The Battle against Antibiotic Resistance: Novel Therapeutic Options for Acinetobacter baumannii

Abstract

Undoubtedly, Acinetobacter baumannii stands out as one of the most effective bacteria responsible for nosocomial infections within the healthcare system. Due to its multidrug-resistant nature and the frequency of outbreaks that it causes the treatment of infections caused by this bacterium is challenging, antimicrobial combination therapy has been utilized to treat multidrug resistance Gram-negatives when monotherapy is ineffective. In contrast to antibiotics or short peptides, which possess only the capacity to bind and regulate a specific target, antibodies exhibit supplementary properties attributed to their Fc region, including opsonophagocytic activity, the agglutination process, and activation of the complement system. The criticality of antibodies is exemplified in triggering immunity against A. baumannii, stimulating protective mechanisms, preventing bacterial attachment to epithelial cells, opsonization, and complement-dependent bacterial destruction. Given antibodies’ significant role in humoral immunity, monoclonal antibodies (mAbs) may be generated to specifically bind to certain targets, thereby providing supplemental defense as a form of immunotherapy or passive immunization. Many encouraging tactics, ranging from phage therapy to immunotherapy, are being scrutinized for their efficacy in treating infectious diseases, thus shaping the future treatment landscape.

1. Introduction

Bacterial infections are the leading cause of death worldwide. Although the discovery of antibiotics successfully controlled bacterial infections, overuse and misuse of antimicrobial agents exacerbated the selection of multidrug-resistant (MDR) organisms. Acinetobacter baumannii is a bacteria that increases infection and mortality in vulnerable patients due to its ability to escape from antibiotic treatments effectively.

The increasing prevalence of nosocomial A. baumannii infections can be largely attributed to the remarkable ability of A. baumannii to colonize and form biofilms. Treatment options for these highly resistant pathogens are very limited. Because of this, clinicians are forced to resort to last-resort antibiotics, including colistin, which may induce nephrotoxicity, and select colistin-resistant A. baumannii.

To effectively treat and limit the spread of MDR A. baumannii (MDR-AB), a thorough understanding of the bacterial virulence factors and host-pathogen interactions is crucial. Therefore, before dealing with the new methods of treating A. baumannii, there is a need for a brief explanation to clarify the interactions between the host and the pathogen.

2. Virulence factors

A. baumannii, a highly antibiotic-resistant pathogen, possesses several virulence factors contributing to its pathogenesis and high mortality rates. Several recent studies have investigated virulence factors associated with the pathogenesis of A. baumannii and could thus serve as novel therapeutic targets.

These factors include the capsular polysaccharide (k-type), a major virulence factor [1]. The prevalent capsular types of A. baumannii include KL2, KL10, KL14, KL22, and KL52, with KL2 being associated with higher drug resistance and virulence [2].

Other virulence factors of A. baumannii include outer membrane proteins (Omps), lipopolysaccharide (LPS), capsular polysaccharide (CPS), phospholipase, nutrient-acquisition systems, efflux pumps, protein secretion systems, quorum sensing, and biofilm production (Table 1) [3]. Understanding these virulence factors is crucial for developing novel therapeutic targets and strategies to combat this multidrug-resistant pathogen [4, 5].

Virulence factor	Functions	Modulation
omps	Induce cell apoptosis, complement resistance, biofilm formation, cell invasion, and OMV biogenesis.	Unknown
CPS	Complement resistance and biofilm formation	Up-regulated upon antibiotic or ROS exposure
OMVs	Transferring OmpA and toxin delivery	Up-regulated upon antibiotic exposure
LPS	Membrane integrity, induce cell apoptosis, and antibiotic resistance	Loss during colistin resistance development
T6SS	Interspecies competition	Activate upon contact with competing bacteria
Micronutrient acquisition systems	Nutrient acquisition	Up-regulated under nutrient-deprived conditions
Type IV pili	Twitching motility	Up-regulated during growth in human serum
Bap	Biofilm formation	Up-regulated while growing under low iron conditions
Csu Pili	Biofilm formation	Antibiotic exposure

Table 1.

A. baumannii components, functions, and regulation conditions of them.

2.1 The mechanisms of A. Baumannii to promote self-survival

A. baumannii outer membrane proteins (Omps) play a versatile role in promoting bacterial survival. Omps in A. baumannii facilitate bacterial acclimatization to antibiotic- and host-induced stresses, aiding in immune evasion, stress tolerance, and resistance to antibiotics and antibacterial [6]. The ability of A. baumannii to adhere to abiotic surfaces and form biofilms, facilitated by Omps, and helps the bacteria survive in harsh environmental conditions such as desiccation, nutrient deficiency, and antibiotic treatment [7]. Additionally, A. baumannii outer membrane vesicles (OMVs), which contain Omps, contribute to the delivery of virulence factors to host cells, enhancing bacterial survival, nutrient acquisition, biofilm formation, and pathogenesis. A. baumannii strains that produce more abundant Omps, such as MDR-AB, exhibit more powerful cytotoxicity, stronger innate immune responses, and contain more virulence factors, potentially leading to worse outcomes [8].

LPS is the main component on the extracellular membrane of Gram-negative bacteria [9]. Mutations in the lipid A biosynthetic pathway can lead to changes in the structure of LPS in A. baumannii, resulting in colistin resistance. LPS-deficient A. baumannii strains show altered activation of the host innate immune inflammatory response, indicating the importance of LPS in interacting with host immune system. In addition, LPS-deficient A. baumannii can have alterations in their lipid A composition, such as the addition of phosphoethanolamine (pEtN) and galactosamine (GalN), which can affect the binding affinity of colistin. Loss of LPS in A. baumannii can lead to the upregulation of lipoproteins and the accumulation of the capsular polysaccharide poly-β-1,6-N-acetylglucosamine as compensatory mechanisms for membrane stabilization.

Capsular polysaccharides (CPS) in A. baumannii play a crucial role in bacterial virulence and survival [10]. The CPS structures in A. baumannii are diverse and can vary between strains [5]. These CPS structures often include rare sugars and branched oligosaccharide repeating units. The CPS biosynthesis gene encodes glycosyltransferases that are responsible for the synthesis of CPS structures. The presence of specific CPS structures, such as KL2, has been associated with antibiotic resistance and clinical outcomes in A. baumannii infections. Understanding the CPS structures and the genetics involved in their synthesis is important for developing targeted treatment strategies against A. baumannii infections [11].

Phospholipase functions of A. baumannii play a crucial role in promoting bacterial survival. Multiple studies have identified phospholipases as virulence factors that contribute to the pathogenicity of A. baumannii. The phospholipases are involved in the growth of phosphatidyl choline as a carbon source. These phospholipases involve various processes, such as hemolytic and cytolytic activities [12]. The phospholipases enable A. baumannii to adapt to different host niches and environments, enhance resistance to antimicrobial peptides, and facilitate the invasion of host cells [13].

Nutrient acquisition systems are often crucial for pathogen growth and survival during infection and represent attractive therapeutic targets. The pathogen utilizes various mechanisms to acquire essential nutrients from the host, such as heme and zinc.

A. baumannii has metal homeostatic systems that regulate the levels of essential nutrient metals in bacteria, particularly iron and zinc, that are important for colonizing different tissues and growth within vertebrates [14]. These systems, such as siderophores, heme uptake systems, and zinc uptake systems, enable the bacteria to overcome host-imposed zinc limitation by aiding in zinc uptake into the cells. The hemO locus, including the heme-degrading enzyme and scavenger, is required for high-affinity heme acquisition from host hemoglobin and serum albumin [15]. Additionally, A. baumannii possesses a Zn uptake (Znu) system consisting of an inner membrane ABC transporter and an outer membrane TonB-dependent receptor, which allows the pathogen to overcome host-imposed Zn limitation [16]. The TonB-dependent receptor HphR is an important component of the heme uptake system in A. baumannii and is involved in iron acquisition and cellular processes contributing to virulence [17].

Efflux pumps, such as the RND-type efflux pumps AdeABC and AdeIJK, contribute to resistance against antibiotics and biocides [18]. They are involved in extruding hazardous substances, including antibiotics, from within the bacterial cells [19]. The overexpression of these efflux pumps, particularly AdeABC, has been found to enhance the survival of A. baumannii when exposed to residual concentrations of biocides [20]. Additionally, efflux pump genes, such as adeABC, have been associated with tigecycline resistance in A. baumannii [21]. Efflux pumps have broad substrate specificity and are widely distributed among bacterial species, making them a major contributor to multidrug resistance in A. baumannii [22].

Secretion systems have recently been demonstrated to be involved in the pathogenic process, and five types of secretion systems out of the currently known six from Gram-negative bacteria have been found in A. baumannii. They can promote the bacteria’s fitness and pathogenesis by releasing various effectors. Additionally, antibiotic resistance is found to be related to some types of secretion systems [23]. The type VI secretion system (T6SS) is one such system found in A. baumannii, which is involved in bacterial competition and the delivery of toxic effector proteins [24, 25]. The T6SS in A. baumannii is highly diverse, with significant diversity in the range of encoded T6SS VgrG and effector proteins. There are multiple VgrG genes in A. baumannii strains, with most strains encoding between two and four different VgrG proteins. T6SS structural components of A. baumannii are distinctive from other Gram-negative pathogens, as evidenced by the presence of the Acinetobacter genus-specific protein AsaA. The T6SS in A. baumannii is involved in bacterial competition and secretion of T6SS effectors, such as Hcp is associated, which acts as a virulence factor, transporter of effectors, and chaperone [26]. The putative T6SS effectors in A. baumannii have diverse functions, including peptidoglycan hydrolases, lipases, nucleases, and nucleic acid deaminases [27].

Quorum sensing (QS) in A. baumannii plays a crucial role in bacterial survival and pathogenicity [28]. The QS system coordinates the behavior of individual bacteria in a population by mediating the synthesis, secretion, and binding of auto-inducer signals. The deletion of the auto-inducer synthase gene abaI in A. baumannii resulted in a decrease in biofilm formation and pathogenicity [28]. Additionally, the QS system regulates important virulence-related phenotypes, such as surface-associated motility and biofilm formation [29]. The antibacterial peptide octopromycin inhibited biofilm formation and surface movements in A. baumannii, demonstrating its anti-quorum sensing activity [30]. Furthermore, the abaI/abaR QS system was found to affect growth characteristics, morphology, biofilm formation, resistance, motility, and virulence in A. baumannii [31].

Targeting virulence factors can be an effective strategy for combating A. baumannii and other multidrug-resistant bacteria. Additionally, the development of innovative strategies, such as using bacteriophages and antibiotics in combination, has shown increased efficacy in eradicating biofilms formed by antibiotic-resistant A. baumannii strains. So, targeting virulence factors and biofilm formation can be an effective approach for designing drugs to combat multidrug-resistant bacteria.

3. Antimicrobial drug resistance and overcoming its problems

Antimicrobial drug resistance is one of the three major global threats to public health identified by the World Health Organization (WHO) in the twenty-first century [32]. A. baumannii is one of the main and most successful pathogens responsible for hospital-acquired infections in the modern healthcare system, associated with high mortality rates [33]. According to the reports of the WHO, about 80% of MDR or extensively drug-resistant (XDR) microbes have occurred due to the misuse and overuse of antibiotics, and these infections are associated with severe side effects [34]. Due to the prevalence of infections and outbreaks caused by A. baumannii drug resistance, few antibiotics are effective for treating infections caused by this pathogen [35]. Due to the spread of MDR bacteria and other resistant pathogens, there are limited treatment and prevention options, the failure of most antibiotics necessitates the search for better treatment options, and the need for alternative treatment options to treat these microbial pathogens (Figure 1).

Figure 1.

Newer approaches to tackle of MDR A. baumannii.

3.1 Combined treatment

Antibiotics, such as colistin, carbapenems, and tigecycline, have been widely used to treat A. baumannii [36]. However, the emergence of this bacterium’s multidrug-resistant strains has limited these drugs’ effectiveness. Therefore, new treatment options like combination therapy are an urgent need. The idea behind combination therapy is to use two or more antibiotics to kill the bacteria, which works in different ways [37]. In fact, in this method, two antibiotics with other mechanisms of action are used, such as a β-lactam antibiotic and an aminoglycoside, or two antibiotics that have the same means of action but work on different targets, such as two different carbapenems [38].

There are several notable advantages to using combination therapy to treat A. baumannii. One of these advantages is increasing the effectiveness of treatment [39]. Bacteria are attacked from different angles, making it more difficult for bacteria to develop resistance using multiple drugs. Another advantage is that combination therapy can reduce the risk of treatment failure [40]. Since A. baumannii is very resistant to antibiotics, using one antibiotic may not be effective in treating the infection, using multiple antibiotics reduces the chance of treatment failure, using lower doses of each antibiotic can reduce the risk of side effects in patients.

For example, pairing β-lactam antibiotics with β-lactamase inhibitors has proven effective in combating resistant strains. Additionally, combining antibiotics with different mechanisms of action can target multiple bacterial pathways, increasing treatment efficacy [41].

Combination therapy, including colistin/imipenem, colistin/meropenem, colistin/rifampicin, colistin/teicoplanin, colistin/sulbactam, colistin/tigecycline, and imipenem/sulbactam has been widely studied [42].

Combination therapy has been explored as a potential treatment option for A. baumannii infections. Various combinations have been studied, including polymyxins, rifampicin, fosfomycin, sulbactam, and avibactam. Polymyxin-based combinations, such as with cell-wall acting agents, rifamycins, and fosfomycin, have been extensively studied [43, 44]. Berberine hydrochloride (BBH) has shown synergistic effects with antibiotics against MDR A. baumannii (MDR-AB), including tigecycline, sulbactam, meropenem, and ciprofloxacin [45]. High-dose sulbactam, combined with additional antibacterial agents, including colistin, has shown promise in treating MDR-AB or XDR A. baumannii (XDR-AB) infections [46]. Fosfomycin has also been explored as a potential component of combination therapy against carbapenem-resistant A. baumannii (CR-AB) conditions [47, 48].

The combination of colistin and tigecycline is effective in the treatment of A. baumannii infection that causes pneumonia by ventilator, and the combined therapy of colistin, meropenem, and ampicillin-sulbactam in A. baumannii infection in patients. It was effective in treating blood malignancy [49, 50, 51]. The combined treatment of colistin/rifampicin and ampicillin/sulbactam/carbapenem combination therapy is effective for the treatment of A. baumannii MDR bacteria, causing carbapenem-resistant skin and soft tissue infections [52].

3.2 Repurposing

Repurposing existing drugs is also considered a strategy for treating MDR bacterial infections. Repurposing drugs, drug repositioning, or therapeutic switching is like giving a second life to medication previously used for different purposes [53].

Instead of starting from scratch, drug repurposing allows researchers to tap into a vast library of already approved drugs, saving time and resources in drug development [54]. Moreover, low risk of failure, shorter time frame cycles, high success rates, and less investment are the practicalities of drug repurposing. These drugs have undergone rigorous safety and efficacy testing, making them attractive candidates for new applications [55].

Drug repurposing has emerged as a promising approach to combating drug-resistant A. baumannii infections. Several FDA-approved drugs have shown potential for repurposing in treating A. baumannii. Etoposide and genistein inhibit the synthesis of polyphosphates, a virulence factor in A. baumannii (Table 2) [56, 57, 58]. 5-fluorouracil (5-FU), fluspirilene, and Bay 11–7082 were identified as drugs that resensitize A. baumannii to azithromycin and colistin in combination [59, 60]. Erythromycin, levamisole, chloroquine, and propranolol inhibit quorum sensing and virulence factors in A. baumannii [61, 62]. Tyrothricin, typically active against Gram-positive bacteria, exhibited antimicrobial activity against drug-resistant A. baumannii [63, 64].

	Compound	Activity-alone or in combination with	Approved use or known as
Central Nervous System	Citalopram	Polymyxin B	Antidepressant
	Fluspirilene	Colistin	Antipsychotic
Infectiology	Apramycin	Alone	Antibacterial
	Niclosamide	Colistin	Anti-helminthic
	Oxyclozanide	Alone	Anti-helminthic
	Rafoxanide	Alone	Anti-helminthic
	Ivermectin	Alone	Anti-parasitic
	Zidovudine	Alone	Antiretroviral
	Ciclopirox	Alone	Antifungal
Metabolism	Ebselen	Alone	Anti-inflammatory
	Bay 11–7082	Colistin	Anti-inflammatory
Natural Compound	Resveratrol	Colistin	Stilbene
Oncology	Mitomycin C	Alone	Anti-tumor
	Tamoxifen	Alone	Breast cancer
	5-Fluorouracil (5-FU)	Zidovudine	Antineoplastic (Colon Cancer)
	Mitotane	Polymyxin B	Antineoplastic
	Gallium	Alone	Antineoplastic
	Toremifene	Alone	Breast cancer

Table 2.

Relevant repurposing reports for MDR-AB.

Apramycin, Niclosamide, Oxyclozanide, Rafoxanide, and Ciclopirox are antibacterial, antifungal, and anthelmintic agents that have a therapeutic effect on MDR-AB.

Apramycin is an aminoglycoside approved for veterinary use. Apramycin can potentially be used against highly drug-resistant pathogens [65]. Niclosamide is an anthelmintic drug that has been commercially available in some countries since the 1960s. Niclosamide is usually administered orally and is well absorbed by the intestinal mucosa. High doses of this drug are associated with serious side effects. This drug has recently been suggested to treat other diseases, such as cancer [66].

Niclosamide alone has no antibacterial activity against A. baumannii, but a synergistic interaction between Niclosamide and colistin has been observed against CR-AB. This drug interacts with colistin-resistant strains negatively charged outer membrane, leading to a synergistic effect with colistin.

Oxyclozanide is used in veterinary medicine to treat fluke infections in ruminants. Oxyclozanide enhances the effect of colistin on colistin-sensitive and resistant isolates of A. baumannii, P. aeruginosa, and K. pneumoniae. This effect may be due to disrupting of the bacterial cell envelope [67].

Rafoxanide at a suitable dose had histidine kinase-antagonistic activities, which disrupted the abilities of MDR bacterial and fungal cells to adapt to stress conditions [68].

Ciclopirox, an antifungal drug, has bacteriostatic activity against E. coli, K. pneumonia, and A. baumannii of MDR strains [69]. The ciclopirox mechanism of action is the effect on the galactose and LPS salvage pathways [70].

Mitomycin C, tamoxifen, 5-FU, mitotane, and gallium include anti-tumor and antineoplastic drug agents that have a therapeutic effect on MDR-AB. The anticancer drug Mitomycin C can kill A. baumannii exponential-phase, stationary-phase, and biofilm cells [71].

The tamoxifen metabolites were active against MDR Gram-negative bacilli and might be potential antimicrobial agents to treat infections by these pathogens [72]. Colistin combination therapy with selective estrogen receptor modulators (SERM) as tamoxifen, raloxifene, and toremifen also exhibited good activity against polymyxin-resistant P. aeruginosa, K. pneumonia, and A. baumannii [73].

5-FU, another anticancer drug, despite the overall safety of 5-FU, is toxic in some cases, with toxicities including gastrointestinal (e.g., diarrhea, nausea, vomiting, mucositis/stomatitis, anorexia), hematological (e.g., neutropenia, thrombocytopenia, anemia), and dermal (e.g., hand-foot syndrome) symptoms [74]. The combination of 5-FU with azithromycin was effective against CR-AB; this combination, possibly reducing 5-FU toxicity, has also been found to inhibit the growth of bacterial pathogens and reduce the production of virulence factors [66].

Mitotane, an antineoplastic agent approved for cancer treatment, acts with polymyxin B on carbapenem- or polymyxin-resistant GNB in vitro. These efflux pump inhibitors alone did not affect the bacteria, but their activity was restored when combined with an antibiotic [70].

Gallium’s antibacterial activity dates back many years, but this drug was originally used as an anticancer agent. Due to its chemical similarity to iron, gallium inhibits the reactions or redox pathways of iron and the growth of bacteria [75]. Therefore, gallium compounds show broad-spectrum antibacterial activity and inhibit the growth of important bacterial pathogens such as A. baumannii, P. aeruginosa, S. aureus, K. pneumoniae, and E. coli [76].

3.3 New antibiotics

Acinetobacter is one of the ESKAPE pathogens known for their ability to escape commonly used antibacterial treatments. With the rise of antibiotic resistance and the limited efficacy of current therapeutic options, exploring novel antibiotics offers hope in combating A. baumannii infections. New antibiotics have been developed and tested for treating MDR bacterial strains. These include cephalosporins and carbapenems in combination with new β-lactamase inhibitors, tetracycline derivatives, fourth-generation fluoroquinolones, new combinations of β-lactam and β-lactamase inhibitors, siderophore cephalosporins, and new aminoglycosides that have been approved or are in clinical development [77, 78].

Novel siderophore cephalosporins antibiotics, such as cefiderocol (CFDC) [79] and GT-1 (LCB10–0200), have shown promise for the treatment of A. baumannii infections. CFDC demonstrates strong activity against MDR-AB isolates with lower minimum inhibitory concentration (MIC) values than other Gram-negative agents [80]. GT-1, combined with a β-lactamase inhibitor GT-055, has shown efficacy against many multidrug-resistant pathogens, including A. baumannii [81]. These novel siderophore cephalosporins utilize a “trojan-horse approach” to evade resistance mechanisms in Gram-negative bacteria [82]. However, available clinical data for cefiderocol are conflicting, leaving infectious disease specialists uncertain about its optimal use in clinical practice.

New tetracycline antibiotics, such as eravacycline and TP-6076, have shown promise for treating Acinetobacter infections. Eravacycline has demonstrated higher potency than tigecycline and has been effective against XDR-AB in vitro [83]. It has also been found to have a low propensity to induce Clostridioides difficile infection (CDI) [84]. TP-6076, a fully synthetic fluorocycline, has shown greater activity than other tetracycline-class antimicrobials against CR-AB isolates [85]. These novel tetracyclines can be valuable additions to the limited armamentarium of drugs targeting Acinetobacter [86].

Non-β-lactam β-lactamase inhibitors antibiotics have shown promise for the treatment of Acinetobacter infections. The etx2514/sulbactam combination has demonstrated efficacy against MDR-AB isolates, including those producing class D β-lactamases [87]. The zidebactam/cefepime combination has shown in vitro activity against CR-AB [88]. Wck 4234/meropenem combination has exhibited broad-spectrum activity against MDR Enterobacteriaceae, including NDM, KPC, OXA, CTX-M, SHV, and TEM enzyme-producing isolates [89]. Ln-1-255/meropenem-imipenem combination has demonstrated decreased resistance rates against CR-AB isolates [90]. However, non-β-lactam β-lactamase inhibitors for treating Acinetobacter infections are still ongoing research and development. Further, clinical data is needed to support the efficacy of these inhibitors, and gaps still exist in the treatment of infections caused by MDR Acinetobacter spp.

Novel β-lactam antibiotics, such as AIC-499 and FSI-1671, combined with sulbactam have shown promise for treating Acinetobacter infections. AIC-499 is a member of the diazabicyclooctane class of β-lactamase inhibitors with broad-spectrum activity against Ambler class A, C, and D serine β-lactamases [87]. Sulbactam, a first-generation β-lactamase inhibitor, has limited action against Acinetobacter spp. due to susceptibility to cleavage by β-lactamases [91]. However, when combined with durlobactam, the activity of sulbactam is effectively restored against MDR Acinetobacter strains [92]. FSI-1671, in combination with sulbactam, has also shown enhanced antimicrobial activity against A. baumannii clinical strains in China, with cefoperazone-sulbactam as the most potent compound.

Novel polymyxin B-derived molecules, such as SPR741 and FADDI-287, have shown potential for treating A. baumannii. SPR741 has been found to potentiate several large scaffold antibiotics in Gram-negative pathogens by interacting predominantly with the outer membrane (OM) [93]. FADDI-287, on the other hand, has been shown to induce significant perturbation in glycerophospholipid metabolism and histidine degradation pathways, leading to synergistic bacterial killing in both polymyxin-susceptible and resistant A. baumannii [94]. These findings suggest that these novel polymyxin B-derived molecules can overcome resistance mechanisms and enhance the efficacy of antibiotics against A. baumannii.

A new aminoglycoside called apramycin (EBL-1003) has promising potential for treating A. baumannii infections. It has proven to have wide antibacterial action against A. baumannii strains resistant to various drugs, including standard-of-care aminoglycosides [95, 96]. Because of its distinct chemical makeup, apramycin can circumvent resistance mechanisms frequently present in clinical isolates that produce carbapenemase [97]. Apramycin is quickly bactericidal against A. baumannii according to in vitro experiments [98]. In a mouse lung infection model, apramycin was also discovered to have a high likelihood of target attainment and robust in vivo activity [99]. According to these findings, apramycin may be a potent therapeutic alternative for treating CR-AB lung infections linked to high mortality rates and few therapeutic options.

3.4 Bacteriophage

A. baumannii infections may be treated using bacteriophage therapy, particularly when there is MDR. Bacteriophages are viruses that can target and eradicate particular types of bacteria. The effectiveness and safety of phage therapy against A. baumannii infections have been demonstrated in several studies [91, 100, 101]. Other Gram-negative and Gram-positive bacteria, such as S. aureus, P. aeruginosa, K. pneumoniae, Enterococcus, and Salmonella, have been demonstrated to be sensitive to phage therapy. The effectiveness of phages can be assessed by evaluating their host range, adsorption rate, and growth curve from various sources, such as sewage or wastewater [102]. Bacteriophages, particularly lytic ones, have shown promise as anti-A. baumannii therapeutics [103]. Both options are using phages in monophage therapy, phage cocktails, or in conjunction with antibiotics [104]. Bacteriophages can be administered parenterally, orally, topically, or by inhalation [105]. Endolysins and depolymerases, two phage-derived enzymes, have also been investigated for use against A. baumannii [106]. Antimicrobial therapy can use specific lytic bacteriophages or enzymes generated from phages to treat infections brought on by strains of A. baumannii that are incredibly drug-resistant.

Bacteriophages have multiple mechanisms of action against Acinetobacter. Some phages produce depolymerase, such as the tail spike proteins of phages Fri1, AS12, BS46, and AP22, which specifically recognize and digest the capsular polysaccharide (CPS) of A. baumannii. Other phages can cause mutations in genes that alter the architecture of the bacterial envelope, leading to phage resistance but also increased sensitivity to antibiotics, such as colistin [107]. Additionally, phages can degrade biofilms formed by A. baumannii, as demonstrated by the bacteriophage AB3 and its endolysin LysAB3 [106]. Further, phage therapy can target specific mechanisms of antimicrobial resistance, such as efflux pumps, by using efflux pump inhibitors or phage steering [108]. So, phage therapy, including phage cocktails and combination therapy with antibiotics, as well as phage-derived enzymes, such as endolysins and depolymerase, shows promise in combating MDR-AB [109].

But, bacteriophage therapy for Acinetobacter infections has limitations that must be addressed. Firstly, there needs to be more reliable safety and efficacy data for phage therapy due to the heterogeneity in previously published studies [110]. Secondly, the systemic effects of phage therapy need to be better understood [111]. Thirdly, the optimal application protocol for phage therapy, including the route of administration, frequency of administration, treatment duration, and phage titer, is still [112]. Additionally, the concurrent ecological and evolutionary interplay between phages and host bacteria requires further research to utilize the potential of bacteriophage therapy [103].

3.5 Antimicrobial peptides

Antimicrobial peptides (AMPs) have been demonstrated to prevent the MDR bacteria A. baumannii from growing. AMPs with a strong antibacterial action against A. baumannii strains include PapMA-3, MSI-78, h-Lf1–11, magainin-2, and LL-37. These peptides exhibit potential as antibacterial agents for treating bacteria that are resistant to antibiotics and have a broad spectrum of antimicrobial action [113].

Multiple mechanisms explain how AMPs work against Acinetobacter. The bacterial membrane may be damaged by AMPs, which will cause cell lysis [114, 115]. Additionally, they can infiltrate bacterial cells and engage with internal elements [116]. The mode of action of AMPs can be identified using experimental biophysical methods with model membranes and bacterial cells [117]. AMPs can cause the Acinetobacter membrane to become permeable [113]. The membrane’s interaction with AMPs and lipids may result in soft supramolecular configurations, which can thin and lyse the membrane [118].

A hybrid peptide called PapMA-3 showed low cytotoxicity and strong bacterial selectivity against carbapenem-resistant bacteria [119]. A. baumannii was also significantly resistant to the antibacterial effects of MSI-78, h-Lf1–11, magainin-2, and LL-37 [120, 121]. The anti-A. baumannii potency of Melittin, Histatin-8, Omega76, AM-CATH36, Hymenochirin, (this peptide showed moderate activity against Gram-negative bacteria), and Mastoparan was the highest [113]. In animal models of A. baumannii-induced pneumonia, the AMP derivatives dN4 and dC4 have shown therapeutic effectiveness [119, 120, 121, 122]. Furthermore, it has been demonstrated that larger doses of dN4 and dC4 can suppress and/or remove Acinetobacter biofilms [119].

A. baumannii, the cyclic peptide ZY4, exhibited little potential to induce resistance and outstanding efficacy against A. baumannii, including MDR strains [123]. According to these results, the antimicrobial peptides PapMA-3, MSI-78, h-Lf1–11, magainin-2, LL-37, Melittin, Histatin-8, Omega76, AM-CATH36, Hymenochirin, Mastoparan, dN4, and dC4 are efficient in treating Acinetobacter infections.

For the treatment of Acinetobacter infections, AMPs have limitations. Their toxicity and stability in vivo are two drawbacks that restrict their use [124]. Another drawback is their inherent limitations as peptides, such as stability, cytotoxicity, and bioavailability [125]. Natural AMPs are only useful for topical applications due to their pharmacological characteristics [126]. However, efforts are being made to get around these restrictions by creating novel AMPs and peptidomimetics by clever chemical changes [127]. Despite these drawbacks, AMPs hold potential as alternative therapies for focusing on bacterial infections, such as Acinetobacter, in both extracellular and intracellular contexts [113].

3.6 Monoclonal antibodies

Monoclonal antibodies (MAbs) are synthetic proteins, replicating the immune system’s defense against pathogens like bacteria and cancer cells. These antibodies target molecules on the pathogen’s surface known as antigens. The distinguishing quality of MAbs is their specificity, which enables them to recognize and bind to a certain target with extreme accuracy [128].

Heavy chains and light chains, two different protein chains, make up mAbs. These chains come together to form a Y-shaped structure. The antibody’s variable region, found at the end of each Y-shaped arm and binds to the particular antigen [129]. On the other hand, the antibody’s constant region controls its effector actions, such as triggering the immune system or obstructing the pathogen’s activity [130, 131].

MAbs can use different pathways to exert their therapeutic effects. The process of neutralization, in which antibodies bind to the pathogen and stop it from infecting host cells, is a typical one. In order to enlist immune cells in the fight against the disease, antibodies can potentially trigger antibody-dependent cellular cytotoxicity (ADCC). Furthermore, MAbs can influence the immune system’s response, enhancing the body’s ability to eliminate the infection.

MAbs have shown promise as novel therapeutics for Acinetobacter infections. These antibodies targeting outer membrane protein A (OmpA) of A. baumannii have improved opsonophagocytic killing of the bacteria [132]. Another MAb targeting the capsule of A. baumannii has been found to enhance macrophage opsonophagocytosis and reduce pro-inflammatory cytokines, leading to improved survival in mouse models [133]. A second MAb, developed through hybridoma technology, has also been shown to enhance macrophage opsonophagocytosis and improve survival in murine models of A. baumannii infection alone and combination with antibiotics [134]. Furthermore, MAbs have the advantage of being a narrow spectrum, targeting only the pathogenic species and potentially avoiding microbiome disruption. Additionally, a human MAb targeting a DNABII epitope has demonstrated efficacy in disrupting biofilms formed by Gram-positive and Gram-negative bacteria, including A. baumannii [135]. These findings suggest that MAbs have the potential to be effective therapeutic options for Acinetobacter infections, either alone or in combination with antibiotics.

Modulation of pro- and anti-inflammatory cytokines, such as IL-1, IL-6, TNF, and IL-10, was necessary for MAb treatment to be effective [136]. The FDA has approved three antibacterial MAb medicines, and numerous others are undergoing clinical studies [137].

Treating A. baumannii infections has led to the development of the MAbs C8 and 65. In deadly bacteremic sepsis and aspiration pneumonia models of XDR-AB infection, MAb C8 improves opsonophagocytosis by focusing on the capsular carbohydrate on the bacterial surface [134]. On the other hand, MAb 65 is extremely powerful and effective while expanding the coverage of immunotherapeutic strains. Combined with antibiotics, such as colistin, it improves macrophage opsonophagocytosis and results [135]. These MAbs have demonstrated the ability to decrease cytokine production, blood bacterial density, and sepsis biomarkers, showing their therapeutic potential [132, 133]. Antibacterial MAb treatment works by regulating pro- and anti-inflammatory cytokines and improving germ clearance by opsonophagocytosis.

AR401-mAb is a monoclonal antibody developed for the treatment of A. baumannii infections. It is highly effective against a broad range of clinical isolates and has been shown to improve outcomes when combined with antibiotics [134]. AR401- MAb is synergistic with colistin, a commonly used antibiotic, further enhancing its protective effects [138]. These findings suggest that using MAbs, such as AR-401, in treating A. baumannii infections may effectively improve outcomes and reduce bacterial burden.

Another study produced MAbs against the outer membrane protein A (OmpA) of A. baumannii, with one MAb, 1G1-E7, showing high reactivity and opsonophagocytic killing activity [132].

3.7 Nanoparticles

Nanoparticles are exceedingly small particles, usually measuring between 1 and 100 nanometers. Various substances, including metals, metal oxides, lipids, and polymers, can be used to create them. Nanoparticles differ from their bulk counterparts in multiple ways due to their small size and frequently show improved reactivity and physical features.

Due to their unique characteristics, nanoparticles hold considerable potential for infection control. The ability of nanoparticles to carry antimicrobial drugs or to naturally have antimicrobial features makes them useful in fighting drug-resistant bacteria. They also interact with bacterial cells well due to their large surface area-to-volume ratio, strengthening their antimicrobial actions.

The bacterial cell membrane can be damaged by nanoparticle interaction, which results in cell death. They can pierce bacterial membranes, resulting in structural damage and cellular component release. This condition impairs the bacteria’s capacity to continue performing essential tasks and ultimately results in their death. Acinetobacter is typically treated with nanoparticles by various mechanisms, including membrane damage, ROS production, efflux pump inhibition, and disruption of bacterial growth and biofilm formation. Copper sulfide nanoparticles, gallate-polyvinylpyrrolidone-capped hybrid silver nanoparticles, metal nanoparticles (such as silver, copper, gold, and aluminum), and zinc oxide nanoparticles are the specific types of nanoparticles utilized in treating Acinetobacter. These nanoparticles can potentially be potent therapeutic agents since they have demonstrated antibacterial activity against drug-resistant strains of Acinetobacter.

Nanoparticles, especially silver nanoparticles (AgNPs) and copper sulfide nanoparticles (cN16E-CuS), have shown promise in treating A. baumannii infections.

Silver nanoparticles (AgNPs) prevent the growth of drug-resistant strains by damaging bacterial membranes and producing reactive oxygen species (ROS). Biologically synthesized AgNPs also show efflux pump inhibitory activity, contributing to their antibacterial effect against MDR-AB. In addition, silver nanoparticles can induce apoptosis, inhibit the synthesis of new DNA in bacteria, and contribute to their antibacterial products. The antimicrobial activity of AgNPs is concentration-dependent and effective against extracellular and intracellular A. baumannii. Therefore, silver nanoparticles (AgNPs) have shown potential in treating A. baumannii infection. AgNPs showed good inhibitory activity against MDR-AB isolates, both alone and in combination with certain antibiotics. The combination of AgNPs with colistin, meropenem, or tigecycline significantly increased the sensitivity of MDR-AB to these antibiotics. In addition, silver nanoparticles inhibited the growth of A. baumannii and showed anti-biofilm activity, especially against weak biofilm producers. AgNPs have been found to interfere with the development of A. baumannii and disrupt biofilm formation, leading to a decrease in the expression of virulence and biofilm genes. Biogenic silver nanoparticles (Bio-AgNPs) synthesized by Fusarium oxysporum have also demonstrated antibacterial activity against CR-AB. Combined with polymyxin B, they showed synergistic effects, reducing the viable A. baumannii cells.

Another study reported using cationic antimicrobial lipid-stabilized copper sulfide nanoparticles (cN16E-CuS) for treating CR-AB. cN16E-CuS exhibited excellent antimicrobial activity against A. baumannii, producing excess reactive oxygen species and damaging bacterial membranes [139]. These findings suggest that nanoparticles, such as AgNPs and cN16E-CuS, can be used as alternative treatments for A. baumannii infections.

3.8 Gene editing

The DNA of creatures, including bacteria, can be changed using groundbreaking gene editing. It entails precise genetic manipulations such as adding, deleting, or changing particular genes. By concentrating on and interrupting the genes important for antibiotic resistance and other virulence factors, this method holds enormous potential for fighting bacterial infections.

Several gene editing tools have been developed to target bacterial pathogens, including A. baumannii. These tools include zinc finger nucleases (ZFNs) [140], transcription activator-like effector nucleases (TALENs) [141], and clustered regularly interspaced short palindromic repeats (CRISPR) [142] systems. Each tool offers unique advantages and can be tailored to target specific genes or regions within the bacterial genome.

CRISPR/Cas systems have shown potential as gene-editing tools for treating A. baumannii infections [143]. Genetic manipulation methods for studying A. baumannii pathogenesis and drug-resistance mechanisms are time-consuming and inefficient [144]. However, a detailed protocol for genetic manipulation in A. baumannii, including gene deletion, insertion, and point mutation, has been provided [145]. This protocol can aid in developing more innovative approaches to diagnosing and treating A. baumannii infections [146]. CRISPR/Cas systems can provide useful information about the functions of genes in A. baumannii and help identify potential targets for antimicrobials [140].

The Cas9 enzyme, which functions as molecular scissors, and a tiny RNA molecule known as a guide RNA, which points the Cas9 enzyme to the precise target spot in the bacterial genome, make up the CRISPR-Cas9 system.

Researchers have successfully applied the CRISPR-Cas9 system to target and edit the genes in A. baumannii. By designing appropriate guide RNA molecules, specific genes involved in antibiotic resistance or biofilm formation can be disrupted or modified, rendering the bacteria susceptible to existing antibiotics or impeding their ability to form biofilms.

CRISPR-Cas9 has been used for genetic manipulation in A. baumannii to study pathogenesis and drug-resistance mechanisms [145], which allowed for investigating drug-resistant mechanisms [147]. Additionally, a method for deleting drug-resistant genes in A. baumannii using CRISPR-Cas9 has been developed, providing a novel approach for preventing the spread of drug-resistant genes and treating drug-resistant bacteria [148].

3.9 Other

LpxC inhibitors have shown potential for the treatment of MDR-AB infections. Inhibiting LpxC, an enzyme involved in lipid biosynthesis, can reduce the toxicity of lipopolysaccharide (LPS) and enhance the efficacy of antibiotics [149, 150]. Compounds, such as LpxC-2 and LpxC-4, are synergistic with iron chelators (2,2′-bipyridyl and deferiprone) and gallium nitrate, significantly reducing bacterial counts.

The lipid A production is inhibited by LpxC inhibitors, such PF-5081090, which also increase cell permeability and improve resistance to a range of antibiotics such as rifampin, vancomycin, azithromycin, imipenem, and amikacin [72]. Additionally, LpxC inhibitors can prevent the activation of the Toll-like receptor 4 (TLR4) by A. baumannii LPS, which increases the opsonophagocytic death of the bacterium and decreases inflammation [150]. According to these results, LpxC inhibitors might be a different type of treatment for A. baumannii infections resistant to many drugs [151].

RX-P873, a novel antibiotic from the Pyrrolocytosine series, has shown high binding affinity for the bacterial ribosome and broad-spectrum antibiotic properties. It has demonstrated in vitro activity against MDR Gram-negative and Gram-positive strains of bacteria, including A. baumannii. In a study, RX-P873 was found to be highly active against A. baumannii isolates, with a MIC90 value of 1 μg/ml, which was two-fold more active than colistin and four-fold more active than tigecycline [152]. Additionally, a case report described the successful treatment of XDR-AB peritoneal dialysis-associated peritonitis with combination antibiotics, including intraperitoneal polymyxin B, without the need for catheter removal or switch to hemodialysis [153]. A study on RX-P873’s activity against extracellular and intracellular forms of infection by A. baumannii, and other bacteria found that RX-P873 may be a useful alternative for disorders involving intracellular bacteria, especially Gram-negative species [154]. Therefore, RX-P873 shows potential as a treatment for Acinetobacter infections.

4. Conclusion

Novel therapeutic strategies for antimicrobial therapy of Acinetobacter baumannii include combination therapy, drug repurposing, novel antibiotics, bacteriophage therapy, antimicrobial peptides (AMPs), human monoclonal antibodies (Hu-mAbs), nanoparticles, and gene editing. These strategies aim to overcome drug resistance and improve the efficacy of treatment against extensively drug-resistant Acinetobacter baumannii.

Acknowledgments

We are grateful to the financial support by Burn Research Center, Shiraz University of Medical Sciences, Shiraz, Iran.

Conflict of interest

The authors declare that they have no conflict of interest.