Jpn. J. Infect. Dis., 54, 2001

Laboratory and Epidemiology Communications

Genetic Differentiation of a Measles Virus Isolated froma Case Occurrlng 15 Days after Measles Vaccination
Shinichi Takao*, Takashi Ishidal , Yukie Shimazu, Shinji Fukuda,

Masahiro Noda and Kazuo Miyazaki
Division ofMicrobiology ll ,Hiroshima Prefectural Institute of Health and Environment,
Minami‑machi 1‑6‑29, Minami‑ku, Hiroshima 734‑0007 andJDepartment of Pediatrics, Fukuyama National Hospital,
Okinogami‑cho 41411 7, Fukuyama, Hiroshima 720‑8520
Communicated by Hiroo lnouye(Accepted June 27, 2001)

Measles virus (MV) is an important human pathogen･ Since

DdeI AvaII HaeIII

1 954, when Enders and Peebles succeeded in isolating MV;

IMmany live measles vaccines have been developed, and areNCIt

ClF

I]

CFCF

widely used today (1)･ Liv? measles.Vaccines are highly

immunogenetic and effective ln PreVentlng a natural infection
ofwild‑type MV As some recipients develop vaccine‑induced
side‑reactions such as fever and rash (2‑4), it is becoming

important to differentiate vaccine‑derived viruses from wild‑

type viruses. We report here the results of genetic differentia‑
tion of an MV isolated from a case occurrlng 15 days after
live measles vaccination.
In June 18, 2000, a 1‑year lトmonth‑old girl with a highfTever (39.OoC), cough and rash, was admitted to our hospital･ A
血agnosis ofmeasles was made based on her c血ical symptoms

and K.oplik's spots on her buccal mucosa. For virologlCal
examinations, a throat swab collectedfrom the patient on JuneFig. I. RFLP analysis of the CAM strain and the FO0‑3704l strain･The 635‑ntd long hemagglutinin (H) encoding region (ntd 8067‑870 1 )22 wasinoculated into B95a cells, which is a highly susceptible

was RT‑PCR‑amplified, and the products were then digestedwith

cell line to MV (5). Characteristic cellfusion with multinucle‑
ated giant cells appeared. The isolate was identified as an MV

DdeI, AvaII, orHaeIII. M: 100 bp ladder, NC: non‑cut (635 bp), C:
CAM strain, F: FO0‑3704l straill.

by an indirect immunofluorescent test and micro‑neutralizing

test uslng specific antisera. lt was unknown whether the patient
had been in contact with measles patients in the previous 2‑3

strain were different from those of the CAM strain; the productweeks, but she had received a measles vaccine (CAM strain,of the symptoms. As side‑reactions of the vaccination arewas cleaved into three fragments by Ddel and by Avail. Theseresults suggested that the FO0‑37041 strain was not aBiken, Osaka) on June 7, 2000, about 10 days befわre the onset

frequently observed around 2 weeks a洗er vaccinations (2‑4),

vaccine‑derived strain. To confirm this speculation, We

the origin of the pre.sent isolate (FOOl37041 strain) wasdetemined the nucleotide sequences of the PCR product ofdetermined by a restrlCtion fragment length polymorphism

FO0‑37041 strain by uslng a direct sequenclng method, and(RFLP) assay (6). RNA was extracted from a culturefluid ofcompared the nucleotide sequence with those of the CAM

FO0‑37041 ‑infected B95a cells or from commercial measles

strain, which was obtained from the GenBank database (acces‑CAM vaccin.e (Biken)･ The hemagglutinin (H) protein
encoding reglOn, れtd 8067‑8701, 0f the MV genome wassion No.UO3649). We observed 24 nucleotide differences
between the CAM strain and the FO0‑37041 strain, and 1 1 ofamplified by an RT‑PCR method by using the MH2 (5'‑them were associated with amino acid changes (Table 1 ). The

GGGCTCCGGTGTTCCATATG‑3′) and MH6 (5′‑nucleotide substitution from G to A at 85 16 Observed in theCTTGAATCTGGTATCCACTCCAATl3') primer pair (7).
For RFLP analysis, 5 lil of the PCR product was digested byeither DdeI, Avail, or HaeIII, and the fragments were detectedby electrophoresis in 3% agarose gels with ethidium bromidestainlng. As shown in Figure 1 , the PCR product derived fromisolate F00‑3704 1 was associated with an amino acid changechange is related to the slower mobility of the H protein in

the CAM strain was cleaved into four fragments by DdeI,and two fragments by Avail or HaeIII. In contrast, the diges‑

type MV (4, 7‑10)･ Our analysis indicated that the Gas? Wチs ation patterns of the PCR product derived from FO0‑37041

was compatible with the vaccine‑associated case.

from Asp toAsn creating a new glycosylation site (8). Thisgel electrophoresis and the loss of hemadsorptlOn aCtivlty Of

an MV, both of which are characteristics of a recent wild‑

wild‑type MV infection though the timlng Ofthe vacclnatlOn*Corresponding author: Fax: +81‑82‑254‑1908, E‑mail:takao@urban.nejp
119

Jpn. J. Infect. Dis., 54, 2001
Table 1. Comparison ofnucleotide and predicted amino acid substitu‑

2. Yamaguchi, S., Nakayama, T., Mori, T., Sasaki, K.,
Makino, S. and Sonoda, S. (1994): Genetic differentia‑

tion of HA gene (nucleotide position: 808718676) betweenthe CAM
strain and the FO0‑3704 1 straintion ofAIK‑C measles vaccine strain from wild strains

Diffcrencc ofnuclcotidc (Amino acid)Nuclcotide (Amino acid)in measles outbreak in a nursery. Clin. Ⅵro1., 22, 271‑

CAM FO0‑3704 1

275 (in Japanese).

C (Lcu)T (Phc)3. Ihara, T., Kitamura, K., Kamlya, H., Ochiai, H. and lto,

8105

C (Pro)T (Scr)M･ (1994): Virological studies and clinical featureP in8120

C (LCu)T (Phc)children with fever within 2 weeks a洗er measles vacclna‑

8124

G (Scr)A (As°)

tion. Clin. Viro1., 22, 276‑280 (in Japanese).

8131

T (Cys)C (*)2

4. Kobune, F., Funatu, M., Takahashi, H., Fukushima, M.,8096】( ー ー (ー ー( ー( ー( ー2 2 2 2 つJ つJ つJt J つJ つJt J 4 4 47 0 0 ノ 9 n7 0 ‑ つJI 4 7 0 8 0 1 2∠U 07 4 5 7 1 エU 07 2 8 0ノ 7 ‑ 3 7 0 2 4 人V つJ 9ー ヽノ ′ヽノ ー ヽノ ー ) ー ヽノ ー 1 ー 1 ー8143

G (Lcu)A(*)

Kawamoto, A., Iizuka, S., Sakata, H., Yamazaki, S., Arita,

8156

C (Lcu)T (Phc)M., Wenbo, X. and Zhang, LB. (1995): Characteriza‑

8167

T(Lcu)

C(*)

tion of measles viruses isolated after measles vaccina‑

8174

G (Gly)A (Arg)

8206

A(Gly)G(*)

8282

T (Scr)C (Pro)8285

T(Lcu)C(*)

83日

C (Asp)T (*)

6. Mori, T. (1994): A simple method for genetic differentia‑

8383

A(九rg)

G (*)

tion of the AIKIC vaccine strain from wild strains of8419

A(Gln)

G (*)

8429

A (Lys)C (Gln)tion. Vaccine, 13, 370‑372.

5. Kobune, F., Sakata, H. and Sugiura, A. (1990): Marmoset
lymphoblastoid cells as a sensitive host fわr isolation ofmeasles virus. ∫. Viro1., 64, 700‑705.

measles virus. BiologlCals, 22, 179‑185.

7. Nakayama, T., Mori, T., Yamaguchi, S., Sonoda, S.,

8434

T (Gly)C (*)

Asamura, S., Yamashita, R., Takeuchi, Y. and Urano, T.

8474

T(Leu)C (*)(1995): Detection ofmeasles virus gepome directlyfrom

8482

T (Asp)C (*)

clinical samples by reverse transcrlptaSe‑pOlymerase

G (Asp)A (Asn)chain reaction and genetic variability. Ⅵms Res., 35, 1‑T(Leu)C (*)

G (Ser)A (*)

8 0 8 (XU O 5 5 5 ′lU ′0 1 tJ 5 tJ 3

∠U Oノ 7 4 5(45 5)

16.

8. Rota, J. S., Hummel, K. B., Rota, P. A. and Bellini, W. J.
(1 992): Genetic variability of the glycoprotein genes ofcurrent wild‑type measles isolates. Virology, I 88, 1351

TA(Asn) ≡(Thr)142.

1 The number of the nucl￠otide is based onthe genomic location of the

9. Saito, H., Sato, H., Abe, M., Harata, S., Amano, K., Suto,
T and Morita, M. ( 1992): Isolation and characterization

Edmonston strain.

2Asterisks represent an identical amino acidwiththc CAM strain.

or the measles vims strain with low hemagglutination
activity. Intervirology, 33, 57‑60.

10. Sakata, H., Kobune, F., Sato, T. A., Tanabayashi, K.,
Yamada, A. and Sugiura, A. (1993): Variation in field

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120