Jpn. J. Infect. Dis., 53, 2000

Laboratory and Epidemiology Communications

Plasmid Encoded Enterotoxin (Pet) Gene in Enteroaggregative
Escherichia coli Isolated from Sporadic Diarrhea Cases
Mitsugu Yamazaki*, Kazuhisa Inuzuka. , Hironori Matsui, Kenji Sakae, Yasumoto Suzuki,
Ⅶtaka Miyazaki and Kenitiro lto2

A ichi Prefectural Institute of Public Health,
Nagare 7‑6. Tsuji‑machi, Kita‑ku, Nagoya 462‑85 76,)Anjo Kosei Hospital,

Miyukihonmachi 12‑38, Anjo 446‑8602 and2National Institute ofPublic Health,

Shirokanedai 4‑6‑I, Minato‑ku. To&0 1 08‑8638
Communicated by Yutaka Miyazaki(Accepted February 5, 2001 )Table･ Detection ofpetand asIA genes in 65 aggR gene positive isolates

Nataro et al.first reported that some Escherichia coli isolates

from diarrhea patients had a capacityto adhere to HEp‑2 cells

petand the surface of glass Petri dishes, and named them

(+)

SerO

pettypes(‑)

SerOtypeSEnteroaggregative E. colt (EAggEC) (I). Its relation to0126:H27

persistent diarrhea in infants and HIV patients and to someastA (+)

diarrheasinadults has been indicated, thoughnot conclusively

120126:HUT

30111:H21

110111:HUT

10127a:H212(2).
Total

The aggregative adherence of EAggEC was due to the15Total

086a: H27

presence of aggregative adherence fimbriae (AAF), AAF/ 1
and AAF/lI, whose expression lS positively controlled by theaggR gene (2)･ This aggR gene is present in many EAggECs,
but some aggR‑positive E. coli isolates have no AAFs (3,4).
EAggEC was found to produce EAggEC heat‑stable entero‑toxin (EASTl encoded by astA, molecular weight 4.1 kDa)I

424 1 2 7 2086a:H9086a:HUT

asLA (‑)

055:H100126:H19
Total(2) and EAggEC plasmid encoded toxin (Pet encoded by pet,Total

molecular weight 108 kDa) (5). Among them, EASTl is
widely distributed in pathogenic E. coli regardless of theaggregative adherence phenotype (2). Nataro et al. reported

01

Tota15(23%)

3650(77%)

The figures indicate number of isolates.

that two strains of EASTトpositive EAggEC produced

55oC for 45 see, and at 72oC for 60 see. The strain 042 (5)different results when administered to volunteers (6); thewas used as apet gene positive control. Pet negative controls

difference between the two strains was later found to residein thepet gene (5).

were I 7‑2 and JM22 1 (092:H33) strainsshowin.g aggregative

Asthe above data appeared to suggest that the pet geneadhesion (7), and 25 1 (OUT:HUT) strain showlng dimISed

adhesion (5,7), 886L (0111:H2) strain showlでg localized

was involved in the pathogenlClty Of EAggEC, and as noadhesion (7)･ The genes ofaggRand asIA were detected using

epidemiological investlgation has beenperformed on the gene

the method described by Moriya et al. (8).

in Japan, We studied the incidence ofthepet gene in E･ Colt

Among the 364 isolates which reacted to the Enteropath0‑

isolated from diarrhea patients treated in a hospital in Aichi

genic E･ colt 0 Typing Sera, 65 had the aggR gene.Asshown

Prefecture from January 1 989 to December 1 992.

Among isolates kom 9,684 patients (7,597from the pediatricin the Table, thepet gene was detected in 15 isolates (23%)
among aggR positives. Eslava et al. reported that 15% ofE.

ward and 2,087from the intemal medicine Yard), 364 isolates

Coli of the EAggEC phenotype isolatedfrom diarrhea patientsfrom different patients were aggregated using commerciallyavailable Enteropathogenic E. coli 0 Typing Sera (Denka

werepet gene‑positive (5). Czeczulin et al. reported that 1 8%

Seiken, Tokyo) (7)･ The 364 isolates were screened for theof EAggEC isolated from Thailand, Mexico, Peruand ninepresence ofthepet gene by PCR using PnmerS Which were
designed based on the pet sequence in GenBank accession

other countries were pet gene‑positive and that, among aggR‑in six countries including Thailand, Peru, and the PhilipplneS

positive EAggECs, 24% were pet gene‑positive (3). Therefore,

No. AFO56581. The sense prlmer Was 5'‑TTTCCAG

Our data was qulte comparable to those ofpreviousreports･

CACTTCCTGTTCC‑3', and the anti‑sense prlmer Was 5'‑

All the 15 pet‑positive isolates were astA‑positive, andATTTCCAACGTCTACGCCAT‑3 I. PCR amplification

belonged to the 0126 serotype.AsMoriya et al. (8) reported

consisted of30 cycles of incubation at 94oC fわr 60 sec, atthat 0 I 26 E･ Coli isolated kom healthy people possessed aggRand astA, it remains to be elucidated whether or notpet is*Co汀eSpOnding author: E‑mail :mitsugu｣′amazaki@mail.prer

directly associated with the pathogeniclty Of 0 126. Strains
ofserotypes ofO33:H16, 044:H18, 01 I 1 :H2, and OUT:HIO

aichijp, Fax: +81‑52‑913‑3641

248

Jpn. J. Infect. Dis., 53, 2000
isolated from the diarrhea of children have been reported tocoli. Infect. Immun., 67, 2692‑2699.

possess pet (3)･ These serotypes †ere not included in this4･ Okeke, Ⅰ. N., Lamikanra, A., Czeczulin, J･, Dubovsky,

study because none of them were lSOlated in this study. In

ln conclusion, 17.9% (65/364 isolates) of 0 sero‑typable

F･, Kaper･ J･ B･ and Nataro･ J･ P･ (2000): Heterogene?us
virulence of enteroaggregative Escherichia coli strains
isolated from children in southwest Nigeria. J. Infect.
Dis., 181, 252‑260.
5. Eslava, C., Navarro‑Garcia, F. Czeczulin, J. R., Henderson,Enteropathogenic E. coli isolates from diarrhea patients had

Ⅰ. 良., Cravioto, A. and Nataro, ∫. P. (1998): Pet, anorder to clarifythe role of pet in pathogeniPity,further study
will be required by examinlng thepet gene ln a Wide range ofdia汀hea patient isolates including the above serotypes.

aggR, and 23% (15/65) of these aggR‑positive E. coli had

autotransporter enterotoxin from enteroaggregative

pet gene. This frequency was qulte comparable to the previousEscherichia colt. Infect. Immun., 66, 3 155‑3 163.

studies perfomed in developing countries.

6･ Nataro, J･ P., Yikang, D., Cookson, S., Cravioto, A.,
Savarino, S. J., Guers, L D., Levine, M. M. and Tacket,
C. 0. (1995): Heterogeneity or enteroaggregative

Strains042 and 1 7‑2 were kindly obtained from Dr. James

Nataro, and JM221, 886L, and 251 from Dr. Blay Kay.

Escherichia colt virulence demonstrated in volunteers.

J. Infect. Dis., 171, 465‑468.
7. Yamazaki, M., Saito, M., huzuka, K., Shima, S., Taniwaki,REFERENCES

H. and Ito, K. (1997): eaeA genes in Escherichia coltderived from Japanese patients with sporadic diarrhea.

1. Nataro, ∫. P., Kaper, ∫. 良., Robins‑Browne, 良., Plado, V,J. Jpn.Assoc. Infect. Dis., 71, 105911065.

Vial, P. and Levine, M. M. (1987): Pattems of adherence

8. Moriya, K., Sumi, N., Nakao, M., Yamazaki, M., Saito,
M. and lto, K. (2000): The prevalence of virulence‑of diarrheagenic Escherichia colt to HEp‑2 cells. Pediatr.

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Garcia, F. and Nataro, J. P. (1999): Phylogenetic analysis

with diarrhea. J. Jpn.Assoc. Infect. Dis., 74, 134‑141(inof enteroaggregative and di軌sely adherent Escherichia

Japanese).

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