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Sexually Transmitted Diseases
Published in Victor A. Bernstam, Pocket Guide to GENE LEVEL DIAGNOSTICS in Clinical Practice, 2019
A DNA probe assay for this organism using radiolabeled DNA fragments from a virulent strain of H. ducreyi could identify the organism with a sensitivity of 103 colony-forming units (cfu) in colony hybridization assays in pure or mixed cultures.
Protein Subunit Vaccines and Recombinant DNA Technology
Published in F. Y. Liew, Vaccination Strategies of Tropical Diseases, 2017
Most frequently bacteriophage lambda vectors are used.24 Lambda combines many of the useful properties of plasmids together with the ability to accept long fragments of DNA. Lambda can accept up to 20 to 25 kb of DNA. Cosmid vectors (plasmids containing the cos site of lambda) can accept even more DNA, up to 45 kb but are more difficult to use. In the region of 1000 clones may be required in order to obtain a DNA library representative of an entire bacterial genome. This would increase to 105 to 106 for a more complex eukaryotic genome. Finally the same process of in situ colony hybridization can be used to identify relevant DNA sequences.
Loop-mediated isothermal amplification assay as a point-of-care diagnostic tool for Vibrio parahaemolyticus: recent developments and improvements
Published in Expert Review of Molecular Diagnostics, 2019
Karanth Padyana Anupama, Anirban Chakraborty, Iddya Karunasagar, Indrani Karunasagar, Biswajit Maiti
The study by Yamazaki et al. [33] represents the first report of LAMP assay for V. parahaemolyticus, which targeted the tlh gene. The study revealed that as less as 5.3 × 102 CFU/ml V. parahaemolyticus could be detected in less than 40 min, which was ten-fold more sensitive than conventional PCR. Subsequently, the LAMP assay targeting the toxR gene of V. parahaemolyticus was also reported [76]. These assays, although highly sensitive, were not specific to the virulent strains. Expectedly, LAMP assays targeting the virulent strain-specific genes such as tdh and trh were also developed [77,78]. These pioneering studies proved that the sensitivity of the LAMP assay is higher than PCR and colony hybridization. In addition, it was also clear that LAMP takes lesser time compared to the existing methods. Considering these favorable qualities, attempts have been made to improve upon the underlying methodology. Over the years, different versions of LAMP assays have been developed aimed at proving its potential as the point-of-care diagnostic tool for rapid and efficient detection of V. parahaemolyticus. The conventional approach of LAMP assay is more practical and straightforward compared to other LAMP assays. So far, LAMP assay has not been assessed for the detection of V. parahaemolyticus from clinical samples. Although reports are available that used clinical isolates to demonstrate the efficacy of the technique, LAMP assay is yet to find an application in the detection of V. parahaemolyticus in a routine clinical setup. Usually, environmental samples show a higher prevalence of V. parahaemolyticus, which includes both pathogenic and non-pathogenic strains. In contrast, the prevalence of V. parahaemolyticus is usually low in clinical samples, but it includes only the pathogenic strains. This further emphasizes the need for a robust and sensitive technique that could allow the detection of the pathogenic strains to prevent possible outbreaks.