The development of modern molecular detection methods has led to imaging techniques becoming less significant for the detection of pathogens. They still, however, have an important role to play in the detection of unknown pathogens. The Bundeswehr Institute of Microbiology has therefore established a special focus on imaging techniques, covering conventional light microscopy and fluorescence-microscopy, confocal laser scanning microscopy, and electron microscopy.
Light microscopy, which is usually applied in combination with staining procedures, is commonly used to obtain initial indications of the presence of bacteria in specimens. With the use of special stains, Rickettsiae can also be visualised by light microscopy in cell culture. Using phase contrast microscopy, cytopathic effects can be made visible in living cells, providing clues as to the species of the cultivated virus. Dark field microscopy shows particularly small or low-contrast pathogens that have a small cell diameter and are difficult or impossible to visualise using conventional methods.
The Bundeswehr Institute of Microbiology uses fluorescence microscopy to detect antibodies against various viruses, Rickettsiae and Coxiellae. It is also employed to identify unknown viruses, non-lytic viruses and Rickettsiae in cell cultures. For this purpose, defined specific sera are used.
Fluorescence in situ hybridisation (FISH)
A special fluorescence microscopy technique is hybridisation with fluorescence-labelled DNA probes. It allows the identification and localization of pathogens in tissue sections. The technique was used at the Bundeswehr Institute of Microbiology to identify, for example, Francisella tularensis in lymph node tissue.
Confocal laser scanning microscopy
Confocal laser scanning microscopy is a special technique that combines the principle of fluorescence microscopy with the contrast-intensifying mechanism of confocal microscopy using laser and digital image scanners. By selecting suitable fluorescent dyes, individual structures in the cell can be visualised at high resolution and, if applicable, further investigations at a sub-cellular level can be performed. This method is used at the Bundeswehr Institute of Microbiology to investigate the growth characteristics and cell alterations caused by various TBE virus strains, Rickettsia and Francisella in various cells, including neuronal cells.
Electron microscopy is still the only technique available to obtain sufficiently detailed images of particles and structures on the micro- and nanoscales. It is an important “open view” procedure in medical biodefence which allows samples to be examined for viruses in a non-selective way. This may allow the preliminary identification of some pathogens associated with biothreats such as smallpox viruses or filoviruses. A Working Group for Electron Microscopy has been set up at the Bundeswehr Institute of Microbiology that focuses on scanning and transmission electron microscopy. In cooperation with the Centre for Electron Microscopy at the University of Ulm, it was possible to study the structure of a new sandfly fever virus that was isolated at the Institute.