Simultaneous detection of four protozoan parasites on leafy greens using a novel multiplex PCR assay

Shapiro, Karen; Kim, Minji; Rajal, Verónica Beatriz; Arrowood, Michael J.; Packham, Andrea E.; Aguilar, Beatriz; Wuertz, Stefan

doi:10.1016/j.fm.2019.103252

Cited by 28 publications

(32 citation statements)

References 41 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In recent years, several molecular techniques have been developed for the detection of C. cayetanensis in fresh produce, including qPCR and various multiplex qPCR methods (Steele et al, 2003;Gajadhar, 2011, 2016;Murphy et al, 2017;Shapiro et al, 2019) (Table 1). qPCR can steadily identify a few oocysts in the fresh produce, such as three oocysts in a gram of fruit, or five oocysts in a gram of herbs or green onions (Lalonde and Gajadhar, 2016).…”

Section: Fresh Producesmentioning

confidence: 99%

“…Another qPCR assay targeting the internal transcribed spacer 1 region was developed for the detection of C. cayetanensis in berries, and proved to be an effective approach that may be a suitable option for use in food-testing laboratories (Temesgen et al, 2019b). A multiplex PCR assay has been developed for simultaneous detection of four protozoan oocysts via a rapid, inexpensive, and simple protocol (Shapiro et al, 2019). Robertson et al (2000) validated the use of lectin-coated paramagnetic beads for the isolation of Cyclospora oocysts from fruits and vegetables.…”

Section: Fresh Producesmentioning

confidence: 99%

See 1 more Smart Citation

Advances in Cyclosporiasis Diagnosis and Therapeutic Intervention

Cui

et al. 2020

Front. Cell. Infect. Microbiol.

View full text Add to dashboard Cite

Cyclosporiasis is caused by the coccidian parasite Cyclospora cayetanensis and is associated with large and complex food-borne outbreaks worldwide. Associated symptoms include severe watery diarrhea, particularly in infants, and immune dysfunction. With the globalization of human food supply, the occurrence of cyclosporiasis has been increasing in both food growing and importing countries. As well as being a burden on the health of individual humans, cyclosporiasis is a global public health concern. Currently, no vaccine is available but early detection and treatment could result in a favorable clinical outcome. Clinical diagnosis is based on cardinal clinical symptoms and conventional laboratory methods, which usually involve microscopic examination of wet smears, staining tests, fluorescence microscopy, serological testing, or DNA testing for oocysts in the stool. Detection in the vehicle of infection, which can be fresh produce, water, or soil is helpful for case-linkage and source-tracking during cyclosporiasis outbreaks. Treatment with trimethoprim-sulfamethoxazole (TMP-SMX) can evidently cure C. cayetanensis infection. However, TMP-SMX is not suitable for patients having sulfonamide intolerance. In such case ciprofloxacin, although less effective than TMP-SMX, is a good option. Another drug of choice is nitazoxanide that can be used in the cases of sulfonamide intolerance and ciprofloxacin resistance. More epidemiological research investigating cyclosporiasis in humans should be conducted worldwide, to achieve a better understanding of its characteristics in this regard. It is also necessary to establish in vitro and/or in vivo protocols for cultivating C. cayetanensis, to facilitate the development of rapid, convenient, precise, and economical detection methods for diagnosis, as well as more effective tracing methods. This review focuses on the advances in clinical features, diagnosis, and therapeutic intervention of cyclosporiasis.

show abstract

Section: Fresh Producesmentioning

confidence: 99%

Section: Fresh Producesmentioning

confidence: 99%

Advances in Cyclosporiasis Diagnosis and Therapeutic Intervention

Cui

et al. 2020

Front. Cell. Infect. Microbiol.

View full text Add to dashboard Cite

show abstract

“…Otherwise, Shields et al (2012) 22 compared various buffers as deionized water, 1 M Glycine, pH 5.5, a detachment solution, and 0.1% Alconox®, to recover C. parvum and Cyclospora cayetanensis from lettuce, herbs and raspberries; there was no significant difference between the deionized water and 1 M glycine, pH 5.5, while Alconox gave the best recovery rate. On the other hand, Shapiro et al (2019) 23 described a method for simultaneous detection of C. parvum, G. duodenalis, T. gondii and C. cayetanensis. They found that leaf-washing followed by a multiplex-PCR allowed higher recoveries and more consistent detection of parasites compared to stomaching processing.…”

Section: Toxoplasma Oocysts Detectionmentioning

confidence: 99%

Cryptosporidium spp., Giardia duodenalis and Toxoplasma gondii detection in fresh vegetables consumed in Marrakech, Morocco

Berrouch¹,

Escotte-Binet²,

Amraouza³

et al. 2020

Afr H. Sci.

View full text Add to dashboard Cite

Background: Protozoan parasites such as Toxoplasma gondii, Giardia duodenalis, and Cryptosporidium spp., can be transmitted to humans via accidental consumption of contaminated water, fresh produce and foodstuffs. There is a lack of epidemiological data about these pathogens in Morocco. Hence the aim of this study, which is the determination of their prevalence in some leafy greens and root vegetables sold in Marrakech. Methods: A total of 132 vegetable samples including carrot, coriander, lettuce, parsley and radish were purchased monthly from three different markets in Marrakech from March 2017 to January 2018, pre-treated and subjected to microscopic and molecular analyses. Results: Of the 132 samples of vegetables analyzed by qPCR, the overall rate of protozoan was 21.21% (28/132); 22 samples were found to be contaminated with T. gondii, 6 with G. duodenalis, and none was positive for C. parvum/hominis. Whereas, modified Ziehl-Neelsen staining allowed the detection of Cryptosporidium spp. in 3% (4/132) of examined samples. Conclusion: This survey on the presence of protozoan parasites in fresh vegetables revealed that vegetables sold in Marrakech are contaminated by these protozoan parasites, as it showed that leafy green vegetables were more susceptible for parasitic contamination than root ones. Keywords: Fresh vegetables; protozoan parasites; Marrakech; qPCR.

show abstract

“…However, this technique is time-consuming, non-specific, and lacks 2 of 11 sensitivity [9]. To overcome these issues, molecular methods have been developed to detect C. cayetanensis in clinical and environmental samples [10][11][12]; however, the food producing industry requires a molecular method able to detect a low oocyst concentration (40-1500 oocyst per gram) as found in food and environmental samples [13]. Also, it is important to have a tool available to perform molecular traceability and identification of contamination sources.…”

Section: Introductionmentioning

confidence: 99%

A Molecular Tool for Rapid Detection and Traceability of Cyclospora cayetanensis in Fresh Berries and Berry Farm Soils

Reséndiz-Nava

Orozco-Mosqueda

Mercado-Silva

et al. 2020

Foods

View full text Add to dashboard Cite

Due to recent outbreaks of cyclosporiasis associated with consumption of fresh berries, producers are demanding modern microbiological tools for the rapid and accurate identification of the human pathogen Cyclospora cayetanensis in berries and environmental samples. The aim of the present work was to develop a molecular tool based on a PCR approach for the rapid and accurate detection of C. cayetanensis. A nested PCR assay was validated for the amplification of a 294 bp size region of the 18S rRNA gene from C. cayetanensis. The limit of detection for the nested PCR assay was validated using 48 berry samples spiked with ~0, 10, 100, and 1000 oocyst per gram of sample. With this assay, it was possible to detect as few as 1 oocyst per gram of berry, in a 50 g sample. Sanger DNA sequencing and phylogenetic analysis were carried out to confirm the presence of C. cayetanensis in berry (n = 17) and soil (n = 5) samples. The phylogenetic analysis revealed that the C. cayetanensis sequences obtained from Mexico clustered within a group recovered from China, Peru, Guatemala-Haiti, and Japan. The PCR protocol designed in the present study could be an important tool for the rapid and accurate detection of this human pathogen in environmental and food samples.

show abstract

Simultaneous detection of four protozoan parasites on leafy greens using a novel multiplex PCR assay

Cited by 28 publications

References 41 publications

Advances in Cyclosporiasis Diagnosis and Therapeutic Intervention

Advances in Cyclosporiasis Diagnosis and Therapeutic Intervention

Cryptosporidium spp., Giardia duodenalis and Toxoplasma gondii detection in fresh vegetables consumed in Marrakech, Morocco

A Molecular Tool for Rapid Detection and Traceability of Cyclospora cayetanensis in Fresh Berries and Berry Farm Soils

Contact Info

Product

Resources

About