Predominant SARS-CoV-2 variant impacts accuracy when screening for infection using exhaled breath vapor.

BACKGROUND: New technologies with novel and ambitious approaches are being developed to diagnose or screen for SARS-CoV-2, including breath tests. The US FDA approved the first breath test for COVID-19 under emergency use authorization in April 2022. Most breath-based assays measure volatile metabolites exhaled by persons to identify a host response to infection. We hypothesized that the breathprint of COVID-19 fluctuated after Omicron became the primary variant of transmission over the Delta variant. METHODS: We collected breath samples from 142 persons with and without a confirmed COVID-19 infection during the Delta and Omicron waves. Breath samples were analyzed by gas chromatography-mass spectrometry. RESULTS: Here we show that based on 63 exhaled compounds, a general COVID-19 model had an accuracy of 0.73 ± 0.06, which improved to 0.82 ± 0.12 when modeling only the Delta wave, and 0.84 ± 0.06 for the Omicron wave. The specificity improved for the Delta and Omicron models (0.79 ± 0.21 and 0.74 ± 0.12, respectively) relative to the general model (0.61 ± 0.13). CONCLUSIONS: We report that the volatile signature of COVID-19 in breath differs between the Delta-predominant and Omicron-predominant variant waves, and accuracies improve when samples from these waves are modeled separately rather than as one universal approach. Our findings have important implications for groups developing breath-based assays for COVID-19 and other respiratory pathogens, as the host response to infection may significantly differ depending on variants or subtypes.

In recent decades, scientists have found we exhale thousands of compounds that reveal much about our health, including whether we are sick with COVID-19. Our team asked whether the breath profile of someone infected with the Delta variant of COVID-19 would match the breath profile caused by the Omicron variant­a version of the virus that is more transmissible. We analyzed breath samples from 142 people, some sick with either the Delta or Omicron variant of COVID-19, and others who were negative for COVID-19. Our results indicate that the Delta variant altered the contents of our breath in a different way than the Omicron variant, and breath-based tests improved when optimized to detect only one of the variants. These findings might impact the design of future breath-based tests for COVID-19.

A low cost, easy-to-assemble, open-source modular mobile sampler design for thermal desorption analysis of breath and environmental VOCs.

Exhaled breath vapor contains hundreds of volatile organic compounds (VOCs), which are the byproducts of health and disease metabolism, and they have clinical and diagnostic potential. Simultaneous collection of breath VOCs and background environmental VOCs is important to ensure analyses eliminate exogenous compounds from clinical studies. We present a mobile sampling system to extract gaseous VOCs onto commercially available sorbent-packed thermal desorption tubes. The sampler can be connected to a number of commonly available disposable and reusable sampling bags, in the case of this study, a Tedlar bag containing a breath sample. Alternatively, the inlet can be left open to directly sample room or environmental air when obtaining a background VOC sample. The system contains a screen for the operator to input a desired sample volume. A needle valve allows the operator to control the sample flow rate, which operates with an accuracy of -1.52 ± 0.63% of the desired rate, and consistently generated that rate with 0.12 ± 0.06% error across repeated measures. A flow pump, flow sensor and microcontroller allow volumetric sampling, as opposed to timed sampling, with 0.06 ± 0.06% accuracy in the volume extracted. Four samplers were compared by sampling a standard chemical mixture, which resulted in 6.4 ± 4.7% error across all four replicate modular samplers to extract a given VOC. The samplers were deployed in a clinical setting to collect breath and background/environmental samples, including patients with active SARS-CoV-2 infections, and the device could easily move between rooms and can undergo required disinfection protocols to prevent transmission of pathogens on the case exterior. All components required for assembly are detailed and are made publicly available for non-commercial use, including the microcontroller software. We demonstrate the device collects volatile compounds, including use of chemical standards, and background and breath samples in real use conditions.

Devices Engineered to Collect Exhaled Breath Condensate (EBC) and their Applications

IMPACT: Human exhaled breath is rich in metabolomic content that represents pulmonary function and gas exchange with blood, which can provide insights into an individual’s state of health. OBJECTIVES/GOALS: Human exhaled breath is rich in metabolomic content that represents pulmonary function and gas exchange with blood. It contains a mixture of compounds that offer insight into an individual’s state of health. Here, we present two novel non-invasive breath sampling devices for use in basic medical practice. METHODS/STUDY POPULATION: The two breath samplers have a disposable mouthpiece, a set of inhale and exhale one-way flap valves to allow condensation of exhaled breath only, and a saliva filter. The housing is constructed out of Teflon®, a chemically inert material to reduce chemical absorbance. The first device condenses exhaled breath into a frozen condensate using dry ice pellets and the other is a miniaturized design that liquifies exhaled breath on a condenser surface with micropatterned features on a cooling plate. Both designs have individual strategic and analytical advantages: frozen exhaled breath condensate (EBC) has high retention of analytes and sample volume;EBC collected in liquid phase offers facilitated sample collection and device portability. RESULTS/ANTICIPATED RESULTS: We investigated if breath aerosol size distribution affects the types or abundances of metabolites. We modified the geometry of the first device to redirect aerosol trajectories based on size. The trapping of larger aerosols increases with filter length, thus altering the aerosol size distribution although no significant changes in the metabolite profiles were found. With the miniaturized device, metabolite abundances were measured in a small cohort of healthy control and mild asthmatic subjects. Differences among subjects were found, as well as main differences between control and asthmatic groups. All analyses of EBC were performed with liquid chromatography - mass spectrometry. Inflammatory suppression found in asthmatic subjects can be explained by prescribed daily use of inhaled corticosteroids. DISCUSSION/SIGNIFICANCE OF FINDINGS: Breath collection devices can be used in intensive care units, outpatient clinics, workplaces, and at home. EBC analysis has been used to monitor asthma and chronic obstructive pulmonary disease. It can be applied to infectious respiratory diseases (e.g. influenza, COVID-19) and for monitoring environmental and occupational chemical exposures.

Inactivation of SARS-CoV-2 in clinical exhaled breath condensate samples for metabolomic analysis.

Exhaled breath condensate (EBC) is routinely collected and analyzed in breath research. Because it contains aerosol droplets, EBC samples from SARS-CoV-2 infected individuals harbor the virus and pose the threat of infectious exposure. We report for the first time a safe and consistent method to fully inactivate SARS-CoV-2 in EBC samples and make EBC samples safe for processing and analysis. EBC samples containing infectious SARS-CoV-2 were treated with several concentrations of acetonitrile. The most commonly used 10% acetonitrile treatment for EBC processing failed to completely inactivate the virus in samples and viable virus was detected by the assay of SARS-CoV-2 infection of Vero E6 cells in a biosafety level 3 laboratory. Treatment with either 50% or 90% acetonitrile was effective to completely inactivate the virus, resulting in safe, non-infectious EBC samples that can be used for metabolomic analysis. Our study provides SARS-CoV-2 inactivation protocol for the collection and processing of EBC samples in the clinical setting and for advancing to metabolic assessments in health and disease.