Thus, its distinct chemical features and alternative mode of action may contribute to the unique activity of indolicidin against N. brasiliensis. Conclusions Selected AMPs are capable to contribute
to the first line of defense against Nocardia, yet, susceptibility appears to vary across different Nocardia species. Interestingly, our finding of neutrophil-derived check details AMPs to possess a broad antinocardial spectrum is paralleled by the characteristic feature of a neutrophil-rich infiltrate in histopathological specimens of nocardiosis. Moreover, the observed resistance of N. brasiliensis is remarkable, since N. brasiliensis is frequently reported to cause buy Temozolomide cutaneous and lymphocutaneous disease in otherwise immunocompetent hosts. Further studies should address in more detail the differential activity of AMPs, its causes and pathophysiologic
significance. Methods Bacterial strains and culture conditions Four strains of the genus Nocardia were investigated: Nocardia farcinica (ATCC Vadimezan 3318), Nocardia nova (ATCC 33726), Nocardia asteroides (ATCC 19247) and Nocardia brasiliensis (ATCC 19296). Strains were grown on Columbia blood agar for at least 72 hours at 37°C. Then 30 ml of Mueller-Hinton-broth (MHB) supplemented with 1% Tween 80 (Serva, Heidelberg, Germany) was inoculated with one loop of bacteria scraped off the agar plates. MHB was incubated in a shake incubator (220 rpm at 37°C). 10 ml of the culture was transferred
to a 50 ml tube which contained 1 mm glass beads (BioSpec Products, Bartlesville, USA). After vortexing for 10-15 seconds a homogenous suspension could be gained. A few millilitres of the suspension were used to inoculate another 50 ml of MHB (also supplemented with 1% Tween 80). Cultures were incubated until mid-logarithmic PJ34 HCl phase was reached. Incubation times were different for each Nocardia species (N. farcinica 12 h, N. nova 24 h, N. asteroides 16 h, N. brasiliensis 72 h). Innate defense antimicrobial peptides The activities of major human and bovine AMPs belonging to different families of AMPs were tested (summarized in Table 2): human cathelicidin LL-37, human α-defensins human neutrophil peptides 1-3 (HNP 1-3) and human β-defensin-3 (hBD-3), bovine indolicidin and bovine β-defensins lingual antimicrobial peptide (LAP) and tracheal antimicrobial peptide (TAP). Human cathelicidin LL-37, bovine indolicidin, LAP and TAP were synthesized using standard Fmoc/tBu chemistry on a multiple peptide synthesizer Syro II (MultiSynTech, Witten, Germany). Oxidation of the reduced LAP and TAP was achieved by dissolving the prepurified peptide with 2 M acetic acid and dilution to a peptide concentration of 0.