Abstract
Cathelicidin-BF, derived from the banded krait (Bungarus fasciatus), is a typically cationic, amphiphilic and α-helical antimicrobial peptide (AMP) with 30 amino acids that exerts powerful effects on multidrug-resistant (MDR) clinical isolates, including Pseudomonas aeruginosa, Acinetobacter baumannii, and Klebsiella pneumoniae, but whether it targets plasma membranes or intracellular targets to kill bacteria is still controversial. In the present study, we demonstrated that the disruption of bacterial membranes with high concentrations of cathelicidin-BF was the cause of bacterial death, as with conventional antibiotics at high concentrations. At lower concentrations, cathelicidin-BF did not cause bacterial plasma membrane disruption, but it was able to cross the membrane and aggregate at the nucleoid regions. Functional proteins of the transcription processes of P. aeruginosa and A. baumannii were affected by sublethal doses of cathelicidin-BF, as demonstrated by comparative proteomics using isobaric tags for relative and absolute quantification and subsequent gene ontology (GO) analysis. Analysis using the Kyoto Encyclopedia of Genes and Genomes showed that cathelicidin-BF mainly interferes with metabolic pathways related to amino acid synthesis, metabolism of cofactors and vitamins, metabolism of purine and energy supply, and other processes. Although specific targets of cathelicidin-BF must still be validated, our study offers strong evidence that cathelicidin-BF may act upon intracellular targets to kill superbugs, which may be helpful for further efforts to discover novel antibiotics to fight against them.
Highlights
The bacterial pathogens Pseudomonas aeruginosa and Acinetobacter baumannii are the top two causes of pneumonia acquired in intensive care units (ICUs) and ventilator-associated pneumonia (VAP), with mortality rates of 37.4 and 34.5%, respectively (Zhang et al, 2014)
Cathelicidin-BF was stable after incubation in sterile deionized water at 37◦C for nearly 24 h, reflected by the fact that the minimal inhibitory concentration (MIC) against DH5α increased slightly from 8 mg/L to 16 mg/L, it almost completely lost its antibacterial activity after 1 h incubation in mouse serum, reflected by the fact that the MIC against DH5α increased to more than 128 mg/L (Figure 1)
We demonstrated that the disruption of the bacterial membrane at high concentrations of cathelicidin-BF was the result of bacteria death, as is the case for conventional antibiotics at high concentrations (Figure 3)
Summary
The bacterial pathogens Pseudomonas aeruginosa and Acinetobacter baumannii are the top two causes of pneumonia acquired in intensive care units (ICUs) and ventilator-associated pneumonia (VAP), with mortality rates of 37.4 and 34.5%, respectively (Zhang et al, 2014). The well-known targets of AMPs are negatively charged prokaryotic cell membranes Their “selective toxicity” induces transmembrane pores that cause the leakage of intracellular components and bacterial death while leaving the electrically neutral membranes of eukaryotic cells untouched (Matsuzaki, 1999). This hypothesis is challenged by the fact that some AMPs kill bacteria and viruses, fungi, protozoa, parasites, and cancer cells, and some AMPs have hemolytic activities (Wang et al, 2016). Because some clinical isolates have gained resistance to most of the antibiotics available yet some AMPs still work, in particular through non-membrane targets, elucidation of their unique modes of action is highly anticipated
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