Detection and Differentiation of Malaysian Newcastle Disease Virus Isolates by RNA-Polymerase Chain Reaction and Cycle Sequencing
This study was undertaken to develop diagnostic tests for Newcastle disease virus (NDV) based on RNApolymerase chain re action (RNA-PCR) and cycle sequencing techniques as a supplement to the presently available tests. Two RNA-PCR cycle sequencing systems each targeted at the haemagglutinin-ne...
Saved in:
| Main Author: | |
|---|---|
| Format: | Thesis |
| Language: | English English |
| Published: |
1995
|
| Subjects: | |
| Online Access: | http://psasir.upm.edu.my/id/eprint/8597/1/FSAS_1995_3_A.pdf |
| Tags: |
Add Tag
No Tags, Be the first to tag this record!
|
| Summary: | This study was undertaken to develop diagnostic
tests for Newcastle disease virus (NDV) based on RNApolymerase
chain re action (RNA-PCR) and cycle sequencing
techniques as a supplement to the presently available
tests.
Two RNA-PCR cycle sequencing systems each targeted
at the haemagglutinin-neuraminidase (HN) and fusion (F)
genes were developed. RNA-PCR amplification of a 398
base pairs (bp) HN gene fragment was performed on
total RNAs extracted from infected allantoic fluid of 9
Malaysian NDV field isolates, vaccine strain V4-UPM and
velogenic strain AF2240. Sequence analysis over 113
bases in the amplified fragment showed variations among
these isolates/strains. However, identical sequences
were obtained from some of the field isolates within a
particular pathotype and these were thought to be of the
same strain of NDV. RNA-PCR amplification of a 242 bp F gene fragment
was performed on total RNAs which were isolated from
several types of tissues (spleen, brain and lung) from
six non-vaccinated chickens which were challenged with
the velogenic strain AF2240. The spleen was found to
have the highest number of samples positive by PCR.
Spleens from uninfected chickens as well as those from
vaccinated and challenged chickens which were
slaughtered were all PCR negatives. The challenged virus
was thought to have been neutralised by the antibodies
produced by the chickens. The identity of the PCR
products amplified from the spleens w'ere confirmed by
cycle sequencing. Similarly, this test was also
performed on RNAs from infected allantoic fluid of 11
different NDV strains/isolates. These samples were also
PCR positives. |
|---|