Jpn. J. Infect. Dis., 55, 14‑18, 2002

Original Article

Prevalence ofEscherichia coli Possesslng the eaeA Gene of Enteropathogenic
E. coli (EPEC) Or the aggR Gene of Enteroaggregative E･ coli (EAggEC) in
Traveler's Diarrhea Diagnosed in Those Retumlng tO Tama, Tokyo

from Other Asian Countries
Kazue Ogatal, Rei Katol, Kenichiro lto3 and Sumio Yamadal.2*

J Tama Branch Laboratory, Too,o Metropolitan Research Laboratory OfPublic Health,Shibazaki‑cho 3‑16‑25, Tachikawa‑shi, To砂0 190‑0023,
20nice ofQualiO) Assurance, To砂o Metropolitan Research Laboratory Qflpublic Health,

Hyakunin‑cho 3‑24‑1, Shinjuku‑ku, To&0 16910073 and3National Institute of Public Health,

Shirokanedai 4‑6‑1, Minato‑ku, Too)o l08‑8638, Japan(Received June 1 1, 2001. Accepted January 29, 2002)

SUMMARY: To elucidate the importance of enteropathogenic Escherichia colt (EPEC) and enteroaggregative
E. colt (EAggEC) as etiological agents in traveler's diarrhea, the detection of the eaeA and aggR gene in E･ coli
strains isolated from overseas travelers with diarrhea in Tama, Tokyo was carried out uslng a PCR method. Of

192 travelers who were mostly adults and had visited Asian countries from April 1998 to March 1999, aggR‑
positive E･ colt straiTs were detected in 26 ( 1 3 ･5%)･ These stra空represent the second predonlinant enteropathogen

following enterotoxlgenic E. colt (ETEC), whereas eaeA‑posltlVe E･ coli strains were confi‑ed in seven subjects(3.6%). Tn 1 3 cases with aggR and four caseswith eaeA, the organisms were detected in stool samples of patientsas the only potential enteric pathogen. The clinical symptoms of these patients were similar to those in patientswith ETEC; however, the severlty Ofillness was milder than that associated with ETEC alone. Three strains with

eaeA and flVe Strains with aggR were typed as six different kinds of 0 serogroups, of which four strains
belonged to the classical EPEC serogroups (055, 01 14, 0119, and O127a)I These finding苧suggestthat aggR‑

positive E. coli (EAggEC) is a signiflCant Causative agent in traveler's diarrhea. ln addition, lt Was demonstrated
that eaeA‑positive E. colt (EPEC) is markedly correlated with diarrhea in adults.

associated with specific agg operons ( I 1 , 1 2). Tn recent studies,

INTRODUCTION

it was demonstrated that dgg operons consisted of several

Escherichia coIL'can cause gastrointestinal infectiops ( I )I

potential reading frames, e.g., aggA, aggB, aggC, aggD, andaggR (13,14). Furthermore, it was revealed that certain
EAggEC strains produce an STJlike toxin (enteroaggregative

Diarrheagenic E. coIL'can be grouped into five categorleS (2)Pn the basis of the types ofvirulence, i･e･, production of toxin,

ST, EAST)(15).

1nVaSiveness, patterns ofbacterial attachment to host cells,

and effects of attachment on host cells: enterotoxlgenic E.

ln some case‑Control studies of infant and childhood diarrhea,

coli (ETEC), enterohemorrhagic E. colt (EHEC), enteroinvasive

the pathogenic role ofEPEC in infants younger than 12 months

E. coli (EIEC), enteropathogenic E. colt (EPEC), and entero‑

old has been well described (16‑20); EAggEC has been

aggregative E. coli (EAggEC). Tn addition, it has been proposed

associated with persistent diarrhea among children (20‑22).

that difhsely adhering E. colt (DAEC) is diarrheagenic, but

In contrast, the slgnirlCanCe Ofthese organisms as etiologlCal

its role in acute diarrheal disease is controversial (3). In Japan,

agents in adult disease remains unclear. EAggEC was evaluatedstrains belonglng tO Classical EPEC serotypes have also been

as a slgnificant etiologlCal agent in recent studies of diarrhea

included as a. group or enteropathogens (4)･

diseases in North American travelers to Mexico (23) andEPEC stralnS usually exhibit localized adherence (LA) toepithelial cells and induce attaching and effacing (A/E) lesions

the role ofEPEC in traveler's dia汀hea has not been thoroughly

on enterocytes (5,6)･ EPEC virulence genes eafand eae areinvestlgated. ln Japan, traveler's diarrhea has also been arespectively associated with LA and A/E lesions (7‑9), andpublic health problem and has affected the actual states ofamong Spanish travelers to developing countries (24). However,recently, bundle‑forming pi]us (Bfp) has been reported as?negastrointestinal infection (25). To determine the role played byadherence factor (10). EAggEC strains show aggregatlVeadherence (AA) to HEp‑2 0r HeLa cells; such adherence ismediated by aggregative adherence fimbriae 1 (AAF/1)

EPEC and EAggEC as causative agents of traveler's dia汀hea

in Japanese travelers to developlng countries, the presence orabsence of the eaeA or aggR gene ofE. colt strains isolated

from overseas travelers in Tama, Tokyo, was examined by
PCR. ln addition, the clinical symptoms oftravelers infectedwith EPEC and EAggEC were investlgated.

*Corresponding author: Mailing address: O作ce of quality

Assurance, Tokyo Metropolitan Research Laboratory of Public
Health, Hyakunin‑cho 3124‑1, Shinjuku‑ku, Tokyo 169‑0073.

Tel: +81‑313363‑3231, Fax: +81‑3‑3368‑4060, E‑mail:yamada@tokyo‑eiken. go jp
14

Themixture was amplified by 25 cycles of denaturation at

MATERIALS AND METHODS

94oC f♭r 0.5 min, annealing at 50oC fわr 1.0 min, and exten‑

Stool samples: A total of 192 fecal specimens obtained
from travelers retumng to Japan from other countries, mostly

PC‑700 (ASTEC Co., Ltd., Tbkyo). Ten microliters of the

Asian countries, during the period from April 1998 to Marchreaction mixture were then analyzed by electrophoresis onsion at 72oC for 1.5 min with Program Temp Control System

1999 were examined. Stool specimens collected fTrom persons

1.5% agarose gels, and the reaction products were visualized

under quarantine or receivlng medical care were received in

by stainlng With ethidium bromide.

Cary and BlaiトtranSpOrtation medium at public health centers

0‑serotyping ofE･ coli isolates and eaeA‑ or aggR‑ ppsトin Tama, Tokyo, and were transferred within 2 days to ourtive strains: 0 antlgenS Were determined by slide agglutlna‑

laboratory. Health cards of the travelers accompanied thespecimens; these cards included infbmation such as countries
visited and clinical symptoms. Infbmation on the cards wastion tests with kits using E. coli antisera (I) (Denka‑Seiken).
Adherence assay: For the eaeAl 0r aggR‑ positive E. coli
isolates, marlnOSe‑resistant adhesion to HEp‑2 cells was tested

glVen by the traveler at the time of stool sample collection.

by the method described by Cravioto et al. (29).

Standard procedures: Stool samples were cultured bypreviously described methods f♭r bacterial pathogens (26).

RE StJIJTS

For the diagnosis of diarrheagenic E. colt, three suspectedcolonies on DHL agar (Nissui Phamaceutical Cot, Ltd., Tbkyo)

The travelers examined were mostly adults, and were acuteplates were randomly selected and screened biochemically.

or convalescent‑phase patients. According to the standard

The E. colt strains thus identified were tested for a classicalprocedure (4), enteropathogens were detected in 87 0日92

EPEC serotype, ETEC, and Shiga‑toxin (STX) producing‑E.
colt (STEC)･ In brief, boiled cells ofEI COli strains werecases (45.3%). As shown in Table 2, by the introduction ofserogrouped by a slide agglutination test with the fbllowlng

the aggRIPOSitive E. coli was detected in 26 cases (13.5%),

types of commercial antisera from Denka‑Seiken Co., Ltd.

whereas eaeA‑positive E. Colt was confirmed in only seven

PCR assays to detect eaeA or aggR genes ofEPEC or EAggEC,

0126, 0127a, 0128, 0146, and 0166. Forthe toxin tests, E.

cases (3.6%). Three cases with eaeA‑E. colt and 1 3 with aggR‑
E. coliinvolved a possible mixed infection. Consequently, acolt strains were inoculated into casamino acid‑yeast extract

tota一 of 1 04 cases were pathogen‑positive, and the positivlty

broth (Nissui) and were shaken at 370C fわr overnight. The

ratio increased to 54.2%. Number orenteropathogens isolated(Tokyo): 026, 044, 055, 086, 0111, 0114, 0日9, 0125,

culture was centrifuged for 20min at 23,000 × g, and thewere 1 62 strains, in which most prevalent one was the ETEC.

supernatant was examined for heat‑labile toxin (LT) by CT‑

RPLA (Denka‑Seiken), for heat‑stable toxin (ST) by COuST

Next most prevalent pathogen isolated was the aggRIPOSitive
E. colt, followed by Plesiomonas shigelloides, Aeromonas

EIA (Denka‑Seiken), and fわr STX by VT‑RPLA (Denka‑

spp･, Campylobacter spp., classical EPEC serotype, Salmonella

Seiken).

spp., Vibrio parahaemolyticus, and eaeA‑positive E. colt.

PCR assay: The various prlmerS used for the detection of
the virulent genes ofEPEC and EAggEC are listed in Table 1

infection with PlesEbmonas, A e710mOnaS SPP. and Campylobacter

Three of the patientswith eaeAIPOSitive E. Coli had mixed(27,28), The determination of EPEC and EAggEC in the

spp., or AelVmOnaS SPP. and aggR‑positive E. colt'. One‑halfstrains tested was perfonned by the identification of the eaeA

(13 cases) ofall aggRIPOSitive E. Colt cases were detected

gene, and aggR gene. Furthennore, the eaeA I 0r aggRIPOSitive

together with one to fわur other enteropathogens. They werestrains were tested for the presence of the eafand bfpA genes,

classified in ten patterns, and involved combinations oror the astA gene. ln brief, stock cultures were hoculated onto
nutrient agar slants and incubated at 370C ovemight. A small

Aeromonas spp･, V. parahaemolyticus, and eaeA‑positive E.

cluster orbacteria was suspended in 100 〝l ofdistiHed water,

L･OIt'.

Camplyobacter spp., Plesiomonas, Salmonella spp., ETEC,

The suspected countries where infectionwith eaeA‑positiveand boiled at 94oC for 5 min. After a short centrifugation (Imin, 18,000 × g), 4.8 lil oftemplate were mixedwith 5 Lilof 10 XPCRbuffer (Takara Shuzo Co. Ltd., Kyoto), 3 LEI of
Tablc 2. Detection ofenteropathogens from overseas travelers

25 mM MgCl, 2 LEI of2.5 mM dNTP (Takara Shuzo), 0.1 Lilwith diarrhea jn Tama, Tokyo (April 1998‑March 1999)

of50 uM sense prlmer and antisense prlmer, 0.2 〃l ofTaq
Numbcr examined

DNA polimerase (Takara Shuzo), and 35 LLl of distilled water.

Number positive for pathogens (%)

1 92

1 04 (54.2)

No. orcases with (%):
Table L PCR prlmerS used in this study

Gene

PrimereaeA eaek 1 *eaek4*

Sequence

Location

GCTTAG TGCTGGTTTA GGAT
TCGCCGTTCAG A GATCGC

66‑85

554‑537

EAF EAF‑B* *TGG ATCG C CAATGTTCTTGG

14ト160EAF‑SR**

ATGGGG ACCATGTATTATC A

922‑941

E!rpA bfps**

Enterotoxlgenic E. coli

36 (18.7)

aggR gene‑positive E. colt26(13.5)

P/eLu'(,m()naS Shigelloides

25 (13,0)

Aeromonas sppI

18( 9.4)

Campy/Obacter spp ,12( 6.2)

Classical EPEC serotypell( 5.7)*

Salmonella spp.

10( 5.2)

V. parahaemolvticus

10( 5.2)

G AAGTAATGAGC GCAACGTC

154‑173

ACATGCCGCTTTATCCAACC

387‑368

eaeA gene‑positive E. colt7( 3.6)

GTATACACAAAAGAAG GAAGC

loo‑120

Shigella spp.

3( 1.6)

aggRkas2* *ACAG AATC G TCAGCATC AG C

353‑334

Verotoxin‑producing E. coli2( 1.0)astA EASTOS 1 **G CCATC AACACAGTATATC CG

3‑23

V. cholerae 0‑ I ttox十)1( 0.5)CGCGAGTGACGGCTTTGTAG

11ト92

V. cholerae 0‑1 (tox一)1( 0.5)

bfpas**

aggR aggRksl**

EASTOAS2*** : Reference28.

* : One eaeA gene‑positive E. C(Jli and three aggR gene‑positive** : Reference 27.

E. cL7/i were observed.

15

E. coIL'occurred were Thailand (2 cases), Indonesia (2 cases),Viet Nam ( 1 case), and others (2 cases). Those countries where(2 strains each), and 055, 01 19, 0125, 0126, 0127a, 0142,
or 0146 (1 strain each). Four of the strains mentioned above

aggR‑positive E. Colt infection occurred were Thailand (8were positive for either the eaeA or aggR gene. In contrast,cases), Indonesia (5 cases), India (5 cases), Viet Nan (2 cases),Philippines (2 cases), Iran (1 case), and South Africa (I case).

the ETEC and verotoxin‑producing E. coli (VTEC) strainsthat were isolated did not possess either of the genes.

The remainlng two Cases Were detected in travelers who had

visited other Asian countries. There was no special seasonal

DISCUSSION

variation associated with either of the E. coli isolations.
The eaeA‑ and aggR‑positive strains were characterizedEnteric infection due to EPEC or EAggEC is characterized

serologlCally, phenotyplCally, and genetically; on一y threeby acute diarrhea in infants or chronic diarrhea in children in

eaeA‑positive strains and five aggR‑positive strains weredeveloping countries. In previous studies (23,24), EAggEC

typed to particular serogroups, namely, 0153 (2 strains) andstrains have been involved in the etiology or traveler's dia汀hea

055 (I strain) in the fbmer, and 015 (2 strains), 01 14, 0119,
or 0 1 27a (each I strain) in the latter. Among.serotyped strains,
eaeA‑positive E. coli 055, and aggRIPOSltlVe E. coli 01 14,
01 19, and O127a had already been detemined as one of theclassic EPEC groups. In the HEp‑2 cell adherence assay, atypical LA pattern was observed in only two of the eaeAIin North American travelers to Mexico and in Spaniards traveレ

mg to developlng COuntries. However, an association between

the presence orEAggEC or EPEC strains in stool samples ofdia汀hea in adults has not been conclusively demonstrated,
In order to dete‑in° the potentially causal role orthe EAggEC

and EPEC strains in traveler's diarrhea, genetic diagnosis bypositive strains, whereas 1 4 aggR‑positive strains showed the

PCR assay was performed on the E. colt strains isolated fTrom

typlCal AA pattem. Furthermore, none of the eaeA‑positive

overseas travelers presentlng With diarrhea in Tam乱, Tokyo.

strains possessed the eqfand bhA gene, but 1 2 aggR‑positive
To clarify the severityofillness and the clinical symptoms

The aggRIPOSitive E. coli was detected in 26 (13.5%) of the
1 92 cases oftraveler's diarrhea; thus, this strainwasthe second
most prevalent strain after ETEC. Similarly, Mathewson etof patients infected with eaeA‑, or aggR‑positive E. co/i,al. (23) reported that E. colt strains showing mannose‑resistantstrains were asIA‑positives.

retrospective investlgations were ca汀ied out on 1 6 patients,

focal adherence were detected in 1 7 cases (9.0%) of the North

among whom either of the two strains was fわund as the only

American travelers studied. Recent work by Gascon et al.

potential enteric pathogen; these cases were compared with(24) showed that the plasmid DNA fわr EAggEC was present

those in which patterns were infected with ETEC alone. As

in 23 cases (13.9%) among the Spanish travelers included inshown in Table 3, the severlty Ofthe illness and the clinicalthat study. Typically, aggRIPOSitive E. co/i strains were

symptoms of patients infected with aggR‑positive E. coli anddetected among travelers to Southeast Asian countries or tothose witll ETEC were almost identical, although a predomi一the Indian subcontine叫and geographic distribution was

れance of mild cases with diarrhea occurrlng less than fわur

similaJ･ tO that observed in the case ofETEC. On the othertimes per day was observed in aggR‑positive E. colt infec‑

hand, EAggEC strains confirmed in the study of Spanish

tions. In addition, abdominal cramps were obseⅣed in 69%

travelers were isolated in travelers retumlngfrom all geographicof the cases; a few of the cases were accompanied by feverareas, especially from the Indian subcontinent, West Africa,(3 1%), nausea, and/Or vomiting (38%). A fTew patients infectedwith eaeA‑positive E. colt also experiencedmildly wateryand Central America.
Among the cases in which aggR‑positive E, coli wasdiarrhea and headache or fever, but none reported abdominal

detected, 1 3 cases involved this organism as the only potential

Cramps ･enteropathogen. The clinical symptoms associated with theClassic EPEC serotype strains were detected in ll cases;
these which serotypes belonged to serogroups 0 1 14 and 0 1 28

aggR‑positive strain were watery diarrhea (6 I %), abdominal
cramps (69%), fever (31%), and nausea or vomiting (39%)･
How㌣er, the severlty Ofillness was milder than that obseⅣed

in patlentS With ETEC infection alone. Tn the investlgation of

Spanish travelers, Gascon et al, reported that five of 23
EAggEC strains caused chronic diarrhea (>14 days); theclinical symptoms of patients with other EAggEC strains wereTable 3. Symptoms and severlty Oftraveler's diarrhea in patientsfrom whom eaeA or aggR gene positive E. colt‑ was isolated as
the only potential enteric pathogen% of symptoms with:similar to those reported in our results. The present andeaeA‑E. Colt aggRIE. COILI ETEC*

previous findings indicate that EAggEC is important as an

Characteristics
of patients(ll‑4)(n‑13)

etiologlCal agent in travelers retumlng With gastrointestinal(a‑25)

illness from a developlng country.

Character Of stool

Watery1 00 0 0

As shown in the report byAlbert et al. (20), EPEC is a′n ヽヽ.

l 1 0nLooseslgnificant enteric pathogen in infants of up to 12 months ofBloodyage; the prevalence of this pathogen tends to diminish withage. ln the present study, jt was suggested that the eaeA‑positive

Maximum nulllber of diarrhea episodes per day

Mild(1‑3)10054Moderrate (4‑6)

0

3fS

Severe (>7)

O

8

0

69

Abdolllinal cramps

E. Colt strain is markedly correlated withadult diarrhea,

2 5 2 ′nU て｣ 4 2 4 4 ∠U

although the occu汀enCe Of such strains was lower than thatof EAggEC, which accounted fわr seven cases (3.6%). Fourof seven eaeA‑positive E. colt strains were identified as only
potential enteropathogens, and their patients had mild cases

Nausea/vomitlng

50

39

Fever

50

31

Additional clinical symptoms were nausea and/Or vomltlng,

H eadache50

15and fever (2 cases e,ach); however, no patients in this groupwith watery dia汀hea occumng less than fわur times per day.

experienced abdomlnal cramps.

* : Patients infected with enterotoxigenic E. colL‑ (ETEC) alone during

Although it is or interest to study the correlations between

the sallle period.

lbcertain serotypes and particular gene‑positive E. coli strains,mucosa. Infect. Immun., 55, 69‑77.

only three eaeA‑positive strains and five aggR‑positive strains

7. Donnenberg, M. S. and Kaper, ∫. (1992): Enteropath0‑

could be typed in the present study･ Of which one eaeA‑

genic Escherichia coli. Infect. Immun･, 60, 3953‑3961 ･

positive E. coli strain and three aggR‑positive E･ colt strains
belonged to the classic EPEC serogroups; 055 in the case of
eaeA and Ol14, 0119, and O127a in aggR. Among theremainlng four strains, two eaeA‑positive strains were 0 1 53,
and two aggR‑positive strains were 01 5. These results agree

8. Giron, ∫.A., SucYue Ho,A. and Schoolnik, G. K. (1991):

An inducible bundle‑formlng Pilus of enteropathogenic
Escherichia coli. Science, 254, 710‑713.

9. Jerse,A. E., Yu, J., Tall, B. and Kaper, J. B. (1990): A
genetic locus of enteropathogenic Escherichia coli neces‑with those ofMathewson et al. (23) and Gascon et al. (24),sary for the production of attaching and effacing lesions

who also described that only two of28 enteroadherent E. colion tissue culture cells. Proc. Nath. Acad. S°i., 87, 7839‑

strains isolated from North American travelers belonged tothe classical serogroup 044, and none of the EAggEC strains

10. Grion, ∫. A., Donnenberg, M. S., Martin, W. C., JaⅣis,7843.

K. G. and Kaper, J. B. (1993): Distribution of the bundle‑
forming pilus structural gene (bfpA) among enteropath0‑

isolated from the Spanish cases was positive for the classic

serotypes. On the other hand, in a previous study on acuteinfantile diarrhea (1 8), 70% of LA‑E. coli strains belonged

genic Escherichia coli. J. Infect. Dis., 168, 1037‑1041･
1 1. Vial, P. A., Robins‑Browne, R., Lior, H., Prado, V, Kaper,to the classical EPEC 0 serogroups. Moreover, C血an et al･(30) demonstratedthat the predominant serogroups of AA‑E･J. B., Nataro, ∫. P., Maneval, D., EIsayed, A. E. andcoli strains from acute and chronic diarrhea were 044, 0 I 1 I ,Levine, M. M. ( 1 988): Characterization ofenteroadherent

and 0126. These differences may indicate that there arebroader serogroups of EPEC or EAggEC than are currently
recognized and that differences in the specific 0 serogroups areaggregative Escherichia coli, a putative agent of diarrheal

disease. J. Infect. Dis., 158, 70179.
12. Nataro, J. P., Deng, Y., Maneval, D. R., German, A. L.,

predominant in infant, childhood, and adult types of diarrhea･Martin, W. C. and Levine, M. M. (1992): Aggregative

The identification ofEPEC and EAggEC has been performed

adherence fimbriae I of enteroaggregative Escherichia

by obseⅣlng the adherence pattems to HEp‑2 cells, or by ac(フJf mediate adherence to HEp‑2 cells and hemagglutina‑

combination of cell assay and hybridization with DNA probesof virulence factors. However, several studies have demon‑
strated that 37% of adherent strains ofE. colt exhibit atypicalor non‑characteristic pattems ( I 7), andthatthere are differencesamong the results ofcell assays and DNA‑hybridization tests
as regards LAIE. coli strains (17, 19,20). In present study, twoeaeA‑positive E. coli strains and 14 aggR‑positive E. colttion of human erythrocytes. Infect. Immun., 60, 2297‑
2304.

13. Savarino, S. J., Fox, P., Yikang, D. and Nataro, J. P.
(1994): Identification and characterization of a genecluster mediating enteroaggregative Escherichia coli
aggregative adherence fimbria I biogenesis. ). Bacterio1.,
176, 4949‑4957.

strains showedtypical adherence patterns, but the other strahs

14. Nataro, J. P., Yikang, D., Yingkang, D. and Walker, K.

could not be identified. In addition, none of the eaeA‑positive(1 994): AggR, a transcriptional activator of aggregative

E･ colt str.aims possessed the bhA or eafgene, whereas 12

adherence fimbria I expression in enteroaggregative

aggR‑posltlVe E. colt strains were astA‑positive. Further

Escherichia colt. J. Bacterio1,, 176, 4691‑4699.

studies will be necessary ln Order to determine whether EPEC

15, Savarino, S, JリFasano,A., Robertson, D. C. and Levine,

or EAggEC strains play a slgnificant role in infantile andM. M. (199 1): Enteroaggregative Escherichia coli elabo‑

childhood diarrhea, or in gastroenteritis in adults.

rate a heat‑stable enterotoxin demonstrable in an in vitro
rabbit intestinal model. ∫. Clin. invest., 87, 1450‑1455.

16. Comes, T. A. T., Rassi, V, MacDonald, K.し, Romos, S.

REFERENCES

R. T. S., Trabulsi,し. 良., Vieira, M.A. M., Guth, B. E. C.,l･ Gray, L D･ (1995): Escherichia, Salmonella, Shigella,
and Yersinia. p.4501456. In P.R. Murray, E. J. Baron, M.
A. Pfaller, et al. (eds.), Manual ofClinical Microbilogy.
6th ed. AMS Press, Washington.

Candieas, ∫. A. N., lvery, C., Tbledo, M. 良. 已 and Blake,

P･ A･ (1991)‥ Enteropathogens.associated with acutediarrheal disease in urban infantlS in S畠o Paulo, Brazil.

J. Infec. Dis., 164, 3311337.

2. Salyers, A. A. and Whitt, D. D. (1994): Escherichia colt

17. Forestier, C., Meyer, M., Favre‑Bonte, S., Rich, C.,gastrointestinal infection. p. 1 901204. In A. A. Salyers andMalpuech, G., Le Bougenec, C., Sjrot, ∫., Joly, 良. and de

D･ D Whitt (eds･), Bacterial Pathogenesis‑a molecularapproach･ AMS Press, Washington.

Champs, C. ( 1 996): Enteroadherent Escherichia colt anddiarrhea in children: a prospective case‑control shdy. ∫.

Clin. Microbio1., 34, 2897‑2903.

3･ Comes, T･ A., Blake, P. A. and Trabulsi, L 良. (1989):

Prevalence of Escherichia coli strains with localized,

18. Gonzalez, 良., Diaz, C., Marino, M., Cloralt, 良., Pequeneze,di凪ISe, and aggregative adherence to Hela cells in infants

M･ and Perez‑Schael, I. (1 997): Age‑Specific prevalencewith diarrhea and matched controls. J. Clin. Microbio1.,of Escherichia colt with localized and aggregative
adherence in Venezuelan infants with acute diarrhea. J.

27, 266‑269.

Clin. Microbio1., 35, 1103‑1107.
19･ Comes, T･A･ T･, Vieira, M.A. M.,Abe, C. M., Rodrigus,

4･ Zen‑Yozi, H., Sakai, S., Terayama, T., Kudoh, Y. and Ito,

T･ ( 1 985): Manual for Examination of Enteropathogenic
Bacteria･ 4th ed･ Igakushoin, Tokyo (in Japanese).

D･, Grifrln, P･ M･ and Ramos, S･ R･ T･ S･ (1998): Adheren?e5. Nataro, J. P,, Kaper, J. B., Robins‑Browne, R., Prado, V.,patterns and adherence‑related DNA sequences ln

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

Escherichia colt isolates from children with and withoutof diarrheagenic Escherichia coli to HEp‑2 cells･ Pediatr.
Infect. Dis., 6, 8291831.

diarrhea in S孟O Paulo clty, Brazil. J. Clin. Microbio1.,

36, 3609‑3613.

6. Knutton, S., Lloyd, D. R. and McNeish,A. S. (1992):

20. Albert, M. ∫., Faruque, A. S. G., Sack, 良. B. andAdhesion of enteropathogenic Escherichia coIL'to human

Mahalanabis, D. (1999): Case‑Control study ofLintestinal enterocytes and cultured human intestinal

enteropathogens associated with childhood diarrhea in17

21. Baqui,A. H., Sack, R. B., Black, R. E., Haider, K., Hossain,

Tnfect. Agents Surveillance Rep., 16, 217'‑218'･
26. Yamada, S., Matsushita, S., Dejsirilert, S. and Kudoh, Y.

A.,Alim,A. R. M. A., Yunus, M., Chowdhury, H. R. and( I 997): Incidence and clinical symptoms ofAeromonas‑

Dhaka, Bangladesh. ∫. Clin. Microbio1., 37, 3458‑3464･

Siddique, A. K. (1 992): Enteropathogens associated withassociated travellers'diarrhea in Tokyo. Epidemiol.

acute and persistent diarrhea in Bangladish children <5

Tnfect., 119, 1211126.

years of age. J. lnfec. Dis., 166, 792‑796･
22. Fang, G. D., Lima,A.A. M., Martins, C. V., Nataro, J. P.

27. Ratchtrachenchai, 0‑A., Subpasu, S. and lto, K. (1997):
Tnvestlgation on enteroaggregative Escherichia colt

and Guemnt, 良.し. (1995): Etiology and epidemiology of

infection by multiplex PCR. Bull. Dept･ Med･ Sci･, 39,
211‑220.

persistent dia血ea in northeastem Brazil: a hospital‑based,

28. Yamazaki, M., Saito, M., 1nuzuka, K., Shima, S., Taniwaki,

prospective, case‑control study. ∫. Pediatr. Gastroenterol.

Nutr., 21, 137‑144.

H. and Ito, K. (1997): eaeA genes in Escherichia coli

23. Mathewson, J. J,, Jolmson, P. C., DuPont, H. L., Morgan,
D. R., Thomton, S. A., Wood, L V and Ericsson, C. D.

derived from Japanese patients with sporadic diarrhea.

J. Jpn. Assoc. Infect. Dis., 71, 105911065.
29. Cravioto, A., Tello, A., Navarro, A., Ruiz, J･, Villajan, H.,
Uribe, F. and Eslava, C. (1 979): An adhesive factor foundin strains of Escherichia coli belonglng tO the traditional
infantile enteropathogenic serotypes. Curr. Microbiol.,(I 985): A new recognized cause of travelers'diarrhea:
enteroadherent Escherichia colt. J. Infec. Dis., 15 I, 471‑
475.

241 Gascon, J･, Vargas, M･, Quinto, L･, Corachan, M･, JimeTez

3, 95‑99.

de Anta, M. T. and Vila, ∫. (1998): EnteroaggregatlVe

Escherichia coli strains as a cause of traveler's diarrhea:

30. °han, K. N., Phillips,A. D., Knutton, S., Smith, H. 良. anda case‑control study. J. Infec. Dis., 177, 1409‑1412.
25. National Institute of Health and Infectious Diseases

coli: another cause of acute and chronic diarrhoea in

Walker‑Smith, J. A. ( I 994): Enteroaggregative Escherichia

ControI Division, Ministry of Health and Welfare (1995):

England? ∫. Pediatr. Gastroenterol. Nutr., 18, 87‑91.

Traveler's diarrhea, Japan, January 1992‑July 1995.

18