Isolation and characterization of two plasmids in a clinical Acinetobacter nosocomialis strain.

BACKGROUND: Acinetobacter species are recognised as important nosocomial pathogens that have become a major cause of invasive opportunistic infections in hospitalised patients. Their clinical significance is largely due to the rapid development of antimicrobial resistance among strains. The development of antibiotic resistance among bacterial strains occurs frequently by the acquisition of resistance genes by gene transfer systems such as bacterial plasmids. METHOD: Multi-antibiotic resistant Acinetobacter nosocomialis strain 178 was isolated from a hospital in Melbourne, Australia. This strain was screened for the presence of plasmids. The two plasmids isolated were sequenced and annotated. RESULTS: Two plasmids isolated from a single clinical Acinetobacter nosocomialis strain were sequenced. One plasmid, designated pRAY*-v3, appears to have evolved via the same lineage as the pRAY plasmid isolated from an Acinetobacter baumannii in South Africa. The other plasmid, designated pAB49-v1, appears to be an evolutionary descendent from a cryptic plasmid isolated from an A. baumannii almost 20 years ago. Both of the plasmid sequences here share a high level of sequence similarity with their ancestors, however differences are noted. CONCLUSION: The isolation of these plasmid-lineages across different decades and continents suggests their global dissemination.

Insertions or deletions (Indels) in the rrn 16S-23S rRNA gene internal transcribed spacer region (ITS) compromise the typing and identification of strains within the Acinetobacter calcoaceticus-baumannii (Acb) complex and closely related members.

To determine whether ITS sequences in the rrn operon are suitable for identifying individual Acinetobacter Acb complex members, we analysed length and sequence differences between multiple ITS copies within the genomes of individual strains. Length differences in ITS reported previously between A. nosocomialis BCRC15417T (615 bp) and other strains (607 bp) can be explained by presence of an insertion (indel 13i/1) in the longer ITS variant. The same Indel 13i/1 was also found in ITS sequences of ten strains of A. calcoaceticus, all 639 bp long, and the 628 bp ITS of Acinetobacter strain BENAB127. Four additional indels (13i/2-13i/5) were detected in Acinetobacter strain c/t13TU 10090 ITS length variants (608, 609, 620, 621 and 630 bp). These ITS variants appear to have resulted from horizontal gene transfer involving other Acinetobacter species or in some cases unrelated bacteria. Although some ITS copies in strain c/t13TU 10090 are of the same length (620 bp) as those in Acinetobacter strains b/n1&3, A. pittii (10 strains), A. calcoaceticus and A. oleivorans (not currently acknowledged as an Acb member), their individual ITS sequences differ. Thus ITS length by itself can not by itself be used to identify Acb complex strains. A shared indel in ITS copies in two separate Acinetobacter species compromises the specificity of ITS targeted probes, as shown with the Aun-3 probe designed to target the ITS in A. pitti. The presence of indel 13i/5 in the ITS of Acinetobacter strain c/t13TU means it too responded positively to this probe. Thus, neither ITS sequencing nor the currently available ITS targeted probes can distinguish reliably between Acb member species.