Use of next‐generation sequencing for the identification and characterization of M aize chlorotic mottle virus and S ugarcane mosaic virus causing maize lethal necrosis in K enya
The diagnosis of novel unidentified viral plant diseases can be problematic, as the conventional methods such as real‐time PCR or ELISA may be too specific to a particular species or even strain of a virus, whilst alternatives such as electron microscopy ( EM ) or sap inoculation of indicator specie...
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Published in | Plant pathology Vol. 62; no. 4; pp. 741 - 749 |
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Main Authors | , , , , , , , , , , , , , , , , , , , |
Format | Journal Article |
Language | English |
Published |
01.08.2013
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Online Access | Get full text |
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Abstract | The diagnosis of novel unidentified viral plant diseases can be problematic, as the conventional methods such as real‐time
PCR
or
ELISA
may be too specific to a particular species or even strain of a virus, whilst alternatives such as electron microscopy (
EM
) or sap inoculation of indicator species do not usually give species level diagnosis. Next‐generation sequencing (
NGS
) offers an alternative solution where sequence is generated in a non‐specific fashion and identification is based on similarity searching against
G
en
B
ank. The conventional and
NGS
techniques were applied to a damaging and apparently new disease of maize, which was first identified in
K
enya in 2011.
ELISA
and
TEM
provided negative results, whilst inoculation of other cereal species identified the presence of an unidentified sap transmissible virus.
RNA
was purified from material showing symptoms and sequenced using a
R
oche 454
GS
‐
FLX
+. Database searching of the resulting sequence identified the presence of
M
aize chlorotic mottle virus
and
S
ugarcane mosaic virus
, a combination previously reported to cause maize lethal necrosis disease. Over 90% of both viral genome sequences were obtained, allowing strain characterization and the development of specific real‐time
PCR
assays which were used to confirm the presence of the virus in material with symptoms from six different fields in two different regions of
K
enya. The availability of these assays should aid the assessment of the disease and may be used for routine diagnosis. The work shows that next‐generation sequencing is a valuable investigational technique for rapidly identifying potential disease‐causing agents such as viruses. |
---|---|
AbstractList | The diagnosis of novel unidentified viral plant diseases can be problematic, as the conventional methods such as real‐time
PCR
or
ELISA
may be too specific to a particular species or even strain of a virus, whilst alternatives such as electron microscopy (
EM
) or sap inoculation of indicator species do not usually give species level diagnosis. Next‐generation sequencing (
NGS
) offers an alternative solution where sequence is generated in a non‐specific fashion and identification is based on similarity searching against
G
en
B
ank. The conventional and
NGS
techniques were applied to a damaging and apparently new disease of maize, which was first identified in
K
enya in 2011.
ELISA
and
TEM
provided negative results, whilst inoculation of other cereal species identified the presence of an unidentified sap transmissible virus.
RNA
was purified from material showing symptoms and sequenced using a
R
oche 454
GS
‐
FLX
+. Database searching of the resulting sequence identified the presence of
M
aize chlorotic mottle virus
and
S
ugarcane mosaic virus
, a combination previously reported to cause maize lethal necrosis disease. Over 90% of both viral genome sequences were obtained, allowing strain characterization and the development of specific real‐time
PCR
assays which were used to confirm the presence of the virus in material with symptoms from six different fields in two different regions of
K
enya. The availability of these assays should aid the assessment of the disease and may be used for routine diagnosis. The work shows that next‐generation sequencing is a valuable investigational technique for rapidly identifying potential disease‐causing agents such as viruses. |
Author | Boonham, N. Skelton, A. Wangai, A. Smith, J. Nixon, T. Miano, D. W. Kimani, E. Kinyua, Z. M. Harju, V. Reeder, R. Hany, U. Thwaites, R. Phiri, N. Deb Nath, P. Adams, I. P. Mumford, R. Barnes, A. Glover, R. Souza‐Richards, R. Fox, A. |
Author_xml | – sequence: 1 givenname: I. P. surname: Adams fullname: Adams, I. P. organization: Food and Environment Research Agency Sand Hutton York YO41 1LZ UK – sequence: 2 givenname: D. W. surname: Miano fullname: Miano, D. W. organization: Kenya Agricultural Research Institute Nairobi PO Box 14733–00800 Kenya – sequence: 3 givenname: Z. M. surname: Kinyua fullname: Kinyua, Z. M. organization: Kenya Agricultural Research Institute Nairobi PO Box 14733–00800 Kenya – sequence: 4 givenname: A. surname: Wangai fullname: Wangai, A. organization: Kenya Agricultural Research Institute Nairobi PO Box 14733–00800 Kenya – sequence: 5 givenname: E. surname: Kimani fullname: Kimani, E. organization: Kenya Plant Health Inspectorate Service PO Box 49592‐00100 Nairobi Kenya – sequence: 6 givenname: N. surname: Phiri fullname: Phiri, N. organization: CAB International Africa Regional Centre PO Box 633‐00621 Nairobi Kenya – sequence: 7 givenname: R. surname: Reeder fullname: Reeder, R. organization: CAB International Egham TY20 9TY UK – sequence: 8 givenname: V. surname: Harju fullname: Harju, V. organization: Food and Environment Research Agency Sand Hutton York YO41 1LZ UK – sequence: 9 givenname: R. surname: Glover fullname: Glover, R. organization: Food and Environment Research Agency Sand Hutton York YO41 1LZ UK – sequence: 10 givenname: U. surname: Hany fullname: Hany, U. organization: Food and Environment Research Agency Sand Hutton York YO41 1LZ UK – sequence: 11 givenname: R. surname: Souza‐Richards fullname: Souza‐Richards, R. organization: Food and Environment Research Agency Sand Hutton York YO41 1LZ UK, University of Nottingham Sutton Bonington LE12 5RD UK – sequence: 12 givenname: P. surname: Deb Nath fullname: Deb Nath, P. organization: Food and Environment Research Agency Sand Hutton York YO41 1LZ UK, Assam Agricultural University Jorhat‐785013 Assam India – sequence: 13 givenname: T. surname: Nixon fullname: Nixon, T. organization: Food and Environment Research Agency Sand Hutton York YO41 1LZ UK – sequence: 14 givenname: A. surname: Fox fullname: Fox, A. organization: Food and Environment Research Agency Sand Hutton York YO41 1LZ UK – sequence: 15 givenname: A. surname: Barnes fullname: Barnes, A. organization: Food and Environment Research Agency Sand Hutton York YO41 1LZ UK – sequence: 16 givenname: J. surname: Smith fullname: Smith, J. organization: Food and Environment Research Agency Sand Hutton York YO41 1LZ UK – sequence: 17 givenname: A. surname: Skelton fullname: Skelton, A. organization: Food and Environment Research Agency Sand Hutton York YO41 1LZ UK – sequence: 18 givenname: R. surname: Thwaites fullname: Thwaites, R. organization: Food and Environment Research Agency Sand Hutton York YO41 1LZ UK – sequence: 19 givenname: R. surname: Mumford fullname: Mumford, R. organization: Food and Environment Research Agency Sand Hutton York YO41 1LZ UK – sequence: 20 givenname: N. surname: Boonham fullname: Boonham, N. organization: Food and Environment Research Agency Sand Hutton York YO41 1LZ UK |
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Cites_doi | 10.1007/s007050050362 10.1111/j.1364-3703.2009.00545.x 10.1006/viro.1999.0107 10.1007/s11262-011-0644-2 10.1093/nar/gkn721 10.1094/PD-65-39 10.1093/nar/gkq1079 10.1016/j.jviromet.2010.11.002 10.1111/j.1439-0434.2010.01745.x 10.1186/1471-2105-10-421 10.1094/Phyto-77-162 10.1093/bioinformatics/btp666 10.1016/j.virol.2009.03.024 10.1093/nar/17.8.3163 10.1016/j.virol.2009.02.028 10.1111/j.1365-3059.2012.02622.x 10.1101/gr.5969107 10.1093/molbev/msr121 10.1099/0022-1317-34-3-475 10.1007/s00705-001-0799-6 10.1094/PD-64-944 10.1007/s00705-008-0069-y |
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References | e_1_2_6_10_1 Hill SA (e_1_2_6_11_1) 1984 Morales FJ (e_1_2_6_16_1) 1999; 44 e_1_2_6_19_1 e_1_2_6_13_1 e_1_2_6_14_1 e_1_2_6_12_1 e_1_2_6_17_1 e_1_2_6_18_1 e_1_2_6_15_1 e_1_2_6_21_1 e_1_2_6_20_1 Valverde R (e_1_2_6_24_1) 1990; 74 e_1_2_6_9_1 e_1_2_6_8_1 e_1_2_6_5_1 e_1_2_6_4_1 e_1_2_6_7_1 e_1_2_6_6_1 e_1_2_6_25_1 e_1_2_6_3_1 e_1_2_6_23_1 e_1_2_6_2_1 e_1_2_6_22_1 e_1_2_6_26_1 |
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Title | Use of next‐generation sequencing for the identification and characterization of M aize chlorotic mottle virus and S ugarcane mosaic virus causing maize lethal necrosis in K enya |
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