How many bacteria?

Tree of Life I’ve asked the question How many different bacterial species are there on this blog before. It seems that we may be approaching an answer.

A census of species of Archaea and Bacteria published recently showed that, despite ever-increasing sequencing efforts, the PCR-based retrieval of 16S rRNA genes is approaching saturation. On average, 95% of the genes analyzed today are identical to those present in public databases, with rarefaction analysis indicating that about one-third of the bacterial and archaeal diversity has already been covered. Despite previous estimates of up to 1012 microbial species, the option should be considered that the census of Archaea and Bacteria on planet Earth might yield only millions of species after all.

After All, Only Millions? MBio. 2016 Jul 5; 7(4). pii: e00999-16. doi: 10.1128/mBio.00999-16

Posted in Microbiology | Tagged , , , | Leave a comment

Emerging Paramyxoviruses – Receptor Tropism and Zoonotic Potential

MERS-CoV Emerging infectious disease events are dominated by zoonoses: infections that are naturally transmissible from animals to humans or vice versa. A worldwide survey of ~5,000 bat specimens identified 66 novel paramyxovirus species – more than double the existing total within this family of viruses. Also, novel paramyxoviruses are continuously being discovered in other species, such as rodents, shrews, wild and captivated reptiles and farmed fish, as well as in domestic cats and horses. Paramyxoviruses exhibit one of the highest rates of cross-species transmission among RNA viruses, and paramyxoviral infection in humans can cause a wide variety of often deadly diseases. Thus, it is important to understand the determinants of cross-species transmission and the risk that such events pose to human health. Whilst pathogen diversity and human encroachment play important roles, This paper focuses on receptor tropism and envelope determinants for zoonosis of emerging paramyxoviruses.

 

Emerging Paramyxoviruses: Receptor Tropism and Zoonotic Potential. (2016) PLoS Pathog 12(2): e1005390. doi: 10.1371/journal.ppat.1005390

Posted in Microbiology | Tagged , , , , , , | Leave a comment

HTLV-1 Replication: An Update

HTLV-1 Replication I spent more than 10 years working on HTLV, the first human retrovirus to be discovered, but it was a long time ago (propably before most of the people reading this now were born :-)
It’s good to get an update on recent research in this field, although as the authors of this article point out:

“As the first human retrovirus discovered in the early 1980s, HTLV-1 has been studied extensively, yet there is still no treatment or vaccine for HTLV-1 infection.”

Molecular Studies of HTLV-1 Replication: An Update. (2016) Viruses 8(2), 31; doi: 10.3390/v8020031
Human T-cell leukemia virus type 1 (HTLV-1) was the first human retrovirus discovered. Studies on HTLV-1 have been instrumental for our understanding of the molecular pathology of virus-induced cancers. HTLV-1 is the etiological agent of an adult T-cell leukemia (ATL) and can lead to a variety of neurological pathologies, including HTLV-1-associated-myelopathy/tropical spastic paraparesis (HAM/TSP). The ability to treat the aggressive ATL subtypes remains inadequate. HTLV-1 replicates by (1) an infectious cycle involving virus budding and infection of new permissive target cells and (2) mitotic division of cells harboring an integrated provirus. Virus replication initiates host antiviral immunity and the checkpoint control of cell proliferation, but HTLV-1 has evolved elegant strategies to counteract these host defense mechanisms to allow for virus persistence. The study of the molecular biology of HTLV-1 replication has provided crucial information for understanding HTLV-1 replication as well as aspects of viral replication that are shared between HTLV-1 and human immunodeficiency virus type 1 (HIV-1). Here in this review, we discuss the various stages of the virus replication cycle—both foundational knowledge as well as current updates of ongoing research that is important for understanding HTLV-1 molecular pathogenesis as well as in developing novel therapeutic strategies.

Posted in Microbiology | Tagged , , , , , , | Leave a comment

Towards a cure for herpesvirus infection with CRISPR/Cas9

Bovine herpesvirus Most adults carry multiple herpesviruses. Following the initial acute infection, these viruses establish life-long infections in their hosts and cause cold sores, keratitis, genital herpes, shingles, infectious mononucleosis, and other diseases. Some herpesviruses can cause cancer in man. During the latent phase of infection, the viruses remain dormant for long periods of time, but retain the capacity to cause occasional reactivations, that may lead to disease. This new study suggests that attacking herpesvirus DNA with CRISPR/Cas9 genome editing technology can suppress virus replication and, in some cases, lead to elimination of the virus from infected cells.

The snag? Well this is a purely in vitro study in cultured cell, not even an animal model. Will this approach be safe and effecting in humans? It will take some years to find that out.

 

CRISPR/Cas9-Mediated Genome Editing of Herpesviruses Limits Productive and Latent Infections. (2016) PLoS Pathog 12(6): e1005701. doi: 10.1371/journal.ppat.1005701

Herpesviruses infect the majority of the human population and can cause significant morbidity and mortality. Herpes simplex virus (HSV) type 1 causes cold sores and herpes simplex keratitis, whereas HSV-2 is responsible for genital herpes. Human cytomegalovirus (HCMV) is the most common viral cause of congenital defects and is responsible for serious disease in immuno-compromised individuals. Epstein-Barr virus (EBV) is associated with infectious mononucleosis and a broad range of malignancies, including Burkitt’s lymphoma, nasopharyngeal carcinoma, Hodgkin’s disease, and post-transplant lymphomas. Herpesviruses persist in their host for life by establishing a latent infection that is interrupted by periodic reactivation events during which replication occurs. Current antiviral drug treatments target the clinical manifestations of this productive stage, but they are ineffective at eliminating these viruses from the infected host. Here, we set out to combat both productive and latent herpesvirus infections by exploiting the CRISPR/Cas9 system to target viral genetic elements important for virus fitness. We show effective abrogation of HCMV and HSV-1 replication by targeting gRNAs to essential viral genes. Simultaneous targeting of HSV-1 with multiple gRNAs completely abolished the production of infectious particles from human cells. Using the same approach, EBV can be almost completely cleared from latently infected EBV-transformed human tumor cells. Our studies indicate that the CRISPR/Cas9 system can be effectively targeted to herpesvirus genomes as a potent prophylactic and therapeutic anti-viral strategy that may be used to impair viral replication and clear latent virus infection.

Posted in Microbiology | Tagged , , , , , , | Leave a comment

Virtual Simulations as Preparation for Lab Exercises

Robot As an alternative to tutorials, virtual laboratory simulations represent a new way of preparing students for hands on exercises, such as laboratory work, and various computer based technologies are now being recognized as enabling reform of laboratory teaching practice. The aim of this paper was to investigate if a virtual laboratory can be used to replace a face to face tutorial to prepare students for a laboratory exercise – more specifically, if a group of students who prepare for a laboratory exercise in microbiology by using a virtual laboratory have comparable outcomes to a group who are given a face to face tutorial by an experienced teacher.

 

Virtual Simulations as Preparation for Lab Exercises: Assessing Learning of Key Laboratory Skills in Microbiology and Improvement of Essential Non-Cognitive Skills. (2016) PLoS ONE 11(6): e0155895. doi: 10.1371/journal.pone.0155895
A total of 189 students who were participating in an undergraduate biology course were randomly selected into a vLAB or demonstration condition. In the vLAB condition students could use a vLAB at home to ‘practice’ streaking out bacteria on agar plates in a virtual environment. In the demonstration condition students were given a live demonstration from a lab tutor showing them how to streak out bacteria on agar plates. All students were blindly assessed on their ability to perform the streaking technique in the physical lab, and were administered a pre and post-test to determine their knowledge of microbiology, intrinsic motivation to study microbiology, and self-efficacy in the field of microbiology prior to, and after the experiment.The results showed that there were no significant differences between the two groups on their lab scores, and both groups had similar increases in knowledge of microbiology, intrinsic motivation to study microbiology, as well as self-efficacy in the field of microbiology. Our data show that vLABs function just as well as face to face tutorials in preparing students for a physical lab activity in microbiology. The results imply that vLABs could be used instead of face to face tutorials, and a combination of virtual and physical lab exercises could be the future of science education.

 

Cann, AJ (2016) Increasing Student Engagement with Practical Classes Through Online Pre-Lab Quizzes. Journal of Biological Education, 50(1), 101-112.

Posted in Microbiology | Tagged , , ,

How soil bacteria escape sticky root traps

Longitudinal section of a root tip Plant root tips are covered by a protective sleeve of loosely attached border cells that can release a matrix containing proteins, polysaccharides, and DNA. In animal immune systems, extracellular DNA forms the backbone of neutrophil extracellular traps (NETs) deployed by immune cells to immobilize and kill invading microbes. Some animal pathogens can secrete DNases to degrade NETs and facilitate infection. Plant border cells release DNA-containing extracellular traps in response to the high-impact plant pathogenic bacterium Ralstonia solanacearum. R. solanacearum secretes two DNases that free the pathogen from these extracellular traps. The bacterium needs these DNases for full virulence and normal colonization of its host plants. This work reveals that, like animal pathogens, the plant pathogen R. solanacearum can overcome a DNA-based host defense system with secreted enzymes.

 

Escaping Underground Nets: Extracellular DNases Degrade Plant Extracellular Traps and Contribute to Virulence of the Plant Pathogenic Bacterium Ralstonia solanacearum. (2016) PLoS Pathog 12(6): e1005686. doi: 10.1371/journal.ppat.1005686
Plant root border cells have been recently recognized as an important physical defense against soil-borne pathogens. Root border cells produce an extracellular matrix of protein, polysaccharide and DNA that functions like animal neutrophil extracellular traps to immobilize pathogens. Exposing pea root border cells to the root-infecting bacterial wilt pathogen Ralstonia solanacearum triggered release of DNA-containing extracellular traps in a flagellin-dependent manner. These traps rapidly immobilized the pathogen and killed some cells, but most of the entangled bacteria eventually escaped. The R. solanacearum genome encodes two putative extracellular DNases (exDNases) that are expressed during pathogenesis, suggesting that these exDNases contribute to bacterial virulence by enabling the bacterium to degrade and escape root border cell traps. We tested this hypothesis with R. solanacearum deletion mutants lacking one or both of these nucleases, named NucA and NucB. Functional studies with purified proteins revealed that NucA and NucB are non-specific endonucleases and that NucA is membrane-associated and cation-dependent. Single ΔnucA and ΔnucB mutants and the ΔnucA/B double mutant all had reduced virulence on wilt-susceptible tomato plants in a naturalistic soil-soak inoculation assay. The ΔnucA/B mutant was out-competed by the wild-type strain in planta and was less able to stunt root growth or colonize plant stems. Further, the double nuclease mutant could not escape from root border cells in vitro and was defective in attachment to pea roots. Taken together, these results demonstrate that extracellular DNases are novel virulence factors that help R. solanacearum successfully overcome plant defenses to infect plant roots and cause bacterial wilt disease.

Posted in Microbiology | Tagged , , , , ,

The Feat of Packaging Eight Unique Genome Segments

The Feat of Packaging Eight Unique Genome Segments Influenza A viruses harbor a segmented RNA genome that is organized into eight distinct viral ribonucleoprotein (vRNP) complexes. Although a segmented genome may be a major advantage to adapt to new host environments, it comes at the cost of a highly sophisticated genome packaging mechanism. Newly synthesized vRNPs conquer the cellular endosomal recycling machinery to access the viral budding site at the plasma membrane. Genome packaging sequences unique to each RNA genome segment are thought to be key determinants ensuring the assembly and incorporation of eight distinct vRNPs into progeny viral particles. Recent studies using advanced fluorescence microscopy techniques suggest the formation of vRNP sub-bundles (comprising less than eight vRNPs) during their transport on recycling endosomes. The formation of such sub-bundles might be required for efficient packaging of a bundle of eight different genomes segments at the budding site, further highlighting the complexity of IAV genome packaging.

The Feat of Packaging Eight Unique Genome Segments. Viruses 2016, 8(6), 165; doi: 10.3390/v8060165

Posted in Microbiology | Tagged , , , , ,

Zika virus, the new kid on the block

Zika virus - NIAID As Europe “welcomes” Zika virus*, this short introduction by Maria Zambon is highly readable, and essential for anyone who’s been hiding under a rock for the past six months.

 

Zika virus, the new kid on the block. Euro Surveill. 2016; 21(23): pii=30255. DOI: http://dx.doi.org/10.2807/1560-7917.ES.2016.21.23.30255
According to Tolstoy, happy families were all alike, whereas unhappy families were each unhappy in their individual ways. So it is with the emergence of new virus infections. Each new virus epidemic brings misery to affected human populations, in unique ways. In the last 15 years, we have experienced the emergence and spread of Severe Acute Respiratory Syndrome (SARS), H5N1 and H7N9 influenza A viruses, pandemic influenza A(H1N1), Middle Eastern Respiratory Syndrome (MERS) and Ebola virus disease, and most recently in 2015–16, Zika virus. The wider societal impact that such infectious disease events can cause has been amply demonstrated with Ebola virus in West Africa, which was responsible for over 11,000 deaths and has inhibited economic growth in this war-torn region of the world. Each of the viruses mentioned above occupies a different ecological niche, with diverse impact on the human population (magnitude of the epidemic, disease severity) as a result of transmission characteristics, host immune response and disease pathogenesis. Serious complications and deaths from Zika virus infection have not been common: most infections are asymptomatic or very mild, although there is an association with neurological complications such as Guillain–Barré syndrome. The key issue, however, is the impact of infection on pregnancy. For most emerging viruses, classical control measures of contact tracing and quarantine will eventually break chains of transmission between humans following zoonotic infection, when human-to-human transmission occurs and infectiousness is related to symptomatic illness. However, when infection is through a vector-borne route and sexual transmission can occur from a minimally symptomatic person, such as with Zika virus infection, additional population-based control measures must be undertaken. Vector control requires sustained and determined efforts to achieve a measurable impact and may involve a range of interventions at a personal level (e.g. avoidance, mosquito nets and insecticide) and at population level (e.g. breeding genetically resistant mosquitoes). Steering towards other rational interventions requires evidence from well-documented individual case studies.

* Sexual transmission of Zika virus in an entirely asymptomatic couple returning from a Zika epidemic area, France, April 2016
Sexual transmission of Zika virus in Germany, April 2016

Posted in Microbiology | Tagged , , , , , , ,

How prions kill neurons

Hippocampal neuron Prion diseases are fatal neurodegenerative disorders that cause memory loss, impaired coordination, and abnormal movements. The molecular culprit in prion diseases is PrPSc, an infectious isoform of a host-encoded glycoprotein (PrPC) that can propagate itself by a self-templating mechanism. Whether PrPSc itself is toxic to neurons, and if so, the cellular mechanisms by which it produces neuronal pathology is unknown, in part because of the absence of suitable cell culture models.

This paper describes a hippocampal neuronal cultural system to detect the toxic effect of PrPSc on dendritic spines, which are postsynaptic elements responsible for excitatory synaptic transmission, and which are implicated in learning, memory, and the earliest stages of neurodegenerative diseases. Purified, exogenously applied PrPSc causes acute retraction of dendritic spines, an effect that is entirely dependent on expression of PrPC by target neurons, and on the on the presence of a nine-amino acid, polybasic region at the N-terminus of the PrPC molecule. Both protease-resistant and protease-sensitive forms of PrPSc cause dendritic retraction.

This culture system provides new insights into the mechanisms responsible for prion neurotoxicity, and it provides a platform for characterizing different pathogenic forms of PrPSc and testing potential therapeutic agents. Because dendritic spine loss is a common theme in many neurodegenerative conditions, including Alzheimer’s, Huntington’s, and Parkinson’s diseases, and has been suggested to contribute to clinical symptoms in patients, the researchers also suggest that their system allows direct comparisons between pathogenic mechanisms involved in prion diseases and other neurodegenerative disorders.

 

A Neuronal Culture System to Detect Prion Synaptotoxicity. (2016) PLoS Pathog 12(5): e1005623. doi: 10.1371/journal.ppat.1005623
Synaptic pathology is an early feature of prion as well as other neurodegenerative diseases. Although the self-templating process by which prions propagate is well established, the mechanisms by which prions cause synaptotoxicity are poorly understood, due largely to the absence of experimentally tractable cell culture models. Here, we report that exposure of cultured hippocampal neurons to PrPSc, the infectious isoform of the prion protein, results in rapid retraction of dendritic spines. This effect is entirely dependent on expression of the cellular prion protein, PrPC, by target neurons, and on the presence of a nine-amino acid, polybasic region at the N-terminus of the PrPC molecule. Both protease-resistant and prote- ase-sensitive forms of PrPSc cause dendritic loss. This system provides new insights into the mechanisms responsible for prion neurotoxicity, and it provides a platform for character- izing different pathogenic forms of PrPSc and testing potential therapeutic agents.

Posted in Microbiology | Tagged , , , ,