The incidence of multi-drug-resistant (MDR) bacteria strains increases yearly; thus, alternative therapies are required to compensate for increasingly ineffective antimicrobials that are currently available. An increasingly clinically relevant MDR pathogen is Acinetobacter baumannii, an opportunistic nosocomial Gram-negative bacterium, and member of the ESKAPE group of pathogens which include Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Enterobacter species (1). The ESKAPE designation refers to an extensive drug-resistant phenotype, and ability to overcome most current antimicrobial therapies. The prevalence of cases of A. baumannii infection is increasing worldwide and is a leading cause of morbidity and mortality (2, 3). This has prompted the World Health Organization (WHO) to assign carbapenem-resistant A. baumannii the highest critical priority ranking emphasizing the urgent need to identify new and effective drug therapies to combat this pathogen (4).

The opportunistic nature of A. baumannii primarily affects at-risk hospitalized patients with stays of longer-duration, especially immunocompromised individuals (5). Additionally, it is increasingly the source of infection in combat-wounded military personnel (6, 7). A. baumannii infection commonly presents as ventilator-associated pneumonia and catheter-related bloodstream/urinary infection, in addition to that of wounds (8–10). Due to their MDR nature, empiric treatment is generally ineffective and results in poor clinical outcomes (11). If untreated, rampant bacterial growth can ensue and give rise to TLR4-mediated septic shock and death (12, 13).

The incidence of MDR A. baumannii co-infection has increased in intensive care units (ICU) due to prolonged hospitalization of COVID-19 patients (14). One retrospective study of A. baumannii co-infection in COVID-19 hospitalized patients in Wuhan, China found that nearly half of the individuals who developed secondary bacterial infection died, with A. baumannii accounting for 36% of these infections (15). In another study, nearly 12% of ICU patients admitted with severe COVID-19 pneumonia developed secondary bacterial infections with K. pneumoniae or A. baumannii. Both pathogens exhibited extensive MDR resulting in more than double the mortality rate (from 38 to 83%) typically observed in COVID-19 patients (16). The incidence of A. baumannii infection in the past decade is associated with increased ICU admission, and longer hospital stays (17, 18). The nosocomic nature of this pathogen arises from its resistance to desiccation, and its ability to thrive in environments under selective pressures (19).

Investigation of A. baumannii pathogenic mechanisms and development of effective therapeutics against this pathogen has been a long-standing clinical and laboratory challenge. Initially, clinical isolates exhibited a hypovirulent phenotype necessitating the use of immunosuppressed mice as hosts or mixing of bacteria with porcine mucin to inhibit initial host clearance to study its pathophysiological mechanisms (20). However, recent MDR clinical isolates exhibit a hypervirulent phenotype, which rendered immunocompetent wild-type mice more readily susceptible to bacterial challenge (21–23). Thus, immunocompetent hosts now afford useful models to assess the efficacy of novel therapeutics with these more virulent A. baumannii isolates.

In order to develop novel and effective therapies to combat this pathogen it is first necessary to elucidate the underlying immune mechanisms by which A. baumannii evades the host’s adaptive and innate immune system. In this review we examine the current understanding of antibody-mediated and cell-mediated immune responses and key elements of protective immunity derived from recent therapeutic studies.

The success of passive immunization and use of monoclonal antibodies underscores the essential role of antibody-mediated immunity in protection against A. baumannii (24–33). Consistent with this notion, A. baumannii vaccinated B cell knockout mice were not protected in a murine-pneumonia model (34). Compared to vaccinated wild-type animals, these mice developed higher bacterial loads and subsequent extrapulmonary bacterial dissemination and bacteremia.

Moreover, some studies show correlation of survival with specific antibody titers (26, 35). Dose variation in subunit vaccines has shown larger doses of recombinant A. baumannii protein elicit higher antibody titers and higher protective efficacy (30). Interestingly, some studies observed higher protection in passive than in active immunization models (28, 36, 37). Across most studies, passive immunization has been shown to be an effective therapeutic route with only one study using an intranasal challenge model showing no improvement in survival albeit a delayed time to death (34). Thus, a focus of this review is that of the protective role of antibody-mediated immunity to A. baumannii.

Much of the vaccine development efforts for A. baumannii have focused on antibody-mediated immunity, and few studies have investigated the cell-mediated aspects of protection. As a result, most studies have utilized the IgG isotype ratio to determine whether vaccine-induced protection was Th1 or Th2 driven, and few studies have investigated direct T cell responses to A. baumannii, e.g., in T cell recall and adoptive transfer.

With the increasing drug-resistance of A. baumannii and few new antibiotic prospects on the horizon, vaccines have been looked to as a potential preventative measure for the most ‘at-risk’ patients, i.e., those developing infection due to prolonged hospitalization. Many vaccine studies utilized live-attenuated A. baumannii strains, inactivated whole-cell vaccines, or OMV preparations due to their robust immunogenicity and ability to induce protection (24, 28, 32, 34, 52). With the emergence of transcriptomics and identification of essential A. baumannii virulence factors, more emphasis has been placed on the development of recombinant protein subunit vaccines with improved safety profiles (26, 29, 30, 37). Along these lines, next-generation vaccine technology has been applied to engineering DNA vaccines against A. baumannii virulence factors that have proven difficult to produce for subunit vaccines (57, 58).

Vaccine-induced protective immunity tends to correlate with lower bacterial burdens 12 to 24 hours post-challenge in models using hypervirulent strains of A. baumannii that typically lead to acute death within 24 to 48 hours (12, 29, 31, 32, 52, 53). These studies utilize intravenous and intraperitoneal injections of bacterial inoculum to simulate bacteremia and septic shock and have demonstrated vaccine-induced protection through rapid clearance of bacteria ( Figure 3 ). Additionally, an A. baumannii-associated pneumonia model showed reduced bacterial burden in the lungs as well as limiting bacterial dissemination to extrapulmonary areas in vaccinated mice 24 hours after intranasal challenge (34). The acute nature of these models underscores the central role that preexisting antibody-mediated immune responses play in clearance of A. baumannii prior to bacterial dissemination. The abrogation of cytokine storm as a result of rapid bacterial clearance resulted in the drastic reduction in proinflammatory cytokine levels in protected mice, and a significant decrease was noted in IL-1β, TNF-α, and IL-6 in serum 12 and 24 hours post-challenge in protected mice (12, 24, 29, 34, 36, 52, 53). This was observed with both active and passive immunization.

The effect of vaccine-induced protective immunity on reducing A. baumannii pathology underscores the importance of early control and rapid clearance of the infection. Specifically, early time points post-challenge displayed pathology similar to mock-vaccinated animals followed by drastic reduction in inflammation 24 hours post-challenge (28, 31). Moreover, vaccinated mice exhibited limited infiltration of neutrophils and macrophages in lung infection models (21, 59), and active and passive immunization studies documented the importance of controlling bacterial burden within 24 hours of challenge.

Interestingly, a recent study utilizing a protective mAb in combination with complement, macrophage, and neutrophil depletion to evaluate the protective mechanisms showed that correlation between survival and bacterial burden does not always apply (46). In this study, double and even triple depleted mice did not survive subsequent challenge, but still exhibited strikingly reduced bacterial burdens compared to placebo mice. Use of TNF-α knockout mice indicated that the cytokine was not required for mAb efficacy. Moreover, IL-10 knockout mice were not protected which is in accordance with previous studies that have demonstrated its importance in A. baumannii infection (60, 61). However, macrophage depletion or treatment with an anti-TNFα antibody restored the efficacy of the mAb in IL-10 knockout mice. Efficacy of the mAb appeared to be linked to induction of IL-10 from neutrophils which balanced the macrophage’s TNF-α rather than simply reduction in bacterial burden observed in other therapeutic studies. The authors concluded that the protection conferred by the mAb was a result of inflammatory cytokine modulation suggesting that the delicate equilibrium of the innate and adaptive immune response is required for A. baumannii clearance, and that imbalance elicits a cytokine storm detrimental to the host. This interconnection remains to be elucidated as alternative therapeutics for A. baumannii infection are developed.

Acinetobacter baumannii is an increasingly relevant opportunistic pathogen due to development of MDR. The World Health Organization has assigned carbapenem-resistant A. baumannii as the ‘rank one’ critical pathogen requiring new therapeutic identification and development. With few new antibiotics on the horizon, novel therapies like vaccines and monoclonal antibodies must be developed to deal with this urgent microbial threat. Immunotherapeutic development has shed light on the potential mechanisms of adaptive protection against A. baumannii. Current research has demonstrated the importance of antibody-mediated protection through passive transfer of serum antibodies, B cell knockouts, and generation of monoclonal antibodies. The efficacy of this protection appears to be primarily a result of antigen-specific coating of the pathogen leading to opsonophagocytosis by neutrophils and macrophages. The role of cell-mediated immunity in this protection remains incompletely understood and will require further study to determine the exact underlying mechanisms. Protective efficacy correlates with lower bacterial burdens in addition to reduced proinflammatory cytokine production. However, recent studies have investigated immunomodulation of these therapeutics in maintaining immune homeostasis and dampening of cytokine storm. A greater understanding of how the adaptive immune system counters A. baumannii virulence factors in secondary infection can give much needed insight into the most effective immune response required to clear this pathogen.

The advances and research outlined in this review are expected to lead to novel immunotherapeutics to combat A. baumannii infections. An effective therapy would impact real world patient mortality and hospital costs arising from nosocomial infection. With few novel antimicrobials in development, this microbe’s ability to become resistant to current therapies outpaces current antibiotic treatment options. The key to the future development and implementation of vaccines and mAbs to A. baumannii may be establishing of a conserved immune response across multiple strains. Current research into A. baumannii will benefit from using clinically relevant bacterial strains and experimental models. Despite antibody-mediated protection demonstrating success in acute challenge models, this may only incompletely address the importance of cell-mediated immunity to A. baumannii. Moreover, it remains to be seen if the protection displayed in animal challenge models is indicative of protection against natural infection in humans. Studies have demonstrated rapid antibody-mediated neutralization of large doses of bacteria, but achieving these antibody titers early during initial bacterial exposure may not be realistic in a clinical setting.

In the laboratory setting, challenges remain in elucidating the mechanisms underlying the adaptive immune responses to A. baumannii, for example due to the use of different bacterial strains between laboratories and variations in experimental models used to investigate therapeutic efficacy. A. baumannii strains used to induce protection in certain experimental animal models may not provide protection against infection with other strains, which underscores the importance of searching for universally protective strains or antigens/antigenic epitopes. Additionally, bacterial challenge routes vary across current studies with some evaluating protection in only a blood or lung infection model. We posit that it will be paramount to evaluate protection in both clinically relevant models.

Novel immunotherapeutics against A. baumannii will inform a deeper understanding of its virulence factors and pathogenesis, and elucidating how this microbe evades the host’s innate but not adaptive immune response will further advance therapeutic development. The primary protective correlate of these therapies in current acute infection animal models is antibody-based. Of critical importance, however, is the development of alternative A. baumannii challenge models. Current acute models using hypervirulent strains afford little time for the host to mount a robust cell-mediated immune response; models need to be developed to allow A. baumannii to successfully infect immunocompetent wild type animals at lower challenge doses without first manipulating the host’s innate immunity. This could come from developing new animal models, or via isolating and evaluating new clinical bacterial strains. Extending the time from infection to mortality will facilitate elucidating the cell-mediated aspects of the adaptive immune response and inform the optimal antigen/adjuvant for use in future vaccine formulations.

Recent advancements in vaccine design will provide a novel edge in immunotherapeutic development against this everchanging pathogen. Along these lines, utilization of next-generation genomics, transcriptomics, and proteomics tools will aid identification of conserved virulence factors that can be assessed for their immunoprotective potential. Moreover, in silico models will accelerate development of safer protein-based subunit vaccines. We posit that the future will bring safer protein, DNA, and RNA based vaccines constructed to target conserved virulence factors and be multivalent affording broader protection. With antibody-mediated protection seemingly the most effective approach, there will also be an emphasis on monoclonal antibody development for clinical use. The road of translational science from the bench to the clinic is long but is paramount to improve patient outcomes against this multidrug resistant pathogen.

SJ searched literature and wrote the manuscript. J-JY searched literature and edited the manuscript. JC, C-YH and TF edited the manuscript. BA conceived and edited the manuscript. All authors contributed to the article and approved the submitted version.

This study was partially funded by the U.S. National Institutes of Health grants NS117742 to TF and AI135005 to C-YH. This work was also supported by The Jane and Roland Blumberg Endowment fund to BA.

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The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

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