Interview with Dr. Wolfgang Fritzsche, Department head Nanobiophotonics at the Leibniz Institute of Photonic Technology Jena e.V.
Instant treatment is absolute vital for patients developing sepsis. Providing a specific therapy early on is key. To manage this the pathogenic organisms need to be identified accurately. But a fungal sepsis can still be a hard nut to crack.
A new method was developed in the ImSpec project that is now capable to identify pathogenic fungi not only faster, but more reliably. Dr. Wolfgang Fritsche explains MEDICA-tradefair.com exactly how this method works.
Dr. Fritzsche, what exactly is ImSpec and what research methods does it imply?
Dr. Wolfgang Fritzsche: Mainly ImSpec is a research platform to establish methods to detect and characterize biomolecules in parallel. We demonstrated this within this project by identifying sepsis-relevant fungi with DNA-based techniques. The common approach would be to only grow single pathogens with classic breeding techniques and use procedures of standard molecular biology. This project should demonstrate that it is possible to simultaneously detect different pathogenic organisms which nicely showed in the sepsis trials that were done at University Hospital Jena in collaboration with SIOS Messtechnik GmbH, ABS GmbH, Molidax GmbH and Nanopartica GmbH and were funded by the Federal Ministry of Education and Research.
Why exactly is sepsis so dangerous?
Fritzsche: Simply put your organs fail in a complex but systematic manner because of a bacterial or fungal infection. So, it is absolutely vital for diagnosis to know as fast as possible with what kind of pathogen you are dealing with. A targeted therapy early on greatly increases the odds of survival for the patient.
In this special case we dealt with fungal infections. Fungi are much more difficult to diagnose than bacteria, mainly because they take more time to be bred – we are talking of days here. Also, it is mostly not reasonable to use broadband fungicides. Antimycotics are very expensive compared to antibiotics and additionally weaken the patient. You do not want to give this medication just on suspicion, so you try to accurately identify the fungi you are dealing with in order to start the right treatment.
How exactly does this method you and your project partners developed work?
Fritzsche: It is a two-step method. First, we need to multiply the fungal DNA retrieved in the sample. This is done by polymerase chain reaction (PCR). The second step is filling a DNA micro array with the retrieved DNA fragments. This array consists of many measurement spots with different DNA affinities. This means that only specific strands of DNA will be bound to a certain spot and produce a signal.
In our case certain fungal DNA. To detect a signal the amplified DNA does not need to carry a marker molecule. We are using an effect called surface plasmon resonance. Light can be used to induce local resonance in metallic nanoparticles which then act as a sensor in each spot. To lock the pathogenic DNA to their surface the particles themselves need to be coated in a layer of single-standed complementary DNA. If any target DNA is present it will bind to the nanoparticle sensor and effectively alter the refraction index in the medium around this spot. This shifted plasmon resonance can be measured very accurately. With our new imaging technique it is now possible to read a whole array at once. We can retrieve all spectral information for all given pixels instantaneously by producing an image stack with a Michelson interferometer. We then use Fourier transformation on this stack to determine spectroscopic properties. Duration of analysis does not increase with the quantity of measured spots anymore and is only limited by optical resolution.
Fritzsche: We established a demonstrator in this project. So the basic technique of imaging spectroscopy for PCR based DNA detection is sound. There are two objectives that need to be met for this to become a clinical application. Firstly, on the clinical side of things, the sample treatment and standardization of conveyance and procedures preceding actual detection need to be put in a standardized protocol.
Secondly, our detection instruments used as a demonstrator are not an appliance yet and cannot be used in the lab or at bedside in a clinic. It is still a laboratory setup which needs automation and implementation of the according protocols. But all main principles and techniques could be demonstrated to be working so far.
Could this technique also be used on septic bacteria?
Fritzsche: Yes. It all starts with the amplification of pathogenic DNA. We did use fungal DNA, but you can adopt the protocol for septic bacteria by adjusting the DNA sequences during PCR. All following steps are the same and can be done the same way. In principle, our method can be used both with different DNA sequences and antibody based immunoassays.
The interview was conducted by Olga Wart. MEDICA-tradefair.com