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Paralytic shellfish poisoning (PSP) caused by consumption of shellfish that have fed on toxic algae is a major health issue worldwide. Dinoflagellates of the genus Alexandrium can produce high amounts of paralytic shellfish toxins (PSTs) at such low densities that water discoloration often associated with Harmful Algal Blooms (HABs) is not always evident. Detection of blooms during early stages is extremely important for human health issues, effects on shellfish populations, bioaccumulation of toxin within the food web, and maintenance of fisheries. Species that produce PSTs are difficult to distinguish morphologically from non-PST producing species, and current identification methods are expensive, time consuming, and require special training. Development of a rapid, low-cost and easy-to-use device to detect and monitor Alexandrium would be an important advancement as HABs vary interannually in location, intensity, and duration, making detection and prediction areas of necessary research.

Surface plasmon resonance (SPR) is a label-free, optical detection method that measures the change in refractive index after binding (hybridization) of target to probe on a surface. As target molecules bind to a probe on a metal surface, the refractive index shifts, causing a change in the surface plasmon wave. SPR can monitor biological interactions in real-time, providing a distinct advantage over other types of detection. Peptide nucleic acids (PNA) are short-sequenced DNA mimics where the negatively charged sugar-phosphate backbone is replaced by a neutral peptide chain. PNA probes have a high discrimination for mismatches, are resistant to protease and nuclease degradation, and will hybridize in low salt concentrations, making them ideal for use in field settings. Thiolated PNA probes with spacer molecules inserted before the probe sequence hybridize readily to unmodified gold surfaces forming self-assembling monolayers (SAMs). SAMS of PNA bind to target sequences following Watson-Crick rules for base parings and with heightened specificity.

Our research combines the use of SPR and PNA probes to develop direct detection sensors not currently feasible for field use or not sensitive enough with presently available molecular probes. This technology will fill a current gap in the ability to easily monitor potential HABs on-site with little sample processing.

From Bracher, A., L. Connell and R. Smith. Development of a direct detection method for Alexandrium spp. Using surface plasmon resonance and peptide nucleic acid probes. 2009. [VIEW PDF]

Relevant Literature:

• S. Shumway, S. Sherman-Caswell, J. Hurst. (1988) Paralytic shellfish poisoning in Maine: monitoring monster. J. Shellfish Res. vol .7, p. 175.
• D. Anderson (1997) Bloom dynamics of toxic Alexandrium species in the northeastern U.S. Limnol. Oceanogr. vol. 42 issue 5/2, pp. 1009-1022.
• J. Geraci, D. Anderson, R. Timperi, D. St. Aubin, G. Early, J. Prescott, C. May.(1989) Humpback whales (Megaptera novaeangliae) fatally poisoned by dinoflagellate toxin. Can. J. Fisheries and Aquatic Sciences vol. 46, pp. 1895-1898.
• D. McGillicuddy, D. Anderson, D. Lynch, D. Townsend. (2005) Mechanisms regulating large-scale seasonal fluctuations in Alexandrium fundyense populations in the Gulf of Maine: Results from a physical-biological model,” Deep-Sea Res. II vol. 52 issue 19-21, pp. 2698-2714.
• K. Jensen, H. Orum, P. Nielson, B. Norden. (1997) Kinetics for hybridization of peptide nucleic acids (PNA) with DNA and RNA studied with the BIAcore technique. Biochemistry vol. 36.
• P. Nielsen, M. Egholm, O. Buchardt (1994) Peptide Nucleic Acid (PNA). A DNA Mimic with a Peptide Backbone. Bioconjugate Chem. vol. 5, pp. 3-7.
• P. E. Nielsen, M. Egholm, R. Berg and O. Buchardt. (1991) Sequence selective recognition of DNA by strand displacement with a thyminesubstituted polyamide. Science, 254, 1497–1500.
• Connell Lab publications [READ MORE]

Project funded by: NOAAs CSCOR MERHAB

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