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Wyświetlanie 1-2 z 2
Tytuł:
Toxoplasma gondii infection in selected species of free-living animals in Poland
Autorzy:
Sroka, J.
Karamon, J.
Wójcik-Fatla, A.
Dutkiewicz, J.
Bilska-Zając, E.
Zając, V.
Piotrowska, W.
Cencek, T.
Powiązania:
https://bibliotekanauki.pl/articles/2085130.pdf
Data publikacji:
2019
Wydawca:
Instytut Medycyny Wsi
Tematy:
Polska
Toxoplasma gondii
nested PCR
genotyping
free-living animals
Opis:
Introduction and objective. Free-living animals can play an important role as a reservoir of Toxoplasma gondi;, however, data concerning this issue in Poland are still limited.The aim of study was to assess the occurrence of T. gondii infection by using molecular methods in free-living animals in selected regions of Poland. Materials and method. Tissues samples of 396 animals (foxes, muskrats, birds, martens, badgers, polecats, raccoons, minks, raccoon dogs, otters, small rodents and insectivores, and grass snakes were collected from various regions of Poland. After samples digestion, DNA was isolated using QIAmp DNA Mini Kit (Qiagen). DNA extraction from small rodents and insectivores samples was performed without digestion. Next, nested PCR (B1 gene) and, for a part of nested PCR positive amplicons, RFLP PCR, were performed according to the method by Grigg and Boothroyd (2001). The other part of nested PCR positive DNA isolates were genotyped using 5 genetic markers: SAG1, SAG2 (5’- and 3’), SAG3, BTUB and GRA6, based on the method by Dubey et al. (2006). These PCR products were sequenced and compared with the NCBI database using Blast. Results. In total, in 50 of the 396 examined animals DNA of T. gondii was detected (12.6%). The highest percentages of positive results in PCR was obtained in martens (40.9%) and badgers (38.5%), lower in birds (27.3%) and the lowest in foxes (7.4%). The RFLP and multilocus PCR analysis showed the dominance of T. gondii clonal type II (or II/III). Conclusions. The results of this study indicate the frequent T. gondii infection among free-living animals in Poland, especially martens and badgers, which may indirectly indicate that these animals contribute to the spread of the parasite in the sylvatic environment in Poland. The genotyping analysis showed the dominance of T. gondii clonal type II (or II/III).
Źródło:
Annals of Agricultural and Environmental Medicine; 2019, 26, 4; 656-660
1232-1966
Pojawia się w:
Annals of Agricultural and Environmental Medicine
Dostawca treści:
Biblioteka Nauki
Artykuł
Tytuł:
Optimization of flotation, DNA extraction and PCR methods for detection of Toxoplasma gondii oocysts in cat faeces
Autorzy:
Sroka, J.
Karamon, J.
Dutkiewicz, J.
Wójcik-Fatla, A.
Cencek, T.
Powiązania:
https://bibliotekanauki.pl/articles/2081971.pdf
Data publikacji:
2018
Wydawca:
Instytut Medycyny Wsi
Tematy:
flotation
Toxoplasma gondii
faeces
Real time PCR
nested PCR
cats
oocysts detection
Opis:
Introduction and objective. The aim of the study was to compare the effectiveness of selected oocysts concentration methods, DNA extraction protocols and PCR assays targeting the B1 gene, for the development of procedures which would be effective and useful in laboratory practice for the detection of T. gondii in faecal samples from cats. Materials and method. In order to compare the influence of the flotation fluids on microscopy and PCR detection of T. gondii, saturated solutions of saccharose, MgSO4, ZnSO4 and NaNO3 were used. To determine the sensitivity of PCR tests used: Real time PCR (RT) and nested PCR, water samples spiked with T. gondii tachyzoites and oocysts were tested. DNA was extracted using DNeasy Blood and Tissue Kit (Qiagen) (K1). The same PCR tests were used to assess the efficacy of T. gondii DNA detection in samples of cat faeces spiked with oocysts, using DNA extraction by a K1 set and a K2 set (QIamp DNA Stool Mini Kit, (Qiagen). Results. The initial results showed that NaNO3 was most useful as a flotation fluid due to the lack negative effect on the oocysts and amplification efficacy in PCR. The level of detection for water samples (100 μl) was determined as 100 tachyzoites and 1–50 oocysts in RT, and 2–20 oocysts in nested PCR. The limit of detection (LD) for stool samples (250 mg) spiked with oocysts, where the K1 set was used, determined as 250 and 5 oocysts in RT and nested PCR, respectively. For samples extracted with the K2 set, LD in RT was determined as 1–50 oocysts (depending on the variant) and 50 oocysts in nested PCR. Conclusions. The most effective methods for detection of T. gondii in cat faeces seem to be centrifugal flotation with NaNO3, followed by DNA extraction with removing of inhibitors (K2 set) and Real Time PCR targeting B1 gene.
Źródło:
Annals of Agricultural and Environmental Medicine; 2018, 25, 4; 680-685
1232-1966
Pojawia się w:
Annals of Agricultural and Environmental Medicine
Dostawca treści:
Biblioteka Nauki
Artykuł
    Wyświetlanie 1-2 z 2

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