DNA containing plant viruses - Cauliflower mosaic virus & Geminivirus

1. FUNDAMENTALS OF MICROBIOLOGY
SEMINAR ON : DNA CONTAINING PLANT
VIRUSES
PRESENTED BY : SUMESH. M (21MBTB24)
1 M.Sc. MICROBIOLOGY
DEPARTMENT OF MICROBIAL
BIOTECHNOLOGY
BHARATHIAR UNIVERSITY

2. CONTENTS
● INTRODUCTION TO DNA CONTAINING PLANT VIRUSES
● CAULIFLOWER MOSAIC VIRUS
● GEMINI VIRUS
● SUMMARY AND CONCLUSION
● REFERENCES

3. INTRODUCTION
● A virus is a small parasite that cannot reproduce by itself.
● Most viruses have either RNA or DNA as their genetic material. The nucleic acid may be
single- or double-stranded. The entire infectious virus particle, called a virion, consists of the
nucleic acid and an outer shell of protein called capsid.
● A DNA virus is a virus with DNA as its genetic material and replicates using a
DNA-dependent DNA polymerase. DNA viruses belong to either Group I (double-stranded
DNA; dsDNA) or Group II (single-stranded DNA; ssDNA) of the Baltimore classification
system for viruses.
● DNA viruses are relatively rare in plants, compared to their RNA counterparts.

4. ● Like most viruses, the genomes of most single stranded DNA viruses are small, encoding
only a few proteins, and are therefore dependent on host cell factors for replication.
● Double stranded DNA viruses only infect lower species of plants, such as algae. These
viruses are huge dsDNA viruses with genomes ranging from 160 to 560 kb with up to 600
protein-encoding genes, making them distinctly different from viruses infecting higher
plants.
● Plant viruses are generally spread through vectors, such as insects, but can also be passed
from generation to generation.

5. CAULIFLOWER MOSAIC VIRUS
(CaMV)

6. INTRODUCTION
● Cauliflower mosaic virus (CaMV) is a type of plant virus of the caulimoviruses which falls
under the family Caulimoviridae. According to Baltimore’s Classification of viruses CaMV
is classified in the Group VII dsDNA-RT viruses. The CaMV are the only plant viruses,
which have a dsDNA genome.

7. MORPHOLOGY
● The CaMV consists of open circular dsDNA as genetic material with single strand
discontinuity like hepadna Virus.
● CaMV shows icosahedral symmetry which means almost spherical shaped particle with a
diameter of 50 nm. It is a non enveloped virus.
● It consists of more than one protein shell. The protein subunits surround a solvent-filled
central cavity. Each virion consists of a circular dsDNA molecule of about 8.0 kb which is
interrupted by site-specific discontinuities resulting from its replication by reverse
transcription.
● The nucleic acid is present inside the capsid.

8. GENOME
● The genome of CaMV is a mono-partite, The CaMV genome consists of approximately
8,000 base-pairs of circular, double - stranded DNA with one discontinuity in one strand,
and one or more discontinuity in the other strand
● There are six major coding regions (I, III, IV, V, VI, VI11) and 2 minor coding regions (II.
VII) in the genome. The minor coding regions act as the store house of the non- essential
genes. The mechanism of expression of these proteins is very special.
The 7 or 8 Open Reading Frames are :-
➢ ORF I – P1: movement protein
➢ ORF II – P2: aphid/insect transmission factor

9. ➢ ORF III – P3: virion-associated protein : structural protein,
DNA-binding capabilities
➢ ORF IV – P4: capsid protein
➢ ORF V – P5: pro-pol : protease, bifunctional reverse transcriptase and RNaseH
➢ ORF VI – P6: transactivator/viroplasmin : inclusion body Formation/trafficking; possibly
other functions
➢ ORF VII/VIII – unknown

10. TRANSMISSION
● CaMV is transmitted in a non-circulatory manner by aphid species such as Myzus persicae.
● CaMV is transmitted between host plants by more than 27 aphid species in a non-persistent
and non-circulative manner, meaning that after an aphid acquires the virus from an infected
plant, the virus does not circulate or replicate within the insect. The virus is retained for a
short period (a few hours) in the aphid stylets (mouthparts), where CaMV receptor
candidates have recently been identified. The virus can then be released to initiate a new
infection during aphid feeding on healthy plants.
● Once CaMV is introduced within a plant host cell, virions migrate to the nuclear envelope of
the plant cell for replication. There are no known cases of CaMV transmission via seeds.

11. ● CaMV can also be transmitted mechanically by sap inoculation using contaminated hands
and pruning tools It has been shown that CaMV can stay for hours on surfaces such as doors,
phones, and gloves, and can be exchanged by hand-shaking, so disinfecting tools,
equipment, and anything that contacts plants will reduce the chance of infection.

12. REPLICATION
CaMV replicates by reverse transcription: the steps involved in the replication of CaMV are
the following.
1. Viral particles enter a plant cell and are uncoated. Which means viruses are made of genetic
material and a protein coat called capsid. The function of the capsid is to protect the genetic
material of the virus. The breaking of capsid and releasing of the genetic material is called as
uncoating. At this stage the viral DNA consists of three fragments, one on the – strand (α)
and two on the + strand (β and γ) which are imperfectly assembled into a circular genome
with three gaps or discontinuities (D1, D2, and D3).
2. The viral DNA enters the nucleus where the replication of genetic material takes place. .

13. 3. The host DNA-dependent RNA polymerase transcribes from the 35S promoter all the way
around the viral genome, surpassing the 35S promoter. (This creates two copies of the 35S
promoter in the resulting RNA.) Transcription also initiates at the 19S promoter
4. The viral RNAs pass into the host cytoplasm where the reverse transcription takes place.
5. The 3′ end of a tRNA anneals to a site corresponding to discontinuity 1 (D1) near the 5′
end of the 35S RNA.
6. The tRNA primes synthesis, by the viral reverse transcriptase (encoded by ORF V), of a
new α strand.
7. RNase H removes the RNA from the DNA–RNA duplex, leaving the component to the
terminal repeat sequence.

14. 8. This anneals to the 3′ end of the 35S RNA, and DNA synthesis continuous
9. Synthesis of the α strand completes. RNase H activity exposes purine-rich regions at
the position of discontinuity 3 (D3), which primes the synthesis of the γ DNA strand.
10. RNase H activity exposes purine-rich regions at the position of discontinuity 2 (D2),
which primes the synthesis of the β DNA strand. When the new γ strand of DNA
reaches the 5′ end of the new α strand it switches to the 5′ end of the new α strand,
recreating discontinuity 1 (D1). When the new γ strand of DNA reaches the 5′ end of
the new β strand, it displaces the primer and some of the newly synthesized β strand,
resulting in the recreation of discontinuity 2 (D2). When the new β strand of DNA
reaches the 5′ end of the new γ strand, it displaces the primer and some of the newly
synthesized γ strand, resulting in the recreation of discontinuity 3 (D3).

15. 11. At this point the new viral genome can either be packaged into capsids and released from the
cell or they can be transported by movement proteins into an adjacent, uninfected cell. The newly
produced viruses are passed to uninfected cells via a small hole between two plant cells called
plasmodesmata.

16. PATHOGENESIS & SYMPTOMS
● It mostly infects members of the Brassicaceae family, including radish, mustard,
cauliflower, broccoli, turnip, canola, and cabbage. Some CaMV strains (D4 and W260) are
also able to infect Solanaceae species, such as devil’s trumpets (genus Datura) and tobacco
plants (genus Nicotiana). CaMV genetic variants have been described in different host
species with different symptoms, virulence, and transmission rates.

17. Symptoms include :
➢ Mosaic
➢ Necrotic lesions
➢ Chlorotic mottling
➢ Vein clearing
➢ Vein banding
➢ Chlorotic spotting
➢ Leaf stunting
➢ Stunted growth, and deformation of the overall plant structure.
● Symptoms may vary depending on the viral strain, host ecotype, and environmental
conditions.

18. PREVENTION AND TREATMENT
● There is no cure for cauliflower mosaic virus. Once a plant is infected, then it is too late.
Prevention is the only way to prevent agricultural or gardening losses. Since cauliflower
mosaic virus is usually spread by aphids, so any sort of pest control can be beneficial in
reducing infection.
● CaMV can also be transmitted mechanically by sap inoculation using contaminated hands
and pruning tools, so disinfecting tools, equipment, and anything that contacts plants will
reduce the chance of infection.

19. GEMINI VIRUS

20. INTRODUCTION
● Gemini virus is a DNA virus, known to infect a wide range of economically important and
agriculture crops of monocotyledonous and dicotyledonous plants. Several diseases such as
maize streak virus disease and curly top virus disease are caused by Gemini virus.
● Geminiviridae is a family of plant viruses which has a circular genome of single-stranded
(ss) DNA. There are 520 species in this family,
● According to the Baltimore classification they are considered class II viruses. It is the largest
known family of single stranded DNA viruses

21. MORPHOLOGY
● Geminivirus have a circular single stranded DNA. Their genome is either segmented or non
segmented. The non segmented genome is 2500-3000 nucleotides long and the segmented
genome is 4800-5600 nucleotides long. This genome encodes for both structural and non
structural proteins. In Gemini virus, both segments must be transmitted to the host for a full
systemic infection to occur
● Geminiviruses are non enveloped and Icosahedral virions that consist of a capsid. The capsid
is elongated, germinate, or twinned, and has 22 capsomers. The capsid is 30nm long and has
a diameter of 18-20 nm.

22. GENOME
● Geminivirus have a Single stranded closed circular DNA. Many species of Geminiviridae have a bipartite
genome, which means, the genome is segmented into two segments (ie : DNA A and DNA B) that are
packaged into separate particles. Both these segments are generally required for successful symptomatic
infection in a host cell.

23. ● The DNAA segment encodes five to six proteins, that are :-
1) V1 (R1)—positive sense : Coat protein
2) V2—positive sense : Movement protein (precoat ORF)
3) C1 (L1)—negative sense: Replication initiation protein (Rep)
4) C2: (L2)—negative sense : Transcription activator protein (TrAP)
5) C3: (L3)—negative sense : Replication enhancer
6) C4:—negative sense : May determine symptom expression
● The DNA B segment encodes two different movement proteins. These proteins have
functions in intra- and intercellular movement in host plants. The Two movement proteins
are :-
1) V1 (R1)—positive sense : Nuclear shuttle protein
2) C1 (L1)—negative sense : Movement protein

24. TRANSMISSION
● In nature, Geminiviruses are transmitted by phloem-feeding insects, including various
species of leafhoppers, a treehopper and whiteflies of the species Bemisia tabaci.
● Members of the genus Begomovirus are transmitted by whiteflies
● Those of the genera Mastrevirus, Curtovirus, Becurtovirus, Mulcrilevirus and
Turncurtovirus are transmitted by leafhoppers
● Those of the genus Capulavirus are transmitted by aphids
● Members of species in the genera Topocuvirus and Grablovirus are transmitted by
treehoppers.

25. REPLICATION
● Geminivirus replicates by rolling circle mechanism: the steps involved in the replication of
Geminivirus are the following
➢ Virus penetrates into the host cell.
➢ After removal of capsid protein, The viral ssDNA genome penetrates into the nucleus.
➢ The ssDNA is converted into dsDNA with the participation of cellular factors.
➢ Bidirectional dsDNA transcription from the IR promoter produces viral mRNAs and
translation of viral proteins.
➢ Replication is initiated by cleavage of the(+)strand by REP, and occurs by rolling circle
producing ssDNA genomes. The virus has a circular, single stranded, DNA that replicates in
host plant cells.

26. ➢ The entire process is initiated by the geminiviral replication initiator protein, Rep, which is
also responsible for altering the host environment to act as part of the replication machinery.
During the rolling circle replication, the ssDNA of geminivirus is converted to dsDNA and
Rep is then attached to the dsDNA at the origin of replication.
➢ After Rep, along with other replication proteins, binds to the dsDNA it forms a stem loop
where the DNA is then cleaved at the nanomer sequence causing a displacement of the
strand. This displacement allows the replication fork to progress in the 3’ to 5’ direction
which ultimately yields a new ssDNA strand and a concatemeric DNA strand.
➢ These newly synthesized ssDNA can either be converted to dsDNA and serve as a template
for transcription/replication OR can be encapsidated by capsid protein and form virions
released by cell lysis. OR can be transported outside the nucleus, to a neighboring cell
through plasmodesmata (cell-cell movement) with the help of viral movement proteins.

27. ROLLING CIRCLE MECHANISM

29. PATHOGENESIS AND SYMPTOMS
● Viruses belonging to the family Geminiviridae are plant viruses that are obligate intracellular
parasites. The species of geminiviridae family is known to infect a wide range of plants and
crops, including both monocotyledons and dicotyledonous plants. Include spinach, maize,
tomato, pepper, okra, turnip etc.

30. General symptoms of diseases caused by Geminivirus are
➢ Stunting
➢ Curling
➢ Twisting of the leaves
➢ Short internodes and stunted appearance
➢ No apical growth
➢ Yellowing of the infected plants
➢ Down-curled tips and margins of the youngest leaves
➢ Chlorosis
➢ Yellowing of veins
➢ Yellow mosaic patterns, and
➢ Dwarfing of leaves.

31. PREVENTION
● The species of geminivirus are mainly transmitted by insect vectors. Mainly including
Leafhoppers, treehoppers, whiteflies, and aphids. Geminiviruses are not transmitted through
seeds, whereas many are graft-transmissible and some are mechanically (sap) transmissible.
● Applying insecticides in plants will prevent these leafhopper, treehopper, whiteflies like
insects so that it will reduce the chance of infection. Geminivirus can also be transmitted
mechanically by sap inoculation using contaminated hands and pruning tools, so disinfecting
tools, equipment, and anything that contacts plants will reduce the chance of infection.

32. TREATMENT
● There is no cure for gemini virus. Once a plant is infected, then it is too late. Prevention is
the only way to prevent agricultural or gardening losses. Gemini virus is usually spread by
Leafhoppers, treehoppers, whiteflies and aphids, so any sort of pest control can be beneficial
in reducing infection.

33. CONCLUSION
In this seminar we discussed two DNA viruses, ie, Cauliflower mosaic virus and Gemini
virus. Cauliflower Mosaic virus is a Double-Stranded DNA virus and Gemini virus is a
Single-Stranded DNA virus. Both cause very harmful effects on various crops and plants. These
viruses are generally spread through vectors, such as insects and can also be transmitted by
mechanical vectors. These viruses can be passed from generation to generation.

34. REFERENCES
● Aurélie Bak and Joanne B. Emerson Cauliflower mosaic virus (CaMV) Biology,
Management, and Relevance to GM Plant Detection for Sustainable Organic Agriculture
● Bio.libretexts.org/Bookshelves/Microbiology/ plant_DNA_viruses
● Biologydiscussion.com/plants/list-of-viruses-found-in-plants-microbiology
● Bio.libretexts.org/plant dna viruses
● Cauliflower mosaic virus/wikipedia
● Covey et al., 1990; Al-Kaff and Covey, 1994.
● Geminiviridae/wikipedia
● Gupta S. K et al 2017 Diseases of vegetable ornamental and spice crops.

35. ● Bioibretexts.org/General Microbiology (Boundless)
● Lodish H, Berk A, Zipursky SL, et al. Molecular cell biology 2000
● Open reading frame/wikipedia
● Haas et al., 2002
● Talk.ictvonline.org/ictv-reports/ictv_online_report/ssdna-viruses/w/geminiviridae
● T.Sathya/plant_viruses/slideshare
● Vincent N. Fondong/protein structure and function.2013
● Viralzone/geminiviridae
● Yasaka et al., 2014

36. THANK YOU