bioRxiv Science⌕ Search
Home/Archive/bioRxiv

bioRxiv · 10.1101/2021.02.16.431476

Capturing actively produced microbial volatile organic compounds from human associated samples with vacuum assisted sorbent extraction

Phan, J.·Kapcia, J.·Rodriguez, C. I.·Vogel, V. L.·Dunham, S. J. B.·Whiteson, K.
View original source ↗Export metadata ↓

Abstract

Volatile organic compounds (VOC) from biological samples have unknown origins. VOCs may originate from the host or different microbial organisms from a microbial community. In order to disentangle the origin of microbial VOCs, we performed volatile headspace analysis of bacterial mono- and co-cultures of Staphylococcus aureus, Pseudomonas aeruginosa, and Acinetobacter baumannii, and stable isotope probing in biological samples of feces, saliva, sewage, and sputum. We utilized mono- and co-cultures to identify volatile production from individual bacterial species or in combination with stable isotope probing to identify the active metabolism of microbes from the biological samples. To extract the VOCs, we employed vacuum assisted sorbent extraction (VASE). VASE is an easy-to-use commercialized solvent-free headspace extraction method for semi-volatile and volatile compounds. The lack of solvents and the near vacuum conditions used make developing a method relatively easy and fast when compared to other extraction options like tert-butylation and solid phase microextraction. However, VASE does not work on nonvolatile compounds, thus excluding many protein analyses and heavy stable isotope labelling experiments. Using the workflow described here, we identified volatile signatures from mono- and co-cultures indicating there were volatiles specific to certain microbes or co-cultures. Furthermore, analysis of the stable isotope probing of biological samples identified VOCs that were either commonly or uniquely produced from the different human derived biological samples. Here we present the general workflow and experimental considerations of VASE. SUMMARYFrom this protocol, readers will be able to extract volatile organic compounds from a biological sample with the vacuum assisted sorbent extraction method, run samples on a GC-MS with the use of the Entech autosampler, and analyze data.

Source connections

Explore related subjects

biochemistryPreprint

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

Phan, J., Kapcia, J., Rodriguez, C. I., Vogel, V. L., Dunham, S. J. B., Whiteson, K.. 2021-02-18. Capturing actively produced microbial volatile organic compounds from human associated samples with vacuum assisted sorbent extraction. https://doi.org/10.1101/2021.02.16.431476

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

The structures of two intermediates en route to DNA repair within a (6-4) photolyase

Pyrimidine (6-4) pyrimidone photoproducts (6-4PPs) are mutagenic DNA lesions produced when UV light crosslinks two adjacent pyrimidine bases. These lesions must be repaired to maintain genome integrity, a function executed by the (6-4) photolyase enzymes. This repair chemistry is complex, involving the transfer of a hydroxyl group from one base to another, accompanied or followed by cleavage of an interbase bond. It remains unclear how the enzyme orchestrates this reaction. Typically, models invoke a stepwise process in which an oxetane intermediate is transiently populated, but despite over thirty years of investigation, such an intermediate has never been definitively observed. Conducting dose-controlled X-ray crystallography on the (6-4) photolyase from Vibrio cholerae, we observed the transformation of 6-4PP into an oxetane species. Furthermore, we identified a previously unanticipated hemi-cleaved intermediate. The progressive transformation of 6-4PP into oxetane, hemi-cleaved, and fully repaired species requires a reexamination of the existing models of DNA repair by (6-4) photolyases.

biochemistry↗
bioRxiv2026

Endothelial protein C receptor is an entry receptor for Langya virus

Langya virus (LayV) was recently discovered in infected individuals with febrile illness and classified as a Parahenipavirus, a group of poorly understood pathogens. LayV is distantly related to Nipah henipavirus which causes recurring outbreaks with case fatality rates reaching up to 90%. No vaccines or therapeutics are approved for use in humans for any parahenipavirus or henipavirus despite their broad geographic distribution and pandemic potential. Here, we discovered that human endothelial protein C receptor (EPCR) is an entry receptor for LayV and that the tetrameric LayV attachment glycoprotein (G) binds EPCR avidly. We reveal the molecular basis of LayV G recognition of the EPCR receptor and provide a blueprint of the determinants of receptor ortholog usage. We found that human and shrew EPCR orthologs promote cell entry of LayV and of the closely related Mojiang pseudoviruses. These results along with the detection of several Parahenipaviruses in shrews set these pathogens apart from Henipaviruses, which use Pteropid fruit bats as reservoir hosts. Our data support future surveillance efforts, by enabling prediction of EPCR utilization among newly discovered Parahenipaviruses, and the development and evaluation of countermeasures targeting this emerging group of human pathogens.

biochemistry↗
bioRxiv2026

RNA-targeted aminopyridine compounds provide a new antifungal strategy

While RNA is an emerging therapeutic target class, progress is limited by a shortage of well-defined targets in which ligand binding is coupled to measurable function. The group I intron, a self-splicing ribozyme essential in fungal mitochondria but absent from humans, offers both a tractable platform for exploring first principles of small molecule-RNA targeting and a selective antifungal target. We recalibrated an intron-splicing assay to approximate cellular conditions and performed an extensive, cryo-EM-guided structure-activity relationship campaign against the Candida albicans mtLSU intron. Ligands containing a flexible diamine outperformed conformationally constrained analogues, suggesting a general principle for engaging dynamic RNA surfaces and yielding lead aminopyridine 56 (IC 21.6 nM). Compound 56 synergizes with amphotericin B against C. albicans, suggesting a path for combination therapies. RT-qPCR confirmed direct target engagement in cells. This work establishes the fungal group I intron as a viable, mechanistically distinct antifungal target and a model platform for RNA-targeted discovery.

biochemistry↗