Microbial metabolomics' latest SICRIT: Soft ionization by Chemical Reaction in-Transfer mass spectrometry.

Microbial metabolomics studies are a common approach to identifying microbial strains that have a capacity to produce new chemistries both in vitro and in situ. A limitation to applying microbial metabolomics to the discovery of new chemical entities is the rediscovery of known compounds, or "known unknowns." One contributing factor to this rediscovery is the majority of laboratories use one ionization source-electrospray ionization (ESI)-to conduct metabolomics studies. Although ESI is an efficient, widely adopted ionization method, its widespread use may contribute to the re-identification of known metabolites. Here, we present the use of a dielectric barrier discharge ionization (DBDI) for microbial metabolomics applications through the use of soft ionization chemical reaction in-transfer (SICRIT). Additionally, we compared SICRIT to ESI using two different Vibrio species-Vibrio fischeri, a symbiotic marine bacterium, and Vibrio cholerae, a pathogenic bacterium. Overall, we found that the SICRIT source ionizes a different set of metabolites than ESI, and it has the ability to ionize lipids more efficiently than ESI in positive mode. This work highlights the value of using more than one ionization source for the detection of metabolites.

A Label-Free Approach for Relative Spatial Quantitation of c-di-GMP in Microbial Biofilms.

Microbial biofilms represent an important lifestyle for bacteria and are dynamic three-dimensional structures. Cyclic dimeric guanosine monophosphate (c-di-GMP) is a ubiquitous signaling molecule that is known to be tightly regulated with biofilm processes. While measurements of global levels of c-di-GMP have proven valuable toward understanding the genetic control of c-di-GMP production, there is a need for tools to observe the local changes of c-di-GMP production in biofilm processes. We have developed a label-free method for the direct detection of c-di-GMP in microbial colony biofilms using matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI). We applied this method to the enteric pathogen Vibrio cholerae, the marine symbiont V. fischeri, and the opportunistic pathogen Pseudomonas aeruginosa PA14 and detected spatial and temporal changes in c-di-GMP signal that accompanied genetic alterations in factors that synthesize and degrade the compound. We further demonstrated how this method can be simultaneously applied to detect additional metabolites of interest from a single sample.

Reducing Mass Confusion over the Microbiome.

As genetic tools continue to emerge and mature, more information is revealed about the identity and diversity of microbial community members. Genetic tools can also be used to make predictions about the chemistry that bacteria and fungi produce to function and communicate with one another and the host. Ongoing efforts to identify these products and link genetic information to microbiome chemistry rely on analytical tools. This tutorial highlights recent advancements in microbiome studies driven by techniques in mass spectrometry.

A label-free approach for relative spatial quantitation of c-di-GMP in microbial biofilms.

Microbial biofilms represent an important lifestyle for bacteria and are dynamic three dimensional structures. Cyclic dimeric guanosine monophosphate (c-di-GMP) is a ubiquitous signaling molecule that is known to be tightly regulated with biofilm processes. While measurements of global levels of c-di-GMP have proven valuable towards understanding the genetic control of c-di-GMP production, there is a need for tools to observe the local changes of c-di-GMP production in biofilm processes. We have developed a label-free method for the direct detection of c-di-GMP in microbial colony biofilms using matrix-assisted laser desorption ionization mass spectrometry imaging (MALDI-MSI). We applied this method to the enteric pathogen Vibrio cholerae, the marine symbiont V. fischeri, and the opportunistic pathogen Pseudomonas aeruginosa PA14 and detected spatial and temporal changes in c-di-GMP signal that accompanied genetic alterations in factors that synthesize and degrade the compound. We further demonstrated how this method can be simultaneously applied to detect additional metabolites of interest in a single experiment.