scholarly journals Comparative aspects of laboratory testing for the detection of Toxoplasma gondii and its differentiation from Neospora caninum as the etiologic agent of ovine abortion

2020 ◽  
Vol 32 (6) ◽  
pp. 898-907
Author(s):  
Nicola Meixner ◽  
Marie F. Sommer ◽  
Nelly Scuda ◽  
Kaspar Matiasek ◽  
Matthias Müller
Keyword(s):  
Toxoplasma Gondii ◽  
Real Time ◽  
Real Time Pcr ◽  
Neospora Caninum ◽  
Placental Tissue ◽  
Etiologic Agent ◽  
Detection Methods ◽  
Tissue Samples ◽  
Specificity And Sensitivity ◽  
Histologic Evaluation

Histologic examination of aborted material is an essential component in the diagnosis of ovine toxoplasmosis. However, the detection of Toxoplasma gondii in histologic sections, and its differentiation from the closely related protozoan Neospora caninum, is challenging. We developed a chromogenic in situ hybridization (ISH) assay for the identification of T. gondii in paraffin-embedded tissue samples. We examined retrospectively the archived placental tissue of 200 sheep abortion submissions for the presence of T. gondii by immunohistochemistry (IHC), ISH, and real-time PCR (rtPCR). All placental samples that tested positive for T. gondii by rtPCR (9 of 200) were also positive by IHC, with inconclusive IHC staining in an additional 7 rtPCR-negative cases. Further testing for N. caninum of all 200 placentas by rtPCR revealed 7 Neospora-positive cases. T. gondii ISH was positive in 4 of 9 IHC-positive samples and 1 of the 7 N. caninum rtPCR-positive samples. Real-time PCR was used as the reference standard for specificity and sensitivity calculations regarding placenta samples. Specificity of ISH and IHC was 99% and 96–100%, respectively. The sensitivity of ISH (44%) was quite low compared to IHC (100%). The exclusive use of ISH for the detection of T. gondii, and thus for the diagnosis of ovine toxoplasmosis, was not acceptable. However, combined with rtPCR, both ISH and IHC can be useful detection methods to improve histologic evaluation by visualizing the parasite within tissue sections.

scholarly journals Ability of Real-time PCR in diagnose Differentiation Various Forms of Cutaneous Leishmaniasis: A Comparative Study with Histopathology

2019 ◽  
Author(s):  
Maryam Fekri Soofi Abadi ◽  
Meisam Fekri ◽  
alireza moradabadi ◽  
Reza Vahidi ◽  
Simin Shamsi Meymandi ◽  
...  
Keyword(s):  
Cutaneous Leishmaniasis ◽  
Real Time ◽  
Real Time Pcr ◽  
Relative Number ◽  
Parasite Load ◽  
Detection Methods ◽  
Tissue Samples ◽  
Microscopic Evaluation ◽  
Histopathological Evaluation ◽  
Histopathological Studies

Abstract objective: Histopathological studies suggest that parasite load is different between acute and chronic forms of cutaneous leishmaniasis (CL). However, highly sensitive detection methods are still needed to distinguish different forms of leishmaniasis. In the present study, we developed a quantitative real-time polymerase chain reaction (PCR) to detect and quantify leishmania tropica parasites in paraffin-embedded tissue samples. Results: The ability of real-time PCR for leishmania detection was higher than histopathological evaluation. The parasite loads were quantified by qPCR assay and microscopic evaluation were highly correlated ( r =0.598; P <0.001). Among patients, the parasite load was inversely correlated with disease duration (acute CL lesions had very higher parasite loads than chronic CL lesions), but there was no difference in parasite load according to the patients’ age and sex as well as location of the lesions. In contrast to Ridley scoring system (P<0.001), there were no statistically significant differences in the relative number of parasites among the lupoid and non-lupoid forms of chronic lesions in real-time PCR (P=0.549), which indicates the superiority of histopathological evaluation in CL forms differentiation.

scholarly journals Examination of Mycobacterium avium subsp. avium distribution in naturally infected hens by culture and triplex quantitative real time PCR

2010 ◽  
Vol 55 (No. 7) ◽  
pp. 325-330 ◽  
Author(s):  
M. Kaevska ◽  
I. Slana ◽  
P. Kralik ◽  
I. Pavlik
Keyword(s):  
Real Time ◽  
Mycobacterium Avium ◽  
Real Time Pcr ◽  
Clinical Symptoms ◽  
Bird Species ◽  
Etiologic Agent ◽  
Mycobacterium Avium Subsp ◽  
Quantitative Real Time Pcr ◽  
Tissue Samples ◽  
Conventional Culture

Mycobacterium avium subsp. avium (MAA) is the etiologic agent of avian tuberculosis, a chronic contagious disease described in a wide variety of domestic and wild bird species. The aims of this study were to assess the advantages of triplex quantitative real time PCR (qPCR) in comparison with culture testing for distribution of MAA in the organs of hens displaying varying degrees of clinical symptoms of the disease. From one small flock of ten hens and one cock with a history of weight loss, 98 tissue samples were examined in total. Pathological lesions were observed in six hens from which two were clinically ill. A total of 12 samples were positive by culture and 16 were positive by IS901 and IS1245 qPCR, confirming MAA infection. In conclusion, qPCR was a faster and more reliable alternative method in comparison with conventional culture analysis. Due to the detection of MAA in the muscle tissue of one hen, consumption of under cooked meat originating from infected fowl could pose a threat to immunosuppressed individuals.

The first detection ofToxoplasma gondiiDNA in environmental air samples using gelatine filters, real-time PCR and loop-mediated isothermal (LAMP) assays: qualitative and quantitative analysis

Parasitology ◽  
2017 ◽  
Vol 144 (13) ◽  
pp. 1791-1801 ◽  
Author(s):  
ANNA LASS ◽  
BEATA SZOSTAKOWSKA ◽  
KRZYSZTOF KORZENIEWSKI ◽  
PANAGIOTIS KARANIS
Keyword(s):  
Toxoplasma Gondii ◽  
Real Time ◽  
Real Time Pcr ◽  
Microscopic Analysis ◽  
Detection Methods ◽  
Type I ◽  
Air Samples ◽  
Two Samples ◽  
Environmental Air ◽  
Tract Infections

SUMMARYToxoplasma gondiiinfections are acquired through the ingestion of oocysts present in the environment. However, there is no data about their occurrence in the air or about airborne transmission of these infections. In the present paper, we report on the identification ofT. gondiiusing rapid molecular detection methods, supported by microscopic analysis, in environmental air samples. A total of 71 samples were collected, using gelatine filters, from kitchen gardens, recreational areas and sandpits located in northern and north-eastern Poland. Material recovered from the filters was analysed using real-time PCR and loop-mediated isothermal assays targeting theT. gondiiB1 gene.Toxoplasma gondiiDNA was found in two samples, as confirmed by both molecular assays. Genotyping at the SAG2 locus showedToxoplasmaSAG2 type I. Moreover, the presence ofT. gondiioocysts was confirmed in one of the positive samples with the use of microscopy. The results showed thatT. gondiimay be present in environmental air samples and that respiratory tract infections may play a role in the high prevalence of toxoplasmosis in humans and animals. To the best of our knowledge, this is the first epidemiological evidence that oro-fecal and foodborne toxoplasmosis may be traceable to an airborne respiratory origin and that this may represent a new, previously unknown transmission route for this disease.

scholarly journals Serological and molecular detection of Neospora caninum and Toxoplasma gondii in human umbilical cord blood and placental tissue samples

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Pâmella Oliveira Duarte ◽  
Leandra Marla Oshiro ◽  
Namor Pinheiro Zimmermann ◽  
Bárbara Guimarães Csordas ◽  
Doroty Mesquita Dourado ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Cord Blood ◽  
Umbilical Cord ◽  
Umbilical Cord Blood ◽  
Molecular Detection ◽  
Neospora Caninum ◽  
Placental Tissue ◽  
Human Umbilical Cord Blood ◽  
Human Umbilical Cord ◽  
Tissue Samples

scholarly journals Real-time PCR-based quantification of Toxoplasma gondii in tissue samples of serologically positive outdoor chickens

2010 ◽  
Vol 105 (7) ◽  
pp. 935-937 ◽  
Author(s):  
Cleiton Paulo Aigner ◽  
Aristeu Vieira da Silva ◽  
Fabiano Sandrini ◽  
Paulo de Sá Osório ◽  
Lilian Poiares ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Real Time ◽  
Real Time Pcr ◽  
Tissue Samples

scholarly journals Ability of Real-time PCR for Differential Diagnosis of Various Forms of Cutaneous Leishmaniasis: A Comparative Study with Histopathology

2019 ◽  
Author(s):  
Maryam Fekri Soofi Abadi ◽  
Meisam Fekri ◽  
alireza moradabadi ◽  
Reza Vahidi ◽  
Simin Shamsi Meymandi ◽  
...  
Keyword(s):  
Cutaneous Leishmaniasis ◽  
Real Time ◽  
Real Time Pcr ◽  
Relative Number ◽  
Parasite Load ◽  
Detection Methods ◽  
Tissue Samples ◽  
Microscopic Evaluation ◽  
Histopathological Evaluation ◽  
Histopathological Studies

Abstract objective: Histopathological studies suggest that parasite load is different between acute and chronic forms of cutaneous leishmaniasis (CL). However, highly sensitive detection methods are still needed to distinguish different forms of leishmaniasis. In the present study, we developed a quantitative real-time polymerase chain reaction (PCR) to detect and quantify leishmania tropica parasites in paraffin-embedded tissue samples. Results: The ability of real-time PCR for leishmania detection was higher than histopathological evaluation. The parasite loads were quantified by qPCR assay and microscopic evaluation were highly correlated ( r =0.598; P <0.001). Among patients, the parasite load was inversely correlated with disease duration (acute CL lesions had very higher parasite loads than chronic CL lesions), but there was no difference in parasite load according to the patients’ age and sex as well as location of the lesions. In contrast to Ridley scoring system (P<0.001), there were no statistically significant differences in the relative number of parasites among the lupoid and non-lupoid forms of chronic lesions in real-time PCR (P=0.549), which indicates the superiority of histopathological evaluation in CL forms differentiation.

scholarly journals Ability of Real-time PCR for Differential Diagnosis of Various Forms of Cutaneous Leishmaniasis: A Comparative Study with Histopathology

2019 ◽  
Author(s):  
Maryam Fekri Soofi Abadi ◽  
Meisam Fekri ◽  
alireza moradabadi ◽  
Reza Vahidi ◽  
Simin Shamsi Meymandi ◽  
...  
Keyword(s):  
Cutaneous Leishmaniasis ◽  
Real Time ◽  
Real Time Pcr ◽  
Relative Number ◽  
Parasite Load ◽  
Detection Methods ◽  
Tissue Samples ◽  
Microscopic Evaluation ◽  
Histopathological Evaluation ◽  
Histopathological Studies

Abstract objective: Histopathological studies suggest that parasite load is different between acute and chronic forms of cutaneous leishmaniasis (CL). However, highly sensitive detection methods are still needed to distinguish different forms of leishmaniasis. In the present study, we developed a quantitative real-time polymerase chain reaction (PCR) to detect and quantify leishmania tropica parasites in paraffin-embedded tissue samples. Results: The ability of real-time PCR for leishmania detection was higher than histopathological evaluation. The parasite loads were quantified by qPCR assay and microscopic evaluation were highly correlated ( r =0.598; P <0.001). Among patients, the parasite load was inversely correlated with disease duration (acute CL lesions had very higher parasite loads than chronic CL lesions), but there was no difference in parasite load according to the patients’ age and sex as well as location of the lesions. In contrast to Ridley scoring system (P<0.001), there were no statistically significant differences in the relative number of parasites among the lupoid and non-lupoid forms of chronic lesions in real-time PCR (P=0.549), which indicates the superiority of histopathological evaluation in CL forms differentiation.

scholarly journals Validation of Nonnested and Real-Time PCR for Diagnosis of Sheep-Associated Malignant Catarrhal Fever in Clinical Samples

2007 ◽  
Vol 19 (4) ◽  
pp. 405-408 ◽  
Author(s):  
Donald L. Traul ◽  
Naomi S. Taus ◽  
J. Lindsay Oaks ◽  
Donal O' Toole ◽  
Fred R. Rurangirwa ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Nested Pcr ◽  
High Specificity ◽  
Clinical Samples ◽  
Malignant Catarrhal Fever ◽  
Fatal Disease ◽  
Tissue Samples ◽  
Specificity And Sensitivity ◽  
Experimentally Induced

Sheep-associated malignant catarrhal fever (SA-MCF), a frequently fatal disease primarily of certain ruminants, is caused by ovine herpesvirus 2 (OvHV-2). Molecular diagnosis of SA-MCF in affected animals has relied on detection of OvHV-2 DNA using a nested PCR, which has significant potential for amplicon contamination as a routine method in diagnostic laboratories. In this report, a nonnested and a previously developed real-time PCR were validated for detection of OvHV-2 DNA in samples from clinically affected animals. Three sets of blood or tissue samples were collected: 1) 97 samples from 97 naturally affected animals with evidence of clinical SA-MCF; 2) 200 samples from 8 animals with experimentally induced SA-MCF; and 3) 100 samples from 100 animals without any evidence of clinical SA-MCF. Among 97 positive samples defined by nested PCR from clinically affected animals, 95 (98%) were positive by nonnested PCR and 93 (96%) were positive by real-time PCR, respectively. One hundred percent of the samples from the animals with experimentally induced MCF were positive by real-time PCR, while 99% were positive by nonnested PCR. Neither nonnested PCR nor real-time PCR yielded a positive result on any of the 100 nested PCR-negative samples from animals without evidence of clinical MCF. The data confirmed that both nonnested and real-time PCR maintained high specificity and sensitivity for the detection of OvHV-2 DNA in clinical samples.

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