The scFv antibody towards the Burkholderia pseudomallei exotoxin was previously constructed by phage display and exhibited good specificity towards the exotoxin. We report here the optimization of the scFv expression in an E. coli expression system. Four different E. coli strains (ER2537, TG1, HB2151, and XL1-Blue) were examined for optimal expression of the scFv protein. Two types of carbon source (i.e. 0.2% glucose and 0.2% glycerol) were also tested for their ability to induce the scFv expression. Cells that carried the scFv construct were grown at 30 degrees C and induced with 0.05 mM IPTG. The expression was then monitored by SDS-PAGE, Western blotting, and indirect ELISA. The Western blot profile showed different levels of the scFv expression among the host strains; XL1-Blue exhibited the highest level of the scFv protein expression. Glycerol at a concentration of 0.2% (v/v) significantly increased the scFv protein expression level when compared to 0.2% (w/v) glucose. Further optimization demonstrated that the scFv protein expression in XL1-Blue was the most optimal with a glycerol concentration as low as 0.05%. However, by indirect ELISA, only the scFv protein that was expressed in 0.2% (v/v) glycerol exhibited high specificity towards the Burkholderia pseudomallei exotoxin.
Eighty-six clinical isolates of Aeromonas hydrophila were studied for their ability to produce four exotoxins: a haemolysin active against rabbit erythrocytes, cytotoxin and enterotoxin detectable with Vero cell cultures, and the cholera toxin-like factor detected by an enzyme-linked immunosorbent assay. At least one exotoxin was produced by 80% of enteric and 96% of non-enteric isolates. The exotoxin profiles of non-enteric isolates were more restricted than those of enteric isolates, with haemolysin and cytotoxin producers preponderant. Although haemolysin and cytotoxin were produced by isolates from all sources, the enterotoxin and cholera toxin-like factor were more common amongst enteric isolates. The production of haemolysin and cytotoxin were closely related but the association between the enterotoxin and the cholera toxin-like factor was not significant.
A 61-year-old male presented with community-onset pneumonia not responding to treatment despite given appropriate antibiotics. Computed tomography scan of the thorax showed large multiloculated pleural effusion with multiple cavitating foci within collapsed segments; lesions which were suggestive of necrotising pneumonia. Drainage of the effusion and culture revealed methicillin-resistant Staphylococcus aureus, which had the same antibiotic profile with the blood isolate and PVL gene positive.
Panton-Valentine leukocidin (PVL) is a cytotoxin which causes leukocyte destruction and tissue necrosis. Although it is produced by fewer than 5% of Staphylococcus aureus strains, PVL-producing S. aureus is emerging as a serious problem worldwide. There has been a marked increase in the incidence of necrotizing lung infections with a very high mortality associated with these strains. This report describes a fatal case of hospital-acquired necrotizing pneumonia caused by PVL-positive methicillin-susceptible S. aureus in a patient with a brain tumor.