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Pseudomonas aeruginosa possesses at least two multiple drug efflux systems which are defined by the outer membrane proteins OprM and OprJ. We have found that mutants overexpressing OprM were two- and eightfold more resistant than their wild-type parent to sulfamethoxazole (SMX) and trimethoprim (TMP), respectively. For OprJ-overproducing strains, MICs of TMP increased fourfold but those of SMX were unchanged. Strains overexpressing OprM, but not those overexpressing OprJ, became hypersusceptible to TMP and SMX when oprM was inactivated. The wild-type antibiotic profile could be restored in an oprM mutant by transcomplementation with the cloned oprM gene. These results demonstrate that the mexABoprM multidrug efflux system is mainly responsible for the intrinsic resistance of P. aeruginosa to TMP and SMX.

Knockout mutations were constructed in the arcA gene of a virulent type b strain of Haemophilus influenzae, and the behavior of the resulting mutants was investigated in a number of conditions that mimicked distinct steps in the natural infection pathway. In arcA mutants, synthesis of capsule and lipooligosaccharide (LOS) and growth in synthetic media were unaltered compared to synthesis of capsule and LOS and growth in synthetic media in the wild-type H. influenzae type b parent strain. However, the virulence of the arcA mutants for BALB/c mice was significantly reduced. Upon exposure to human blood or serum, the arcA mutants showed markedly reduced survival compared with the survival of its wild-type parent. Serum resistance could be fully restored by complementation in cis with the H. influenzae arcA gene but not by complementation in cis with the homologous gene from Escherichia coli. The proteomes of wild-type and mutant bacteria were markedly different, especially under anaerobic conditions, underscoring the global regulatory role of ArcAB in H. influenzae. Evaluation of antibody titers and classical complement activities in various serum samples pointed to complement-mediated bactericidal activity as the factor that distinguishes between the arcA mutant and wild-type phenotypes. Comparative analysis of the membrane fractions of the arcA mutants and the wild-type strain revealed several ArcA-regulated proteins, some of which may be implicated in the serum hypersensitivity phenotype.The only known natural host of Haemophilus influenzae is humans. Carriage of unencapsulated H. influenzae in the nasopharyngeal area is common, especially among children, and is considered a probable source of infection in otitis media, sinusitis, and pneumonia (51). Life-threatening H. influenzae meningitis is caused mainly by encapsulated type b strains; this is attributed to several factors, including the resistance of these strains to bactericidal activities of blood. Passage from the upper respiratory mucosa via the general circulation to the meninges requires successive adaptations of a bacterium's physiology in order to cope with the environmental changes that it encounters. Although the roles of the type b capsule (61) and lipooligosaccharide (LOS) (57) in invasive disease have been clearly demonstrated, we know little about the roles of other virulence factors in H. influenzae infections, not in the least because of the lack of reliable animal models.We hypothesized that the capacity of H. influenzae to swiftly adapt its physiology to match environmental conditions, such as changes in oxygen availability, is likely a virulence-associated trait. Two-component systems that are regulators of gene transcription in response to environmental signals have been implicated in virulence in a number of bacterial species, including Bordetella pertussis, Salmonella enterica serovar Typhimurium, and Shigella flexneri (5,18,53). No such role has yet been demonstrated for the ArcAB system involved in oxygendependent regulation of gene ...

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