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BackgroundActinomyces are a common part of the residential flora of the human intestinal tract, genitourinary system and skin. Isolation and identification of Actinomyces by conventional methods is often difficult and time consuming. In recent years, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) has become a rapid and simple method to identify bacteria.ObjectiveThe present study evaluated a new in-house algorithm using MALDI-TOF-MS for rapid identification of different species of oral Actinomyces cultivated from subgingival biofilm.DesignEleven reference strains and 674 clinical strains were used in this study. All the strains were preliminarily identified using biochemical methods and then subjected to MALDI-TOF-MS analysis using both similarity-based analysis and classification methods (support vector machine [SVM]). The genotype of the reference strains and of 232 clinical strains was identified by sequence analysis of the 16S ribosomal RNA (rRNA).ResultsThe sequence analysis of the 16S rRNA gene of all references strains confirmed their previous identification. The MALDI-TOF-MS spectra obtained from the reference strains and the other clinical strains undoubtedly identified as Actinomyces by 16S rRNA sequencing were used to create the mass spectra reference database. Already a visual inspection of the mass spectra of different species reveals both similarities and differences. However, the differences between them are not large enough to allow a reliable differentiation by similarity analysis. Therefore, classification methods were applied as an alternative approach for differentiation and identification of Actinomyces at the species level. A cross-validation of the reference database representing 14 Actinomyces species yielded correct results for all species which were represented by more than two strains in the database.ConclusionsOur results suggest that a combination of MALDI-TOF-MS with powerful classification algorithms, such as SVMs, provide a useful tool for the differentiation and identification of oral Actinomyces.

Actinomyces species play an important role in the pathogenesis of oral diseases and infections. Susceptibility testing is not always routinely performed, and one may oversee a shift in resistance patterns. The aim of the study was to analyze the antimicrobial susceptibility of 100 well-identified clinical oral isolates of Actinomyces spp. against eight selected antimicrobial agents using the agar dilution (AD) and E-Test (ET) methods. We observed no to low resistance against penicillin, ampicillin-sulbactam, meropenem, clindamycin, linezolid and tigecycline (0–2% ET, 0% AD) but high levels of resistance to moxifloxacin (93% ET, 87% AD) and daptomycin (83% ET, 95% AD). The essential agreement of the two methods was very good for benzylpenicillin (EA 95%) and meropenem (EA 92%). The ET method was reliable for correctly categorizing susceptibility, in comparison with the reference method agar dilution, except for daptomycin (categorical agreement 87%). Penicillin is still the first-choice antibiotic for therapy of diseases caused by Actinomyces spp.

Actinomyces naeslundii and A. oris are dental plaque formers involved in the pathogenesis of periodontitis. The aim of the study was to investigate the clonal relationship within two oral Actinomyces populations collected from plaque of patients with chronic periodontitis. The 223 clinical strains of A. naeslundii and A. oris were isolated from biofilm samples collected supra and subgingivally from teeth with shallow (probing depth (PD) = 3-4 mm), deep (PD = 5-6 mm) and very deep (PD ≥7 mm) pockets from 20 chronic periodontitis patients. All strains were submitted to repetitive sequence-based PCR typing using DiversiLab (BioMerieux,Marcy l´Étoile, France). Seven patients harboured only unrelated (<95% similarity) multiple isolates, while 13 harboured both similar (>95% similarity) and unrelated isolates at different sites. Identical (>98% similarity) strains were found to be present in the subgingival shallow depths more often than in the other subgingival depths. The number of clones in individual patients varied from 2 to 17 different rep-PCR genotypes. The clonal relationship within the oral populations of A. naeslundii and A. oris in an individual was unpredictable, ranging from the presence of multiple genotypes with no clonal similarity to only two different clones supra or subgingivally at different sites.