Difficulty in detecting low levels of polymyxin resistance in clinical Klebsiella pneumoniae isolates: evaluation of Rapid Polymyxin NP test, Colispot Test and SuperPolymyxin medium.

Polymyxins are important therapeutic options for treating infections, mainly those caused by carbapenem-resistant Klebsiella pneumoniae. Specific chemical characteristics of polymyxins make it difficult to perform antimicrobial susceptibility testing, especially within the clinical laboratory. Here we aimed to evaluate the performance of three phenotypic methods: Rapid NP Polymyxin Test, ColiSpot test and the SuperPolymyxin medium. To accomplish this, 170 non-duplicate clinical K. pneumoniae isolates were analysed (123 colistin-resistant and 47 susceptible). The sensitivity and specificity obtained for Rapid Polymyxin NP Test, Colispot and SuperPolymyxin medium were, respectively, 90% and 94%, 74% and 100%, and 82% and 85%. Very major errors occurred more frequently in low-level colistin-resistant isolates (MICs 4 and 8 µg/mL). Rapid Polymyxin NP proved to be a method capable of identifying colistin-resistant strains in acceptable categorical agreement. However, major errors and very major errors of this method were considered unacceptable for colistin-resistance screening. Although the Colispot test is promising and easy to perform and interpret, the results did not reproduce well in the isolates tested. The colistin-containing selective medium (SuperPolymyxin) showed limitations, including quantification of mucoid colonies and poor stability. Nevertheless, Colispot and SuperPolymyxin medium methods did not present acceptable sensitivity, specificity and categorical agreement. It is essential to use analytical tools that faithfully reproduce bacterial resistance in vitro, especially in last-line drugs, such as polymyxins, when misinterpretation of a test can result in therapeutic ineffectiveness.

Identification of a 43-kDa outer-membrane protein as an adhesin in Aeromonas caviae.

Aeromonas spp. are associated with intestinal and extra-intestinal infections. However, the virulence factors of A. caviae remain, for the most part, poorly known. This study examined the interactions involved in the adherence of A. caviae isolates Ae56, Ae391 and Ae398 to HEp-2 cells. All strains expressed high levels of aggregative adherence. Maximum adhesion occurred with bacteria grown at 22 degrees C, but transmission electron microscopy did not reveal the presence of fimbrial structures on the bacterial cell surface. Outer-membrane proteins (OMPs) extracted from isolate Ae398, grown at 22 degrees C and 37 degrees C, showed similar SDS-PAGE protein profiles. Most proteins were < 60 kDa. A major 43-kDa protein was seen only in the boiled OMP extract. The biotinylated 43-kDa protein bound specifically to HEp-2 cells. Microbeads coated with the 43-kDa protein were also adherent to HEp-2 cells, and anti-43-kDa protein antibody blocked adherence of 43-kDa protein-coated latex beads. These data suggest that the 43-kDa OMP functions as an adhesin in A. caviae.