Genotyping of environmental and clinical Stenotrophomonas maltophilia isolates and their pathogenic potential

Abstract

Stenotrophomonas maltophilia is a highly versatile species with useful biotechnological potential but also with pathogenic properties. In light of possible differences in virulence characteristics, knowledge about genomic subgroups is therefore desirable. Two different genotyping methods, rep-PCR fingerprinting and partial gyrB gene sequencing were used to elucidate S. maltophilia intraspecies diversity. Rep-PCR fingerprinting revealed the presence of 12 large subgroups, while gyrB gene sequencing distinguished 10 subgroups. For 8 of them, the same strain composition was shown with both typing methods. A subset of 59 isolates representative for the gyrB groups was further investigated with regards to their pathogenic properties in a virulence model using Dictyostelium discoideum and Acanthamoeba castellanii as host organisms. A clear tendency towards accumulation of virulent strains could be observed for one group with A. castellanii and for two groups with D. discoideum. Several virulent strains did not cluster in any of the genetic groups, while other groups displayed no virulence properties at all. The amoeba pathogenicity model proved suitable in showing differences in S. maltophilia virulence. However, the model is still not sufficient to completely elucidate virulence as critical for a human host, since several strains involved in human infections did not show any virulence against amoeba.

Figure 1. Dendrogram based on (GTG)5 and BoxA1R fingerprint profile similarities.

Analysis was performed by using Pearson's correlation with UPGMA clustering. Cophenetic correlation values are shown for each branch. Grey bars at the branches are representing the error bars. In each condensed cluster assigned as genetic grouop similarity values (%) ± standard deviation (%) are shown. Reference strains from previous AFLP/MLST groupings were highlighted with specific symbols. Reference isolates accounted to 16S rRNA groups E2 were shown with a black point. Isolates characterized additionally by gyrB gene sequencing (Figure 2) were marked with coloured boxes representing the gyrB genetic group. Strains belonging to no gyrB group were highlighted in grey.

Figure 2. Neighbour-joining tree based on partial gyrB gene sequences.

Neighbour-joining tree of Stenotrophomonas strains with Xanthomonads and Xylella fastidiosa used as outgroups. Reference strains from previous AFLP/MLST groupings were highlighted with specific symbols. Reference isolates accounted to 16S rRNA groups E2 were shown with a black point.

Figure 3. Dictyostelium discoideum plate killing assay.

Bars are representing the number of amoeba necessary to form a plaque on the bacterial lawn of the 59 different isolates. The mean of at least three experiments was determined for each strain. Standard deviation is indicated with error bars. Genetic groups as determined by gyrB gene sequencing are pointed out as A–J.

Figure 4. Acanthamoeba castellanii plate killing assay.

Bars are representing the number of amoeba necessary to form a plaque on the bacterial lawn of the 59 different isolates. The mean of at least three experiments was determined for each strain. Standard deviation is indicated with error bars. Genetic groups as determined by gyrB gene sequencing are pointed out as A–J.