Assessment of Shiga toxin-producing Escherichia coli isolates from wildlife meat as potential pathogens for humans

Abstract

A total of 140 Shiga toxin-producing Escherichia coli (STEC) strains from wildlife meat (deer, wild boar, and hare) isolated in Germany between 1998 and 2006 were characterized with respect to their serotypes and virulence markers associated with human pathogenicity. The strains grouped into 38 serotypes, but eight O groups (21, 146, 128, 113, 22, 88, 6, and 91) and four H types (21, 28, 2, and 8) accounted for 71.4% and 75.7% of all STEC strains from game, respectively. Eighteen of the serotypes, including enterohemorrhagic E. coli (EHEC) O26:[H11] and O103:H2, were previously found to be associated with human illness. Genes linked to high-level virulence for humans (stx(2), stx(2d), and eae) were present in 46 (32.8%) STEC strains from game. Fifty-four STEC isolates from game belonged to serotypes which are frequently found in human patients (O103:H2, O26:H11, O113:H21, O91:H21, O128:H2, O146:H21, and O146:H28). These 54 STEC isolates were compared with 101 STEC isolates belonging to the same serotypes isolated from farm animals, from their food products, and from human patients. Within a given serotype, most STEC strains were similar with respect to their stx genotypes and other virulence attributes, regardless of origin. The 155 STEC strains were analyzed for genetic similarity by XbaI pulsed-field gel electrophoresis. O103:H2, O26:H11, O113:H21, O128:H2, and O146:H28 STEC isolates from game were 85 to 100% similar to STEC isolates of the same strains from human patients. By multilocus sequence typing, game EHEC O103:H2 strains were attributed to a clonal lineage associated with hemorrhagic diseases in humans. The results from our study indicate that game animals represent a reservoir for and a potential source of human pathogenic STEC and EHEC strains.

FIG. 1.

Dendrogram based on PFGE patterns of EHEC O103:H2 strains (Tables 3 and 4). Game isolates are marked in boldface characters. The 22 strains fall in three major clusters, namely, clusters A, B, and C (>68% similarity), and form subclusters in clusters A (A1 and A2) and B (B1 and B2). Strain 327/98 from a mixed game meat assortment grouped in subcluster B1 (88% similarity) together with four strains from cattle, milk, and two human patients. Strain 327/98 shows 95% similarity to a bovine strain (GSO740-1) and to a strain from a HUS case (CB5500). Two EHEC O103:H2 strains from wild boar (351/04) and roe deer (105/03) grouped in subcluster B2 (90% similarity) together with three strains from cattle and humans. Strain 351/04 is 100% similar in its PFGE profile to the human strains CB08223 and D242/02-3. In cluster C, strain RL0523/05 from hare meat shares 79% similarity with a bovine isolate (RW2198) and a human isolate (CB8198).

FIG. 2.

Dendrogram based on PFGE patterns of EHEC O26:H11 strains (Tables 3 and 4). Game isolates are marked in boldface characters. Ten strains subdivided into major clusters A and B (82% similarity). Strain D618/98 from cheese was different with respect to its stx_2d gene and showed a nonclustering PFGE pattern. Cluster A (84% similarity) consists of five strains originating from deer meat, cattle meat, and a human patient. The PFGE patterns of strain 331/02 (deer meat) and bovine strains D316/04 and CB07505 were 92% similar. Cluster B (85% similarity) consists of two strains from hare meat and two human strains. The strains from hare meat were 88% similar to the human RL 06/0524 strain.

FIG. 3.

Dendrogram based on PFGE patterns of STEC O113:H21 strains (Tables 3 and 4). Game isolates are marked in boldface characters. Twenty-two of the 24 strains grouped in major clusters A and B (82% similarity). Subcluster A1 is composed of seven strains from deer meat, beef, and cattle. The strain from deer meat (001/05) shows 91% similarity in its PFGE profile to the strains from cattle. Subcluster A2 harbors three strains with 93% similarity: one each from deer meat (140/04), a beef-pork isolate (RL0464/05), and a human strain (CB08578). Subcluster A3 (89% similarity) was composed of a strain each from wild boar meat, from cattle, and from beef. Five strains from red deer meat (206/99, 539/99, RL 06/0377, 641/99, and 014/04) grouped in subcluster B1 together with strain D366/99 from lamb meat (90% similarity).

FIG. 4.

Dendrogram based on PFGE patterns of STEC O91:H21 strains (Tables 3 and 4). Game isolates are marked in boldface characters. Fourteen of the 17 strains grouped into four clusters designated A (82% similarity), B (85%), C (89%), and D (94%). Three strains (170/00 from wild boar meat, RL06/0393 from deer meat, and RL06/0468 from pork) presented individual PFGE patterns. Strains 092/02 and RL 06/0303 from deer meat grouped in cluster A together with a human strain (CB07753) and a bovine strain (D392/99) (84 and 82% similarity). No strain from game was present in clusters B, C, and D.

FIG. 5.

Dendrogram based on PFGE patterns of STEC O128:H8 strains (Tables 3 and 4). Game isolates are marked in boldface characters. All but 1 (RL0529/05) of the 34 STEC O128:H8 strains were arranged in clusters A and B. Strain RL0529/05 represents the only isolate lacking the stx_1c gene and shows a nonclustering PFGE pattern. Cluster A (79% similarity) was formed exclusively by strains from deer meat isolated between 1999 and 2006. Subcluster B1 (84% similarity) was composed of a strain from deer meat (RL 06/0367) and three STEC strains from sheep. Subcluster B2 (85% similarity) was formed by 21 strains; 10 of these, including 2 deer, 2 lamb, 1 sheep, and 5 human strains, showed 91% similarity in their PFGE profiles. One strain from game (467/01) showed 95% similarity to all of the nongame strains grouped in subcluster B2.

FIG. 6.

Dendrogram based on PFGE patterns of STEC O146:H21 strains (Tables 3 and 4). Game isolates are marked in boldface characters. Two strains from game (264/04 and RL06/0466) present in this group formed PFGE cluster A (73% similarity). All other STEC O146:H21 strains except DG074/2 grouped into cluster B (>74% similarity) and were from sheep, beef, and humans. The bovine DG074/2 strain was different from all other STEC O146:H21strains with respect to its stx_1d gene and showed a nonclustering PFGE profile. The two STEC strains from game were less than 69% related to the other STEC O146:H21 strains and lacked the stx_1c gene which was present in all but one (CB07743) cluster B strains.

FIG. 7.

Dendrogram based on PFGE patterns of STEC O146:H28 strains (Tables 3 and 4). Game isolates are marked in boldface characters. The 26 O146:H28 strains divided into major clusters A, B, and C (>72% similarity). High-level similarities between strains from game and strains from patients were found in clusters B2 (RL0510/05 and CB08026 [92% similarity]) and B4 (RL0160/05 and CB07787 [90% similarity]). In cluster B3, one strain from roe deer (RL06/0123) and one from beef (D166/00) showed 92% similarity. Lower levels of similarity between strains from game and other sources were found in cluster A (75%) and cluster C (81%).