Genomic, Transcriptomic and Metabolomic Studies of Two Well-Characterized, Laboratory-Derived Vancomycin-Intermediate Staphylococcus aureus Strains Derived from the Same Parent Strain

Abstract

Complete genome comparisons, transcriptomic and metabolomic studies were performed on two laboratory-selected, well-characterized vancomycin-intermediate Staphylococcus aureus (VISA) derived from the same parent MRSA that have changes in cell wall composition and decreased autolysis. A variety of mutations were found in the VISA, with more in strain 13136p−m+V20 (vancomycin MIC = 16 µg/mL) than strain 13136p−m+V5 (MIC = 8 µg/mL). Most of the mutations have not previously been associated with the VISA phenotype; some were associated with cell wall metabolism and many with stress responses, notably relating to DNA damage. The genomes and transcriptomes of the two VISA support the importance of gene expression regulation to the VISA phenotype. Similarities in overall transcriptomic and metabolomic data indicated that the VISA physiologic state includes elements of the stringent response, such as downregulation of protein and nucleotide synthesis, the pentose phosphate pathway and nutrient transport systems. Gene expression for secreted virulence determinants was generally downregulated, but was more variable for surface-associated virulence determinants, although capsule formation was clearly inhibited. The importance of activated stress response elements could be seen across all three analyses, as in the accumulation of osmoprotectant metabolites such as proline and glutamate. Concentrations of potential cell wall precursor amino acids and glucosamine were increased in the VISA strains. Polyamines were decreased in the VISA, which may facilitate the accrual of mutations. Overall, the studies confirm the wide variability in mutations and gene expression patterns that can lead to the VISA phenotype.

Challenges and emergent solutions for LC‐MS/MS based untargeted metabolomics in diseases

“…143 Integration of transcriptomics and metabolomics is another form of multi-tier 'omics. [144][145][146] This transcriptomics-based method uses a "guilt-by-association" approach-an assumption that relies on coexpression of genes, known and unknown, that operate in the same…”

VraR Binding to the Promoter Region of agr Inhibits Its Function in Vancomycin-Intermediate Staphylococcus aureus (VISA) and Heterogeneous VISA

“…A link has also been implied between the acquisition of vancomycin resistance in S. aureus and attenuated virulence (10,26,27,29,32). In our study, vancomycin resistance was induced in three strains of S. aureus, including Mu3 and Mu50, which both carry the mutation in vraS(I5N) and result in replacement of isoleucine at residue 5 of the protein with asparagine (31,33).…”

“…Our previous report has also demonstrated that VISA/hVISA strains have reduced coagulase activity and reduced or no hemolysis (28). Hattangady et al performed complete genome comparison, along with transcriptomic and metabolomic studies, of two laboratory-selected VISA strains and found that expression of surface-associated virulence determinants was decreased in VISA isolates (29). Majcherczyk et al and Peleg et al used a rat model and a Galleria mellonella model, respectively, to show that virulence and infectivity of VISA/hVISA strains attenuate as the vancomycin MIC for the strains increases (26,27).…”

Proteomic and Metabolomic Analyses of a Tea-Tree Oil-Selected Staphylococcus aureus Small Colony Variant

“…Metabolites were extracted from both SH1000 and SH1000-TTORS-1 in triplicate as previously described [73]. Briefly, overnight cultures were used to inoculate 125 mL of LB (initial OD 580 = 0.01) and incubated (37 • C, 200 rpm) until an OD 580 = 0.07 was reached.…”

“…Metabolic quenching was achieved by adding 500 µL of cold methanol (−20 • C) and all samples were stored at −80 • C. Metabolite analysis was carried out by the Roy J. Carver Biotechnology Center, University of Illinois at Urbana-Champaign, Urbana, IL, USA as previously described [73]. Metabolite relative concentrations were normalized using 100 mg of dry cell weight and statistically significant alterations in metabolite levels were based upon a Student's t-test p-value < 0.05 and reported as the average ± standard error of the mean for all three biological replicates.…”