Borna Disease in a Dog with Lethal Meningoencephalitis

1
Borna Disease in a Dog with Lethal
Meningoencephalitis
?????? ????? ???????

• Received 4 February 1998/Returned for
  modification 9 April 1998/Accepted 20 April 1998

2
Introduction

• Borna ?????????????????,???????????????????????
  ??????,???????????????????????????????????????????
  ???????????????????????? ?

3
Borna ???????

• Borna ???????????????,????????????????????????????
  ,????????????????????????(????????????)?Borna?????
  ???,?????????????BDV????????????,??BDV????????????
  ,??????????

4
????

• Fig 1
• ?Cresyl echt violet?,??????????????????,??????
  ????(??110?)

5

• Fig 1
• ?Cresyl echt violet????,???????????(??110?)
• ??????????????,???????????? Joest-Degen??????????
  (??510?)

6

• ??????????(Vorarlberg)???????????,????????????????
  ????????????????????????????????
  ???????????????????????????,??????????,??????????
  ?????????????????????????????????????,????????????
  ??

7
?????

• ???????(T7?SP6)clone?????????????(pGEM 3z
  )?,???RNA???????,?DNA???,??????NTP???,????????????
  ?,?????????????????????(T7?SP6)?????,????RNA????SP
  6?????????????,?????????,?4?dNTP???????RNA????????
  ?,????RNA???????????dNTP,???RNA?????

8
(No Transcript)
9
in situ hybridization(????)

• ???????????,??digoxigenin labelle
  RNA??ISH??????????BDV ORF1(open reading frame
  1) RNA,???RNA?digoxigenin labelle,??RNA?probe?????
  ???????????RNA

10
????

1. ?0.1 N HCl for 10 min??
2. ?0.1 (wt/vol) pepsin(????) in 0.1 HCl at 37C
   for 20 min
3. hybridization with partially hydrolyzed(??)
   probe at 50C overnight
4. ? 13 SSC for 15 min ?? (2?)?0.13 SSC for 20 min
   50C (2?)
5. ?????????antidigoxigenin(????)?hybrid?????IHC

11
??????(Immunochistochemistry,IHC)

• ???????????????,???????????????
  (??????????????) ????????????(??????),????????????
  ???,?????????

12
????????????????

• ??????????????????????,??????,????,??????,???????
  ????
• ???????????????????????????????,?????????????????
  ??????????????????????????????????,??????????

13
???????(IHC)????? (ISH)????

• ???????(IHC)????? (ISH)????, (IHC)????????,?4-6???
  ???,?ISH?????????(ISH)????????,?(IHC)?????????????
  ??????,(ISH)????????DNA,?(IHC)?????????????
  (ISH)??????????????????DNA?RNA????????????,???????
  ?????????????,???????????DNA??,?ISH?????????

14
RT-PCR

• RT-PCR????Reverse Transcriptase PCR
  (????PCR),?????????????????RNA?????????????cDNA,??
  ?PCR????????????????????????,????PCR????????Ethidi
  um bromide(????)???????,?????DNA??,?UV????????????
  ?,??????????????????PCR??????(212bp)?

15
???????????H????????????
Astrocytes (????)
16
Neurites(????)
???????????H????????????
17
???????????H????????????
white matter(??)
18
???????????H????????????
????(BO18)?????BDV (p38/40)
19
??ISH?RNA probe???????????RNA, ???????????????(??
570?)
20
??ISH?RNA probe???????????RNA??????????
21
AF054275(??BDV,212bp)

• CCCAGCCTTGTGTTTCTATGTTTGCTAATCCCAGGACTGCACGCTGCATT
  TGTTCACGGAGGGGTGCCTCGTGAATCTTACCTGTCGACGCCTGTTACGC
  GTGGGGAACAGACTGTCGTTAAGACTGCGAAGTTTTACGGGGAAAAGACA
  ACACAGCGTGATCTCACCGAGCTGGAGATCTCTTCTATATTCAGCCATTG
  TTGCTCATTAC

22
AF054276(??BDV,212bp)

• CCCAGCCTTGTGTTTCTATGTTTGCTAATCCCAGGACTGCACGCTGCATT
  TGTTCACGGAGGGGTGCCGCGTGAATCTTACCTGTCGACGCCTGTTACGC
  GTGGGGAACAGACTGTCGTTAAGACTGCGAAGTTTTACGGGGAAAAGACA
  ACACAACGTGATCTCACCGAGCTGGAGATCTCTTCTATATTCAGCCATTG
  TTGCTCATTAC

23
??

• BDV????????????????
• ??immunohistochemistry(???????)?RT-PCR??in situ
  hybridization(????)???????????BDV????BDV???RNA?