Primer on Agar-Based Microbial Imaging Mass Spectrometry

Yang, Jianping; Phelan, Vanessa V.; Simkovsky, Ryan; Watrous, Jeramie D.; Trial, Rachelle M; Fleming, Tinya C.; Wenter, Roland; Moore, Bradley S.; Golden, Susan S.; Pogliano, Kit; Dorrestein, Pieter C.

doi:10.1128/jb.00823-12

Cited by 146 publications

(204 citation statements)

References 73 publications

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“…V isualizing microbial interactions with mass spectrometry imaging (MSI) has provided insights into many important biological processes, where it has captured molecular snapshots of metabolic exchange, antibiotic resistance, microbial competition, and more [1][2][3][4][5][6][7]. There are a number of different MSI methods reported for chemical mapping of microbial systems [3,4,6].…”

Section: Introductionmentioning

confidence: 99%

Utilizing a Robotic Sprayer for High Lateral and Mass Resolution MALDI FT-ICR MSI of Microbial Cultures

Anderton

Chu

Tolić

et al. 2016

J. Am. Soc. Mass Spectrom.

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Abstract. The ability to visualize biochemical interactions between microbial communities using MALDI MSI has provided tremendous insights into a variety of biological fields. Matrix application using a sieve proved to be incredibly useful, but it has many limitations that include uneven matrix coverage and limitation in the types of matrices that could be employed in studies. Recently, there has been a concerted effort to improve matrix application for studying agar plated microbial cultures, many of which utilized automated matrix sprayers. Here, we describe the usefulness of using a robotic sprayer for matrix application. The robotic sprayer has twodimensional control over where matrix is applied, and a heated capillary that allows for rapid drying of the applied matrix. This method provided a significant increase in MALDI sensitivity over the sieve method, as demonstrated by FT-ICR MS analysis, facilitating the ability to gain higher lateral resolution MS images of Bacillus subtilis than previously reported. This method also allowed for the use of different matrices to be applied to the culture surfaces.

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Section: Introductionmentioning

confidence: 99%

Utilizing a Robotic Sprayer for High Lateral and Mass Resolution MALDI FT-ICR MSI of Microbial Cultures

Anderton

Chu

Tolić

et al. 2016

J. Am. Soc. Mass Spectrom.

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“…Thin-layer BG11 and BG11 0 agar plates of N. punctiforme wild-type and pks2 − mutant monocultures or bacterial interactions were inoculated from fresh liquid cultures and incubated for 10 d (29). Sample preparation, matrix application, and dehydration were conducted as previously described (30). For MALDI imaging experiments of plant cyanobacteria symbiosis, G. manicata plants harboring the symbiotic glands were cut into thin slices.…”

mentioning

confidence: 99%

Nostopeptolide plays a governing role during cellular differentiation of the symbiotic cyanobacterium Nostoc punctiforme

Liaimer

Helfrich

Hinrichs

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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Nostoc punctiforme is a versatile cyanobacterium that can live either independently or in symbiosis with plants from distinct taxa. Chemical cues from plants and N. punctiforme were shown to stimulate or repress, respectively, the differentiation of infectious motile filaments known as hormogonia. We have used a polyketide synthase mutant that accumulates an elevated amount of hormogonia as a tool to understand the effect of secondary metabolites on cellular differentiation of N. punctiforme. Applying MALDI imaging to illustrate the reprogramming of the secondary metabolome, nostopeptolides were identified as the predominant difference in the pks2 − mutant secretome. Subsequent differentiation assays and visualization of cell-type-specific expression of nostopeptolides via a transcriptional reporter strain provided evidence for a multifaceted role of nostopeptolides, either as an autogenic hormogonium-repressing factor or as a chemoattractant, depending on its extracellular concentration. Although nostopeptolide is constitutively expressed in the free-living state, secreted levels dynamically change before, during, and after the hormogonium differentiation phase. The metabolite was found to be strictly down-regulated in symbiosis with Gunnera manicata and Blasia pusilla, whereas other metabolites are up-regulated, as demonstrated via MALDI imaging, suggesting plants modulate the fine-balanced cross-talk network of secondary metabolites within N. punctiforme.

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“…The matrix was applied to wet agar by sieving through a 53 μm stainless steel sieve until the surface was covered with matrix powder as previously described [12]. A second round of matrix was applied after 1 h of drying at 40°C followed by a final drying step of 2 h at 40°C.…”

Section: Sieving Techniquementioning

confidence: 99%

“…Consequently, several sample preparation techniques may be used, mainly based on methods established for tissue imaging [9]. Following the approach of applying dry matrix on a tissue sample [10,11], sieving matrix powder onto wet agar samples has become a common sample preparation technique for microbial IMS in our lab [12]. This sieving method has been proven to be useful in most studies and certainly benefits from its simple and straight-forward procedure [12].…”

Section: Introductionmentioning

confidence: 99%

“…Following the approach of applying dry matrix on a tissue sample [10,11], sieving matrix powder onto wet agar samples has become a common sample preparation technique for microbial IMS in our lab [12]. This sieving method has been proven to be useful in most studies and certainly benefits from its simple and straight-forward procedure [12]. Vergeiner et al have recently shown a modification to this approach by spraying a matrix suspension onto wet agar, which resulted in a more homogeneous matrix coverage and better sensitivity for a specific metabolite [13].…”

Section: Introductionmentioning

confidence: 99%

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Homogeneous Matrix Deposition on Dried Agar for MALDI Imaging Mass Spectrometry of Microbial Cultures

Hoffmann

Dorrestein

2015

J. Am. Soc. Mass Spectrom.

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Abstract. Matrix deposition on agar-based microbial colonies for MALDI imaging mass spectrometry is often complicated by the complex media on which microbes are grown. This Application Note demonstrates how consecutive short spray pulses of a matrix solution can form an evenly closed matrix layer on dried agar. Compared with sieving dry matrix onto wet agar, this method supports analyte cocrystallization, which results in significantly more signals, higher signal-to-noise ratios, and improved ionization efficiency. The even matrix layer improves spot-to-spot precision of measured m/z values when using TOF mass spectrometers. With this technique, we established reproducible imaging mass spectrometry of myxobacterial cultures on nutrient-rich cultivation media, which was not possible with the sieving technique.

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Primer on Agar-Based Microbial Imaging Mass Spectrometry

Cited by 146 publications

References 73 publications

Utilizing a Robotic Sprayer for High Lateral and Mass Resolution MALDI FT-ICR MSI of Microbial Cultures

Utilizing a Robotic Sprayer for High Lateral and Mass Resolution MALDI FT-ICR MSI of Microbial Cultures

Nostopeptolide plays a governing role during cellular differentiation of the symbiotic cyanobacterium Nostoc punctiforme

Homogeneous Matrix Deposition on Dried Agar for MALDI Imaging Mass Spectrometry of Microbial Cultures

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