AdeRS is the first characterized and also the most studied TCS in A. baumannii. It was first described in a clinical strain A. baumannii BM4454, when the inactivation of adeS resulted in an increased susceptibility to aminoglycosides due to the downregulation of the RND efflux pump AdeABC (Marchand et al., 2004) (Figure 3). Since it was first identified, a number of mutations in either adeR, adeS, or both have been shown to be directly responsible for the overexpression of the AdeABC pump (Ruzin et al., 2007; Yoon et al., 2013; Sun et al., 2016). Considering AdeRS system's role in the expression of AdeABC, it can be said that it plays a role in the susceptibility of A. baumannii to antibiotics that are substrates of the AdeABC pump. Further, the overexpression of AdeABC efflux pump has been associated with the decreased susceptibility to tigecycline observed in some clinical isolates of A. baumannii (Sun et al., 2014; Yuhan et al., 2016) thus implicating an indirect role of AdeRS in the susceptibility toward tigecycline. This is important since tigecycline is one of the last resort antibiotics for the treatment of multidrug resistant A. baumannii infections (Ni et al., 2016). However, there needs to be further investigations into this due to the possibility of involvement of other factors for the observed tigecycline susceptibility (Yoon et al., 2013).

Recent transcriptomics data suggest that the role of AdeRS extends well-beyond the expression of AdeABC efflux pump. A study in A. baumannii AYE showed that AdeRS controls the expression of almost 600 different genes (Richmond et al., 2016). Products of a number of these genes are believed to play a role in virulence, biofilm formation and multi drug efflux activity. However, deletion of adeB in the same strain resulted in similar phenotypes as deletion of adeRS. This suggests that at least some phenotypic changes observed upon the adeRS deletion may be a result of the decreased expression of the AdeABC efflux pump (Richmond et al., 2016).

The multifaceted regulon of AdeRS remains to be explored further, especially in clinically relevant phenotypes of A. baumannii. Further, environmental signals that activate the sensor kinase, AdeS, remain mostly unknown. However, we recently uncovered evidence that AdeRS system maybe responding to the NaCl concentrations in the growth medium (De Silva and Kumar, 2017). This work links adaption to environmental conditions, such as NaCl concentration to antibiotic susceptibility (as a result of expression of the AdeABC pump) as well as virulence factors, such as biofilm formation and surface-associated motility. It is therefore obvious that AdeRS plays a role in the antibiotic susceptibility of A. baumannii but also possibly in its virulence. However, it's role in antibiotic susceptibility and virulence is likely to be more strain-specific, as it is not uncommon to find disrupted copies of adeRS genes in clinical isolates of A. baumannii, such as LAC-4 and AB031 (Table 1).

BaeSR, named such because of its homology with an E. coli TCS (Leblanc et al., 2011), mediates a possible “cross–talk” with other TCSs. It has been shown to regulate overlapping regulons with other TCSs in A. baumannii. BaeSR was initially thought to be associated with the regulation of AdeABC RND efflux pump expression (Lin et al., 2014) (Figure 3). This is indicative of a possible cross–talk between BaeSR and AdeRS. Further investigations into the BaeSR revealed that it may also modulate the expression of AdeIJK and MacAB-TolC efflux pumps (Henry et al., 2012). However, efforts to determine the DNA binding sites in the promoters corresponding to the observed target genes remain unsuccessful, leaving room for further explorations (Lin et al., 2015). A phenotypic microarray screen revealed that the deletion of baeR resulted in reduced tolerance of A. baumannii to tannic acid (Lin et al., 2015), a diverse group of natural antibacterial compound (Henis et al., 1964). Tannic acids has been used as a topical agent in burn patients (Hupkens et al., 1995) as they display effective antibacterial activity against various bacteria, including E. coli, Staphylococcus aureus, Staphylococcus epidermidis, Salmonella spp. etc (Kim et al., 2016). Tannic acid has also been shown to inhibit biofilm formation in Staphylococcus aureus (Payne et al., 2013). In A. baumannii, Tannic acids are being explored as adjuvants for antimicrobial therapy. They were shown to synergize the activity of novobiocin, rifampicin, and fusidic acid against MDR A. baumannii (Chusri et al., 2009). However, the role of BaeSR TCS in tannic acid as well as the expression of efflux pumps controlled by BaeSR will have to be considered to in order to explore the clinical usage of tannic acid as an adjuvant therapy options against A. baumannii.

Studies on the environmental signals that BaeSR responds to remain limited. However, expression of baeR and baeS in A. baumannii is induced by sucrose (20% w/v) (Lin et al., 2014), suggesting that BaeSR may be involved in A. baumannii's response to osmotic stress.

A. baumannii's resistance to commonly used antibiotics has led to an increased use of “last resort” antibiotics, such as colistin (Karaiskos et al., 2017; Jiménez-Guerra et al., 2018). As a result, emergence of colistin resistance is becoming more common in A. baumannii (Cai et al., 2012; Lean et al., 2015). Investigations into the mechanisms of resistance to colistin in A. baumannii have revealed the involvement of PmrAB resistance (Park et al., 2011; Rolain et al., 2013), named so for its role in polymixin (Figure 3). PmrAB has been described in various Gram-negative pathogens including E. coli (Quesada et al., 2015), Salmonella enterica (Gunn, 2008), Klebsiella pneumoniae (Cheng et al., 2010), and Pseudomonas aeruginosa (Lee and Ko, 2014) and has been shown to have a similar function colistin resistance. Observations of mutations in both pmrA (RR) and pmrB (HK) leading to decreased susceptibility to colistin presented preliminary evidence of the connection between PmrAB and colistin susceptibility in A. baumannii (Adams et al., 2009). Further, both colistin-resistant clinical isolates as well as laboratory generated spontaneous mutants showed phosphoethanolamine modification of lipid A of lipopolysaccharide (LPS) within the outer membrane (Arroyo et al., 2011; Beceiro et al., 2011). The modification of lipid A is mediated by PmrC which is generally part of the same operon as pmrAB (Raetz et al., 2007). PmrC can add phosphoethanolamine to either 4′ or 1′ phosphate of lipid A (Da Silva and Domingues, 2017). This modification of LPS results in a positively charged phosphate groups and prevents the binding of the cationic colistin (Tamayo et al., 2005a,b; Arroyo et al., 2011). Mutations in both pmrA and pmrB cause the overexpression of the pmrCAB operon.

Observations that low pH or supplementation of Fe³⁺ in the growth medium (Adams et al., 2009) lead to colistin resistance may suggest that PmrB could be responding to those signals (Gunn, 2008). However, growth of A. baumannii under low pH or in iron supplemented growth media failed to alter the expression of pmrA (Adams et al., 2009). Therefore, the environmental signals to which PmrAB responds to in A. baumannii remain elusive.

GacSA is a TCS that is well-characterized in Pseudomonas sp. (Gooderham and Hancock, 2009). GacSA in A. baumannii ATCC19606 was identified when the transposon insertions in the gacS sensor kinase gene rendered the mutants incapable of utilizing citrate as the sole carbon source (Dorsey et al., 2002). This suggests that GacSA is involved in citrate metabolism. Since the initial characterization of GacSA in A. baumannii ATCC19606, a number of subsequent studies have carried out the functional characterization of GacSA TCS in A. baumannii ATCC17978. Interestingly, in A. baumannii ATCC17978, the gacS gene is not linked to the response regulator-encoding gene. Rather, it has both a HisKA domain and a REC domain suggesting that it could function as a hybrid sensor kinase. Although, there is also a possibility that in A. baumannii ATCC17978, the response regulator for GacS is encoded elsewhere in the genome. This indicates that the organization of the gacSA genes may vary from strain to strain in A. baumannii.

In addition to the initially observed metabolic role of gacS, a transposon mutant with a disrupted gacS gene displayed significantly reduced A. baumannii's ability to inhibit Candida albicans (Peleg et al., 2008b). gacS deletion mutant also displayed attenuated virulence in a mouse infection model (Cerqueira et al., 2014). Deletion of gacS also led to the revelation of its involvement in a number of other virulence related functions. These include control of pili synthesis, motility, and biofilm formation, resistance against human serum, and metabolism of aromatic compounds (Cerqueira et al., 2014) (Figure 3). GacSA is involved in the regulation of the aromatic compound catabolism through the paa operon that encodes the components of the phenylacetic acid catabolic pathway. The paa gene cluster is significantly downregulated in the gacSA deletion mutants which may explain their attenuated virulence in a mouse septicaemia model (Cerqueira et al., 2014). The attenuated virulence of gacSA deletion mutants was observed in a later study involving a zebra fish virulence model as well (Bhuiyan et al., 2016) adding to the repertoire of studies that suggest that GacSA may function as a global virulence regulator in A. baumannii.

Biofilm formation is an important virulence factor of pathogens, such as A. baumannii that helps them survive harsh conditions present in hospital environments. The ability of A. baumannii to form biofilms starts with its attachment to surfaces that is mediated by the expression of pili. The expression of pili is mediated by the csu operon in A. baumannii and is under the regulatory control of BfmRS (Tomaras et al., 2008). Deletion of the response regulator bfmR in A. baumannii ATCC19606 resulted in the complete abolishment of biofilm formation (Tomaras et al., 2008) (Figure 3). While of the role of csu operon in the attachment of A. baumannii on abiotic surfaces is well-established (Tomaras et al., 2003; Moon et al., 2017; Pakharukova et al., 2018), its role in the adherence of A. baumannii to human epithelial cells remains ambiguous. It was observed that A. baumannii ATCC19606 strain lacking csuE in fact adhered to bronchial epithelial cells better than the wild-type parent making the role pili in adherence to epithelial cells unclear (de Breij et al., 2009). It is possible that this was a strain specific outcome and further investigations are required to draw definitive conclusions.

In addition to regulating biofilm formation, BfmRS also plays a role in regulating the exopolysaccharide production (Geisinger et al., 2018). Exopolysaccharides play an important role in virulence of A. baumannii as they are a component of the capsule, which protects A. baumannii against serum killing and increasing the virulence in animal models. Further, antibiotic exposure leads to an increase in capsule production in A. baumannii mediated by increased expression of genes in K-locus, which in turn is regulated by the BfmRS system (Geisinger and Isberg, 2015).

Crystal structure of BfmR shows that it binds to its own promoter with higher affinity in an inactive (dephosphorylated) state compared to the active (phosphorylated) state (Draughn et al., 2018). This is unusual behavior highlights a unique self-regulation strategy of BfmRS system Therefore, BfmRS system is an excellent candidate to study not only the mechanisms that regulate virulence factors in A. baumannii but also the functioning of the TCSs systems in general.

A1S_2811 is a recently characterized hybrid sensor kinase possessing four histidine–containing phosphotransfer domains as well as a regulatory CheA-like domain and a CheY-like receiver domain (Chen et al., 2017). CheA and CheY homologs in E. coli and P. aeruginosa are associated with regulatory roles in controlling motility via regulating either pili or flagella (Li et al., 1995; Alon et al., 1998; Bertrand et al., 2010). Interestingly, in A. baumannii, this hybrid sensor kinase is expressed in an operon composed of five genes where the four other genes upstream are pilJ, pilI, pilH, and pilG. Phenotypic analysis of the deletion mutant of A1S_2811 revealed a significant reduction in surface motility and biofilm formation at the gas-liquid interface. More intriguingly, abaI, which encodes a N-acylhomoserine lactone involved in quorum sensing, was also significantly downregulated. Supplementation with synthetic homoserine lactone complemented the biofilm and motility phenotypes (Figure 3). This suggests that A1S_2811 regulates biofilm formation and surface motility through an AbaI-associated quorum sensing pathway rather than the conventional pili associated pathway (Chen et al., 2017). This is in contrast to the BfmRS mediated regulon controlling biofilm formation in A. baumannii. Association of both BfmRS and A1S_2811 with biofilm formation is also an example of one phenotype being under the control of multiple regulatory networks formed by different TCSs.

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.