β-lactams
Since the first β-lactam antibiotic was discovered (penicillin), they have become incorporated as a core part of clinical practice as treatments for various bacterial infections; β-lactam antibiotics are chosen as the antibacterial drug of choice (16). β-lactam antibiotics act on the peptidoglycan in the cell walls of fungi and bacteria, and they work by suppressing bacterial cell division or inducing bacterial rupture (24). However, bacteria can produce β-lactamase to enzymatically break down β-lactam antibiotics, which is the most prevalent mechanism of drug resistance. In the Ambler classification, β-lactamases can be grouped into one of four classes (A-D) according to the sequences of the amino acids that make up the enzyme (25).
Ambler class A enzymes. The serine β-lactamases of molecular class A are the most important enzymatic source of both natural and acquired resistance to β-lactams, particularly in Acinetobacter baumannii (26). TEM, SHV, CTX-M and KPC are the primary Ambler class A enzymes (27). TEM, CTX-M and KPC can hydrolyze penicillin, cephalosporin and carbapenem. Additionally, the use of antibiotics allows these enzymes to evolve and develop stronger drug resistance (28).
Ambler class B enzymes. Zinc-dependent metallo-β-lactamases (MBLs) are typically associated with gene cassettes of integrons and thus spread easily amongst bacteria (29). MBLs are classified into 3 subclasses. B1 and B3 are catalytically inactivated by two Zn2+ ions, and B2 is catalytically inactivated by one Zn2+ ion (14). NDM, VIM, SPM and IMP are the primary Ambler class B enzymes. The presence of the plasmid enables the rapid spread of the MBL gene, and the spread of NDM-1 is closely associated with drug resistance in Acinetobacter baumannii (30,31). Since the discovery of NDM-1 in India, over 24 NDM variants have been identified (32). NDM enzymes, composed of 270 amino acids, hydrolyze most β-lactams (including carbapenems) but not monobactams. However, NDM enzymes cannot be countered by clinically available β-lactamase inhibitors, including avibactam, clavulanate, sulbactam and tazobactam (33). Studies have shown that the percentage of NDM-1-positive isolates tends to be the highest, and Acinetobacter baumannii with the NDM gene show resistance to ampicillin (34). The acquisition of the NDM-1 gene is likely facilitated by the action of Tn125(35).
Ambler class C enzymes. Acinetobacter-derived cephalosporinases (ADCs) are responsible for resistance to cephalosporin antibiotics. ADC is the primary Ambler class C enzyme (27). ADC-mediated drug resistance is achieved through overexpression of ADC, and this overexpression itself is achieved through an ISAba1 insertion sequence, which is located in close proximity to the genes which confer resistance (36). The production of AmpC β-lactamases may be either chromosomally mediated or plasmid-mediated. AmpC β-lactamases are not inhibited by clavulanic acid, but are inhibited by cloxacillin or boronic acid (37). Acinetobacter baumannii can rapidly develop drug resistance due to the chemical similarity of the molecules between β-lactamase inhibitors and β-lactams, thus β-lactamase inhibitors, such as sulbactam and clavulanic acid, eventually become ineffective against Acinetobacter baumannii (38).
Ambler class D enzymes. Amongst the D-type β-lactamases, oxacillinase (OXA) is associated with resistance to carbapenems (39). The primary reason for carbapenem resistance is the presence of oxacillinase, which belongs to class D Ambler β-lactamases. To date, >400 OXA enzymes encoded by chromosomal or plasmid-localized genes have been characterized (40). The hydrolytic activity of OXA-type groups is more potent for oxacillin than benzylpenicillin; however, OXA-type enzymes are not considered extended-spectrum β-lactamases (ESBLs) as they do not hydrolyze broad-spectrum cephalosporins (7). The OXA-23 enzyme is encoded by a chromosomal gene or located on a plasmid, and it confers resistance to several antibiotics including ticarcillin, meropenem, amoxicillin and imipenem. The OXA-40 enzyme can hydrolyze penicillin; however, its ability to hydrolyze cephalosporins and carbapenems is weak, and it is resistant to inhibitors such as tazobactam, sulbactam, clavulanic acid and NaCl. The OXA-51 gene is generally non-transferable, encoded by chromosomal DNA. Clavulanic acid, tazobactam, or NaCl effectively blocks the activity of OXA-51. OXA-58 is found on a non-transferable 30k plasmid. When this plasmid is incorporated into the gene chain of Acinetobacter baumannii, carbapenem susceptibility is reduced (7). Because of certain insertion sequences, such as ISAbaI, ISAba125 and ISAba825, the overproduction ADC and OXA-51 confer high-level resistance to third- and fourth-generation cephalosporins (41). Carbapenem antibiotics, as the most commonly used antibiotics for nosocomial infections in the world, have successfully led to the enhancement of drug resistance in microorganisms such as Acinetobacter baumannii (16). The prevalence of carbapenem-resistant Acinetobacter Baumannii (CRAB) is increasing rapidly in many countries and regions, and this has complicated treatment choices (42). Carbapenem resistance is primarily mediated by B-type and D-type. The most common OXA-type carbapenemases include OXA-23, OXA-24, OXA-48, OXA-51 and OXA-58. Among them, OXA-23, OXA-24, OXA-48 and OXA-58 are acquired carbapenemases, whereas OXA-51 is intrinsic to Acinetobacter baumannii (43). The genes encoding these enzymes are regulated by upstream insertion sequences (IS), specifically ISAba1, ISAba2, ISAba3, ISAba9 and IS18. They lead to increased resistance to carbapenems through the expression of the blaOXA gene. In addition to OXA carbapenemases, the transferable MBL family, including VIM, IMP, GIM, SIM and NDM enzymes are also associated with the drug-resistant phenotype of Acinetobacter baumannii (44).
Other aspects aside from enzymes. Outer membrane proteins (OMPs) in general are β-barrel-shaped monomeric or trimeric porins that allow the diffusion of small molecules into and out of the periplasmic space of Gram-negative bacteria (45). The outer membrane of Acinetobacter baumannii contains several OMPs, including OmpA, CarO, OprD-like OMPs, Omp 33-36 kDa, AbuO, TolB, DcaP, Oma87/BamA, NmRmpM, CadF and OprF, amongst others. OMP has multiple functions, which confers bacterial resistance to threats such as harsh environments and antibiotics (46). OmpA is the most abundant outer membrane porin in Acinetobacter baumannii, and it functions by binding to efflux pumps and expelling antimicrobial compounds from the periplasm (47). OmpA increases the sensitivity of Acinetobacter baumannii to nalidixic acid, chloramphenicol, aztreonam, imipenem and meropenem, this feature is inseparable from its C-terminal region and Acinetobacter baumannii peptidoglycan (PG) coupling regulates outer membrane vesicle (OMV) stability (48). In addition, OmpA also actively siphons extracellular drugs to mediate antibiotic resistance and isogenic mutants, which in turn leads to a loss of cell wall integrity that sensitizes bacteria to colistin and also confers virulence (46). The outer membrane protein CarO is a carbapenem drug resistance-related OMP encoded by the CarO gene. CarO is divided into two subgroups, namely CarOa and CarOb. Facing different environments and hosts, the rapid adaptation of Acinetobacter baumannii results in alterations of the CarO gene (46). The resistance-nodulation-cell division (RND) efflux pump system is also associated with resistance in Acinetobacter baumannii. The efflux pump can extrude a variety of antibacterial agents, reducing the accumulation of antibiotics (49). The overexpression of adeABC plays an important role in acquired resistance to antibiotics. Cefepime, cefpirome and cefotaxime are the β-lactams most affected by the adeABC efflux system (50). Moreover, penicillin-binding protein 7/8 increases susceptibility to complement and contributes either directly or indirectly to serum resistance (51).
Novel options for resistance against β-lactam-based antibiotics. In recent years, researchers have found that the amino acid sequence of OmpA is highly conserved (>89%) in various clinical isolates, and OmpA mediates the adhesion and invasion of Acinetobacter baumannii to epithelial cells. OmpA can stimulate the innate immune response and induce biofilm formation, thus OmpA is a potential therapeutic target, although it has been shown that OmpA is not necessary for bacterial survival (22). It has been shown that increased iron content enhances OmpA protein expression in Acinetobacter baumannii, and strains with high OmpA protein expression are more aggressive, thus iron control strategies can be used in the management of Acinetobacter baumannii to reduce drug resistance (52). Cefiderocol, a member of the β-lactam antibiotics family, inhibits the synthesis of Gram-negative bacterial cell walls by binding to penicillin-binding proteins. However, due to its siderophore-like properties, it can enter the periplasmic space in bacteria and exhibits high stability to various β-lactamases such as AmpC and ESBLs (53,54). In an in vitro study, Cefiderocol was shown to be effective against OXA-23, OXA-40 and OXA-58. as well as NDM and IMP-producing Acinetobacter baumannii isolates (55). Efflux pumps are an important part of drug resistance. The efflux pump inhibitor carbonyl cyanide m-chlorophenylhydrazone can enhance the susceptibility of Acinetobacter baumannii to imipenem and cefepime (56). Equally effective efflux pump inhibitors include Quercetin, particularly when combined with imipenem, and it has a significant inhibitory effect on NDM and mexB/adeB (57). Acinetobacter baumannii vaccine studies has shown that the most effective vaccines tend to be multiplexed (consisting of outer membrane vesicles, bacterial ghosts, or multi-subunits) and are usually composed of antigens from OmpA, OmpW, OmpK and Omp22(58). For resistance to carbapenem antibiotics, the development of β-lactamase inhibitors has shown favorable results. β-lactamase inhibitor diazabicyclooctanes combined with sulbactam restored the sensitivity of sulbactam to carbapenem-resistant Acinetobacter (59). QPX7728 is a boric acid-lactamase inhibitor, which was shown to inhibit class A ESBLs, class B carbapenemases (NDM, VIM and IMP), class C and class D (OXA-23), and it enhanced its action against carbapenem-resistant Acinetobacter baumannii when combined with meropenem, ceflorazone, piperacillin and cefepime (60). Additionally, the combination of ampicillin and sulbactam (18 g per day) is an effective regimen for reducing the mortality of patients with CRAB (61). In addition, in terms of vaccine development, vaccines against BauA and OmpA that are vital virulence factors in pathogenicity of Acinetobacter baumannii play a certain role and combination of these antigens that can bind BauA and OmpA enhanced clearance of bacteria in liver and spleen (62). TCM ingredients can also be used to treat drug-resistant Acinetobacter baumannii. Possibly due to the synergistic action with antibiotics on efflux pump AdeB, Berberine hydrochloride combined with sulbactam can improve the antibacterial efficiency against Acinetobacter baumannii (63). Cilantro oil combined with piperacillin or cefoperazone can enhance the efficacy of the latter (64); however, the mechanism underlying the improved efficacy when combined needs to be determined. Other TCMs such as Piper betle combined with antibiotics are also worthy of research (65).
Polymyxins
The resistance of Acinetobacter baumannii to Polymyxins include: i) modification of the lipid A structure, ii) complete loss of Lipopolysaccharide (LPS) via mutations in the genes that synthesize lipid A, iii) reduction in the expression of cofactors involved in LPS synthesis, and iv) downregulation of proteins that participate in the export and/or stabilization of outer membrane precursors (66). LPS is part of the outer membrane of bacteria. Polymyxins inhibit bacterial membranes after binding to LPS, interact with lipid A of the bacterial outer membrane, and cause cell permeability and death by destroying membrane phospholipids. However, polymyxins antibiotics take a long time to work, and the use of colistin may increase the probability of nephrotoxic and neurotoxic complications (67). Colistin resistance in Acinetobacter baumannii is primarily caused by mutations in the PmrBTCS sensor kinase resulting in overexpression of PmrC. It has been shown that by knocking out the colistin PmrA mutant, its MICs is reduced by 64 to 1,024-fold, thereby restoring sensitivity to polymyxins (68). In Acinetobacter baumannii, various mutations and small fragments in the PmrB region are the primary cause of colistin resistance, and the most common PmrB mutation is A138T (69). The mutation of PmrA and PmrB of Acinetobacter baumannii can lead to resistance to polymyxins, and its virulence and fitness are also reduced. In addition, impaired virulence and fitness are also related to the lpx gene (70). Mutations in lpxA, lpxC and lpx affect lipid A synthesis. These spontaneous mutations include single-base changes, large deletions, and insertions of IS elements, all of which contribute to the high resistance exhibited by Acinetobacter baumannii (71). In addition, the induction of endogenous production of reactive oxygen species (ROS) by polymyxins, thus leading to oxidative killing of bacteria via hydroxyl radicals. Acinetobacter baumannii via inhibiting the formation of hydroxyl radicals attenuates polymyxin killing (72).
In the face of increasing drug resistance, it is a novel direction to identify new targets for use in combination with multiple drugs, such as the development of inhibitors against the targets of the modified bacterial outer membrane LPS two-component signal transduction system (73). Solving the problem of drug resistance should not only rely on antibiotics, instead, it may be favorable to combine current therapeutics with silver nanoparticles. Silver nanoparticles can penetrate microbial cell walls and alter cell membrane structure; this may reduce the MIC by 8-32X when used in combination with colistin (74). In addition, macolacin, a chemically synthesized substance that targets the plasmid-borne polymyxin resistance gene mcr-1, is also effective for Gram-negative pathogens expressing mcr-1 including Acinetobacter baumannii (75). The combination of polymyxin B with imipenem, meropenem, tigecycline and rifampicin in the treatment of Acinetobacter baumannii was superior to any of these alone, and the combination with rifampicin had the best effect (76). In terms of TCMs, the extract of Scutellaria barbata was shown to exhibit a good inhibitory effect on Acinetobacter baumannii, and the mechanism may be related to ROS (77), and the combination of Scutellaria barbata and polymyxin may have unexpected effects.