σ P -NagA-L1/L2 Regulatory Circuit Involved in ΔompA 299-356 -Mediated Increase in β-Lactam Susceptibility in Stenotrophomonas maltophilia.

OmpA, the most abundant porin in Stenotrophomonas maltophilia KJ, exists as a two-domain structure with an N-terminal domain of β-barrel structure embedded in the outer membrane and a C-terminal domain collocated in the periplasm. KJΔOmpA 299-356 , an ompA mutant of S. maltophilia KJ with a truncated OmpA devoid of 299 to 356 amino acids (aa), was able to stably embed in the outer membrane. KJΔOmpA 299-356 was more susceptible to β-lactams than wild-type KJ. We aimed to elucidate the mechanism underlying the Δ ompA 299-356 -mediated increase in β-lactam susceptibility (abbreviated as "ΔOmpA 299-356 phenotype"). KJΔOmpA 299-356 displayed a lower ceftazidime (CAZ)-induced β-lactamase activity than KJ. Furthermore, KJ2, a L1/L2 β-lactamases-null mutant, and KJ2ΔOmpA 299-356 , a KJ2 mutant with truncated OmpA devoid of299 to 356 aa, had comparable β-lactam susceptibility. Both lines of evidence indicate that decreased β-lactamase activity contributes to the ΔOmpA 299-356 phenotype. We analyzed the transcriptome results of KJ and KJΔOmpA 299-356 , focusing on PG homeostasis-associated genes. Among the 36 genes analyzed, the nagA gene was upregulated 4.65-fold in KJΔOmpA 299-356 . Deletion of the nagA gene from the chromosome of KJΔOmpA 299-356 restored β-lactam susceptibility and CAZ-induced β-lactamase activity to wild-type levels, verifying that nagA -upregulation in KJΔOmpA 299-356 contributes to the ΔOmpA 299-356 phenotype. Furthermore, transcriptome analysis revealed that rpoE (Smlt3555) and rpoP (Smlt3514) were significantly upregulated in KJΔOmpA 299-356 . The deletion mutant construction, β-lactam susceptibility, and β-lactamase activity analysis demonstrated that σ P , but not σ E , was involved in the ΔOmpA 299-356 phenotype. A real-time quantitative (qRT-PCR) assay confirmed that nagA is a member of the σ P regulon. The involvement of the σ P -NagA-L1/L2 regulatory circuit in the ΔOmpA 299-356 phenotype was manifested. IMPORTANCE Porins of Gram-negative bacteria generally act as channels that allow the entry or extrusion of molecules. Moreover, the structural role of porins in stabilizing the outer membrane by interacting with peptidoglycan (PG) and the outer membrane has been proposed. The linkage between porin deficiency and antibiotic resistance increase has been reported widely, with a rationale for blocking antibiotic influx. In this study, a link between porin defects and β-lactam susceptibility increase was demonstrated. The underlying mechanism revealed that a novel σ P -NagA-L1/L2 regulatory circuit is triggered due to the loss of the OmpA-PG interaction. This study extends the understanding on the porin defect and antibiotic susceptibility. Porin defects may cause opposite impacts on antibiotic susceptibility, which is dependent on the involvement of the defect. Blocking the porin channel role can increase antibiotic resistance; in contrast, the loss of porin structure role may increase antibiotic susceptibility.