Clinical significance of Escherichia albertii.
We collected 278 eae-positive strains that were originally identified by routine diagnostic protocols as EPEC or EHEC. They were isolated from humans, animals, and the environment in Japan, Belgium, Brazil, and Germany during 1993-2009 (Table 1; online Technical Appendix, wwwnc.cdc.gov/pdfs/11-1401-Techapp.pdf). To characterize the strains, we first determined their intimin subtypes by sequencing the eae gene as described (online Technical Appendix). Of the 275 strains examined, 267 possessed 1 of the 26 known intimin subtypes (4 subtypes-[eta], v, [tau] and a subtype unique to C. rodentium--were not found). In the remaining 8 strains, we identified 5 new subtypes; each showed <95% nt sequence identity to any known subtype, and they were tentatively named subtypes N1-N5. For subtype N1, 3 variants were identified (N1.1, N1.2, and N1.3, with >95% sequence identity among the 3 variants) (Figure 1, panel A).
To determine the phylogenetic relationships of the strains, we performed multilocus sequencing analysis of 179 strains that were selected from our collection on the basis of intimin subtype and serotype (see online Technical Appendix for selection criteria and analysis protocol). Among the 179 strains, 26 belonged to the E. albertii lineage (Figure 2). The 26 E. albertii strains were from 14 humans (13 from symptomatic patients), 11 birds, and 1 cat. All of the 5 new intimin subtypes were found in the E. albertii strains. Intimin subtypes found in other E. albertii strains were also rare subtypes found in E. coli (10). This finding suggests that more previously unknown intimin subtypes may exist in the E. albertii population.
We next analyzed thepheV, selC, andpheUloci of the 26 E. albertii strains for the presence of LEE elements as described (online Technical Appendix). These 3 genomic loci are the known LEE integration sites in E. coli. By this analysis, all E. albertii strains except 1 (EC05-44) contained the LEE in the pheU locus (the integration site in EC05-44 was not identified). This finding indicates that despite the remarkable diversity of intimin subtypes, the LEE elements are preferentially integrated into the pheU tRNA gene in E. albertii.
Because all E. albertii strains isolated so far contained the cdtB gene encoding the cytolethal distending toxin B subunit (8,9), we examined the presence and subtype of the cdtB gene as described (online Technical Appendix). This analysis revealed that all E. albertii strains except 1 (CB10113) possessed the cdtB gene belonging to the II/ III/V subtype group (Figure 1, panel B); this finding is consistent with published findings (9). In addition, 2 strains (E2675 and HIPH08472) each of which was subtype I, possessed a second cdtB gene, (Figure 1, panel B).
We used PCR to further investigate the presence of Shiga toxin genes (stx) and their variants (online Technical Appendix) and found that 2 E. albertii strains possessed the stx2fgene (Figure 2, panel B). Stx2 productionby these strains was confirmed by using a reverse-passive latex agglutination kit (online Technical Appendix). The 2 stx2f-positive strains were those containing the subtype I cdtB gene in addition to the II/III/V subtype group gene: 1 (HIPH08472) was isolated from a patient with diarrhea and the other (E2675) was from a healthy Corvus sp. bird (Figure 2).
Last, we examined the phenotypic and biochemical properties of the 26 E. albertii strains and compared the results with those obtained in a previous study (9) and with those of E. albertii type strain LMG20976 (Table 2). To identify features that could discriminate E. albertii from E. coli, the results were further compared with those of E. coli (11). Consistent with findings in previous reports (5-7,9), the lack of motility and the inability to ferment xylose and lactose and to produce [beta]-D-glucuronidase were common biochemical properties of E. albertii that could be used to discriminate E. albertii from E. coli, although 1 E. albertii strain was positive for lactose fermentation. The inability of E. albertii to ferment sucrose has been described as a common feature (9); however, a positive reaction to this test was found for 5 (19.2%) E. albertii strains. Moreover, approximately half of E. coli strains are positive for sucrose fermentation. Thus, the inability to ferment sucrose is not informative. Rather, the inability to ferment dulcitol (all E. albertii strains were negative, 60% of E. coli strains are positive) may be a useful biochemical property for differentiation.
In the current clinical laboratory setting, a substantial number of E. albertii strains are misidentified as EPEC or EHEC. Because 13 of the isolates were from patients with signs and symptoms of gastrointestinal infection, E. albertii is probably a major enteric human pathogen. In addition, E. albertii should be regarded as a potential Stx2f-producing bacterial species, although the clinical significance of Stx2f-producing strains is unknown.
Notable genetic, phenotypic, and biochemical properties of E. albertii, which were identified by analyzing the confirmed E. albertii strains, are 1) possession of intimin subtypes rarely or previously undescribed in E. coli, 2) possession of the II/III/V subtype group cdtB gene, 3) LEE integration into thepheUtRNA gene, 4) nonmotility, and 5) inability to ferment xylose, lactose, and dulcitol (but not sucrose) and to produce [beta]-D-glucuronidase. These properties could be useful for facilitating identification of E. albertii strains in clinical laboratories, which would in turn improve understanding of the clinical significance and the natural host and niche of this newly recognized pathogen. In this regard, however, current knowledge of the genetic and biological properties of E. albertii might be biased toward a certain group of E. albertii strains because, even with this study, only a limited number of strains have been analyzed. To more precisely understand the properties of E. albertii as a species, further analysis of more strains from various sources is necessary.
This work was supported by the following KAKENHI (Grants-in-Aid for Scientific Research) grants from the Ministry of Education, Culture, Sports, Science and Technology of Japan: Applied Genomics, 17019058, to T.H.; Kiban-B, 20310116, to T.H.; and Wakate-B, 23790480, to T.O.
Dr Ooka is an assistant professor at the Department of Infectious Diseases, Faculty of Medicine, University of Miyazaki. His research interests include the genomics and pathogenicity of pathogenic bacteria, especially attaching and effacing pathogens.
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Author affiliations: University of Miyazaki, Miyazaki, Japan (T. Ooka, Y. Ogura, T. Hayashi); Osaka Prefectural Institute of Public Health, Osaka, Japan (K. Seto); Miyazaki Prefectural Institute for Public Health and Environment, Miyazaki (K. Kawano); National Institute of Animal Health, Ibaraki, Japan (H. Kobayashi); Fukuoka Institute of Health and Environmental Sciences, Fukuoka, Japan (Y. Etoh, S. Ichihara, K. Horikawa); Yamagata Prefectural Institute of Public Health, Yamagata, Japan (A. Kaneko); Toyama Institute of Health, Toyama, Japan (J. Isobe); Hokkaido Institute of Public Health, Hokkaido, Japan (K. Yamaguchi); Universidade Federal de Sao Paulo, Sao Paulo, Brazil (T.A.T. Gomes); University of Liege, Liege, Belgium (A. Linden, M. Bardiau, J.G. Mainil); and Federal Institute for Risk Assessment, Berlin, Germany (L. Beutin)
Address for correspondence: Tetsuya Hayashi, Division of Bioenvironmental Science, Frontier Science Research Center, University of Miyazaki, Kihara 5200, Kiyotake, Miyazaki 889-1692, Japan; email: email@example.com
Table 1. Summary of 275 eae-positive strains originally identified by routine diagnostic protocols as EPEC or EHEC * Origin No. strains Human, n = 193 Symptomatic 154 Asymptomatic 7 No information 32 Animal, n = 76 Bird 38 Pig 31 Cat 1 Deer 1 Bovid 1 Sheep 1 No information 3 Environment, n = 6 6 * EPEC, enteropathogenic Escherichia coli; EHEC, enterohemorrhagic E. coli. Table 2. Comparison of biochemical properties of Escherichia spp. strains 26 E. albertii E. albertii strains (this LMG20976 Agent or test study) ([dagger]) (type strain) Indole 96.2 - Motility 0 - Urea 0 - ONPG 88.5 + MUG 0 - Citrate 0 - Acetate 92.3 + Malonate 0 - [H.sub.2]S on triple sugar iron 0 - Voges-Proskauer 0 - Lysine decarboxylase 100 + Ornithine decarboxylase 100 + Arginine dihydrolase 0 - Glucose, acid 100 + Glucose, gas 100 + Acid from Adonitol 0 - L-arabinose 100 + Cellobiose 0 - Dulcitol 0 - Myo-inositol 0 - Lactose 3.9 - Maltose 88.5 + Mannitol 100 + L-rhamnose 0 - Salicin 26.9 - D-sorbitol 57.7 - Sucrose 19.2 - Trehalose 96.2 + D-xylose 0 - E. albertii E. coli Agent or test strains (9) (11) ([dagger]) Indole 100 98 Motility 0 95 Urea 0 1 ONPG ND ND MUG ND (+) ([double dagger]) Citrate 0 1 Acetate ND 90 Malonate ND 0 [H.sub.2]S on triple sugar iron ND 1 Voges-Proskauer ND 0 Lysine decarboxylase 100 90 Ornithine decarboxylase 100 65 Arginine dihydrolase 0 17 Glucose, acid 100 100 Glucose, gas 100 95 Acid from Adonitol ND 0 L-arabinose 100 99 Cellobiose ND 2 Dulcitol ND 60 Myo-inositol ND 1 Lactose 0 95 Maltose ND 95 Mannitol 100 100 L-rhamnose 0 0 Salicin ND 40 D-sorbitol V 94 Sucrose 0 50 Trehalose ND 98 D-xylose 0 95 * ONPG, ortho-nitrophenyl-fi-galactoside; MUG, methylumbelliferyl- [beta]-D-glucuronide; -, negative; +, positive: ND, not determined. ([dagger]) Average (%) of positive strains. ([double dagger]) Most E. coli strains produce [beta]-D-glucuronidase.
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|Author:||Ooka, Tadasuke; Seto, Kazuko; Kawano, Kimiko; Kobayashi, Hideki; Etoh, Yoshiki; Ichihara, Sachiko; K|
|Publication:||Emerging Infectious Diseases|
|Date:||Mar 1, 2012|
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