Genetic Characterization of Toxoplasma gondii DNA Samples Isolated From Humans Living in North America: An Unexpected High Prevalence of Atypical Genotypes

Pomarès, Christelle; Devillard, Sébastien; Holmes, Tyson H.; Olariu, Tudor Rareș; Press, Cynthia; Ramirez, Raymund; Talucod, Jeanne; Estran, Remy; Su, Chunlei; Dubey, J. P.; Ajzenberg, Daniel; Montoya, José G.

doi:10.1093/infdis/jiy375

Cited by 42 publications

(21 citation statements)

References 42 publications

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“…In a recent retrospective study on 76 clinical samples, the relative sensitivity of B1 qPCR was shown to be only 54% of that of Rep 529 qPCR (4). However, whereas there is high genetic homogeneity of strains circulating in Europe, particularly in France, much greater heterogeneity has been recently highlighted in the United States, but less than in South America (26). The sensitivity of the Rep 529 target was questioned for non-type II T. gondii isolates.…”

Section: Discussionmentioning

confidence: 99%

“…Selected samples were sent to laboratory 1 as part of routine diagnosis and treatment with no unnecessary invasive procedures. Genotyping data obtained by microsatellite analysis were available for 34 samples from a previous study (26). Thirteen isolates belonged to type II, 5 to type III, and 16 to atypical lineages.…”

Section: Methodsmentioning

confidence: 99%

“…In contrast, in South America, T. gondii strains show high genetic diversity, with mainly atypical strains (20)(21)(22)(23). Recently, an unexpectedly high prevalence of non-type II strains has been highlighted in the United States (24)(25)(26). Therefore, it is very important to determine the performance of Rep 529 PCR for the diagnosis of non-type II strains.…”

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confidence: 99%

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Is Real-Time PCR Targeting Rep 529 Suitable for Diagnosis of Toxoplasmosis in Patients Infected with Non-Type II Strains in North America?

Pomarès

Estran

Press³

et al. 2020

J Clin Microbiol

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Toxoplasma gondii DNA detection is essential to antenatally diagnose a congenital infection and reactivation of a past infection in an immunocompromised patient. Initially, PCR methods targeted the 35-fold repetitive B1 gene, and more recently, coding sequence Rep 529 has been preferred, as it was reported to be repeated 200- to 300-fold and yielded far better sensitivity than amplification of the B1 sequence. To date, few data are available in regard to the efficacy of Rep 529 for non-type II genotypes. In this study, we compared the results of B1 quantitative PCR (qPCR) with those of two different Rep 529 qPCRs performed on 111 samples in two different laboratories (Rep 529-1 and Rep 529-2). The performances of the 3 qPCRs were also compared according to the genotypes of the isolates for 13 type II and 21 non-type II samples. The performance of the Rep 529 target was superior to that of the B1 target regardless of the genotype (threshold cycle [CT] values for the Rep 529-1 and Rep 529-2 qPCRs were lower than those for the B1 qPCR [P < 0.001 and P < 0.01, respectively]). The same results were observed when a comparison was made according to the genotype of the strain (type II and non-type II genotypes). To our knowledge, these results provide the first relative quantitative data revealing that the efficiency of Rep 529 qPCR does not depend on the genotype of T. gondii isolates and that, in fact, it is superior to B1 qPCR.

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

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Is Real-Time PCR Targeting Rep 529 Suitable for Diagnosis of Toxoplasmosis in Patients Infected with Non-Type II Strains in North America?

Pomarès

Estran

Press³

et al. 2020

J Clin Microbiol

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“…Most infections in humans are asymptomatic. However, severe complications may occur during (i) congenital Toxoplasma infection, such as abortion, stillbirth and hydrocephalus in new-borns [1,4,5]; (ii) ocular toxoplasmosis, with retinochoroidal lesions leading to chronic ocular disease [1,2]; (iii) encephalitis in immunosuppressed patients [1,4,5]; and (iv) multivisceral toxoplasmosis due to atypical genotypes in South America [6][7][8]. A possible implication of T. gondii genetic diversity on the pathogenesis of toxoplasmosis has been postulated [9][10][11].…”

Section: Introductionmentioning

confidence: 99%

Detection of Toxoplasma gondii oocysts in fresh vegetables and berry fruits

et al. 2020

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Background: Toxoplasma gondii is the third most important contributor to health burden caused by food-borne illness. Ingestion of tissue cysts from undercooked meat is an important source of horizontal transmission to humans. However, there is an increasing awareness of the consumption of fresh fruit and vegetables, as a possible source for oocyst transmission, since this stage of the parasite can persist and remain infective in soil and water for long time. Herein, we outline findings related with detection of T. gondii oocysts in vegetables and berry fruits, which are usually raw consumed. The procedure includes the estimation of the number of oocysts. Methods: Food samples were collected from local producers and supermarket suppliers. Toxoplasma gondii oocysts were concentrated after washing the samples by applying high resolution water filtration and immunomagnetic separation (method 1623.1: EPA 816-R-12-001-Jan 2012), in order to (i) remove potential Cryptosporidium spp. oocysts and Giardia spp. cysts present in the samples; and (ii) select T. gondii oocysts. Toxoplasma gondii oocyst detection and an estimation of their numbers was performed by conventional PCR and real time qPCR, using specific primers for a 183-bp sequence of the T. gondii repetitive DNA region. All PCR-positive DNA samples were purified and sequenced. Restriction enzyme digestion with EcoRV endonuclease confirmed the presence of the T. gondii DNA fragment. In addition, the presence of the parasite was observed by fluorescent microscopy, taking advantage of the oocysts autofluorescence under UV light. Results: Forty percent of the analysed samples (95% CI: 25.5-56.5%) presented the expected PCR and digested DNA fragments. These fragments were confirmed by sequencing. Microscopic autofluorescence supported the presence of T. gondii-like oocysts. The estimated mean (± SE) oocyst concentration was 23.5 ± 12.1 oocysts/g, with a range of 0.6-179.9 oocysts/g. Conclusions: Our findings provide relevant evidence of contamination of fresh vegetables and berry fruits with T. gondii oocysts.

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“…Supplemental feeding of cats may alter T. gondii transmission by changing cat distribution, demographics and exposure to infected prey (Schmidt et al, ; VanWormer et al, ). Furthermore, free‐roaming cats in central California, USA are known reservoirs of atypical T. gondii genotypes (e.g., Type X) (VanWormer et al, ), which are associated with more severe disease in people (Blaizot et al, ; Carme et al, ; Pomares et al, ). Understanding the risks of exposure to T. gondii from contaminated soil will help inform management of free‐roaming cat colonies and assess the risk of human exposure to this zoonotic pathogen.…”

Section: Introductionmentioning

confidence: 99%

Seasonal and spatial variation in Toxoplasma gondii contamination in soil in urban public spaces in California, United States

Wit

Kilpatrick

VanWormer

et al. 2019

Zoonoses and Public Health

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Background: Toxoplasma gondii is a zoonotic parasite that can have severe implications for human health. Acutely infected cats shed environmentally resistant T. gondii oocysts in their faeces that contaminate soil, and soil can serve as a reservoir of infection for humans. Free-roaming domestic cats are thought to play an important role in environmental contamination with T. gondii, but few studies have directly measured the direct contribution of free-roaming cats to T. gondii in soil. Methods:Our goals were to determine whether T. gondii soil contamination occurs in public areas with free-roaming cat colonies in central California and examine spatial and temporal variation in soil contamination. We initially performed spiking experiments to compare the limit of T. gondii detection in soil using three conventional nested PCR assays and one real-time quantitative PCR. The nested PCR targeting the internal transcribed spacer (ITS-1) of the small subunit ribosomal RNA was the most sensitive assay, with a limit of detection between 20 and 200 oocysts per gram of soil. We applied the ITS1 PCR assay on soil from sites in city and state parks, public playgrounds and community gardens in central California, USA. Samples were collected during spring, summer and fall and in sites located along the coast and inland. Results:We detected and sequence-confirmed T. gondii in 5.6% of all of our soil subsamples, but with large seasonal and spatial variation in soil contamination: we only detected T. gondii during fall and only in coastal sites (44.3% soil prevalence), despite similar sampling intensity across space and time. Conclusions:Our results suggest that free-roaming cat colonies are an important source of T. gondii in spaces where people recreate and grow food and that soil contamination is highly seasonal and spatially variable. Management of free-roaming cats could prevent T. gondii infections by reducing environmental contamination with this zoonotic pathogen. K E Y W O R D Sfree-roaming cats, management, molecular detection, public health, sea otters | 71 de WIT eT al.

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Genetic Characterization of Toxoplasma gondii DNA Samples Isolated From Humans Living in North America: An Unexpected High Prevalence of Atypical Genotypes

Abstract: Clinicians should be aware that atypical genotypes are common in North America and have been associated with severe ocular and systemic disease and unusual presentations of toxoplasmosis in immunocompetent patients.

Cited by 42 publications

References 42 publications

Is Real-Time PCR Targeting Rep 529 Suitable for Diagnosis of Toxoplasmosis in Patients Infected with Non-Type II Strains in North America?

Is Real-Time PCR Targeting Rep 529 Suitable for Diagnosis of Toxoplasmosis in Patients Infected with Non-Type II Strains in North America?

Detection of Toxoplasma gondii oocysts in fresh vegetables and berry fruits

Seasonal and spatial variation in Toxoplasma gondii contamination in soil in urban public spaces in California, United States

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