Tobacco mosaic virus tomato

Contents

Tobacco mosaic virus

Assorted References

  • agent of infection
    • In plant virus

      …of the most well-studied viruses, tobacco mosaic virus (TMV), is spread mechanically by abrasion with infected sap. Symptoms of virus infection include colour changes, dwarfing, and tissue distortion. The appearance of streaks of colour in certain tulips is caused by virus.

  • discovery by Ivanovsky
    • In Dmitry Ivanovsky

      …study of mosaic disease in tobacco, first detailed many of the characteristics of the organisms that came to be known as viruses. Although he is generally credited as the discoverer of viruses, they were also independently discovered and named by the Dutch botanist M.W. Beijerinck only a few years later.

  • place in virology
    • In virology

      …to be a virus) of tobacco mosaic disease could pass through a porcelain filter impermeable to bacteria. Modern virology began when two bacteriologists, Frederick William Twort in 1915 and Félix d’Hérelle in 1917, independently discovered the existence of bacteriophages (viruses that infect bacteria).

    • In virus

      …by an agent, later called tobacco mosaic virus, passing through a minute filter that would not allow the passage of bacteria. This virus and those subsequently isolated would not grow on an artificial medium and were not visible under the light microscope. In independent studies in 1915 by the British…

    • In virus: The protein capsid

      …helical rod-shaped virus is the tobacco mosaic virus, which was crystallized by Wendell Stanley in 1935. The tobacco mosaic virus contains a genome of single-stranded RNA encased by 2,130 molecules of a single protein; there are 161/3 protein molecules for each turn of the RNA helix in the ratio of…

  • tobacco diseases
    • In tobacco: Diseases and pests

      …black root rot, Fusarium wilt, tobacco mosaic virus (TMV), bacterial leaf spot, downy mildew, black shank, broomrape, and witchweed. These may be controlled by sanitation, crop rotation, the use of fungicide and herbicide sprays and fumigants, and breeding of disease-resistant strains. Some resistant

Tobacco Mosaic Virus

6.3.1 Tobacco Mosaic Virus

Tobacco mosaic virus (TMV; Tobamovirus, Virgaviridae) is a rodlike virus with a length of 300 nm and diameter of 18 nm. TMV capsids are composed of 2130 identical protein subunits, which assemble around the viral ssRNA to form a helical structure, with a hollow central cavity of 4 nm diameter. The structural chirality and inherent asymmetry of the virus allowed chemical or genetic modifications spatially at one or the other end of the helical rod. TMV has a well-defined in vitro assembly system controlled by pH, ionic strength, and protein concentration. TMV particles can be produced in large quantities from infected tobacco plants (Young et al., 2008; Ma et al., 2012). TMV is both chemically and physically stable to solvents, pH (3.5–9.0), temperature (up to 90°C), and reducing agents. Due to this flexibility, tubelike TMV capsids were biochemically modified on both interior and exterior surfaces for the synthesis of metal nanoparticles, nanowires, and layer-by-layer assemblies that can be organized into higher order 2-D and 3-D structures through a variety of reactions (Young et al., 2008; Ma et al., 2012; Culver et al., 2015; Narayanan and Han, 2017b).

Blood clearance kinetics and biodistribution studies of radiolabeled TMV in both normal and tumor-bearing mice revealed their rapid clearance and nontoxicity in both models (Wu et al., 2013). TMV rods can also undergo thermal transition to form RNA-free densely packed spherical nanoparticles (SNPs) of size ~ 50 nm (Atabekov et al., 2011). Bruckman et al. (2014a) studied the biodistribution, pharmacokinetics, and blood compatibility of native and PEGylated TMV rods and spheres in mice. Both versions exhibited comparable in vivo profiles with few differences, where rods circulate longer than spheres illustrating the effect of shape on circulation. PEGylation increased circulation times. Spheres are more rapidly cleared from tissues compared with rods with no overt toxicity. The impact of aspect ratio on the biodistribution and tumor homing of PEGylated or receptor-targeted RGD-displaying TMV formulations was investigated, and it was found that the aspect ratio has a profound impact on the behavior of the nanoparticles in vivo and in vitro (Shukla et al., 2015a). In vivo implantation of RGD-TMV hosted bioadhesive peptide (RGD) hydrogels indicated that TMV is less immunogenic, nontoxic, and biodegradable in mice (Luckanagul et al., 2015).

TMV hosts well-defined amino acids (tyrosines and glutamates) at specific locations on its outer surface. Natural and genetically modified TMV particles with nonnative amino acids (cysteines and lysines) were used to attach new functionalities such as fluorophores, PEGs, and other nanoparticles using orthogonal synthetic methods to obtain biosensors, optoelectronic devices, catalysts, and imaging agents (reviewed in Culver et al., 2015; Wen et al., 2015a, 2016; Narayanan and Han, 2017b). The scope for genetic fusions to the termini of TMV capsid proteins is limited, as this can inhibit virus particle formation (Steele et al., 2017). TMV particles have been loaded with paramagnetic gadolinium ions selectively at the interior (iGd-TMV) or exterior (eGd-TMV) surface, and it was found that interior-labeled TMV rods can undergo thermal transition to form 170 nm-sized SNPs. These functionalized TMV rods and spheres showed high relaxivities to act as MRI contrast agents (Bruckman et al., 2013). Coupling of TMV particles to the two-photon dye BF3-NCS has been demonstrated to allow visualization of diseased brain vasculature in mice (Niehl et al., 2015). In a comparative study, fluorescently labeled CPMV and TMV were used to determine the impact of dye density, dye localization, conjugation chemistry, and microenvironment on the optical properties of the virus-based optical probes. The results indicated that both are extraordinarily robust under ultrashort, pulsed laser light conditions with a significant amount of excitation energy, maintaining structural and chemical stability (Wen et al., 2015b,c). In another study, CPMV and TMV were anchored with fibrin-binding peptides (CREKA and GPRPP) and contrast agents for dual-modality magnetic resonance (MR) and optical imaging. The resultant particles are found to be specific to fibrin binding in vitro. Preclinical studies in a carotid artery photochemical injury model of thrombosis confirmed greater attachment of elongated TMV rods to thrombi than CPMV (Wen et al., 2015b,c). Hu et al. (2017) prepared a bimodal contrast agent by loading the internal cavity of TMV nanoparticles with a dysprosium (Dy3 +) complex and NIR fluorescent dye Cy7.5, and the external surface of TMV was conjugated with an Asp-Gly-Glu-Ala (DGEA) peptide via a PEG linker to target integrin α2β1. The resulting Dy-Cy7.5-TMV-DGEA was useful for ultra-high-field magnetic resonance and near-infrared fluorescence imaging of prostate cancer in vivo.

TMV has also been used as a vaccine vehicle to display various heterogeneous antigenic epitopes and immunization of animals with each of these chimeras induced a strong humoral protective immune response against the respective pathogen challenge (reviewed in Hefferon, 2017). A tumor associated carbohydrate antigen (Tn; target for antitumor vaccine development) conjugated to TMV can elicit antigen-specific IgG and IgM responses (Yin et al., 2012). Engineering of TMV to target vascular cell adhesion molecule (VCAM)-1 (which is highly expressed on activated endothelial cells at atherosclerotic plaques) resulted in molecular targeting of atherosclerotic plaques by functionalized TMV particles in an atherosclerotic ApoE(−/−) mouse model (Bruckman et al., 2014b). Cysteine-inserted TMV mutants were cross-linked to methacrylated hyaluronic acid (MeHA) polymers by “click” chemistry and formed hydrogels under physiological condition, which could promote in vitro chondrogenesis of bone marrow mesenchymal stem cells (BMSCs) (Maturavongsadit et al., 2016). TMV was tailored with peroxidase-like inorganic nanoparticles (platinum nanoparticles) and cancer cell target groups (folic acid) to obtain TMV-FA-Pt nanoparticles for cancer cell detection using a cell-based ELISA (CELLISA) (Guo et al., 2018).

Self-assembling nanoscale disks derived from a double arginine mutant of recombinantly expressed TMV (RR-TMV) was functionalized with the dox and further modified with PEG for improved solubility. RR-TMV-DOX-PEG displayed cytotoxic properties like those of dox alone when incubated with U87MG glioblastoma cells (Finbloom et al., 2016). The cationic porphyrin-based photosensitizer was successfully and stably loaded into the interior channel of TMV via electrostatic interactions. The resulting TMV-photosensitizer exhibited improved cell uptake and efficacy when compared with free photosensitizer, making it a promising platform for improved PDT of melanoma (Lee et al., 2016a). Bruckman et al. (2016) demonstrated the loading of dox into rods and spheres by chemical conjugation and encapsulation approaches for targeted delivery to breast cancer. Tumor delivery and efficacy of TMV loaded with phenanthriplatin (a cationic monofunctional DNA-binding platinum (II) anticancer drug) was confirmed by using a triple-negative breast cancer mouse model (Czapar et al., 2016). The TMV-cisplatin conjugate (TMV-cisPt) was synthesized using a charge-driven reaction and it showed superior cytotoxicity and DNA double-strand breakage (DSB) in platinum-sensitive (PS) and platinum-resistant (PR) cancer cells when compared with free cisplatin (Franke et al., 2017). TMV conjugates loaded with streptokinase (STK) via intervening PEG linkers were synthesized, and the resultant TMV-STK formulations showed thrombolytic activity in vitro using static phantom clots (Pitek et al., 2017). TMV was used as a multivalent carrier for the delivery of the antimitotic drug valine-citrulline-monomethyl auristatin E (vcMMAE) targeting non-Hodgkin’s lymphoma (Kernan et al., 2017).

Interaction studies between VNPs derived from TMV and plasma revealed that the VNP protein corona is significantly less abundant compared with the corona of synthetic particles. A library of functionalized TMV rods with PEG and peptide ligands targeting integrins or fibrin(ogen) showed different dispersion properties, cellular interactions, and in vivo fates depending on the properties of the protein corona, influencing target specificity, and nonspecific scavenging by macrophages (Pitek et al., 2016a). Serum albumin (SA) conjugation to TMV-based nanocarriers resulted in a “camouflage” effect more effective than PEG coatings. SA-“camouflaged” TMV particles exhibited decreased antibody recognition and enhanced pharmacokinetics in a BALB/c mouse model (Pitek et al., 2016b).

Tobacco Mosaic Virus Uses In Pharmaceutical Research

Tobacco Mosaic Virus Uses in Pharmaceutical Research

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By Michael Itschner II

Introduction

The Tobacco Mosaic Virus (TMV) has been well researched since it has been defined a virus in 1898 by Martinus Beijerinck. TMV has been used as a model virus for a teaching tool in biology classes and as a scaffold for drug delivery mechanisms. Since there has been much research done on this virus previously, the structure of TMV has been extensively studied and is well understood. The scaffolding of this virus is of particular interest because its protein base serves as a template for precise targeting of surface groups on cells. Plant viruses have drawn more attention since they are unable to infect mammalian hosts and have good blood and tissue compatibility in mammals. TMV is also biodegradable and able to be cleaned out of non-target tissues and organs, making TMV a promising candidate for the targeting and delivery of therapeutics to mammals.

Discovery

The discovery of the Tobacco Mosaic Virus (TMV) and the mosaic disease it caused can be attributed to three individuals, Adolf Mayer, Dmitrii Iwanowski, and Martinus Beijerinck. In 1886 Adolf Mayer (1843-1942) found in his research that extracts from ground up diseased tobacco plants could infect healthy tobacco plants. He followed Koch’s postulates moving forward in his research and was able to produce cultured organisms from the extracts but would not be able to reproduce the disease. Using filter paper to screen out cellular pathogens, Mayer concluded that a bacterial species was responsible. He reasoned that the infectious agent passed through the initial filtration paper, but after multiple filtrations the agent stopped infecting other plants. Replicated research on the filtration of the microbe causing the tobacco mosaic disease by Dmitrii Iwanowski (1864-1920) found contrasting results to Mayer. In 1892 Iwanowski reported that the Chamberland filter candles, which were fine filer that retained bacteria, did not trap the disease-causing microbe, it passed through the filter. This led Iwanowski to believe that a toxin released from a bacterium, which was dissolved in sap, caused the disease or the bacterium was small enough to pass through the filter candles . Martinus Beijerinck (1851-1931) also worked with filtration experiments and also found that the material, absent of microorganisms, that passed through the porcelain filters continued to be infectious to healthy plants. In 1898 Beijerinck called this material a virus and continued to research its transmission and growth. Although these three individuals were not completely correct in their assumptions about the TMV, their discoveries were crucial to its classification in the microbial world.

Structure

(Figure 1) Loading scheme of the Tobacco Mosaic Virus .

The crystal structure of TMV was discovered in 1935 by Wendell Stanley (1904-1971), which led him to win the 1946 Nobel Prize in Chemistry for the first crystallization of a virus.TMV is rod-shaped and is arranged in a helix of repeating protein subunits with its single stranded RNA embedded within it, resembling a nanotube.One TMV particle has 2,130 copies of the protein that forms the coat of the virus, which covers the 6,400 single stranded RNA nucleotides. The TMV RNA contains the genetic information for four genes, two of which are replicase-associated proteins, a movement protein and a coat protein. The RNA is coiled in a positive sense orientation acting as an mRNA in the host using the host ribosomes to translate its proteins and continue its reproduction.(Figure 1)

Transmission

It is easy for TMV to spread between tobacco plants, and other related plants, through direct contact such as leaf to leaf, or through indirect contact such as tools touching an infected plant and then touching a healthy one. One damaged tobacco plant cell is all it takes for a TMV particle to enter the organism and begin replication. Tobacco seed coats can also be contaminated with this virus, infecting the germination of the plant. TMV is also very stable and can still infect plants causing the mosaic disease for many years after its release. A study found that a 50 year old TMV that had been in storage at 4 °C could still infect tobacco plants. Once the TMV has used the host ribosomes to synthesize the replicase-associated proteins, they attach themselves to the 3’ end of the positive sense strand of the RNA and begin the synthesis of the negative sense strand of RNA. The negative sense RNA acts as the template strand for the synthesis of a full positive sense genome and subgenomic RNA (sgRNA). TMV uses its sgRNA and the host translational machinery to translate its movement protein (MP) and coat protein (CP), weighing approximately 30 kDa and 17.5 kDa respectively. When the coat proteins are synthesized they associate with the new positive sense RNA producing viable virions that can infect other healthy tobacco cells within the plant. Movement proteins are used to transport the viable TMV virons to neighboring cells through the plasmodesmata between the plant cell walls. Under normal conditions the plasmodesmata is too small for the whole TMV complex to pass through, but when bound to the movement proteins the plasmodesmata expands its diameter and allows the TMV virion to pass through the junction and infect the neighboring cell. This process is repeated for cell-to-cell infection, but once the virus particles reach the plants phloem, the vascular tissue in plants, the infection becomes systemic, spreading from the roots to the leaves.

Epidemiology

(Figure 2) Tobacco Mosaic Virus infection of neighboring cells .

Viruses need to have a host organism in order to continue having progeny and passing on their genetic code, TMV is no different. The host plants can be infected by TMV entering the plant through a damaged cell or contamination of virions on healthy seed coats, possibly from farmers not being careful when handling TMV contaminations. The virus can spread via clothing and tools on farms and in greenhouses. Since TMV particles are stable, they can survive in dead plant material over many months and through harsh weather conditions. This virus can also be a recurring problem if farmers plant the same crops year to year.(Figure 2)

Using TMV in Drug Delivery

Investigations looking into viruses as suitable carriers for cancer therapeutics have been gaining in popularity in recent years. Viruses have evolved to effectively spread their genes into host organisms for millions of years, making them logical candidates to use as delivery vectors of anticancer drugs to specific tissues. TMV has been increasing in popularity as delivery tool because plant based viruses are unable to infect or replicate in mammalian cells, and TMV is biocompatible in human blood and tissue. TMV is also biodegradable, clearing from non-target organs in a few hours. In the field of nanocarriers, aspect-ratio (AR), the length of a molecule divided by its width, is a statistic that is used to determine the efficiency of a nanocarrier to transport its drug cargo its destination. The dimensions of TMV used in the nanocarrier studies are 18 nm wide and 300 nm long with a 4 nm wide channel in the middle of the tubule . A high AR indicates that the carrier will have favorable targeting interactions and better distribution of its payload. The nanotube structure of TMV offers a high AR, which is good for evading the immune system, and has more efficient targeting interactions with tumor cells. Many people who receive chemotherapy are given drugs that are platinum based agents. Toxicity and drug resistance are major drawbacks for this category of drug. Off target toxicity from therapeutics results in the damage of healthy tissue, causing patients to fall ill to preventable diseases, fatigue, nausea and many more side effects. Cancer cells that do not fully terminate can persist on living causing recurring cancers and mutate into drug resistant forms. Using a carrier to hold the therapeutic drugs can reduce the toxic side effects of the drug. Carriers can be used as a targeting apparatus to bring the drugs to the cancerous tissue without any degradation of the drug to biological processes. Phenanthriplatin is a drug that is up to 40 fold more potent that other platinum agents, but its unable to be more effective than this class of drug because there is no efficient delivery mechanism established.

TMV as a Treatment for Melanoma

(Figure 3) TMV nanocarrier plus photosensitizer lead to cell death A) cell viability after 8hr of Zn-EpPor TMV B) Images of LIVE / Dead cells, green=live, red=dead .

Photodynamic therapy (PDT) is a different approach to treating melanoma, cancer of the skin. There are three requirements of this method and they are all nontoxic, a photosensitizer, light, and oxygen. The combination of these three requirements in a cell will lead to cell death. When the photosensitizer is exposed to a certain wavelength of light in the presence of oxygen it will begin to react, resulting in the release of a reactive oxygen species (ROS). The ROS will cause damage to the internal cellular components by oxidative stress causing the cell to undergo apoptosis. Photosensitizers are not soluble under biological conditions and do not accumulate in high enough concentrations in tumor tissues to be effective, and patients have to avoid sunlight after treatment to prevent unwanted activation the drug. The use of TMV as a delivery system for the photosensitizers offers a promising method to increase concentrations in tumor tissues and lower off target side effects.

The interior of the TMV has a high concentration of negative charges due to the glutamic acids that are exposed in this area. Research done by Lee et al. (2016) has capitalized on this detail using a photosensitizer, Zn-EpPor, which has positive charge molecules to load into the TMV based on charge-charge interactions. The positive Zn-EpPor will bind favorably to the negatively charged internal TMV tube. They found that approximately 800 molecules of their photosensitizer were successfully loaded into the TMV structure. Using fluorescence analysis, they found that there was a 40% increase in cellular uptake of the Zn-EpPor TMV than the Zn-EpPor alone in B16F10 melanoma cell lines. They report that when the melanoma cells take up the Zn-EpPor TMV, they are transported to the endolysosomes. The acidic environment in these organelles case the protonation of the carboxylic acids in the interior of the TMV and giving it a more positive charge and triggering the release of the Zn-EpPor drug. When this drug is then exposed to light, it begins to release ROS molecules causing the cell to die. After the release of the drug, the TMV shell will then be degraded by the proteases and hydrolyases that are contained within the lysozymes. When the melanoma cells were exposed to free Zn-EpPor and TMV Zn-EpPor, only in the light exposed samples was massive cell death observed (Figure 3) . The degraded parts are then excreted with the other waste products from cellular function. TMV works well as a nanocarrier in this field because of its effective delivery to target cells and release its payload while being able to be degraded and excreted without having any harmful side effects.

Tobacco Mosaic Virus (TMV) as a Vaccine

Francisella tularensis is a pathogenic bacterium that is responsible for the disease tularemia, also known as rabbit fever. The Center for Disease Control (CDC) has documented this pathogen as a biological threat. Several countries have studied this bacterium as a bioweapon for biological terrorism. The grounds for F. tularensis to be considered a biological weapon can be attributed to its rapid infection rate, infectious as a vapor, incapacitates its hosts, and causes death in a short period of time. Previous attempts to develop a vaccine have not been successful, with the closest usable vaccine was the Russian Live Vaccine Strain (LVS) that was made using F. holartica. While this vaccine did offer some resistance to tularemia, it still caused health problems and negative reactions in its patients, forcing the US Food and Drug Administration (FDA) to turn down approval for its development. Currently, a vaccine has not been developed yet for the prevention of this disease. Research by Banik et al. (2015) has investigated the potential of using the tobacco mosaic virus as a delivery apparatus to transport a potential vaccine molecule to combat tularemia.

(Figure 4) Two different designs for the TMV nanocarrier .

The failure of these previous vaccines can be attributed to their design. Several only having a single subunit of a larger structure aimed at targeting the bacteria, but was not able to defend against all the possible strains ofF. tularensis. Another possible explanation for the vaccine failures could have been that single targeting proteins were unable to be affective against all the possible orientations of the bacterial proteins. Or a lack of the proper antigens to stimulate an immune response that would target the bacteria. A new approach to developing a vaccine against this disease is two fold; the first being to develop a vaccine that is effective against F. tularensis, and second is to use TMV as its delivery mechanism.

TMV is of particular interest as an antigen carrier because its structure allows for it to be ingested by cells easily and able to display antigens on its surface. It’s possible that TMV is able to simulate antibody production due to the exposed repeated antigens on its surface, this mimics other viral coats that cause the human body view it as threatening. Or the other possibility, TMV RNA stimulates the cell-mediated immunity cascade . Another positive of using TMV as a vector for the vaccine in drugs is the fact that TMV cannot infect mammals nor does it have negative affects on host antibodies. TMV can be used several times as a booster for multistep vaccinations.

To test this method, Banik et al. (2015) used recombined proteins from F. tularensis to stimulate an immune response in mice. They used two different combinations, three different recombinant proteins from F. tularensis inside an TMV (multiconjugate), and a TMV with one type of recombinant F. tularensis protein (monofonjugate)(Figure 4). They found in their research that when TMV was paired with these recombinant proteins, an immune response was activated. Antibodies formed against the three recombinant proteins that were used in the experiment. The TMV-monoconjugate had less antibodies being produced than the amount of antibodies produced in the TMV-multiconjugate response. The implications of these results could lead to the formation of a vaccine against bacteria that can be used to inoculate an entire population. This result would also provide humanity with a method to combat an outbreak of a deadly biological weapon in the event of military hostilities escalating to chemical warfare.

TMV Scaffolds Used to Make Other Nanoparticles

Nanoparticles are well sought into because of their applications in drug delivery and in other nanotechnologies. The tobacco mosaic virus is a symmetrical rod of repeating subunits of protein stacked on top of each other, which has the ability to be reformed into other nanoparticle shapes. Research into diverse nanoparticle shapes are being looked into to see their potential use in in vivo studies. Research has found that when TMV rods are heated up they can collapse to take on different structures and sizes of spherical nanoparticles. These different sized nanoparticles have the potential to be used in the medical field to find a variety of delivery and imagining agents. The spherical nanoparticles form as a result of heating the TMV to 94 °C for at least ten seconds. The size of the nanoparticles can vary depending on the concentration of the TMV that is in solution at one time. The reported sizes of these spherical nanoparticles are between 50-800 nm in diameter and can range in morphology by changing the temperature and time under heat conditions. For the complete conversion of TMV into the spherical nanoparticles, 125 seconds at 100 °C is required. Shorter times at 100 °C will result in a mix of TMV rods and spheres. Longer times at 100 °C will not change the morphology of the spherical nanoparticles. Further investigation into these different nanoparticle shapes could lead to novel structures that can be used for medical applications in drug delivery that researchers are not yet aware of.

TMV Applications in Micromachines

Small-scale machines and batteries, micromachines and microbatteries, have modern day applications in medical devices, commercial electronics, and wireless sensors that are used in environmental, homeland security, and structural features. Like all other technology, energy is required to make it function, but with the miniaturization of technology, the source of power for these devises also needs to get smaller. The most desired batteries or energy generators are able to have a large energy output with minimal area taken up within the device. Research that has been investigating the improvement of batteries at the microscale level has focused heavily on microelectromechanical systems (MEMS). This technology uses electrodes that are three-dimensional shapes, opposed to two-dimensional films, to increase the surface area of the electrodes for more reaction space. Another approach to improving battery function at the nanoscale level is the use of nanomaterials to increase the area of space that the electrode and electrolyte meet, increasing the reaction area. Increasing the stability of these chemicals on the small scale also increases the efficiency of the microbatteries. The tobacco mosaic virus is useful in this technology as a scaffold for the production of nanomaterials.

(Figure 5) Micropillers of the TMV on electrodes in a microbattery .

The TMV is more effective in this role when it’s modified with extra cysteine amino acid residues. The thiol functional groups improve TMV’s ability to bind (self assemble) to metal surfaces of the electrodes. When the TMV adheres to the surface of the electrodes they form a strong network that increases the surface area of the electrode for more reactions to take place with the electrolytes. This structure also helps to stabilize the microbattery as a whole. TMV is able to conduct electrons when a metallic film covers the virus to enhance its conductivity. V2O5 was one of these materials used to increase TMV’s conductivity. Underneath the TMV structure, a metal such a gold, nickel or copper, is used to collect the electrons of the reactions from the surface between the conductive TMV and electrolyte solution. In this battery, gold was used as the material of choice to form the electrode pillars (Figure 5). The reasoning for this choice was due to the fact that TMV was able to self assemble onto the gold pillars easily, and it’s inert at the voltage range that the battery was operated in (2.6-3.6 V)(Figure 5). The implications of these experiments can lead to the development of microscale batteries that have the capability to power small machines; which can be used to for commercial, medical, and military applications.

Conclusions

The tobacco mosaic virus has been studied and documented for well over a century. Its origins of research are based on its infectious pathogenic nature in several species of plants. The structure of TMV, hollow rod of repeating protein subunits surrounding a single RNA strand, allows for researchers to take advantage of nature’s creation and utilize it for other purposes. The hollow nanotube structure of the TMV offers a place for therapeutic drugs to be held and then delivered to its target destination, such as tumors and cancer cells. It’s fascinating that this virus started its relationship with humankind as a killer of crops, but now its being used for many different applications to solve human problems.

Authored for BIOL 238 Microbiology, taught by Joan Slonczewski, 2018, Kenyon College.

A typical mosaic pattern on flue-cured tobacco leaves infected with tobacco mosaic virus.
Photo: Courtesy of JP Krausz

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Tobacco mosaic virus (TMV) was the first virus discovered.

In 1889, Martinus Beijerinck, found that ‘tobacco mosaic disease’ was caused by a pathogen able to reproduce and multiply in the host cells of the plant. He called it ‘virus’ (from the Latin virus, meaning poison) to differentiate this form of disease from those caused by bacteria.

Tobacco yield losses
due to TMV are currently estimated at only 1%, because resistant tobacco varieties are routinely grown. However, TMV affects other crops, and losses of up to 20% have been reported in tomatoes.

TMV can be a major problem because, unlike most other viruses, it does not die when the host plant dies and can withstand high temperatures. Thus, the virus can survive on implements, trellis wires, stakes, greenhouse benches, containers and contaminated clothing for many months.

It can also survive in crop debris on the soil surface and infect a new crop planted on contaminated land.

Tobacco products, particularly those containing air-cured tobacco, may carry TMV too.

The virus cannot be transmitted in the smoke of burning tobacco, but smokers, especially those who roll their own cigarettes, could possibly carry the virus on their hands and transmit it to healthy plants.

Sap-feeding insects such as aphids cannot transmit TMV. However, chewing insects such as grasshoppers and caterpillars do occasionally transmit the virus. They are not considered important vectors, however.

Transmission
Tobacco mosaic virus is usually spread from plant to plant via ‘mechanical’ wounds caused by contaminated hands, clothing or tools such as pruning shears and hoes. This is because TMV occurs in very high concentrations in most plant cells. When plants are handled, the tiny leaf hairs and some outer cells are inevitably damaged and leak sap onto hands, tools and clothing.

Seeds from infected plants can also carry the virus on their seed coats. The earlier the age at which the mother plant is infected, the more likely it is that the virus will contaminate the seed coat during seed harvesting. When the seed germinates, the virus may enter the seedling through small cuts caused by transplanting and handling, or during the germination/emergence process.

Once inside the plant, the virus releases its genetic code (RNA). The plant mistakes this for its own RNA, and starts to produce viral proteins.

The virus then spreads to neighbouring cells through microscopic channels in the cell walls (plasmodesmata), and eventually enters the translocation system of the plant (xylem and phloem). From here, it spreads to the entire plant.

Signs and symptoms
Symptoms first appear about 10 days after infection. The plants do not usually die, but growth can be seriously stunted. In the case of tomatoes, certain TMV strains can cause deformed fruit, non-uniform fruit colour and delay ripening.

Specific symptoms depend on the host plant, age of the infected plant, environmental conditions, the virus strain and the genetic background of the host plant.

However, common signs include mosaic-like patches (mottling) on the leaves, curling of leaves and the yellowing of plant tissues.

Managing the virus
No chemicals can cure a plant infected with a virus, and TMV is no exception. As mentioned before, however, resistant plant varieties are available.

You will need to consider adaptability, potential yield and resistance to other important diseases when selecting varieties.

Ultimately, effective TMV management involves using virus-free seedlings or plants and implementing strict hygiene procedures:

  • Use new potting mix and new or thoroughly cleaned seedling trays when growing seedlings;
  • If infected plants are discovered, either remove and destroy the plants and restrict access to the area, or always work in the affected area last and decontaminate yourself and your equipment afterwards;
  • Remove all crop debris from the land, seedling production beds and benches in greenhouses;
  • Place tools in a disinfectant solution for at least 10 minutes and rinse thoroughly with tap water;
  • Disinfect door handles and other greenhouse structures that may have become contaminated by wiping thoroughly with recommended disinfectants;
  • Thoroughly wash your hands with recommended disinfectants, such as carbolic soap, or a mixture of non-fat milk powder at 20% weight/vol, 10% bleach, and 70% ethanol, after handling tobacco products or TMV-infected plants. Make sure that the solutions are fresh, and replace regularly (it is recommended that the bleach solution be replaced every four hours).

If you are a seedling producer, ensure that greenhouses are within a clean zone and control the movement of people, plants, vehicles and materials into the greenhouse areas.

Treat each greenhouse as a separate unit, with protective clothing, tools, gloves and bins in each. These items should not be moved between units.

The ARC’s Industrial Crops unit (Rustenburg) has a virus diagnostic laboratory and conducts diagnostic services for nurseries and farmers.

Email Phillip Mphuti at .

Infection and coaccumulation of tobacco mosaic virus proteins alter microRNA levels, correlating with symptom and plant development

Results

Virus Infections Cause Different Types of Symptoms and Alter Accumulation of miRNAs.

Groups of ≥20 N. tabacum plants were mechanically inoculated with TMV, ToMV, PVX, TEV, or PVX in two independent experiments. All plants were placed in the same greenhouse for the duration of each experiment, and the percentage of plants exhibiting disease symptoms and severity of symptoms was recorded . We also recorded the number of infected plants with flowers and the height of the plants 30 days after inoculation.

Under the conditions used in this study, TMV and ToMV developed disease symptoms in a shorter period than TEV, PVY, and PVX. TMV and ToMV infection caused delays in flowering, and plants were taller than noninfected plants. Infection by TEV and PVX produced symptoms in an intermediate time period compared with TMV and PVY; plants infected with PVY showed symptoms later in time than did infection with TMV. Symptoms caused by PVX infection were relatively severe; TEV infection produced chlorosis in leaves with mild leaf distortion, and PVY cause mild mottling of infected leaves (SI Fig. 5).

Based on these data, a disease severity rating was created (Table 1). In sum, in this study, TMV and ToMV were the most aggressive, PVX and TEV were less aggressive, and PVY was the least aggressive.

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Table 1.

Relative index of disease symptom severity

To determine whether infection by TMV, ToMV, PVX, PVY, or TEV altered miRNA accumulation, groups of 12 N. tabacum plants were separately infected with each virus. At 10 days after infection, sRNAs were isolated from six to eight leaves per group, and accumulation of a selection of miRNAs was analyzed by Northern blots by using sequences from Arabidopsis thaliana that are correlated with plant development. Fig. 1 shows the relative accumulation of miRNAs in two independent biological replicates. Hybridization with miRNA was measured by using a radioactivity-scanning device and normalized based on the amount of rRNA quantified by using ethidium bromide staining of gels. The amount of miRNA species in noninfected plants was arbitrarily set at 1.0, and other data were computed relative to these plants. We observed that infection by TMV and ToMV caused highly significant increases in the levels of most of the 10 miRNAs tested. TEV and PVX caused moderate changes in the miRNAs tested, whereas infection with PVY caused the fewest changes in miRNAs (Fig. 1). miRNAs 156, 160, 164, 166, 169, and 171 were most severely affected. miR171* was the only complementary strand (miRNA*) detected, although complementary strands of all miRNAs were tested (data not shown). Failure to detect a sequence is not conclusive evidence of lack of presence and can be explained by low levels of miRNA* and/or problems with detection.

Fig. 1.

miRNAs accumulation is altered by viral infections. (Left) Northern blot analysis to detect the accumulation of various miRNAs and miRNA* after infection with selected viruses. Ethidium-bromide-stained rRNA shown below each blot was used to normalize data. (Right) Average and standard error of miRNA level of two independent biological replicates. The data were derived based on hybridization of RNAs derived from noninfected plants, established as 1.0.

Suppression of PTGS by Viral Infections.

To analyze PTGS suppressor activity by these viruses, 3-week-old transgenic N. benthamiana plants that constitutively express a gene encoding GFP were used. When plants of this line were infiltrated with Agrobacterium tumefaciens carrying an inverted repeat of the gfp construct (27), the bright green fluorescence normally produced in these plants suppressed by 25–30 days, as expected, as a consequence of gene silencing (14). Silenced plants were then separately inoculated with the five viruses. When systemic symptoms were observed, the plants were analyzed under UV illumination. As expected, most of the plants infected with PVY and TEV recovered green fluorescence, indicating that infection with either virus suppressed PTGS (Table 1) (15). In contrast, the majority of plants inoculated with TMV, ToMV, and PVX did not show a GFP signal, indicating these viruses did not have a strong suppressor of PTGS (Table 1). However, small areas of GFP fluorescence, mainly around leaf veins, were observed in a low number of ToMV and TMV infected plants (Table 1). The level and tissue specificity of PTGS suppression activity of the viruses reported here are in agreement with previous studies that used similar assays (Table 1) (15).

Molecular Characterization of Transgenic Plants Expressing MP and/or CP.

To study the effect of transgenic expression of TMV MP and CP on CP-mediated resistance. and miRNAs, we crossed a transgenic line that produces the TMV MP (plant line 277; refs. 19 and 28) with a transgenic line that produces a mutant of TMV CP, CPT42W (23); both plant lines were developed in N. tabacum cv Xanthi and have been extensively characterized (17, 22, 24, 26). F1 progeny of the cross were normal in appearance and were selfed to obtain double-homozygous lines; three F3 lines were selected for further study.

The presence of both transgenes in F3 plants was confirmed by genomic PCR (SI Fig. 6), and accumulation of MP and CP mRNAs and proteins was established via Northern and Western blot assays (SI Fig. 6). Two of the three F3 homozygous lines accumulated similar levels of CP and MP RNA and protein (lines nos. 21 and 22). In the third line (no. 18), neither MP nor CP RNA nor protein was detected, suggesting that both genes were silenced. This hypothesis was supported by an analysis that detected the accumulation of sRNA that includes CP gene sequences (SI Fig. 6). The silenced line, named mpxcpT42W*, and line 22, referred to as MPxCPT42W, were selected for further studies.

Coexpression of TMV MP and CPT42W Alters Plant Development.

MPxCPT42W lines 22 and 21 exhibited severe morphological changes and poor fertility. F3, F4, and F5 progeny exhibited mild mosaic patterns on leaves, reduced number of plants that produce flowers, and reduced number of flowers per flowering plant (summarized in Table 2 and Fig. 2). Other phenotypes include reduced seed germination, deformed seedlings, reduced plant height, and abnormal flower morphology (Table 2 and Fig. 2). Plant line 277 (accumulates MP) may exhibit mild chlorosis and narrow leaves under conditions of high light and temperature (M. Deom and R.N.B., unpublished data). The CPT42W parental line is indistinguishable from the nontransgenic plants or line mpxcpT42W*. Based on these observations, we proposed that accumulation of MP + CPT42W was responsible for the phenotypes described above.

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Table 2.

Quantitative description of altered development of MPxCPT42W plants

Fig. 2.

Abnormal phenotyes of line MPxCPT42W. (A)WT tobacco (N. tabacum, cv Xanthi nn; Sx) and (B and C) flowers of plant line MPxCPT42W. (D) Leaves of nontransgenic (Left) and of MPxCPT42W plants highlighting the rounded shape of the leaves (Right). (E) Normal bipartite stigma of WT plants and (F) tripartite stigma of line MPxCPT42W. (G and H) MPxCPT42W seedlings showing cup-shaped or partially fused cotyledons. (I) WT stamens and pistils. (J) Stamens of MPxCPT42W. (K) Symmetric leaves of WT seedlings and (L) asymmetric leaves of line MPxCPT42W. (M) Normal morphology of flowers of Sx, MP, CPT42W, and mpxcpT42W*, respectively. (N) Phenotypes of flowers of MPxCPT42W. (O–Q) Scanning electron micrographs of abaxial surfaces of MP, CPT42W, and MPxCPT42W leaves. P, Q, and R, respectively, show altered shapes of epidermal cells, as well as altered sizes. (Scale bars: 50 μm).

We tested this hypothesis by a grafting study to silence expression of the transgenes. PTGS produces a mobile signal that can cross graft junctions and induce silencing of a homologous transgene in the grafted scion (28, 29). When line MPxCPT42W was used as scion and line mpxcpT42W* as a rootstock, the abnormal phenotypes in the scion were partially abrogated (Table 2). Height, percentage of plants that produce flowers or buds, and numbers and shape of flowers at 2 mo after grafting were not significantly different from WT plants or transgenic lines 277 (MP), CPT42W, or mpxcpT42W* (Table 2). Western blot assays of scion tissues of grafted plants were performed to monitor the accumulation of MP and CP; most samples did not accumulate detectable levels of MP and CP. However, MP and CP were detected in one particular plant, showing that the PTGS was not established in this scion. As expected, this plant showed phenotypes similar to MPxCPT42W. The reciprocal graft (i.e., MPxCPT42W, used as rootstock) did not show unusual phenotypes (Table 2).

As controls in this study, we developed grafted plants comprising MPxCPT42W as scion with rootstocks of transgenic plants that produce CPT42W, MP or nontransgenic plants. None of these grafted plants restored the normal phenotype to the scion (data not shown). We concluded from these studies that coexpression of MP + CP is responsible for the abnormal development observed on MPxCPT42W lines.

The phenotypes exhibited in line MPxCPT42W are similar to those exhibited by a group of transgenic A. thaliana plants in which either miRNAs or targets of miRNA were altered (10, 13, 30–32). As described in Fig. 2, flowers of line MPxCPT42W exhibit a loss of symmetry (Fig. 2 C), altered number and shape of reproductive organs (Fig. 2 B, J, and N), and stigmas were frequently tripartite (Fig. 2 F and Table 1). Other changes were also observed (Fig. 2) and may be the cause of low fertility in this plant line (compare Fig. 2 J–I).

A high percentage of seedlings of the F3 progeny of MPxCPT42W (see Table 2) produce abnormal cotyledons (cup-shaped or partially fused cotyledons) (Fig. 2 G and H) and asymmetrically shaped leaves (Fig. 2 L) compared with nontransgenic plants (Fig. 2 K). Similar phenotypes were described in A. thaliana with increased miR164 levels (31, 32).

Leaves of MPxCPT42W have an unusual round shape: the length/width ratio of the leaves was 0.68 and is statistically different from WT plants (Table 2; Fig. 2 D). In addition, MPxCPT42W leaves appear more waxy with a rough/hard texture (Fig. 2 D) and epidermal cells are larger than nontransgenic leaves (Table 2; compare Fig. 2 Q and P with Q). Similar changes were observed in transgenic A. thaliana plants with changes in miRNAs (30, 31). Recently, it was reported that miR160 regulates genes that alter epidermal cell shape in A. thaliana (33). These observations led us to consider whether or not the activity of this and/or other miRNAs are affected in plant line MPxCPT42W (33).

Accumulation of miRNAs Is Altered in MPxCPT42W Plants.

We investigated the accumulation of selected 21-nt miRNAs that are involved in plant development in A. thaliana in MPxCPT42W plants by using miRNA probes designed from A. thaliana sequences. Fig. 3 shows the results of Northern blot hybridization that detects specific miRNAs. In each experiment, we also included samples of leaf tissues 15 days after inoculation with TMV, as well as tissues from parent plant lines, and line mpxcpT42W*. The radioactive signal was normalized to the amount of ribosomal RNA in each sample. Numbers presented above each subfigure represent the average relative accumulation of each miRNA from two biological replicates, compared with noninfected, nontransgenic plant tissues (set at 1.0). Standard errors are given in parentheses.

Fig. 3.

Effects of TMV MP and CPT42W and TMV infection on accumulation of miRNAs. (A and B) Northern blot analyses to detect the accumulation of miRNA and miRNA*. Ethidium-bromide-stained rRNA shown below each blot was used to normalize data. Relative accumulation of miRNA compared with noninoculated and nontransgenic N. tabacum plants (Sx = 1.0) is shown. The average of two independent biological replicas and standard errors (in parentheses) is shown.

miRNAs 156, 164, 165, and 167 accumulated to higher levels in MPxCPT42W and TMV-infected plants compared with nontransgenic and noninfected tissue (Fig. 3 A). On the other hand, transgenic plants MP, CPT42W, and mpxcpT42W* and nontransgenic plants accumulated similar amounts of these miRNAs (Fig. 3 A). Therefore, there is a strong correlation between increased miRNA accumulation and the aberrant phenotype exhibited in plant line MPxCPT42W and in TMV-infected plants. Accumulation of miR 156 was also elevated in the MP plant line (line 277) compared with nontransgenic plants, although not to the level of the MPxCPT42W line (Fig. 3 A). Although accumulation of miR160 was also somewhat increased in line 277, as was miRNA 156, 164, 165, and 167, the differences were not considered significant (Fig. 3 B). Accumulation of miRNA 171 and its complement (miR171*) were altered only in TMV-infected plants, in agreement with Fig. 1 (see Fig. 3 B).

MP Interacts with CPT42Win Vivo.

Because MPxCPT42W plants, but not either of the parent lines, exhibit abnormal phenotypes and changes in miRNAs, we investigated the possibility that TMV MP interacts in vivo with CPT42W. For this study, we used bimolecular fluorescence complementation (BiFC). BiFC is based on the formation of a fluorescent complex when two fragments of yellow fluorescent protein (YFP) are brought together by interaction between proteins fused to the fragments (34, 35). Sequences encoding YFP a.a. 1 – 155 YFP and a.a. 156–239 (YFPN and YFPC, respectively) were fused to sequences encoding the MP or CPT42W to produce MP-YFPN, MP-YFPC, CP-YFPN, and CP-YFPC. Constructs encoding YFPN and YFPC (E-YFPN and E-YFPC) and constructs encoding MP-YFPN, MP-YFPC, CP-YFPN, and CP-YFPC were developed (SI Fig. 7).

Leaves of N. benthamiana were infiltrated with a suspension of A. tumefaciens harboring the BiFC constructs, and sites were examined via fluorescence microscopy. Controls that induced coexpression of E-YFPN + E-YFPC or constructs encoding only MP or CP did not produce fluorescence (Fig. 4). In contrast, YFP fluorescence was detected after infiltration that caused coexpression of MP-YFPN + MP-YFPC, and CP-YFPN + CP-YFPC, indicating interactions between each protein. Coassembly of CP monomers is well known (36), and coassembly of MP in vitro was previously demonstrated (37) (Fig. 4).

Fig. 4.

TMV MP interacts with CPT42W in vivo by BiFC. YFP epifluorescence microcroscope images of N. benthamiana epidermal leaf cells in leaves agroinfiltrated with a mixture of Agrobacterium strains harboring constructs encoding the indicated fusion proteins. Each image is a representative picture of several experiments.

CP and MP colocalize with each other during TMV infection (23). When the BiFC assay was applied to coexpression of the CP-YFPN + MP-YFPC, fluorescence was observed (Fig. 4); we did not detect signal when CP-YFPC and MP-YFPN were coexpressed (Fig. 4). These studies provide evidence that MP interacts with CP in vivo, and that such interaction is assembly-specific and/or that orientation of the interacting proteins interferes with fluorophore assembly. Similar results were described for other plant proteins (38, 39).

Because plants that produce both MP and CP exhibit abnormal development, we suggest that complexes comprising MP + CPT42W possess functions not inherent in either protein alone, including altering miRNA accumulation.

TMV MP and CP Do Not Suppress PTGS.

Virus proteins that function as suppressors of PTGS can alter plant development and miRNA accumulation or/and activities (10–12). Therefore, we conducted experiments to determine whether TMV MP, CPT42W, or MPxCPT42W suppress PTGS to elicit the phenotypes in plant line MPxCPT42W.

Local Silencing.

Leaves of nontransgenic N. benthamiana were infiltrated with A. tumefaciens carrying a GFP gene construct (35S-GFP), resulting in transient GFP expression (SI Fig. 8). Coinfiltration with Agrobacterium strains that carry genes encoding 35S-GFP and an inverted repeat of GFP sequences (35S-dsGFP) does not result in green florescence, because PTGS is triggered by the 35-dsGFP gene (SI Fig. 8) (40). Similarly, transient expression of the PTGS suppressor HC-Pro (35S-HC-Pro) simultaneously with 35-GFP + 35-dsGFP inhibits silencing of the 35S-GFP gene, resulting in fluorescence comparable to that induced by 35S-GFP (SI Fig. 8).

Leaves of N. benthamiana plants were coagroinfiltrated with genes encoding 35S-GFP + 35S-ds-GFP + 35S-MP (35S-MP produces TMV MP), 35S-GFP + 35S-ds-GFP + 35S-CPT42W (35S-CPT42W produces TMV CPT42W), or 35S-GFP + 35S-ds-GFP + 35S-MP + 35S-CPT42W (SI Fig. 8). In none of these experiments did we observe GFP fluorescence. These data, summarized in Table 3, suggest that transient expression of MP and/or CPT42W did not suppress PTGS in this system and led us to propose that these proteins do not suppress PTGS in transgenic lines of N. tabacum used in this study.

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Table 3.

MP and CPT42W do not suppress local and systemic PTGS

Systemic Silencing.

We conducted experiments to determine whether TMV MP and/or CPT42W prevent spread of the gene silencing signal as does p25 of PVX (16). Three week old transgenic N. benthamiana plants expressing GFP (14) were agroinfiltrated with 35S-dsGFP or coinfiltrated with 35S-dsGFP + 35S-HC-Pro, 35S-p25, 35S-MP, 35S-CPT42W, or 35S-MP + 35S-CPT42W. Systemic silencing of GFP was obtained when plants were inoculated with 35S-dsGFP alone, and when 35S-dsGFP was coinfiltrated with 35S-MP, 35S-CPT42W, or 35S-MP + 35S-CPT42W (Table 3). As expected, coinfiltrating genes encoding HC-Pro or p25 prevented the spread of gene silencing (Table 3) (16). In contrast, expression of genes encoding MP, CPT42W, or both did not prevent systemic silencing of GFP in transgenic N. benthamiana.

New Users

Tobacco mosaic virus (TMV) results in losses in North Carolina of about 1 to 2 percent of the crop by reducing the yield and quality of flue-cured tobacco. The ideal way to control mosaic is by the use of resistant varieties; several new hybrids, which have acceptable yield and quality, are now available. Strict sanitation procedures are necessary to prevent the virus from becoming established in the crop and to prevent spread of the virus if efforts to keep it out of the crop are unsuccessful. Crop rotation helps to keep losses minimal in fields where mosaic has occurred.

Symptoms

Mosaic is so common that most tobacco growers know the symptoms of the disease in the field. The most characteristic symptom is a “mottled” appearance of the leaf (alternate areas of light and dark green tissue) (Figure 1). The tissue may be rough (Figure 2), and will burn on hot, sunny days (Figure 3). Several other viruses cause symptoms on flue-cured tobacco that look like mosaic. Symptoms on seedlings are much milder and easily overlooked. Stunting and mild mottling may be observed (Figure 4). The first step in controlling mosaic, therefore, is to be sure that the virus causing the mosaic symptoms is TMV. Growers who have a mosaic problem should get assistance from their County Extension Center if there is any doubt as to the identity of the causal virus.

Pathogen biology and epidemiology

Properties of the Virus: Tobacco mosaic virus, like other viruses, is a very small chemical particle that can multiply only in a living host and only can be seen with an electron microscope. It differs from other viruses that infect tobacco in two ways that are important in its control. First, TMV is very resistant to destruction. It will survive for at least 50 years in dead, dried tissue while other viruses become inactive when their host plant dies. Second, TMV is primarily transmitted mechanically while insects primarily transmit the other viruses.

Transmission: Tobacco mosaic virus is transmitted mechanically by any means that results in the virus coming in contact with injured cells of a host plant. The primary mechanism for this is contaminated worker’s hands or equipment that comes into contact with a healthy plant. Contaminated hands can be freed of the virus by washing with a detergent. The virus can be inactivated on equipment by scrubbing it with a brush using detergent or by steaming. Although the virus is transmitted primarily on worker’s hands and equipment, anything that mechanically moves the virus from a source to a healthy plant can transmit it. Chewing insects, such as flea beetles and grasshoppers, are capable of transmitting the virus, but such transmission is very rare in nature. Seed may be infested with the virus and transmit the virus.

Source of the Virus: Tobacco mosaic virus overwinters in a number of ways, and these must be understood for successful control through sanitation practices. Infection of tobacco from overwintering sources of the virus is known as “primary infection.”

A. Tobacco Products – All forms of tobacco may carry TMV so it is advisable that these products not be used by workers, especially around greenhouses and during transplanting. Spraying plants with milk prior to pulling and transplanting will reduce the number of plants that become infected. It should always be used during transplanting in situations where tobacco use by workers cannot be prevented. Transplants should be sprayed until runoff with one pound of dried milk in a gallon of water immediately prior to pulling.

B. Tobacco Trash – Tobacco mosaic virus will survive for years in dried tobacco tissue, so anything that may be contaminated with pieces of leaf, stem, or root tissue should be cleaned prior to use in the crop. This includes float trays that may have infected roots in the walls of the tray.

C. Soil-Borne Virus – tobacco mosaic virus overwinters in infected stalk and root. Experimental data in North Carolina indicates that infection occurs at transplanting when a plant is pushed against a piece of virus-infected tissue. The number of transplants that become infected in this way will depend on quantity of overwintering tissue surviving. The virus will overwinter in dead as well as living plant tissue, but dead tissue contains less active virus than living tissue. Numerous studies during the past 40 years in North Carolina has shown that from 0.1 to 5.5% of the plants planted to a field that contained mosaic the previous year will become infected with TMV.

D. Other Crops – Tomato, pepper, and eggplant are hosts of the TMV and in addition to not being used in rotation should not be handled prior to working in tobacco. Fruit from these crops also contain active virus and should not be handled while working in tobacco. There is, for example, enough active TMV in one infected tomato to infect every tobacco plant grown in North Carolina.

E. Weeds – A number of weeds are known to be hosts of TMV, but horsenettle is the only one found thus far to be infected in North Carolina. This weed is common where flue-cured tobacco is grown and frequently is found infected with TMV. The virus can be transmitted from infected horsenettle to tobacco mechanically, and this weed is suspected to be the source of TMV found in many tobacco fields. The only way horsenettle can be eliminated as a source of TMV is to eradicate it. Growers should consult with their County Extension Center for eradication procedures.

Secondary Spread: Most of the mosaic plants in heavily diseased fields were infected by virus that was spread from a few tobacco plants infected from overwintering sources of the virus. Secondary spread can be reduced by removing primarily infected plants from the field or by cultivating in a manner to prevent spread of the virus. In seedling production, clipping is a very effective means of spreading TMV. Seedlings should be scouted closely for TMV at each clipping.

There are a number of factors that must be considered when deciding if roguing in the field is feasible, so the final decision must be based on each situation. As a general guideline, however, the two most important factors are the time that primary infection occurred and the number of infected plants.

Although primary infection can occur anytime during the growing season, the most critical time is during transplanting. Plants infected at this time from any of the overwintering sources will show mosaic symptoms in 2 to 4 weeks. Generally, if fewer than 100 plants per acre are showing symptoms at this time it is feasible to remove them. Roguing should, of course, be done prior to cultivation. Plants used to replant rogued plants frequently become infected from virus in the roots left in the soil when plants are removed so replanting is not recommended. If too many plants are infected to make roguing feasible, secondary spread can be reduced by carrying out cultivation operations so that equipment does not touch the plants. Where this is possible, cultivation should be done when the plants are dry and preferably partially wilted because less virus transmission occurs under these conditions.

It is seldom feasible or of value to rogue plants after layby because most secondary spread occurs during or before this operation. Losses can be reduced by keeping a good supply of water to the crop because mosaic burn, the most damaging form of the disease, usually occurs only when infected plants come under water stress. Irrigation of a field containing a significant amount of mosaic may be worthwhile under conditions of moisture stress that would not be of value to rogue after layby if fewer than 10-20 plants are showing symptoms by the time the crop is knee-high. This will reduce losses on the current year’s crop somewhat, and perhaps more importantly, prevent extensive spread to the remainder of the crop and thus overwintering of virus in stalks and roots.

Management

The problem of TMV carry-over in stalks and roots can be reduced by crop rotation or by using a mosaic resistant variety.

Crop Rotation: Crop rotation for mosaic control consists of planting a crop that is not susceptible to mosaic in alternate years. Crops planted in North Carolina that are susceptible to TMV are tobacco, tomato, pepper, and eggplant. Tobacco varieties carrying mosaic resistance are essentially non-hosts of the virus and can be used as a rotation crop. Growers who do not want to plant their entire crop to mosaic resistant varieties might at least consider planting them in fields where a mosaic problem occurred the previous year.

Growers who find rotation unfeasible and who do not want to plant any of their crop to a mosaic resistant variety can reduce virus carry-over by doing a thorough job of stalk and root destruction. This will significantly reduce, and may in some situations eliminate, infection during transplanting from infested old crop debris. Plants that do become infected should be removed prior to the first cultivation to prevent spread of the virus.

Resistant Varieties: Mosaic resistant varieties have historically been lower in yield and quality than non-resistant varieties. Some of the new releases compare favorably with non-resistant varieties, however. Even when resistant varieties may not perform as well as resistant varieties on a given farm, there are two situations where they would be of benefit. The first is on problem mosaic farms where losses to mosaic exceed the differential income potential between a mosaic resistant and a susceptible variety. The other situation is where monoculture is practiced and a mosaic resistant variety can be grown on a problem field to break the cycle of virus carry-over. Varieties with TMV resistance are available in the Tobacco Production Guide:

Other Links

North Carolina Ag Chem Manual:

Creation date: 2001

Revision date: April 2010

Key words: tobacco, mosaic, rotation, resistance, variety

Glossary terms: mottling, transmission, infection, spread

Common questions and answers about tobacco mosaic virus

Tobacco mosaic virus (TMV) is highly transmittable through routine greenhouse operations. If you have found TMV on plants in your greenhouse this season, Michigan State University Extension recommends their immediate disposal. We have compiled common questions from growers and their answers.

What is TMV?

TMV is a single-stranded RNA virus that commonly infects Solanaceous plants, which is a plant family that includes many species such as petunias, tomatoes and tobacco.

What are the hosts of TMV?

Pathologists estimate that there could be up to 350 plant species susceptible to TMV. According to Spence et al. in the European Journal of Plant Pathology, some of the more susceptible species that show symptoms are petunia, bacopa, verbena, scaevola, diascia, calibrachoa and lobelia. Some species can be a host for the virus, but not show symptoms.

How stable is TMV?

TMV is an incredibly stable virus. In fact, it is so stable that it can remain in tobacco plants after the extensive processing necessary to make tobacco products.

Why do symptoms differ between infected plants?

Symptoms differ between infected plants depending on the stage of disease severity, the genetic line of the virus and the host plant.

Can tobacco products carry TMV?

Yes, tobacco products can carry the virus and using them without washing your hands afterwards can potentially spread TMV. For that reason, do not use tobacco in the greenhouse or without washing your hands prior to handling plant material.

Can TMV stay viable in plant debris or dead plant material?

Yes, TMV can stay active in dead plant material for long periods of time. It can even stay viable without the presence of a host on surfaces such as greenhouse benches, floors and worker’s clothes.

How effective is spraying plants with milk to prevent the virus from spreading?

Spraying plants with 20 percent nonfat dry milk has been shown to be somewhat effective in preventing the spread of the virus from TMV-infected tobacco plants to uninfected tobacco plants. We recommend spraying plants prior to transplanting to reduce the risk of spreading TMV as part of a methodical management strategy.

How “full-proof” is spraying plugs or liners with milk?

While spraying milk on plugs or liners may have some effectiveness in reducing the spread of TMV, spraying milk should not be the primary management tool for TMV in your greenhouse. In order for milk to inactivate TMV, it must be liquid. Remember to continue scouting, testing, disinfecting and implementing the best sanitation possible in your facility.

How does the milk work to inactivate the virus?

Milk coats the virus and inactivates it.

Is it possible to receive a positive and a negative TMV test result from two different samples on the same plant?

TMV may not be spread equally throughout the plant tissue. Therefore, it is possible to test one leaf on a plant and get a positive TMV result, while another leaf may yield a negative TMV result.

If one plant is infected in a combo pot, will the others become infected?

Yes, it is possible for one plant to spread TMV to a neighboring plant just by growing together as their leaves come in contact with one another.

Is TMV spread by insects?

No, TMV is not spread by the most common greenhouse insects that often vector other viruses, like thrips and aphids. In addition, beneficial insects have not been linked to spreading TMV. However, there are a couple minor exceptions that may only be applicable to certain production facilities. First, pollinators such as bumble bees used in the pollination of some greenhouse crops, like cucumbers and tomatoes, can spread TMV. Also, larger chewing insects – not common in greenhouse production – such as grasshoppers can spread TMV.

Can simply brushing an infected plant and then a non-infected plant spread TMV?

Yes, the slightest brush of clothing infected with TMV was sufficient to spread the virus to uninfected plants, according to a study by Losenge et al.

How can I wash my clothing between work days to ensure that the cloth is not harboring TMV?

Washing clothes with standard amounts of laundry detergent or in milk was effective to inactivate TMV on clothing to prevent spread, according to Losenge et al.

Is there a preferred hand sanitizer on transplant lines?

To our knowledge, there has not been widely published evidence that there is a preferred type of hand sanitizer for TMV. According to the Centers of Disease Control and Prevention, alcohol-based hand sanitizers are effective against human viruses with a membrane, such as Rhinovirus, also known as the common cold. Since TMV does not have a membrane, there is minimal evidence that alcohol-based hand sanitizers will inactive it. We recommend washing your hands with soap and water as frequently as possible.

On a transplanting line, we recommend that the plants be sprayed with a milk solution before going through the machine or transferred by hand. The milk solution on the plants should still be wet as they are transplanted. Employees on a transplanting line should wear gloves and periodically dip their hands in milk solutions for the most effective control.

If a grower needs to trim a basket, how do you recommend sterilizing the scissors?

We recommend dipping your scissors in a container of liquid (20 percent dry powdered milk, 80 percent water) milk to inactivate TMV and prevent spread of the virus. Milk has been established to be a good disinfectant on cutting tools. Consider plug and liner dips into plant growth regulators for aggressive species in combination pots for next season.

How often should you remix a 1:10 bleach solution for disinfecting the greenhouse floors and benches?

We recommend that a grower remix a bleach solution every four hours.

How important are the at-home TMV tests in the overall management strategy for TMV?

We recommend using the at-home TMV strips as a tool in a methodical approach to test and rogue all plants that test positive, whether they have symptoms or not. We also recommend sending in samples of plants to a diagnostic lab, such as MSU Diagnostic Services, to have documentation of your results should you need it in the future.

What should I do with my weed mats if I have had the virus in my greenhouse?

We recommend you dispose and replace the weed mat if you have had TMV in your greenhouse this season.

What if I have potentially-infected petunias in baskets above tomato transplants?

Tomato plants can also be a host of TMV, so we recommend moving the tomatoes into a less risky area or switching the location of your baskets, if possible.

A grower performed a TMV test and it did not come back positive in 30 minutes, but came back positive only after 24 hours. Which result is accurate?

Always follow the manufacturer’s instructions for virus testing kits. The positive result after 24 hours is likely a false positive.

Tobacco Mosaic Virus or TMV was the first virus discovered. In 1886 Adolf Mayer described the plant disease. Back then bacterias were the main culprit for diseases. Mayer noted the disease and everyone thought it was a bacteria. This changed in the 1930’s. It became obvious that TMV was not a bacteria. An agent (the virus) was shown to infect plants after being crystallized, something bacteria can not do. One lab used a filtration system to prevent bacteria from reaching the plants. In this lab TMV-infected plants. These experiments proved that TMV was not a bacteria. If not a bacteria what was it? Enter the electron microscope. Photos of TMV were taken with an electron microscope and viruses were discovered.

Positive-sense single-stranded RNA plant virus that infects many members of Solanaceae family. It can infect over 350 different species. The virus causes stunted growth and leaf discoloration.

TMV infects tobacco, many members of Solanaceae (nightshade family). This includes tomatoes, peppers, and potatoes. All economically important and tasty. According to Penn State Extension TMV can infect over 350 different species. It is difficult to measure the economic impact of Tobacco Mosaic Virus or any plant virus. A 2010 estimate of plant disease damage in George found that virus diseases cost $559,100 worth of damages to producers of Blackberry (Williams-Woodward, J. 2010.). For one crop in one state viruses cost half a million dollars. Imagine what the number would be for all crops in the world. Finding solutions for plant viruses would increase the amount of food we can produce. How does it spread?

Can Tobacco Mosaic Virus be spread from Cigarettes?

Yes. If infected tobacco leaves were used in the cigarettes then touching the cigarette can lead to spreading TMV.

That’s another reason not to smoke. How else does TMV spread? Tobacco Mosaic Virus spreads from contact. Humans are the primary vector, or way it spreads. We touch an infected plant or surface that has the virus and then touch a susceptible plant. The virus can stay viable for a long time which is how a cigarette can spread the disease. Other items such as tools, lab coats, clothing, or gloves can also spread TMV.

Cultural Practices

In integrated pest management, one of the first steps to take after you have identified a problem and decide to act is cultural practices. This includes hand washing practice, cleaning tools, and organizing your workspace. All of these sound simple but they can lead to a huge reduction in spending diseases. A team (Lewandowski, D. J., Hayes, A. J., and Adkins, S. 2010) looked at disinfectants for TMV and other viruses on petunias.

The team found that often petunia plants do not show symptoms of TMV even if they were infected. Clippers were shown to transfer the virus to up to 20 plants after being used on an infected plant. Plants not showing signs or symptoms and Clippers transferring the diseases to plants makes it clear for a need of disinfectant for tools. They tested many chemical disinfectants including milk. They found that Non-fat Dried Milk was easy to get, safe, and one of the best disinfectants they tested.

Targets

When scientist develops antibiotics they look for differences between humans and bacteria. These differences are called targets. The antibiotics focus on these targets. Here is an example of how targets work. Antibiotics block essential proteins in bacteria. Humans have different proteins so the antibiotics don’t affect us. What are the targets for TMV? The virus is made up of two components: genetic information and a protein coat. The protein coat for Tobacco Mosaic Virus is made from only one protein. This protein is only found in this virus, not found in plants or other organisms. The protein that makes up the coat would be an ideal target.

Virus Predator

In ecology, if this was an invasive animal we would look to introduce a predator. The predator would then control the population. What is the predator of a virus? A virus is only genetic information and a protein coat. As shown the protein coat is a target and the weak point to attack. There is something that could attack a protein coat: a prion. Prions are deformed proteins that can cause other proteins to be deformed. If you think of the proteins that make up the coat as a resource. When a prion is introduced it would reduce that resource and reduce the ability of the virus to reproduce. What would a protein coat prion look like? What does the virus look like? Here is an illustration of the virus.

Author: Thomas Splettstoesser (www.scistyle.com)

The proteins are stacked very closely together. They twist at an angle making a spiral. For a prion all that would be needed would be to make the proteins to twist at a different angle when assembling. This would cause them to assemble to another vessel that the genetic information can not use, preventing the virus from being made. The infected cell would then be killed and release non-viable protein coats. Engineering a system that does this will take time. What is another way to provide protection to the plant with tools we already have? Genetic engineering is one way.

Digestive Enzymes

In the case of Papaya genetic information was inserted to save it from a virus. This method can be used with single strained viruses like Tobacco Mosaic Virus. The opposite strain of the genetic information of the virus is inserted into the plant. The plant produces the genetic material. When a virus inserts the genetic information to take over the cell it is blocked by the other stain copy. The two copies combine and prevent the virus from reproducing. The problem with this techniques is that it only works for a specific virus and the whole plant is wasting resources by continuously producing a negative copy of genetic information of the virus. How can this be improved?

Instead of the whole plant, only parts of the plant should be focused on defense. Tobacco Mosaic Virus spreads through contact and through the phloem. Companion cells in the plant could be designed to express digestive enzymes to target the protein coat of TMV. This would keep the phloem free from the virus and prevent the virus from spreading to other parts of the plant.

Summary

Ideas covered:

  • Using Disinfectant such as milk on tools between cuttings
  • Developing a prion for the protein coat
  • Inserting genes to produce digestive enzymes in companion cells in the phloem

When suggesting ideas I like to suggest one idea that can be implemented right away, one idea that would be a long term solution. For Tobacco Mosaic Virus the idea that can be applied right now is using milk or Non-fat Dried Milk as a disinfectant. The long-term solutions are typically inspired by ecology. In this case, it is finding a predator for a virus which is a prion. Developing a prion for the protein coat of a virus is a challenge, however, if done would provide long-term passive control of the virus.

You can read more of what I wrote about plant science

Work Cited:

Tobacco Mosaic Virus. Wikipedia. https://en.wikipedia.org/wiki/Tobacco_mosaic_virus

Tobacco Mosaic Virus – How To Protect Your Cannabis Plants

Tobacco mosaic virus, as the name suggests, is a virus common to tobacco plants. TMV causes splotchy and twisted leaves, leaving a strange mottling or mosaic pattern in its wake. It can also slow growth and reduce yields. TMV was the first plant virus to be discovered.

Worse, TMV appears to have spread to other kinds of plants. These include tomatoes, peppers, eggplant, spinach, and marigold. It also appears that cannabis is susceptible to TMV. And although TMV cannot hurt the grower, it can prevent your plants from being successful.

SPOTTING SYMPTOMS

Plants with TMV have a very distinctive look to them. Leaves will be twisted and curved in ways unnatural to the plant. The leaves will also feature yellow stripes, spots, and a strange mottled, mosaic pattern.

Symptoms can be observed on several leaves or on just a few. Some plants are just carriers and never display symptoms themselves. It is also easier to see the leaf mottling if the affected plant is partly in the shade.

Specific visual cues that your plants have been infected with TMV include:

  • Strange leaf colour: Brown leaves with “burnt” edges, pale or yellow stripes in old and new growth, and dark purple or black patches are one sign. So is the yellowing of the leaves between the veins. A mottled, mosaic pattern is a major mark of TMV.
  • Stagnation in growth: Both old and new growth can be affected by TMV. If you plant appears not to be growing as it should be, it could be that TMV is slowing it down. Wilting is another possible sign of TMV infection, as is slowed root spread.
  • Abnormal growth: Leaves grow in a strange, twisted pattern. They also appear webbed, curling under or upwards in odd ways.
  • Strange Stems: The stems can be either significantly weakened or appear in strange colours like red or purple.
  • Anaemic buds: Your buds will not get nice and fat, but will stay small.

TREATING INFECTED PLANTS

So far, the incidence of this condition have not been proven – only reported – in cannabis plants. However, the news is not good if you suspect your plant has become infected. There is no cure. An infected plant will have TMV forever. Your main goal, in other words, is to find the infected plants and remove them from the grow. TMV appears to spread via contact. As viruses can also be present in pollen and seeds, they can live for a long time in a grow room. They can survive on grow room equipment, carpets, soil, and dead plant matter.

That is why it is also essential to immediately quarantine and remove any plant you suspect is infected, pronto. Be sure to fully sanitise all grow room surfaces of any TMV traces before starting your next grow cycle.

A NOTE ON PREVENTING INFECTION IF YOU SMOKE

If you smoke cannabis or tobacco, you run the risk of carrying the virus on your hands. As such, it is a good idea to always wash your hands before coming into contact with any plants.

Tomato mosaic virus and tobacco mosaic virus

Quick Facts

  • Tomato mosaic virus (ToMV) and Tobacco mosaic virus (TMV) are hard to distinguish.
  • Tomato mosaic virus (ToMV) can cause yellowing and stunting of tomato plants resulting in loss of stand and reduced yield.
  • ToMV may cause uneven ripening of fruit, further reducing yield.
  • Tobacco mosaic virus (TMV) was once thought to be more common on tomato.
  • TMV is usually more of a tobacco pathogen than a tomato pathogen.

Host and pathogen

ToMV infects tomato most commonly, but the virus can also infect pepper, potato, apple, pear, cherry and numerous weeds, including pigweed and lamb’s quarters.

Symptoms may differ on different hosts. TMV has a very wide host range, affecting numerous crops, ornamentals and weeds including cucumber, lettuce, beet, pepper, tomato, petunia, jimson weed and horsenettle.

Signs and symptoms

Green and yellow mosaic pattern on leaf infected with TMV

Overall, tomato mosaic virus symptoms can be varied and hard to distinguish from other common tomato viruses. A definitive diagnosis can be accomplished by submitting a sample to the University of Minnesota Plant Disease Clinic.

  • Mottled light and dark green on leaves. Tobacco Mosaic virus symptoms on a tomato seedling
  • If plants are infected early, they may appear yellow and stunted overall.
  • Leaves may be curled, malformed, or reduced in size.
  • Spots of dead leaf tissue may become apparent with certain cultivars at warm temperatures.
  • Fruits may ripen unevenly.
  • Reduced fruit number and size.
  • Yellowish rings may form if fruit ripens in warm weather.
  • Fruits may show internal browning just under the skin (brownwall).

Environment

  • Symptoms may be suppressed during cool temperatures. As a result, infected seedlings may not display symptoms until moved to a warm environment.

Biology and disease cycle

Tomato mosaic virus and tobacco mosaic virus can exist for two years in dry soil or leaf debris, but will only persist one month if soil is moist. The viruses can also survive in infected root debris in the soil for up to two years.

Seed can be infected and pass the virus to the plant but the disease is usually introduced and spread primarily through human activity. The virus can easily spread between plants on workers’ hands, tools, and clothes with normal activities such as plant tying, removing of suckers, and harvest.

The virus can even survive the tobacco curing process, and can spread from cigarettes and other tobacco products to plant material handled by workers after a cigarette. Proper hand washing and sterilization of tools and equipment is essential to preventing spread this disease.

Once inside a plant, the virus multiplies resulting in the symptoms described above.

Resistant varieties

There are numerous tomato varieties that are resistant to one or the other of the viruses. These are usually denoted in seed catalogs, often with the code ToMV after the variety name if resistant to tomato mosaic virus and TMV if resistant to tobacco mosaic virus. There are only a few varieties that are resistant to both viruses.

Several popular rootstocks for grafted tomatoes can also confer resistance to varieties that may not normally be resistant.

Tomato varieties resistant to ToMV and TMV

ToMV Resistant TMV Resistant Resistant to both Resistant Rootstock
Bolseno Big Beef BHN-444 Estamino (ToMV)
Tomimaru Muchoo Celebrity Health Kick DRO138TX (ToMV)
Pink Wonder BHN-871 Sophya Colossus (TMV)
Beorange Clermont Talladega Maxifort (TMV)
Pozzano Geronimo SuperNatural (TMV)
Sunpeach Sungold RST-04-105-T (TMV)

A more extensive list of resistant tomato varieties can be found at Cornell University’s Vegetable MD Online.

Cultural control

  • Use certified disease-free seed or treat your own seed.
    • Soak seeds in a 10% solution of trisodium phosphate (Na3PO4) for at least 15 minutes.
    • Or heat dry seeds to 158 °F and hold them at that temperature for two to four days.
  • Purchase transplants only from reputable sources. Ask about the sanitation procedures they use to prevent disease.
  • Inspect transplants prior to purchase. Choose only transplants showing no clear symptoms.
  • Avoid planting in fields where tomato root debris is present, as the virus can survive long-term in roots.
  • Wash hands with soap and water before and during the handling of plants to reduce potential spread between plants.
  • Disinfect tools regularly — ideally between each plant, as plants can be infected before showing obvious symptoms.
    • Soaking tools for 1 minute in a 1:9 dilution of germicidal bleach is highly effective.
    • Or a 1-minute soak in a 20% weight/volume solution of nonfat dry milk and water is also very effective.
    • When pruning plants, have two pruners and alternate between them to allow proper soaking time between plants.
  • Avoid using tobacco products around tomato plants, and wash hands after using tobacco products and before working with the plants.
    • Tobacco in cigarettes and other tobacco products may be infected with either ToMV or TMV, both of which could spread to the tomato plants.
  • Scout plants regularly. If plants displaying symptoms of ToMV or TMV are found, remove the entire plant (including roots), bag the plant, and send it to the University of Minnesota Plant Diagnostic Clinic for diagnosis.
  • If ToMV or TMV is confirmed, employ stringent sanitation procedures to reduce spread to other plants, fields, tunnels and greenhouses.
    • Completely pull up and burn infected plants. Do not compost infected plant material.
    • After working with diseased plants, thoroughly disinfect all tools and hands as outlined above.
    • For added security against spread, keep separate tools for working in the diseased area and avoid working with healthy plants after working in an area with diseased plants.
    • At the end of the season, burn all plants from diseased areas, even healthy-appearing ones, or bury them away from vegetable production areas.
    • Disinfect stakes, ties, wires or any other equipment between growing seasons using the methods noted above.

Chemical control

There are currently no chemical options that are effective against either virus.

Anna Johnson; Michelle Grabowski, Extension educator and Angela Orshinsky, Extension plant pathologist

Reviewed in 2015

Integrated Pest Management

Fact Sheets > Vegetables > Crop Specific Articles > Tomatoes

Mosaic Diseases of Tomatoes

Common Mosaic and Cucumber Mosaic

Pathogens. Tobacco mosaic virus (TMV) and the closely related tomato mosaic virus (ToMV) cause common mosaic; cucumber mosaic virus (CMV) causes cucumber mosaic. Both diseases cause stunting of the plants and a lowering of yield. For both diseases, symptoms can vary widely, depending on the age of the plant, the variety of tomato, the strain of the virus causing the disease, and the environmental conditions.

TMV is a worldwide pathogen and one of the first plant viruses that scientists described. It has been important in Europe since the mid-1800s. In the United States, it was first reported in Connecticut on tobacco in 1899 and on tomato in 1909. It has a very wide range of hosts, including tomato and the related plants of eggplant, nightshade weeds, pepper, potato, and tobacco. TMV is seen on apple, beet, sugar beet, buckwheat, currant, grape, pear, spinach, and turnip, as well. In addition, ornamentals, foxglove, phlox, snapdragon, zinnia, and weeds of the amaranth and goosefoot families are affected.

CMV is another widespread virus. It was first reported in the 1900’s in several places in North America. It is now considered to be worldwide. It has a very wide host range, which includes tomato, carrot, celery, cucurbits, legumes, lettuce, spinach, pepper, dahlia, delphinium, columbine, geranium, petunia, phlox, zinnia, viola, and many weeds, such as chickweed, pokeweed, and milkweed.

Symptoms. Common mosaic (TMV/ToMV) often causes leaves to be stunted or elongated, in a condition called “fernleaf.” This name is due to the strong resemblance of these leaves to leaves of many kinds of ferns. The youngest leaves may be curled. Leaves may be mottled yellow and dark green. This is the symptom which gives the disease the name “mosaic.” The dark green areas may be raised. Mottling usually occurs most severely on plants grown under low light and low temperature, conditions which may exist in a greenhouse during the winter. Leaf stunting and distortion are usually worse under these conditions, as well.

Stem streaking occasionally occurs with dark streaks that are either sparse and short or prevalent and long. Such stems are easily broken and have brown areas inside. Fruit is rarely affected. It may be mottled or a brownish bronze color inside, which can be seen through the thin skin of the fruit. Fruit may show uneven ripening or yellow rings, as well.

Severe strains of TMV/ToMV cause the lower leaves to turn downward at the petiole, become rough and crinkled or corrugated, and possibly cause the leaflets to curl downward at the edges. Younger leaves may have extensive yellow to white areas with dark green blisters.

When some tomato varieties are infected with TMV/ToMV and kept at high temperature conditions (80o to 85o F) for a prolonged time, they develop dead areas on leaves, stems and roots.

Cucumber mosaic virus (CMV) causes plants to become yellow, bushy and very stunted. Leaves may be extremely distorted and malformed. Leaflets are often very narrow – this is called “shoestring”. Often the leaves on one portion of the plant (e.g., the top or the bottom) show severe symptoms, while those higher or lower in the plant are less affected. Other leaf symptoms include a yellow and green mottling similar to tobacco mosaic symptoms. Severely affected plants produce few fruit.

Identification of the Diseases. It is difficult to diagnose which virus is present without the assistance of an experienced diagnostician. The fernleaf and the shoestring symptoms are very similar, and the mosaic symptoms are indistinguishable. Control and prevention measures are very different for the two diseases, so accurate diagnosis is important.

Prevention. TMV is spread readily by touch. The virus can survive on clothing in bits of plant debris for about two years, and can easily enter a new plant from a brief contact with a worker’s contaminated hands or clothing. Tobacco products can carry the virus, and it can survive on the hands for hours after touching the tobacco product. Ensure that workers do not carry or use tobacco products near the plants, and wash well (with soap to kill the virus) after using tobacco products. Ensure that workers wear clothing not contaminated with tomato, tobacco or other host-plant material. Exclude non-essential people from greenhouses and growing areas.

Choose resistant varieties. Use disease-free seed and transplants, preferably certified ones. Avoid the use of freshly harvested seed (2 years old is best if non-certified seed is used). Seed treatment with heat (2 to 4 days at 158o F using dry seed) or trisodium phosphate (10% solution for 15 minutes) has been shown to kill the virus on the outside of the seed and, often, most of the virus inside the seed as well. Care must be taken to not kill the seeds, though. Use a two-year rotation away from susceptible species. In greenhouses, it is best to use fresh soil, as steaming soil is not 100% effective in killing the virus. If soil is to be steamed, remove all parts of the plant from the soil, including roots. Carefully clean all plant growing equipment and all greenhouse structures that come into contact with plants.

When working with plants, especially when picking out seedlings or transplanting, spray larger plants with a skim milk solution or a solution made of reconstituted powdered or condensed milk. Frequently dip hands, but not seedlings, into the milk. Wash hands frequently with soap while working with plants, using special care to clean out under nails. Rinse well after washing. Tools should be washed thoroughly, soaked for 30 minutes in 3% trisodium phosphate and not rinsed.

Another method for control of this disease is to artificially inoculate plants with a weak strain of the virus. This will not cause symptoms on the plants but protect them against disease-causing strains of the virus. This is used commonly in Europe, but strains of the virus are not yet available in the United States due to concerns about the possibility of the weak virus strains causing disease on the plants.

Cucumber mosaic is spread in a nonpersistant manner by aphids. It is not spread by seed. Control weeds, many of which are host species. Surrounding tomato fields with a taller, non-susceptible plant, such as corn, may help shield the plants from aphids blowing in from other areas. See current recommendations for control of aphids, although it is generally considered that insecticides will not control this disease. The aphids pick the virus up from the plants in about a minute and are able to spread it immediately. Insecticides take longer than this to kill the aphids. Mineral oil sprays can be used to prevent the virus from being transmitted.

At this time there are no tomato varieties resistant to CMV.

References.

By: Pamela S. Mercure, IPM Program Assistant, University of Connecticut, 1998

Reviewed by: T. Jude Boucher, IPM, University of Connecticut. 2012

The information in this document is for educational purposes only. The recommendations contained are based on the best available knowledge at the time of publication. Any reference to commercial products, trade or brand names is for information only, and no endorsement or approval is intended. The Cooperative Extension System does not guarantee or warrant the standard of any product referenced or imply approval of the product to the exclusion of others which also may be available. The University of Connecticut, Cooperative Extension System, College of Agriculture and Natural Resources is an equal opportunity program provider and employer.

Tobacco Mosaic Virus (TMV)

by Nebula Haze

Tobacco Mosaic Virus (TMV) is a virus that is commonly found in tobacco plants which causes splotchy or twisted leaves, strange mottling symptoms (a “mosaic”), slowed growth, and reduced yields. Mosaic virus has spread to several other species of plants, and there is evidence that cannabis plants may be able to catch mosaic virus, too. Although mosaic virus can’t hurt you (the grower), it can prevent plants from growing as fast or yielding as well as they could have.

These mottled leaves could potentially be signs of mosaic virus in plants

Can Cannabis Plants Catch Mosaic Virus?

These pictures show the symptoms of what several marijuana growers believe to be the result of mosaic virus, including twisted, curved leaves, yellow stripes, spots and a mosaic pattern. However, it’s important to note that these symptoms could potentially be caused by a mutation or other genetic factor. It’s also possible that other plant problems such as heat, root rot, stress, nutrient deficiencies, etc. could cause similar symptoms. You may need to worry about TMV if the affected plants are growing slowly, seem sickly, and generally aren’t producing normally.

The leaves may look like they have uneven stripes of light and dark green. Yellowing is worse on the parts of the leaves that are deformed and twisted. For many plants suspected to have mosaic virus, the dark green areas tend to be somewhat thicker than the lighter portions of the leaf.

Curved leaves with yellow stripes or mottling are usually considered the main symptom of mosaic virus.

In my experience, some growers swear their crops have been greatly affected by TMV, while others deny that it’s actually even spread to cannabis plants at all. The mosaic virus can be difficult to test for, even in a lab. The main problem with TMV is that it may cause plants to grow slowly and produce poorly. If your plant is growing fast and healthy, with no other symptoms, and you’re not noticing it spreading from plant to plant, you probably shouldn’t get too worried.

You probably don’t need to worry about TMV if…

  • Affected plants are otherwise healthy and fast-growing
  • You’re not noticing the symptoms spread from plant to plant
  • It seems to be genetic (for example common among all plants of a strain) but you’re not seeing symptoms on unrelated plants
  • You think another problem may be causing the symptoms, such as nutrient deficiencies, root problems, heat stress, etc.

When it comes to cannabis plants, curved leaf “fingers” can be caused by many things, including a random mutation, incorrect pH, watering problems, root problems or other deficiencies. However, with TMV the twisted growth is accompanied by a speckling/mosaic pattern.

Several leaves throughout the plant can display symptoms, or it may just be one or two leaves. With other species that get mosaic virus, some plants are silent carriers and may never show any symptoms.

It’s easier to see the leaf mottling if the affected plant surface is partly in the shade.

Mosaic virus can potentially stunt young plants that display a lot of symptoms, and may also cause odd, fern-like growth on young marijuana leaves.

There are sometimes yellow spots or speckles that appear on the unhealthy parts of the leaves.

A plant virus can be hard to pin down, since many factors can cause similar symptoms. Unfortunately, a virus can also transfer from plant to plant, so a grower might think they’re dealing with an environmental factor instead of an infectious disease.

However, when you have unexplained symptoms that look like pictures of TMV, it’s definitely something to consider! Luckily, mosaic virus appears to be pretty rare in marijuana plants!

Fun Fact: Tobacco Mosaic Virus was the first virus ever to be discovered!

The mosaic virus can attack a wide range of plants, including tomato, pepper, eggplant, tobacco, spinach, petunia, marigold, and maybe even our beloved herb marijuana.

Here’s a pic of a tobacco plant with confirmed TMV – the mottled leaves are the main symptom of the virus besides overall slow growth.

Here’s a pic of a squash plant that has caught Mosaic Virus

Euphorbia viguieri plant infected with mosaic virus

How can cannabis plants catch mosaic virus?

The mosaic virus has spread from tobacco plants and is known to infect at least 125 species of plants, including tomatoes, peppers, cucumbers, and many types of flowers. But as of yet, it’s not widely accepted that it can infect cannabis.

The pictures presented today are all possibly cannabis plants with mosaic virus. They appear to follow the symptoms of mosaic virus in other plant species, and the problem seems to spread from plant to plant like a virus. What do you think? Just regular plant problems or something more?

Do these cannabis plants have mosaic virus?

At this point, cannabis growers haven’t confirmed for certain that these leaf symptoms are caused by mosaic virus!

The thing that really stinks about TMV is if it does affect cannabis plants, that also means it can travel from plant to plant by direct contact, just like the virus does in tobacco plants. It’s known that the mosaic virus can also live in the soil, or be transferred from one plant to another via your hands. Some growers have claimed to see the symptoms after exposing their plants to tobacco or tobacco smoke.

Unfortunately, few cannabis growers have the equipment or the means to test if a plant actually has TMV, and a lot of the leaf symptoms could possibly be caused by other things!

Without proper testing, we pretty much have to go by comparing and guessing! I’ve never seen these symptoms myself in real life, but I would love to see it because I could test if I could get other plants to “catch” it via physical contact!

How do you treat a cannabis plant has mosaic virus?

We’re not sure if marijuana can catch TMV, but we likely should treat it the same way we do with other types of plants that catch mosaic virus.

Unfortunately, when it comes to mosaic virus, there is no cure. An infected weed plant will have TMV forever, though it may not always be actively showing symptoms. If you believe you have a marijuana plant with TMV, your main goal is to prevent it from spreading to other plants!

In greenhouse and commercial operations, the main way to deal with mosaic virus is to dispose of all affected plants, including any soil they were growing in, and enforce a strict policy of hand washing between touching plants. Luckily TMV probably won’t kill your plants, and there’s no evidence it will hurt you if you harvest the plant, but infected plants grow slower and end up producing smaller yields so you definitely want to keep it out of your marijuana garden!

Have you ever seen cannabis plants infected by Mosaic Virus? Let us know!

Could the Symptoms be Caused by Something Else?

Some cannabis plants may show mutations such as variegation (two-toned leaves), and this normal and natural phenomenon may be confused for TMV. One difference is the plant otherwise grows fast and healthy.

Two-toned leaves (variegation) is a common mutation. Nothing to worry about if plants are otherwise healthy and fast-growing.

There are other plant problems that can cause similar symptoms. It’s always a good idea to investigate and see if it might be something else!

Diagnose Your Sick Plant!

How Does TMV Spread?

“Mosaic” disease is caused by a virus. The tobacco mosaic virus is very stable and can persist in contaminated soil, in infected plant debris, on or in the seed coat, and in manufactured tobacco products. The virus is transmitted readily from plant to plant by mechanical means.

This may simply involve picking up the virus while working with infected plant material, then inoculating healthy plants by rubbing or brushing against them with contaminated tools, clothing, or hands.

Virus diseases cannot be “cured” once a plant is infected!

Therefore, every effort should be made to prevent introduction of virus diseases into the garden.

Sanitation and cleanliness is the primary means of controlling virus diseases. Infected plants should be removed immediately to prevent spread of the pathogens. The use of tobacco products during cultural practices should be avoided to prevent inoculation of plants with the tobacco mosaic virus.

Anyone who uses tobacco or works with infected plant material should wash their hands thoroughly in soapy water before handling marijuana plants!

More Cannabis Pests, Bugs & Viruses

Jump to…

Cannabis bugs, mold and other annoying pests!

Diagnose Your Sick Plant!

How to stop marijuana nutrient deficiencies

7 Steps to Cure Most Marijuana Growing Problems

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