Adherence of Actinobacillus pleuropneumoniae to swine-lung collagen.

Actinobacillus pleuropneumoniae serotype 1 adhered to immobilized swine-lung collagen. Bacteria bound to collagen type I, III, IV and V. At 5 min incubation, 30 % of bacteria adhered to collagen, reaching saturation in around 90 min. Treatment of bacteria with divalent-metal chelators diminished their attachment to collagen, and Ca(2+) but not Mg(2+) increased it, suggesting Ca(2+) dependence for adherence. Proteolytic enzymes drastically reduced bacterial adherence to collagen, showing that binding involved bacterial surface proteins. Porcine fibrinogen, haemoglobin and gelatin partially reduced collagen adhesion. A 60 kDa outer-membrane protein of A. pleuropneumoniae recognized the swine collagens by overlay. This membrane protein was apparently involved in adhesion to collagen and fibrinogen, but not to fibronectin and laminin. Antibodies against the 60 kDa protein inhibited the adhesion to collagen by 70 %, whereas pig convalescent-phase antibodies inhibited it by only 40 %. Serotypes 1 and 7 were the most adherent to pig collagen (taken as 100 %); serotypes 6 and 11 were the lowest (approximately 50 %), and neither showed the 60 kDa adhesin to biotinylated collagens. By negative staining, cells were observed initially to associate with collagen fibres in a polar manner, and the adhesin was detected on the bacterial surface. The results suggest that swine-lung collagen is an important target for A. pleuropneumoniae colonization and spreading, and that the attachment to this protein could play a relevant role in pathogenesis.

Adherence of actinobacillus pleuropneumoniae serotype 1 to swine buccal epithelial cells involves fibronectin.

The swine pathogen Actinobacillus pleuropneumoniae serotype 1 was investigated for its ability to adhere to swine, rat, and human buccal epithelial cells (BEC). The highest number of bacteria adhered was to swine BEC. This binding ability was affected by heating, extreme pH, treatment with sodium dodecyl sulfate, ethylenediamine tetra-acetate, or periodate, and proteolysis, suggesting that cell-surface glycoproteins participate in adherence and that adherence is based mostly on ionic interactions. Mannose and swine fibronectin may play a direct role in this interaction. Convalescent-phase serum from naturally infected pigs inhibited the adhesion. There was a correlation between bacterial pathogenicity as well as host specificity and the capacity for adherence to swine BEC. Adhesion to swine BEC provides a convenient method to study in vitro the adherence of A. pleuropneumoniae and other pathogens of the pig respiratory tract.

Actinobacillus pleuropneumoniae: virulence and gene cloning

Actinobacillus pleuropneumoniae is the causal agent of porcine contagious pleuropneumonia(PCP). The infection produces important economic losses in porciculture due to its high morbidity and mortality. Survivors are asymptomatic carriers infectious to other pig and have low alimentary conversion. The causative agent possesses several virulence factor: adhesion fimbriae, lipopolysaccharide of the outer membrane, capsule, and cytolysins. In addition, our group has reported secretion proteases of a wide pH range of activity. These proteases degrade different substrates such as porcine gelatin, hemoglobin and IgA, and bovine or human hemoglobin. To control PCP dissemination, farmers require serodiagnostic tests which detect carriers and discriminate between vaccinated and infected animal. Bacterines used as immunogens are serotype specific and do not prevent the infection. Genes have been cloned that codify a cohemolysin, cytolysins, and an iron-binding protein. We have cloned A. pleuropneumoniae genes using the expression plasmids pUC19 and Bluescript, in Escherichia coli Q358 and DH5alpha; the screening for antigen production was made in four gropus of pigs (vaccinated, experimentally infected, naturally infected, and from alaughterhouses); two E. coli clones expressed polypeptides recognized by sera from all the groups