AI Article Synopsis
- A TaqMan MGB assay targeting conserved regions of Tritrichomonas foetus was developed to improve diagnostic sensitivity compared to traditional methods.
- The assay showed 2500-fold increased sensitivity over microscopy and 500-fold over conventional PCR when tested on smegma or mucus samples.
- Urine as a specimen offered similar detection levels as other methods but created inconclusive results in field studies, highlighting the assay's effectiveness in detecting T. foetus in clinical specimens.
Article Abstract
A Tritrichomonas foetus-specific 5' Taq nuclease assay using a 3' minor groove binder-DNA probe (TaqMan MGB) targeting conserved regions of the internal transcribed spacer-1 (ITS-1) was developed and compared to established diagnostic procedures. Specificity of the assay was evaluated using bovine venereal microflora and a range of related trichomonad species. Assay sensitivity was evaluated with log(10) dilutions of known numbers of cells, and compared to that for microscopy following culture (InPouch TF test kit) and the conventional TFR3-TFR4 PCR assay. The 5' Taq nuclease assay detected a single cell per assay from smegma or mucus which was 2500-fold or 250-fold more sensitive than microscopy following selective culture from smegma or mucus respectively, and 500-fold more sensitive than culture followed by conventional PCR assay. The sensitivity of the conventional PCR assay was comparable to the 5' Taq nuclease assay when testing purified DNA extracted from clinical specimens, whereas the 5' Taq nuclease assay sensitivity improved using crude cell lysates, which were not suitable as template for the conventional PCR assay. Urine was evaluated as a diagnostic specimen providing improved and equivalent levels of T. foetus detection in spiked urine by both microscopy following culture and direct 5' Taq nuclease detection, respectively, compared with smegma and mucus, however inconclusive results were obtained with urine samples from the field study. Diagnostic specimens (n=159) were collected from herds with culture positive animals and of the 14 animals positive by 5' Taq nuclease assay, 3 were confirmed by selective culture/microscopy detection (Fisher's exact test P<0.001). The 5' Taq nuclease assay described here demonstrated superior sensitivity to traditional culture/microscopy and offers advantages over the application of conventional PCR for the detection of T. foetus in clinical samples.
Download full-text PDF |
Source |
|---|---|
| http://dx.doi.org/10.1016/j.vetpar.2006.06.012 | DOI Listing |
Publication Analysis
Top Keywords
Similar Publications
Construction of an end-repairing-engineered quadratic in vitro transcription machine for single-molecule monitoring of alkaline phosphatase in human cancers.
Talanta
February 2025
School of Chemistry and Chemical Engineering, State Key Laboratory of Digital Medical Engineering, Southeast University, Nanjing, 211189, China. Electronic address:
Alkaline phosphatase (ALP) is an essential hydrolase widely present in humans, and it extensively acts as a biomarker for multiple human diseases. Conventional ALP assays suffer from complicated synthesis, tedious operation, low sensitivity, and large sample consumption. Herein, we construct an end-repairing-engineered quadratic in vitro transcription machine for single-molecule monitoring of ALP in diverse cancers with 3'-phosphoryl (PO) nucleic acid as a macromolecular substrate.
View Article and Find Full Text PDFStructural Basis for C2'-methoxy Recognition by DNA Polymerases and Function Improvement.
J Mol Biol
October 2024
Division of Biological Sciences and Medicine, University of Science and Technology of China, Hefei, Anhui 230027, PR China. Electronic address:
DNA modified with C2'-methoxy (C2'-OMe) greatly enhances its resistance to nucleases, which is beneficial for the half-life of aptamers and DNA nanomaterials. Although the unnatural DNA polymerases capable of incorporating C2'-OMe modified nucleoside monophosphates (C2'-OMe-NMPs) were engineered via directed evolution, the detailed molecular mechanism by which an evolved DNA polymerase recognizes C2'-OMe-NTPs remains poorly understood. Here, we present the crystal structures of the evolved Stoffel fragment of Taq DNA polymerase SFM4-3 processing the C2'-OMe-GTP in different states.
View Article and Find Full Text PDFSimultaneous Detection of Influenza A/B, Respiratory Syncytial Virus, and SARS-CoV-2 in Nasopharyngeal Swabs by One-Tube Multiplex Reverse Transcription Polymerase Chain Reaction.
Trop Med Infect Dis
June 2023
Department of Pulmonary Medicine, Government Medical College, Srinagar 190001, India.
The treatment and outcome of respiratory virus infections differ. SARS-CoV-2, as well as other respiratory viruses such as influenza virus (A and B) and respiratory syncytial virus (RSV), require simultaneous, cost-effective, and rapid differential detection. We used a gold standard five-target single-step RT-PCR to detect influenza viruses, RSV, and SARS-CoV-2, and this method can be extended to detect influenza virus subtypes.
View Article and Find Full Text PDFCRISPR-Cas9 recognition of enzymatically synthesized base-modified nucleic acids.
Nucleic Acids Res
February 2023
KU Leuven, Department of Pharmaceutical and Pharmacological Sciences, Rega Institute for Medical Research, Medicinal Chemistry, Herestraat 49, Box 1041, 3000 Leuven, Belgium.
An enzymatic method has been successfully established enabling the generation of partially base-modified RNA (previously named RZA) constructs, in which all G residues were replaced by isomorphic fluorescent thienoguanosine (thG) analogs, as well as fully modified RZA featuring thG, 5-bromocytosine, 7-deazaadenine and 5-chlorouracil. The transcriptional efficiency of emissive fully modified RZA was found to benefit from the use of various T7 RNA polymerase variants. Moreover, dthG could be incorporated into PCR products by Taq DNA polymerase together with the other three base-modified nucleotides.
View Article and Find Full Text PDFFirst report of Tobacco rattle virus infecting Brassica oleracea var. botrytis (cauliflower) in India.
Plant Dis
November 2022
Indian Agricultural Research Institute, 28802, Division of Plant Pathology, New Delhi, Delhi, India;
Article Synopsis
- Cauliflower, an important vegetable crop, showed virus-like symptoms such as stunted growth and leaf deformities during the summer of 2022 at the Indian Agricultural Research Institute (IARI) in New Delhi.
- The symptoms were linked to a potential tobacco rattle virus (TRV) infection, confirmed through transmission electron microscopy which revealed rod-shaped particles.
- Reverse transcription PCR was used to amplify a specific gene associated with TRV, confirming the presence of the virus in three cauliflower samples collected from the research institute.
Want AI Summaries of new PubMed Abstracts delivered to your In-box?
Enter search terms and have AI summaries delivered each week - change queries or unsubscribe any time!