Highly Specific Ligation-dependent Microarray Detection of Single Nucleotide Polymorphisms

Abstract

The fast detection and characterization of pathogens are essential for an efficient treatment of infectious diseases. However, the development of improved and reliable diagnostic methods is still an ongoing process because not only pathogens but also their antibiotic resistances have to be identified. The gold standard today is, however, a cultivation-based characterization approach, which takes days until results can be evaluated. In patients with, for example, severe sepsis, the diagnostic test duration is a very critical parameter because a delay of treatment optimization increases the mortality rate significantly. In contrast, DNA-based molecular techniques can obtain results within a few hours. A further challenge in diagnostic laboratories is that patient samples have to be screened for hundreds of potential pathogens, antibiotic resistance genes, and virulence factors, which is achieved by using a number of specialized tests at the moment. Microarrays are outstandingly good for the simultaneous analysis of thousands of different genes and have become a popular tool in biological studies. Nevertheless, further optimizations of the microarray technology are required due to the obligatory DNA labeling and/or amplification steps and the effects of nonspecific DNA hybridization. Here, we describe a fast and highly specific solid-support-based DNA characterization method for pathogens and antibiotic resistance genes.