Surface-Enhanced Raman Scattering (SERS) in Microbiology: Illumination and Enhancement of the Microbial World

The microbial world forms a huge family of organisms that exhibit the greatest phylogenetic diversity on Earth and thus colonize virtually our entire planet. Due to this diversity and subsequent complex interactions, the vast majority of microorganisms are involved in innumerable natural bioprocesses and contribute an absolutely vital role toward the maintenance of life on Earth, whilst a small minority cause various infectious diseases. The ever-increasing demand for environmental monitoring, sustainable ecosystems, food security, and improved healthcare systems drives the continuous search for inexpensive but reproducible, automated and portable techniques for detection of microbial isolates and understanding their interactions for clinical, environmental, and industrial applications and benefits. Surface-enhanced Raman scattering (SERS) is attracting significant attention for the accurate identification, discrimination and characterization and functional assessment of microbial cells at the single cell level. In this review, we briefly discuss the technological advances in Raman and Fourier transform infrared (FT-IR) instrumentation and their application for the analysis of clinically and industrially relevant microorganisms, biofilms, and biological warfare agents. In addition, we summarize the current trends and future prospects of integrating Raman/SERS-isotopic labeling and cell sorting technologies in parallel, to link genotype-to-phenotype in order to define community function of unculturable microbial cells in mixed microbial communities which possess admirable traits such as detoxification of pollutants and recycling of essential metals.

Generalized Implementation of Rapid-Scan Fourier Transform Infrared Spectroscopic Imaging

We describe a novel, generalized data acquisition sequence to allow rapid-scan Fourier transform infrared (FT-IR) spectroscopic imaging using focal plane array (FPA) detectors. This technique derives its applicability from the reproducible performance of modern FT-IR instrumentation and the availability of FPAs with simultaneous, full array acquisition, or snapshot electronics. Instead of sampling the entire interferogram in one mirror sweep over a predetermined retardation, as in traditional continuous-scanning techniques, the modulated light from the interferometer is recorded over several mirror sweeps. The FPA detector is synchronized for data acquisition after a specified delay with respect to the initiation of the mirror motion to provide a highly under-sampled interferogram. By incorporating appropriate delays in subsequent interferometer mirror scans, the entire interferogram is sampled and reconstructed. The signal-to-noise ratios (SNR) of the resulting interferograms are analyzed and are compared with step-scan spectroscopic imaging data.

Time-Resolved Step-Scan FT-IR Spectroscopy of the Photodynamics of Carbonmonoxymyoglobin

The kinetics of protein response and of CO recombination after photolysis of the Fe-CO bond in carbonmonoxymyoglobin have been monitored via time-resolved step-scan FT-IR absorption difference spectroscopy in D2O solution. Although the initial photodissociation is too fast to observe with currently available FT-IR instrumentation, we have been able to correlate the CO recombination kinetics with protein secondary structural changes via changes in the amide I band of the polypeptide chain with microsecond time resolution. The spectral and kinetic data corroborate and confirm previously published single-frequency infrared studies. This is the first application of time-resolved step-scan FT-IR spectroscopy in the absorbance difference mode to study the photodynamics of an aqueous protein solution at room temperature. This work also demonstrates the potential of the technique for the sub-microsecond kinetic analysis of other biological molecules of interest.

The Near-Infrared Capabilities of LEST

The Large Earth-based Solar Telescope (LEST) will be a powerful, next-generation telescope with unprecedented angular resolution, capable of highly accurate polarimetry of the Sun, covering the optical spectral range from about 300 nm into the near infrared to about 2.5 μm.The telescope is a 2.4-m aperture, “polarization-free” concept based on a modified Gregorian optical system. A fast polarization modulator will be located close to the secondary focus of the system. An actively controlled NTT-type main mirror, a high precision pointing and tracking system, a helium-filled light path and a thin entrance window, together with an integrated adaptive optics system, will give the telescope near diffraction-limited performance in the visible. LEST will be sited on La Palma, in the Canary Islands, near the caldera rim on the Roque de los Muchachos Observatory, which often offers excellent seeing. A frequently occurring seeing parameter of ro = 15–20 cm in the visible will correspond to ro ≥ 1 m in the near IR.The construction of LEST will begin in 1993, and the telescope is to be ready for “first light” in 1997. The telescope facility will accommodate a large number of focal plane instruments on a spacious instrument table. LEST will be made available for near-IR instrumentation from the start of its regular operation.