Diagnosis of intestinal protozoan infections in patients in Cuba by microscopy and molecular methods: advantages and disadvantages

Puebla, Luis Enrique Jerez; Núñez, Fidel A.; Nodarse, Jorge Fraga; Millán, Iraís Atencio; Rodríguez, Iredys Cruz; Silva, Isabel Martínez; Valdés, Lucía Ayllón; Robertson, Lucy J.

doi:10.1016/j.mimet.2020.106102

Cited by 18 publications

(21 citation statements)

References 35 publications

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“…and D . fragilis have clearly demonstrated that molecular screening methods are needed in accurately detecting the presence of these protists in stool samples [ 12 , 56 , 57 , 62 , 63 ]. The use of molecular methods to improve detection of Blastocystis sp.…”

Section: Introductionmentioning

confidence: 99%

Investigation of neglected protists Blastocystis sp. and Dientamoeba fragilis in immunocompetent and immunodeficient diarrheal patients using both conventional and molecular methods

et al. 2021

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Introduction The clinical significance of Blastocystis sp. and Dientamoeba fragilis in patients with gastrointestinal symptoms is a controversial issue. Since the pathogenicity of these protists has not been fully elucidated, testing for these organisms is not routinely pursued by most laboratories and clinicians. Thus, the prevalence of these organisms and the subtypes of Blastocystis sp. in human patients in Turkey are not well characterized. This study aimed to determine the prevalence of Blastocystis sp. and D. fragilis in the diarrheic stool samples of immunodeficient and immunocompetent patients using conventional and molecular methods and to identify Blastocystis sp. subtypes using next generation sequencing. Material and methods Individual stool specimens were collected from 245 immunodeficient and 193 immunocompetent diarrheic patients between March 2017 and December 2019 at the Gazi University Training and Research Hospital in Ankara, Turkey. Samples were screened for Blastocystis sp. and D. fragilis by conventional and molecular methods. Molecular detection of both protists was achieved by separate qPCRs targeting a partial fragment of the SSU rRNA gene. Next generation sequencing was used to identify Blastocystis sp. subtypes. Results The prevalence of Blastocystis sp. and D. fragilis was 16.7% and 11.9%, respectively as measured by qPCR. The prevalence of Blastocystis sp. and D. fragilis was lower in immunodeficient patients (12.7% and 10.6%, respectively) compared to immunocompetent patients (21.8% and 13.5%, respectively). Five Blastocystis sp. subtypes were identified and the following subtype distribution was observed: ST3 54.4% (n = 37), ST2 16.2% (n = 11), ST1 4.4% (n = 3), ST6 2.9% (n = 2), ST4 1.5% (n = 1), ST2/ST3 11.8% (n = 8) and ST1/ST3 8.8% (n = 6). There was no statistically significant difference in the distribution of Blastocystis sp. subtypes between immunocompetent and immunodeficient patients. Conclusion and recommendation Our findings demonstrated that Blastocystis sp. and D. fragilis are commonly present in immunocompetent and immunodeficient patients with diarrhea. This study is the first to use next generation sequencing to address the presence of Blastocystis sp. mixed subtypes and intra-subtype variability in clinical samples in Turkey.

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Section: Introductionmentioning

confidence: 99%

Investigation of neglected protists Blastocystis sp. and Dientamoeba fragilis in immunocompetent and immunodeficient diarrheal patients using both conventional and molecular methods

et al. 2021

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“…In low-income regions, surveillance of D. fragilis is probably a low priority, so that sensitive techniques in routine diagnostic laboratories are scarcely used, contributing to a largely incomplete epidemiological picture. Few studies on D. fragilis prevalence have been published in Latin America, and to date, investigations were performed in Argentina 13 Mexico 14 , Venezuela 15 , Cuba 16 and Brazil 17 – 20 . Infection rates of <2% to 40% were found, which may be explained by differences in the surveyed populations, geographic locations and diagnostic methods 2 – 4 .…”

Section: Discussionmentioning

confidence: 99%

“…Recently, surveys based on microscopy and/or molecular assays have been carried out in low-resource communities in Asia 10 , Middle East 11 and Africa 12 . In Latin America, information is still limited 13 – 16 , including in Brazil, where there have been only four studies on this subject 17 – 20 . Among these studies, two focused on HIV patients 17 , 18 , one on dwellers of fisher villages 19 and the other on subjects referred to a clinical laboratory 20 .…”

Section: Introductionmentioning

confidence: 99%

Prevalence and genetic characterization of Dientamoeba fragilis in asymptomatic children attending daycare centers

Oliveira-Arbex

David

Cacciò

et al. 2021

Rev. Inst. Med. trop. S. Paulo

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In order to provide additional data on the prevalence and genetic diversity of Dientamoeba fragilis in human populations, we conducted a study in children from low-income communities in Sao Paulo State, Brazil. Fecal samples from daycare center attendees up to 6 years old (n=156) and staff members (n=18) were submitted to PCR and sequencing of D. fragilis as well as to microscopic examination for the presence of other intestinal parasites. All children assessed were asymptomatic and 10.3% (16/156) were positive for D. fragilis . No worker was found to be positive. An association between Dientamoeba and coinfection with other intestinal parasites was observed. Concerning the genetic diversity, 14 and only two isolates were genotype 1 and genotype 2, respectively. Our findings outline interesting aspects: (1) asymptomatic children as carriers of Dientamoeba in communities in which environmental conditions ensure parasite transmission and, (2) association between Dientamoeba infection in young children and coinfection with other enteric parasites, reinforcing its transmission via the fecal–oral route.

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“…Published data from the French CNR-LE for cryptosporidiosis show that: i) even with around 250 notified cases each year, cryptosporidiosis is still largely underestimated in France, ii) cryptosporidiosis is predominant in immunocompetent individuals and especially in young children and young adults, and iii) cryptosporidiosis is over-represented in the summer [ 5 , 6 ]. The routine diagnosis of cryptosporidiosis still relies on light microscopy examination for many laboratories [ 7 , 8 , 9 , 10 ]. However, light microscopy examination lacks sensitivity, is time-consuming, and requires skilled technicians, making it an inefficient method for laboratories which are able to switch to PCR analysis [ 8 , 10 , 11 , 12 ].…”

Section: Introductionmentioning

confidence: 99%

“…The routine diagnosis of cryptosporidiosis still relies on light microscopy examination for many laboratories [ 7 , 8 , 9 , 10 ]. However, light microscopy examination lacks sensitivity, is time-consuming, and requires skilled technicians, making it an inefficient method for laboratories which are able to switch to PCR analysis [ 8 , 10 , 11 , 12 ]. Currently, several PCR methods are available to screen Cryptosporidium spp.…”

Section: Introductionmentioning

confidence: 99%

Comparative Performance of Eight PCR Methods to Detect Cryptosporidium Species

et al. 2021

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Diagnostic approaches based on PCR methods are increasingly used in the field of parasitology, particularly to detect Cryptosporidium. Consequently, many different PCR methods are available, both “in-house” and commercial methods. The aim of this study was to compare the performance of eight PCR methods, four “in-house” and four commercial methods, to detect Cryptosporidium species. On the same DNA extracts, performance was evaluated regarding the limit of detection for both C. parvum and C. hominis specificity and the ability to detect rare species implicated in human infection. Results showed variations in terms of performance. The best performance was observed with the FTD® Stool parasites method, which detected C. parvum and C. hominis with a limit of detection of 1 and 10 oocysts/gram of stool respectively; all rare species tested were detected (C. cuniculus, C. meleagridis, C. felis, C. chipmunk, and C. ubiquitum), and no cross-reaction was observed. In addition, no cross-reactivity was observed with other enteric pathogens. However, commercial methods were unable to differentiate Cryptosporidium species, and generally, we recommend testing each DNA extract in at least triplicate to optimize the limit of detection.

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Diagnosis of intestinal protozoan infections in patients in Cuba by microscopy and molecular methods: advantages and disadvantages

Cited by 18 publications

References 35 publications

Investigation of neglected protists Blastocystis sp. and Dientamoeba fragilis in immunocompetent and immunodeficient diarrheal patients using both conventional and molecular methods

Investigation of neglected protists Blastocystis sp. and Dientamoeba fragilis in immunocompetent and immunodeficient diarrheal patients using both conventional and molecular methods

Prevalence and genetic characterization of Dientamoeba fragilis in asymptomatic children attending daycare centers

Comparative Performance of Eight PCR Methods to Detect Cryptosporidium Species

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