Malaria is one of the leading killer diseases in sub-Saharan Africa. Although the disease is curable, early and accurate diagnosis is key to effective therapy. Existing malaria diagnostic techniques have low detection accuracy especially when the parasite load in the blood is low. In this paper, we report on a simple photo-thermal based technique for detection of the Plasmodium parasites’ biomarker (hemozoin) in blood smear samples. The technique has demonstrated 100% Plasmodium detection sensitivity and specificity from the ten blood smear test samples used.
Malaria is a tropical disease that has been a leading cause of deaths in sub-Saharan Africa with children below five years being the most affected. According to World Health Organization (WHO) 2015 malaria report, the disease caused about 0.5 million deaths globally and infected approximately 200 million people during the reported period [
Early and accurate diagnosis of the malaria is key to effective treatment of the disease. However existing malaria diagnostic technique such as optical microscopy (which is the gold standard method) and Plasmodium parasites antigen detection assays also known as Rapid Diagnostic Techniques (RDTs) lack high detection sensitivity and specificity [
Limited studies have been reported on development of non-invasive techniques for malaria diagnosis [
the absorber’s intrinsic optical properties (specifically the refractive index) from the equilibrium (steady) state which in turn causes divergence and change of intensity of the probe beam.
The reported photoacoustic based malaria diagnostic technique on the other hand suffers from a number of limitations which has hampered its clinical adoption. One major challenge has been the problem of wide inter- and intra-variability of detected PA signals. Detected acoustic signals also suffer from low SNR due to acoustic attenuation before the signal reaches the transducer. The conversion efficiency from optical energy to acoustic energy in tissue (water) is also significantly low due to the low value of Gruneisten parameter for water which is estimated to be 0.2 [
In this paper, we report development of a novel technique termed as Photothermal Induced Optical Scattering Modulation (PTIOSM) for detection of hemozoin in malaria infected blood. The technique is premised on the fact that photon absorption by chromophores is followed by a relaxation session where the absorber’s molecules move from excited state to ground state. If the relaxation process is non-radiative, the absorbed optical energy is released in form of heat. The emitted heat energy takes some time before it diffuses away from where it is deposited (Thermal conferment time). During this phase, the intrinsic optical properties such as the absorber’s refractive index and its optical absorption coefficient are temporarily modified.
In the case of PTIOSM technique, modulation in the probe beam signal is detected in the reflection (backward) mode instead of transmission (forward) mode as is the case for thermal lens deflection. Modulation intensity with respect to optical excitation wavelength is expected to carry information on the composition as well as the concentration on chromophores in the probed sample.
A common anode, 3 W - 9 W RGB Light Emitting Diode (LED) (product ID 2524, from Adafruit Industries, USA [
Light from the RGB LED (PB and EB) irradiated samples (stained blood smear slides) and the transmitted light passed through an optical filter (Omega Optical, Inc. 450 nm SP (450 SP) RapidEdge 25 mm Optical Shortpass Filter) that blocked the EB (the red and green pulsed light) but transmitted the probe beam (the blue light) after interacting with the photo-thermally excited sample. A photodiode and trans-impedance amplifier module (OPT101) was used to detect light from the filter and the generated photodiode current was converted to a voltage signal and then pre-amplified using an inbuilt trans-impedance amplifier in the module. To boost the bandwidth of the detected signal, a 100 ohms resistor was externally connected to the trans-impedance module (between pin 2 and 5 while pin 4 was left unconnected) to provide the negative feedback instead of using the inbuilt 1 MΩ resistor that offers a limited bandwidth of 12 KHz [
The aim of this stage was to reduce or possibly eliminate the noise riding in the acquired PTIOSM signal. There were two possible main sources of noise in the acquired signals; the radiative pick-up noise from adjacent switching equipment such as the function generator, and the background (stray) light detected by the photodiode. Noise correction was performed by coherently subtracting the PTIOSM signal due to a blank glass slide (also termed as the reference sample) from a PTIOSM signal obtained from a stained blood smear sample. The resultant differential signal was free of existing common mode noise.
Fast Fourier Transform (FFT) algorithm implemented in Matlab was used to convert the acquired time domain signals to frequency domain. The amplitude and phase spectrums of the frequency domain signals were plotted in Matlab and visually analyzed with a view of identifying the frequency bands which best differentiated infected samples from non-infected samples based on their signal intensities at different frequency bands.
Two sets of Geimsa stained blood smear samples were used to test the developed PTIOSM probe. One was Plasmodium falciparum infected blood smears samples (acquired from Carolina biological company [
After conversion from time domain to frequency domain and noise filtering of PTIOSM signals, scatter plots for the amplitude spectra belonging to the five Plasmodium positive and five Plasmodium negative samples were made (
PTIOSM technique ability to correctly classify malaria infected blood smear samples from the non-infected samples can be attributed to two factors; change of hemozoin’s refractive index during optical excitation hence causing a significant modulation of the PTIOSM signal at frequency band corresponding to its size. Another possible explanation could be that the excitation beam causes generation of photoacoustic waves, which in turn induces vibration of optical scatterer in the medium leading to modulation of diffuse reflected light from the sample. Variations of signal intensity from samples having the same infection status (either infected or non-infected) can be attributed to two possible factors: the concentration of responsible chromophores (in this case hemozoin and hemoglobin) and the optical excitation intensity.
From the reported results, the developed PTIOSM probe has demonstrated excellent capability of malaria diagnosis in blood smear samples. The simple instrumentation involved and its real time operation makes it a suitable candidate for mass screening of the disease in malaria endemic regions. However, further testing using more samples prepared under different conditions (both thin and thick blood smears, stained and non-stained) are required to conclusively determine the techniques sensitivity and specificity. Besides, further experimentation is required to investigate the capability of the probe in quantifying the parasite load (parasitemia) in the samples and the limit of detection of the technique.
A novel, simple photo-thermal based technique (PTIOSM) for detection of Plasmodium parasites in infected blood smear samples has been described. A RGB LED was used as the optical source to supply both the PB and the EB. Diffusely reflected PB light from the sample was detected by a photodiode, amplified and the signal preprocessed for noise cancellation. Fourier transform was used to convert the signals from time domain to frequency domain and then some specific frequency bands were used to differentiate Plasmodium infected samples from non-infected samples based on an empirically determined intensity threshold. The technique yielded 100% detection sensitivity and specificity using red LED light as the excitation beam.
The authors declare no conflicts of interest regarding the publication of this paper.
Memeu, D.M., Sarroney, M.A. and Maina, C. (2018) Photo-Thermal Induced Optical Scattering Modulation Sensor for Malaria Diagnosis. Open Journal of Biophysics, 8, 185-193. https://doi.org/10.4236/ojbiphy.2018.84014