Research Task 11 Development of biosensors for use in marine systems

PhD position. Host: Aarhus with secondments to MPI and Unisense. Supervisors: Prof. Niels Revsbech, Dr Jens Gundersen and Dr Dirk De Beer

The development of biosensors for methane and nitrate was initiated a decade ago,^(1,2) and in 2004 a paper on a NO^₂- biosensor was published.⁽³⁾ Especially the nitrate and nitrite biosensors are of great interest to the aquatic sciences,⁽⁴⁾ and they often function well with resolutions down to 0.1 µM and with great short-term (hours to days) stability. The sensors are based on immobilized bacteria transforming nitrite + nitrate or nitrite alone to nitrous oxide that is subsequently detected by an electrochemical sensor. The 90% response time for a macroscale sensor at room temperature is about 2 min, and for microscale sensors it is about 30 s. Bacteria (Pseudomonas sp.) isolated from Greenland now allow work with biosensors down to 0°C and at salinities ranging from about 0.1 to 40‰. We are thus in principle able to perform routine work in the deep sea at >2°C and <37‰ salinity, and the first deep sea nitrate determinations have been performed with an autonomous deep sea lander off the Japanese coast (R.N. Glud and N.P. Revsbech, unpublished data from 2008). The various components of the biosensors are well characterized (well-functioning membranes, almost ideal electrochemical sensor), but we have problems with the immobilization of the bacteria in the tip, and often the bacterial biomass moves away and into the depth of the nutrient reservoir inside the sensor. If this problem can be solved prolonged marine deployments for nitrite and nitrate measurements can be performed, as the lifetime of such sensors could be several months. Already with the present methodology a Ph.D. student can get sufficient data for a successful Ph.D. project, but in addition the student will improve microbial immobilization inside the biosensor using extracellular stabilizing constituents of the inner medium (Fe³⁺, Al³⁺), gels that are compatible with a phosphate-containing inner medium, mutanegenesis to inhibit flagellar movement, etc. If successful this will lead to a routinely used tool for the aquatic sciences. In the project focus will be on in-situ measurements, although the fieldwork will be supplemented with laboratory investigations.

⁽¹⁾ Larsen, L.H., et al., 1997. Anal. Chem. 69: 3527-3531. ⁽²⁾ Damgaard, L.R., & N.P. Revsbech. 1997. Anal. Chem. 69: 2262-2267. ⁽³⁾ Nielsen, M., et al., 2004. Appl. Env. Microbiol. 70: 6551-6558.⁽⁴⁾ Revsbech, N.P., et al., 2006. Antonie van Leeuwenhoek Int. J. Gen. Mol. Microbiol. 90 (4): 361-375.