Dataset Information

Testing long-term (up to 4 weeks) RNAlater preservation of field collected animal-microbe symbioses at room temperature for metaproteomic analyses

ABSTRACT: When studying gene expression in microbe-animals symbioses collected in the field it is essential to quickly and efficiently preserve in situ symbiont and host gene expression patterns. One of the most commonly used sample preservation methods for samples targeted for proteomic analyses is flash freezing, however, liquid nitrogen or dry ice needed for flash freezing are often not available at remote field sites. We first tested if RNAlater allows to preserve proteins in animal-microbe symbioses as efficiently as flash freezing and without introducing issues with downstream processing (see PXD014591). Second, for the data in this PRIDE submission we tested if RNAlater preserves protein expression patterns over time at room temperature. We used the marine gutless oligochaete Olavius algarvensis as a test case. Olavius algarvensis lives in shallow water sediments off the coast of Elba, Italy. It has no digestive and excretory system and harbors five bacterial symbionts that fulfill its nutritional and waste recycling needs (Kleiner et al., 2012, PNAS 109(19):1173-82). For this dataset, we fixed a total of 33 worms and incubated them in RNAlater for up to 4 weeks. We then evaluated proteome preservation quality in terms of the number of identified proteins, abundances of individual proteins and potential biases against specific protein or taxonomic groups. Out of this 33 samples, eleven worms were incubated for 24 hours in RNAlater at 4°C (t0), while the other worms were incubated in RNAlater at room temperature (21-23°C) for additional 24 hours (t1, 6 worms), one week (t2, 8 worms), and four weeks (t3, 8 worms). We removed RNAlater from the worms after incubation and froze the samples at -80°C.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Olavius Algarvensis Gamma 3 Endosymbiont Olavius Algarvensis Chromatiales Bacterium Oalggamma1 Olavius Algarvensis Gamma 1 Endosymbiont Olavius Algarvensis Delta 4 Endosymbiont Spirochaeta Sp. Elba Olavius Algarvensis Sulfate-reducing Endosymbiont Olavius Algarvensis Spirochete Endosymbiont Olavius Algarvensis Delta 1 Endosymbiont

SUBMITTER: Marlene Jensen

LAB HEAD: Dr. Manuel kleiner

PROVIDER: PXD014881 | Pride | 2021-11-03

REPOSITORIES: Pride

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Publications

Evaluation of RNAlater as a Field-Compatible Preservation Method for Metaproteomic Analyses of Bacterium-Animal Symbioses.

Jensen Marlene M Wippler Juliane J Kleiner Manuel M

Microbiology spectrum 20211027 2

Field studies are central to environmental microbiology and microbial ecology, because they enable studies of natural microbial communities. Metaproteomics, the study of protein abundances in microbial communities, allows investigators to study these communities "in situ," which requires protein preservation directly in the field because protein abundance patterns can change rapidly after sampling. Ideally, a protein preservative for field deployment works rapidly and preserves the whole ...[more]

PMID: 34704828

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Project description:BACKGROUND: Stool metabolites provide essential insights into the function of the gut microbiome. The current gold standard for storage of stool samples for metabolomics is flash-freezing at - 80 °C which can be inconvenient and expensive. Ambient temperature storage of stool is more practical, however no available methodologies adequately preserve the metabolomic profile of stool. A novel sampling kit (OMNImet.GUT; DNA Genotek, Inc.) was introduced for ambient temperature storage and stabilization of feces for metabolomics; we aimed to test the performance of this kit vs flash-freezing. To do this stool was collected from an infant's diaper was divided into 2 aliquots: 1) flash-frozen and 2) stored in an OMNImet.GUT tube at ambient temperature for 3-4 days. Samples from the same infant were collected at 2 different timepoints to assess metabolite changes over time. Subsequently, all samples underwent metabolomic analysis by liquid chromatography-tandem mass spectrometry (LC-MS/MS).RESULTS: Paired fecal samples (flash-frozen and ambient temperature) from 16 infants were collected at 2 timepoints (32 individual samples, 64 aliquots). Similar numbers of metabolites were detected in both the frozen and ambient temperature samples (1126 in frozen, 1107 in ambient temperature, 1064 shared between sample types). Metabolite abundances were strongly correlated between storage methods (median Spearman correlation Rs = 0.785 across metabolites). Hierarchical clustering analysis and principal component analysis showed that samples from the same individuals at a given timepoint clustered closely, regardless of the storage method. Repeat samples from the same individual were compared by paired t-test, separately for the frozen and OMNImet.GUT. The number of metabolites in each biochemical class that significantly changed (p < 0.05) at timepoint 2 relative to timepoint 1 was similar in flash-frozen vs ambient temperature storage. Changes in microbiota modified metabolites over time were also consistent across both methodologies.CONCLUSION: Ambient temperature storage and stabilization of stool in the OMNImet.GUT device yielded comparable metabolomic results to flash freezing in terms of 1) the identity and abundance of detected biochemicals 2) the distinct metabolomic profiles of subjects and 3) changes in metabolites over time that are plausibly microbiota-induced. This method potentially provides a more convenient, less expensive home collection and storage option for stool metabolomic analysis.

Dataset Information

Testing long-term (up to 4 weeks) RNAlater preservation of field collected animal-microbe symbioses at room temperature for metaproteomic analyses

Publications

Evaluation of RNA<i>later</i> as a Field-Compatible Preservation Method for Metaproteomic Analyses of Bacterium-Animal Symbioses.

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