Category A and B biothreat agents are deemed to be of great concern by the US Centers for Disease Control and Prevention (CDC) and include the bacteria Francisella tularensis, Yersinia pestis, Burkholderia mallei, and Brucella species. Underscored by the impact of the 2020 SARS-CoV-2 outbreak, 2016 Zika pandemic, 2014 Ebola outbreak, 2001 anthrax letter attacks, and 1984 Rajneeshee Salmonella attacks, the threat of future epidemics/pandemics and/or terrorist/criminal use of pathogenic organisms warrants continued exploration and development of both classic and alternative methods of detecting biothreat agents. Volatile organic compounds (VOCs) comprise a large and highly diverse group of carbon-based molecules, generally related by their volatility at ambient temperature. Recently, the diagnostic potential of VOCs has been realized, as correlations between the microbial VOC metabolome and specific bacterial pathogens have been identified. Herein, we describe the use of microbial VOC profiles as fingerprints for the identification of biothreatrelevant microbes, and for differentiating between a kanamycin susceptible and resistant strain. Additionally, we demonstrate microbial VOC profiling using a rapid-throughput VOC metabolomics method we refer to as 'simultaneous multifiber headspace solid-phase microextraction' (simulti-hSPME). Finally, through VOC analysis, we illustrate a rapid non-invasive approach to the diagnosis of BALB/c mice infected with either F. tularensis SCHU S4 or Y. pestis CO92. Category A and B biothreat agents include several bacteria, viruses, and toxins deemed to be of great concern by the US Centers for Disease Control and Prevention (CDC), due to their ease of dissemination/transmission, morbidity/mortality rates, and ability to cause public panic and disruption. Underscored by the impact of the 2020 SARS-CoV-2 outbreak, the 2016 Zika pandemic, the 2014 Ebola outbreak, the 2001 anthrax letter attacks, and the 1984 Rajneeshee Salmonella attacks, the threat of future epidemics/pandemics and/or terrorist use of pathogenic organisms warrants continued exploration and development of both classic and alternative methods of rapidly detecting agents of threat. Although sensitive and selective detection techniques such as polymerase chain reaction (PCR), microbial dichotomous keying, and/or enzyme-linked immunosorbent assays (ELISA) are well established 1 , these techniques are typically time consuming, laborious, and costly. Conversely, bacterial metabolome fingerprints, such as those derived through the use of techniques like Matrix-Assisted Laser Desorption Ionization (MALDI) coupled with mass spectrometry (MS), are rapid, reproducible, and cost effective 2,3. Using gas chromatography coupled with mass spectrometry (GC-MS), we have previously demonstrated the diagnostic potential of volatile organic compounds (VOCs) emanating from biological samples 4-7. VOCs comprise a large and highly diverse group of carbon-based molecules, generally related by their volatility at ambient temperature. A VOC-b...

The US Centers for Disease Control and Prevention (CDC) estimates that at minimum, 2 million people in the United States are infected with antimicrobial resistant (AMR) bacteria, resulting in 23,000 deaths each year. With no projected decrease in the number of deaths and the evergrowing threat of multi‐drug resistant bacteria, the development of a rapid and accurate diagnostic test is crucial, not only to reduce the burden on the public healthcare system, but to save countless lives.Microbial volatile organic compounds (mVOCs) are a structurally diverse group of microbial‐derived metabolites, generally related by their volatility at ambient temperature. There is great diagnostic potential for mVOCs, as numerous studies have identified unique mVOC‐fingerprints indicative of specific bacterial pathogens. In addition, specialized headspace sampling methods, such as solid‐phase microextraction (hSPME), coupled to gas chromatography, have enhanced the isolation, preconcentration, and analysis of mVOCs from biological specimens. Herein, we describe the development and assessment of our newly developed and patented metabolomics device and technique known as “simultaneous multifiber headspace solid‐phase microextraction (simulti‐hSPME),” for the rapid and minimally invasive preparation of mVOC fingerprints. This method is evaluated against antimicrobial susceptible and resistant strains of Yersinia pestis and Francisella tularensis.Support or Funding InformationFunding for this project is provided by DTRA (Grant Number: HDTRA 1‐16‐1‐0040).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.