Laser-assisted indirect detection of bacterial endospores in suspicious macroscopic samples

Abstract

Globalisation, climate change, population growth, geopolitical tensions, radicalisation - these factors influence the security and stability of our society in a variety of ways and pose an increasing challenge in different areas. This also includes the threat posed by pathogenic microorganisms to our society and its security. In addition to multiple scenarios based on the natural spread of new or dangerous pathogens and their resistance towards antibiotics, the threat of deliberate release of pathogens is also of increasing importance. The first year of the Corona pandemic highlighted how sensitive modern society is to new and dangerous microorganisms and the impact this can have on various critical sectors and infrastructures. The general knowledge of society's sensitivity increases the risk of a terrorist act involving biological agents, especially as in bioterrorist activities the panic caused is an important effect in addition to the actual health threat. Rapid, reliable detection of organisms released during a terrorist attack or present in a clandestine laboratory as an indication of an imminent terrorist act are of the highest priority for the protection of the population and the emergency services. However, the specificity of currently used pathogen detection methods often correlates inversely with the detection speed. Furthermore, the detection principle cannot be automated in most cases, which would further increase the protection of involved and uninvolved persons during the operation. The aim of the present work was therefore to develop a test set-up in which certain groups of pathogens were to be detected or excluded at high detection speed. The concept focuses on the rapid detection of bacterial endospores in suspicious samples as an indication of the presence of pathogens of the genera Bacillus (e.g. Bacillus anthracis, Bacillus cereus) or Clostridium (e.g. Clostridium perfringens). The aim here is to quickly obtain a first statement about the biological hazard potential of the suspect sample and thus to be able to take initial protective measures at a very early stage. This technique should then be supplemented by more precise but longer-lasting identification techniques. Dipicolinic acid (DPA) serves as a marker molecule for endospores in the examined sample, which is released from the spore core by means of laser energy prior detection. A downstream reaction produces an antenna effect resulting from the interaction of DPA with lanthanides. Due to this effect, a shift in the wavelength of the emitted light can be measured after excitation in the UV range if DPA is present in the suspicious sample, from which conclusions can also be drawn about the concentration of DPA. The design of the detection concept is aimed at simplified automation at a later stage, both in terms of sampling and the possibility of digital optical evaluation.