Tandem mass spectrometry proteotyping of extremophiles sampled from high-altitude Andean lakes
Ontology highlight
ABSTRACT:
INSTRUMENT(S): Q Exactive HF
ORGANISM(S): Unclassified Chromatiaceae
SUBMITTER: 
Jean ARMENGAUD
LAB HEAD: Jean Armengaud
PROVIDER: PXD044759 | Pride | 2024-05-02
REPOSITORIES: Pride
Publications
Exploring Andean High-Altitude Lake Extremophiles through Advanced Proteotyping.
Runzheimer Katharina K Lozano Clément C Boy Diana D Boy Jens J Godoy Roberto R Matus Francisco J FJ Engel Denise D Pavletic Bruno B Leuko Stefan S Armengaud Jean J Moeller Ralf R
Journal of proteome research 20240220 3
Quickly identifying and characterizing isolates from extreme environments is currently challenging while very important to explore the Earth's biodiversity. As these isolates may, in principle, be distantly related to known species, techniques are needed to reliably identify the branch of life to which they belong. Proteotyping these environmental isolates by tandem mass spectrometry offers a rapid and cost-effective option for their identification using their peptide profiles. In this study, we ...[more]
Similar Datasets
Project description:Tandem mass spectrometry-based proteotyping allows characterizing microorganisms in terms of taxonomy and is becoming an important tool for investigating microbial diversity from several ecosystems. Fast and automatable sample preparation for obtaining peptide pools amenable to tandem mass spectrometry is necessary for enabling proteotyping as a high-throughput method. First, the protocol to increase the yield of lysis of several representative bacterial and eukaryotic microorganisms was optimized by using a long and drastic bead-beating setting with 0.1 mm silica beads, 0.1 and 0.5 mm glass beads, in presence of detergents. Then, three different methods to obtain greater digestion yield from these extracts were tested and optimized for improve efficiency and reduce application time: denaturing electrophoresis of proteins and in-gel proteolysis, suspension-trapping filter-based approach (S-Trap) and, solid-phase-enhanced sample preparation named SP3. The latter method outperforms the other two in terms of speed and delivers also more peptides and proteins than with the in-gel proteolysis (2.2 fold for both) and S-trap approaches (1.3 and 1.2 fold, respectively). Thus, SP3 directly improves tandem mass spectrometry proteotyping.
| S-EPMC6742703 | biostudies-literature
Project description:Mass spectrometry (MS) and proteomics offer comprehensive characterization and identification of microorganisms and discovery of protein biomarkers that are applicable for diagnostics of infectious diseases. The use of biomarkers for diagnostics is widely applied in the clinic and the use of peptide biomarkers is increasingly being investigated for applications in the clinical laboratory. Respiratory-tract infections are a predominant cause for medical treatment, although, clinical assessments and standard clinical laboratory protocols are time-consuming and often inadequate for reliable diagnoses. Novel methods, preferably applied directly to clinical samples, excluding cultivation steps, are needed to improve diagnostics of infectious diseases, provide adequate treatment and reduce the use of antibiotics and associated development of antibiotic resistance. This study applied nano-liquid chromatography (LC) coupled with tandem MS, with a bioinformatics pipeline and an in-house database of curated high-quality reference genome sequences to identify species-unique peptides as potential biomarkers for four bacterial pathogens commonly found in respiratory tract infections (RTIs): Staphylococcus aureus; Moraxella catarrhalis; Haemophilus influenzae and Streptococcus pneumoniae The species-unique peptides were initially identified in pure cultures of bacterial reference strains, reflecting the genomic variation in the four species and, furthermore, in clinical respiratory tract samples, without prior cultivation, elucidating proteins expressed in clinical conditions of infection. For each of the four bacterial pathogens, the peptide biomarker candidates most predominantly found in clinical samples, are presented. Data are available via ProteomeXchange with identifier PXD014522. As proof-of-principle, the most promising species-unique peptides were applied in targeted tandem MS-analyses of clinical samples and their relevance for identifications of the pathogens, i.e. proteotyping, was validated, thus demonstrating their potential as peptide biomarker candidates for diagnostics of infectious diseases.
| S-EPMC7050107 | biostudies-literature
Project description:Zoonotic pathogens that can be transmitted via food to humans have a high potential for large-scale emergencies, comprising severe effects on public health, critical infrastructures, and the economy. In this context, the development of laboratory methods to rapidly detect zoonotic bacteria in the food supply chain, including high-resolution mass spectrometry proteotyping are needed. In this work, an optimized sample preparation method for liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based proteome profiling was established for Francisella isolates, and a cluster analysis, as well as a phylogenetic tree, was generated to shed light on evolutionary relationships. Furthermore, this method was applied to tissues of infected hare carcasses from Germany. Even though the non-informative data outnumbered by a manifold the information of the zoonotic pathogen in the resulting proteome profiles, the standardized evaluation of MS data within an established automated analysis pipeline identified Francisella (F.) tularensis and, thus, could be, in principle, an applicable method to monitor food supply chains.
| S-EPMC7225524 | biostudies-literature
Project description:A comparison of the performance when using X!Tandem with a 1) large, exhaustive, reference database; and 2) small, specialized, database.
2017-04-21 | PXD005924 | Pride
Project description:Background: Methane yield and biogas productivity of biogas plants depend on microbial community structure and functionality, substrate supply, and general process parameters. Little is known, however, about the correlations between microbial community function and the process parameters. To close this knowledge gap the microbial community of 40 industrial biogas plants was evaluated by a metaproteomics approach in this study. Results: Liquid chromatography coupled to tandem mass spectrometry (Elite Hybrid Ion Trap Orbitrap) enabled the identification of 3138 metaproteins belonging to 162 biological processes and 75 different taxonomic orders. Therefore, database searches were performed against UniProtKB/Swiss-Prot and several metagenome databases. Subsequent clustering and principal component analysis of these data allowed to identify four main clusters associated to mesophilic and thermophilic process conditions, upflow anaerobic sludge blanket reactors and sewage sludge as substrate. Observations confirm a previous phylogenetic study of the same biogas plant samples that was based on 16S-rRNA gene by De Vrieze et al. (2015) (De Vrieze, Saunders et al. 2015). Both studies described similar microbial key players of the biogas process, namely Bacillales, Enterobacteriales, Bacteriodales, Clostridiales, Rhizobiales and Thermoanaerobacteriales as well as Methanobacteriales, Methanosarcinales and Methanococcales. In addition, a correlation study and a Gephi graph network based on the correlations between the taxonomic orders and process parameters suggested the presence of various trophic interactions, e.g. syntrophic hydrogen transfer between Thermoanaerobacteriales and Methanomicrobiales. For the elucidation of the main biomass degradation pathways the most abundant 1% of metaproteins were assigned to the KEGG map 1200 representing the central carbon metabolism. Additionally, the effect of the process parameters (i) temperature, (ii) organic loading rate (OLR), (iii) total ammonia nitrogen (TAN) and (iv) sludge retention time (SRT) on these pathways was investigated. For example high TAN correlated with hydrogenotrophic methanogens and bacterial one-carbon metabolism, indicating syntrophic acetate oxidation. Conclusion: This study shows the benefit of large-scale proteotyping of biogas plants, enabling the identification of general correlations between the process parameters and the microbial community structure and function. Changes in the level of microbial key functions or even in the microbial community type represent a valuable hint for process problems and disturbances.
2016-04-19 | PXD003526 | Pride
Project description:Campylobacter fetus is a causative agent of intestinal illness and, occasionally, severe systemic infections and meningitis. C. fetus currently comprises three subspecies: C. fetus subspecies fetus (Cff), C. fetus subspecies venerealis (Cfv), and C. fetus subspecies testudinum (Cft). Cff and Cfv are primarily associated with mammals whereas Cft is associated with reptiles. To offer an alternative to laborious sequence-based techniques such as multilocus sequence typing (MLST) and polymerase chain reaction (PCR)-ribotyping for this species, the purpose of the study was to develop a typing scheme based on proteotyping. In total, 41 representative C. fetus strains were analyzed by intact cell mass spectrometry and compared to MLST results. Biomarkers detected in the mass spectrum of C. fetus subsp. fetus reference strain LMG 6442 (NCTC 10842) as well as corresponding isoforms were associated with the respective amino acid sequences and added to the C. fetus proteotyping scheme. In combination, the 9 identified biomarkers allow the differentiation of Cft subspecies strains from Cff and Cfv subspecies strains. Biomarkers to distinguish between Cff and Cfv were not found. The results of the study show the potential of proteotyping to differentiate different subspecies, but also the limitations of the method.
| S-EPMC6563684 | biostudies-literature
Project description:Respiratory-tract infections are a predominant cause for clinical treatment, although, clinical assessments and standard clinical laboratory protocols are time-consuming and often inadequate for reliable diagnoses. Novel methods, preferably applied directly to clinical samples, excluding cultivation steps, are needed to improve diagnostics, provide adequate treatment and reduce the use of antibiotics and associated development of antibiotic resistance. This study applied nano-liquid chromatography coupled with tandem MS, with a bioinformatics pipeline and an in-house database of curated high-quality reference genome sequences to identify species-unique peptides as potential biomarkers for bacterial pathogens commonly found in respiratory tract infections: Staphylococcus aureus; Moraxella catarrhalis; Haemophilus influenzae and Streptococcus pneumoniae. The species-unique peptides were initially identified in pure-culture bacterial reference strains, reflecting the genomic variation in the four species and, furthermore in clinical respiratory tract samples, without prior cultivation, elucidating proteins expressed in clinical conditions of infection. Peptide biomarker candidates for each species is presented. As a proof-of-principle, the most promising species-unique peptides were applied in targeted tandem MS-analyses of clinical samples and their relevance for identifications of the pathogens, i.e. proteotyping, was validated, demonstrating their potential as effective peptide biomarker candidates for diagnostics of infectious diseases.
2020-01-17 | PXD014522 | Pride
Project description:A range of methodologies may be used for analyzing bacteria, depending on the purpose and the level of resolution needed. The capability for recognition of species distinctions within the complex spectrum of bacterial diversity is necessary for progress in microbiological research. In clinical settings, accurate, rapid and cost-effective methods are essential for early and efficient treatment of infections. Characterization and identification of microorganisms, using, bottom-up proteomics, or "proteotyping", relies on recognition of species-unique or associated peptides, by tandem mass spectrometry analyses, dependent upon an accurate and comprehensive foundation of genome sequence data, allowing for differentiation of species, at amino acid-level resolution. In this study, the high resolution and accuracy of MS/MS-based proteotyping was demonstrated, through analyses of the three phylogenetically and taxonomically most closely-related species of the Mitis Group of the genus Streptococcus: i.e., the pathogenic species, Streptococcus pneumoniae (pneumococcus), and the commensal species, Streptococcus pseudopneumoniae and Streptococcus mitis. To achieve high accuracy, a genome sequence database used for matching peptides was created and carefully curated. Here, MS-based, bottom-up proteotyping was observed and confirmed to attain the level of resolution necessary for differentiating and identifying the most-closely related bacterial species, as demonstrated by analyses of species of the Streptococcus Mitis Group, even when S. pneumoniae were mixed with S. pseudopneumoniae and S. mitis, by matching and identifying more than 200 unique peptides for each species.
| S-EPMC6287849 | biostudies-literature
Project description:Flash proteotyping is a methodology for ultra-fast identification of microorganisns by tandem mass spectrometry. Here, we obtained results on five reference strains and ten new bacterial isolates. The methodology is based on direct sample infusion into the mass spectromete and an original, highly sensitive procedure for data processing and taxonomic identification.
2023-12-21 | PXD045510 | Pride
Project description:Azalea belongs to Rhododendron, which is one of the largest genera of flowering plants and is well known for the diversity and beauty in its more than 1000 woody species. Rhododendron contains two distinct groups: the most high-altitude and a few low-altitude species; however, the former group is difficult to be domesticated for urban landscaping, and their evolution and adaptation are little known. Rhododendron ovatum has broad adaptation in low-altitude regions but possesses evergreen characteristics like high-altitude species, and it has floral fragrance that is deficient in most cultivars. Here we report the chromosome-level genome assembly of R. ovatum, which has a total length of 549 Mb with scaffold N50 of 41 Mb and contains 41 264 predicted genes. Genomic micro-evolutionary analysis of R. ovatum in comparison with two high-altitude Rhododendron species indicated that the expansion genes in R. ovatum were significantly enriched in defence responses, which may account for its adaptability in low altitudes. The R. ovatum genome contains much more terpene synthase genes (TPSs) compared with the species that lost floral fragrance. The subfamily b members of TPS are involved in the synthesis of sesquiterpenes as well as monoterpenes and play a major role in flora scent biosynthesis and defence responses. Tandem duplication is the primary force driving expansion of defence-responsive genes for extensive adaptability to the low-altitude environments. The R. ovatum genome provides insights into low-altitude adaptation and gain or loss of floral fragrance for Rhododendron species, which are valuable for alpine plant domestication and floral scent breeding.
| S-EPMC8633516 | biostudies-literature