclusions: This study confirmed that B. pilosicoli infections occur in Polish pig herds, but the prevalence is at a low level and the presence of B. pilosicoli is not associated with the development of diarrhea in pigs. B. hyodysenteriae is still a common cause of diarrhea among pigs from Polish herds.

scholarly journals Cryptosporidium spp. during chemotherapy: a cross-sectional study of 94 patients with malignant solid tumor

2021 ◽  
Vol 41 (5) ◽  
pp. 293-298
Author(s):  
Mehmet Karabey ◽  
Hüseyin Can ◽  
Tülay Öncü Öner ◽  
Mert Döşkaya ◽  
Sedef Erkunt Alak ◽  
...  
Keyword(s):  
Sample Size ◽  
Real Time ◽  
Solid Tumors ◽  
Real Time Pcr ◽  
Tertiary Care ◽  
Diagnostic Methods ◽  
Cross Sectional ◽  
Cryptosporidium Spp ◽  
Stool Samples ◽  
Pcr Targeting

BACKGROUND: Cryptosporidium spp . is a protozoan parasite that infects many vertebrate animals, including humans. Since Cryptosporidium spp . can cause chronic life-threatening diarrhea and severe malabsorption in immunocompromised patients, we investigated the prevalence of this parasite among patients undergoing chemotherapy for malignant solid tumors. OBJECTIVE: Investigate the prevalence of Cryptosporidium spp . in stool samples. DESIGN: Cross-sectional. SETTING: Tertiary care. PATIENTS AND METHODS: Stool samples were collected from adult patients with malignant solid tumors receiving chemotherapy and diarrhea. Cryptosporidium spp . prevalence was determined using Ziehl–Neelsen staining, ELISA, and real-time PCR targeting of the COWP gene. MAIN OUTCOME MEASURE: The prevalence of Cryptosporidium spp . in patients undergoing chemotherapy for malignant solid tumors. SAMPLE SIZE: 94 RESULTS: The prevalence was 2.1% (2/94), 5.3% (5/94), and 5.3% (5/94) as detected by Ziehl–Neelsen staining, real-time PCR and ELISA, respectively. The prevalence reached 8.5% (8/94) using all results obtained from the three methods. Among eight positive stool samples, four were positive by at least two different methods (Ziehl–Neelsen staining-ELISA or ELISA-real-time PCR) whereas the remaining four were positive by either ELISA or real-time PCR. CONCLUSION: These findings show the risk of cryptosporidiosis in cancer patients and the necessity to use at least two diagnostic methods during the diagnosis of cryptosporidiosis to reach more accurate and trustworthy results. LIMITATIONS: Further studies with a larger sample size are recommended. CONFLICT OF INTEREST: None.

scholarly journals Development of Conventional Multiplex PCR: A Rapid Technique for Simultaneous Detection of Soil-Transmitted Helminths

Pathogens ◽  
2019 ◽  
Vol 8 (3) ◽  
pp. 152 ◽  
Author(s):  
Vivornpun Sanprasert ◽  
Ruthairat Kerdkaew ◽  
Siriporn Srirungruang ◽  
Sarit Charuchaibovorn ◽  
Kobpat Phadungsaksawasdi ◽  
...  
Keyword(s):  
Multiplex Pcr ◽  
Real Time ◽  
Real Time Pcr ◽  
Intestinal Parasites ◽  
Simultaneous Detection ◽  
Diagnostic Tools ◽  
Multiple Infections ◽  
Parasitic Infections ◽  
Soil Transmitted Helminths ◽  
Stool Samples

Soil-transmitted helminths (STHs) are the most common intestinal parasites infecting humans worldwide. STH infections are a major cause of morbidity and disability. Accurate diagnostic tools are pivotal for assessing the exact prevalence of parasitic infections. Microscopic examination and culture techniques have been used to observe the presence of eggs or larvae of parasites in stool samples, but they are time-consuming and have low sensitivity. Therefore, accurate, simple, and inexpensive diagnostic techniques are still required for simultaneous detection of STH infections. Although molecular-based techniques, such as real-time PCR and multiplex real-time PCR, have been developed, they are not suitable for routine diagnosis due to the requirement for expensive reagents and instruments. In this study, we established a conventional multiplex PCR for simultaneous rapid detection of Ascaris lumbricoides, Necator americanus, and Strongyloides stercoralis in stool samples. Our results show that the multiplex PCR could detect the DNA of STHs at a very low target gene concentrations (lower than 1 pg) with no cross-amplification. Multiplex PCR had five times higher sensitivity than the formalin–ethyl acetate concentration technique (FECT) in the detection of multiple infections, and two times higher for detection of S. stercoralis. However, multiplex PCR was comparable to FECT in the detection of A. lumbricoides and N. americanus. In conclusion, this method could be used as an alternative method for the detection of STHs, especially for S. stercoralis.

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