Minor Differences in the Proteome of Bacillus subtilis and Bacillus mojavensis Based upon High Abundance/ Conserved Protein Mass Spectra; Implications for Rapid, Improved Identification of Two Pathogen Genetically Closely Related

Chambers, Timothy; Culak, Renata; Gharbia, Saheer E.; Shah, Haroun N.

doi:10.4172/2470-1289.1000119

Cited by 5 publications

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“…This technique is elusive about the quantitative detection and distribution of surface carbohydrates at the single cell level; however, it is limited to targets that have available specific binding ligands and it might be more promising if targets in addition to carbohydrates are considered. Regarding fatty acid profiles, B. cytotoxicus and B. pseudomycoides in particular contain different amounts of certain major fatty acids, 2,29,65 whereas no specific pattern or diagnostic markers could be identified among the other B. cereus group species.Our data support findings from studies that have revealed limits of classical MALDI-TOF MS biotyping to discriminate species belonging to the Bacillus genus45,46 (Table 3). Studies that have focused on Bacillus classification based on MALDI-TOF MS analysis examined strains predominantly classified as B. anthracis, B. cereus, B. thuringiensis, and B. subtilis species.…”

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confidence: 90%

“…44 Furthermore, mass spectra quality depends on the physiological state of the bacilli as progression of sporulation; that is, the presence of spores, can suppress several of the few characteristic mass ions. 45,46 The objective of this study was to assign type strain-specific diagnostic peptides to the various members of the B. cereus group. We analyzed in silico-generated tryptic peptides to estimate the potential number of type strain-specific diagnostic peptides and reconstructed a phylogenetic tree based on the clustering of theoretical precursor masses.…”

Section: ■ Introductionmentioning

confidence: 99%

“…Matrix-assisted laser desorption/ionization-time-of-flight (MALDI-TOF) biotyping is a valuable and well-established technique to rapidly identify bacterial species. − As the members of the B. cereus group are genetically very similar, MALDI biotyping results are reliable on genus but not on species level . Furthermore, mass spectra quality depends on the physiological state of the bacilli as progression of sporulation; that is, the presence of spores, can suppress several of the few characteristic mass ions. , …”

Section: Introductionmentioning

confidence: 99%

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Bacillus cereus Group-Type Strain-Specific Diagnostic Peptides

et al. 2016

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The Bacillus cereus group consists of eight very closely related species and comprises both harmless and human pathogenic species such as Bacillus anthracis, Bacillus cereus, and Bacillus cytotoxicus. Numerous efforts have been undertaken to allow presumptive differentiation of B. cereus group species from one another. However, methods to rapidly and accurately distinguish these species are currently lacking. We confirmed that classical matrix-assisted laser desorption/ionization-time-of-flight (MALDI-TOF) biotyping cannot achieve reliable identification of each type strain. We therefore assigned type strain-specific diagnostic peptides to the B. cereus group based on comparisons of their proteomic profiles. The number of diagnostic peptides varied remarkably in a type strain-dependent manner. The accuracy of the reference database was crucial to validate candidate diagnostic peptides and led to a noteworthy reduction of verified diagnostic peptides. Diagnostic peptides ranged from one for B. weihenstephanensis to 62 for B. pseudomycoides and were associated with proteins involved in diverse biological processes, e.g. amino acid biosynthesis, cell envelope, cellular processes, energy metabolism, and transport processes. However, 45.6% of all diagnostic peptides comprised currently unclassified proteins or proteins of unknown function. In addition, a phylogenetic tree based on clustering of theoretical precursor masses deduced from in silico-generated tryptic peptides was reconstructed.

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confidence: 90%

Section: ■ Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Bacillus cereus Group-Type Strain-Specific Diagnostic Peptides

et al. 2016

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“…As a solution for this problem Lasch et al [42] proposed the protein enrichment protocol where application of combined TFA treatment promoted the protein isolation from spores. However, this analytical solution does not prevent the sample from isolation of protein from mixed bacterial culture (different time and stage of sporulation), on the one hand, leading to an increase of proteins signals in MALDI spectra, but on the other hand causing misidentification [50]. However, there are some reports which suggest that in the case of Bacillus strains, species identification could be also performed based only on the protein profiling of their spores using both top-down and bottom-up approaches which was proved for such species as B .…”

Section: Discussionmentioning

confidence: 99%

Analysis of bacteria associated with honeys of different geographical and botanical origin using two different identification approaches: MALDI-TOF MS and 16S rDNA PCR technique

et al. 2019

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In the presented work identification of microorganisms isolated from various types of honeys was performed. Martix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and 16S rDNA sequencing were applied to study environmental bacteria strains.With both approches, problematic spore-forming Bacillus spp , but also Staphylococcus spp ., Lysinibacillus spp ., Micrococcus spp . and Brevibacillus spp were identified. However, application of spectrometric technique allows for an unambiguous distinction between species/species groups e.g. B . subtilis or B . cereus groups. MALDI TOF MS and 16S rDNA sequencing allow for construction of phyloproteomic and phylogenetic trees of identified bacterial species. Furthermore, the correlation beetween physicochemical properties, geographical and botanical origin and the presence bacterial species in honey samples were investigated.

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“… Huang and Huang (2018) used a MALDI-TOF MS instrument (Microflex LT, Bruker Daltonics) to rapidly identify L. casei group strains at the species and subspecies-level by using ethanol/formic acid/acetonitrile for protein extraction in a routine protocol ( Chambers et al, 2015 ) with MS peaks analyzed using ClinProTools software (Bruker Daltonics). Type strains of the L. casei group were used to construct main spectral profiles (the highest quality spectra) for compiling an in-house database given the optimized culture conditions [cultivated anaerobically at 37°C on de Man, Rogosa and Sharpe (MRS) agar for 20 h].…”

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confidence: 99%

Identification and Classification for the Lactobacillus casei Group

Huang

et al. 2018

Front. Microbiol.

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Lactobacillus casei, Lactobacillus paracasei, and Lactobacillus rhamnosus are phenotypically and genotypically closely related, and together comprise the L. casei group. Although the strains of this group are commercially valuable as probiotics, the taxonomic status and nomenclature of the L. casei group have long been contentious because of the difficulties in identifying these three species by using the most frequently used genotypic methodology of 16S rRNA gene sequencing. Long used as the gold standard for species classification, DNA–DNA hybridization is laborious, requires expert skills, and is difficult to use routinely in laboratories. Currently, genome-based comparisons, including average nucleotide identity (ANI) and digital DNA–DNA hybridization (dDDH), are commonly applied to bacterial taxonomy as alternatives to the gold standard method for the demarcating phylogenetic relationships. To establish quick and accurate methods for identifying strains in the L. casei group at the species and subspecies levels, we developed species- and subspecies-specific identification methods based on housekeeping gene sequences and whole-cell matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) spectral pattern analysis. By phylogenetic analysis based on concatenated housekeeping gene sequences (dnaJ, dnaK, mutL, pheS, and yycH), 53 strains were separated into four clusters corresponding to the four species: L. casei, L. paracasei and L. rhamnosus, and Lactobacillus chiayiensis sp. nov. A multiplex minisequencing assay using single nucleotide polymorphism (SNP)-specific primers based on the dnaK gene sequences and species-specific primers based on the mutL gene sequences provided high resolution that enabled the strains at the species level to be identified as L. casei, L. paracasei, and L. rhamnosus. By MALDI-TOF MS analysis coupled with an internal database and ClinProTools software, species- and subspecies-level L. casei group strains were identified based on reliable scores and species- and subspecies-specific MS peaks. The L. paracasei strains were distinguished clearly at the subspecies level based on subspecies-specific MS peaks. This article describes the rapid and accurate methods used for identification and classification of strains in the L. casei group based on housekeeping gene sequences and MALDI-TOF MS analysis as well as the novel speciation of this group including L. chiayiensis sp. nov. and ‘Lactobacillus zeae’ by genome-based methods.

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Minor Differences in the Proteome of Bacillus subtilis and Bacillus mojavensis Based upon High Abundance/ Conserved Protein Mass Spectra; Implications for Rapid, Improved Identification of Two Pathogen Genetically Closely Related

Cited by 5 publications

References 0 publications

Bacillus cereus Group-Type Strain-Specific Diagnostic Peptides

Bacillus cereus Group-Type Strain-Specific Diagnostic Peptides

Analysis of bacteria associated with honeys of different geographical and botanical origin using two different identification approaches: MALDI-TOF MS and 16S rDNA PCR technique

Identification and Classification for the Lactobacillus casei Group

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