MicrobiologyBytes

Rapid advances of genomics and metagenomics lead not only to the rapid growth of sequence databases but to discovery of fundamentally novel types of genetic elements. The discovery and characterization of giant viruses that infect unicellular eukaryotes, in particular members of the family Mimiviridae infecting amoeba, over the last decade revealed a remarkable new class of agents that are typical viruses by structure and reproduction strategy but exceed many parasitic bacteria in size and genomic complexity. Like bacteria, the giant viruses (sometimes called giruses) possess their own parasites and their own mobilomes, i.e. communities of associated mobile genetic elements.

The first virus infecting a giant virus, the Sputnik virophage, was isolated from a mimivirus-infected acanthamoeba and shown to replicate within the mimivirus factories and partially inhibit the reproduction of the host mimivirus. The second virophage, named Mavirus, is a parasite of the Cafeteria roenbergensis virus (CroV), a distant relative of the mimiviruses. The third virophage genome was isolated from the Antarctic Organic Lake (hence OLV, Organic Lake Virophage) where it apparently controls the reproduction of its virus host classified as a phycodnavirus. The three well-characterized virophages possess small isocahedral virions and genomes of 20 to 25 kilobase encoding 21 to 26 proteins each. Although the virophages are similar in genome size and structure and are generally construed as related, only a minority of the virophage genes are homologous.

Analysis of the Mavirus genome resulted in the discovery that this virophage shared 5 homologous genes with the large, self-replicating eukaryotic transposable elements of the Maverick/Polinton class (hereinafter Polintons). The Polintons that are scattered among genome of diverse eukaryotes and reach high abundance in some protists, such as Trichomonas vaginalis, have long been considered ‘virus-like’ transposons because of their large size (20 kb and larger) and the presence of several genes that are common in viruses but not in other transposable elements. The Mavirus shows by far the closest affinity with the Polintons among the currently known viruses, and accordingly, it has been proposed that the Polintons evolved from the virophages.

In addition to the virophages, the giant viruses host several other groups of mobile elements. These include self-splicing introns, inteins, putative bacterial-type transposons and the most recently discovered novel linear plasmids named transpovirons. The transpovirons are highly abundant genetic elements associated with several giant viruses of the Mimiviridae family that contain only 6 to 8 genes two of which are homologous to genes of the Sputnik virophage, indicating multiple gene exchanges within the giant virus mobilome.

Virophages, polintons, and transpovirons: a complex evolutionary network of diverse selfish genetic elements with different reproduction strategies. (2013) Virology Journal, 10:158 doi: 10.1186/1743-422X-10-158
Recent advances of genomics and metagenomics reveal remarkable diversity of viruses and other selfish genetic elements. In particular, giant viruses have been shown to possess their own mobilomes that include virophages, small viruses that parasitize on giant viruses of the Mimiviridae family, and transpovirons, distinct linear plasmids. One of the virophages known as the Mavirus, a parasite of the giant Cafeteria roenbergensis virus, shares several genes with large eukaryotic self-replicating transposon of the Polinton (Maverick) family, and it has been proposed that the polintons evolved from a Mavirus-like ancestor. We performed a comprehensive phylogenomic analysis of the available genomes of virophages and traced the evolutionary connections between the virophages and other selfish genetic elements. The comparison of the gene composition and genome organization of the virophages reveals 6 conserved, core genes that are organized in partially conserved arrays. Phylogenetic analysis of those core virophage genes, for which a sufficient diversity of homologs outside the virophages was detected, including the maturation protease and the packaging ATPase, supports the monophyly of the virophages. The results of this analysis appear incompatible with the origin of polintons from a Mavirus-like agent but rather suggest that Mavirus evolved through recombination between a polinton and an unknownvirus. Altogether, virophages, polintons, a distinct Tetrahymena transposable element Tlr1, transpovirons, adenoviruses, and some bacteriophages form a network of evolutionary relationships that is held together by overlapping sets of shared genes and appears to represent a distinct module in the vast total network of viruses and mobile elements. The results of the phylogenomic analysis of the virophages and related genetic elements are compatible with the concept of network-like evolution of the virus world and emphasize multiple evolutionary connections between bona fide viruses and other classes of capsid-less mobile elements.

The human JC polyomavirus is a bit of a mystery. Many people are infected with it, but few become ill as a result. This virus bides its time, waiting for your immune systen to let its guard down, then wham! People infected with HIV, those who have AIDS, or those receiving immunomodulatory therapies for autoimmune diseases are at serious risk for progressive multifocal leukoencephalopathy (PML), where the virus can spread from the kidney to the central nervous system and cause a fatal, demyelinating disease.

Recent reports have shown that virus isolates from PML patients often have distinct changes within the major capsid protein. This paper shows that that these mutations result in abolished engagement of the carbohydrate receptor motif necessary for infection. Viruses with PML-associated mutations are not infectious in glial cells, suggesting that they may play an alternative role in PML. Interesting stuff, suggesting that interaction with cell surface receptors is an important determinant of tissue tropism and JC virus pathogenesis for PML, even though the best defence remains a healthy immune system.

Progressive Multifocal Leukoencephalopathy-Associated Mutations in the JC Polyomavirus Capsid Disrupt Lactoseries Tetrasaccharide c Binding. (2013) mBio 4(3): e00247-13 doi: 10.1128/mBio.00247-13
The human JC polyomavirus (JCPyV) is the causative agent of the fatal, demyelinating disease progressive multifocal leukoencephalopathy (PML). The Mad-1 prototype strain of JCPyV uses the glycan lactoseries tetrasaccharide c (LSTc) and serotonin receptor 5-HT2A to attach to and enter into host cells, respectively. Specific residues in the viral capsid protein VP1 are responsible for direct interactions with the α2,6-linked sialic acid of LSTc. Viral isolates from individuals with PML often contain mutations in the sialic acid-binding pocket of VP1 that are hypothesized to arise from positive selection. We reconstituted these mutations in the Mad-1 strain of JCPyV and found that they were not capable of growth. The mutations were then introduced into recombinant VP1 and reconstituted as pentamers in order to conduct binding studies and structural analyses. VP1 pentamers carrying PML-associated mutations were not capable of binding to permissive cells. High-resolution structure determination revealed that these pentamers are well folded but no longer bind to LSTc due to steric clashes in the sialic acid-binding site. Reconstitution of the mutations into JCPyV pseudoviruses allowed us to directly quantify the infectivity of the mutants in several cell lines. The JCPyV pseudoviruses with PML-associated mutations were not infectious, nor were they able to engage sialic acid as measured by hemagglutination of human red blood cells. These results demonstrate that viruses from PML patients with single point mutations in VP1 disrupt binding to sialic acid motifs and render these viruses noninfectious.

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.

A nice short article by John Coffin looking at pathogen-host co-evolution.

Virions at the Gates: Receptors and the Host–Virus Arms Race. (2013) PLoS Biol 11(5): e1001574. doi:10.1371/journal.pbio.1001574
All viruses need to bind to specific receptor molecules on the surface of target cells to initiate infection. Virus–receptor binding is highly specific, and this specificity determines both the species and the cell type that can be infected by a given virus. In some well-studied cases, the virus-binding region on the receptor has been found to be unrelated to the receptor’s normal cellular function. Resistance to virus infection can thus evolve by selection of mutations that alter amino acids in the binding region with minimal effect on normal function. This sort of positive selection can be used to infer the history of the host–virus “arms race” during their coevolution. In a new study, Demogines et al. use a combination of phylogenetic, structural, and virological analysis to infer the history and significance of positive selection on the transferrin receptor TfR1, a housekeeping protein required for iron uptake and the cell surface receptor for at least three different types of virus. The authors show that only two parts of the rodent TfR1 molecule have been subject to positive selection and that these correspond to the binding sites for two of these viruses—the mouse mammary tumor virus (a retrovirus) and Machupo virus (an arenavirus). They confirmed this result by introducing the inferred binding site mutations into the wild-type protein and testing for receptor function. Related arenaviruses are beginning to spread in human populations in South America as the cause of often fatal hemorrhagic fevers, and, although Demogines et al. could find no evidence of TfR1 mutations in this region that might have been selected as a consequence of human infection, the authors identified one such mutation in Asian populations that affects infection with these viruses.

Bacteria communicate with one another via quorum-sensing signal molecules. This paper describes the first example of quorum-sensing molecules participating in interspecies bacterial cell death. This is an interesting observation in its own right – but think about this: these peptides potentially provide the basis for a new class of antibiotics which trigger death by acting from outside the cell.

Novel Quorum-Sensing Peptides Mediating Interspecies Bacterial Cell Death. (2013) mBio 4(3): e00314-13 doi: 10.1128/mBio.00314-13
Escherichia coli mazEF is a toxin-antitoxin stress-induced module mediating cell death. It requires the quorum-sensing signal (QS) “extracellular death factor” (EDF), the penta-peptide NNWNN (EcEDF), enhancing the endoribonucleolytic activity of E. coli toxin MazF. Here we discovered that E. coli mazEF-mediated cell death could be triggered by QS peptides from the supernatants (SN) of the Gram-positive bacterium Bacillus subtilis and the Gram-negative bacterium Pseudomonas aeruginosa. In the SN of B. subtilis, we found one EDF, the hexapeptide RGQQNE, called BsEDF. In the SN of P. aeruginosa, we found three EDFs: the nonapeptide INEQTVVTK, called PaEDF-1, and two hexadecapeptides, VEVSDDGSGGNTSLSQ, called PaEDF-2, and APKLSDGAAAGYVTKA, called PaEDF-3. When added to a diluted E. coli cultures, each of these peptides acted as an interspecies EDF that triggered mazEF-mediated death. Furthermore, though their sequences are very different, each of these EDFs amplified the endoribonucleolytic activity of E. coli MazF, probably by interacting with different sites on E. coli MazF. Finally, we suggest that EDFs may become the basis for a new class of antibiotics that trigger death from outside the bacterial cells.

Hmm, this sounds like a good exam question for next year’s paper. I wonder if any of my students read this blog? ;-)

The modulation of apoptosis by oncogenic viruses. (2013) Virology Journal, 10: 182 doi:10.1186/1743-422X-10-182
Transforming viruses can change a normal cell into a cancer cell during their normal life cycle. Persistent infections with these viruses have been recognized to cause some types of cancer. These viruses have been implicated in the modulation of various biological processes, such as proliferation, differentiation and apoptosis. The study of infections caused by oncogenic viruses had helped in our understanding of several mechanisms that regulate cell growth, as well as the molecular alterations leading to cancer. Therefore, transforming viruses provide models of study that have enabled the advances in cancer research. Viruses with transforming abilities, include different members of the Human Papillomavirus (HPV) family, Hepatitis C virus (HCV), Human T-cell Leukemia virus (HTLV-1), Epstein Barr virus (EBV) and Kaposi’s Sarcoma Herpesvirus (KSHV).Apoptosis, or programmed cell death, is a tightly regulated process that plays an important role in development and homeostasis. Additionally, it functions as an antiviral defense mechanism. The deregulation of apoptosis has been implicated in the etiology of diverse diseases, including cancer. Oncogenic viruses employ different mechanisms to inhibit the apoptotic process, allowing the propagation of infected and damaged cells. During this process, some viral proteins are able to evade the immune system, while others can directly interact with the caspases involved in apoptotic signaling. In some instances, viral proteins can also promote apoptosis, which may be necessary for an accurate regulation of the initial stages of infection.

My Leicester colleague Catherine Pashley has done a lot of work in this area, so I was interested in this recent minireview in PLOS Pathogens.

• What Is Asthma?
• Why Do Fungi Make Spores? And a Guide to Terminology
• Do Fungal Spores Cause Asthma?
• Which Species Are Associated with Asthma?
• If Identification to Species Matters, Will New Tools Provide Needed Data?

Asthma and the Diversity of Fungal Spores in Air. (2013) PLoS Pathog 9(6): e1003371. doi:10.1371/journal.ppat.1003371
The diversity of fungal spores in air is vast, but research on asthma focuses on a handful of easily identified, culturable species. Ecologists are developing new tools to probe communities and identify the full complement of fungi in habitats. These tools may enable identification of novel asthma triggers, but scientists involved in public health or medicine rarely interact with mycologists focused on ecology. With this primer, my aim is to facilitate communication by providing doctors with a basic, modern guide to spores, by teaching mycologists the essential facts of asthma, and by providing patients with a succinct summary of what is known about spores and asthma. By highlighting the use of emerging metagenomics technologies in ecology, I intend to illustrate how these tools might be used to more thoroughly understand the potential diversity of fungi involved in asthma.

See: 10 things you should know about novel coronavirus (nCoV)

I’ve spent quite a bit of time in the last week marking exam essays about papillomaviruses, so it’s good to relax by reading a few recent journal articles about …. papillomaviruses ;-)

Animal papillomaviruses. Virology. 24 May 2013 pii: S0042-6822(13)00266-3. doi: 10.1016/j.virol.2013.05.007
We provide an overview of the host range, taxonomic classification and genomic diversity of animal papillomaviruses. The complete genomes of 112 non-human papillomavirus types, recovered from 54 different host species, are currently available in GenBank. The recent characterizations of reptilian papillomaviruses extend the host range of the Papillomaviridae to include all amniotes. Although the genetically diverse papillomaviruses have a highly conserved genomic lay-out, deviations from this prototypic genome organization are observed in several animal papillomaviruses, and only the core ORFs E1, E2, L2 and L1 are present in all characterized papillomavirus genomes. The discovery of papilloma-polyoma hybrids BPCV1 and BPCV2, containing a papillomaviral late region but an early region encoding typical polyomaviral nonstructural proteins, and the detection of recombination breakpoints between the early and late coding regions of cetacean papillomaviruses, could indicate that early and late gene cassettes of papillomaviruses are relatively independent entities that can be interchanged by recombination.

Papillomavirus E6 oncoproteins. 24 May 2013 Virology pii: S0042-6822(13)00248-1. doi: 10.1016/j.virol.2013.04.026
Papillomaviruses induce benign and malignant epithelial tumors, and the viral E6 oncoprotein is essential for full transformation. E6 contributes to transformation by associating with cellular proteins, docking on specific acidic LXXLL peptide motifs found on these proteins. This review examines insights from recent studies of human and animal E6 proteins that determine the three-dimensional structure of E6 when bound to acidic LXXLL peptides. The structure of E6 is related to recent advances in the purification and identification of E6 associated protein complexes. These E6 protein-complexes, together with other proteins that bind to E6, alter a broad array of biological outcomes including modulation of cell survival, cellular transcription, host cell differentiation, growth factor dependence, DNA damage responses, and cell cycle progression.