Postdoc, Monash University
A field study that brings ATR-FTIR for malaria diagnosis from the lab to where it is actually needed
In the current research project, we used attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy for field studies in Laos to diagnose malaria and determine the applicability of this method for the detection of asymptomatic carriers of malaria. Malaria is one of the most devastating diseases worldwide. Its distribution in Southeast Asia is hard to control as many patients have a very low number of the malaria parasite Plasmodium in their blood. These patients often do not show classical malaria symptoms. Thus they are called asymptomatic carriers. Due to the low parasitemia, they are often very hard to diagnose. Recent experiments using (ATR-FTIR) spectroscopy for the detection of Plasmodium in human red blood cells (RBCs) were carried out in our lab on RBCs that had been spiked with Plasmodium falciparum (3D7 strain) to different percentages of parasitemia. ATR-FTIR spectra of RBCs were recorded after fixation with methanol. This study enabled to detect the parasite down to 0.00001% parasitemia, a value that would cause asymptomatic malaria in humans. The previously developed protocols had to be adapted to be applied in the field. Warm temperatures and high humidity as well as limited capacity for the storage or cooling of chemicals characterize the tropical environment in malaria regions. Therefore, sample preparation should not require further chemicals such as methanol for fixation. Furthermore, in order to increase the number of samples that can be detected per time, as few sample preparation steps as possible were desired. Preliminary experiments: Whole blood samples that were spiked with P. falciparum infected RBCs were tested after lysis with distilled water. Lysing RBCs before drying them on a glass slide lead to increased sensitivity combined with uncomplicated sample preparation. Field study: Blood samples of 596 participants from six different villages in the province Champasak in Laos were tested for malaria using microscopy, rapid diagnosis tests, polymerase chain reaction and ATR-FTIR spectroscopy on lysed dried blood samples. The evaluation of the thus obtained data is currently in process.
Abstract: New diagnostic modalities for malaria must have high sensitivity and be affordable to the developing world. We report on a method to rapidly detect and quantify different stages of malaria parasites, including ring and gametocyte forms, using attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FT-IR) and partial least-squares regression (PLS). The absolute detection limit was found to be 0.00001% parasitemia (<1 parasite/μL of blood; p < 0.008) for cultured early ring stage parasites in a suspension of normal erythrocytes. Future development of universal and robust calibration models can significantly improve malaria diagnoses, leading to earlier detection and treatment of this devastating disease.
Pub.: 04 Apr '14, Pinned: 29 Aug '17
Abstract: The mechanical properties of agarose-derived hydrogels depend on the scaffolding of the polysaccharide network. To identify and quantify such higher order structure, we applied Raman optical activity (ROA)-a spectroscopic technique that is highly sensitive toward carbohydrates-on native agarose and chemically modified agarose in the gel phase for the first time. By spectral global fitting, we isolated features that change as a function of backbone carboxylation (28, 40, 50, 60, 80, and 93 %) from other features that remain unchanged. We assigned these spectral features by comparison to ROA spectra calculated for different oligomer models. We found a 60:40 ratio of double- and single-stranded α-helix in the highly rigid hydrogel of native agarose, while the considerably softer hydrogels made from carboxylated agarose use a scaffold of unpaired β-strands.
Pub.: 24 Mar '17, Pinned: 29 Aug '17
Abstract: Vibrational circular dichroism (VCD) spectra of aqueous solutions of proline were recorded in the course of titrations from basic to acidic pH using a spectrometer equipped with a quantum cascade laser (QCL) as an infrared light source in the spectral range from 1320 to 1220 cm(-1). The pH-dependent spectra were analyzed by singular value decomposition and global fitting of a two-pK Henderson-Hasselbalch model. The analysis delivered relative fractions of the three different protonation species. Their agreement with the relative fractions obtained from performing the same analysis on pH-dependent Fourier transform infrared (FT-IR) and QCL-IR spectra validates the quantitative results from QCL-VCD. Global fitting of the pH-dependent VCD spectra of L-proline allowed for extraction of pure spectra corresponding to anionic, zwitterionic, and cationic L-proline. From a static experiment, only pure spectra of the zwitterion would be accessible in a straightforward way. A comparison to VCD spectra calculated for all three species led to assignment of vibrational modes that are characteristic for the respective protonation states. The study demonstrates the applicability of QCL-VCD both for quantitative evaluation and for qualitative interpretation of dynamic processes in aqueous solutions.
Pub.: 25 Mar '14, Pinned: 29 Aug '17
Abstract: Changes in vibrational circular dichroism (VCD) were recorded on-line during a chemical reaction. The chiral complex nickel-(-)-sparteine chloride was hydrolyzed to free (-)-sparteine base in a biphasic system of sodium hydroxide solution and chloroform (CHCl(3)). Infrared (IR) and VCD spectra were iteratively recorded after pumping a sample from the CHCl(3) phase through a lab-built VCD spectrometer equipped with a tunable mid-IR quantum cascade laser light source, which allows for VCD measurements even in the presence of strongly absorbing backgrounds. Time-dependent VCD spectra were analyzed by singular value decomposition and global exponential fitting. Spectral features corresponding to the complex and free (-)-sparteine could be clearly identified in the fitted amplitude spectrum, which was associated with an exponential decay with an apparent time constant of 127 min (t(½) = 88 min).
Pub.: 14 Mar '14, Pinned: 29 Aug '17