Jpn. J. Infect. Dis., 52, 1999

Laboratoryand Epidemiology Commumications

Expedences of Microbial Contamination of Animal Colonies Maintainedin the National Institute of Infectious Diseases, Japan (NIID)
Kazuhiro Takimoto* , Yasuko K. Yamada, Yasushi Ami, Yuriko Suzaki,
Mikiko Yabe and Toshihiko AsanODivision of Experimental Animal Research, National Institute oflnjTectious Diseases,
7Toyaf7∽ 1123‑1, Shinjuku‑ku, 7Tokyo 162‑8640

Communicated by Hiroshi Ybshikura(Accepted January 14, 2000)

Viraland bacterialcontamination of laboratory animals main‑tained in the NIID is reported here. BoTdetella bronchiseptica,ll 2 0 8 6 4mouse hepatitis virus (MHV),and the hemagglutinating virusof Japan(HVJ) are the infectious agents;all of them aJ,e known

uJnJaSJOhaq∈nN

to contaminate laboratoryanimals ( 1 ,2).
Rabbits maintained in the Murayama Branch of the NIID
are regularly examined by nasal swabs for pathogens. In

September 1 996, B. bronchiseptica was detected in the nasal
swab of a rabbit during a regular check of microbial infec‑tion of laboratory animals Crable I ). Necessary measures suchas restriction of entry into animal laboratories were taken

promptly to prevent further spread. Examination revealed that<252550100( negativ e)

rabbits purchased from one particular breeder were infectedwith the bacteria. All of thel18 rabbits kept in the same
animal room were sacrificed. All rabbits entenngthe laboratory

200

400

800

1600

≧3200

ELISA liter

Fig･ 1. MHV‑Specific lgG ELISA titers of sera of mice in MHV‑con‑taminated mouse colony･

thereafter were checked for the pathogen during the next onemonth. Wth these measures, we were able to free the rabbitcolony of B. bT10nChiseptlCa.

MHV infection occurred in a mouse breeding colony ln
the Tbyama main campus of the NIID in October 1995. As
several randomly chosen mice were fわund positive for MHV

infection during lmmunOlogical tests performed during theregular microbiologiCalcheck (Table I), entry into the room
was immediately restricted. A total of 63 mice (A/WySnJ and

1

2

3

4

5

6

N

M

12lst PCR

3456

N

2nd PCRI̲5: Feces of mouse contaminated withMHV

BALB/cXid strains), each of which was taken randomly froma cage, was examined by ELISA and immunonuorescent

N : Feces of BALB/c mouse (negative control)

M : MarkerDNA

antibody (IPA) techniques for anti‑MHV antibody. Forthe
test, we used MHV antigen, which was purchased from
DENKA SEIKEN Co. Ltd., Tokyoandthe affinity‑purified

Fig･ 2･ Representative results of Rr/PCR assays･ The six left lanes

show the first PCR andthe sixright lanes showthe second PCR･ 1‑

5: FecalSpeclmenS Of MHV‑infected mice･ N: fecalSpeclmen fromnomal BALB/c mice. M: Molecular weight marker･Fab'portion of goat anti‑mouse IgG (H+L‑chain) (MBLNagoya) was used as a secondary antibody. Sixty mice (95%)

were found to be MHV‑infected (Fig. I). Three months after
the event, detection of the MHV genome from fresh stool
specimenswas attempted by reverse transcription (RT)/nested

PCR (3) by using primers constmCted on the sequence of theJHM strain of MHV MHV was detected in 20 out of 37 micetested (Fig. 2). The sequenceanalysis revealed homology with*Corresponding author: Fax: +81 ‑3‑5285‑ 1267, E‑mail: takimoto

RI or Y strains, enteric MHV strains reported in the USA･However, our isolate was not identical to these strains. There‑@ nih. go.Jp255

Jpn. J. Infect. Dis., 52, 1999
Table l･ Examinations for regularmicrobiologiCalmonitonng of mouse, rabbit and guinea plg COIomies in NIID
Pathogcn

Test

Mouse

Rabbit

Gui

nea

plg

Bacteda & Mycoplasma
Pseudomonas aeruglnOSa

Cultivadon (NAC agar)

SaLmonelkz spp.

Cultivation (DHL agar)

Escherichia colt Ol 15a, C:K(B)

Cultivation (DHL agar)

PasEeureLkz pneumoEropICaCultivadon (Horse blood agar)

CoTynebacterium kuEscheri

Cultivation (FNC agar)

00000000TIZZCr 's organism

Serological examinadon (CF)MycopEasma puLmon is

Cuhivation (PPLO agar)

MycopLasma pulmon is

SerologiCalexaminadon (ELISA)

StaphyLococcus aweus

Cultivation (Mannit‑Salt agar)

PasteureLLa muLtoci血

Cultivadon (Horse blood agar)

BoTdeEeLhz bronchisepEica

Cultivation (DHL agar)

StrepEococcus zooepidemicus

Cultivation (Horse blood agar)

StrepEococcus pneumoniae

Cultivation (Horse blood agar)

0000○000000Viruses

Mouse hepatitis virus (MHV)

Serological examination (ELISA)

Sendaivirus (HVJ)

SerologiCalexaminadon (ELISA)

SendaiviruS (HVJ)

SerologiCalcxamination (CF)

Ectromeria virus

Serological examination (ELISA)○ ○○0 0 00 0

Parasi tes

GiaTldia muTis

Microscopic examination

SpironucLeus muTis (HeJdmita mum‑S)

MicroscoplC examination

SyphacL'a spp.

MicroscoplC examination

CysEicencus fzzscioLaris

VisualMyobia muscuLi

MicroscopIC examination

Radfondia aがnis

MicroscopIC examination

EimeTia spp.

Microscopic examination

CysticeTCuS Pislformis

Vi sual examination

Psoroptes cunicuLi

Mi croscopl C eXami nation
Mi croscopl C eXami nation

El'meria cavise

examinadon

Table 2. Detection of antibody in sera ofmiCc infcctedwithA, B or C strain of virus
No. of positive/No. of animals examined

Virus strain Mouse strainAB

BALB/C

Anti‑A And‑B Anti ‑D Anti ‑SendaiviruS

6/6

BALB/cNT0/5C3HNTBALD/cI)

NT

5/5

NT

NT

5/5

NT

6/6

NT

5/5

0/5

0/50/5l )Not testedfore, our isolate was referred to as the TY strain (unpublishedin rabbits by uslng UV‑inactivated Sendai virus. Sendai virusdata)･ The data indicated the continued persistence of MHV

infection of the mouse colony has not reoccurred.

Thus, these expenences underscorethe importance of regu‑

in the colony.
Contamination of a mouse colony withSendaivirus o∝urred

lar microbiological monitoring (1もble 1 ) of laboratory animals

duringanexpenment on the adaptation of influenza virus toand of prompt reactions to prevent the spread of pathogens･

mice･ The virus was serially passaged in mice by pulmonary

infection･ During the experiments, Control mice suddenly

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256