Detection of SARS-CoV-2 using RT-PCR and other advanced methods can achieve high accuracy. However, their application is limited in countries that lack sufficient resources to handle large-scale testing during the COVID-19 pandemic. Here, we describe a method to detect SARS-CoV-2 in nasal swabs using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) and machine learning analysis. This approach uses equipment and expertise commonly found in clinical laboratories in developing countries. We obtained mass spectra from a total of 362 samples (211 SARS-CoV-2-positive and 151 negative by RT-PCR) without prior sample preparation from three different laboratories. We tested two feature selection methods and six machine learning approaches to identify the top performing analysis approaches and determine the accuracy of SARS-CoV-2 detection. The support vector machine model provided the highest accuracy (93.9%), with 7% false positives and 5% false negatives. Our results suggest that MALDI-MS and machine learning analysis can be used to reliably detect SARS-CoV-2 in nasal swab samples. The outbreak of coronavirus disease 2019 (COVID-19) is a crisis that affects rich and poor countries alike 1. Detection of SARS-CoV-2 in patient samples is a critical tool for monitoring spread of the disease, guiding therapeutic decisions and devising social distancing protocols 2. Detection assays based on RT-PCR are the most effective and sensitive method for diagnosis of SARS-CoV-2 infection and are used in laboratories around the world 3. However, some countries lack the laboratory resources and access to PCR kits to conduct testing at the required levels. Therefore, other reliable diagnostic techniques are needed. Most clinical diagnostic laboratories have MALDI-MS equipment, which is used to identify bacterial and fungal infections. We propose to leverage the ease-of-use and robustness of MALDI-MS pathogen identification for large-scale SARS-CoV-2 testing in developing countries. MALDI-MS-based assays rely on reference spectra of strains and bioinformatics for high-sensitivity and high-specificity species identification through proteomic profiling. This approach is well established and accepted in many countries for routine diagnostics of yeast and bacterial infections. However, no spectral libraries for SARS-CoV-2 identification using MALDI-MS are publicly available to our knowledge. We first acquired MALDI mass spectra of nasal swab samples that had been tested for SARS-CoV-2 by RT-PCR and analyzed them using machine learning (ML). In this experiment (Fig. 1a), a total of 362 samples (211 SARS-CoV-2-positive and 151 negative, unequivocally confirmed by PCR), which came from three different countries, Argentina (Lab 1), Chile (Lab 2) and Peru (Lab 3), were placed on the MALDI plate without prior sample purification.

A new method that allows a linear drift tube to be operated as a continuous ion mobility filter is described. Unlike conventional ion mobility instruments that use an electrostatic gate to introduce a packet of ions into a drift region, the present approach uses multiple segmented drift regions with modulated drift fields to produce conditions that allow only ions with appropriate mobilities to pass through the instrument. In this way, the instrument acts as a mobility filter for continuous ion sources. By changing the frequency of the applied drift fields it is possible to tune this instrument to transmit ions having different mobilities. A scan over a wide range of drift field frequencies for a single ion species shows a peak corresponding to the expected resonance time of the ions in one drift region segment and a series of peaks at higher frequencies that are overtones of the resonant frequency. The measured resolving power increases for higher overtones, making it possible to resolve structures that were unresolved in the region of the fundamental frequency. We demonstrate the approach by examining oligosaccharide isomers, raffinose and melezitose as well as a mixture of peptides obtained from enzymatic digestion of myoglobin. S eparation of a mixture of ions by ion mobility spectrometry (IMS) in linear drift tubes is initiated by gating short packets of ions into the front of a drift region containing a static buffer gas. Different species then migrate through the drift region under the influence of an applied field according to their mobilities in the buffer gas, such that ions with high mobilities reach the detector before those with low mobilities [1][2][3][4][5][6]. This approach has attracted considerable attention as a means of analyzing complex mixtures [7][8][9][10][11][12][13][14][15][16]. Additionally, measured mobilities can be converted to experimental cross sections and comparison of these values with cross sections for model geometries that are calculated can be used to characterize ion structures [17][18][19][20][21][22][23][24][25][26][27][28].In this report we introduce a new approach for isolating ions having specific mobilities (or collision cross sections). Ions from a continuous source enter a drift tube with segmented drift regions. The drift fields are modulated at a frequency that allows only those ions having mobilities that are resonant with the experimental conditions to be transmitted through all drift regions. In this way, this device filters away all ions except those with mobilities over the selected narrow range. An unanticipated feature of this approach is the observation that ions can be passed at overtone frequencies; moreover, the resolving power in higher overtone regions is greater than that observed in the fundamental frequency range. Because of the ability to select ions in different frequency regions, including those that are associated with higher overtones, we refer to the approach as overtone mobility spectrometry (OMS).To demonstrate this approach we have examin...

The transport of ions through multiple drift regions is modeled in order to develop an equation that is useful for an understanding of the resolving power of an overtone mobility spectrometry (OMS) technique. It is found that resolving power is influenced by a number of experimental variables, including those that define ion mobility spectrometry (IMS) resolving power: drift field (E), drift region length (L), and buffer gas temperature (T). However, unlike IMS, the resolving power of OMS is also influenced by the number of drift regions (n), harmonic frequency value (m), and the phase number (ϕ) of the applied drift field. The OMS resolving power dependence upon the new OMS variables (n, m, and ϕ) scales differently than the square root dependence of the E, L, and T variables in IMS. The results provide insight about optimal instrumental design and operation.

Overtone mobility spectrometry (OMS) is examined as a means of determining the collision cross sections for multiply-charged ubiquitin and substance P ions, as well as for singly-charged rafinose and melezitose ions. Overall, values of collision cross section measured by OMS for stable ion conformations are found to be in agreement with values determined by conventional ion mobility spectrometry (IMS) measurements to within ~1%, relative uncertainty. The OMS spectra for ubiquitin ions appear to favor different conformations at higher overtones. We propose that the changes in the distributions as a function of the overtone region in which they are measured arise from the elimination of ions that undergo structural transitions in the drift regions. Kinetics simulations suggest that structural transitions occurring on the order of a few ms and resulting in a ~4% change in ion collision cross sections are detected by OMS measurements. The unique method of distinguishing ion mobilities with OMS reveals these structural transitions which are not readily apparent from traditional IMS measurements.

Highlights This study with MALDI-TOF comprises, as far as we know, the first report describing the performance of this technology with COVID-19 diagnosis. This work would encourage researchers to explore the potential of MALDI-TOF MS to assess the feasibility of this technology, as a rapid and reproducible screening tool for diagnosis of SARS-CoV-2. According to our preliminary results, mass spectrometry-based methods combined with multivariate analysis showed potential as a complementary diagnostic tool.

A new, two-dimensional overtone mobility spectrometry (OMS-OMS) instrument is described for the analysis of complex peptide mixtures. OMS separations are based on the differences in mobilities of ions in the gas phase. The method utilizes multiple drift regions with modulated drift fields such that only ions with appropriate mobilities are transmitted to the detector. Here we describe a hybrid OMS-OMS combination that utilizes two independently operated OMS regions that are separated by an ion activation region. Mobility-selected ions from the first OMS region are exposed to energizing collisions and may undergo structural transitions before entering the second OMS region. This method generates additional peak capacity and allows for higher selectivity compared with the one-dimensional OMS method. We demonstrate the approach using a three-protein tryptic digest spiked with the peptide Substance P. The [M+3H]3+ ion from Substance P can be completely isolated from other components in this complex mixture prior to introduction into the mass spectrometer.

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Alzheimer's disease (AD) is the most common cause of dementia, affecting more than 36 million people worldwide. Octodon degus, a South American rodent, has been found to spontaneously develop neuropathological signs of AD, including amyloid-β (Aβ) and tau deposits, as well as a decline in cognition with age. Firstly, the present work introduces a novel behavioral assessment for O. degus - the burrowing test - which appears to be a useful tool for detecting neurodegeneration in the O. degus model for AD. Such characterization has potentially wide-ranging implications, because many of these changes in species-typical behaviors are reminiscent of the impairments in activities of daily living (ADL), so characteristic of human AD. Furthermore, the present work characterizes the AD-like neuropathology in O. degus from a gene expression point of view, revealing a number of previously unreported AD biomarkers, which are found in human AD: amyloid precursor protein (APP), apolipoprotein E (ApoE), oxidative stress-related genes from the NFE2L2 and PPAR pathway, as well as pro-inflammatory cytokines and complement proteins, in agreement with the known link between neurodegeneration and neuroinflammation. In summary, the present results confirm a natural neuropathology in O. degus with similar characteristics to AD at behavioral, cellular and molecular levels. These characteristics put O. degus in a singular position as a natural rodent model for research into AD pathogenesis and therapeutics against AD.