Multidrug resistant (MDR) pathogen infections are serious threats to hospitalized patients because of the limited therapeutic options. A novel group of antibiotic candidates, antimicrobial peptides (AMPs), have recently shown powerful activities against both Gram-negative and Gram-positive bacteria. Unfortunately, the viability of using these AMPs in clinical settings remains to be seen, since most still need to be evaluated prior to clinical trials and not all of AMPs are potent against MDR clinical isolates. To find a connection between the characteristics of several of these AMPs and their effects against MDR pathogens, we selected 14 AMPs of animal origin with typical structures and evaluated their in vitro activities against clinical strains of extensive drug-resistant Acinetobacter baumannii, methicillin-resistant Staphylococcus aureus, extended spectrum β-lactamase-producing Pseudomonas aeruginosa and extended spectrum β-lactamase-producing Escherichia coli. Our results showed that these peptides' hydrophilic/hydrophobic characteristics, rather than their secondary structures, may explain their antibacterial effects on these clinical isolates. Peptides that are amphipathic along the longitudinal direction seemed to be effective against Gram-negative pathogens, while peptides with hydrophilic terminals separated by a hydrophobic intermediate section appeared to be effective against both Gram-negative and Gram-positive pathogens. Among these, cathelicidin-BF was found to inhibit all of the Gram-negative pathogens tested at dosages of no more than 16 mg/L, killing a pandrug-resistant A. baumannii strain within 2 h at 4×MICs and 4 h at 2×MICs. Tachyplesin III was also found capable of inhibiting all Gram-negative and Gram-positive pathogens tested at no more than 16 mg/L, and similarly killed the same A. baumannii strain within 4 h at 4×MICs and 2×MICs. These results suggest that both cathelicidin-BF and tachyplesin III are likely viable targets for the development of AMPs for clinical uses.