Table 1.
Characteristics of the bacterial strains used in this study.
Figure 1.
Functional distribution of the genes found in all six A. baumannii strains included in this study.
Figure 2.
Distribution of genes in individual strains of the A. calcoaceticus – A. baumannii complex.
(A) Venn diagram showing the number of overlapping genes between the four strains that make up the A. calcoaceticus – A. baumannii complex. (B) Pairwise comparisons of the number of genes present in A. baumannii ATCC 19606T but absent in each of A. calcoaceticus (blue), A. pittii (green) and A. nosocomialis (red).
Figure 3.
Genetic organisation and conservation of the siderophore clusters found in A. baumannii ATCC 19606T and not in A. calcoaceticus.
(A) Siderophore cluster 1 (operons 36–39) is known as the acinetobactin chromosomal cluster, and (B) siderophore cluster 2 (operon 17) (See Table 2 for details about the operons). The presence of homologues for each gene in A. pittii, A. nosocomialis, and A. calcoaceticus is shown.
Table 2.
Select operons with putative virulence function found in A. baumannii ATCC 19606T and not in A. calcoaceticus.
Figure 4.
Metabolic diversity of specific strains of the A. calcoaceticus – A. baumannii complex.
(A) Phenotype Microarray (PM) comparative results showing the number of compounds used (green) or not used (red) by A. baumannii ATCC 19606T [A], A. nosocomialis [B], A. pittii [C] and A. calcoaceticus [D]. The external circle and PM number represent the Biolog plate number. (B) A. baumannii ATCC 19606T is able to metabolise the carbon source, D–glucarate and produce α–Ketoglutarate through the functional enzymes, D–glucarate dehydrogenase and KDG dehydratase. α–Ketoglutarate is then utilized in the citrate cycle. These enzymes are not found in A. calcoaceticus.
Figure 5.
Virulence attributes of individual strains belonging to the A. calcoaceticus – A. baumannii complex.
(A) Adherence of A. baumannii ATCC 19606T, A. pittii, A. nosocomialis and A. calcoaceticus to human bronchial epithelial cells after 1 hour. Results are expressed as mean number of bacteria per 100 epithelial cells ± standard deviation (SD) of two independent experiments performed in duplicate. The asterisk signifies statistical significance (P<0.05) between A. pittii and A. baumannii. The comparison between A. pittii and A. calcoaceticus was also significant (P<0.05). (B) Levels of IL–6 and (C) IL–8 in the culture medium of human bronchial epithelial cells after 24 hour stimulation with specific strains of the A. calcoaceticus – A. baumannii complex. Results are expressed as mean levels of IL–6 and IL–8 (in ng/ml) ± SD of three independent experiments. Asterisk signifies statistical significance (P<0.05) between A. pittii and A. baumannii. The comparison between A. pittii and A. calcoaceticus was also significant (P<0.05).. (D) Persistence and biofilm formation of A. baumannii ATCC 19606T (squares), A. pittii (upward triangles), A. nosocomialis (downward triangles) and A. calcoaceticus (diamonds) on three–dimensional human skin constructs. Results are expressed as mean CFU per skin construct ± SD of three independent experiments. Dotted line represents the lower limit of detection. Asterisk signifies statistical significance comparing A. calcoaceticus with A. baumannii ATCC 19606T (P<0.05) (E) Alcian–blue PAS staining shows biofilm formation (black arrow) on human skin constructs by A. baumannii ATCC 19606T but not by (F) A. calcoaceticus. Scale bar is equivalent to 20 µm. (G) Approximately 1×104 CFU were injected in the thigh muscles of neutropenic mice and the number of viable bacteria was determined after 48 hrs. Results are expressed as mean number of bacteria (in CFU/muscle) ± SD from three animals. Dotted line represents lower limit of detection. Asterisk signifies statistical significance (P<0.05) between A. baumannii and A. nosocomialis or A. calcoaceticus.