Recognition of Staphylococcus aureus isolates as small colony variants applying Fourier-transform infrared spectroscopy

Abstract number: p1730

Becker  K., Al Laham  N., Fegeler  W., Proctor  R.A., Peters  G., von Eiff  C.

Objectives: 

Small colony variants (SCVs) are an emerging infectious disease problem, presenting as a naturally occurring, slow-growing subpopulation of Staphylococcus aureus that are characterized by tiny colonies on solid media. Studies on SCVs recovered from patients with persistent infections are hampered due to their frequent unstable phenotype. In particular, SCVs are not easily distinguishable from the normal phenotype in broth media and a reversion of SCVs into the normal phenotype is not traceable.

Methods: 

A set of isogenic S. aureus isolates comprising the (i) normal and the (ii) SCV phenotype (isogenic to the isolate with normal phenotype) recovered from clinical specimens, as well as (iii) corresponding mutants mimicking the SCV phenotype (knock-out of hemB), and (iv) their complemented mutants were used to investigate the feasibility of Fourier-transform infrared (FTIR) spectroscopy to trace the expressed phenotype in broth media. The respective isolates cultured on solid media served as controls. In addition, all isolates were genotyped by pulsed-field gel electrophoresis and spa typing.

Results: 

Using first-derivative infrared spectra to calculate spectral distances, hierarchical clustering based on spectral information in three different spectral ranges resulted in a dendrogram that showed a clear discrimination between both staphylococcal phenotypes. Distinct clusters comprising the clinical and mutant SCV phenotype on one hand and the normal phenotype (isolate with normal phenotype and complemented mutant) on the other hand were found. Thus, SCVs from different clonal lineages gave spectra that were more similar to one another than to their normal growth parent. FTIR was also shown to be able to trace the switch of the phenotypes in broth when the medium was supplemented.

Conclusion: 

FTIR spectroscopy allows a rapid, reproducible and clear discrimination of different phenotypes of S. aureus in fluid media for diagnostic and research purposes. In contrast to genotyping approaches, FTIR staphylococcal fingerprinting is only reliable for typing purposes if the isolates exhibit the same phenotype. In future studies, this technique may also provide an approach for tracing the SCV phenotype in infected tissues.