Relatedness of Streptococcus suis isolates of various serotypes and clinical backgrounds as evaluated by macrorestriction analysis and expression of potential virulence traits

Abstract

We evaluated the genetic diversity of Streptococcus suis isolates of different serotypes by macrorestriction analysis and elucidated possible relationships between the genetic background, expression of potential virulence traits, and source of isolation. Virulence traits included expression of serotype-specific polysaccharides, muramidase-released protein (MRP), extracellular protein factor (EF), hemolysin activity, and adherence to epithelial cells. Macrorestriction analysis of streptococcal DNA digested with restriction enzymes SmaI and ApaI allowed differentiation of single isolates that could be assigned to four major clusters, named A1, A2, B1, and B2. Comparison of the genotypic and phenotypic features of the isolates with their source of isolation showed that (i) the S. suis population examined, which originated mainly from German pigs, exhibited a genetic diversity and phenotypic patterns comparable to those found for isolates from other European countries; (ii) certain phenotypic features, such as the presence of capsular antigens of serotypes 2, 1, and 9, expression of MRP and EF, and hemolysin activity (and in particular, combinations of these features), were strongly associated with the clinical background of meningitis and septicemia; and (iii) isolates from pigs with meningitis and septicemia showed a significantly higher degree of genetic homogeneity compared to that for isolates from pigs with pneumonia and healthy pigs. Since the former isolates are considered highly virulent, this supports the theory of a clonal relationship among highly virulent strains.

FIG. 1

Macrorestriction analysis of S. suis isolates by PFGE. (A) PFGE of SmaI-digested DNA of S. suis serotype 9 isolates (lanes 1 to 20 and 23) and, in addition, serotype 2 hemolysin activity-positive control strain P1/7 (lane 21) and serotype 2 MRP- and EF-positive control strain D282 (lane 22). Lane M, size standard (ApaI-digested genomic DNA from S. aureus strain DSM 1104) indicating molecular sizes (in kilobases). (B) PFGE of ApaI-digested DNA of S. suis serotype 2 isolates (lanes 1 to 24) including the type 2 reference strain (lane 5), MRP- and EF-positive control strains 4005 (lane 7), T15 (lane 8), and D282 (lane 23), as well as hemolysin activity-positive control strain P1/7 (lane 20). Lane M, size standard (ApaI-digested genomic DNA from S. aureus, strain DSM 1104) indicating molecular sizes (in kilobases).

FIG. 2

Dendrogram of similarity among the observed PFGE macrorestriction patterns of SmaI-digested DNAs from 99 S. suis isolates. Serotypes (ST) and clinical backgrounds (CB) are indicated on the right. Isolates originated either from pigs with typical invasive disease (meningitis [M], septicemia [S], arthritis [A]), from pigs with pneumonia (P), or from healthy pigs (H). Major clusters are indicated by A1, A2, B1, and B2. nt, nontypeable.

FIG. 3

Comparison of combinations of different features of S. suis isolates with their clinical backgrounds. Additional features of serotype 2 isolates (A), isolates expressing MRP and EF (MRP/EF isolates) (B), hemolysin-positive isolates (Hly isolates) (C), and cluster A1 isolates (A1 isolates) (D) are shown. The leftmost pair of bars in each graph (labeled Total) refers to all isolates expressing the indicated feature; labeling of the remaining pairs of bars indicates the additional feature found in isolates of the respective group. Each pair of bars shows the proportion of isolates from pigs with meningitis-septicemia (gray bars) compared to the total proportion of isolates with this combination of features (black bars). Results are expressed as a percentage of the total number for each group (N), which is indicated in boxes above the respective groups.