Posts Tagged ‘plants’

Fungal killers. Or not.

Monday, September 9th, 2013

Histoplasma capsulatum A couple of nice recent papers about fungi and disease – or not:

 

Pathogens are often described by the nature of their relationship with their hosts. At one extreme are species that are entirely dependent on their host to complete their life cycle (often called obligate parasites). At the other are opportunistic species, which live as saprobes on dead organic matter, but can also invade living organisms (often called facultative pathogens). In between lies an array of combinations ranging in their degree of host dependency and ability to cause disease.

Another way to categorize pathogens is according to their pathogenic lifestyle and disease characteristics. In this case, different terminology is used for plant and animal pathogens: plant-attacking fungi are usually categorized according to the way they feed on the host, e.g., biotrophic or necrotrophic pathogens. Fungi that cause disease in animals are usually described according to the type of disease they cause, e.g., superficial or invasive mycoses. Therefore, we tend to think about fungal pathogens of plants and animals in different terms and treat them separately. Yet fungi attacking animals or plants are actually closely related. Moreover, close examination of animal and plant pathosystems reveals that fungal pathogens in both groups share similar infection strategies and sometimes even cause similar symptoms (although similarity in symptoms doesn’t necessarily indicate similar mechanism). For example, pH-lowering molecules, such as oxalic acid, are virulence factors against plant, animal, and insect hosts. This warrants revisiting the terminology and the way in which we think about fungal pathogens of animals and plants.

Fungi Infecting Plants and Animals: Killers, Non-Killers, and Cell Death. (2013) PLoS Pathog 9(8): e1003517. doi:10.1371/journal.ppat.1003517

 

And on to:

 

The yeast Candida albicans is well known as the most common agent of symptomatic fungal disease, but its more typical role is as a permanent resident of the healthy gastrointestinal microbiome. Longitudinal molecular typing studies indicate that disseminated C. albicans infections originate from individuals’ own commensal strains, and the transition to virulence is generally thought to reflect impaired host immunity. Nevertheless, the ability of this commensal pathogen to thrive in radically different host niches speaks to the existence of functional specializations for commensalism and disease. To investigate the C. albicans commensal lifestyle, researchers developed a mouse model of stable gastrointestinal candidiasis in which the animals remain healthy, despite persistent infection with high titers of yeast. Using this model, they found that a C. albicans mutant lacking the Efg1 transcriptional regulator had enhanced commensalism, such that mutant cells strongly outcompeted wild-type cells in mixed infections.

Passage through the mammalian gut triggers a phenotypic switch that promotes Candida albicans commensalism. (2013) Nature Genetics 45, 1088–1091. doi:10.1038/ng.2710
Among ∼5,000,000 fungal species, C. albicans is exceptional in its lifelong association with humans, either within the gastrointestinal microbiome or as an invasive pathogen. Opportunistic infections are generally ascribed to defective host immunity but may require specific microbial programs. Here we report that exposure of C. albicans to the mammalian gut triggers a developmental switch, driven by the Wor1 transcription factor, to a commensal cell type. Wor1 expression was previously observed only in rare genetic backgrounds, where it controls a white-opaque switch in mating. We show that passage of wild-type cells through the mouse gastrointestinal tract triggers WOR1 expression and a novel phenotypic switch. The resulting GUT (gastrointestinally induced transition) cells differ morphologically and functionally from previously defined cell types, including opaque cells, and express a transcriptome that is optimized for the digestive tract. The white-GUT switch illuminates how a microorganism can use distinct genetic programs to transition between commensalism and invasive pathogenesis.

 

Extreme virus resistance in plants

Friday, June 14th, 2013

Extreme resistance When a virus infects a plant, a slient war rages, fought with RNA weapons.

 

Multiple and complex layers of defense help plants to combat pathogens. A first line of defense relies on the detection, via dedicated host-encoded receptors, of signature molecules (so called pathogen-associated molecular patterns, PAMPs) produced by pathogens. In turn, this PAMP-triggered immunity (PTI) may be itself antagonized by adapted pathogens that have evolved virulence effectors to target key PTI components. Host plants react to PTI suppression by producing disease resistance (R) proteins that recognize virulence effectors and activate highly specific resistance called Effector Triggered Immunity (ETI). It has been noted that RNA silencing, a sequence-specific antiviral defense response based on the production of virus-derived 21–24 nt small RNAs on the one hand, and its suppression by virulence effectors, called viral suppressors of RNA silencing (VSRs) on the other, are conceptually similar to PTI. A new paper in PLOS Pathogens supports this hypothesis by showing that extreme resistance is indeed activated following detection, in specific host species, of the VSR activity of a viral virulence effector. The ensuing antiviral immunity displays many characteristics of ETI, suggesting that one or several R proteins must sense the integrity of the host silencing machinery.

 

Extreme Resistance as a Host Counter-counter Defense against Viral Suppression of RNA Silencing. (2013) PLoS Pathog 9(6): e1003435. doi:10.1371/journal.ppat.1003435
RNA silencing mediated by small RNAs (sRNAs) is a conserved regulatory process with key antiviral and antimicrobial roles in eukaryotes. A widespread counter-defensive strategy of viruses against RNA silencing is to deploy viral suppressors of RNA silencing (VSRs), epitomized by the P19 protein of tombusviruses, which sequesters sRNAs and compromises their downstream action. Here, we provide evidence that specific Nicotiana species are able to sense and, in turn, antagonize the effects of P19 by activating a highly potent immune response that protects tissues against Tomato bushy stunt virus infection. This immunity is salicylate- and ethylene-dependent, and occurs without microscopic cell death, providing an example of “extreme resistance” (ER). We show that the capacity of P19 to bind sRNA, which is mandatory for its VSR function, is also necessary to induce ER, and that effects downstream of P19-sRNA complex formation are the likely determinants of the induced resistance. Accordingly, VSRs unrelated to P19 that also bind sRNA compromise the onset of P19-elicited defense, but do not alter a resistance phenotype conferred by a viral protein without VSR activity. These results show that plants have evolved specific responses against the damages incurred by VSRs to the cellular silencing machinery, a likely necessary step in the never-ending molecular arms race opposing pathogens to their hosts.

 

 

Plant Virus Ecology

Sunday, May 26th, 2013

Plant Viruses

The latest in the “Pearls” series from PLOS Pathogens is about plant virus ecology. In this short and highly accessible article, Marilyn Roossinck provides an excellent primer for the non-initiated:

  • Plant Virus Biodiveristy
  • Plant Viruses and Invasive Species
  • Viruses, Plants and Insects
  • Persistent Plant Viruses
  • Mutualistic Viruses of Plants

 

Plant Virus Ecology. (2013) PLoS Pathog 9(5): e1003304. doi:10.1371/journal.ppat.1003304
Viruses have generally been studied either as disease-causing infectious agents that have a negative impact on the host (most eukaryote-infecting viruses), or as tools for molecular biology (especially bacteria-infecting viruses, or phage). Virus ecology looks at the more complex issues of virus-host-environment interactions. For plant viruses this includes studies of plant virus biodiversity, including viruses sampled directly from plants and from a variety of other environments; how plant viruses impact species invasion; interactions between plants, viruses and insects; the large number of persistent viruses in plants that may have epigenetic effects; and viruses that provide a clear benefit to their plant hosts (mutualists). Plants in a non-agricultural setting interact with many other living entities such as animals, insects, and other plants, as well as their physical environment. Wild plants are almost always colonized by a number of microbes, including fungi, bacteria and viruses. Viruses may impact any of these interactions.

 

BTV VLPs, OMG

Thursday, May 23rd, 2013

BTV VLPs Bluetongue is a severe disease of ruminants, notably sheep and cattle. The causal agent, the dsRNA Bluetongue virus, is spread by an insect vector and occurs in its vector’s habitat in temperate climates throughout much of the world. BTV is the type member of genus Orbivirus in the family Reoviridae, with 26 known serotypes. When bluetongue first broke out in the United Kingdom in autumn of 2007, the disease was already rapidly spreading throughout continental Europe, causing high mortality rates in sheep and having a detrimental effect on the livestock trade through trade restrictions and loss of stock. The only effective weapon against the disease is control of the spread of BTV through rigorous vaccination programmes. Currently available commercial vaccines are based on both inactivated virus and live, attenuated strains and protect against a single serotype or multiple serotypes when provided as a cocktail. However, the possibility of recombination between the live vaccine strain(s) and wild-type virus in infected animals, leading to the emergence of new infectious strains has motivated efforts to develop safer vaccines.

One approach in the development of an inherently safe vaccine has been the production of Bluetongue virus-like particles (VLPs). BTV has a nonenveloped icosahedral structure, with four main structural proteins (VP3, VP7, VP5 and VP2) arranged in concentric shells around the segmented double-stranded RNA genome and minor structural and nonstructural proteins involved in virus replication. French et al. have shown that these four structural proteins, expressed in insect cells using a baculovirus expression system, assemble into virus-like particles devoid of nucleic acid.

This paper describes plant-based high-level expression of assembled subcore-, core- and virus-like particles of BTV serotype 8. Purified preparations of the VLPs, consisting of all four structural proteins, elicited an immune response in sheep and provided protective immunity against challenge with a South African BTV-8 field isolate. This demonstrates that plant expression provides an economically viable method for producing complex VLPs, such as those of BTV, with the desired biological properties. It represents a significant advance in the use of plant-based systems for the production of complex biopharmaceuticals. The methods employed could also be applied to other situations where the expression of multiple proteins is required, such as the reconstruction of metabolic pathways.

 

A method for rapid production of heteromultimeric protein complexes in plants: assembly of protective bluetongue virus-like particles. Plant Biotechnol J. 06 May 2013 doi: 10.1111/pbi.12076
Plant expression systems based on nonreplicating virus-based vectors can be used for the simultaneous expression of multiple genes within the same cell. They therefore have great potential for the production of heteromultimeric protein complexes. This work describes the efficient plant-based production and assembly of Bluetongue virus-like particles (VLPs), requiring the simultaneous expression of four distinct proteins in varying amounts. Such particles have the potential to serve as a safe and effective vaccine against Bluetongue virus (BTV), which causes high mortality rates in ruminants and thus has a severe effect on the livestock trade. Here, VLPs produced and assembled in Nicotiana benthamiana using the cowpea mosaic virus-based HyperTrans (CPMV-HT) and associated pEAQ plant transient expression vector system were shown to elicit a strong antibody response in sheep. Furthermore, they provided protective immunity against a challenge with a South African BTV-8 field isolate. The results show that transient expression can be used to produce immunologically relevant complex heteromultimeric structures in plants in a matter of days. The results have implications beyond the realm of veterinary vaccines and could be applied to the production of VLPs for human use or the coexpression of multiple enzymes for the manipulation of metabolic pathways.

 

Plant viruses as gene delivery vehicles

Wednesday, May 1st, 2013

Cowpea Chlorotic Mottle Virus The use of animal virus-like particles (VLPs) as vectors for the delivery of genes to mammalian cells has been explored for many years. Plant viruses are almost without exception “just” genetic material (DNA or RNA) surrounded by a shell composed of the capsid protein (CP) – with no membrane envelope. Plant-derived VLPs have not been used for direct gene delivery and expression. There have been no attempts to use a spherical plant viral capsid to deliver heterologous genes for expression in mammalian cells, even though there are several independent demonstrations of the internalization of plant virus by cells.

In light of there being no direct demonstration of spherical plant viruses disassembling and thereby releasing their contents in animal cells, this paper asks the question: Can heterologous genes in spherical plant VLPs be made available to a mammalian cell and their protein products synthesized?

Getting RNA into cells is a major barrier to future theraputic approaches, so robust systems are urgently needed.

 

Reconstituted plant viral capsids can release genes to mammalian cells. Virology. 19 April 2013 doi: 10.1016/j.virol.2013.03.001
The nucleocapsids of many plant viruses are significantly more robust and protective of their RNA contents than those of enveloped animal viruses. In particular, the capsid protein (CP) of the plant virus Cowpea Chlorotic Mottle Virus (CCMV) is of special interest because it has been shown to spontaneously package, with high efficiency, a large range of lengths and sequences of single-stranded RNA molecules. In this work we demonstrate that hybrid virus-like particles, assembled in vitro from CCMV CP and a heterologous RNA derived from a mammalian virus (Sindbis), are capable of releasing their RNA in the cytoplasm of mammalian cells. This result establishes the first step in the use of plant viral capsids as vectors for gene delivery and expression in mammalian cells. Furthermore, the CCMV capsid protects the packaged RNA against nuclease degradation and serves as a robust external scaffold with many possibilities for further functionalization and cell targeting.

How to stop the bugs eating your lunch

Tuesday, March 26th, 2013

Nepenthes gracilis Like other carnivorous plants, Nepenthes species grow on poor soil. They need to complement their mineral nutrients – primarily with nitrogen and phosphorus – from caught and digested prey. When visiting the pitfall traps, the attracted prey, mainly arthropods, falls into the trap, drowns and is digested by the enzyme cocktail of the pitcher fluid.

Due to the fact that closed Nepenthes pitchers have no direct contact with the environment, it has been widely claimed that their pitcher fluid is sterile and that all proteins and compounds identified in this pitcher fluid are solely plant-derived. But only two experiments had been conducted to demonstrate the sterility of pitcher liquid: fluid taken from a closed pitcher was plated either on plain nutrient agar (Hepburn, 1918) or on meat agar plates (Lüttge, 1964) and incubated for several days. In no case were any bacterial colonies detected and the authors concluded that the pitcher fluid is sterile. However, the presence of microbes cannot be excluded by such simple experiments because most micro-organisms cannot be grown in culture.

Researchers have now analysed the composition of Nepenthes digestive fluid from closed pitchers to reveal whether or not pitchers are really sterile inside and how these plants manage to keep microbial growth under control. Thecontent of proteins, inorganic ion compositions and secondary metabolites were studied. In addition, the effect of pitcher fluid on microbial growth was investigated. The results reveal that the fluid of closed Nepenthes pitchers is composed provides anti-microbial conditions. Thus these plants can avoid, at least to some extent, the growth of microbes that compete with the plant for the prey-derived nutrients available in the pitcher.

 

Secreted pitfall-trap fluid of carnivorous Nepenthes plants is unsuitable for microbial growth. (2013) Annals of Botany 111 (3): 375-383

 

Plant virus expression vectors and production of biopharmaceutical proteins

Wednesday, September 26th, 2012

Potyvirus It is a sad fact that a leading cause of infant mortality in developing countries continues to be from infectious diseases which are quite preventable. New technologies which can both deliver vaccines en mass and without the need for syringes, as well as induce a strong mucosal immune response, could revolutionize the accessibility of much needed vaccines in developing countries. Approaches range from the use of inhalation devices which deliver the vaccine antigen in the form of an aerosol spray, to patches containing microneedles which can deliver the desired vaccine antigen across the skin barrier. A third tactic involves the development of plant production platforms as delivery systems for oral vaccines.

 

Plant virus expression vectors set the stage as production platforms for biopharmaceutical proteins. (2012 ) Virology 433(1): 1-6. doi: 10.1016/j.virol.2012.06.012
Transgenic plants present enormous potential as a cost-effective and safe platform for large-scale production of vaccines and other therapeutic proteins. A number of different technologies are under development for the production of pharmaceutical proteins from plant tissues. One method used to express high levels of protein in plants involves the employment of plant virus expression vectors. Plant virus vectors have been designed to carry vaccine epitopes as well as full therapeutic proteins such as monoclonal antibodies in plant tissue both safely and effectively. Biopharmaceuticals such as these offer enormous potential on many levels, from providing relief to those who have little access to modern medicine, to playing an active role in the battle against cancer. This review describes the current design and status of plant virus expression vectors used as production platforms for biopharmaceutical proteins.

How do plant viruses induce disease?

Tuesday, February 28th, 2012

Potyvirus Plant viruses are biotrophic pathogens that need living tissue for their multiplication and thus, in the infection-defence equilibrium, they do not normally cause plant death. In some instances virus infection may have no apparent pathological effect or may even provide a selective advantage to the host, but in many cases it causes the symptomatic phenotypes of disease. These pathological phenotypes are the result of interference and/or competition for a substantial amount of host resources that can disrupt host physiology to cause disease. This interference/competition affects a number of genes, which seems to be greater the more severe the symptoms that they cause. Induced or repressed genes belong to a broad range of cellular processes, such as hormonal regulation, cell cycle control and endogenous transport of macromolecules, among others. In addition, recent evidence indicates the existence of interplay between plant development and antiviral defence processes, and that interference among the common points of their signalling pathways can trigger pathological manifestations. This review provides an update on the latest advances in understanding how viruses affect substantial cellular processes, and how plant antiviral defences contribute to pathological phenotypes.

 

How do plant viruses induce disease? Interactions and interference with host components. (2011) J Gen Virol. 92(12): 2691-2705

Top 10 of Top 10′s ?

Thursday, January 12th, 2012

TMV I recently wrote about the top 10 plant viruses in molecular plant pathology. Well Ed didn’t like that list, so he’s published his own:

  1. Tobacco mosaic virus (TMV)
  2. Tomato spotted wilt virus (TSWV)
  3. Tomato yellow leaf curl virus (TYLCV)
  4. Cucumber mosaic virus (CMV)
  5. Potato virus Y (PVY)
  6. Cauliflower mosaic virus (CaMV)
  7. African cassava mosaic virus (ACMV)
  8. Plum pox virus (PPV)
  9. Brome mosaic virus (BMV)
  10. Potato virus X (PVX)

“I see only ONE virus in the major list – African cassava mosaic begomovirus (ACMV) – that infects and causes severe losses in one of the four major food crops grown on this planet: all the rest, excepting viruses infecting the also-ran potato, are pathogens of fruits, vegetables or horticulturally-important plants. Or hardly pathogenic at all, as in the case of BMV – and before anyone argues, I probably have the best collection of African (and other) isolates of the virus in the world, and a lot of experience of it in the field.”

I have a feeling this could go on for some time :-)