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

gene (tdh) and the thermostable direct hemolysin‑related

T. and Shoji, T. (1 999): Isolation of thermostable direct
hemolysin‑producing Vibrio parahaemolyticusfrom food
implicated in food poISOnlng in Mie Prefecture. Infect.
Agents Surveillance Rep., 20, 1 89‑ 192 (in Japanese).
4. Arakawa, E., Murase, T., Shimada, T., Okitsu, T., Yamai, S.

hemolysingene (blh) of Vibrio parahaemolyticus by poly‑and Watanabe, H. (1999): Emergence and prevalence ofmerase chain reaction. Mol. Cell. Probes, 6, 477‑487.

a novel Vibrio parahaemolyticus 03 :K6 cloneinJapan.
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363.

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Y. (1 992): Detection of the thermostable direct hemolysin

3. Sugiyama, A., Nakano, Y., Iwade, Y., Yamauchi, A.,Nakayama, 0., Matsumoto, T., Kou, Y., Ito, T., Nishinaka,

Laboratory and Epidemiology Communications
Occurrence of Scrub Typhus (Tsutsugamushi) in Kanagawa Prefectureand Types of Orientia tsutsugamushi Involved
Yumiko Furuya* , Takashi Katayama, Miyuki Hara, Yoshiya Yoshida,
Mitsunobu lmai and Tbshikatsu Hagiwaral
Kanagawa Pylefectural Public Health Laboratonl,
Nakao I‑Ill, Asahi‑ku, Yokohama 241‑0815 Kanagawa andLNational Institute of lnfectious Diseases,

Toyama 1‑23‑1, Shinjuku‑ku 16218640 Too/0
Communicated by Shigetou Suzuki(Accepted May 9, 2000)

and Kuroki, but, with PCR using type‑specific primers, theScmb typhus, or Tsutsugamushi disease, is caused byOrientia tsutsugamushi. Kanagawa Prefecture near Tokyo

flrSt and the last specimens were positive for only the Kuroki

experienced a large epidemic of scrub typhusin198811989. It
then wanned:in1996 only 9 cases were reported. However,type, while the second and third were positive for only thethe epidemic now appears to be in resurgence. From April

tended to show higher antibody titers for both Gilliam andKawasaki types (#98043, #98044, and #98045). This is

Kawasaki type. Sera PCR‑positive for the Kawasaki type1997 to March 1998, 17 clinical cases were reported. FromApril 1998 to March 1999, 21 cases were reported,and fromApril 1999 to March 2000, 43 cases were reported. The

probably becausethe 56 kDa proteins of the bothtypes shareda corrmon epitope (5). However, some sera (ex., #9033) wereinfection was most frequent in November. In 1997, 8 cases

positive for onlyantibody against the Kawasakitype. Use ofoccurred in November and 1 case occurred in December. In1998, 13 cases occurred in November and 2 cases occurred

all the five type antigens, i.C., the Kawasaki and Kuroki typeantigens in addition to the three standard antigens, Gilliam,in December. In 1999, 1 case occurred in May, 28 cases in

Karp and Kato typeantigens, is recommended to increase

November, and 6 cases in December.

the sensitivity of antibody detection.

All these patients were examined for serum antibody (1)and for the 0. tsutsugamushi genome whenever the blood

Acute phase sera were generally positive for the genome
in PCR. However, some sera, such as #98047, which attained

from patients in the disease's the acute phase was available

highantibody levels alreadyinthe acute phase were negative(2)･ The serumantibody yas detected using the immunofluo‑
rescent teclmique employlng antigens of Gilliam, Karp, Kato,for the genome.

Kawasaki, and Kuroki types (1). Sera showing more than a

80% of the cases were due to the Kawasaki type, a tendency
which is in agreement with the data obtainedinMiyazaki (1)and Chiba Prefectures (6). But 3 cases (6.4%) wereuntypable;
this may be due to mutadons which madethe genome detecdon

Table 2 summarizes immunological and PCR data. About

4‑foldincrease ofIgM or IgG titers in the disease's convales‑
cent phaseincomparison withthe acute phase orthose showing
IgM titers exceeding X 80 in the acute phase were consid‑
ered positive. The 0. tsutsugamushi genome was detecteduslng neSted PCR using genus Orientia‑Specific pnmers andimpossible uslng the five sets of primers used.

The work was supported by a grant‑in‑aid by the Ministryof Health and Welfare of Japan.

type‑Specific primers (3, 4).
A portion of the data for individual patients is reproducedin Table 1. Sera #98038, #8044, #98045, and #98046 reactedwith all of the antigens of Gilliam, Karp, Kato, Kawasaki,

REFERENCES*ComSpOnding author: Fax: +8 1 ‑45‑363‑1 037

1. Yamamoto, S., Kawabata, N., Ooura, K., Murata, M.and77

Jpn. J. Infect. Dis., 53, 2000
Table 1. Representative data of serodiagnosisand typing by PCR

Antibody Detection
PCR3)
sampleDays))

Gilliam

KarpKatoKawasaki

Kuroki

Diag2)(Type)

IgM IgG IgM IgG IgM IgG IgM IgG IgM IgG
OO OO

彬 脚 000 ∠UO 8 5 4 0 7 4 ′LU 2 5 4 2 つJ 8

つJ つJ

7 n■UtJ 1 l 1 2 2

鵬 紳

00

つJ 4

00

004

4<10<10<10<10<10<10<104040<1040<1020160

00

00

040

45/▲U(XU Oノ紳 紳 M 007 ∠U#9903340<10<10160640

160

160

40

320

80

160

80

160

80

640

<10<10<10<10<10<10<10<10<10<1040<10<10<10<10<1080

160<10<1040<10<10<10<10<10160<10<10320320

4080

2080

640

640

2040

<10<10<10<10<10<10<10<10<10<10160

320

10160

320 1280

1080<1010208080320408 %<1010<1040160
20<103204020<10103202020160160160160160160

4040<10<10<10<10<10<10<10<10<10<10<10<10<10<1080<10<10<10<10<10<10<10<10<10<10<10<10<10<10<1040<10血4)+(Kw)++( Kr)++(Kw)++(Kw)++(Kw)++( Kr)

40 1280
160 1280<10+<10<10<10+<10<10+(ut)<10++(Kw)<101)Days after orLSet Ofillness. 2)Finalresult ofserodiagnosis. 3)Result ofPCR (Types: Kw, Kawasaki; Kr, Kuroki; ut,untypable). 4)Finalresults not obtained.

Table 2. Irrmunofluorescent Antibody Detection (IF) and Typing by PCR
TotalIF+PCR‑

IF+PCR+samples
19̲9̲9 恥9 7粥99 山KwTyping

Krbyut

PCRCon丘mIedcases′0 0 ′0 2tJ 4 5 2

1 2 4 0 7 1 つJ 1

IfLPCR+

Kp014 5584001

0 3 つJ1029̲ 535 59

12 42●■l一137

63(2) (79) (13) (6)'Figures indicate number of samples. lF+PCR‑: positive for IF only. IF+PCR+: positive for bothIF and PCR.
IF‑PCR+: positive for PCR only. Typing by PCR: Kp stands for Karp, Kw for Kawasaki, Kr for Kuroki, andut for untypable. * Percentage of each type.

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tsutsugamushi isolatedfrom patients withTsutsugamushifeverand their disbibution in Miyazaki Prefecture. J. Jpn.

ofRickettsia tsutsugamushi DNAfrom clinical specimens

by nested polymerase chainreaction. J. Jpn.Assoc. Infect.
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Rinsho DNA Shindanho. Kanehara Press, Tbkyo (inKawamura, A., Jr.(1991 ): Use ofmonoclonal antibodies
against Rickettsia tsutsugamushi Kawasaki for sero‑diagnosis by enzyme‑linked immunosorbent assay. J. Clin.
Microbio1., 29, 340‑345.
6. Kaiho, I., Tokieda, M., Yoshida, Y., Furuya, Y., Murata,M., Tanaka, Il. and Kawamura, A., Jr. (1993): Epidemi‑Japanese).

3. Furuya, Y., Yoshida, Y., Katayama, T., Yamamoto, S. and

Kawamura, A., Jr.(1 993): Serotype‑Specific amphficationof Rickettsia tsutsugamushi DNA by nested polymerase

ology of Tsutsugamushi disease and typing of isolated

chain reaction. ∫. Clin. Microbio1., 3 1, 1637‑1640.

Ricketlsia in Chiba Prefecture. J. Jpn.Assoc. Infect. Dis.,

4. Yoshida, Y., Furuya, Y., Katayama, T., Kaiho, I. andYamamoto, S. (1994): Serotype‑Specific amplification

67, 1961201 (in Japanese).

78