unraveling virus complexes in plants/ ciat apr 2015
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Wilmer J. Cuellar
UNRAVELING VIRUS COMPLEXES IN PLANTS
2015Palmira, Colombia
E-mail w.cuellar@cigar.org
Viruses
A virus is basically a protein shell with a few genes inside.
They occur as part of fast evolving and complex communities and they are ecologically important components of the environment.
It is not longer practical for one laboratory to focus on single strains grown in pure culture.
They are historically associated with disease.
Mosaics Yellowing
Ring SpotLeaf Curling
Most of plant pathology in the early 1900s consisted of symptom description
In the beginning we had… ‘mosaics’, ‘yellowings’, ‘ringspots’, etc.
TBT Early plant virology days
‘This masking of symptoms or development of immunity, or whatever it is, seems to hold under greenhouse conditions for practically all the plants tested.’
‘Attempts have been made to produce symptoms on these leaves from other plants, but without success (...). The virus is still present even though the disease symptoms fail to appear.’
Wingard, 1928. J. Agric. Res. 37, 127-153.
But if you leave the plants to grow older…
a.k.a. ‘immunization’, ‘Protective inoculation’, ‘interference’, ‘reciprocal protection’, ‘mutual antagonism’, ‘prophylactic inoculation’, etc. Worked
with similar (related) viruses and under laboratory conditions.
McKinney, 1929. Mosaic diseases in the Canary Islands, west Africa, and Gibraltar. J. Agric. Res. 39, 557-578.Salaman, R.F. 1933. Protective inoculation against a plant virus. Nature. 131, 468-468.
You will get: Cross protection
Things are different in nature: latent and mild viruses
In 1925 Johnson (US) analyzed healthy-looking potatoes and found a ‘mild mottle virus’ in almost all US potato varieties (Potato virus X).
The X virus was also found in Europe and described as ‘simple mosaic’ in some varieties or ‘latent virus’ in others.
In nature:
Johnson J. 1925. Wisc. Res. Bull. 63:12pp. Smith K. 1931. Proc. Royal Soc London. 109, 251-267.Salaman RF. 1937. Proc. Royal Soc London. 229, 137-217.Dykstra T.P. 1938. Phytopathology. 629:40-67.
X + YYX
Mixed infections are common in cassava worldwide
CBSD mosaic CMD mosaic
Caribbean mosaicCommon mosaic Frogskin mosaic
No mosaic
AFRI
CAAM
ERIC
A
Mixed infected Single infected
In the search of ‘Frogskin’ aethiologyIt was first reported in 1971 from southern Colombia (Pineda, et al. 1983)
Cauca1971
• In 1980, flexous viral particles were observed in cassava plants displaying FSD (Pineda B., et al., 1980)
• In parallel, mycoplasma related structures were found. Complex infection virus-mycoplama (Pineda B. & Lozano J.C., 1981)
• The landrace Secundina (MCol 2063) expressed mosaic symptoms where FSD was endemic (CIAT Cassava Program Annual Reports, 1984,1985)
• The presence of a potexvirus (CsVX?) in endemic regions of FSD was reported (Harrison B. D. & Lennon, A. M., 1989)
• Multiple dsRNA (viral?) were associated to FSD (Cuervo M., 1989)
• In 2007, it is reported the a phytoplasma in FSD-affected plants (Alvarez E., et. al. 2009).
• In 2008, a REOvirus is found with cassava plants affected by FSD (Calvert L., et. al.
2008). • FSD plants giving negative results to REOvirus and to Phytoplasma• In 2014 we showed that all affected plants contain a complex infection and
there was not enough evidence to associate previous pathogens separately (Carvajal-Yepes et al., 2014)
Sites of study: North Coast, Valle del Cauca, Eastern Plains
Improving diagnostics to reduce risk -e.g.Cassava
H RS H RS H RS
Symptoms vary depending on the root variety
Venezolana (MCOL-2215)
Mtai8 MCOL-2737
Reproducing motling symptoms
infectedControl
Reproducing leaf deformation symptoms
infectedControl
Updated list of virus species infecting cassava
NAME TAXONOMY DIAGNOSTIC
America
CsCMV Potexvirus ELISA/RT-PCR
CsVX Potexvirus ELISA
CsNAV Potexvirus RT-PCR
CsVMV Cavemovirus PCR
CsFSaV Reoviridae RT-PCR
CsPLV Polerovirus RT-PCR
CsTLV Torradovirus RT-PCR
AsiaCsGMV Nepovirus RT-PCR
ICMV Geminvirius PCR
SLCMV Geminivirus PCR
Africa
ACMV Geminivirus PCR
EACMV Geminivirus PCR
SACMV Geminivirus PCR
ICVM Geminivirus PCR
CBSV Ipomovirus RT-PCR
UCBSV Ipomovirus RT-PCR
Let’s take a look
Cassava Virologists
Phylogenetic tree (nt)
CM4919-1 Sec94-2
Sec92-1 CM523-7 CM6740-7_1
CM4574-1 Sec92-2
CM6740-7 CM6740-7_1
CM6740-7_2 CM6740-7 CM4919-1 CM4574-7
Sec13 Nataima
Lyophilized 19FSD23
Mcol2215 1
6FSD5
SM909-25 Mcol2737-7s
546010-113FSD86
FSD80 OUT
87
6960
88
70
62
7360
0.01
in-vitro
Field collected
Improving diagnostics tools by surveying virus diversity
AAAAAA
AAAAAARNA1
RNA2
Full genome characterization CsTLV
Tools for evaluating virus cleaning protocols
Positive Negative
% o
f inf
ectio
n
N: 246
CsFSaV
CsFSaV: 42%CsPLV: 4.7%CsTLV: 4.7%
5.1 5.2 5.3 5.4
To detect early build up of viruses in the field
Virus free plants became infected in the first cycle45%
60%
Tools to evaluate viruses in roots
Transgenic Line L17Transgenic Line L10
H
VIRUS 2VIRUS 1
NT H
VIRUS 2VIRUS 1
NT
H
LEGEND VIRUS 1 and VIRUS 2: Different virus ifnections.H: Transgenic line grafted with not-infected ‘Secundina’.NT: Non-transgenic line.
Tools to identify their pathogenicity proteins important in disease
AAAAAA
Normal light Ultraviolet light
1 2 3
C 1
2 3
Three examples of reinforcing diagnostics
Cassava common mosaic disease in the South: Re-emerging mixed infections
Photos: Ovidio Antonio UsetINTA, Misiones, Argentina
CsCMD-2014 Reports:Misiones (Arg)Corrientes (Arg)Itapúa (Par)
CsCMV: Early identification, early response
Cordoba-COL
PRO29-COL GU09-COL
Arg24-4
Corrientes-ARG
Mcol1505-1
U23414-Brazil
Mcol22-1
Arg126-6
Arg120-3
Arg128-1
Arg104-3
Arg22-1
Bra456-4
Par92-1
Arg25-1
Arg28-4
Arg97-4
JF913280-Parana
Arg127-1
Arg113-6
Arg121
Arg34-4
PAr94-2
Outgroup
99
94
95
100
82
61100
71
77
70
98
100
91
0.05
2013-2014
p22 can suppress the defense of the plant
p22 enhances the accumulation of the virus
p22 by itself Induce severe symptoms in other plants
(-) (+) (-) (-)
P-Pro MTR HEL RdRp p26 p7 p22
Uganda
Sweetpotato: Sometimes viruses change strategies
P-Pro MTR HEL RdRp p26 p7Rest of the world
East Africa strain
West Africa strain
Citrus: Re-evaluation of cleaning systems
Main citrus regions in Colombia (2014) 17 ICA-registered nurseries 5 comercial greenhouses 80 samples
99% infected by known pathogens (Mixed infections) CEVd 46% HSVd 81% CTV 60%
Main drivers of disease
Anderson PK., et al. 2004. Emerging infectious diseases of plants (...) TRENDS in Ecol and Evol. 19:535-544.
Early identification should include the survey for potential pathogens before they jump into crops.
Take home messages and Future directions
A high diversity of hosts implies a even wider biodiversity of potential pathogens. We need to understand the biological significance of that diversity and their interactions.
It is not longer practical to focus on diagnosis of single isolated pathogens grown in pure culture, especially with diseases affecting RTB crops.
An “early identification, early response” strategy. Emphasis basic field research and biological tests. Sequences are needed but not enough. Including the diversity of potential pathogens in wild plants.
We suggest the establishment of a Surveillance Network which should incorporate information on disease-conductive environments and considering historical information on previous epidemics.
Carvajal-Yepes M, Olaya C, Lozano I, Cuervo M, Castaño M, Cuellar WJ. (2014) Unravelling complex virus infections in cassava. Virus Research 186, 76-86.
Legg J, Kumar L, Makeshkumar T, Ferguson M, Kanju E, Ntawuruhunga P, Cuellar WJ. (2015) Cassava virus diseases: Biology, Epidemiology and Management. Advances in Virus Research. 91, 85-142.
DiFeo L, Zanini A, Rodriguez P, Cuervo M, Carvajal-Yepes M, Cuellar WJ. (2015) First report of Cassava common mosaic virus and Cassava frogskin associated virus infecting cassava in Argentina. Plant Disease. 9, 733.
Van der Vlugt R, Verbeek M, Dullemans AM, Wintermantel WM, Cuellar WJ, Fox A, Thompson JR. (2015) Torradoviruses. Annual Review of Phytopathology. 53, 23.1-23.28.
Cassava Virology - Publications
Cassava virology – CIAT
Ivan Lozano Monica Carvajal Jenyfer Jimenez
MSc Thesis completed:Cristian Olaya, 2014. MSc-Thesis. UNAL-Palmira, Colombia.Ana M. Leiva, 2015. MSc-Thesis. UNAL-Palmira, Colombia.
THANK YOU FOR YOUR ATTENTION
Alejandro Quintero Bertha Garcia
We inherited a strong Research Laboratory, founded by Dr. Francisco Morales.
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