Actinobacillus pleuropneumoniaeIron Transport and Urease Activity: Effects on Bacterial Virulence and Host Immune Response

Abstract

Actinobacillus pleuropneumoniae, a porcine respiratory tract pathogen, has been shown to express transferrinbinding proteins and urease during infection. Both activities have been associated with virulence; however, their functional role for infection has not yet been elucidated. We used two isogenic A. pleuropneumoniae single mutants (⌬exbB and ⌬ureC) and a newly constructed A. pleuropneumoniae double (⌬ureC ⌬exbB) mutant in aerosol infection experiments. Neither the A. pleuropneumoniae ⌬exbB mutant nor the double ⌬ureC ⌬exbB mutant was able to colonize sufficiently long to initiate a detectable humoral immune response. These results imply that the ability to utilize transferrin-bound iron is required for multiplication and persistence of A. pleuropneumoniae in the porcine respiratory tract. The A. pleuropneumoniae ⌬ureC mutant and the parent strain both caused infections that were indistinguishable from one another in the acute phase of disease; however, 3 weeks postinfection the A. pleuropneumoniae ⌬ureC mutant, in contrast to the parent strain, could not be isolated from healthy lung tissue. In addition, the local immune response-as assessed by fluorescenceactivated cell sorter and enzyme-linked immunosorbent spot analyses-revealed a significantly higher number of A. pleuropneumoniae-specific B cells in the bronchoalveolar lavage fluid (BALF) of pigs infected with the A. pleuropneumoniae ⌬ureC mutant than in the BALF of those infected with the parent strain. These results imply that A. pleuropneumoniae urease activity may cause sufficient impairment of the local immune response to slightly improve the persistence of the urease-positive A. pleuropneumoniae parent strain.Actinobacillus pleuropneumoniae is the etiologic agent of porcine pleuropneumonia, a highly infectious disease of fattening pigs occurring worldwide (12). A number of putative virulence factors, such as Apx toxins, capsule, lipopolysaccharide (LPS), the ability to utilize transferrin-bound iron, and urease, have been described elsewhere (18). To date, conclusive evidence obtained by challenge experiments has been presented to confirm the role of Apx toxins and capsular material. A spontaneous Apx toxin-negative A. pleuropneumoniae strain was shown to be avirulent (14), and this result was supported later by using transposon mutagenesis (36) as well as by an isogenic A. pleuropneumoniae apxC insertion mutant (29). Also, capsule-deficient A. pleuropneumoniae strains obtained by chemical mutagenesis were shown to be attenuated (22), and this result was confirmed by reconstituting virulence properties and capsule formation upon transformation with a recombinant plasmid (39). Also, it was shown recently that the [Cu,Zn]-superoxide dismutase is not required for virulence (35). For other putative virulence factors, such as LPS (1, 3, 4), and the utilization of transferrin-bound iron (15,17,40), no conclusive challenge experiments have been performed to date. With respect to urease, data are inconclusive; ureasenegative A. pleuropneumoniae mutants hav...

Functional characterization ofActinobacillus pleuropneumoniaeurea transport protein, ApUT

“…We have therefore performed a detailed functional characterization of a bacterial urea transporter, ApUT, from the swine pathogen A. pleuropneumoniae, because it bears considerable sequence identity to mammalian UTs. Although this microbe is a strict respiratory tract pathogen, urease activity has been implicated in its virulence (1). Since the pathogenetic site for this organism, the respiratory tract, does not have an acidic pH, it is not clear whether urea uptake and metabolism Fig.…”

Nontypeable Haemophilus influenzae Gene Expression Induced In Vivo in a Chinchilla Model of Otitis Media

“…Urease has been shown to be involved in acid tolerance in Helicobacter (54) and A. pleuropneumoniae infection models (8,11). It is notable that, in one study, 87% of NTHI isolates from middle ear effusions were urease positive (15).…”

Glycoprotein analysis of porcine bronchoalveolar lavage fluid reveals potential biomarkers corresponding to resistance toActinobacillus pleuropneumoniaeinfection

“…Serum-Werk-Bernburg AG, Bernburg, Germany). BALF was obtained by lavaging the bronchus trachealis with 100 mL of 154 mM NaCl solution using a bronchoscope [4]. The yield of recovery normally ranged from 60 to 80 mL BALF.…”