Subinhibitory Concentrations of Trimethoprim and Sulfamethoxazole Prevent Biofilm Formation by Acinetobacter baumannii through Inhibition of Csu Pilus Expression

Abstract

Acinetobacter baumannii is emerging as a multidrug-resistant nosocomial pathogen of increasing threat to human health worldwide. Pili are important bacterial virulence factors, playing a role in attachment to host cells and biofilm formation. The Csu pilus, which is assembled via the chaperone-usher secretion system, has been studied in A. baumannii ATCC 19606. Here we show that, in opposition to previous reports, the common laboratory strain ATCC 17978 produces Csu pili. We found that, although ATCC 17978 was resistant to sulfamethoxazole (Smx) and trimethoprim (Tmp), subinhibitory concentrations of these antibiotics abolished the expression of Csu and consequently produced a dramatic reduction in biofilm formation by ATCC 17978. Smx and Tmp acted synergistically to inhibit the enzymatic systems involved in the bacterial synthesis of tetrahydrofolate (THF), which is required for the synthesis of nucleotides. The effects of these antibiotics were partially relieved by exogenous THF addition, indicating that Smx and Tmp turn off Csu assembly by inducing folate stress. We propose that, for Acinetobacter, nanomolar concentrations of Smx and Tmp represent a "danger signal." In response to this signal, Csu expression is repressed, allowing biofilm dispersal and escape from potentially inhibitory concentrations of antibiotics. The roles of antibiotics as signaling molecules are being increasingly acknowledged, with clear implications for both the treatment of bacterial diseases and the understanding of complex microbial interactions in the environment.KEYWORDS Acinetobacter, folate biosynthesis, folate stress, pilus assembly P ili formed by Gram-negative bacteria play an important role in the ability of these organisms to adhere to surfaces, to form biofilms, and to invade host cells and can affect motility (1). The chaperone-usher (CU) family of pili has been the focus of many studies, and the P-pili and type I pili of uropathogenic Escherichia coli play particularly important roles in establishing infection (2-4). A CU pathway for pilus biogenesis is present in most medically relevant Acinetobacter species (5). The genes for this CU pathway are encoded in a single six-gene operon, termed csu, and include csuA /B, csuA, csuB, csuC, csuD, and csuE (6). In the presence of the CsuC chaperone, CsuA/B polymerizes to form the major pilus subunit (7). Additional experimental and bioinformatic analyses have provided evidence that CsuD functions as the usher and CsuE as a tip adhesin, while CsuA and CsuB may play roles as minor pilin subunits (5,6,8). Despite its being conserved in many sequenced Acinetobacter baumannii strains, the role of Csu has been characterized only for A. baumannii ATCC 19606, an isolate used in many laboratories (5, 6). Genetic studies in this strain have shown that mutation of csuC or

The structure of PilA from Acinetobacter baumannii AB5075 suggests a mechanism for functional specialization in Acinetobacter type IV pili

“…Additionally, we found that biofilm formation was accompanied by the formation of crosslinking pilus-like fibers for all strains (Fig. S8), which we attribute to chaperone-usher (Csu) pili, consistent with their role in Acinetobacter biofilm formation (66,67).…”

The lytic transglycosylase MltB connects membrane homeostasis and in vivo fitness of Acinetobacter baumannii

“…To test this hypothesis, transmission electron microscopy was performed on the WT, ∆mltB mutant, and complemented strains. The WT and the complemented strains presented long appendages, consistent with pili (Alvarez-Fraga et al, 2016;Moon et al, 2017) at their cell surface. The numbers of pili-like structures were drastically reduced in the ∆mltB mutant ( Fig.…”

Broad-spectrum suppression of bacterial pneumonia by aminoglycoside-propagated Acinetobacter baumannii

“…pilus expression, biofilm formation, and capsule production in response to various antibiotics [20,21]. These findings demonstrate that antibiotics can have complex effects on host-pathogen interactions that extend beyond simply inhibiting bacterial growth.…”

“…concentrations of trimethoprim and sulfamethoxazole results in inhibition of Csu pilus expression and thereby reduces biofilm formation. Therefore, trimethoprim and sulfamethoxazole may serve as a signal for A. baumannii to disperse biofilms and thereby escape antibiotic pressure [20]. Many strains of A. baumannii constitutively express a type VI secretion system (T6SS), which is employed to kill competing bacteria within the environment [46].…”