We extracted a granule-rich sediment from normal human neutrophils and subjected it to chromatographic, electrophoretic, and functional analysis. The extract contained three small (molecular weight < 3,500) antibiotic peptides that were named human neutrophil peptide (HNP)-1, and

We tested the in vitro susceptibility of Candida albicans to three defensins from human neutrophilic granulocytes (HNP-1, 2, and 3), a homologous defensin from rabbit leukocytes (NP-1), and four unrelated cationic peptides. Although the primary amino acid sequences of HNP-1, 2, and 3 are identical except for a single amino-terminal amino acid alteration, HNP-1 and HNP-2 killed C. albicans but HNP-3 did not. C. albicans blastoconidia were protected from HNP-I when incubations were performed in the absence of oxygen or in the presence of inhibitors that blocked both of its mitochondrial respiratory pathways. Neither anaerobiosis nor mitochondrial inhibitors substantially protected C. albicans exposed to NP-1, poly-L-arginine, poly-L-lysine, or mellitin. Human neutrophilic granulocyte defensin-mediated candidacidal activity was inhibited by both Mg2+ and Can, and was unaffected by Fe2". In contrast, Fe2" inhibited the candidacidal activity of NP-i and all of the model cationic peptides, whereas Mg2+ inhibited none of them. These data demonstrate that susceptibility of C. albicans to human defensins depends both on the ionic environment and on the metabolic state of the target cell. The latter finding suggests that leukocyte-mediated microbicidal mechanisms may manifest oxygen dependence for reasons unrelated to the production of reactive oxygen intermediates by the leukocyte.

Six antimicrobial peptides, corresponding to the family of "Ilysosomal cationic proteins" described previously by Zeya and Spitznagel (H. I.

Six related cysteine-rich, low-molecular-weight peptides were purified from rabbit peritoneal granulocytes and tested in vitro for fungicidal activity against Candida albicans. Two peptides (NP-1 and NP-2) were highly effective, one (NP-3a) was moderately active, and three (NP-3b > NP-4 >> NP-5) had substantially less potency. There was a general, but imperfect, correlation between the candidacidal potency of each peptide and its net cationic charge. Candidacidal activity by NP-1 was concentration and time dependent and occurred rapidly under optimal low-ionic-strength conditions. It was inhibited by increasing either the ionic strength or Ca2+ concentration of the incubation mixtures, but was relatively unaffected by Mg2+. Candidacidal activity was independent of H+ concentrations between pH 5 and 8, but decreased below pH 5. Candidacidal activity was temperature sensitive and was virtually abolished when NP-1 was incubated with C. albicans at 0°C. Cysteine-rich antimicrobial peptides such as NP-1 and NP-2 may equip leukocytes to deal with infections caused by C. albicans and other fungi that are susceptible to their microbicidal effects.

Microbicidal cationic proteins 1 and 2, peptides derived from rabbit lung macrophages, were tested for bactericidal activity against various bacterial species. Both were highly active against diverse gram-positive and gram-negative organisms under conditions of near-neutral pH (between 7 and 8) and relatively low ionic strength. Susceptible species included Staphylococcus aureus, Staphylococcus epidermidis, Streptococcus pneumoniae, Listeria monocytogenes, Pseudomonas aeruginosa, Klebsiella pneumoniae, Haemophilus influenzae, Escherichia coli, and Serratia marcescens. Streptococcus agalactiae, type 1A, was less susceptible than the aforementioned organisms or S. agalactiae, type 3. Bordetella bronchiseptica, a common commensal and pathogen of the rabbit respiratory tract, was completely resistant to both peptides.